ADIOLOGY ,.,.11 --­ NCOLOGY September 2001 Vol. 35 No. 3 Ljubljana ISSN 1318-2099 ADIOLOGY AND NCOLOGY Editorial office Radiologij and OncologiJ Institute of Oncology Zaloška 2 SI-1000 Ljubljana Slovenia Phone: +386 1 4320 068 Plwne/Fnx: +386 1 4337 410 E-mail: gsersa@onko-i.si Aims and scope .TI. September 2001 Vol. 35 No. 3 Pages 161-235 ISSN 1318-2099 UDC 616-006 CODEN: RONCEM Radiologij and Oncology is a joumal devoted to publication of original contributions in diagnostic and interventional radiology, computerized tomography, ultrasound, magnetic resonance, nuclear medicine, radiotherapy, clinical and experimental oncology, radiol1iology, radiop/zysics and radia/ion pralec/ion. Editor-in-Chief Gregor Serša Ljubljana, Slovenia Executive Editor Viljem Kovac Ljubljana, S/ovenia Editorial board Marija Auersperg Ljubljana, Slovenia Nada Bešenski Zagreb, Croatia Karl H. Bohuslavizki Hamburg, Germany Haris Boka Zagreb, Croatia Nataša V. Budihna Ljubljana, Slovenia Maijan Budihna Ljubljana, Slovenia Malte Clausen Hamburg, Gemzany Christoph Clemm Miinchen, Germany Mario Corsi Udine, Italy Ljubomir Diankov Sofia, Bulgaria Christian Dittrich Vienna, Austria Ivan Drinkovic Zagreb, Croatia Gillian Duchesne Melbourne, Australia Editor-in-Chief Erneritus Tomaž Benulic Ljubljana, Slovenia Editor Uroš Smrdel Ljul1ljana, S/ovenia BelaFornet Budapest, Hungary Tullio Giraldi Trieste, Italy Andrija Hebrang Zagreb, Croatia Laszl6 Horvath Pecs, Hungary Berta Jereb Ljubljana, Slove11ia Vladimir Jevtic Ljubljana, S/ovenia H. Dieter Kogelnik Salzburg, Austria Jurij Lindtner Ljubljana, Slovenia Ivan Lovasic Rijeka, Croatia Marijan Lovrencic Zagreb, Croatia Luka Milas Houston, USA Metka Milcinski Ljubljana, Slovenia Maja Osmak Zagreb, Croatia Branko Palcic Vancouver, Canada Jurica Papa Zagreb, Croatia Dušan Pavcnik Portland, USA Stojan Plesnicar Ljublimza, S/ovenia Ervin B. Podgoršak Montreal, Canada Jan C. Roos Amsterdam, Netherlands Slavko Šimunic Zagreb, Croatia Lojze Šmid Ljubljana,Slovenia Bonit Štabuc Ljubljana, Slovenia Andrea Veronesi Avimw, Italy Živa Zupancic Ljubljana, S/ovenia Publisher Association of Radiology and Oncology Affiliated with Slovenian Medica/ Association S/ove11ian Association of Radiology, Nuclear Medicine Society, Slovenian Society far Radiotherapy and Oncology, and Slovenian Cancer Society Croatian Medica/ Association -Croatian Society of Radiology Societas Radiologorum Hungarorum Friuli-Venezia Giulia regional groups of S.I.R.M. (Ita/ian Society of Medica/ Radiology) Copyright © Radiology and Oncology. Ali rights reserved. Reader for English Mojca Cakš Key words Eva Klemencic Secretaries Milica Harisch Mira Klemencic Design Monika Fink-Serša Printed by lmprint d. o. o., Ljubljana, Slovenia Pub/ished quarterly in 700 copies Bank account number 02010-0090006751 Foreig11 currency account number 010-7100-900067 /4 NLB d.d., PodruŽllica Ljubljana Center, Ljubljana S. W.I.F. T. Code L]BASI2X Subscription fee far institutions $ 100 (16000 SIT), individua/s $ 50 (5000 SIT) The publication of this joumal is subsidized by the Ministry of Science and Tec/1110/ogy of tl1e Republic of Slovenia. Indexed and abstracted by: BIOMEDICINA SLOVENICA CHEMICAL ABSTRACTS EMBASE / Excerpta Medica Sci Base This journal is printed on acid- free paper Radiology and Oncology is avai/able on the internet at: http://www.onko-i.si/radiolog/mo.html Ljubljana, Slovenia September 2001 Vol. 35 No. 3 CONTENTS DIAGNOSTIC AND INTERVENTIONAL RADIOLOGY .TI. ISSN 1318-2099 UDC 616-006 CODEN: RONCEM Breath-hold times in patients undergoing radiological examinations: comparison of expiration and inspiration with and without hyperventilation Groell R, Schaffler GJ, Schloffer S 161 Percutaneous drainage of abdominal fluid collections that require laparatomy or relaparatomy with ultrasound gudance Miletic D, Uravic M, Fuckar Ž, Glavaš R, Topijak-Polic D 167 Ultrasound-guided central venous cannulation in patient with radical dissection on both sides of neck: case report Šustic A, Cerovic R, Juretic M 175 Bifocal primary intracranial germinoma in a child. Case report Koren A 179 NUCLEAR MEDICINE Artifacts and non-osseous uptake in bone scintigraphy. Imaging reports of 20 cases Weiner G M, Jenicke L, Miiller V, Bohuslavizki K H EXPERIMENTAL ONCOLOGY Celi electropermeabilization to small molecules in vitro: control by pulse parameters Macek-Lebar A, Miklavcic D Electroporator for in vitro cell permeabilization Puc M, Flisar K, Reberšek S, Miklavcic D CLINICAL ONCOLOGY Bone metastases from malignant melanoma: a retrospective review and analysis of 28 cases Brountzos E, Panagiotou I, Bafaloukos D, Kelekis D RADIOPHYSICS Ranking radiotherapy treatment plans: physical or biological objectives? Ebert M 215 SLOVENIAN ABSTRACTS 225 NOTICES 231 Radiology and Oncology is covered in Biomedicina Sloveni en, Chemicnl Abstmcts, EMBASE / Excerpta Medica, and Sci Base Breath-hold times in patients undergoing radiological examinations: comparison of expiration and inspiration with and without hyperventilation Reinhard Groell, Gottfried J. Schaffler, Stephan Schloffer Department of Radiology, University Hospital Graz, Austria Background. Breath-holding is necessary for imaging studies of the thorax and abdomen using computed tomography, magnetic resonance imaging or ultrasound examinations. The purpose of this study was to compare the breath-hold times in expiration and inspiration and to evaluate the effects of hyperventilation. Patients and methods. Thirty patients and 19 healthy volunteers participated in this study after informed consent was obtained in all. The breath-hold times were measured in expiration and inspiration before and after hyperventilation. Results. The mean breath-hold times in expiration (patients: 24±9sec, volunteers: 27±7sec) were signifi­cantly shorter than those in inspiration (patients: 41±20sec, p<0.001; volunteers: 62±18sec, p<0.001). Additional hyperventilation resulted in a significant increase (range: 40-60%, pŁ0.005) of the mean breath-hold times either in expiration and in inspiration and for both patients and volunteers. Conclusions. Although breath-holding in expiration is recommended for various imaging studies particu­larly of the thorax and of the abdomen, suspending respiration in inspiration enables the patient a consid­erable longer breath-hold time. Key words: tomography, X-ray computer; magnetic resonance imaging; ultrasonography; breath holding Introduction Respiratory motion may degrade imaging studies particularly of the thorax and of the abdomen. Therefore breath-holding is crucial during thoracic and abdominal examinations Received 5 June 2001 Accepted 19 June 2001 Correspondence to: Reinhard Groell, MD. Department of Radiology, University Hospital Graz, Auenbruggerplatz 9, A-8036 Graz, Austria. Phone: +43 316 385 81611; Fax: ++43 316 385 3231; E-mail: reinhard.groell@kfunigraz.ac.at using different modalities such as computed tomography, magnetic resonance imaging, digital angiography, or Doppler sonography. Breath-holding in expiration is regarded to result in more consistent organ positioning and is therefore recommended for various im­aging studies particularly of the abdomen.1 However, in the clinical experience it is easi­er to hold the breath in inspiration than in ex­piration. This is of clinical importance when the clinician has to make the decision whether the examination should be per­formed in expiration or inspiration. Previous reports have shown that the max­imum breath-hold time may be increased by hyperventilation and by administration of oxygen.2,3 However, a comparison of breath-hold capabilities between expiration and in­spiration has - to our knowledge - not been performed yet. This prompted us to prospec­tively evaluate and compare the breath-hold capabilities of patients and of healthy volun­teers in expiration and inspiration without and after hyperventilation. Patients and methods The study population consisted of 30 outpa­tients and 19 healthy volunteers. The patients (15 female, 15 male; mean age: 64±15years, range: 31-85 years) were referred to abdomi­nal ultrasound for various clinical reasons. The majority (n=17) of them were examined in routine screening. Six of the patients were smokers with a smoking history of more than ten pack-years. Four patients had a medical history of chronic obstructive pulmonary dis­ease (COPD) and two of them additionally had chronic heart failure (CHF), the patients received medical therapy for these condi­tions. The healthy volunteers (8 female, 11 male; mean age: 32±5years, range: 23­43years) were employees of our institution. Two of them were smokers with a smoking history of more than ten pack-years. None of the volunteers had known diseases of the cardio-respiratory system. All patients and healthy volunteers gave informed consent to the performance of this study. For the patients the measurements were performed while the patients were wait­ing for their ultrasound examination. During the study all participants were lying in the supine position. The patients were instructed to hold their breath in expiration ("breathe in, breathe out, hold your breath") and in inspi­ration ("breathe in, breathe out, breathe in, hold your breath"). Then the same respiratory maneuvers were performed following six deep inhalations of room air (corresponding to approximately 20-30 seconds of hyperven­tilation). A time span of at least 2 minutes Figure 1. Mean breath-hold times in patients and healthy volunteers without and after hyperventilation. was kept between the breath-holds. The order of inspiration and expiration was changed al­ternately among different subjects to mini­mize the effects of training. Statistical comparison of different respira­tory maneuvers was performed using a Student t-test with a 5% level of statistical se­curity. Multivariate analysis was performed to evaluate the influence of age, sex, smoking history, cardio-pulmonary diseases, or of the order of examinations (expiration performed before inspiration or vice versa) on the meas­ured breath-hold times. Results The mean breath-hold times in expiration were significantly shorter than those in inspi­ration both without hyperventilation (pa­tients: 24±9sec vs. 41±20sec, p<0.001; volun­teers: 27±7sec vs.62±18sec, p<0.001) and after hyperventilation (patients: 37±18sec vs. 59±29sec, p<0.001; volunteers: 42±11sec vs.87±28sec, p<0.001) (Figures 1,2). Hyper­ventilation resulted in a significant increase of the measured mean breath-hold times in expiration (patients: 24±9sec vs. 37±18sec, p<0.001; volunteers: 27±7sec vs.42±11sec, p=0.005) and inspiration (patients: 41±20sec vs. 59±29sec, p<0.001; volunteers: 62±18sec vs. 87±28sec, p=0.002). In expiration the mean breath-hold times were not statistically different (p>0.23) between patients and healthy volunteers either without or after hy­perventilation. However, in inspiration the breath-hold times of the healthy volunteers were generally longer than those of the pa­tients (p<0.003). Multivariate analysis re­vealed that in patients with COPD or CHF the breath-hold times without hyperventilation were not statistically different from those of patients without such diseases (Figure 2). After hyperventilation, the mean breath-hold times were lower in the patients with COPD or CHF (expiration: 21±10sec vs. 40±18sec, p=0.02; inspiration: 37±16sec vs. 62±30sec, p=0.05), however this comparison is limited by the small number of patients with COPD or CHF (total: n=4). While there were no sig­nificant sex differences in breath-holding without hyperventilation, after hyperventila­tion lower breath-hold times were observed in women than in men (p<0.03). Age (p>0.2), smoking history (p>0.4), or the order of ex­aminations (expiration performed before in­spiration or vice versa, p>0.1) showed no sig­nificant influence on the measured breath-hold times. Discussion In various radiological modalities the optimal image quality is achieved when the patients hold their breath during the entire study. With the advent of fast imaging modalities such as spiral computed tomography or rapid magnetic resonance imaging more and more studies may potentially be acquired within a single breath-hold. Frequently the study time lies in the order of the breath-hold time with some variations in either direction. These variations, however, may considerably influ­ence the success of the examinations. Thus, to optimize the quality of the examination, the radiologist has to consider the breath-hold capabilities of the patient. Breath hold­ing in expiration is reported to allow a more reproducible organ positioning than breath holding in inspiration.1 That is why many studies, particularly of the abdomen, should primarily be performed in expiration. Although thoracic studies usually benefit from maximal distention of the lungs as it oc­curs in inspiration, expiratory scans may be necessary e.g. in patients with obstructive lung diseases to document possible air trap­ping.4 Reviewing the literature, we found three radiological studies that evaluated the breath-hold capabilities of adults either in expiration Figure 2a,b. Scatter graphs showing the breath-hold times in expiration/inspiration without (Figure 2a) and after (Figure 2b) hyperventilation. The data points are connected for each of parameter of ventilation. The graphs indi­cate smokers (S) with a history of more than 10 pack-years, patients with COPD (P), and patients with chronic heart insufficiency (H). or in inspiration and two of these studies documented the benefit of hyperventilation and of administration of oxygen.2,3,5 We found no report that compared breath-hold­ing capabilities between expiration and inspi­ration that could be transformed to the con­ditions in a radiological setting. Some reports have investigated the physiological changes that occur during suspended respiration with special interest in oxygen saturation and heart rate, most of these studies were per­formed in divers.6-8 As part of the physiolo­gic diving reflex a decrease in the heart rate can be observed during breath-holding which was also observed by Gay and Marks.2,5 With and without hyperventilation the breath-hold times in inspiration exceeded those in expiration by approximately 50­130%. Although it is a wide-held belief that it is easier to hold the breath in inspiration than in expiration, the amount of these differences exceeded our expectations. All of the patients investigated in this study were outpatients and none of them was severely pulmonary-compromised, although four patients had a medical history of COPD and two of them additionally had CHF. The healthy volunteers were generally consider­ably younger than the patients, and most of them were physically active. This may ex­plain the longer mean times in the group of volunteers than in the patient group. In our study population hyperventilation increased the maximum breath-hold capabilities in ex­piration and inspiration, however these ef­fects were less pronounced in patients with cardiac or pulmonary diseases (Figure 1). This confirms the results of Marks et al. who demonstrated that the effects of hyperventi­lation were less beneficial in pulmonary-com­promised patients while administration of oxygen resulted in increased breath-hold times even in pulmonary-compromised pa­tients.2 Similar to the observations of Gay et al. we found no significant influence of smok­ing on the maximum breath-hold times.5 In conclusion, suspending respiration in inspiration results in considerably longer breath-hold times when compared to breath-holding in expiration. The radiologist has to decide which respiratory maneuver is best suitable to optimize the performance of the specific imaging studies. Acknowledgement We thank Mag. Friedrich J. Kofler for his as­sistance in editing the manuscript. References 1. Donovan PJ. Technique of examination and nor­mal pancreatic anatomy. In: Siegelman SS, editor. Computed tomography of the pancreas. New York: Churchill-Livingstone; 1983. p. 1-32. 2. Marks B, Mitchell DG, Simelaro JP. Breath-holding in healthy and pulmonary comprised populations: effects of hyperventilation and Oxygen Inspiration. J Magn Reson Imaging 1997; 7: 595-7. 3. Sadatoh N, Hatabu H, Takahashi M, Imanaka K, Sano A. Oxygen-assisted breath-holding in com­puted tomography. J Comput Assist Tomogr 1987; 11: 742-4. 4. Arakawa H, Webb WR. Air trapping on expirato­ry CT scans in the absence of inspiratory scan ab­normalities: correlation with pulmonary function tests and differential diagnosis. Am J Roentgenol 1998; 170: 1349-53. 5. Gay SB, Sistrom CL, Holder CA, Suratt PM. Breath-holding capabilities of adults. Invest Radiol 1994; 9: 848-51. 6. Liner MH. Cardiovascular and pulmonary re­sponses to breath-hold diving in humans. Acta Physiol Scand Suppl 1994; 620: 1-32. 7. Sterba JA. Lundgren CE. Breath-hold duration in man and the diving response induced by face im­mersion. Undersea Biomed Res 1988; 15: 361-75. 8. Butler PJ, Woakes AJ. Heart rate in humans during underwater swimming with and without breath-hold. Respir Physiol 1987; 69: 387-99. Percutaneous drainage of abdominal fluid collections that require laparotomy or relaparotomy with ultrasound guidance Damir Mileti“1, Miljenko Uravi“2, ëeljko Fu‰kar2, Robert Glavaä3, Dubravka Topljak-Poli“1 1Department of Radiology, 2Clinic for Surgery, 3Department of Gastroenterology, Clinical Hospital Rijeka, Croatia Background. The aim of the study was to determine efficacy and reliability of percutaneous abdominal drainage in surgical patients and to evaluate intercostal approach to drain subphrenic collections. Material and methods. Eighty-seven patients aged from 29 to 84 years (mean, 55.5 years) were percuta­neously drained under the sonographic guidance due to the postoperative or nonoperated abdominal collec­tion that would otherwise require laparotomy. Intercostal, subcostal, lateral and anterior approach with eight to 14 French catheters were used to evacuate abdominal collection. Results. The intercostal approach was used to drain 31 (60.8%) of 51 subphrenic collections. The mean du­ration of drainage was independent of the intercostal or subcostal drainage route, but was significantly pro­longed (p<0.05, Mann-Whitney U test) for purulent collections (median, 18 days; range 7-73 days) in com­parison to hematomas, bilomas and other nonpurulent collections (median, 11 and 6 days, respectively). Sonographically guided percutaneous drainage was a definitive method in 92% patients, with 9.2% minor complications. Successful rate for subphrenic collections was even greater (96%). Conclusions. Sonographically guided percutaneous drainage is the method of choice in the treatment of ab­dominal collections that require laparotomy. If the puncture site is at least two intercostal spaces lower than the dome of diaphragm and catheter is not introduced through the pleural effusion, intercostal drainage is equally efficient and not less secure than subcostal approach. Key words: sonography; abdomen; drainage Received 2 June 2001 Accepted 28 June 2001 Correspondence to: Damir Mileti“, MD, PhD, Department of Radiology, Clinical Hospital Rijeka, Kreäimirova 42, Croatia. Phone: + 385 51 65 83 85; E­mail: Damir.Miletic@medri.hr Introduction Despite the initial skepticism, percutaneous catheter drainage of abdominal collections is well established and widely accepted proce­dure by an interventional radiologist and sur­geons.1,2,3 Fluid collections may be drained under the sonographic, CT or fluoroscopic guidance. The intercostal or subcostal ap­proach may be used to drain subphrenic col­lections. Subcostal drainage route is general­ly recommended to avoid pleural transgression.4 The intercostal approach is preferred to drain left subphrenic collections after splenectomy.5 The aim of our study is to determine the ef­ficiency and reliability of the percutaneous abdominal drainage in surgical patients, as well as to compare the transpleural and ex-trapleural approach to drain both right and left subphrenic collections. Material and methods In five-years' period we planned 101 sono-graphically guided percutaneous drainages of the suspected abdominal collections in pa­tients who were admitted at the surgery clin­ic. Only in 3 patients the intestinal interposi­tion could not be avoided in any potential approach route, and percutaneous drainage was postponed or withdrawn. Percutaneous aspiration of the fluid content with or without instillation of antibiotics was a definitive method in 6 patients. In other five patients initial diagnostic needle aspiration confirmed solid lesion instead, and the aspiration biop­sy followed. Our study includes 87 patients, 46 male and 41 female, with percutaneous catheter drainage of abdominal fluid collection. The mean age of the patients in our study were 55.5±10.9 (SD), ranging from 29 to 84. Laparotomy or relaparotomy was planned due to clinical symptoms such as fever, ab­dominal pain, respiratory or intestinal prob­lems, palpable mass, and CT or sonographic visualization of the fluid collection. Seventy-five patients (86.2%) underwent recent abdominal surgery: of those, biliary surgery was performed in 33, gastrointestinal in 19, splenectomy in 10, liver surgery in 6, and other in 7 patients. The postoperative sonography within the first 10 days detected intraperitoneal collection. In other 12 pa­tients (13.8%) without previous laparotomy intraperitoneal or retroperitoneal fluid collec­tion was found by ultrasound or computed to­mography. In this group we detected paraco­lic abscesses due to diverticulitis in 6 patients, and retrocecal appendicitis in one patient. The retroperitoneal penetration of the de­scendent colon carcinoma caused a huge ab­scess formation with compression to the bow­el in one patient. Two collections appeared liquefied neoplasms; one was induced by the gastric ulcer perforation and the other by the missile penetration. All percutaneous drainages were per­formed under the sonographic guidance. All specimens obtained by needle aspiration were submitted for cytology, aerobic and anaerobic cultures. Nonpurulent materials were also chemically analysed. Eight to 14 French catheters were used to evacuate ab­dominal collections. Generally, we used 8 to 10 French catheters for serous collections in­cluding bile, and for intercostal approach. Twelve to 14 French catheters were used for purulent and viscous collections. The com­plete procedures consumed 10 to 30 minutes of time, depending mostly on localization. Patients were regularly followed-up the 2nd and 7th day from percutaneous catheter placement, as well as immediately after the drainage stopped. The catheter was not re­moved before sonography or CT confirmed the complete evacuation of the infected col­lection. Incomplete drainage was tolerated in particular sterile hematomas if evacuation stopped and clinical response was favorable. In other noninfected collections we also in­sisted on complete evacuation before the catheter removal. In a case of catheter mal­function or absence of clinical improvement, the catheter position and function were re­vised with sonographically controlled instilla­tion of 5-10 ccm of normal saline. We used intercostal, subcostal, lateral, and anterior approach with the patient supine, in the left or right decubitus position. The ac­cess route was elected by the previous sono-graphic examination. Intercostal drainage was performed as lower as possible to reach the collection, at least 2 intercostal spaces lower than the dome of the diaphragm was vi­sualized. If the puncture line included pleural effusion, the catheter was not applied. The linear array transducers of 5 MHz with the central canal to guide the needle were used in our series. The needle position was constantly monitored during the insertion, di­agnostic aspiration and the guide wire place­ment if the Seldinger technique was used. We also used a trocar technique, especially in the intercostal approach. When properly posi­tioned, catheters were secured in place by su­turing to the skin. Statistical differences were calculated with the Mann-Whitney U (MWU) test, and the analysis of variance (ANOVA). The procedures were conducted in accor­dance with the ethical standards of the Helsinki Declaration of 1975. Results In five-years' period we performed 89 percu­taneous catheter drainages of abdominal flu­id collections in 87 patients under the sono-graphic guidance. We detected 34 fluid collections (38.2%) in the right subphrenic space with or without subhepatic extension, 8 collections (9.0%) iso­lated in the subhepatic space, 17 collections (19.1%) in the left subphrenic space, 11 col­lections (12.4%) between the intestinal loops, 10 collections (11.2%) in the right/left para-colic gutter, and 6 collections (6.7%) in the retroperitoneum. Other rare locations includ­ed preperitoneal collections beneath the ab­dominal wall in 2 cases, and lesser sac ab­scess in one patient. In two patients we drained 2 different collections consecutively. Aspirated material was macroscopically purulent in 31, sanguineous in 11, serous in 9, and undetermined in 21 collections. In 17 collections we aspirated bile. A Gram stain, aerobic and anaerobic cultures confirmed in­fected material in 47 (52.8%) specimens. Namely, 5 hematomas, one seroma, and 10 undetermined collections were infected. Fifty-eight specimens were submitted for a chemical analysis to confirm the origin of a fluid collection. Cytologic examination indi­cated malignant tumor in four specimens. Of 51 subphrenic collections, intercostal approach was used to drain 31 (60.8%) collec­tions, 20 out of 34 (58.8%) in the right and 11 out of 17 (64.7%) in the left subphrenic space. We selected intercostal approach (Figure 1) whenever it was the shortest route to reach the collection, but only if pleural effusion could be avoided on the puncture line. In 20 (39.2%) subphrenic collections the extrapleur­al access route was preferred. We preferred the lateral approach for per­cutaneous drainage of the collections in para-colic gutters, the posterolateral approach for the retroperitoneal collections, and the ante­rior approach for preperitoneal and lesser sac collections. The access route for the collec­tions between the intestinal loops was select­ed individually to avoid small or large bowel transgression. Intestinal interposition was ex­cluded by the real-time sonographic monitor­ing of peristalsis. The median number of days of drainage for all abdominal collections was 13 days (range, 2-73 days). Significant variability re­garding the duration of drainage of different fluid qualities was detected (Figure 2, ANO­VA). The median drainage time for purulent collections was 18 days (range, 7-73 days), significantly longer (p<0.05, MWU) in com­parison to hematomas (median, 11 days; range, 5-28 days). The drainage time for bilo-mas (median, 6 days; range, 2-17 days), sero-mas and other nonpurulent collections (medi­an, 6 days; range, 3-13 days) was significantly shorter (p<0.05, MWU) compared to hemato-mas, and purulent collections. Patients stay­ed in hospital up to 20 days after the catheter insertion. If the drainage had to be prolonged (Figure 2), the outpatient care with regular weekly controls was preferred. Although 8 and 10 French catheters were used in the intercostal approach, the mean duration of drainage was not significantly dif­ferent in comparison to the subcostal drainage route where larger catheters (up to 14 French) were placed (median, 15 and 12 days, respectively). Intra-abdominal collections were com­pletely evacuated in 76 (87.4%) patients, in­cluding both patients with two different col­lections in the abdomen. In 4 (4.6%) patients with sterile hematoma evacuation was incom­plete, but percutaneous drainage was a defin­itive method. Further resolution of residual coagulated blood products after the catheter removal was followed-up on ultrasound con­trols until the resorption is completed (up to 95 days). Sonographically guided percu­tanous drainage was entirely successful in 80 patients (92%) with 82 collections. Forty-nine Figure 2. Drainage time variability for different qualities of abdominal collections. of 51 patients with subphrenic collections (96%) were cured by percutaneuos catheter drainage. Only in four patients the surgical proce­dure was necessary, but even those patients benefited from the parcutaneous drainage. Namely, body temperature and white blood cell count decreased, while the overall condi­tion improved. In the first patient with the complete bowel obstruction due to the huge extraluminal abscess formation and compres­sion we evacuated 900 ccm of purulent and necrotic material. This procedure facilitated the normal evacuation of the colon, and a pa­tient became fit for surgery. Carcinoma of the descendent colon penetrating to the retroperitoneal space was diagnosed with the barium enema after the percutaneous drainage (Figure 3). Right subphrenic collec­tion in the second patient was a result of an intestinal perforation due to carcinomatous peritoneal dissemination, and percutaneous drainage preceded the operative resection. In both cases previously existed ileus subsided. Continuous daily drainage of 500-700 ccm of bile in the third patient indicated postopera­tive biliary fistula that required a surgical re­vision. Retrocecal abscess due to perforated appendicitis in the fourth patient was drained, and appendectomy followed. In another two patients abdominal collec­tions appeared liquefied neoplasms on cytol­ogy, and percutaneous drainage was only a palliative procedure with the evident clinical improvement. Finally, one patient with he-matemesis died the 3rd day after the catheter placement. The abscess adjacent to the left hepatic lobe was diagnosed on CT and US, and sonographically guided percutaneous drainage without hepatic transgression fol- Figure 3. Percutaneous drainage of the retroperitoneal collection due to colon carcinoma invasion. Note irregular segmental stenosis of the colon. Catheter was introduced above the inguinal ligament to the left iliac fossa. lowed. We evacuated 600 ccm of purulent material. At autopsy, a bleeding gastric ulcer with localized perforation was detected. The abscess cavity was localized to the lesser sac, and a catheter was placed in the proper posi­tion. We detected 8 (9.2%) minor complications in our series, including superficial skin infec­tion in one case, chills immediately after the catheter placement due to bacteremia in 2 pa­tients, and intestinal content leakage near the subcostally introduced catheter in one patient with intestinal fistula. In one patient left sub-phrenic abscess after splenectomy recurred, and percutaneous drainage had to be per­formed again. The second attempt was fol­lowed by complete resolution of the fluid col­lection. In two patients with intercostal approach concomitant pleural effusion increased in size, without clinical impairment. Aspirated pleural fluid was sterile. Only in one patient asymptomatic pneumothorax was detected after the intercostal catheter insertion. Life-threatening complications of the sonographically guided PAD were not detect­ed in our patients. Discussion In our series 80 out of 87 patients with ab­dominal fluid collections were cured by per­cutaneous drainage. Even 96% of all sub-phrenic collections were cured without further surgical intervention. Complications of the percutaneous catheter drainage of ab­dominal collections at surgery department in our patients were minor, incomparable with the reported morbidity and mortality rate (19.4% and 4.8%, respectively) of the surgical drainage of subphrenic abscesses.6 High successful and low complication rate in comparison to the previous reports2,5 could be explained by the following: (1) five pa­tients with the incomplete evacuation of the sterile hematoma who did not need further intervention were included in the success group (2) in three patients we canceled the procedure due to the equivocal bowel inter­position (3) in 6 patients where needle inser­tion through the pleural effusion was un­avoidable, we elected percutaneous aspiration of the subphrenic collection and instillation of antibiotics if the material was purulent. Percutaneous aspiration was re­peated in 2 patients with the reaccumulation of the fluid content. Sonographic guidance provided a reliable differentiation between subphrenic and pleural accumulation due to the multiplanar imaging and real-time moni­toring of diaphragmatic movement. According to McNicholas et al. subcostal (extrapleural) drainage of subphrenic collec­tions could be difficult only on the left side, especially after splenectomy.7 In our experi­ence this problem appears on both right and left side. Namely, subphrenic inflammation usually causes the ipsilateral diaphragmatic relaxation and diminishes its respiratory movement. Therefore, liver and spleen are withdrawn from the costal arch, and the in­spiration is frequently not very helpful in subcostal approach. Although McNicholas et al. consider right-sided subphrenic collec­tions easier to drain subcostaly, liver parenchyma was frequently interposed and unavoidable in subcostal approach in our pa­tients, especially if subphrenic collection did not extend to the Morison's pouch. We elect­ed the intercostal approach whenever the col­lection could be reached avoiding pleural ef­fusion, if it was the shortest drainage route. In our experience, definitive diagnosis did not need to be established before the percuta­neous puncture of an abdominal fluid collec­tion visualized on CT or sonography. The ini­tial aspiration confirms the presence of a collection, determines whether the collection is infected and the material liquefied enough to be drainable, and establishes a safe route for the subsequent catheter insertion.4 We di-agnosed diverticulitis, large bowel carcinoma and retrocecal appendicitis on barium studies after the drainage was completed. According to our results, the drainage time for hematomas was significantly longer than for bilomas and other nonpurulent collec­tions, and in this group we had 4 incomplete evacuations. One could speculate if catheter drainage of hematomas is really justifiable, or needle aspiration should be sufficient. We recommend the catheter placement when a symptomatic collection is visualized even if sanguineous content is aspirated. Namely, five out of 11 hematomas in our series were infected, and even patients with sterile hematomas benefited from the procedure. Small, asymptomatic early postoperative col­lections are probably seromas or hematomas, requiring only ultrasound follow-up. Conclusively, sonographically guided per­cutaneous drainage is the method of choice for the treatment of symptomatic postopera­tive abdominal fluid collections. This method is also successful in nonoperative treatment of abdominal collections that otherwise re­quires explorative laparotomy. In our experi­ence, it was a definitive method in the treat­ment of the perforative diverticulitis. If the puncture site is at least two intercostal spaces lower than the dome of diaphragm and a catheter is not placed through pleural effu­sion, the intercostal drainage route for sub-phrenic collections is not less secure than the subcostal approach. If pleural effusion is un­avoidable, only percutaneous puncture and aspiration without the catheter placement should be performed. References 1. Mueller PR, Simeone JF, Butch JR. Percuteneous drainage of subphrenic abscess: a review of 62 pa­tients. AJR 1986; 147: 1237-40. 2. VanSonnenberg E, Ferruci JT Jr, Mueller PR, Wittenberg J, Simeone JF. Percuteneous drainage of abscesses and fluid collections: technique, re­sults, and applications. Radiology 1982; 142: 1-10. 3. Gerzof SG, Johnson WC. Radiologic aspects of di­agnosis and treatment of abdominal abscesses. Surg Clin North Am 1984; 64(1): 53-65. 4. Mueller PR, vanSonnenberg E, Ferrucci JT Jr: Percutaneous drainage of 250 abdominal abscess­es and fluid collections. Part II. Current procedur­al concepts. Radiology 1984; 151: 343-7. 5. VanSonnenberg E, Mueller PR, Ferruci JT Jr. Percuteneous drainage of 250 abscesses and fluid collections. Part I: Results, failures, and complica­tions. Radiology 1984; 151(2): 337-41. 6. Haluk GI, Ismail K, Salim D, Levent B, Huerkar K. Subdiaphragmantic abscesses: myths and reali­ties. A report on sixty-two cases. Int Surg 1991; 76: 84-6. 7. McNicholas MMJ, Mueller PR, Lee MJ, Echeverri J, Gazelle GS, Boland GW, et al. Percutaneous drainage of subphrenic fluid collections that occur after spenectomy: efficacy and safety of transpleural versus extrapleural approach. AJR 1995; 165: 355-9. Case report Ultrasound-guided central venous cannulation in patient with radical dissection on both sides of neck: case report Alan áusti“1, Robert Cerovi“2, Mirna Jureti“2 1Department of Anesthesiology and ICU, 2Department of Maxillo-facial surgery, University Hospital Rijeka, Croatia Background. Difficult and complicated surgical procedures in elective oncologic patients often require cen­tral venous (CV) catheterization. In certain number of cases, relevant anatomical changes, caused by basic disease, impede, or even prevent the use of standard "blind" CV cannulation. In such cases ultrasonography can be used successfully as an adjunctive method during the CV cannulation. Case report. We present the patient with a radical dissection on both sides of neck and consequent impor­tant anatomical changes; the ultrasound-guided CV cannulation is successfully performed in this patient. Conclusions. Ultrasound as a supporting method for CV catheterization in oncological patients has poten­tial benefits. Sonography as an important adjunct method in central venous cannulation is recommended in difficult cases. Key words: sonography; catheterization, central venous Introduction Difficult and complicated surgical procedures in elective patients require the central venous (CV) cannulation for monitoring and for par­enteral alimentation in the intraoperative and postoperative period. In certain number of cases relevant anatomical changes, caused by basic disease, impede, or even prevent the Received 14 July 2001 Accepted 31 July 2001 Correspondence to: Alan áusti“, M.D., Ph.D., Department. of Anesthesiology and ICU, Clinical Hospital Rijeka, T. Striěi“a 3, Rijeka 51 000, Croatia. Phone: + 385 51 21 84 07; Fax: + 385 51 21 84 07; E­mail: Alan.Sustic@mamed.medri.hr use of the standard CV cannulation. In such cases ultrasonography can be used success­fully as an adjunctive method during the CV cannulation.1-3 We present the patient with a radical dissection on both sides of the neck and consequent important anatomical chan­ges; the ultrasound guided CV cannulation is successfully performed in this patient. Case report In a 37 years old male patient, planocellular carcinoma of tongue was diagnosed six months before the present hospitalization. At that time the radical dissection on the right side of the neck was performed, as well as the suprahyoid dissection on the left side, with the extirpation of a tumor and the resection of mandible. The plastic operation of the de­fect was done with a free skin-bone flap tak­en from the left leg. Now the patient was hos­pitalized due to multiple palpable lymph nodes on the left side of the neck. The preoperative computerized tomogra­phy (CT) and the ultrasonography (US) de­tected multiple metastatic deposits on the left side of the neck with a complete obliteration and destruction of the left internal jugular vein in the middle part of the neck, and the displacement of the left carotid artery caused by metastatic processes. A somewhat smaller secondary deposit was found immediately above and beside the entrance of jugular vein into the subclavian vein. By the above-mentioned technique an im­portant segment of the right-sided subclavian artery and vein, displaced upwards cran­iodorsaly, can be visualized. As with subsequent diagnostic methods no other metastatic deposits were found, the radical dissection of the left side of the neck was indicated. The preanaesthetic examina­tion informed us of the patient's relatively good state of health, with normal laboratory findings, ASA III. Because of the magnitude of surgical intervention and the necessity of the perioperative and the postoperative mon­itoring, as well as parenteral nutrition, the central venous catheterization was indicated. Because of important anatomical and pathoanatomical changes, the preoperative right-sided subclavial catheter, under sono-graphic monitoring, was decided upon. Ultrasonic apparatus Hitachi 405 EUB, with linear transducer (5 MHz) was used sterilized as recommended by the manufacturer. Immediately before the cannulation by supra-clavial approach, the displaced subclavial vein was visualized with a few transverse and vertical scans. Finding the best vertical sec­tion through the vein, we located the direc­tion, as well as the route of a punction needle, and so we could begin the cannulation with two lumens CV catheter (16 Gauge, Arrow Inc.) with the introductor obeying the rules of a Seldinger technique. Without displacing the transducer we monitored the whole pro­cedure and after the completed cannulation, again visualized the vein with CV catheter in it, by a few transversal and vertical scans (Figure 1). With the convex transducer (5 MHz) via suprajugular access, we visualized vena anonyma (Figure 2) and with the convex transducer (3 MHz) by a standard echocar-diographic approach, right heart, respective­ly. Subsequently, through a catheter, a bolus of 10 ml 0,9% NaCl was applicated; presented us with so-called "hand-made ultrasonic con­trast" in the right atrium and we confirmed the right position of a central venous catheter. Once the position of a catheter was deter­mined, the affixation followed, without the need of radiologic control. Discussion The central venous catheterization in our pa­tient was imperative because it permitted the intra- and postoperative monitoring of CVP, as well as the parenteral nutrition with high­ly caloric parenteral infusions and high-os­molality drugs. Blood samples for laboratory analysis could be taken from CV catheter as well. Although brachial veins were in considera­tion for catheterization, due to the expected long-term need for the CV catheterization as well as potential thrombosis of the brachial or femoral vein, we decided for a standard sub-clavial route. In this very high-risk patient for "blind" (anatomically guided) cannulation we performed the CV cannulation by using the ultrasound guidance. With the ultrasound-guided technique an anesthesiologist (opera­tor) can be oriented in anatomic relationships immediately before the cannulation.1-4 He is able to monitor the position of the needle, guide-wire and catheter in the central vein during the procedure as well. By the de­scribed technique, the whole catheter, its loops or eventual knots, as well as the tip, can be visualized and there is no need for the postoperative confirmation by radiography. Besides, a real-time ultrasound guidance tech­nique, which is cheap, quick and easy to per­form, improves the success rate, reduces the number of passes and gives us the possibility of the early detection of some later complica­tions in the CV cannulation (e.g. arterio-ve­nous fistulas or pseudo-aneurysms).2,5 This report highlights the potential bene­fits of ultrasound as a supporting method for the CV catheterization in oncologycal pa­tients. Summing the personal experience and the data from relevant literature, we can rec­ommend sonography as an important adjunct method in central venous cannulation in dif­ficult cases.1-4 References 1. Slama M, Novara A, Savafin A, Ossart M, Safar M, Fagon J.Y. Improvement of internal jugular vein cannulation using an ultrasound-guided tech­nique. Intensive Care Med 1997; 23: 916-9. 2. Hatfield A, Bodenham A. Portable ultrasound for difficult central venous access. Br J Anaesth 1999; 82: 822-6. 3. Skonick ML. The role of sonography in the place­ment and management of jugular and subclavian central venous catheters. AJR 1994; 163: 291-5. 4. Benter T, Teichgraber UK, Kluhs L, Papadopoulos S, Kohne CH, Felix R, et al. Anatomical variations in the internal jugular veins of cancer patients af­fecting central venous access. Anatomical varia­tion of the internal jugular vein. Ultraschall Med 2001; 22: 23-6. 5. áusti“ A, Stan‰i“ M, Eäkinja N, Matana A, Fu‰kar ë. Iatrogenic pseudo-aneurysm of the axillary ar­tery: the role of color Doppler sonography. J Clin Ultrasound 1996; 24: 323-5. Case report Bifocal primary intracranial germinoma in a child. Case report Aleä Koren Clinical Institute of Radiology, Clinical Center Ljubljana, Slovenia Background. Bifocal primary intracranial germinal tumors are rare. Only 5-10% of all germ cell tumors are found both in the suprasellar and pineal region. Case report. In presented patient we found two primary intracranial germinomas in pituitary and pineal gland that were successfully operated. Radiological properties of germinomas and differential diagnosis are discussed. Conclusions. Although the definite histological diagnosis cannot be achieved by computer tomography and/or magnetic resonance images alone, a detailed analysis of neuroradiological images is useful for pre­dicting the histological diagnosis. Key words: computed tomography; magnetic resonance; germinoma, intracranial Introduction Germ cell tumors located in the central nerv­ous system (CNS) represent less than 4% of the intracranial tumors and affect primarily children and young adults. These tumors fre­quently arise in the suprasellar and pineal re­gion and in the midline structures around the third ventricle. Germ cell tumors can be di­vided as germinoma and nongerminomatous tumors regarding the histology. Only 5-10% of all germ cell tumors are found both in the suprasellar and pineal region1 mostly as ger­minomas. Received 31 July 2001 Accepted 8 August 2001 Correspondence to: Aleä Koren, M.D., Clinical Institute of Radiology, Clinical Center Ljubljana, Zaloäka 7, 1000 Ljubljana, Slovenia; Phone +386 1 543 15 30; Fax: +386 1 433 1044. We present a case of isolated primary bifo­cal germinoma of CNS in a child. Case report An eleven-year-old boy with nocturnal dis-uria was admitted to our hospital because of vomitting and frontal and retro orbital headache, which was depended on the posi­tion of the head. There were no vertigo or vi­sual disturbances. We found signs of prema­ture puberty, bilateral papillary edema and raised serum concentrations of PRL. The computed tomographic (CT) scan showed homogenous, well-delineated suprasellar tumor with obstructive hydrocephalus. An additional pineal region tumor was found on magnetic resonance images (MRI) of the brain. Both tumors had similar neuroradio-logical characteristics (Figures 1-4). We didn't find similar lesions in testicles or elsewhere in the body. The patient had two operations. Surgeons resected first pituitary lesion. The macroscop­ically gray tumor had a peripheral solid vas­cular part and a central soft avascular one. The tumor compressed both optical nerves and the inferior part of optical chiasm and grew into infundibulum and through sellar diaphragm into sella turcica (Figure 5). Hypophysis was completely destroyed. The pineal tumor was resected by the second op­eration (Figure 6). This tumor grew from the third ventricular wall posteriorly in the pineal region to both basal veins and was macro­scopically similar to the previously reseceted suprasellar tumor (Figures 7,8). The histologi­cal examination of both specimens showed mature germinoma with the positive reaction to alkaline phosphatase. All other tumor markers were negative. The patient had upper gaze failure and di­abetes insipidus postoperatively. Cerebros­pinal liquor for malignant cells was negative. No tumor was found on postoperative MRI (Figures 9,10). The additional radiotherapy of the third ventricular region with 4000 cGy was performed. The boy was treated with a substitute hormonal therapy and antiepilep-tic prophylaxis. The child is in complete re­mission 15 months after the diagnosis. Discussion Intracranial germ cell tumors are a heteroge­neous group of lesions that occur in children and young adults. Within the classification of intracranial germ cell tumors, there are a va­riety of different tumor types which carry dif­ferent prognoses. The most recent World Health Organization classification of germ cell tumors is as noted in Table 1. The most frequent histological type is germinoma (65%) and it holds the best prognosis2 with over 90% 5 year survival rate.3-5 They are located usually in the pineal or the suprasellar re­gion.6 Lesions in infundibulum alone are also described.7 Rarely they appear in thalamus and basal ganglia or other intracranial loca- Table 1. WHO classification of intracranial germ cell tumors 5.0 Germ cell tumors 5.1 Germinomas 5.2 Embryonal carcinoma 5.3 Yolk sac tumor 5.4 Choriocarcinoma 5.5 Teratoma 5.5.1 Immature 5.5.2 Mature 5.5.3 Teratoma with malignant transformation 5.6 Mixed germ cell tions. Pineal germinomas have strong male predominance in contrast to suprasellar ger­minomas that are more frequent in females. Bifocal lesions in these regions are found in 5­10%.1 It is unclear whether they represent ac­tual spread of the tumor or the simultaneous development of tumor in two sites. Clinical presentations of germinoma are dependent on the localization of lesions. Tumors in pineal area can present with hy­drocephalus, visual symptoms, obtundation, pyramidal tract signs and ataxia. Suprasellar tumors often produce diabetes insipidus and a pituitary hormonal dysfunction. Germinomas are macroscopically solid, quite homogenous tumors with a possible soft or partly cystic central part. They can seed by cerebrospinal liquor to other part of brain or meningeal surface.8,9 Germinomas are composed of more than one cellular type in 10%,10 their unspecific histological tumor marker is placental alkaline phosphatase.11 The neuroimaging characteristics of germi­nomas and nongerminomatous germ cell tu­mors are similar enough to limit diagnostic certainty, and either tissue confirmation or measurement of specific tumor markers are needed for the diagnosis. In addition, germ cell tumors in the pineal region cannot be de­finitively separated on the basis of neu­roimaging characteristics from other tumors such as pineoblastomas, pineocytomas or gliomas. In the suprasellar region germino-mas may be difficult to separate from other lesions which infiltrate the surrounding brain mimicking gliomas and histiocytomas. Germinomas are radiologicaly well delin­eated, oval or lobular and expansive or partly infiltrative tumors.6 The proportion of water to tumor cells determines their radiological morphology. The tumor can be isodense to hyperdense on CT and isointense to hyperin-tense on T1WI and T2WI on MRI. The solid part of the tumor shows isointense signal to cortex and intense opacification after the ap­plication of gadolinium contrast media Gd CM3 (Figures 1-8). Germinomas are quite ho-mogenous and uncapsular with no calcina­tions and other inclusions or important cystic and/or hemorrhagic areas.12 The treatment for germ cell tumors has be­come somewhat divergent as recommenda­tions for the treatment differ between pure germinomas and other forms of germ cell tu­mors.13 The craniospinal radiation of germi­nomas together with local doses of 4000 cGy showed excellent results. Although a definite histological diagnosis cannot be achieved by CT and/or MRI alone, the detailed analysis of neuroradiological im-ages is useful for predicting the histological diagnosis. References 1. Jennings MT, Gelman R, Hochberg F. Intracranial germ-cell tumors: natural history and pathogene­sis. J Neurosurg 1985; 63(2): 155-67. 2. Felix I, Becker LE. Intracranial germ cell tumors in children: an immunohistochemical and electron microscopic study. Pediatr Neurosurg 1990-91; 16(3): 56-62. 3. Packer RJ, Sutton LN, Rosenstock JG, Rorke LB, Bilaniuk LT, Zimmerman R, et al. Pineal region tu­mors of childhood. Pediatrics 1984;74(1): 97-102. 4. Allen JC. Controversies in the management of in-tracranial germ-cell tumors. Neurol Clin 1991; 9: 441-52. 5. Ballesteros MD, Duran A, Arrazola J, Redondo MJ, Bordiu E, de Abajo S, et al. Primary intrasellar ger­minoma with synchronous pineal tumor. Neuroradiology 1997; 39(12): 860-2. 6. Osborn AG. Diagnostic neuroradiology. St. Louis: Mosby, 1994. 7. Hoffman HJ, Otsubo H, Hendrick EB, Humphreys RP, Drake JM, Becker LE, et al. Intracranial germ-cell tumors in children. J Neurosurg 1991; 74(4): 545-51. 8. Yasue M, Tanaka H, Nakajima M, Kamio M, Nakamura N, Numoto T, et al. Germ cell tumors of the basal ganglia and thalamus. Pediatr Neurosurg 1993; 19(3): 121-6. 9. Sumida M, Uozumi T, Kiya K, Mukada K, Arita K, Kurisu K, et al. MRI of intracranial germ cell tu­mours. Neuroradiology 1995; 37(1): 32-7. 10. Barkovich J. Intracranial, orbital and neck tumors of chilhood. In: Barkovich J, editor. Pediatric imag­ing. New York: Raven Press; 2000. p. 523-32. 11. Baumgartner JE, Edwards MS. Pineal tumors. Neurosurg Clin N Am 1992; 3(4): 853-62. 12. Kollias SS, Barkovich AJ, Edwards MS. Magnetic resonance analysis of suprasellar tumors of child­hood. Pediatr Neurosurg 1991-92; 17(6): 284-303. 13. Paulino AC, Wen BC, Mohideen MN. Contro­versies in the management of intracranial germi­nomas. Oncology 1999; 13(4): 513-21, discussion 521-2, review 528-3. Artifacts and non-osseous uptake in bone scintigraphy. Imaging reports of 20 cases Guido M. Weiner, Lars Jenicke, Vika M.ller, Karl H. Bohuslavizki Department of Nuclear Medicine, University Hospital Eppendorf, Hamburg, Germany Background. Numerous possible artifacts may render the evaluation of bone scans difficult. Case reports. This article provides a pictorial survey of both typical and extraordinary pitfalls in bone scintigraphy, which are caused by increased or reduced tracer accumulation of soft tissue or bone. Conclusions. One should be aware, that in individual patients, "artifacts" in bone scintigraphy lead to for­merly unknown diagnoses, and the diagnostic and therapeutic procedure may be influenced decisively. Key words: radionuclide imaging; bone; 99mTc-diphosphonates; artifacts; benign bone diseases; malignant bone diseases Introduction Bone scintigraphy is commonly used for the diagnosis and staging of both benign and ma­lignant bone diseases.1 After the intravenous injection and the initial distribution through the whole body by simple perfusion, the ra­diotracers used commonly, i.e. 99mTc-labelled diphosphonates such as HDP or MDP in a standard dose of 600-800 MBq, diffuse into the extracellular space. The initial local distri­bution is predominately influenced both by the blood flow and the vascularization of the Received 27 July 2001 Accepted 2 August 2001 Correspondence to: Karl H. Bohuslavizki, M.D., Ph.D., Department of Nuclear Medicine, University Hospital Eppendorf, Martinistr. 52, D-20246 Ham­burg, Germany. Phone: +49 40 42803-4047; Fax: +49 40 42803-9330; E-mail: bohu@uke.uni-hamburg.de perfused region. Thus, early acquisitions re­flect perfusion and blood pool images. Due to its affinity to calcium, diphosphonates are then bound in a simple physicochemical way to bone structures within some 3 hours. Then, the regional skeletal uptake is deter­mined both by the initial perfusion and the metabolism of the bone.2 Bone scans taken after a three to five hours interval after injec­tion usually provide a good contrast between soft tissue and skeletal structures, since the background activity of the soft tissue is as less as 2-10%.3 In order to ensure proper scan reading a thorough indication and a detailed medical history are mandatory prior to the injection. All relevant past illnesses and therapies must be included (drainages, biopsies, surgery). An inspection and an examination at least of the region under investigation should be per­formed routinely. In most cases these simple things help to avoid misinterpretations. If there still remains doubt on the interpreta­tion, additional spot images or SPECT-imag­ing may be required for clarification. Apart from physiological, age-related or typical pathological patterns, numerous caus­es of artificial tracer distribution patterns are known, which have to be taken into account when reading bone scans.4 The main causes for artifacts are contaminations of the skin or of the clothes, paravenous or subcutaneous injection sites.5 However, causes for artifacts in bone scans are numerous, and case reports have been published dealing with hyper-hidrosis,6 foreign body7 and constitution,8 etc. McAffee and Silberstein9 published a sur­vey of various causes of non-osseous uptake. Case reports The following bone scans are intended to give an overview of numerous artifacts, which have been collected over the last few years in a large university-based nuclear medicine de­partment. Figures 1-7 show artifacts, e.g. technical causes, reduced tracer uptake etc., Figures 8 and 9 display malignant soft tissue tumors, Figure 10 shows inflammation, Figures 11-17 display non-inflammatory arti­facts, e.g. lymphatic edema, adipositas, trau­ma, anatomical variants etc, and Figures 18­20 depict artifacts associated with the renal system. Unless otherwise noted, all images were acquired 3 hours after injection using a standard dual-headed whole-body gamma camera (Bodyscan, SIEMENS, Erlangen, Germany) equipped with low-energy high-resolution collimators. Figure 3. Bone scintigraphy following 3 hours after Figure 4. Blood pool images acquired 5 min after i.v. salivary glands scintigraphy. Note, uptake of free injection of 99mTc-HDP. Note a ringlike reduced tracer 99mTc-pertechnetate in the stomach. uptake in the anterior view in a patient with ascites. Figure 5. Cold spot in projection to the right proximal Figure 6. Cold spot on the right upper thoracic wall as femur in the anterior view as a result of attenuation a result of attenuation due to an implantated pace-due to a purse in the trouser pocket. maker. Figure 7. Symmetric cold spots on both femoral heads Figure 8. Massive increased tracer uptake in the soft in a patient with bilateral hip prosthesis. tissue of the lung and the liver caused by calcifying metastasis in a patient with breast cancer. Figure 9. Massive soft tissue accumulation in the left Figure 10. Focal tracer accumulation in the right upper breast in a patient with an inflammatory breast cancer thoracic wall caused by an infected tip of a Buelau-detected by bone scanning. drainage. Figure 18. Absent tracer accumulation in both kidneys in loco typico and tracer accumulation in a kidney graft­ed into the right pelvis following bilateral nephrectomy due to renal cancer. Note, tracer uptake in the upper right ribs and in the sternum caused by thoracotomy due to aortocoronary bypass-grafting. Also, note the loss of the left arm in an accident 40 years ago. Conclusions One should be aware, that in individual pa­tients, "artifacts" in bone scintigraphy lead to formerly unknown diagnoses, and the diag­nostic and therapeutic procedure may be in­fluenced decisively. References 1. M.ller V, Steinhagen J, de Wit M, Jenicke L, Bohuslavizki KH. Bone scintigraphy in clinical routine. Radiol Oncol 2001; 35: 21-30. 2. Sch.michen C. Physiologische Grundlagen der Knochenszintigraphie: Me§technik und quantita­tive Auswertung. Nuklearmediziner 1984; 7: 73-88. 3. Peller PJ, Ho VB, Kransdorf MJ. Extraosseous Tc­99m MDP uptake: a pathophysiologic approach. Radiographics 1993; 13: 715-34. 4. Kessler JR, Wells RG, Sty JR. Skeletal scintigra­phy. Radiographic artifacts. Clin Nucl Med 1992; 17: 511-2. 5. Amico S, Lucas P, Liehn JC, Valeyre J. Unusual site of extraosseous uptake of Tc-99m-HMDP due to subcutaneous heparin injections. Report of two cases. Eur J Nucl Med 1989; 15: 670-2. 6. Ajmani SK, Lerner SR, Pircher FJ. Bone scan arti­fact caused by hyperhidrosis: case report. J Nucl Med 1977; 18: 801-2. 7. Turton DB, Silverman ED. Tampon artifact in bone scintigraphy. Clin Nucl Med 1994; 19: 1103-4. 8. Embry RL, Delaplain CB. Scintigraphic pitfall in a patient with steatopygia. Clin Nucl Med 1992; 17: 824-6. 9. McAffee JG, Silberstein ED. Non-osseous uptake. In: Silberstein ED, McAffee JG, editors. Differential diagnosis in nuclear medicine. New York: McGraw-Hill; 1984. p. 300-18. Cell electropermeabilization to small molecules in vitro: control by pulse parameters Alenka Ma‰ek Lebar and Damijan Miklav‰i‰ University of Ljubljana, Faculty of Electrical Engineering, Ljubljana, Slovenia A systematic study concerning the role of the different electric field parameters (pulse number, duration and amplitude) on electropermeabilization of DC3F cells to small molecules (propidium iodide) and on cell via­bility is presented. Cell permeabilization and viability dependence on the pulse amplitude was determined by twenty different sets of electrical parameters. The number of pulses varied between 1 and 64 and pulse duration between 20 ms and 1 ms. The most important parameter was the pulse amplitude because it trig­gered the electropermeabilization process and the process of cell death. Either in the case of electroperme­abilization as well as in the case of cell viability experiments, the parameter U50 (the pulse amplitude lead­ing to permeabilization or to the death of 50% of cell population) was not changed if the set of electrical parameters consisted of more than 16 pulses. This was independent of the pulse duration. The efficiency of permeabilization was enhanced by using of longer pulses. Such a systematic study of the influence of dif­ferent electric field parameters on electropermeabilization and cell viability may serve as a base for opti­mization of the electropermeabilization conditions for different applications. Key words: electroporation; electromagnetic fields; cell survival; propidium iodide Introduction The phenomenon of cell membrane electro-permeabilization can be described as a dra­matic increase in the transmembrane perme­ability induced by an externally applied elec­tric field. This transient state has important Received 17 July 2001 Accepted 10 August 2001 Correspondence to: Damijan Miklav‰i‰, Faculty of Electrical Engineering, University of Ljubljana, Trěaäka 25, 1000 Ljubljana, Slovenia. Phone: +386 1 4768 456; Fax: +386 1 4264 658; E-mail: damijan@ svarun.fe.uni-lj.si practical applications like the fusion of cells 1 and the introduction of the biologically active substances like drugs 2 and genetic material 3 into cells. Electropermeabilization is nowa­days widely used to manipulate biological cells, organelles, cell aggregates and tissue. The clinical applications gain increasing im­portance, particularly in oncology.4,5 Electro-permeabilization can be achieved with differ­ent sets of pulse parameters, i.e. the strength of applied field (voltage of applied pulses), the number and the shape of pulses, their du­ration and repetition frequency. An identical set of electrical parameters is not necessarily efficient for all applications. While eight square-wave electric pulses of 100 ms deliv­ered at 1 Hz is the most common used set of electrical parameters in electrochemotherapy, the pulses must be much longer (eight square-wave electric pulses of 20 ms deliv­ered at 1 Hz or 2 Hz) for DNA electrotransfer in vivo.5 Likewise, it has been shown in vitro that only pulse duration equal to or longer than 1 ms was associated with the detection of macromolecules in pulsed and viable cells, while the permeabilization of cells to small molecules was already detected for the mi­crosecond range.6 Therefore it is important to know how the pulse parameters affect the electropermeabilization process. The role of pulse parameters in obtaining a higher efficiency of electropermeabilization in vitro was investigated in a number of stud­ies.6-9 The permeabilization as a function of the parameters of applied electric field was quantified in two ways. Either the fraction of electropermeabilized cells in suspension was measured using fluorescence optical mi­croscopy 10, or the permeability of each cell was integrated over the whole cell population by measuring radioactive incorporation 11 or ATP leakage.6 Both types of information were gathered in some studies using flow cytome­try.12 Independently of the method, it was shown that the electric field intensity is the crucial parameter for inducing membrane permeabilization. The permeabilization oc­curs only if the electric field intensity is high­er than a certain threshold value. This thresh­old value is a function of the pulse number and the pulse duration. It decreases by in­creasing either the pulse duration or the num­ber of pulses until it reaches a "real" threshold value below which no permeabilization oc­curs even if using longer pulses or higher number of them. The "real" threshold value was obtained using 10 pulses or more with the duration longer than 100 ms.13 The per­meability threshold depends on the molecu­lar size of the probe used for its measure­ment: the larger the test molecule, the higher the apparent threshold.14 It also depends on the cell line because of their differences in size 15 and membrane properties.16,17 In almost all studies, the effect of pulse du­ration was studied at a given field intensity and pulse number. Similarly, the effect of pulse number was studied at a given field in­tensity and pulse duration, and vice-versa the effect of field intensity was studied at a given pulse duration and pulse number.6-8 How­ever, the fraction of electropermeabilized cells as a function of pulse duration at a giv­en number of pulses is strongly dependent on selected field intensity (Fig. 1). Namely, a plateau is reached at shorter pulses if higher field intensity is used. In this paper, we present a systematic study concerning the role of different electric field parameters (field intensity, pulse num­ber, and duration) on electropermeabilization of DC3F cells to small molecules and on cell viability. DC3F cells have been chosen be­cause considerable information about the electropermeabilization of that strain is avail­ able.2,18,19 The fraction of electropermeabi­lized cells was quantified by the penetration of propidium iodide and the viability of the cells by their cloning efficiency. The cells were pulsed with twenty different sets of electrical parameters (Table 1). The number of pulses varied between 1 and 64, the pulse duration between 20 ms and 1 ms and the pulse amplitude from 40 to 600 V. Materials and methods Chemicals Eagle's minimal essential medium (EMEM), trypsin and propidium iodide (PI) were pur­chased from Sigma Chemical Co. (St. Louis, MO). Fetal calf serum (FCS) and L-glutamine were obtained from Gibco BRL (Galthers-burg, MD), penicillin, streptomycin, gentam­icin from Lek (Ljubljana, Slovenia), and Crystal violet from Kemika (Zagreb, Croatia). Figure 1. Fraction of electropermeabilized cells as a function of pulse duration. Cells were pulsed eight times at different pulse amplitudes. The error bars in the fraction of electropermeabilized cells represent standard devia­tions of the data. PI was dissolved in sterile H2O at a concen­tration of 100 mM. Cell culture DC3F cells, a line of spontaneously trans­formed Chinese hamster lung fibroblasts, were grown in monolayers in the culture medium consisting of EMEM supplemented with 10% heat-inactivated FCS, 10mM L-glut-amine, 100 units/ml penicillin, 100 mg/ml streptomycin, and 11 mg/ml gentamicin. The cells were incubated at 37°C in a humidified atmosphere with 5% CO2, and were routinely subcultured every 4 days. Cell exposure to electric field Cells from the exponential growth phase were trypsinized and centrifuged for 5 min at 4°C and 1500 rpm in the culture medium. They were then resuspended in the serum-free medium supplemented with 0.5 mM CaCl2 at a concentration of 2.2 x 107 cells/ml. 90 ml cell suspension was mixed with 10 ml PI for the determination of electropermeabiliza­tion, or with medium supplemented with 0.5 mM CaCl2 for determination of electropulsed cell viability. A 50 ml droplet of the cell sus­pension was placed between two flat, paral­lel, stainless steel electrodes (length = 6 mm, width = 6mm, interelectrode distance = 2 mm). The electrodes were connected to a voltage generator (Jouan GHT 1287 B, France) generating monophasic square-wave electric pulses with independently adjustable electric parameters (voltage, number of pulses and duration). The cells were pulsed at 1 Hz fre­quency. The pulse parameters were moni­tored by an oscilloscope (Hameg HM 205-3, Table 1. Sets of electrical parameters. Repeated puls­es were delivered at 1 Hz frequency Set of electric Number Pulse duration field parameters of pulses [ms] 1.20 1 20 1.100 1 100 1.500 1 500 1.1000 1 1000 4.20 4 20 4.100 4 100 4.500 4 500 4.1000 4 1000 8.20 8 20 8.100 8 100 8.500 8 500 8.1000 8 1000 16.20 16 20 16.100 16 100 16.500 16 500 16.1000 16 1000 64.20 64 20 64.100 64 100 64.500 64 500 64.1000 64 1000 Germany). All experiments were performed under sterile conditions in a laminar flow hood at room temperature. Determination of electropermeabilization Electropermeabilization of cells was quanti­fied by the penetration of impermeant dye PI. When the membrane is permeable, PI binds to nucleic acids and becomes highly fluorescent. Therefore, it is not necessary to wash the cells to eliminate nonincorporated PI as in case of other fluorescent dyes. A se­lected evaluation method avoids the negative consequences of pipetting and centrifuging the cells that have been already pulsed.20 The cells were pulsed and incubated 5 min at room temperature. Thereafter, 25 ml of cell suspension was resuspended in 1 ml of 0.01 M phosphate-buffered saline (PBS, pH 7.4) and kept at 4°C till being analyzed by flow cy­tometry (FACSort, Becton Dickinson, CA). The flow cytometer was used to measure the number of fluorescent and therefore perme­abilized cells. Excitation was set at the wave­length 488 nm and emission was detected at 640 nm. Fluorescence was recorded for 5000 particles. Only particles large enough to qual­ify as cells were taken into consideration. The number of stained cells was determined and normalized to the number of all cells to get the percentage of permeabilized cells. Determination of electropulsed cell viability Cell viability was determined by means of colony-forming assay. After the exposure to electric pulses, the cells were incubated for 5 min at room temperature. They were then di­luted in the culture medium and seeded in triplicate (300 cells per 60 mm diameter petri dish). After five days, the colonies were fixed with 96% ethanol, stained with Crystal violet and counted. The survival of the cells treated with electric pulses was calculated as the per­centage of the colonies obtained from the un­treated control cells. Statistical analysis All experiments were repeated at least three times on different days. For each experimen­tal point, mean and standard deviation were calculated. Using nonlinear regression, a two-parameter sigmoid curve was fitted to the data where U is the pulse amplitude, -is the per­centage of permeabilized or alive cells, and and b are the two parameters of the sig­ U50 moid curve. Parameter U50 is the pulse ampli­tude leading to permeabilization of 50% of cell population in the case of electroperme­abilization and the pulse amplitude leading to the death of 50% of cell population in the case Figure 2. Effect of pulse amplitude on electropermeabilization of cells and cell viability. DC3F cells were pulsed at 1 Hz one time (A - electropermeabilization, F - cell viability), four times (B - electropermeabilization, G - cell via­bility), eight times (C - electropermeabilization, H - cell viability), sixteen times (D - electropermeabilization, I - cell viability) and sixtyfour times (E - electropermeabilization, J - cell viability). Pulse duration was equal to (•) 20 ms, ( ) 100 ms, (•) 500 ms and (•) 1000 ms. The symbols denote the means and the error bars are the standard devia­tions. of cell viability experiments. Parameter b gov­erns the inclination of the sigmoid curve. Smaller absolute value of parameter b means that the process reaches its plateau at smaller interval of pulse amplitudes, while its larger value means that the process reaches its plateau at larger interval of pulse amplitudes. In the literature, parameter b was paralleled with the efficiency of permeabilization.13 Results In this study we focused on the effect of pulse amplitude, number of pulses and pulse dura­tion on the cell permeabilization and viabili­ty. Repeated pulses were delivered at 1 Hz frequency. The cell permeabilization and via­bility dependence on the pulse amplitude was determined using twenty different sets of electrical parameters (Table 1). Figure 2 shows the results of these measurements. The symbols denote the means and the error bars are standard deviations. A two-parame­ter sigmoid curve is fitted to the data of each set of electrical parameters. For easier com­parison of different sets of electrical parame­ters, the parameter U50 and b of all electrop­ermeabilization curves and curves presenting cell viability are collected in Table 2 and pre­sented in Figures 3 and 4. Electropermeabilization The fraction of electropermeabilized cells in population is under control of the field inten­sity, the pulse duration and the number of pulses. Electropermeabilization occurs only for pulse amplitudes higher than a certain threshold value. This value is lower if longer pulses are used, or if the number of pulses is higher. Permeabilization curves are complete­ly shifted to the lower pulse amplitudes (Fig. 2A-E). If the set of electrical parameters con­sisting of 16 and 64 pulses, 0.5 ms electroper­meabilization curve more or less coincides with 1 ms electropermeabilization curve (Fig. 2D, E). It means that, in the sets of electrical parameters that consist of 16 pulses or more, the usage of the pulses longer than 0.5 ms does not change the fraction of electroperme­abilized cells in population at certain pulse amplitude. Parameter U50 is not changed if the set of electrical parameters consists of more than 16 pulses (Fig. 3A). This is independent of the pulse duration. The relation between the pulse duration and the number of pulses at a given parameter U50 is linear on logarithmic scale, if less than 16 pulses are used. In the literature, parameter b was paral­leled with the efficiency of permeabiliza- Table 2. Parameters U50 and b at different sets of elec­trical parameters. Parameters are determined by sig­moid curves which are fitted to the experimental data of electropermeabilization and cell viability using non­linear regression. Set of Electropermea-Cell viability electric field bilization parameters |b| |b| U50 U50 [V] [V] [V] [V] 1.20 354 53 664 143 1.100 249 59 345 36 1.500 200 33 328 50 1.1000 156 29 265 28 4.20 273 58 392 54 4.100 206 43 309 34 4.500 141 43 210 18 4.1000 118 27 180 24 8.20 252 41 326 43 8.100 173 32 253 30 8.500 122 20 159 32 8.1000 99 13 148 18 16.20 211 35 290 24 16.100 140 23 191 23 16.500 108 17 193 17 16.1000 98 19 110 11 64.20 211 27 274 23 64.100 157 27 182 25 64.500 77 19 77 17 64.1000 57 25 80 6 tion.13 Parameter b of the sigmoid curve is smaller, if the duration of the pulses is longer (Fig. 3B). Therefore, the efficiency of perme­abilization is enhanced by the usage of longer pulses. The enhancement is less pronounced if the set of electrical parameters consists of higher number of pulses. Cell viability Also the fraction of viable cells in population is under control by the field intensity, the pulse duration and the number of pulses as well. Cell death occurs at the field intensities higher than a certain threshold value. This value is lower if the pulses are longer or their number is higher. In that case cell viability curves are completely shifted to the lower pulse amplitudes (Fig. 2 F-J). Likewise, in the case of electropermeabi­lization, the relation between the pulse dura­tion and the number of pulses at a given pa­rameter U50 is linear on logarithmic scale if less than 16 pulses are used. Parameter U50 is not changed if the set of electrical parameters consists of more than 16 pulses (Fig. 4A). This is evident for the pulses shorter than 500 ms. Absolute value of the parameter b of the sigmoid curve is smaller if the duration of the pulses is longer (Fig. 4B). The viability of cells is changed on smaller interval of pulse ampli­tudes if longer pulses are used. Discussion The application of electric field pulses to DC3F cells results in the permeabilization of their plasma membrane. Electrical parame­ters, i.e. pulse amplitude, pulse duration and the number of pulses, have an important role in electropermeabilization process as well as an effect on cell viability. The most important parameter is the pulse amplitude because it triggers the electropermeabilization process and the process of cell death. Both processes have their characteristic threshold values. Either pulse duration or the number of pulses can modulate these threshold values. In our study, we quantified the fraction of Figure 3. Values of parameter U50 and b in the field of pulse duration and number of pulses in the case of elec­tropermeabilization experiments. Each curve represents one value of the parameter U50 (A) or b (B). The value is written on the curve. Scales are logarithmic. Figure 4. Values of parameter U50 and b in the field of pulse duration and number of pulses in the case of cell vi­ability experiments. Each curve represents one value of the parameter U50 (A) or b (B). The value is written on the curve. Scales are logarithmic. electropermeabilized cells by the penetration of PI. Because PI also leaks into healthy cells over time 21, we noticed some fluorescence cells also in the untreated control cells. Therefore, it was difficult to determine the pulse amplitude, which triggers electroper­meabilization process, i.e. threshold value. That is why our conclusions were made by observing electropermeabilization curves, their parameters U50, i.e. the pulse amplitude leading to permeabilization of 50% of cell population in the case of electropermeabiliza­tion and the pulse amplitude leading to the death of 50% of cell population in the case of cell viability experiments, and parameters b, i.e. parameter which governs the inclination of the sigmoid curve. Our conclusions can be summarized as follows: (1) In the sets of elec­trical parameters consisting of 16 or more pulses, the usage of pulses longer than 0.5 ms does not change the fraction of electroperme­abilized cells in the population at a selected pulse amplitude. (2) Both in the electroper­meabilization as well as in the cell viability experiments, parameter U50 is not changed if the set of electrical parameters consists of more than 16 pulses, which is independent of the pulse duration. (3) The efficiency of per-meabilization is enhanced by the usage of longer pulses. (4) The fraction of electroper­mabilized cells and viability of cells vary at a smaller interval of pulse amplitudes if longer pulses are used. In spite of a variety of studies investigating the role of pulse parameters in the electroper­meabilization efficiency, the studies analyz­ing the control of the cell viability by pulse parameters are rare.9,18 This is probably due to the effects of pulsing media which can con­tain a variety of undesirable and even toxic substances. In our study, these undesirable substances are Ca2+ ions. We performed electropermeabilization experiments in EMEM medium, which is a culture medium of DC3F cells. Because of the compatibility of results the pulsation of the cells in case of the cell viability, experiments were made in EMEM like in electropermeabilization experi­ments. Although EMEM Ca2+ concentration (1.8 mM) is in the range of approximate harmless limits of the extracellular fluids con­centrations for short periods (0.5 - 2.0 mM) 22, it can be toxic due to the impairment of Ca2+ cellular transports during cell injury. During and some time after the cells are exposed to electrical pulses, Ca2+ easily diffuses through the transiently permeable membrane due to very low cytosolic Ca2+ free concentration (~0.1 mM). An increase in cytosolic Ca2+ con­centration can directly lead to cell lysis by causing disruption of the cytoskeleton, DNA fragmentation or extensive damage to other cell components.23 Therefore, the cell viabili­ty is affected at lower pulse amplitudes as in the experiments prepared in media without Ca2+. However, the effects of electrical pa­rameters have the same trends as if per­formed in Ca2+ free medium (i. e. SMEM) (da­ta not shown). Our results were obtained using DC3F cell line and small test molecules. In one of our previous studies, we showed that electroper­meabilization curves, obtained by PI for a giv­en set of electrical parameters, are compara­ble with the electropermeabilization curves obtained by using anticancer drug bleomycin as a marker of cell permeabilization.19 So, we can conclude that our observations are valid also for bleomycin, which is the drug of choice for electrochemotherapy.4,5 A systematic study of the influence of dif­ferent electric field parameters (field intensi­ty, pulse number, and duration) on electrop­ermeabilization and cell viability may serve as a base for the optimisation of the electroper­meabilization conditions for different appli­cations. References 1. Teissie J, Rols MP. Fusion of mammalian cells in culture is obtained by creating the contact be­tween cells after their electropermeabilization. Biochem Biophys Res Commun 1986; 140: 258-66. 2. Orlowski S, Belehradek J Jr, Paoletti C, Mir LM. Transient electropermeabilization of cells in cul­ture: increase of the cytotoxicity of anticancer drugs. Biochem Pharmacol 1988; 37: 4727-33. 3. Neumann E, Schaefer-Ridder M, Wang Y, Hofschneider PH. Gene transfer into mouse ly­oma cells by electroporation in high electric fields. EMBO J 1982; 1: 841-5. 4. Rudolf Z, átabuc B, ›emaěar M, Miklav‰i‰ D, Vodovnik L, Seräa G. Electrochemotherapy with bleomycin: the first clinical experience in malig­nant melanoma patients. Radiol Oncol 1995; 29: 229-35. 5. Mir LM. Therapeutic perspectives of in vivo cell electropermeabilization. Bioelectrochemistry 2000; 53: 1-10. 6. Rols MP, Teissie J. Electropermeabilization of mammalian cells to macromolecules: control by pulse duration. Biophys J 1998; 75: 1415-23. 7. Rols MP, Teissie J. Electropermeabilization of mammalian cells: Quantitative analysis of the phenomenon. Biophys J 1990; 58: 1089-98. 8. Wolf H, Rols MP, Boldt E, Neumann E, Teissie J. Control by pulse parameters of electric field-medi­ated gene transfer in mammalian cells. Biophys J 1994; 66: 524-31. 9. Gabriel B, Teissie J. Control by electrical parame­ters of short- and long-term cell death resulting from electropermeabilization of Chinese hamster ovary cells. Biochim Biophys Acta 1995; 1266: 171­8. 10. Mir LM, Banoun H, Paoletti C. Introduction of def­inite amounts of nonpermeant molecules into liv­ing cells after electropermeabilization: direct ac­cess to the cytosol. Exp Cell Res 1988; 175: 15-25. 11. Serpersu EH, Kinosita K, Tsong TY. Reversible and irreversible modification of erythrocyte mem­brane permeability by electric field. Biochim Biophys Acta 1985; 816: 332-48. 12. Bartoletti DC, Harrison GI, Weaver JC. The num­ber of molecules taken up by electroporated cells: quantitative determination. FEBS Lett 1989; 256: 4­10. 13. Rols MP, Teissie J. Modulation of electrically in­duced permeabilization and fusion of Chinese hamster ovary cells by osmotic pressure. Biochemistry 1990; 29: 4561-7. 14. Orlowski S, Mir L. Cell electropermeabilization: a new tool for biochemical and pharmacological studies. Biochim Biophys Acta 1993; 1154: 51-63. 15. ›emaěar M, Jarm T, Miklav‰i‰ D, Ma‰ek Lebar A, Ihan A, Kopitar NA et al. Effect of electric-field in­tensity on electropermeabilization and electrosen­sitivity of various tumor-cell lines in vitro. Electro-and Magnetobiology 1998; 17: 263-72. 16. Kanduäer M, Magister S, áentjurc M, Seräa G, ›e-maěar M, Ma‰ek Lebar A et al. Corelation between membrane fluidity and in vitro electropermeabi­lization threshold. 11th International symposium Spectroscopy in theory and practice, Book of ab­stracts, Bled 11-15 April, 1999; EPR-6. 17. Rols MP, Dahhou F, Mishra KP, Teissie J. Control of electric field induced cell membrane permeabi­lization by membrane order. Biochemistry 1990; 29: 2960-6. 18. Ma‰ek Lebar A, Kopitar NA, Ihan A, Seräa G, Miklav‰i‰ D. Significance of treatment energy in cell electropermeabilization. Electro- and Magneto-biology 1998; 17: 255-62. 19. Kotnik T, Ma‰ek Lebar A, Miklav‰i‰ D, Mir LM. Evaluation of cell membrane electropermeabiliza­ tion by means of a nonpermeant cytotoxic agent. Biotechniques 2000; 28: 921-26. 20. Rols MP, Teissie J. Flow cytometry quantification of electropermeabilization. In: Jaroszeski MJ, Heller R, editors. Flow cytometry protocols. Methods Mol Biol, 1998; 91: 141-8. 21. Weaver JL. Estimation of cell viability by flow cy­tometry. In: Jaroszeski MJ, Heller R, editors. Flow cytometry protocols. Methods Mol Biol 1998; 91: 77­84. 22. Guyton AC, Hall JE. Textbook of medical physiology. Philadelphia: W.B. Saunders Company; 1996. 23. Orrenius S, McConkey DJ, Bellomo G, Nicotera P. Role of Ca2+ in toxic cell kiling. TiPS 1989; 10: 281-5. Electroporator for in vitro cell permeabilization Marko Puc, Karel Flisar, Stanislav Reberäek and Damijan Miklav‰i‰ University of Ljubljana, Faculty of Electrical Engineering, Ljubljana, Slovenia The use of high voltage electric pulse technology, electroporation, in cell biology, biotechnology and medi­cine has attracted an enormous interest. Electroporation is a transient phenomenon that increases the per­meability of cell plasma membrane. In the state of high permeability, the plasma membrane allows small and large molecules to be introduced into the cytoplasm, although the cell plasma membrane represents a con­siderable barrier for them in its normal state. The effectiveness of electroporation depends on many param­eters that can be divided into the parameters of the electric field and the parameters that define the state of cells and their surrounding, i.e. temperature, osmotic pressure, etc. In this article, we present a prototype electroporator GT-1 for in vitro electropermeabilization that we have developed. Our electroporator offers a vast flexibility of parameters and can generate high and low voltage pulses, of which the latter ones are used for electrophoretic transfer of charged molecules through permeabilized cell plasma membrane. Key words: electroporation - instrumentation - methods; cell membrane permeabilization Introduction Viability of a cell depends on the integrity of its plasma membrane. The plasma membrane prevents the exchange of substances between intracellular and extracellular spaces. Exposing the cell to the electric field can in­crease the permeability of the cell plasma membrane. The increased permeability of the plasma membrane allows small and large molecules to be introduced into the cyto­plasm. This phenomenon is transient and is Received 6 July 2001 Accepted 3 August 2001 Correspondence to: dr. Damijan Miklav‰i‰, University of Ljubljana, Faculty of Electrical Engineering, Trěaäka 25, SI-1000 Ljubljana, Slovenia. Phone: +386 1 4768456; Fax: +386 1 4264658; E-mail: damijan@ svarun.fe.uni-lj.si termed electropermeabilization and often al­so referred to as electroporation. The electro-poration is widely used in different medical and biological applications i.e. electrochemo-therapy, transdermal drug delivery and gene transfection. The effectiveness of all these ap­plications depends on many parameters that can be divided into the parameters of the electric field and the parameters that define the state of cell and its surrounding, i.e. tem­perature, osmotic pressure, conductivity of cytoplasm and extracellular fluids etc. With respect to this, optimal parameters of electro-poration have to be used to achieve best effi­ciency of the method.1 For in vitro experiments, where most com­monly used electrodes are parallel plates with a 2 mm inner distance, the threshold voltages typically range from 120 V to 300 V1,2, with the pulse durations from several microsec­onds to several milliseconds.1,3 Beside this, it is often necessary to deliver more then one pulse to increase efficiency of permeabiliza­tion. In that case, pulses must be delivered in a certain period requiring repetition frequen­cies from 1 Hz to several hundred Hz. All these demands are fulfilled by special devices, often referred to as electroporators. Nowadays, there are a lot of commercially available electroporators that are designed for in vitro experiments. The problem of most of these electroporators is that the flexibility of the parameters of electric pulses is not suffi­cient, especially if we want to study the ef­fects of different pulse parameters on the cell permeabilization, survival or average uptake of different molecules. In this paper, we present the design of electroporator for in vitro cell plasma mem­brane permeabilization. The device is operat­ed by an internal computer that allows the user to choose the parameters of electric field. The computer drives the pulse genera­tor that is composed of a digital pulse genera­tor generating the signal, high voltage ampli­fier amplifying the signal, and current amplifier that provides the signal with suffi­cient energy to prevent the voltage amplitude from dropping during the pulse delivery. Beside this, the last version of the developed electroporator also includes a special unit that generates low voltage pulses usually used for the electrophoretic transfer of the charged molecules through the permeabilized cell plasma membrane. System design Figure 1 shows the basic system design of the electroporator. It consists of a computer, pulse generator, voltage amplifier, current amplifier and low voltage pulse generator. Besides this, the device comprises two high voltage power supplies and several low volt­age devices that are necessary for normal op­eration but are not drawn on the figure. The internal computer of the device is in the first place used for selecting pulse param­eters. Therefore, the computer includes a user interface composed of a display and a key­board. The second task of the computer is to control pulse generation after activation has Figure 1. Block diagram of in vitro electroporator GT-1. Table 1. Output parameters of the developed in vitro been triggered. All pulse parameters, except electroporator pulse amplitude, are then transferred to the Value pulse generator. This subunit generates digi-Parameter Min. Max. Increment tal signal that is then amplified in the voltage Delay/Pulse ratio* E PULSES 3 65535 1 Pulse amplitude 25V 500V linear Number of pulses HIGH VOLTAG 1 128 1 Pulse current 30A Pulse duration ES 5ms 9999ms 1ms Delay duration E PULS 0ms 9999ms 1ms Pulse amplitude 0V 50V 10V Number of pulses LOW VOLTAG 1 1000 1 Pulse current 1A Pulse duration 5ms 5000ms1ms&50ms amplifier to the value that we set by external potentiometer. The amplified signal is then intensified by the energy from the current amplifier because we have to fulfill the ener­ gy requirement as defined by the load be­ tween electrodes.4 At this point, the genera­ tion of the high voltage signal that is used for permeabilization of cell plasma membrane is finished. The low voltage pulse generator, which we have constructed just recently, is used for the electrophoretic transfer of charged molecules through permeabilized cell plasma mem­brane. The structure of this subunit, con­trolled by the internal computer, is similar to that of the DC power supply. This version al­lows to change the amplitude in 10 V steps in a range from 0 V to 50 V. Performance and experimental results The developed in vitro electroporator, which is still a prototype, has been used in our labo­ratory for more than two years. During that time, we found and repaired some deficien­cies in design and we also made several other improvements that reflect on greater flexibil­ity of the parameters. The current prototype GT-1 allows the user to set high and low volt­age pulses in a range that is given in Table 1. The pulse repetition frequency -is calculated by using the following equation: 1 -= , (1) T.(1+DR) where T is pulse duration and DR is value of the parameter Delay/Pulse ratio. Furthermore, to demonstrate the perform­ance of the device we designed an experiment where we measured voltage and current on the output (Fig. 2). In the experiment, we ex­ * - parameter defines pulse repetition frequency that is calculated by the equation (1). posed 50 ml of pure SMEM, which was placed between 2mm plate electrodes, to five con­secutive 100 ms pulses of 500 V at a repetition frequency of 2.5 kHz (DR = 3). Figure 2. Performance of electroporator loaded with 50 ml drop of SMEM that was placed between 2 mm plate electrodes. Electroporator was programmed to generate five consecutive 100 ms pulses of 500 V and repetition frequency of 2.5 kHz (DR = 3). Top trace (signal 1) presents voltage and bottom trace (signal 2) presents current flowing through the system. The measurements were performed using LeCroy LT9310C digital oscilloscope, a LeCroy AP015 current probe, and a Tektronix P5100 1:100 voltage probe. The voltage was 500 V, while current increased from 14 A to 19 A. It is evident from the Figure 2 that the in­strument was able to deliver all the pulses without any distortion. Even more, it could easily increase the current between two con­secutive pulses that is usually necessary due to the polarization of the sample between the electrodes. To compare the performance of the proto­type GT-1 with that of Jouan GHT 1287B, we evaluated cell permeabilization, using a bleomycin method5, with both devices. The results of experiments that are shown on Figure 3 are comparable. T-test showed no statistically significant difference between both experiments. Besides the performance experiments that we carried out, our colleagues already pub­lished the results of the experiments using the developed electroporator.6,7 The two ex­periments were performed to study the influ­ence of the parameters of electroporation (medium conductivity 6 and pulse repetition frequency 7) on the cell permeabilization, sur­vival and average uptake. Conclusions The comparison of the developed prototype GT-1 with the commercially available device Jouan showed that GT-1 fulfils all demands of the in vitro investigations. Furthermore, it offers a vast flexibility of the parameters and has the ability to generate high and low volt­age pulses, where low voltage pulses can be used for the electrophoretic transfer of charged molecules through permeabilized cell plasma membrane. Acknowledgment Our research was supported through various grants by the Ministry of Education, Science and Sports of the Republic of Slovenia and in part by IGEA s.r.l. Italy. Authors wish to thank to Maäa Kanduäar D.Sc., Tadej Kotnik D.Sc, and Gorazd Pucihar B.S. who per­formed many experiments with the devel­oped prototypes and provided us with a wealth of feedback information about opera­tion of devices. References 1. Mir LM. Therapeutic perspectives of in vivo cell electropermeabilization. Bioelectrochemistry 2000; 53: 1-10. 2. ›emaěar M, Jarm T, Miklav‰i‰ D, Ma‰ek-Lebar A, Ihan A, Kopitar NA, Seräa G. Effect of electric-field intensity on electropermabilization and elec­trosensitivity of various tumor-cell lines in vitro. Electro Magnetobiol 1998; 17: 261-70. 3. Tsong TY. Electroporation of cell membranes. Biophys J 1991; 60: 297-306. 4. Puc M, Reberäek S, Miklav‰i‰ D. Requirements for a clinical electrochemotherapy device - electropo­rator. Radiol Oncol 1997; 31: 368-73. 5. Kotnik T, Ma‰ek-Lebar A, Miklav‰i‰ D, Mir LM. Evaluation of cell membrane electropermeabiliza­tion by means of a nonpermeant cytotoxic agent. Biotechniques 2000; 28: 921-6. 6. Pucihar G, Mir LM, Miklav‰i‰ D. The effect of pulse repetition frequency on Lucifer Yellow up- take by means of cell electropermeabilization in 7. Pucihar G, Kotnik T, Kanduäer M, Miklav‰i‰ D. vitro. In Magjarevi“ R, Tonkovi“ S, Bilas V, The influence of medium conductivity on elec- Lackovi‰ I, editors. IFMBE Proceedings. Pula: tropermeabilization and survival of cells in vitro. Birotisak; 2001. p. 834-6. Bioelectrochemistry 2001; (accepted - in print). Bone metastases from malignant melanoma: a retrospective review and analysis of 28 cases Elias Brountzos1, Irene Panagiotou2, Dimitrios Bafaloukos2, Dimitrios Kelekis1 1Second Department of Radiology, Medical School, Athens University, Eugenidion Hospital, Athens, Greece, 2Second Department of Oncology, Metaxa Cancer Hospital, Piraeus, Greece Background. The aim of the study was to evaluate the clinical characteristics, the radiological findings, and the treatment effect on the patients with bone metastases from malignant melanoma. Patients and methods. Retrospective review of 293 stage IV melanoma patients during a 15-year period was made. Results. Twenty-eight patients (9.5%) with bone metastases were identified; all patients had a thick or in­termediate primary melanoma (Breslow 2.7-9.9). Most of the patients presented with multiple (95.6%), symptomatic (92.6%) skeletal lesions. Imaging depicted 90 bone lesions. Axial metastases were more com­mon (86%); 54% of them were located at the spine. Skeletal radionuclide scintigraphy was non-specific; ra­diographij and computed tomography was diagnostic. Typical bone metastases were osteolytic (92.5%). Sixty-six lesions were treated with radiotherapy; in 79%there was a palliative response. There was no cor­relation between total dose or fraction size and effective palliation. The skeletal lesions did not respond to concurrent chemotherapy and/or biphosphonates. Median response duration to treatment was estimated to 2.6 months and median survival to 4.7 months. Conclusions. Osseous metastases from malignant melanoma occur in the patients with more advanced pri­mary lesions. They are most frequently osteolytic and located in the axial skeleton. Radiographij and com­puted tomography is diagnostic. Radiotherapy still remains the treatment of choice. Key words: bone neoplasms; malignant melanoma, bone metastases; imaging; radiotherapy Received: 18 June 2001 Accepted 21 June 2001 Correspondence to: Elias Brountzos, MD, Second Department of Radiology, Medical School, Athens University, Eugenidion Hospital, 20 Papadiaman­topoulou st, Athens 11528, Greece; Phone: +301 7227488; Fax: +301 729 2280; E-mail: ebrountz@cc. uoa.gr Introduction In most clinical series bone metastases from malignant melanoma are less frequent, rang­ing from 11%-17%.1 Nevertheless, the autopsy series have revealed that skeletal involve­ment is more common (23-49%).2-4 Skeletal metastases generally occur in pa­tients with widespread metastatic disease and usually represent a late site of recur­rence.5 Median survival is estimated to 2-6 months.1-5 The aim of our study was to present our experience concerning this uncommon site of melanoma dissemination. We retrospectively evaluated the clinical course, the radiological abnormalities and the response to treatment in this subgroup of melanoma patients. Patients and methods We reviewed the records of all patients with stage IV malignant melanoma treated in the 2nd Department of Medical Oncology in Metaxa Cancer Hospital from 1985 to 2000. We recorded the patients with disseminated bone disease. The inclusion data were the fol­lowing: age, sex, location and thickness of primary tumor, time interval between pri­mary tumor and bone metastases, distribu­tion of metastases, location of the metastatic lesion, and presenting signs and symptoms. Table 1. Characteristics of our patients' population Age at AJCC stage IV diagnosis, 50, 26 - 86 years (Median, range) Male (%) 59 Female (%) 41 Site of primary melanoma (%) Lower extremity 14.1 Upper extremity 11.1 Trunk 33.3 Head & neck 37.8 Unknown origin 3.7 Breslow's thickness, mm 4.1, 2.7 - 9.9 (Median, range) Stage III diagnosed 70 prior to stage IV (%) Adjuvant treatment 22.2 before stage IV (%) Disease free interval before 37, 0 - 267 stage IV, months (median, range) Bone metastases as sole 3.7 & initial site (%) Multiple bone metastases (%) 95.6 X-ray, computed tomography, magnetic reso­nance and bone scintigraphy images, detect­ing all radiological abnormalities, were care­fully reviewed. The type and response to treatment were clinically and radiologically evaluated. Effective palliation was clinically defined as significant relief from symptoms for at least 1 month. Palliative responses to radiotherapy were correlated with total radia­tion dose and fraction size. Survival was de­fined as the number of months between the diagnosis of bone metastases and death or last follow-up. Statistical analysis was performed using the chi-squared test for the correlation be­tween primary melanoma's thickness and the development of bone metastases. In the com­parison, statistical significance was deter­mined using a p-value level of 0.001. Results From a total of 293 patients with stage IV ma­lignant melanoma, 28 patients (9.5%) with skeletal metastases were identified. The dis­ease dissemination was diagnosed in all 293 patients using computed tomography of the chest and/or abdomen: all patients were sub­mitted to computed tomography during regu­lar follow-up or were evaluated with comput­ed tomography when they became symptomatic. Table 1 shows the 28 patients' characteristics. All our patients' primary melanomas were thick or intermediate (Breslow's thickness ranging from 2.7-9.9). From the 293 stage IV patients, 165 had a thick or intermediate pri­mary melanoma. When correlated, statistical significance was found (p<0.001). Add­itionally, most patients (70%) had AJCC stage III disease before the diagnosis of bone metastases. Twenty-six patients presented with symp­toms of bone metastases at the time of the di­agnosis: bone pain (85.1%), pathologic frac-tures (14.8%) and/or bone marrow infiltration (7.4%) were the most common findings. Four patients (14.8%) presented with hypercal­caemia. There were no compression fractures or neurological symptoms. Only 2 patients (7.4%) were asymptomatic; in these patients, skeletal involvement was found during rou­tine abdominal computed tomography. In all 28 patients, thoracic and/or abdomi­nal computed tomography was initially per-Table 2. Skeletal distribution of the lesions Location Percentage of lesions (%) Spine 54 Pelvis 21 Ribs 11 Femur & tibia 7 Humeral, ulna & clavicle 4 Temporal, maxilla & mandible 3 formed, followed by skeletal radionuclide scintigraphy in the 26 symptomatic patients, and skeletal survey radiographs in 14. Computed tomography of the appendicular skeleton was performed in 9 symptomatic pa­tients, and MRI of the painful skeletal site in 5 patients. All studies yielded positive results. A total of 90 metastatic bone lesions were depicted. Table 2 shows the distribution of the le­sions: axial metastases were more common (86%) than appendicular bone metastases; the lesions were located more commonly at the spine, particularly at the thoracic and lumbar vertebrae. Table 3 demonstrates the radiological fea­tures found in radiographs and computed to­mography. More than 90% of the lesions were osteolytic. A mixed osteolytic-osteoblastic pattern was uncommon (Figure 1). We found Figure 1. An unusual mixed osteolytic-osteoblastic metastatic pattern located at the lumbar vertebrae. Table 3. Imaging features of the metastatic skeletal le­sions (x-ray, CT) Imaging Percentage features of lesions (%) Osteolytic 92.5 Cortical erosion & destruction 36.6 Soft tissue involvement 10.5 Mixed osteolytic-osteoblastic 8.2 pattern (Fig.1) Completely osteoblastic 2.3 Table 4. Palliation depending on total dose and frac­tion size Total dose No palliated lesions (cGy) / No lesions treated (%) < 3000 13/16 81.25 3000-4000 30/37 81.08 > 4000 9/13 69.23 Dose per No palliated lesions fraction (cGy) / No lesions treated (%) < 200 2/3 67.00 201-300 21/26 80.76 301-400 8/9 88.88 401-500 10/15 66.66 > 500 11/13 84.61 that 91.1% of the lesions had a medullary ori­gin. Scintigraphy uniformly depicted metas­tatic lesions as sites of increased uptake of Tc­99m. Magnetic resonance imaging was more specific in depicting the soft-tissue involve­ment. Whenever imaging follow-up was per­formed to evaluate the response to treatment, certain radiological response patterns were identified. Recalcification, sclerotic rim and periostal reaction were the most common findings. The 66 bone lesions -found in the 26 symp­tomatic patients - were treated with irradia­tion, and showed an effective palliative re­sponse rate of 79% (52/66). Table 4 demonstrates that the palliation of bone metastases was not related to total dose or fraction size. In 18 (of the 26) symptomatic patients, 31 symptomatic bone lesions were treated with radiotherapy and concurrent chemotherapy and/or biphosphonates, while 15 asympto­matic lesions were treated only with chemotherapy and/or biphosphonates. Systemic chemotherapy was also given to the two asymptomatic patients with the metasta­tic lesions at the lumbar vertebrae. In all 20 patients, the disease progressed in ex-traskeletal sites. As shown in imaging follow­up, 61% of the symptomatic lesions respond­ed to therapy; all of them were clinically palliated. All asymptomatic lesions treated with chemotherapy and/or biphosphonates did not respond radiologically or clinically, but progressed. Median response duration to treatment was estimated to 2.6 months and median overall survival was estimated to 4.7 months. Discussion In our study, only 9.5% of the stage IV melanoma patients had skeletal involvement. A similar low percentage of antemortem di­agnosis of bone metastases from malignant melanoma is reported in the literature.1-4 Since clinical diagnosis of skeletal involve­ment is infrequent, little has been published describing the pattern and natural history of melanoma metastatic to the bone. The limitation of our study is that we did not perform routine screening, so we do not know the exact incidence of skeletal metas­tases. Based on melanoma surveillance guide­lines,6,7 our patients were not routinely sub­mitted to bone radionuclide scintigraphy. Nevertheless, we evaluated our patients dur­ing the follow-up with chest and abdominal computed tomography, so we were able to de­tect the majority of skeletal metastases. Melanoma bone metastases occur more fre­quently in the axial skeleton, and are there­fore easily diagnosed by computed tomogra­phy.8,9 Nevertheless, the real incidence of osseous disease might be slightly higher than 9.5%, since appendicular bone metastases could not be depicted by thoracic and ab­dominal computed tomography. Only 3.7% of our patients had bone metas­tases as the first and only site of recurrence; the rest of the patients had widespread dis­ease in multiple metastatic sites. This is also in agreement with the literature.5 In our series, skeletal metastases were more frequent in patients with primary melanomas of the trunk and the head and neck area. We also found that all patients with bone metastases had a thick or intermediate pri­mary melanoma (p<0.001). Additionally, most patients had stage III disease prior to the development of skeletal metastases. Our findings may suggest that the patients with more advanced primary lesions are more like­ly to develop bone metastases, and should be more closely monitored. Only 7.4% of our patients were asympto­matic and the skeletal involvement was found during routine imaging follow-up. The metastatic disease was located at the lumbar vertebrae and was detected by an abdominal computed tomography. In our series, axial metastases (86%) were more common than appendicular bone metastases and 54% of them were located in the spine. Our findings are similar to those previously reported, and suggest that whenever a computed tomogra­phy is performed to evaluate metastatic melanoma, the axial skeleton should be care­fully examined.8,9 Bone radionuclide scintigraphy was non­specific; all metastatic lesions were depicted as sites of increased uptake of Tc-99m. Plain radiographs and computed tomography im­ages were diagnostic. In the literature, there are few reports of the imaging findings of skeletal melanoma metastases.9-11 In our se­ries, melanoma bone metastases were oste­olytic with medullary origin. Lesion growth caused cortical erosion and destruction, pathologic fractures and soft-tissue involve­ment. Atypical skeletal metastases exhibited a mixed osteolytic-osteoblastic pattern or, even more infrequently, a completely os­teoblastic pattern. Magnetic resonance imag­ing better depicted the extent of soft-tissue involvement. The response assessment of bone metas­tases to therapy is difficult; in most cases, de­cisions about the efficacy of treatment are based on symptomatic response or change in extraskeletal metastatic disease. In our study, radiographs and computed tomography were useful in evaluating the tumor's response to treatment. One could argue that since the real inci­dence of bone metastases in clinical series is low, the role of computed tomography or oth­er imaging studies is only complementary. Nevertheless, we showed that serial radi­ographs or computed tomography are essen­tial in establishing the diagnosis, guiding the treatment planning and assessing the tumor's response to treatment. In our series, radiotherapy offered an ef­fective palliation rate of 79%. We believe that it still represents the treatment of choice in this subgroup of patients; the treatment with chemotherapy and/or biphosphonates did not appear as effective. We agree with Rate and al. that concurrent chemotherapy has no influence on palliation.12 We found that in palliative treatment of bone lesions from melanoma, the application of high total dose or high fraction size was not advantageous at all. High doses should be avoided since they do not offer more effective palliation and can create greater complications. Similar results were reported by Konefal et al.13 in the analy­sis of dose fractionation in the palliation of bone and brain metastases from malignant melanoma. Despite the palliation offered with radio­therapy, our patients' prognosis was poor. Median survival was similar to the one previ­ously reported in the literature1,2,5 and esti-mated to 4.7 months. Even if patients with thick or intermediate primary cutaneous melanomas could be more closely monitored in order to detect asymptomatic bone metas­tases, no change in palliation or survival would be achieved. In conclusion, we recommend a skeletal evaluation with radiographs or computed to­mography - whenever symptoms develop ­and a careful examination of the axial skele­ton in the patients with advanced primary melanomas. The life expectancy of these pa­tients is short, but conventional fractionation radiotherapy can offer effective palliation to most of these patients. It is therefore worth­while to pursue the diagnosis. References 1. Lee YT. Malignant melanoma: patterns of metas­tasis. CA Cancer J Clin 1980; 30: 137-41. 2. Balch CM, Soong S-J, Murad TM. A multifactorial analysis of melanoma. IV. Prognostic factors in 200 melanoma patients with distant metastases (stage III). J Clin Oncol 1983; 1: 126-9. 3. Budman DR, Camacho E, Wittes RE. The current causes of death in patients with malignant melanoma. Eur J Cancer 1978; 14: 327-9. 4. Amer MH, Al-Sarraf M, Vaitkevicius VK. Clinical presentation, natural history and prognostic fac­tors in advanced melanoma. Surg Gynecol Obstet 1979; 149: 687-90. 5. Balch CM, Soong S-J, Shaw HM, Urist MM, Mc Carthy WH. An analysis of prognostic factors in 8500 patients with cutaneous melanoma. In: Balch CM, Houghton AN, Milton GW, Sober A, Soong S-J, editors. Cutaneous melanoma. Philadelphia: J-B Lippincot; 1992. p. 165-71. 6. Huang CL, Provost N, Marghoob AA, Kopf AW, Levin L, Bart RS. Laboratory tests and imaging studies in patients with cutaneous malignant melanoma. J Am Acad Dermatol 1998; 39: 451-63. 7. Gershenwald JE, Buzaid AC, Ross MI. Classification and staging of melanoma. Hematol/ Oncol Clin N Am 1998; 12: 737-65. 8. Pattern RM, Shuman WP, Teefey S. Metastases from malignant melanoma to the axial skeleton: a CT study of frequency and appearance. AJR 1990; 155: 109-12. 9. Potepan P, Spagnoli I, Danesini GM, Laffranchi A, Gadda D, Mascheroni L, et al. The radiodiagnosis of bone metastases from melanoma. Radiol Med (Torino) 1994; 87: 741-6. 10. Stewart WR, Gelberman RH, Harrelson JM, Seigler HF. Skeletal metastases of melanoma. J Bone Joint Surg Am 1978; 60: 645-9. 11. Fon GT, Wong WS, Gold RH, Kaiser LR. Skeletal metastases of melanoma: radiographic, scinti-graphic, and clinical revue. AJR 1981; 137: 103-8. 12. Rate WR, Solin LJ, Turrisi AT. Palliative radiother­apy for metastatic malignant melanoma: brain metastases, bone metastases, and spinal cord compression. Int J Radiat Oncol Biol Phys 1988; 15: 859-64. 13. Konefal JB, Emami B, Pilepich MV. Analysis of dose fractionation in the palliation of metastases from malignant melanoma. Cancer 1988; 61: 243-6. Ranking radiotherapy treatment plans: physical or biological objectives? Martin Ebert Department of Radiation Oncology, Sir Charles Gairdner Hospital, Western Australia, Australia Background. The ranking of treatment plans in radiotherapy is of importance when there are alternative approaches to treating an individual patient, in assessment of dose information collected during clinical tri­als and in formulation of objectives for optimization routines. Methods. Several physically and radiobiologically-based dose indices were calculated for a series of model dose-volume histograms (DVHs). The ranking of these DVHs according to each dose index was examined. Variation in the ranking of the radiobiological indices with parameters used in the models was also exam­ined. Ranking according to the indices was also examined for DVHs of planning target volumes (PTVs) for a series of 18 patients treated with external beam radiotherapy for prostate carcinoma. Results. It was found for both the model and real DVHs that treatment plan ranking depends explicitly on the model used for ranking target-volume doses (i.e., the dose index used). For the radiobiological models, there is a strong dependence of DVH ranking on the radiobiological parameters used in the models (specif­ically, the 'alpha' value from the linear-quadratic model). Conclusion. When ranking radiotherapy treatment plans during planning or in evaluation of clinical trials, attention should be paid to the models used in dose evaluation. Key words: radiotherapy planning, computer assisted; radiobiology Introduction Many situations arise in radiotherapy treat­ment planning where multiple treatment plans need to be compared in order to evalu- Received 13 July 2001 Accepted 10 August 2001 Correspondence to: Dr Martin A. Ebert, Director of Physics Research, Department of Radiation Oncology, Sir Charles Gairdner Hospital, Hospital Avenue, Nedlands, Western Australia, 6009, Australia, Tel: +61 8 9346 4900; Fax: +61 8 9346 3402; E-mail: Martin. Ebert@health.wa.gov.au ate the optimal plan, or in order to provide a direct comparison of the relative merits of each of the plans. Such situations arise dur­ ing: Ą The treatment planning process for an in­dividual patient. If two or more irradiation strategies are available, it will be necessary to decide which is the best strategy. Ą During evaluation of dosimetric data col­lected during clinical trials. In order to cor­relate treatment outcome with delivered dose, a method of describing dose distri­butions to target volumes must be used. Ą During inverse planning optimization pro­ cedures. Inverse planning requires specifi­ cation of an objective function which de­ scribes the optimality of a given dose distribution. Successive optimization itera­ tions require a comparison of treatment plans on the basis of that objective func­ tion.1-3 In order to perform the plan comparison in terms of the dose distribution delivered to the target volume (the PTV encompassing the tu­mor volume), indices need to be stated which reduce the complex distribution of dose/vol­ume throughout the PTV to a single scalar value. The dose distribution is frequently pre­sented in the form of a dose-volume his­togram (DVH), which can be easily reduced to a single index (a 'dose index') by computa­tional methods. Several alternatives exist for dose indices. In purely physical terms, the delivered radia­tion doses can be treated as quantities, which directly relate to treatment outcome. In this case, 'physical objectives' are used to describe the optimality of a treatment plan. An alter­native is to attempt to relate the physical dose distribution more directly to some actual in­dication of probable response. In this case, we are using a 'radiobiological objective', which will be based upon some hypothesized (possibly validated) model for cellular re­sponse. The usefulness of either physical or radio-biological dose indices depends very much on the correlation of those indices with treat­ment outcome. Such validation requires eval­uation of data from large-scale clinical trials. In relating dose indices to each other, it is im­portant to consider whether individual dose indices will rank alternative treatment plans differently, and whether that ranking will de­pend on the specifics of the models them­selves. This study aimed at examining those differences and dependencies. Methods Dose indices considered A series of physical dose indices were used. These were: Ą mean dose; Ą minimum dose; Ą maximum dose; Ą dose standard deviation; and Ą least-squares deviation from prescription dose. The radiobiolgically-based indices (DVH reduction values) considered were: Ą tumour control probability (TCP); 4,5 and Ą equivalent uniform dose (EUD). 6-8 Both of these models were based on the linear-quadratic approach to describing cell kill, ignoring time and fractionation effects and assuming independence of all tumor cells. Thus, for a DVH described by a distri­bution of N doses, di, at discrete volumes, vi, for a tumor with uniform cell density r, the equations used for TCP are: (1) with (2) Equation (1) provides population averag­ing by sampling TCP over a large range (K ­typically 104) described by a normal distribu­tion of alpha-values defined by a mean alpha value, a , and a standard deviation, a . For ms EUD, population sampling is not necessary (Ebert, 2000) and the equation for EUD is: (3) where N is the number of bins in the DVH. In all calculations, parameter values of a = 0.1 s Gy-1 and r = 108 cells/cm3 were used. Values of ain the range 0.05 Gy-1 to 0.8 Gy-1 were m considered. The physical and radiobiological indices listed above were calculated for a series of model and real DVHs in order to examine how those DVHs were ranked according to each index. Model DVHs A series of artificial DVHs were considered (Figure 1) which represented a large range of possible dose-volume conditions in a PTV. These distributions are: 1. A normal (Gaussian) distribution with a standard deviation of 5% of the prescrip­tion dose. 2. A normal distribution with a standard de­viation of 10% of the prescription dose. 3. A single-sided normal distribution with a standard deviation of 10% of the prescrip­tion dose. 4. Uniform dose delivery except for a hot spot of 150% over a volume of 5%. 5. Uniform dose delivery except for a cold spot of 50% over a volume of 5%. For all model DVHs a tumor volume of 100 cm3 and a mean dose of 60 Gy was used. DVHs for prostate treatments Figure 2 shows DVHs for PTV for 18 pa­tients treated with external beam radio­therapy for prostate carcinoma (follow-up in­formation pending). Results Model DVHs For the model DVHs, data has been sum-marised in Figure 3. For each dose-index, the value has been shown for each DVH. The ranking of the DVHs according to each index is also shown. Figure 4 shows the variation in TCP and EUD for each of the model DVHs with variation in the value of a. m Figure 1. Model DVHs used to represent a broad range of feasible dose distributions. Figure 2. DVHs for PTV for 18 patients treated with external beam radiotherapy for prostate carcinoma. Prescription dose was 66 Gy (100 % level) for all cases. Figure 3. Summarized results for physical dose-indices for the five model DVHs. a) Mean dose, b) Maximum/Minimum dose, c) Dose standard deviation, d) Sum of least-squares. The numerals show the order of DVH ranking according to each dose-index. Figure 4. Variation in values of a) TCP and b) EUD with alpha (Gy-1) value, showing some overlap in the order of DVH ranking. DVHs for prostate treatments cording to one of the indices, and all indices In order to visualize the ranking of the 18 plotted together. Thus in Figure 5a, the DVHs prostate-patient DVHs according to the phys-have been ordered according to their mean ical dose-indices, the DVHs were ordered ac-dose. The figure then shows how the DVHs Figure 5. Variation in physical dose-indices across all 18 patients DVHs. a) Data sorted by increasing mean dose, b) data sorted by increasing minimum dose. EUD values calculated using a = 0.35 Gy-1. m compared according to the other dose-indices. Figure 5 b shows the same information with DVHs ranked according to minimum dose. Figure 6 shows variation TCP and EUD val­ues for all 18 DVHs as they vary with mean alpha value in the respective radiobiological models. In Figure 7, the ranking of the 18 DVHs ac­cording to TCP or EUD has been shown using an intensity scale at each value of a. m Figure 6. Variation in values of a) TCP and b) EUD with alpha (Gy-1) value. Figure 7. Variation in ranking of the 18 patient DVHs according to a) TCP and b) EUD, and according to the alpha (Gy-1) value used in the TCP/EUD models. Patients have been ranked according to TCP/EUD at a = 0.4 Gy-1. The intensity of the image at each alpha value indicates the ranking of each of the 18 DVHs from lowest TCP/EUD (black) to highest TCP/EUD (white). Discussion The results presented above show that if dose-indices are going to be used to rank rival treatment plans, then the resulting ranking is going to depend explicitly on the particular dose-index used. The model DVHs were con­sidered on the basis of the significant differ­ences between them and, as such, it is not surprising that the physical dose-indices give different DVH rankings as shown in Figure 3. What is more surprising is the subtle change in ranking according to EUD as the alpha-pa­rameter was varied (Figure 4b), and the more dramatic change in ranking with TCP as the alpha-parameter was varied (Figure 4a). The change in ranking is also not consistent be­tween TCP and EUD. In Figure 4a the 'under-dose' DVH is seen to jump ranking order quite rapidly with change in alpha-value, whereas in Figure 4b it is the 'single-sided Gaussian' distribution which changes rank­ing most quickly. The strong dependence of TCP on alpha for the underdose DVH is not unexpected as TCP has been shown to be very sensitive to the presence of regions of low dose.8 For the data taken from patient PTV dose distributions, there are relatively smaller dif­ferences between the 18 DVHs. As a result, smaller but more frequent changes in ranking would be expected. In Figure 5 it is seen that, when the 18 DVHs are ordered according to one of the physical dose-indices, there is con­siderable variation in the order of the other dose-indices. In Figure 5b, some correlation is seen between minimum and maximum dose and dose standard deviation as may be ex­pected. In terms of DVH ranking according to the radiobiological dose-indices, Figure 6 shows that there is considerable overlap both for EUD and TCP. This overlap is reflected in Figure 7 as the ranking of individual DVHs (indicated by the intensity of the plot at each combination of patient number and alpha value) changes rapidly with alpha-value indi­cating a strong sensitivity not only to the ra­diobiological models, but this parameter of the radiobiological models. Close examination of Figure 6a a shows some 'noise' in the TCP vs alpha-value curves. This is due to the statistical sampling methods used to incorporate population sampling in the TCP model. Using large val­ues of K in equation (1) leads to significantly long calculation times for TCP and only min­imal smoothing of these curves (due to strong effects of low alpha values on TCP). The result is that there will be some overlap of DVH rankings as a result of the sampling routines used and this will lead to some of the rapid variations in ranking displayed in Figure 7. Conclusions This study has shown that for a variety of DVH conditions, the ranking of DVHs is de­pendent on the model used for both physical and radiobiological dose-indices. In addition, the ranking of the DVHs also depends on the particular characteristics of the model being used (in this case, the alpha-value in TCP and EUD models based on the linear-quadratic equation). Variations in the ranking result from non-linear transformations between the indices. This must be considered whenever scalar indices are being used to present dosi-metric information in treatment planning, plan optimization or in analysis of dosimetric data from clinical trials. The usefulness of the variety of available indices for describing non-uniform dose dis­tributions will depend on the correlation of each index with treatment outcome. This in­formation will only become available follow­ing detailed assessment of data from large-scale clinical trials. References 1. Brahme A. Treatment optimization using physical and biological objective functions. In Smith A, ed­itor. Radiation therapy physics. Springer-Verlag; 1994. 2. Holmes T, Mackie TR. A comparison of three in­verse planning algorithms. Phys Med Biol 1994; 39: 91-106. 3. Ebert MA. Optimisation in radiotherapy I: Defining the problem. Aust Phys Eng Sci Med 1997; 20: 164-76. 4. Webb S, Nahum AE. A model for calculating tu­mour control probability in radiotherapy includ­ing the effects of inhomogeneous distributions of dose and clonogenic cell density. Phys Med Biol 1993; 38: 653-66. 5. Ebert MA, Hoban PW. Some characteristics of tu­mour control probability for heterogeneous tu­mours. Phys Med Biol 1996; 41: 2125-33. 6. Niemierko A. Reporting and analyzing dose distri­butions: A concept of equivalent uniform dose. Med Phys 1997; 24: 103-10. 7. Niemierko A. Response to "Comment on 'Reporting and analyzing dose distributions: A concept of equivalent uniform dose'". Med Phys 1997; 24: 1325-7. 8. Ebert MA. Viability of the EUD an TCP concepts as reliable dose-indicators. Phys Med Biol 2000; 45: 441-57. Slovenian abstmcl Radio/ Oncol 2001; 35(3): 161-5. Zadrževanje diha pri bolnikih, ki so radiološko preiskovani: primerjava casov po izdihu in po vdihu z ali brez hiperventilacije Groell R, Schaffler GJ, Schloffer S Izhodišca. Pri slikovnih preiskavah kot so racunalniška tomografija, magnetna resonanca ali ul­trazvok je potrebno, da bolnik zadrži dihanje. Namen pricujoce šudije je bil primerjati case zadrževanja diha po vdihu in izdihu in ocenti ucinek hiperventilacije. Bolniki in metode. V študiji smo obravnavali 30 bolnikov in 19 zdravih prostovoljcev, vsi so pod­pisali pismeno privolenje. Cas zadrževaja diha je bil merjen po izdihu in izdihu, pred in po hiper­ventilaciji. Rezultati. Srednji cas zadržanega diha po izdihu (bolniki 24±9 sekunde, prostovoljci 27±7 sekunde) je bil znacilno krajši kot po vdihu (bolniki 41±20 sekunde, p<0.001; prostovoljci 62±18 sekunde, p<0.001). Razlika se je znacilno povecala s hiperventilacijo (40-60%, p'.S:0.005) po izdi­hu in vdihu, tako pri bolnikih kot pri prostovoljcih. Zakljucki. Ceprav je zadrževanje diha po izdihu priporoclivo pri razlicnih slikovnih preiskavah, zlasti še pri preiskavah trupa in trebuha, pa zadrževanje dihanja po vdihu omogoca, da bolniki zadržujejo zrak znatno dlje. Radio/ Onco/ 2001; 35(3): 167-73. Ultrazvocno vodena perkutana drenaža trebušne tekocine namesto laparotomije ali relaparotomije Miletic D, Uravic M, Fuckar ž, Glavaš R, Topljak-Polic D Izhodišca. Namen študije je bil ugotoviti ucinkovitost in zanesljivost perkutane trebušne drenaže pri kirurških bolnikih in ovrednotiti medreberno drenažo pri kopicenju tekocine pod trebušno prepono. Material in metode. 87 bolnikov, starih od 29 do 84 let (srednja starost 55,5 let) smo perkutano drenirali pod kontrolo ultrazvoka zaradi kopicenja tekocine v trebuhu po operaciji ali pri še neoperiranih bolnikih, kjer bi bila sicer potrebna laparotomija. Za odstranitev tekocine smo uporabljali katetre števila 8 do 14, ki smo jih uvedli med rebra ali pod njimi z lateralne ali spred­nje strani trebuha. Rezultati. Z medrebernim pristopom smo drenirali 31 (60,8%) od 51 kopicenj tekocine pod tre­bušno prepono. Trajanje drenaže ni bilo odvisno od vrste pristopa, ali smo drenirali med rebri ali pod njimi. Znacilno pa se je podalšalo trajanje drenaže (p<0.05, Mann-Whitney U test) pri gnojnih tekocinah (srednja vrednost 18 dni; razpon 7-73 dni) v primerjavi s hematomi, bilomi in drugimi negnojnimi kopicenji tekocine (srednja vrednost 11 in 6 dni). Ultrazvocno vodena perkutana drenaža je bila uspešna pri 92% bolnikov, manjših zapletov pa je bilo le 9,2%. Uspešnost posega je bila vecja pri drenaži kopicene tekocine pod diafragmo (96%). Zakljucki. Ultrazvocno vodena perkutana drenaža je metoda izbora pri odstranjevanju trebušne tekocine, kjer bi bila sicer potrebna laparotomija. Ce je mesto vboda vsaj dva medreberna pros­tora nižje od oboka prepone in kateter ni uveden skozi pleuralno tekocino, je medreberna drenaža enako ucinkovita in enako varna kot drenaža pod rebri. Radio/ 011co/ 2001; 35(3): 225-30. S/ove11ia11 abstract Radio/ Oncol 2001; 35(3): 175-7. Ultrazvocno vodeno uvajanje centralnega venskega katetra pri bolniku, ki je imel obojestransko radikalno odstranjene vratne bezgavke: prikaz primera Šustic A, Cerovic· R, Juretic M Izhodišca. Obsežne in težavne operacije v onkologiji veckrat zahtevajo uvajanje centarlnega venskega (CV) katetra. Pri nekaterih bolnikih je zaradi anatomskih sprememb, ki jih je povzroci­la bolezen, ovirano ali celo onemogoceno obicajno "slepo" uvajanje CV katetra. V takšnih primerih je uporaba utrazvoka uspešna dodatna metoda pri uvajanju CV katetra. Prikaz primera. Predstavljamo bolnika, ki je imel obojestransko radikalno odstranjene vratne bezgavke in pri katerem je posledicno prišlo do pomembnih anatomskih sprememb ter smo mu ultrazvocno uspešno uvedli CV kateter. Zakljucki. Ultrazvok je lahko koristna dodatna metoda pri uvajanju CV katetra pri onkoloških bolnikih. V primerih, kjer je uvajanje otežkoceno, priporocamo uporabo ultrazvoka. Radio/ 011col 2001; 35(3): 179-83. Bifokalni primarni intrakranialni germinom pri otroku. Prikaz primera KorenA Izhodišca. Bifokalni primarni intrakranialni germinalni tumorji so redki, tako samo 5-10% vseh germinalnih tumorjev odkrijemo supraselarno in v podrocju cešarike. Prikaz primera. Predstavljamo primer bolnika z dvema primarnima intrakranialnima germi­nomoma v podrocju hipofize in cešarike, ki sta bila operativno uspešno odstranjena. Opišemo tudi nekatere radiološke znacilnosti germinomov in nekatere diferencialne diagnosticne možnosti. Zakljucki. Ceprav koncne histološke diagnoze intrakranialnih germinomov ni mogoce napove­dati samo z racunalniško tomografijo ali/in magnetno resonanco, lahko k dokoncni diagnozi ve­liko pripomore natancna analiza nevroradioloških preiskav. Radio/ Oncol 2001; 35(3): 185-91. Artefakti in kopicenje radiof armaka izven kosti pri scintigrafiji okostja. Slikovni prikaz 20 primerov Weiner GM, Jenicke L, Miiller V, Bohuslavizki KH Izhodišca. Številni možni artefakti lahko znatno otežijo oceno scintigrafije okostja. Prikazi primerov. V clanku smo naredili slikovni pregled obicajnih in nenavadnih primerov, ki lahko zavedejo ocenjevalce scintigramov okostja. Zmotne interpretacije nastanejo ob poveca­nem ali zmanjšanem kopicenju radiofarmaka v mehkih tkivih ali v kosteh. Zakljucki. Posebno pozorni moramo biti na stanja, kjer bi morebitni artefakti privedli do nove (napacne) diagnoze in vplivali na nadaljnje (napacno) obravnavanje bolnika. Radio/ 011co/ 2001; 35(3): 193-202. Elektropermeabilizacija celic z majhnimi molekulami in vitro: vpliv elektricnih parametrov Macek Lebar A, Miklavcic D V clanku predstavljamo sistematicno študijo vloge razlicnih elektricnih parametrov (število, tra­janje in amplituda elektricnih pulzov) v procesu elektropermeabilizacije celic DC3F za majhne molekule in vpliva teh parametrov na preživetje celic. Permeabilizacijo celic in njihovo preživet­je smo dolocili za dvajset razlicnih naborov elektricnih pulzov. Število elektricnih pulzov smo spreminjali od 1 do 64, njihovo trajanje pa med 20 µs in 1 ms. Najpomembnejši elektricni pa­rameter je amplituda elektricnih pulzov, saj sproži proces elektropermeabilizacije in proces umi­ranja celic. Tako v primeru elektropermeabilizacije celic kot tudi ob študiju celicnega preživetja smo ugotovili, da se parameter U(tj. amplituda elektricnih pulzov pri kateri je permeabi­ 50 liziranih (ali mrtvih) 50 % celic v populaciji) ne spremeni, ce uporabljeni vlak elektricnih pulzov sestavlja vec kot 16 elektricnih pulzov. Ugotovitev je neodvisna od njihovega trajanja. Permeabilizacija je ucinkovitejša, ce uporabimo daljše elektricne pulze. Tako sistematicno študi­jo vpliva elektricnih parametrov na elektropermeabilizacijo in preživetje celic lahko uporabimo kot osnovo za optimizacijo elektropermeabilizacijskih pogojev pri razlicnih aplikacijah elek­tropermeabilizacije. Radio/ Oncol 2001; 35(3): 225-30. S/ovC11ian abstrac/ Radio/ Onco/ 2001; 35(3): 203-7. Elektroporator za permeabilizacijo celicnih membran in vitro Puc M, Flisar K, Reberšek S in Miklavcic D Elektroporacija je pojav, pri katerem s kratkotrajnim visokonapetostnim elektricnim impulzom v membrani biološke celice povzrocimo strukturne spremembe. Pore, ki nastanejo v plazmalemi, povecajo prepustnost, zato lahko snovi neposredno vstopajo v celico. Ucinkovitost elektroporaci­je je odvisna od številnih parametrov, ki jih razdelimo na elektricne parametre in parametre, ki so povezani s stanjem celic in njihovo okolico npr. osmotski tlak, temperatura itd. V tem delu pred­stavljamo prototip elektroporatorja GT-1 za in vitro elektroporacijo. Napravo smo zasnovali tako, da omogoca spreminjanje elektricnih parametrov na širokem podrocju. Poleg visokonapetostnih impulzov pa naprava proizvaja tudi nizkonapetostne impulze, ki se jih navadno uporablja za elek­troforeticni vnos nabitih molekul preko elektricno permeabilizirane plazmalerne. Radio/ Onco/ 2001; 35(3): 209-14. Zasevki malignega melanoma v kosteh. Retrospektivni pregled in analiza 28 primerov Brountzos E, Panagiotou I, Bafaloukos D, Kelekis D Izhodišca. Študijo srno izvedli z namenom, da bi ocenili klinicne znacilnosti, radiološke izvide in ucinke zdravljenja bolnikov z zasevki malignega melanoma v kosteh. Bolniki in metode. V 15-letnern obdobju smo retrospektivno pregledali 293 bolnikov z melanomom stadij IV. Rezultati. Med pregledanimi primeri srno zasledili 28 bolnikov (9,5%) z zasevki na kosteh. Pri vecini bolnikov je bil primarni melanom z veliko ali srednje veliko globinsko invazijo (Breslow 2.7-9.9), kostni zasevki so bili vecinoma multipli (95,6%) in sirnptomatski (92,6). S slikovno di­agnostiko srno odkrili 90 zasevkov v kosteh. Bolj pogosti so bili zasevki v aksialnem skeletu (86%), kar 54% smo jih odkrili na hrbtenici. Radionuklidna scintigrafija skeleta ni bila dovolj znacilna, da bi lahko z njo diagnosticirali zasevke melanoma v kosteh, tako srno naredili tudi rentgensko slikanje in racunalniško tomografijo kosti. Najbolj tipicni zasevki v kosteh so bili os­teoliticni (92,5%). Z radioterapijo je bilo zdravljenih 66 zasevkov; paliativen ucinek srno dosegli pri 79%. Med velikostjo celokupne doze ali doze na frakcijo obsevanja ter paliativnirn ucinkom nismo našli korelacije. Ce srno bolnike ob obsevanju zdravili tudi s kemoterapijo in/ali z bifos­fonati, nismo dobili vecjega ucinka zdravljenja. Srednji odgovor na zdravljenje je bil 2,6 mese­ca, srednje preživetje pa 4,7 meseca. Zakljucki. Zasevki malignega melanoma v kosteh se najpogosteje pojavljajo pri bolnikih z lokalno napredovalimi primarnimi tumorji. Najveckrat so zasevki osteoliticni in se najraje zase­jejo v aksialni skelet. Diagnozo potrdimo z rentgensko preiskavo in racunalniško tomografijo, najbolj priporocljivo zdravljenje pa je radioterapija. Radio/ Oncol 2001; 35(3): 215-24. Vrednotenje nacrtov za obsevalno zdravljenje: fizikalni ali biološki pristop? Ebert M Izhodišca. Vrednotenje obsevalnih nacrtov (planov) je v radioterapiji posebno pomembno, kadar so možni razlicni nacini zdravljenja posameznega bolnika, ko ocenjujemo podatke o dozah, ki jih prejemajo bolniki v klinicnih študijah in pri dolocanju optimalnih nacinov obicajnega zdravl­jenja. Metode. Naredili smo modele dozno volumskih histogramov (DVH) in pri tem upoštevali vec fizikalnih in radiobioloških pokazateljev. DVH smo vrednotili glede na vsak posamezen dozni indeks. Pri vrednotenju smo spremljali tudi spreminjanje radiobioloških pokazateljev v odvis­nosti od parametrov, ki smo jih v modelih uporabljali. Nadalje smo ocenjevali DVH pri nacrto­vanih tarcnih volumnih 18 bolnikov s karcinomom prostate, ki smo jih zdravili s teleradioter­apijo. Rezultati. Videli smo, da je vrednotenje obsevalnih nacrtov neposredno odvisno od modelov, ki smo jih uporabili pri ocenjevanju doz v tarcnih volumnih oziroma od uporabljenih doznih in­deksov. Pri vrednotenju DVH-ov v radiobioloških modelih opazimo mocno odvisnost od radio­bioloških parametrov, ki smo jih v modelih uporabili (posebno to velja za vrednosti "alfa" iz lin­earno kvadratnega modela). Zakljucki. Ko vrednotimo nacrte za obsevalno zdravljenje, bodisi med nacrtovanjem obsevanja ali pa ob ocenjevanju klinicnih študij, moramo biti posebno pozorni na modele, ki smo jih upora­bili za izracun obsevalne doze. Radio/ Onco/ 2001; 35(3): 225-30. Notices Notices submitted for publication should contain a mailing address, phone and/ or fax number and/or e-mail oj a Contact person or department. Gastro-intestinal malignancies October, 2001 The ESO training course will be offered in Cairo, Egypt. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Depart­ment of Medica! Oncology, University Hospital of Ioannina, 45110 Ioannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Pediatric oncology Octobc1; 2001 The ESO training course will be offered in Trabzon, Turkey. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Department of Medica! Oncology, University Hospital of Ioannina, 45110 Ioannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Chest and head & neck Octobcr 4-6, 2001 The "Second International Chicago Symposium Malihnancies of the Chest and Head & Neck" will take place in Chicago, USA. Contact Center for Continuing Medica[ Education, 61st The University of Chicago, 950 East Street, Chicago, Illinois, USA; or fax +1 773 702 1736; or e­mail marlene@delphi.bsd.unchicago.edu; or see http://www.uchicago.edu/bsd/ C111e/ Breast cancer October 4-6, 2001 The ESO training course will be offered in Sarajevo, Bosnia and Herzegovina. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Department of Medica! Oncology, University Hospital of Ioannina, 45110 Ioannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Head and neck cancer October 4-6, 2001 The "4th European workshop on Basic Biology of Head and Neck Cancer: Recent Advances in Molecular Biology" will take place in Amsterdam, the Netherlands. Contact Marion van Zuilen; e-mail kno@nki.nl Lung cancer Octobcr 5-7, 2001. The "International Conference on Non Small Celi Lung Cancer: Standards and New Trends in Diagnosis and Therapy" will be offered in Bialystok, Poland. Contact Jacek Niklinski, MD, PhD, Dept. of Thoracic Surgery, Bialystok Medica! University, M. Sklodowska-Curie Str. 15-276, Bialystoc, Poland, or call/fax +348 85 742 35 37; or e-mail niklinsj@cksr. ac.bialystok.pl Radiotherapy Octobcr 7-11, 2001 The ESTRO teaching course "Evidence-Based Radiation Oncology: Principles & Methods" will take place in Cairo, Egypt. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www.estro.be Radiotherapy October 7-1.1, 2001 The ESTRO teaching course "Basic Clinical Radiobiology" will take place in Tenerife, Spain. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Rena! carcinoma October 9-11, 2001 The ESO training course "Rena! Carcinoma" will be offered in Moscow, Russia. Contact M. Vukelic, CSC Ltd., Heligenstadter Strasse 395b, 1190 Vienna, Austria; or call +43 1 369 0444; or fax +43 1 369 0444 20 Breast cancer October 12-14, 2001 "7th The Annual Puerto Rico Breast Cancer Conference" will take place in San Juan, Puerto Rico. Contact with e-mail genteinc@prtc.net Malignant lymphoma October 19-20, 2001 The ESO training course will be offered in Nicosia, Cyprus. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica] School, Depart­ment of Medica] Oncology, University Hospital of Ioannina, 45110 loannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Radiation therapy October 21-25, 2001 The "20th Annual ESTRO Meeting / ECCO 11 Meeting" will take place in Lisbon, Portugal. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Lymphoma October 26-28, 2001 The ESO training course "Non Hodgkin's Lymphoma, Patho-Biology, Classification and Clinical Relevance" will be offered in Cairo, Egypt. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Depart­ ment of Medica! Oncology, University Hospital of Ioannina, 45110 loannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Clinical trials November, 2001 The ESO training course will be offered in loannina, Greece. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Depart­ment of Medica! Oncology, University Hospital of Ioannina, 45110 loannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Psycho-oncology November, 2001 The ESO training course will be offered in Nicosia, Cyprus. Contact ESO office for Balkans and Middle East, N. Pavlidis, E. Andreopoulou Medica! School, Depart­ment of Medica! Oncology, University Hospital of Ioannina, 45110 Ioannina, Greece; or call +30 651 99394 or +30 953 91083; or fax +30 651 97505 Radiation therapy November 4-7, 2001 ASTRO Annual meeting will be held in San Francisco, California, USA. Contact American Society for Therapeutic Radiology and Oncology Office, 1891 Preston White Drive, Reston, VA 20191, USA; or see http://www.as­tro.org Neck and thyroid surgery November 5-7, 2001 The mas ter course will be offered in Milan, ltaly. Call P. Lonati +39 (0)257 489 490; or fax +39 (0)257 489 589 491; or e-mail head&neck@ieo.it Cancer risk November 12-13, 2001 The ESO conference "Reducing Cancer Risk. Focus on the four big killers" will take place in New York, USA. Contact ESO Office, Viale Beatrice d'Este 37, 20122 Milan, ltaly; or call +39 0258317850; or fax +39 0258321266; or e-mail esomi@tin.it Radio/ O11co/ 2001; 35(3): 231-5. Notices 233 Radiation oncology November 28-30, 2001 The ISRO teaching course "Radiation Oncology in the New Millenniurn: What have we learnt frorn evi­dence based medicine?" will take place in Mumbai, India. Contact with e-rnail info@>isro.be; or see http://www.isro.be Medical physics November 30 -December 1, 2001 The "17th Symposiurn of the Belgian Physicists Association (BHPA)" will take place in Brussels, Belgium. See http://www.md.ucl.ac.be/rbnt/bhpa/2001 Radiation oncology November 30 -December 1, 2001 "Advanced Seminar on Stereotactic Radiosurgery and Radiotherapy " will take place in London, U.K„ Contact with e-rnail anna.dowe!Q>rmh.nthames. nhs.k; or see http://www. roylmarsden.org/training/ courses Lung cancer December 6-8, 2001 The International Forum for Lung Cancer will be of­ fered in Athens, Greece. Contact Congress Secretariat -Organising Bureau, "MOEL" Ltd, 36, Eleon str. -GR 14564, Nea Kifissia, Greece; or call +301 6203 614; or fax +301 8078 342; or e-mail liagramo@internet.gr Chemotherapy December 12-14, 2001. The "5th International Symposium on Clinical Febrile Neutropenia" will be offered in Brussels, Belgium. Contact Mrs. Martine Hazard and call +32 2 541 3201; or fax +32 2 541 3202; or e-mail martine.haz­ ard@lbordet.be Lung cancer Marc/z 7-9, 2002. The "2nd World Conference on Clinical Cooperative Research for Lung Cancer" will be of­ fered in Brussels, Belgium. Contact European Lung Cancer Working Party, c/o Prof. J.-P. Sculier, Institute Jules Bordet, 1, rue Heger­Bordet, B-1000 Brussels, or call +32 25 39 04 96; or fax +32 25 34 37 56; or e-mail 101473.1044@compuserve. com; or see http://www.elcwp.org Radiotherapy March 10-14, 2002 The ESTRO teaching course "Radiotherapy Treatment Planning: Principles and Practice" will take place in Dublin, Ireland. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-rnail info@>estro.be; or see http://www. estro.be Lung cancer Marc// 14-15, 2002 The IASLC international workshop "Early lnvasive Lung Cancer. New Diagnostic Tools & Treatment Strategies will be offered in Turin, ltaly. Contact Organising Secretariat, CCI Centro Congressi Internazionale srl, Via Cervino 60, 10155 Turin, ltaly; or call +39 011 244 69 16; or fax +39 011 244 69 00; or e-mail info@>congressiefiere.com Brachyherapy Marc/z 24-26, 2002 The ESTRO teaching course "Endovascular Brachytherapy" will take place in Wien, Austria. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info«Destro.be; or see http://www. estro.be Thoracic surgery April 11-12, 2002. The "5th International Meeting on General Thoracic Surgery" will be offered in Barcelona, Spain. Contact RCT, C/ Aulestia i Pijoan, 12 Baixos 98012, Barcelona, Spain, or call +34 93 415 69 38; or fax +34 415 69 04; or e-mail rct«Drct-congresos.com Radiotherapy May 9-11, 2002 The Annual Brachytherapy Meeting GEC/ESTRO will take place in Antalya, Turkey. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 234 Notices 7795494; or e-mail info1!lestro.be; or see http://www. estro.be Radiation therapy Mm; 15-19, 2002 'fhe 7th lnternational Meeting on Progress in Radio­Oncology lCRO/OGRO 7 will take place in Salzburg, Austria. Contact Prof. D.H. Kogelnik, Salzburg, Austria; call +43 662 44823900; or fax +43 662 4482887; or e-mail d.kogelnik@llkasbg.gv.at Radiotherapy Ju11e 2-6, 2002 The ESTRO teaching course "IMRT and Other Conformal Techniques in Practice" will take place in Amsterdam, the Netherlands. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Bronchology and bronchoesophagology J1me 16-19, 2002. Thc "12th World Congress for Bronchology" and the "12th World Congress for Bronchoesophagology" will be offered in Boston, USA. Contact Congress Secretariat. Tufts University School of Medicine. Office of Continuing Education, l 36 Harrison Avenue, Boston, MA 02111, USA, or call +1 617 636 6509; or fax +1 617 636 0472; or see http://www.aabronchology.org Brachytherapy June 16-20, 2002 The ESTRO teaching course "Modem Brachytherapy Techniques" will take place in Lisboa, Portugal. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Oncology June 30 -Ju/1; 5, 2002. The "18th• UICC International Cancer Congress" will be offered in Oslo, Norway. Contact Norwegian Cancer Society, P.O. Box 5327 Majorstua, N-0304 Oslo, Norway, or call +47 22 59 30 00; or fax +47 22 60 69 80; or e-mail cancer@ oslo2002.org Radiation physics August 25-29, 2002 The ESTRO teaching course "Physics for Clinical Radiotherapy" will take place in Leuven, Belgium. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Prostate cancer September 1-3, 2002 The ESTRO teaching course "Brachytherapy for Prostate Cancer" will take place in Utrecht, the N etherlands. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. estro.be Lung cancer Seple111ber 1-4, 2002 The "8th Central European Lung Cancer Conference" will be offered in Vienna, Austria. Contact Conference Secretariat, Mondial Congress, Faulmanngasse 4, A-1040 Vienna, Austria; or call +43 1 588 04 O; or fax +43 1 586 91 85; or e-mail con­gress@)mondial.at Medica! physics September 9-13, 2002 The "10th International Congress on Boron Neutron Capture Therapy" will take place in Essen, Gerrnany. Contact Dr. Ray Moss with e-mail moss@ljrc.nl Radiation therapy September 17-21, 2002 The 21st Annual ESTRO Meeting will take place in Prague, Czech Republic. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@lestro.be; or see http://www. estro.be Radio/ Oncol 2001; 35(3): 231-5. Notices 235 Radiation therapy October 6-9, 2002 ASTRO Annual meeting will be held in New Orleans, Louisiana, USA. Contact American Society for Therapeutic Radiology and Oncology Office, 1891 Preston White Drive, Reston, VA 20191, USA; or see http://www.astro.org Radiation oncology Marc/z 15-19, 2003. The "2nd International Conference on Translation Research and Pre-Clinical Strategies in Radiation Oncolq,>y, ICTR 2003" will be offered in Lugano, Switzerland. See http://www.osg.ch/ictr2003.html Lungcancer Augusl 10-14, 2003. The "lOth World Conference of the Internationac Association for the Study of Lung Cancer" will be of­fered in Vancouver, Canada. Contact 10th World Conference of Lung Cancer, c/o lnternational Conference Services, 604-850 West Hastings, Vancouver BC Canada V6C 1E1, or call +1 604 681 2153; or fax +1 604 681 1049; or e-mail con­ference@2003worldlungcancer.org Radiation therapy September 21-25, 2003 The ESTRO 22 / ECCO 12 Meeting will take place in Copenhagen, Denmark. Contact FECS office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info@estro.be; or see http://www. fecs.be Radiation therapy October 19-23, 2003 ASTRO Annual meeting will be held in Salt Lake City, Utah, USA. Contact American Society for Therapeutic Radiology and Oncology Office, 1891 Preston White Drive, Reston, VA 20191, USA; or see http://www. astro.org Radiation therapy September 12-16, 2004 The 23rd Annual ESTRO Meeting will be held. Contact ESTRO office, Av. E. Mounier, 83/4, B-1200 Brussels, Belgium; or call +32 7759340; or fax +32 2 7795494; or e-mail info«1>estro.be; or see http://www. estro.be Radiation therapy October 3-7, 2004 ASTRO Annual rneeting will be held in Atlanta, USA. Contact Arnerican Society for Therapeutic Radiology and Oncology Office, 1891 Preston White Drive, Reston, VA 20191, USA; or see http:// www.as­tro.org As a service to our readers, notices of meetings or courses will be inserted free of charge. Please sent information to the Editorial office, Radiology and Oncology, Zaloška 2, SI-1000 Ljubljana, Slovenia. FONDACIJA "DOCENT DR. J. CHOLEWA" JE NEPROFITNO, NEINSTITUCIONALNO IN NESTRANKARSKO ZDRUŽENJE POSAMEZNIKOV, USTANOV IN ORGANIZACIJ, KI ŽELIJO MATERIALNO SPODBUJATI IN POGLABLJATI RAZISKOVALNO DEJAVNOST V ONKOLOGIJI. MESESNELOVA 9 1 000 LJUBLJANA TEL 0 1 51 9 1 2 77 FAKS 0 1 251 81 1 3 ŽR: 501 00-620-1 33-05-1 0331 1 5-2 1 4 779 Activity of "Dr. J. Cholewa" Foundation for Cancer Research and Education -A Report f or the Srecond Quarter of 2001 The activity of the members of the "Dr. J Cholewa" Foundation for cancer research and ed­ucation took a respite in the summer months of the year 2001, although a debate on sev­era! important issues continued without interruption. Among the topics discussed the most important were certainly the problems associated with changes in some of the donors' attitude towards the Foundation and the ways how to adapt to the new circum­stances. The Foundation plans to inaugurate some new forms of activity in the coming autumn of 2001 and later. Various oncology themes are to be presented in a newly formed type of sci­entific meeting with a day of lectures on the Medica! Faculty in Ljubljana and dedicated to the memory of Dr. J. Cholewa. Lectures should are planned to be given by the recipients of the "Dr. J. Cholewa" Foundation awards and grants in order to spread their newly acquired knowledge in oncology to interested individuals from Slovenia and abroad. The Foundation continues to support the regular publication of "Radiology and Oncology" international scientific journal that is edited, published and printed in Ljubljana, Slovenia. The support for the publication of the "Challenge Newsletter" is stili to be re-evaluated and the decision should be taken shortly. In the recent months the Foundation also gave financial support to the Second Central European Cancer Conference in Opatija, Croatia; International Congress on Medica! Ethics in Bled, Slovenia; International Congress on Genetics in Bled, Slovenia; EASL International Symposium in Portorož, Slovenia; and to the First Slovenian-Croatian-Austrian Gastroenterology Symposium in Portorož, Slovenia, dedicated to the problems associated with pancreatic cancer. It should also be noted that a number of grants was awarded to ex­perts from various parts of Slovenia in order to attend various oncological conferences and meetings around the world. The Foundation moved back to its original headquarters in Mesesnelova Street No.: 9 in Ljubljana in the beginning of the year 2001. Despite these changes the Foundation will nev­ertheless continue to pursue its stated objectives. Borut Štabuc, MD, PhD Andrej Plesnicar, MD Tomaž Benulic, MD PACS offers big improvements in produccivity and patienc care, buc only when it meets your needs exaccly. To achieve that, choose a partner with IT expertise and a thorough understanding of radiology, workRows and processes. Ar Philips Medica! Systems we have thar knowledge in depth -based on 100 years' experience. So when you're considering PACS, call Philips. 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Nosecnostji:l le, ce je korist zdravljenja za mater vecja od tveg§lri]a :z:c1 s flukonazolom ne dojijo, Stranski ucinki: PovezalJLs napenjanje, bolecine v trebuhu, driska,. zelorec:lkei se: anafilaksija in angioedem v tem primeru tak-:: .:;, Podrobnejše informacije so na voljo pri proizvajalfLi, Sanolabor let zastopa naslednja podjetja 1 - Kottermann (Nemcija): INTEGRA BIOSCIENCES (Svica): laboratorijsko pohištvo, laboratorijska oprema za mikrobiologijo, varnostne omare za kisline, biologijo celic, molek lamo biologijo luge, topila, pline in strupe, i. biotehnologijo ventilacijska tehnika in digestorji CO,NING (ZDA): DAKO (Danska): specialna laboratorijska plastika testi za aplikacijo v imunohistokerrniji, za aplikacijo v imu1 ologiji, mikro­patologiji, mikrobiologiji, virologiji, biologiji-virologiji, ipd., ehanske eno­mono-in poliklonalna protitelesa in veckanalne pi9ete in nastavki SVANOVA Biotech (Švedska): EVL (!'lizozemska): Elisa testi za diagnostiko v veterini diagnosticni tes.i za uporabo v veterinarski medicini NOVODIRECT BIOBLOCK (F.ancija): kompletna oprema in pripomoc i k za delo v laboratoriju HURNER (Nemcija): ventilrcijska tehnika GFL (Nemcija): CSL -Biosciences: laboratorijski aparati, omare in diagnosticni t.sti za uporabo skrinje za globoko zamrzovanje v veterirarski medicini ANGELANTONI SCIENTIFICA (Italija): BIOMfRICA (ZDA): hladilna tehnika in aparati za laboratorije, hitri testi !la diagnostiko, transfuzijo, patologijo in sodno medicino I EIA /R!A testi EHRET (Nemcija): CHARLES I.CHI (Svica): laminar flow tehnika, inkubatorji, specialna oprema za tes iranje izdelkov sušilniki, suhi sterilizatorji in oprema v farmacevtski industriji;aparati za za laboratorijsko vzrejo živali -kletke procesno kontrolo in ko trolo kvalitete ROSYS -ANTHOS (Avstrija): fotometri, avtomatski pralni sistem za mikrotitrine plošce LABORMED d.o.o. LABORMED, razstavni salon Zg. Pirnice 96/c Bežigrajski dvor SI -1215 Medvode Periceva 29, Ljubljana Tel.: (0)13621414 Tel.: (0)1 436 49 01 Fax: (0)136214 15 in f o@ 1 a bor med . si Fax: (0)l 436 49 05 www. labormed . s SIEMENS Rešitve po meri Mammomat 3000 modular Mammomat 3000 modular i l. univerzalni sistem za vse vrste mamografije , '• optimizacija doze in kompresije z OPDOSE in OPCOMP sistema !• modularna zgradba zagotavlja posodabljanje sistema '.)•servis v Sloveniji z zagotovljenimi rezervnimi deli in garancijo \i • izobraževanje za uporabnike SIEMENS d.o.o. Dunajska 22 1511 Ljubljana Telefon 01 / 474 61 00 Telefaks 01 / 474 61 35 no i.. . lram81 .. _ :-Tramadol analgetik • MEDITRADE¦ d.o.o. Vodovodna 100 1000 Ljubljana, Slovenija Tel.: 01 5894 600 Fax: 01 5684 340 www.meditrade.si Zastopamo: Radiološki program firme Kodak Medicinsko opremo firme Marconi CI', MRI, IM Negatoskope firme Ella Roloskopi firme Planilux Mamografska oprema firme Metaltronica PE: Stritarjeva 5, 4000 Kranj, Slovenija tel.: (0)4/ 2015 050, fax: (0)4/ 2015 055 e-mail: kemomed@siol.net KEMOMED Promega SKRINJE IN HLADILNIKI BIOHfT Radiology a11d Oncology Instructions for authors Editorial policy of the journal Radiology and Oncology is to publish original scientific pa­pers, professional papers, review articles, case reports and varia (editorials, reviews, short communications, professional information, book reviews, letters, etc.) pertinent to diag­nostic and interventional radiology, computer­ized tomography, magnetic resonance, ultra­sound, nuclear medicine, radiotherapy, clinical and experimental oncology, radiobiol­ogy, radiophysics and radiation protection. The Editorial Board requires that the paper has not been published or submitted for publica­tion elsewhere: the authors are responsible for all statements in their papers. Accepted arti­cles become the property of the journal and therefore cannot be published elsewhere with­out written permission from the editorial board. Papers concerning the work on hu­mans, must comply with the principles of the declaration of Helsinki (1964). The approval of the ethical committee must then be stated on the manuscript. Papers with questionable jus­tification will be rejected. Manuscript writtert in English should be submitted to the Editorial Office in triplicate (the original and two copies), including the il­lustrations: Radiology and Oncology, Institute of Oncology, Zaloška 2, SI-1000 Ljubljana, Slovenia; (Phone: +386 1 432 00 68, Tel./Fax: +386 1 433 74 10, E-mail: gsersa@onko-i.si). Authors are also asked to submit their manu­scripts on a 3.5" 1.44 Mb formatted diskette. The type of computer and word-processing package should be specified (W ord for Windows is preferred). All articles are subjected to editorial review and review by independent referee selected by the editorial board. Manuscripts which do not comply with the technical requirements stated herein will be returned to the authors for cor­rection before peer-review. Rejected manu­scripts are generally returned to authors, how­ever, the journal cannot be held responsible for their Ioss. The editorial board reserves the right to ask authors to make appropriate changes in the contents as well as grammatical and stylistic corrections when necessary. The expenses of additional editorial work and re­quests for reprints will be charged to the au­thors. General instructions• Radiology and Onco­logy will consider manuscripts prepared accor­ding to the Vancouver Agreement (N Engl J Med 1991; 324: 424-8, BMJ 1991; 302: 6772; JA­MA 1997; 277: 927-34.). Type the manuscript double spaced on one side with a 4 cm margin at the top and Ieft hand side of the sheet. Write the paper in grammatically and stylisti­cally correct language. Avoid abbreviations unless previously explained. The technical da­ta should conform to the SI system. The man­uscript, including the references may not ex­ceed 15 typewritten pages, and the number of figures and tables is limited to 4. If appropri­ate, organize the text so that it includes: Introduction, Material and methods, Results and Discussion. Exceptionally, the results and discussion can be combined in a single sec­tion. Start each section on a new page, and number each page consecutively with Arabic numerals. Title page should include a concise and in­formative title, followed by the full name(s) of the author(s); the institutional affiliation of each author; the name and address of the cor­responding author (including telephone, fax and e-mail), and an abbreviated title. This should be followed by the abstract page, sum­marising in less than 200 words the reasons for the study, experimental approach, the major findings (with specific data if possible), and the principal conclusions, and providing 3-6 key words for indexing purposes. Structured ab­stracts are preferred. If possible, the authors are requested to submit also slovenian version of the title and abstract. The text of the report should then proceed as follows: Jntroduction should state the purpose of the article and summarize the rationale for the study or observation, citing only the essential references and stating the aim of the study. Material and methods should provide enough information to enable experiments to be re­peated. New methods should be described in detail. Reports on human and animal subjects should include a statement that ethical ap­proval of the study was obtained. Results should be presented clearly and concisely without repeating the data in the ta­bles and figures. Emphasis should be on clear and predse presentation of results and their significance in relation to the aim of the inves­tigation. Discussion should explain the results rather than simply repeating them and interpret their significance and draw conclusions. It should review the results of the study in the light of previously published work. Illustrations and tables must be numbered and referred to in the text, with appropriate location indicated in the text margin. Illu­strations must be labelled on the back with the author's name, figure number and orien­tation, and should be accompanied by a de­scriptive legend on a separate page. Line drawings should be supplied in a form suit­able for high-quality reproduction. Photo­graphs should be glossy prints of high quality with as much contrast as the subject allows. They should be cropped as close as possible to the area of interest. In photographs mask the identities of the patients. Tables should be typed double spaced, with descriptive title and, if appropriate, units of numerical meas­urements included in column heading. References must be numbered in the order in which they appear in the text and their cor­responding numbers quoted in the text. Authors are responsible for the accuracy of their references. References to the Abstracts and Letters to the Editor must be identified as such. Citation of papers in preparation, or sub­mitted for publication, unpublished observa­tions, and personal communications should not be included in the reference list. If essen­tial, such material may be incorporated in the appropriate place in the text. References fol­low the style of Index Medicus. Ali authors should be listed when their number does not exceed six; when there are seven or more au­thors, the first six listed are followed by "et al.". The following are some examples of refer­ences from articles, books and book chapters: Dent RAG, Cole P. In vitro maturation of monocytes in squamous carcinoma of the lung. Br J Cancer 1981; 43: 486-95. Chapman S, Nakielny R. A guide to radiolog­ica/ procedures. London: Bailliere Tindall; 1986. Evans R, Alexander P. Mechanisms of ex­tracellular killing of nucleated mammalian cells by macrophages. In: Nelson DS, editor. Immunobiology of macrophage. New York: Academic Press; 1976. p. 45-74. Page proofs will be faxed to the correspon­ding author whenever possible. It is their re­sponsibility to check the proofs carefully and fax a list of essential corrections to the editori­al office within 48 hours of receipt. If correc­tions are not received by the stated deadline, proof-reading will be carried out by the edi­tors. Reprints: Fifty reprints are free of charge, for more contact editorial board. For reprint infomzation contact: International Reprint Corporation, 287 East "H" Street, Benicia, CA 94510, USA. Tei: (707) 746-8740; Fax: (707) 746-1643; E-mail: reprints@intlreprints.com