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Detection of hepatitis C virus (HCV) in the bile of HCV-infected patients
A1252 AASLD ABSTRACTS
• L0219 DIFFERENT IL-12 p40 RESPONSE IN PATIENTS WITH HEPATITIS C VIRUS -INDUCED CIRRHOSIS AND ALCOHOLIC CIRRHOSIS. Ch. Hanck, M.V. Singer, S. Rossol. Dept. of Int. Medicine IV, University of Heidelberg, Mannheim, Germany. Introduction: The exact mechanisms underlying cirrhosis remain unclear. To investigate the immunoregulatory role of IL- 12 in cirrhosis of different origin, we analysed the circulating concentrations of IL-12, T helper 1 (Thl) -type and Th 2-type cytokines in peripheral blood of patients with (VC) compared with both, patients with alcoholic cirrhosis (AC) and healthy controls (HC). Methods: The serum levels of IL-12 heterodimer, heavy chain IL-12 p40 subunit, IL-2, IL-4, IFN-~,and IL-10 were determined by ELISA in 21 patients with VC (Child Pugh stage A n = 11, Child Pugh stage B n =4, Child Pugh stage C n = 7) and 77 patients with AC (Child Pugh stage A n = 21, Child Pugh stage B n = 22, Child Pugh stage C n = 34). 10 healthy individuals without a history of alcohol abuse and with normal liver function tests served as controls. Results: This study demonstrates significantly elevated serum levels of IL-12 p40 in patients with VC compared with patients suffering from AC or HC (p _< 0.01). The IL 12-p40 and IL-2 response is enhanced in VC compared with AC and HC (p<0,0001) and correlates with the serum levels of IFN-T (Spearman's rank correlation factor r=0,79; p=0.0002). Moreover, IL-12 was found to be significantly correlated with the serum levels of IFN-T (r--0.63, p<0.01) and IL-2 (r=0.6, p=0.01) in patients with VC. Conclusions: The data are indicating, that an antigen-specific and cell mediated immune response mediated by IL 12-p40 is operative in patients with VC, but not AC. IL-12 regulates the production of IFN-7 in VC and is suggested to play an essential role to mount a delayed-type hypersensitivity immune reaction. • L0220 PERCUTANEOUS RADIO-FREQUENCY TISSUE ABLATION-FIRST APPLICATIONS ON IN VITRO BOVINE LIVERS. J. H~insler1, D. Becker 1, W. Mtiller2, D. Neureiter 3, E.G. Hahn1 lUniversity of ErlangenNiJrnberg, Department of Medicine 1, Germany 2Berchtold Corp., Tuttlingen, Germany; 3Universityof Erlangen-Niimberg,Department of Pathology, Germany. Sonographically-guided Radio-Frequency Tissue Ablation applied by percutaneously placed needle can be used for the palliative treatment of primary or secondary liver tumors In vitro experiments were carried out to establish the relations of the coagulation zone to histologic findings and sonographic appearence. Bovine livers were obtained freshly and radiofrequency was applied under ultrasound guidance. The coagulation zone was dissected and examined by determining its three-dimensional diameters (macroscopically) and the histologic appearance of the coagulation margin to the surrounding tissue. In our series of 50 experiments the application time ranged from 30 seconds to 10 minutes. Results: The coagulation zone increased with rising application time in a way that can be described as a negativ logarythmic function. The largest volume was 4 x 5 x 5 cm (length, width, depth) approximating 50 ml. It was no problem to judge the coagulation zone sonographically. The macroscopically and sonographically determined width and length correlated with coefficients of 0.8952 and 0.9562. Microscopically the coagulation necrosis showed sharply outlined margins without any vital cells. Conclusion: Radio-Frequency Tissue Ablation applied under sonographic guidance has certain features which seem to make it a promising method for the palliative treatment of primary and secondary tumors of the liver. L0221 DETECTION OF HEPATITIS C VIRUS (HCV) IN THE BILE OF HCVINFECTED PATIENTS. Y, Haruna, T. Miyamoto, T. Kanda, M. Honda, T. Takao, Departments of Gastroenterology & Diabetology and Surgery, Osaka Prefectural General Hospital, Osaka, Japan. The pathobiology of HCV has not been clarified yet. It is suggested that HCV has similarity in genome to picorna viridae including HAV, and also some researchers reported that bile epithelia could be infected with HCV. If HCV is excreted into bile, it will confirm HCV infection in bile duct leading to bile duct damage in chronic hepatitis C, and the excreted HCV might be involved in intestinal immune system. Therefore, we examined existence of HCV in the bile of patients with chronic hepatitis C. Materials and Methods Twelve patients positive for serum HCV antibody were examined in this study. Eight were also positive for serum HCV RNA by PCR, and the other 4 were negative. When they had abdominal operation, bile was aspirated from their gall bladders by fine needles. Written consents to participate in this study were given by all the patients. Serial dilutions of bile were assayed for HCV RNA by PCR not only to determine HCV RNA levels but also to avoid inhibition of HCV RNA extraction or PCR procedures by bile containing substances. Simultaneously, serial dilutions of serum were assayed for HCV RNA in order to compare the HCV RNA level in bile with that in serum of the same patients. Results HCV RNA was detected in 1:100 diluted bile of all the 5 patients whose serum HCV levels were more than 3.0 Meq/ml. Furthermore, we detected HCV RNA in even 1:1000 diluted bile of 2 patients and in 1:500 of one patient out of the 5. However, HCV RNA was not detected in the other 7 patients, who had low serum levels of HCV (less than 1.1Meq/ml) or were seronegative for HCV RNA. In comparison of HCV levels between bile and
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serum, HCV RNA was detected at the same dilution of bile and serum in all the 5 patients, in whom HCV RNA was detected in their bile. Interestingly, no positive signal of HCV RNA was found when undiluted bile was served for PCR assay. Conclusion The same levels of HCV RNA as in serum were detected in bile of HCV infected patients. L0222 EFFECt OF TROGLITAZONE ON GLUCOSE AND LIPID METABOLISMS IN PATI~LNTS w r r H LIVER CIRRHOSIS COMPLICATED BY DIABETES MELL1TUS. T. Hasegawa, M. Yoneda, K. Nakamura, M. Okada, A. Yumino, K. Sato. Dept. of Gastroenterol, Keiwakai Ebetsu Hosp. and Second Dept. of Med., Asahikawa Med. College, JAPAN. Background: Liver cirrhosis patients often develops diabetes mellitus, and their lipid metabolism becomes catabolic tendency because of the insulin resistance. Therefore, nutritional management for liver cirrhosis patients with diabetes mellitus is usually difficult and treatment by oral hypoglycemic agent, such as glibenclamide, for diabetes mellitus of liver cirrhosis patients is not satisfactory. Troglitazone, which is a newly developed hypoglycemic agent, enhances insulin sensitivity. Purpose: To evaluate the effect of troglitazone for glucose and lipid metabolisms in liver cirrhosis patients complicated by diabetes mellitus. Methods: Seven liver cirrhosis patients complicated by diabetes mellitus were included in this study. All patients were compensated liver cirrhosis, indicating no ascites or jaundice. They were diagnosed diabetes mellitus by 75 g oral glucose tolerance test (75 g GTr) and treated by only diet before the study. Patients, whose serum AST and ALT levels were over 80 IU/L during three months before the study, were excluded from this study to avoid possible hepatic complications by troglitazone. Glibenclamide (1.25-2.5 mg before breakfast) was firstly administered for four months, and then administration of troglitazone (200 nag before breakfast) was introduced for another four months instead of glibenclamid. Serum AST, ALT, cholesterol, triglyceride and free fatty acid levels, and HbAlc were determined before and after the each treatment. 75 g G'I'T was also performed before and after the each treatment and plasma insulin levels were measured to assess insulin resistance. Results: Serum AST and ALT levels were not changed by both glibenclamide and troglitazone. Diabetic control was improved by both glibenclamide and troglitazone, assessed by HbAlc. Insulin resistance was improved by troglitazone, but not by glibenclamide, assessed by plasma insulin levels during 75 g GTT. Troglitazone, but not glibenclamide, increased serum cholesterol level and reduced serum free fatty acid level. Table: Changes of laboratory data after the treatment before treatment glibenclamide troglitazone 75g GTr 0 rain PG (IRI) 107 ± 4 (13 ± 1) 100 ± 5 (15 ± 1) 105 ± 4 (8 ± 1) 120 min PG 0RI) 259±5(183±11) 182±5(185±10) 194±5(98±11) Free Fatty acid mEq/l 1.05 ± 0.26 0.95 ± 0.16 0.50 ± 0.21 T-CHO mg/dl 92 ± 5 90 ± 7 129 ± 4 HbA lc % 6.4 ± 0.3 5.6 ± 0.3 5.5 ± 0.1 PG: plasma glucose mg/dl, IRI: plasma insulin laU/ml (Mean _+SD) Conclusion: These data suggest the therapeutic efficacy of troglitazone for insulin resistance, in liver cirrhosis patients with diabetes mellitus.
• L0223 CHARACTERISTICS OF PATIENTS POSITIVE FOR ANTIHEPATITIS C VIRUS (HCV) ANTIBODIES AND NEGATIVE FOR HCV-RNA BY RT-PCR. E Hashimoto, S Noguchi, M Taniai, N Hayashi. Department of Gastroenterology,Tokyo Women's Medical College, Tokyo Japan. Background & Alms: It is not clear whether the absence of serum HCV-RNA by RT-PCR in patients positive for anti-hepatitis C virus (HCV) can exclude HCV infection. Then, we investigated the characteristics of patients positive for anti-HCV but repeatedly negative for serum HCV-RNA by RT-PCR. Methods: 396 consecutive anti-HCV-positive patients admitted to Tokyo Women's Medical College from Januany 1995 to November 1997, were included in this study. There were 192 with hepatocellular carcinoma (HCC) and the others were scheduled for liver biopsy. None of them had received interferon therapy and patients with acute hepatitis C were excluded. Twentyone patients (4 with HCC) were repeatedly negative for serum HCV-RNA by RT-PCR. After diagnostic biopsy, 10 liver samples from patients without HCC were examined for HCV by RT-PCR. Various clinico-pathological factors were compared between patients negative and positive for HCV-RNA. Results: The HCV-RNA-negative patients showed significantly milder liver disease state than the HCV-RNA-positive patients, There were no significant differences with respect to age, sex, blood transfusion, and alcohol abuse. Among the HCV-RNA-negative patients, 13 had normal liver function tests and 13 showed low titers of anti-HCV. Concerning RIBA-3, 9 showed only C22p positivity, 7 were positive for C33c and C22p, 3 showed weak positivity for C33c or C22p or NS5 and 2 were negative for all bands. Among HCV-RNA-negative patients with HCC, 2 had autoimmune hepatitis with obesity, 1 had severe chronic heart failure, and 1 had alcoholic cirrhosis. Noncancerous liver tissue was obtained, except in the patient with severe chronic heart failure, but it did not show the chracteristic histological features of HCV hepatitis. The patients without HCC who were negative for HCV RNA, included 10 with minimal changes, 5 with mild fatty changes, 1 with portal
• L0219 DIFFERENT IL-12 p40 RESPONSE IN PATIENTS WITH HEPATITIS C VIRUS -INDUCED CIRRHOSIS AND ALCOHOLIC CIRRHOSIS. Ch. Hanck, M.V. Singer, S. Rossol. Dept. of Int. Medicine IV, University of Heidelberg, Mannheim, Germany. Introduction: The exact mechanisms underlying cirrhosis remain unclear. To investigate the immunoregulatory role of IL- 12 in cirrhosis of different origin, we analysed the circulating concentrations of IL-12, T helper 1 (Thl) -type and Th 2-type cytokines in peripheral blood of patients with (VC) compared with both, patients with alcoholic cirrhosis (AC) and healthy controls (HC). Methods: The serum levels of IL-12 heterodimer, heavy chain IL-12 p40 subunit, IL-2, IL-4, IFN-~,and IL-10 were determined by ELISA in 21 patients with VC (Child Pugh stage A n = 11, Child Pugh stage B n =4, Child Pugh stage C n = 7) and 77 patients with AC (Child Pugh stage A n = 21, Child Pugh stage B n = 22, Child Pugh stage C n = 34). 10 healthy individuals without a history of alcohol abuse and with normal liver function tests served as controls. Results: This study demonstrates significantly elevated serum levels of IL-12 p40 in patients with VC compared with patients suffering from AC or HC (p _< 0.01). The IL 12-p40 and IL-2 response is enhanced in VC compared with AC and HC (p<0,0001) and correlates with the serum levels of IFN-T (Spearman's rank correlation factor r=0,79; p=0.0002). Moreover, IL-12 was found to be significantly correlated with the serum levels of IFN-T (r--0.63, p<0.01) and IL-2 (r=0.6, p=0.01) in patients with VC. Conclusions: The data are indicating, that an antigen-specific and cell mediated immune response mediated by IL 12-p40 is operative in patients with VC, but not AC. IL-12 regulates the production of IFN-7 in VC and is suggested to play an essential role to mount a delayed-type hypersensitivity immune reaction. • L0220 PERCUTANEOUS RADIO-FREQUENCY TISSUE ABLATION-FIRST APPLICATIONS ON IN VITRO BOVINE LIVERS. J. H~insler1, D. Becker 1, W. Mtiller2, D. Neureiter 3, E.G. Hahn1 lUniversity of ErlangenNiJrnberg, Department of Medicine 1, Germany 2Berchtold Corp., Tuttlingen, Germany; 3Universityof Erlangen-Niimberg,Department of Pathology, Germany. Sonographically-guided Radio-Frequency Tissue Ablation applied by percutaneously placed needle can be used for the palliative treatment of primary or secondary liver tumors In vitro experiments were carried out to establish the relations of the coagulation zone to histologic findings and sonographic appearence. Bovine livers were obtained freshly and radiofrequency was applied under ultrasound guidance. The coagulation zone was dissected and examined by determining its three-dimensional diameters (macroscopically) and the histologic appearance of the coagulation margin to the surrounding tissue. In our series of 50 experiments the application time ranged from 30 seconds to 10 minutes. Results: The coagulation zone increased with rising application time in a way that can be described as a negativ logarythmic function. The largest volume was 4 x 5 x 5 cm (length, width, depth) approximating 50 ml. It was no problem to judge the coagulation zone sonographically. The macroscopically and sonographically determined width and length correlated with coefficients of 0.8952 and 0.9562. Microscopically the coagulation necrosis showed sharply outlined margins without any vital cells. Conclusion: Radio-Frequency Tissue Ablation applied under sonographic guidance has certain features which seem to make it a promising method for the palliative treatment of primary and secondary tumors of the liver. L0221 DETECTION OF HEPATITIS C VIRUS (HCV) IN THE BILE OF HCVINFECTED PATIENTS. Y, Haruna, T. Miyamoto, T. Kanda, M. Honda, T. Takao, Departments of Gastroenterology & Diabetology and Surgery, Osaka Prefectural General Hospital, Osaka, Japan. The pathobiology of HCV has not been clarified yet. It is suggested that HCV has similarity in genome to picorna viridae including HAV, and also some researchers reported that bile epithelia could be infected with HCV. If HCV is excreted into bile, it will confirm HCV infection in bile duct leading to bile duct damage in chronic hepatitis C, and the excreted HCV might be involved in intestinal immune system. Therefore, we examined existence of HCV in the bile of patients with chronic hepatitis C. Materials and Methods Twelve patients positive for serum HCV antibody were examined in this study. Eight were also positive for serum HCV RNA by PCR, and the other 4 were negative. When they had abdominal operation, bile was aspirated from their gall bladders by fine needles. Written consents to participate in this study were given by all the patients. Serial dilutions of bile were assayed for HCV RNA by PCR not only to determine HCV RNA levels but also to avoid inhibition of HCV RNA extraction or PCR procedures by bile containing substances. Simultaneously, serial dilutions of serum were assayed for HCV RNA in order to compare the HCV RNA level in bile with that in serum of the same patients. Results HCV RNA was detected in 1:100 diluted bile of all the 5 patients whose serum HCV levels were more than 3.0 Meq/ml. Furthermore, we detected HCV RNA in even 1:1000 diluted bile of 2 patients and in 1:500 of one patient out of the 5. However, HCV RNA was not detected in the other 7 patients, who had low serum levels of HCV (less than 1.1Meq/ml) or were seronegative for HCV RNA. In comparison of HCV levels between bile and
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serum, HCV RNA was detected at the same dilution of bile and serum in all the 5 patients, in whom HCV RNA was detected in their bile. Interestingly, no positive signal of HCV RNA was found when undiluted bile was served for PCR assay. Conclusion The same levels of HCV RNA as in serum were detected in bile of HCV infected patients. L0222 EFFECt OF TROGLITAZONE ON GLUCOSE AND LIPID METABOLISMS IN PATI~LNTS w r r H LIVER CIRRHOSIS COMPLICATED BY DIABETES MELL1TUS. T. Hasegawa, M. Yoneda, K. Nakamura, M. Okada, A. Yumino, K. Sato. Dept. of Gastroenterol, Keiwakai Ebetsu Hosp. and Second Dept. of Med., Asahikawa Med. College, JAPAN. Background: Liver cirrhosis patients often develops diabetes mellitus, and their lipid metabolism becomes catabolic tendency because of the insulin resistance. Therefore, nutritional management for liver cirrhosis patients with diabetes mellitus is usually difficult and treatment by oral hypoglycemic agent, such as glibenclamide, for diabetes mellitus of liver cirrhosis patients is not satisfactory. Troglitazone, which is a newly developed hypoglycemic agent, enhances insulin sensitivity. Purpose: To evaluate the effect of troglitazone for glucose and lipid metabolisms in liver cirrhosis patients complicated by diabetes mellitus. Methods: Seven liver cirrhosis patients complicated by diabetes mellitus were included in this study. All patients were compensated liver cirrhosis, indicating no ascites or jaundice. They were diagnosed diabetes mellitus by 75 g oral glucose tolerance test (75 g GTr) and treated by only diet before the study. Patients, whose serum AST and ALT levels were over 80 IU/L during three months before the study, were excluded from this study to avoid possible hepatic complications by troglitazone. Glibenclamide (1.25-2.5 mg before breakfast) was firstly administered for four months, and then administration of troglitazone (200 nag before breakfast) was introduced for another four months instead of glibenclamid. Serum AST, ALT, cholesterol, triglyceride and free fatty acid levels, and HbAlc were determined before and after the each treatment. 75 g G'I'T was also performed before and after the each treatment and plasma insulin levels were measured to assess insulin resistance. Results: Serum AST and ALT levels were not changed by both glibenclamide and troglitazone. Diabetic control was improved by both glibenclamide and troglitazone, assessed by HbAlc. Insulin resistance was improved by troglitazone, but not by glibenclamide, assessed by plasma insulin levels during 75 g GTT. Troglitazone, but not glibenclamide, increased serum cholesterol level and reduced serum free fatty acid level. Table: Changes of laboratory data after the treatment before treatment glibenclamide troglitazone 75g GTr 0 rain PG (IRI) 107 ± 4 (13 ± 1) 100 ± 5 (15 ± 1) 105 ± 4 (8 ± 1) 120 min PG 0RI) 259±5(183±11) 182±5(185±10) 194±5(98±11) Free Fatty acid mEq/l 1.05 ± 0.26 0.95 ± 0.16 0.50 ± 0.21 T-CHO mg/dl 92 ± 5 90 ± 7 129 ± 4 HbA lc % 6.4 ± 0.3 5.6 ± 0.3 5.5 ± 0.1 PG: plasma glucose mg/dl, IRI: plasma insulin laU/ml (Mean _+SD) Conclusion: These data suggest the therapeutic efficacy of troglitazone for insulin resistance, in liver cirrhosis patients with diabetes mellitus.
• L0223 CHARACTERISTICS OF PATIENTS POSITIVE FOR ANTIHEPATITIS C VIRUS (HCV) ANTIBODIES AND NEGATIVE FOR HCV-RNA BY RT-PCR. E Hashimoto, S Noguchi, M Taniai, N Hayashi. Department of Gastroenterology,Tokyo Women's Medical College, Tokyo Japan. Background & Alms: It is not clear whether the absence of serum HCV-RNA by RT-PCR in patients positive for anti-hepatitis C virus (HCV) can exclude HCV infection. Then, we investigated the characteristics of patients positive for anti-HCV but repeatedly negative for serum HCV-RNA by RT-PCR. Methods: 396 consecutive anti-HCV-positive patients admitted to Tokyo Women's Medical College from Januany 1995 to November 1997, were included in this study. There were 192 with hepatocellular carcinoma (HCC) and the others were scheduled for liver biopsy. None of them had received interferon therapy and patients with acute hepatitis C were excluded. Twentyone patients (4 with HCC) were repeatedly negative for serum HCV-RNA by RT-PCR. After diagnostic biopsy, 10 liver samples from patients without HCC were examined for HCV by RT-PCR. Various clinico-pathological factors were compared between patients negative and positive for HCV-RNA. Results: The HCV-RNA-negative patients showed significantly milder liver disease state than the HCV-RNA-positive patients, There were no significant differences with respect to age, sex, blood transfusion, and alcohol abuse. Among the HCV-RNA-negative patients, 13 had normal liver function tests and 13 showed low titers of anti-HCV. Concerning RIBA-3, 9 showed only C22p positivity, 7 were positive for C33c and C22p, 3 showed weak positivity for C33c or C22p or NS5 and 2 were negative for all bands. Among HCV-RNA-negative patients with HCC, 2 had autoimmune hepatitis with obesity, 1 had severe chronic heart failure, and 1 had alcoholic cirrhosis. Noncancerous liver tissue was obtained, except in the patient with severe chronic heart failure, but it did not show the chracteristic histological features of HCV hepatitis. The patients without HCC who were negative for HCV RNA, included 10 with minimal changes, 5 with mild fatty changes, 1 with portal