Low-dose alpha-interferon therapy can be effective in chronic active hepatitis C. Results of a...
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Al162 AASLD ABSTRACTS GASTROENTEROLOGY, VOl. 108, NO. 4 INCREMENT OF T CELLS EXPRESSING CD3L°w, LFA-I HIoH, ICAM- 1 HIOH IN INFLAMMATORYLIVER INDUCEDBY PROPIONIBACTERIUM ACNES AND LIPOPOLYSACCHARIDE IN NEONATALLY THYMECTOMIZED A/J MICE. E. Sanada. Y. Watanabe, M. Kamiyasu, T. Masanaga0 M. Myozaki, T. Nakanishi, G. Kajiyama. First Dept. of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan. Purpose: During the healing stage of acute liver injury induced by Propionibacterium acnes (P.aenes) and lipopolysaeeharide (LPS), neonatally thymectomized (NTx) A/J mice show the pathology like.autoimmune hepatitis accompanied by production of aumantibedies to liver spdcific lipopmtein and antinuclear antibodies. We also have showed that nylor~ wool adherent T cells play a critical role in this experimental hepatitis. The degree of CD3 expression on T cell increases according to the maturation in the thymus, and mature T cells express CD3 in high degree (CD3his h) ~d immature T ceils or extrathymicly maturated T cells express CD3 in low degree (CD31°w). To evaluate the involvement of ICAM-I/LFA-1 adhesion pathway and the degree of CD3 expression on T cells in the inflammatory liver of NTx mice, we analyzed liver-infiltrating T cells and spleen T cells by flow cytometry using anti-CD3, anti-LFA-1 and anti-ICAM-l. Methods: Neonatal thymectomy (NTx) was performed between 2 and 3 days after birth: Mice were classified into 4 groups: The mice in group A had an injection of saline without NTx, the mice in group B had an injection of P.acnes/LPS without NTx, the mice in group C had an injection of saline after NTx. and the mice in group D had an injection of P.aenes/LPS after NTx. In group B and D hepatitis was induced by an intravenous injection of 1.5 rag/20 g body weight of P.acnes followed by an injection of 0.05 l.tg LPS 7 days later. Mice were dissected 2 weeks after the LPS injection, and spleen cells and liver-infiltrating mononuclear cells (LIMC) were prepared for flow cytometric analysis. ' Results: 1) CD3 expression: The degree of CD3 expression in spleen dells was low in group C and D, and normally high in group A and B. The degree of CD3 expression in LIMC was low in group B, C and D, and normally high in group A. 2) ICAM-I/LFA-I expression: 75% of spleen cells in group A and C express both ICAM-1 and LFA-1 (double positive), while 50% of spleen ceils in group B and D were double positive, in all groups 67% to 91% of double positive cells in spleen expressed them in low degree (ICAM-II°W/LFA-11°w). 79% to 96% of LIMC were double positive cells in all groups. Regarding the degree of expression, 68% to 76% of double positive cells in group B, C, and D expressed them in high degree (ICAM-lhlsh/LFA-lhish), while 44% of double positive cells in group A were ICAMilhigh/LFA-lhighcells. Conclusions: T cells expressing CD3 l°w, ICAM-lhlg h, LFA-lhia h increased in inflammatory liver induced by P.acnes/LPS. It suggests important involvement of extrathymicly maturated or immature T cells and their high expression of ICAM-I/LFA-I in the pathogenesis of this hepatitis, ~ • • INCREMENT OF VI36-POSITIVE T CELLS IN LIVER OF NEONATALLY THYMECTOMIZED A/J MICE AND EXPERIMENTAL HEPATITIS INDUCED BY PROPIONI- BACTERIUM ACNES AND LIPOPOLYSACCHARIDE. E. Sanada. Y. Watanabe, M. Kamiyasu, M. Naiki, M; Myozaki, M. Kitamom, T. Nakanishi, G. Kajiyama. First Dept, of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan. During the healing stage of acute liver injury induced by administration of sublethal doses of heat-killed Propionibacterium aches (P. aeries) and lipopolysaccharide (LPS), neonatally thymectomized (NTx) mice show the pathology like autoimmune hepatitis accompanied by production of autoantibodies to liver specific lipoprotein (LSP) and antinuclear antibodies (ANA) and mononuclear cell infiltration in the portal area. We also have showed that both CD4+ T ceils and CD8÷ T cells play a critical role in the inflammatory liver in this model. To clarify the involvement of the specific T cell repertoire in this hepatitis, VB usage of liver T cells and spleen T cells was analyzed by flow cytometry using anti-CD3 and the set of mAbs m TCR-VB such as VII3, VB6, VB8.2, VII9, VIII 1, VBI2. Neonatal thymeetomy was performed between 2 and 3 days after birth. Hepatitis was induced in the mice by an intravenous injection of 1.5 rag/20 g body weight of P. aches followed by an intravenous injection of a non-lethal level (0.05 lig) ofLPS 7 days later. Mice were dissected 2 weeks after the LPS injection. Liver-infiltrating lymphocytes were separated by the method of Abo et al. using a gradient centrifugation. The proportion ofT cells expressing various VB TCRs were analyzed in spleen cells and infiltrating lymphocytes of liver by two color flow cytometry. In results, VB6 TCR-expressing T ceils among liver-infiltrating lymphocytes increased in number in NTx A/J mice with inflammatory liver injury compared with non-thymectomizedA/J mice, while there were no difference in the number of that in spleen cells between NTx and non-thymectomized A/J mice. There were no increase in the number of VI]3, V1~8.2, VI~9, VI31 I, V1312in liver-infiltrating lymphocytes and spleen cells in NTx A/J mice. It has been demonstrated that VII6 TCR expression are strongly associated with T cell recognition of MIs-1a determinants. Taken together, it suggests that T cells expressing VII6 TCR are involved in the pathogenesis of this experimental hepatitis in NTx A/J mice, and also suggests possible involvement of superantigens, such as MIs-1 a determinants in the pathogenesis of this hepatitis induced by P. acnes and lipopolysaccharide. • NON-INVOLVEMENT OF FAS ANTIGEN IN APOPTOSIS OCCURRED IN INFLAMMATORY LIVER INJURY INDUCED BY PROPIONIBACTERIUM ACNES AND LIPOPOLYSACCHARIDE IN M.ICE E. Sanada. Y. Watanabe, M. Kamiyasu, T. Masanaga, M. Myozaki, T. Nakanishi, G. Kajiyama. First Department of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan. Purpose: Fas antigen is a cell surface protein, and Fas/Fas ligand is known to be one of important pathways inducing apoptosis. It has been suggested that apoptosis induced by Fas/Fas ligand may be closely related with liver damage such as acute hepatitis, fulminant hepatitis, chronic hepatitis, and other liver damages. Inflammatory liver injury is induced in mice by administration of Propionibacterium acnes (P.acnes) and Lipopolysaccharide (LPS). To clarify involvement of Fas antigen in inflammatory liver injury, we examined the influences of a defect of Fas antigen on the course of liver damage by using MRL-lpr/lpr (lpr/lpr) mice which did not express Fas antigen, and its congenic strain MRL-+/+ (+/+) mice which expressed Fas antigen on the surface of hepatocytes. Methods : 6 week-old mice before developing lymphadenopathy and suffering from a SLE-like autoimmune disease were used for the experiment. Acute inflammatory liver injury was induced in all mice by an intravenous injection of 0.5 mg/20 g body weight of P. aches followed by an intravenous injection of a non-lethal dose (0.2 gg/20 g body weght) of LPS 7 days later. We examined serum ALT values and histology of liver by hematoxylin-eosin staining. To evaluate occurrence of apoptosis in hepatocytes, we stained the liver by nick-end labeling method, and performed agarose gel electrophoresis of nuclear DNA isolated from the liver. Results : Serum ALT values elevated to 2648+366 KU (mean+SE) in lpr/lpr mice and 768_+135 KU (mean+SE) in +/+ mice at 6 hours after LPS injection, and then serum ALT values gradually decreased to almost normal range in 2 weeks after LPS injection. Histological examination showed diffuse mononuclear cell- infiltration and eosinophilic change of hepatocytes in intralobular region in both of lpr/lpr and +/+ mice, and by nick-end labeling method, positive cells were detected in periportal and intralobular region in both of Ipr/lpr and +/+ mice 6 hours after LPS injection. Nuclear DNA isolated from the liver 6 hours after LPS injection was applied to agarose gel electrophoresis. It demonstrated DNA fragmentation into oligosomes in both of tpr/lpr and +/+ mice. Conclusions : These findings provide the evidence that apoptosis of hepatocytes occurs in inflammatory liver injury induced by P. aches and LPS without involvement of Fas antigen. It suggests that other pathways such as TNF/TNF receptor may mediate the apoptosis of hepatocytes in this liver injury. • LOW-DOSE ALPHA-INTERFERON THERAPY CAN BE EFFECTIVE IN CHRONIC ACTIVE HEPATITIS C. RESULTS OF A MLJLTICENTER, RQ~DOMIZED TRIAL. JM Sanchez-Tapias, R Planas, X Forns, S Ampurdanes, L1 Tito, JM Viver, R Morillas, D Acero, P Mas, JM LLovet, J Rodes, and members of the Catalonian Collaborative Study of Hepatitis. Catalonia. SPAIN. Alpha-interferon (~IFN) is currently accepted as a therapy for chronic hepatitis C (CHC). Since the majority of the unwanted side effects of ~IFN are dose-related, treatment with ic~ doses would be preferable. However, it is uncertain if Ic~ dose alFN therapy is effective in CHC. AI__MM: To determine if low-dose eIFN therapy is effective in the treatment of CFC. PATIENTS AND [~ETHODS: One hundred and forty-one consecutive patients with biopsy proven active CHE (bridging fibrosis was present in 52 and cirrhosis in 19) participated in this study. ~-2b IFN was given t.£.w, for 48 ~e~eks. Patients were randc~nized to initial treatment with 5 MU (Group A) or 1.5 MU (Group B) injections. These doses were reduced in Group A or increaseO in Group B according to response. HCV serotype and changes of serum HCv-RNA were studied in the 54 patients enrolled at one of the participating Centers. RESULTS: Normalization of ALT for at least six months after therapy (sustained response) was OtT=erved in 8 of 70 (i2"/.) from Group A and in 15 of 71 (21%) from Group B (N.S.), Increasing the dose of aIFN was able to induce sustained response in only 5 of 58 patients (9/.) who did not respond to 1.5 MU injection s. In contrast, sustained response was obtained in 15 of 21 patients ~ (71%) who maintained ALT normal during treatment with 1.5 M~J injections. The proportion of patients who presented Sustained response was significantly higher in patients with serotybe 3 than in those with serotype 1 or in those with unidentified serotype. Multivariate analysis showed that only age was independently associated with sustained response. CONCLL~ION: In chronic active hepatitis C the efficacy of aIFN given for 42 weeks at low or moderately high doses is similar. Treatment with low dose is therefore preferable. Increasing doses to patients who do not respond to low-dose therapy may be effective in a few additional cases.
Transcript of Low-dose alpha-interferon therapy can be effective in chronic active hepatitis C. Results of a...
Al162 AASLD ABSTRACTS GASTROENTEROLOGY, VOl. 108, NO. 4
INCREMENT OF T CELLS EXPRESSING CD3 L°w, LFA-I HIoH, ICAM- 1 HIOH IN INFLAMMATORY LIVER INDUCED BY PROPIONIBACTERIUM ACNES AND LIPOPOLYSACCHARIDE IN NEONATALLY THYMECTOMIZED A/J MICE. E. Sanada. Y. Watanabe, M. Kamiyasu, T. Masanaga0 M. Myozaki, T. Nakanishi, G. Kajiyama. First Dept. of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan.
Purpose: During the healing stage of acute liver injury induced by Propionibacterium acnes (P.aenes) and lipopolysaeeharide (LPS), neonatally thymectomized (NTx) A/J mice show the pathology like.autoimmune hepatitis accompanied by production of aumantibedies to liver spdcific lipopmtein and antinuclear antibodies. We also have showed that nylor~ wool adherent T cells play a critical role in this experimental hepatitis. The degree of CD3 expression on T cell increases according to the maturation in the thymus, and mature T cells express CD3 in high degree (CD3his h) ~ d immature T ceils or extrathymicly maturated T cells express CD3 in low degree (CD31°w). To evaluate the involvement of ICAM-I/LFA-1 adhesion pathway and the degree of CD3 expression on T cells in the inflammatory liver of NTx mice, we analyzed liver-infiltrating T cells and spleen T cells by flow cytometry using anti-CD3, anti-LFA-1 and anti-ICAM-l. Methods: Neonatal thymectomy (NTx) was performed between 2 and 3 days after birth: Mice were classified into 4 groups: The mice in group A had an injection of saline without NTx, the mice in group B had an injection of P.acnes/LPS without NTx, the mice in group C had an injection of saline after NTx. and the mice in group D had an injection of P.aenes/LPS after NTx. In group B and D hepatitis was induced by an intravenous injection of 1.5 rag/20 g body weight of P.acnes followed by an injection of 0.05 l.tg LPS 7 days later. Mice were dissected 2 weeks after the LPS injection, and spleen cells and liver-infiltrating mononuclear cells (LIMC) were prepared for flow cytometric analysis. ' Results: 1) CD3 expression: The degree of CD3 expression in spleen dells was low in group C and D, and normally high in group A and B. The degree of CD3 expression in LIMC was low in group B, C and D, and normally high in group A. 2) ICAM-I/LFA-I expression: 75% of spleen cells in group A and C express both ICAM-1 and LFA-1 (double positive), while 50% of spleen ceils in group B and D were double positive, in all groups 67% to 91% of double positive cells in spleen expressed them in low degree (ICAM-II°W/LFA-11°w). 79% to 96% of LIMC were double positive cells in all groups. Regarding the degree of expression, 68% to 76% of double positive cells in group B, C, and D expressed them in high degree (ICAM-lhlsh/LFA-lhish), while 44% of double positive cells in group A were ICAMilhigh/LFA-lhighcells. Conclusions: T cells expressing CD3 l°w, ICAM-lhlg h, LFA-lhia h increased in inflammatory liver induced by P.acnes/LPS. It suggests important involvement of extrathymicly maturated or immature T cells and their high expression of ICAM-I/LFA-I in the pathogenesis of this hepatitis, ~ •
• INCREMENT OF VI36-POSITIVE T CELLS IN LIVER OF NEONATALLY THYMECTOMIZED A/J MICE AND EXPERIMENTAL HEPATITIS INDUCED BY PROPIONI- BACTERIUM ACNES AND LIPOPOLYSACCHARIDE. E. Sanada. Y. Watanabe, M. Kamiyasu, M. Naiki, M; Myozaki, M. Kitamom, T. Nakanishi, G. Kajiyama. First Dept, of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan.
During the healing stage of acute liver injury induced by administration of sublethal doses of heat-killed Propionibacterium aches (P. aeries) and lipopolysaccharide (LPS), neonatally thymectomized (NTx) mice show the pathology like autoimmune hepatitis accompanied by production of autoantibodies to liver specific lipoprotein (LSP) and antinuclear antibodies (ANA) and mononuclear cell infiltration in the portal area. We also have showed that both CD4 + T ceils and CD8 ÷ T cells play a critical role in the inflammatory liver in this model. To clarify the involvement of the specific T cell repertoire in this hepatitis, VB usage of liver T cells and spleen T cells was analyzed by flow cytometry using anti-CD3 and the set of mAbs m TCR-VB such as VII3, VB6, VB8.2, VII9, VIII 1, VBI2. Neonatal thymeetomy was performed between 2 and 3 days after birth. Hepatitis was induced in the mice by an intravenous injection of 1.5 rag/20 g body weight of P. aches followed by an intravenous injection of a non-lethal level (0.05 lig) ofLPS 7 days later. Mice were dissected 2 weeks after the LPS injection. Liver-infiltrating lymphocytes were separated by the method of Abo et al. using a gradient centrifugation. The proportion ofT cells expressing various VB TCRs were analyzed in spleen cells and infiltrating lymphocytes of liver by two color flow cytometry. In results, VB6 TCR-expressing T ceils among liver-infiltrating lymphocytes increased in number in NTx A/J mice with inflammatory liver injury compared with non-thymectomized A/J mice, while there were no difference in the number of that in spleen cells between NTx and non-thymectomized A/J mice. There were no increase in the number of VI]3, V1~8.2, VI~9, VI31 I, V1312 in liver-infiltrating lymphocytes and spleen cells in NTx A/J mice. It has been demonstrated that VII6 TCR expression are strongly associated with T cell recognition of MIs-1 a determinants. Taken together, it suggests that T cells expressing VII6 TCR are involved in the pathogenesis of this experimental hepatitis in NTx A/J mice, and also suggests possible involvement of superantigens, such as MIs-1 a determinants in the pathogenesis o f this hepatitis induced by P. acnes and lipopolysaccharide.
• NON-INVOLVEMENT OF FAS ANTIGEN IN APOPTOSIS OCCURRED IN INFLAMMATORY LIVER INJURY INDUCED BY PROPIONIBACTERIUM ACNES AND LIPOPOLYSACCHARIDE IN M.ICE E. Sanada. Y. Watanabe, M. Kamiyasu, T. Masanaga, M. Myozaki, T. Nakanishi, G. Kajiyama. First Department of Internal Medicine, Hiroshima University School of Medicine, Hiroshima 734, Japan.
Purpose: Fas antigen is a cell surface protein, and Fas/Fas ligand is known to be one of important pathways inducing apoptosis. It has been suggested that apoptosis induced by Fas/Fas ligand may be closely related with liver damage such as acute hepatitis, fulminant hepatitis, chronic hepatitis, and other liver damages. Inflammatory liver injury is induced in mice by administration of Propionibacterium acnes (P.acnes) and Lipopolysaccharide (LPS). To clarify involvement of Fas antigen in inflammatory liver injury, we examined the influences of a defect of Fas antigen on the course of liver damage by using MRL-lpr/lpr (lpr/lpr) mice which did not express Fas antigen, and its congenic strain MRL-+/+ (+/+) mice which expressed Fas antigen on the surface of hepatocytes. Methods : 6 week-old mice before developing lymphadenopathy and suffering from a SLE-like autoimmune disease were used for the experiment. Acute inflammatory liver injury was induced in all mice by an intravenous injection of 0.5 mg/20 g body weight of P. aches followed by an intravenous injection of a non-lethal dose (0.2 gg/20 g body weght) of LPS 7 days later. We examined serum ALT values and histology of liver by hematoxylin-eosin staining. To evaluate occurrence of apoptosis in hepatocytes, we stained the liver by nick-end labeling method, and performed agarose gel electrophoresis of nuclear DNA isolated from the liver. Results : Serum ALT values elevated to 2648+366 KU (mean+SE) in lpr/lpr mice and 768_+135 KU (mean+SE) in +/+ mice at 6 hours after LPS injection, and then serum ALT values gradually decreased to almost normal range in 2 weeks after LPS injection. Histological examination showed diffuse mononuclear cell- infiltration and eosinophilic change of hepatocytes in intralobular region in both of lpr/lpr and +/+ mice, and by nick-end labeling method, positive cells were detected in periportal and intralobular region in both of Ipr/lpr and +/+ mice 6 hours after LPS injection. Nuclear DNA isolated from the liver 6 hours after LPS injection was applied to agarose gel electrophoresis. It demonstrated DNA fragmentation into oligosomes in both of tpr/lpr and +/+ mice. Conclusions : These findings provide the evidence that apoptosis of hepatocytes occurs in inflammatory liver injury induced by P. aches and LPS without involvement of Fas antigen. It suggests that other pathways such as TNF/TNF receptor may mediate the apoptosis of hepatocytes in this liver injury.
• LOW-DOSE ALPHA-INTERFERON THERAPY CAN BE EFFECTIVE IN CHRONIC ACTIVE HEPATITIS C. RESULTS OF A MLJLTICENTER, RQ~DOMIZED TRIAL. JM Sanchez-Tapias, R Planas, X Forns, S Ampurdanes, L1 Tito, JM Viver, R Morillas, D Acero, P Mas, JM LLovet, J Rodes, and members of the Catalonian Collaborative Study of Hepatitis. Catalonia. SPAIN.
Alpha-interferon (~IFN) is currently accepted as a therapy for chronic hepatitis C (CHC). Since the majority of the unwanted side effects of ~IFN are dose-related, treatment with ic~ doses would be preferable. However, it is uncertain if Ic~ dose alFN therapy is effective in CHC. AI__MM: To determine if low-dose eIFN therapy is effective in the treatment of CFC. PATIENTS AND [~ETHODS: One hundred and forty-one consecutive patients with biopsy proven active CHE (bridging fibrosis was present in 52 and cirrhosis in 19) participated in this study. ~-2b IFN was given t.£.w, for 48 ~e~eks. Patients were randc~nized to initial treatment with 5 MU (Group A) or 1.5 MU (Group B) injections. These doses were reduced in Group A or increaseO in Group B according to response. HCV serotype and changes of serum HCv-RNA were studied in the 54 patients enrolled at one of the participating Centers. RESULTS: Normalization of ALT for at least six months after therapy (sustained response) was OtT=erved in 8 of 70 (i2"/.) from Group A and in 15 of 71 (21%) from Group B (N.S.), Increasing the dose of aIFN was able to induce sustained response in only 5 of 58 patients (9/.) who did not respond to 1.5 MU injection s. In contrast, sustained response was obtained in 15 of 21 patients ~ (71%) who maintained ALT normal during treatment with 1.5 M~J injections. The proportion of patients who presented Sustained response was significantly higher in patients with serotybe 3 than in those with serotype 1 or in those with unidentified serotype. Multivariate analysis showed that only age was independently associated with sustained response. CONCLL~ION: In chronic active hepatitis C the efficacy of aIFN given for 42 weeks at low or moderately high doses is similar. Treatment with low dose is therefore preferable. Increasing doses to patients who do not respond to low-dose therapy may be effective in a few additional cases.
