April 1995 AASLD A l129

• P O R T A L HYPERTENSION MODIFIES COLONIC MUCOSAL VASCULATURE. H. Nakajima, Y. Sasald, Y. Uno and A. Munakata. First Department of Internal Medicine, Hirosaki University School of Medicine, Hirosaki, Japan. Aims: Colonic involvements occurring secondary to portal hypertension include rectal variees, vascular eetasias and large hemorrhoids. However, no structural abnormality in the vaseulature underlying portal hypertension has yet been described. The aim of this study is to objectively evaluate alterations in the vaseularity and vasoular pattern. Methods: Eight cirrhotic patients with esophageal varices (aged 46 to 69 years), and age- and gender-matched control subjects were studied. The subjects underwent colonoscopy for recording the image of venous unit, which could be defined as a venous network consisting of confluent venous branches draining into one penetrating vein. Three variables representing characters of the venous structure were calculated for each venous unit : area of the vein bed by using an image processor, total number of branchings and anastomoses by visual count based on the theory of topology. Two or three venous units were analyzed in the eeeum, ascending colon, transverse colon, descending colon, sigmoid colon and the rectum. Results: The venous bed area for the eirrhotie patients, 41 _+37 mm ~ (range: 9-231) was significantly (p<0.01) larger than that for the control subjects 25-_12 mm2(range: 5-53). At each eohinie segment, the venous area for the patients was larger than that for the normal subjects. The difference was significant at the the cecum (p<0.01) and ascending colon (p<0.05). As for any equal venous bed area, the total number of branchings was always greater for the patients. The overall mean of the branchings was 68-+29 for the patients versus 4 2 : 1 7 for the controls (p<0.01). Similarly, as for any equal venous bed area, the total number of anastomoses for the patients was significantly larger than that for the controls. The overan mean of the anastomoses was 7. i-+4.3 for the patients versus 2.5-+2.9 for the conlrols ( p<0.01)

Variables Cirrhotic patients Control subjects p value Vein bed area (ram z ) 41-+37 25_+12 <0.01 No. of branchings 68 _+29 42_+ 17 <0.01 No. of anastomoses 7.1 _+4.3 2.5_+2.9 <0.01

Conclusion: These findings may suggest that , in cirrhotic patients, the colonic mueosal veins increase their caliber and increase branchings and/or anastomoses for the compensation of long-lasting portal hypertension.

• CIIRONIC HEPATITIS B VIRUS INFECTION INDUCED THE APOPTOSIS IN PERIPHERAL T LYMPHOCYTES. K.Nakamura,K.Yuh,H.ShlJo,S.Sugyo, T.Iida,T.gohda,N.Kimura,M.Okumura. First Dept.of Internal Medicine, School of Medicine, Fukuoka Univ., Fukuoka, Japan.

Background It is becoming increasingly clear that activation induced cell death (AICD), also termed apnptosis, plays an important role in the regulation of peripheral blood mononu- clear ceils (PBMCs). Aim We tried to find the evidence of AICD in PBMCs from pa-i-l-ents with HBsAg-posltlve chronic liver disease by flowcytometric assay. Sub~ PBMCs from 4 patients with chronic liver disease type B and 5 normal adults. Methods DNA fragmentation assay: PBMCs from the normal ad-~ were enriched for non-B cells, and they were stimulated with Staphylococcus aureus enterotoxln B in vitro culture for different periods (0,i,2,3,4,5d). Then, Protease K, and RNase A were added to each samples. Samples were electrophoresed over a agarose ffel. Flowcytometrle assay: PBMCs from patients and normal adults were cultured in vitro at 3TC for two hours, and then, (1)PBMCs were rinsed wlth O.i~ Triton X-IO0, and stained with propldium iodide for fractional DNA assay. (2)PBMCs were initially labeled with FITC-conJugated anti-CD3 mAb. Cells were treated with bloti- nylated dUTP and terminal deoxynueleotldyl transferase, and then s t a i n e d wi th PE-conJugated a v l d l n (TdT a s s a y ) . These samples were a n a l y z e d by f l o w c y t o m e t r y . R e s u l t s (1)DNA f r agmen ta t i ons were d e t e c t e d in c e l l s more than 3 days a f t e r in v i t r o c u l t u r e wi th Staphylococcus aureus e n t e r o t o x i n B. Likewise, fractional DNA were also detected in the same cell populations. In TdT assay, labeled cells with dUTP were in- creasingly accord to the duration of cultured periods. These results of our detection system for AICD by flowcytometrie assay were consistent with the findings of DNA fragmentation assay. (2)Fractional DNA was detected in only a patient wlth acute exacerbation. But, labeled cells with dUTP were de- tected in all patients, furthermore in two patients,they had clearly visualized pattern of AICD on flowcytometry. The most of these cells were simultaneously with antl-CD3 mAb positive cells. The percentage of CD3 ÷ dUTP positive cells was significantly increased (p<O.01) in patlents(n=4) than in normal adults(n=5) with the mean (±SD) values at 1.01 + 0.23 and 0.52 ± 0.14, respectively. Conclusions Using DNA agarose gel electrophoresis-based flowcytometrle assay for detection of AICD, we demonstrated the evidence of AICD In PBMCs from chronic liver disease type B. These results indicated that antigen-speciflc T ceils are repeatedly proliferate and following death by the stimulation of HBV- r e l a t e d a n t i g e n s in v lvo .

• E F F E C T OF U R S O D E O X Y C H O L I C A C I D (UDCA) ON PATIENTS W I T H TYPE I A U T O I M M U N E HEPATITIS . K. Nakamura. M. Yoneda, K. Tamori, A. Kimura, T. Kato, S. Yokohama, K. Akiyama, H. Saito, M. Fujita, K. Baba and I. Makino. Internal Medicine II, Asahikawa Med. College, Japan

UDCA has been shown beneficial effects in patients with primary biliary cirrhosis and chronic hepatitis. One of the mechanisms by which UDCA improves liver function is proposed to mediate the immunological effects. On the other hand, autoimmune hepatitis (AIH) is chronic liver disease characterized by immunologic and autoimmunologic features. Therefore, the efficacy of UDCA on liver function in AIH is expected, however nothing is known about the effect of UDCA on AIH. Purpose: To evaluate the effect of UDCA therapy on AIH. Methods: Eight patients with type I AIH, 10 with chronic hepatitis B (HB) and 31 with chronic hepatitis C (HC) were treated with UDCA 600 mg per day for one year. AIH patients were diagnosed by criteria of international autoimmune hepatitis group, 5 patients were definite and 3 were probable AIH. In AIH patients, antinucleic (ANA) and antismooth muscle antibodies (ASMA), lgG and "/-globulin were examined. Clinical liver function tests were performed every 4 weeks before and after the induction of UDCA therapy. In four patients of AIH, histological studies were performed before and one year after the induction of UDCA. Results: UDCA drastically improved transaminase in AIH from 146 + 22 and 163 + 16 to 31 + 2 and 25 + 4 (IU/L; Mean + SE) in AST and ALT, respectively. HC was also improved by UDCA from 103 + 6 and 137 + 9 to 77 + 8 and 93 + 10 in AST and ALT, respectively, however UDCA did not have significant effect on transaminase in HB. Although 88% of AIH became normal in ALT by UDCA, only about 20% of HB and HC became normal. In AIH patients, the levels of IgG and ?-globulin also significantly decreased (p<0.05), and ANA titers in 5 of 8 patients and ASMA titers in 4 of 5 patients were reduced. Furthermore all patients of AIH, performed histological study, were improved in intrahepatic inflammation, but not fibrosis. UDCA therapy of two AIH patients was interrupted and the increase of transaminase was observed, however readoministration of UDCA improved transaminase again. Conclusion: These results suggest that UDCA is useful therapeutic agent in type I autoimmune hepatitis.

Overexpression of LevisY antigen in small hepatcellular carcinoma 'compared with nick end labeling technique and proliferating cell nuclear antigen' R. Nakashio. M. Kitamoto, S. Suemori*, K. Tsuji, S. Kira, Y. Watanabe, T. Nakanishi, and G. Kajiyama. First Dept. of Internal Medicine, Hiroshima University School of Medicine,Hiroshima, Japan *Gastrointestinal Unit, Yokohama City Seibu Hospital, St. Marianna University School of Medicine,Kanagawa, Japan

Since dramatic changes of cellular glycosylation pattern are well known to be closely correlated with differentiation, development and oncogenesis, it is reasonable to think that similar specific changes is associated with apoptosis. So carbohydrate antigen LevisY (its biological meaning was not evident yet) was previously claimed to be closely correlated with apoptosis. To investigate the relationship between LevisY antigen and apoptosis in small hepateellular carcinoma (HCC), we stained liver sections from HCC patients immunohistochemcally by using LevisY antibody, and also stained DNA fragmentation pattems(a markers of apoptosis) by using nick end labeling (NEL) technique. Moreover, we counted proliferating cell nuclear antigen labeling index (PCNA LI) in the LevisY or NEL positive area to examine the biological activity in this area. Methods: 22 sections of HCC less than 3cm in diameter resected from 22 patients were stained by anti LevisY antibody (BM1), anti PCNA antibody (PC10) and also stained by in situ hybridization NEL technique (ApopTag). We microscopically examined main tumor of these sections. LevisY antibody stained cytoplasm positvely, and NEL and PCNA were positvely stained in nuclear. Results: 4 of 22 sections were stained by both of LevisY antibody and NEL technique in the same area of main tumor. Their average of PCNA LI was very low (1.0%), so these areas were thought to be occurring apoptosis. 1 of 22 sections, which was poorly differentiated HCC, was stained by LevisY antibody alone in almost whole area of main tumor. Its PCNA LI was very high (52%), so it was not thought to be occurring apoptosis. 6 of 22 sections were stained by NEL technique alone. Their average of PCNA LI was very low (2.8%). 11 of 22 sections showed negative staining by both techniques. Their average of PCNA LI was high (15.4%). Concluslons: These data suggest that the area showing positive carbohydrate antigen LevisY and NEL together was thought to be occurring apoptosis. On the other hand, the area with positive LevisY alone was not thought to express apotosis. In this area, LevisY antigen was thought to expresse on the cancer cells as a carbohydrate antigen of highly activated oncogenesis marker.