Junko Hirohara

Analysis of regulatory T cells and IgG4-positive plasma cells among patients of IgG4-related sclerosing cholangitis and autoimmune liver diseases

ObjectivesPatients with autoimmune pancreatitis (AIP) characteristically show elevated serum levels of immunoglobulin G4 (IgG4) and abundant infiltration of IgG4-positive plasmacytes in the involved organs. The most common involved organ showing extrapancreatic lesions is the bile duct, which exhibits sclerosing cholangitis (SC). However, the role of IgG4 in the development of IgG4-related SC (IgG4-SC) remains unclear. To clarify the role of IgG4 in IgG4-SC, we have performed an immunohistochemical analysis of the bile duct.MethodsLaboratory and immunohistochemical findings of liver biopsy specimens obtained from patients with IgG4-SC, primary sclerosing cholangitis (PSC), autoimmune hepatitis (AIH), and primary biliary cirrhosis (PBC) were compared. The biopsy specimens were first stained with anti-IgG1, anti-IgG4, and anti-Foxp3 (forkhead box P3) antibodies, and the ratio of IgG4-, IgG1-, and Foxp3-positive cells, respectively, to infiltrated mononuclear cells (IgG4/Mono, IgG1/Mono, Foxp3/Mono) was assessed.ResultsThe ratio of IgG4/IgG1-positive plasma cells was significantly higher in specimens obtained from patients with IgG4-SC than in those from patients with PSC, AIH, and PBC. The Foxp3/Mono ratio in patients with PBC was significantly higher than that in patients with IgG4-SC and PSC. In patients with IgG4-SC, the number of Foxp3-positive cells was significantly correlated with the number of IgG4-positive cells; in the other patient groups, there was no correlation.ConclusionsThe IgG4/IgG1 ratio in the liver may be a useful marker for differential diagnosis of IgG4-SC and PSC. In IgG4-SC, abundant infiltration of regulatory T cells (Tregs) may affect the switching of B cells to IgG4-producing plasmacytes, and there is a possibility that the function of Tregs is different in IgG4-SC and other liver diseases (PSC, AIH, and PBC).

Incidence of and risk factors for hepatocellular carcinoma in primary biliary cirrhosis: National data from Japan

Primary biliary cirrhosis (PBC) primarily affects females and is rarely complicated by hepatocellular carcinoma (HCC). Although the HCC incidence in PBC patients is low, several characteristics and risk factors associated with its development have been reported. In this study, national data concerning the current status of carcinogenesis in PBC patients in Japan are reviewed. Using data from two national questionnaire surveys, we investigated the clinicopathological findings associated with HCC in PBC patients. According to the data of all reviewed PBC patients, the HCC incidence was 2.4% (71/2946). The HCC incidence by gender was 5.1% (19/370) in males and 2.0% (52/2576) in females, and the proportion of males was 26.7%. Prognosis was significantly poorer in the PBC patients with HCC than in those without. Multivariate analysis of risk factors associated with HCC by gender revealed histological stage at the time of PBC diagnosis as an independent risk factor associated with the development of HCC in females, but not in males. Furthermore, data from another national survey of 178 PBC patients with HCC (male/female = 49/129; proportion of males 27.5%) revealed that the duration between the diagnosis of PBC and that of HCC was significantly shorter in males than in females. In addition, histological stage at the time of HCC diagnosis was an independent risk factor for HCC in females, whereas no risk factors were identified in males. Conclusion: these data indicate that males are at risk of developing HCC at any histological stage of PBC. Therefore, male PBC patients in particular should be carefully screened for HCC from the early stages of PBC. (HEPATOLOGY 2013)

Long-term prognosis of primary biliary cirrhosis (PBC) in Japan and analysis of the factors of stage progression in asymptomatic PBC (a-PBC)

Objective: Based on data from a national survey of primary biliary cirrhosis (PBC), the pathology and prognosis of PBC in Japan were clarified. In particular, we tried to perform multivariate analysis of factors useful in determining prognosis of asymptomatic PBC (a-PBC). Methods: The survey was performed 10 times. Responses from 3778 of 4361 registered patients (416 institutions) were investigated (survey period: January 1968-December 1998). At the time of diagnosis, patients were classified as a-PBC or symptomatic PBC (s1-PBC; pruritus only, s2-PBC; jaundice and serum bilirubin level above 2 mg/dl). The survival rate was obtained by the Kaplan-Meier method. Logistic regression analysis was used in multivariate analysis of prognostic factors of a-PBC. Results: There were no significant differences in clinical findings from those in previous reports. The 5-year survival rates of patients with a-PBC, s1-PBC, and s2-PBC at the time of diagnosis were 97, 88, and 53%, respectively. Patients with a-PBC at the time of diagnosis were divided into groups: those in whom the disease progressed to s2-PBC (8%) and did not progress to s2-PBC (92%) at the final examination, and the prognosis was compared between groups. The prognosis was significantly poorer in the s2-PBC progression group. As a result of multivariate analysis for prediction of prognosis, levels at diagnosis of total serum bilirubin (T-Bil), albumin (Alb), total cholesterol (T-Cho), histological stage, and presence or absence of ursodeoxycholic acid (UDCA) administration were selected as significant factors (P<0.00001). Conclusion: Serum T-Bil, Alb, T-Cho, and histological stage at the time of diagnosis and presence or absence of UDCA administration were considered useful early prognostic indicators in patients diagnosed as having a-PBC whose prognosis may deteriorate with progression to s2-PBC.

Guidelines for the management of primary biliary cirrhosis: The Intractable Hepatobiliary Disease Study Group supported by the Ministry of Health, Labour and Welfare of Japan Guidelines for the management of primary biliary cirrhosis: The Intractable Hepatobiliary Disease Study Group supported by the Ministry of Health, Labour and Welfare of Japan

THE JAPANESE VERSION of the clinical practice guidelines for primary biliary cirrhosis (PBC) was developed in 2012 by the Intractable Hepatobiliary Disease Study Group, with the support of the Ministry of Health, Labour and Welfare of Japan, for the use of general physicians, gastroenterologists and hepatologists who treat patients with PBC. In preparation for developing the guidelines, the study group reviewed recent studies that provided important evidence or that were published in leading journals with a high impact factor, in addition to considering the formal consensus of experts on PBC or related subjects. Using the core keywords “primary biliary cirrhosis,” a PubMed search was conducted for English-language clinical trials, randomized clinical trials (RCTs) and meta-analyses that were published from January 1998 to December 2009 and that addressed treatment of PBC and its complications, follow-up, indication of and time of consultation for liver transplantation, or time of consultation with specialists. Medical systems and other culture-specific factors in Japan were also taken into account. Members of the task force exchanged ideas frequently during the drafting process to try and establish a consensus. The final draft was made after collecting comments from the public and all the committee members. The level of evidence (LE; Table 1) and the grade of recommendation (GR; Table 2) were based on the Medical Information Network Distribution Service in Japan (MINDS). After being modified by recent literatures published since 2010, the present English version of the guidelines was developed in order to spread our ideas and exchange opinions with physicians who are involved in the management of PBC patients overseas. *Working Subgroup (English version) for Clinical Practice Guidelines for Primary Biliary Cirrhosis (in alphabetical order): Atsumasa Komori, Clinical Research Center, National Hospital Organization Nagasaki Medical Center; Atsushi Tanaka, Department of Medicine, Teikyo University School of Medicine; Hajime Takikawa, Department of Medicine, Teikyo University School of Medicine; §Hirohito Tsubouchi, Digestive Disease and Life-style Related Disease, Kagoshima University Graduate School of Medical and Dental Sciences and Kagoshima City Hospital; †Hiromi Ishibashi, International University of Health and Welfare/Fukuoka Sanno Hospital and Clinical Research Center, National Hospital Organization Nagasaki Medical Center; Hiroto Egawa, Department of Surgery, Institute of Gastroenterology, Tokyo Women’s Medical University; Junko Hirohara, Third Department of Internal Medicine, Kansai Medical University; Ken Shirabe, Department of Surgery and Science, Kyushu University; Kenichi Harada, Department of Human Pathology, Kanazawa University Graduate School of Medicine; Makoto Nakamuta, Department of Gastroenterology, National Hospital Organization Kyushu Medical Center; Mikio Zeniya, Department of Gastroenterology, Jikei University Graduate School of Medicine; Minoru Nakamura, Clinical Research Center, National Hospital Organization Nagasaki Medical Center and Department of Hepatology, Nagasaki University Graduate School of Biomedical Sciences; Nobuyoshi Fukushima, Department of Gastroenterology, National Hospital Organization Kyushu Medical Center; Shinji Shimoda, Medicine and Biosystemic Science, Graduate School of Medical Sciences, Kyushu University; Shotaro Sakisaka, Department of Gastroenterology and Medicine, Fukuoka University Faculty of Medicine; Toshio Morizane, Japan Council for Quality Health Care; Yasuaki Takeyama, Department of Gastroenterology and Medicine, Fukuoka University Faculty of Medicine; ‡Yasuni Nakanuma, Department of Human Pathology, Kanazawa University Graduate School of Medicine; Yoshiyuki Ueno, Department of Gastroenterology, Yamagata University Faculty of Medicine (†Chairperson of the Working Group, ‡Chairperson of the PBC Subcommittee, §Chairperson of the Intractable Hepatobiliary Disease Study Group). **Correspondence: Hiromi Ishibashi, Fukuoka Sanno Hospital, International University of Health and Welfare, 3-6-45 Momochihama, Sawara-ku, Fukuoka, 814-0001, Japan. Email: hiishibashi-gi@umin.ac.jp bs_bs_banner

Properties and localization of phospholipase A2 activity in rat stomach.

In the glandular stomach of rat, phospholipase A2 activity was detected and characterized by the use of sonicated 1-acyl-2-[1-14C]oleoyl-sn-glycero-3-phosphocholine (1 mM in 200 mM glycylglycine buffer/2 mM CaCl2) as substrate. The specific activity was 4.9 X 10(-2) mumol/min per mg protein in the whole glandular stomach homogenate (n = 8). It showed individual variation among rats. The optimal pH was 8.0. The activity was relatively heat-resistant and calcium-dependent. More than 90% of phospholipase A2 activity was located in the corpus and less than 10% was in the antrum. In the corpus wall, which consists of mucosa, submucosa, muscularis externa and serosa, the mucosal part (mucosa and submucosa) contained 80-90% of total activity. The effect of some clinical agents (ulcerogenic and anti-ulcer agents) on the phospholipase A2 activity was examined. The activity in the corpus was inhibited by cimetidine (50% inhibition at 5 X 10(-3) M) and stimulated by indomethacin (50% stimulation at 5 X 10(-5) M). Cetraxate hydrochloride (10(-6)-10(-3) M), 16,16-dimethylprostaglandin E2 (10(-7)-10(-4) M) and dexamethasone (10(-7)-10(-3) M) had no effect.

Purification and characterization of phospholipase A2 from rat stomach

Phospholipase A2, which is localized in the mucosal part of the corpus of rat stomach (Hirohara et al. (1987) Biochim. Biophys. Acta 919, 231-238), was purified 990-fold from the supernatant of a tissue homogenate by heat treatment at acidic pH, ammonium sulfate fractionation, ion-exchange chromatography, gel-filtration and reverse-phase high-performance liquid chromatography (reverse-phase HPLC). The purified enzyme gave a single protein band on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis with a molecular mass of approx. 17 kDa. The enzyme had a pH optimum of 8.0 and hydrolyzed the 2-arachidonoyl residue of phosphatidylcholine preferentially to the 2-oleoyl residue, the Vmax and Km values for the two being 227 and 29 mumol/min per mg protein and 0.037 and 0.019 mM, respectively. The activity was calcium-dependent and was markedly increased by SDS and dimethyl sulfoxide (DMSO). The enzyme showed typical product inhibition. Free unsaturated fatty acids (oleic, arachidonic and docosahexaenoic acids), which are supposedly the main enzymatic products in vivo, inhibited the activity. Arachidonic acid caused noncompetitive inhibition and its concentration for its maximal inhibition (50% inhibition) was 5 x 10(-5) M. Lysophosphatidylcholine, free saturated fatty acids (palmitic and stearic acids) and arachidonic acid metabolites (leukotrienes and prostaglandins) had no effect on the activity.

A Prospective Randomized Controlled Trial of Preoperative Whole-Liver Chemolipiodolization for Hepatocellular Carcinoma

BackgroundWe previously reported that preoperative chemolipiodolization of the whole liver is effective for reducing the incidence of postoperative recurrence and prolonging survival in patients with resectable hepatocellular carcinoma (HCC). The present randomized controlled trial was performed to evaluate the influence of preoperative transcatheter arterial chemoembolization (TACE) on survival after the resection of HCC.MethodsOperative results and long-term outcome were prospectively compared among 42 patients who received only selective TACE targeting the tumor (selective group), 39 patients who received TACE targeting the tumor plus chemolipiodolization of the whole liver (whole-liver group), and 43 patients without preoperative TACE or chemolipiodolization (control group).ResultsThere were no serious side effects of TACE or chemolipiodolization and the operative outcomes did not differ among the three groups. Even though preoperative TACE induced complete tumor necrosis, there were no significant differences in the pattern of intrahepatic recurrence or the time until recurrence among the three groups. There were also no significant differences in disease-free survival or overall survival among the three groups, even among patients with larger tumor size.ConclusionThese results indicate that preoperative selective TACE and whole-liver chemolipiodolization plus TACE do not reduce the incidence of postoperative recurrence or prolong survival in patients with resectable HCC.

Living donor liver transplantion for primary biliary cirrhosis with autoimmune hemolytic anemia: a case report.

K. Tanaka Insitute of Biomedical Research and Innovation, Foundation of Biomedical Research and Innovation, 2-2 Minatojima-Minamimachi, Chuo-ku, Kobe 650-0047, Japan quently, it is considered a prototypical autoimmune disease [1]. Hemolysis can be detected in more than half of all patients with cirrhosis, regardless of the etiology [2]. However, little has been published regarding any association between PBC and autoimmune hemolytic anemia (AIHA) or other severe hemolytic states [3]. To our knowledge, liver transplantion in a PBC patient with AIHA has not been described previously. Here we report a case of living donor liver transplantation for PBC associated with AIHA.

Recurrent Hepatitis C After Living Donor Liver Transplantation Detected by Tc-99m GSA Liver Scintigraphy

Recurrence of hepatitis C virus (HCV) after living donor liver transplantation was investigated using technetium-99m- diethylenetriaminepentaacetic acid-galactosyl human serum albumin (Tc-99m-GSA) liver scintigraphy. Four patients with decompensated cirrhosis due to HCV infection were retrospectively reviewed in this study. Scintigraphy was performed to determine the hepatic uptake ratio of the tracer corrected for disappearance from the blood, as well as the maximal removal rate of the tracer by hepatocytes, as parameters of hepatic functional reserve. In all patients, serum HCV ribonucleic acid (RNA) was detected 3 months after transplantation. The corrected hepatic uptake ratio and removal rate showed little change after transplantation in two patients without the recurrence of HCV infection. In another two patients, these levels were decreased at 3 months after transplantation. In one patient, recurrent HCV infection was diagnosed by confirmatory histologic examination at 12 months after transplantation. In the other patient, both levels declined further at 8 months. Although treatment was initiated with a combination of interferon plus ribavirin, this patient died of progressive hepatic failure. In conclusion, a decrease in scintigraphic parameters at 3 months after transplantation suggests recurrent HCV infection affecting the graft. Tc-99m-GSA liver scintigraphy is a useful noninvasive method for evaluating graft functional reserve.

Colchicine‐induced Inhibition of Fat Globule Development in Hepatocytes of Rats Injected with Ethionine

To examine the effect of colchicine on ethionine induced fatty liver, adult female rats were starved overnight and then injected i.p. with 1 g kg ethionine at 11th hour of fasting; then a half of the rats were also injected i.p. with 2.5 mg kg colchicine twice at 3 and 6 h after the single administration of ethionine. Similarly, fasted control rats were injected i.p. with vehicle alone at the above times. All of the rats were sacrificed after a 20 h fast, and the hepatocytes in periportal areas were observed ultra‐structurally. In addition, total lipids in the liver tissue were extracted and determined biochemically. Although similar significant increases of triglyceride were observed in the liver tissue of all ethionine‐injected rats, the hapatocytes in the group treated with both chemicals had fewer cytoplasmic fat globules (CFG) than those in the group treated with ethionine only. On the other hand, the diameters of markedly increased membrane‐bound lipid particles (MLP) in the double treated group were distributed mainly in the range 0.2–0.4 μm, compared with those (0.1‐0.2 μm) in the other groups. These findings indicate that colchicine inhibits the development of CFG in ethionine injured hapatocytes. Acta Pathol Jpn 39: 281∼288, 1989.