Junko Kumagai

Sex- and Age-Specific Carriers of Hepatitis B and C Viruses in Japan Estimated by the Prevalence in the 3,485,648 First-Time Blood Donors during 1995–2000

Objective: Carriers of hepatitis B virus (HBV) and hepatitis C virus (HCV) in Japan were estimated on a national basis. Methods: Sera from the first-time blood donors aged 16–64 years in eight jurisdictions of the Japanese Red Cross Blood Center during 1995–2000 were tested for hepatitis B surface antigen (HBsAg) and antibody to HCV (anti-HCV). Viremia with HCV was estimated to be present in 70% of donors with anti-HCV. Results: HBsAg was detected in 22,018 of 3,485,648 (0.63%) blood donors including 12,990 of 1,780,149 (0.73%) men and 9,028 of 1,705,499 (0.53%) women, and anti-HCV in 17,010 (0.49%) including 8,504 (0.48%) men and 8,506 (0.50%) women. Multiplying the carrier rate by the population registered in the Census 2000, the total HBV carriers aged 15–65 years were estimated at 967,753 (95% confidence interval 806,760–1,128,745), of whom 571,210 (479,267–663,152) were men and 396,543 (327,494–465,593) were women. Likewise, the total HCV carriers were estimated at 884,954 (95% confidence interval 725,082–1,044,826), of whom 464,363 (377,927–550,799) were men and 420,591 (347,156–494,027) were women. Conclusion: Estimated numbers of HBV and HCV carriers would help plan to prevent the development of hepatocellular carcinoma in Japan.

Titration of Hepatitis C Virus in Chimpanzees for Determining the Copy Number Required for Transmission

Objective: To determine the copy number of hepatitis C virus (HCV) RNA, determined by nucleic acid amplification test (NAT) for screening blood units in Japan, that can transmit infection to chimpanzees. Methods: Fresh-frozen plasma with markers of HCV infection, as well as inocula pedigreed from 1 of them, were evaluated for the infectious activity in chimpanzees. Results: One unit each (273–282 ml) of fresh-frozen plasma from 2 blood donors or a pool from 13 donors to make a unit, which contained high-titered antibody to HCV but without HCV RNA detectable by NAT, did not infect any of 3 chimpanzees. Two chimpanzees were infected, however, when they were inoculated with 1 ml of serum from a blood donor in the ‘window period’ of HCV infection and containing 7.0 × 106 copies/ml of HCV RNA. The preacute phase serum from 1 of them harvested 7 weeks after the inoculation was titrated in 2 chimpanzees, and an inoculum containing approximately 2 × 101 copies of HCV RNA could transmit infection to both of them. Conclusion: Approximately 20 copies of HCV can transmit infection to recipients, which needs to be taken into consideration in planning the screening of blood units for HCV RNA by NAT. Although the sensitivity of present NAT could be improved further, there would be a limit of it in detecting a low-level HCV RNA in the window period of donors with the infectious capacity in recipients.

High incidence of extrahepatic manifestations in an HCV hyperendemic area.

We previously investigated the incidence of extrahepatic manifestations including oral precancerous disease among the inhabitants in a hepatitis C virus (HCV) hyperendemic area in Fukuoka in Japan. The present study design was based on a prospective cohort at the other HCV hyperendemic area. One oral surgeon examined the oral lesions of 59 adult inhabitants (21 men, 38 women; mean age of 70.7 years), of a hyperendemic area of HCV infection. Furthermore, all subjects were interviewed regarding the natural history of extrahepatic manifestations. All sera were examined for antibodies to HCV (anti-HCV), serum HCV RNA, HCV genotype, antinuclear antibody (ANA), rheumatoid factor (RF) activity, and anti-SS-A and-B antibodies. Anti-HCV or HCV RNA was detected in sera from 59 (100%) or 57 (96.7%) of all subjects. Oral lichen planus (OLP), leukoplakia with leukoedema, or only leukoedema was observed in 8 (8.5%), 1 (1.7%), or 2 (3.4%) subjects, respectively. The incidence of all subjects with one or more HCV-related extrahepatic manifestation was 66.1% (39/59). The subjects with dry mouth were 25.4% (15/59). There was no relation among these autoantibodies, symptoms of dry mouth, or prevalence of HCV-associated extrahepatic manifestations. These findings demonstrate that the inhabitants with HCV infection showed various extrahepatic manifestations, and was not always limited to specific HCV areas.

Interleukin-6 Localization and the Prognosis of IgA Nephropathy

Various cytokines and growth factors may be involved in IgA nephropathy. To clarify whether interleukin-6 was a prognostic factor for this disease, we investigated interleukin-6 positivity of renal biopsy specimens and its relationship with the prognosis. The subjects were 90 patients with IgA nephropathy (42 males and 48 females with a median age of 32.7 ± 13.8 years). Renal biopsy specimens were stained for interleukin-6 using an enzyme-antibody method. Fifty-two of 90 patients showed glomerular positivity for interleukin-6. Among the patients positive for interleukin-6, 24-hour urinary protein excretion and serum creatinine levels were significantly higher at the time of biopsy than in the patients without interleukin-6 positivity, while creatinine clearance was significantly lower. In the interleukin-6-positive patients without steroid therapy, serum creatinine increased significantly after 1 year (Δs-Cr; 1.04 ± 0.45 mg/dl) and creatinine clearance decreased significantly (ΔCcr; –11.7 ± 3.2 ml/min) compared to the interleukin-6-negative patients without steroid therapy. Steroid therapy improved 24-hour urinary protein excretion, serum creatinine, and creatinine clearance in the interleukin-6-positive patients, while these parameters worsened without steroid therapy. On the other hand, the IL-6-negative patients showed no differences of clinical parameters irrespective of the presence or absence of steroid therapy. In conclusion, glomerular interleukin-6 positivity may be a prognostic factor and an indicator of the need for steroid therapy in IgA nephropathy.

Precise ultrastructural localization of endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1 in patients with IgA nephropathy

Using light and electron microscopy, we performed an immunohistochemical study of endothelial leukocyte adhesion molecule-1 (ELAM-1), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in 15 patients with IgA nephropathy to clarify the localization of these adhesion molecules. The normal portions of 2 kidneys removed due to localized carcinoma and 3 biopsies from patients without glomerular disease were used as a control. By light microscopy, ELAM-1, VCAM-1, and ICAM-1 all showed positive staining in IgA nephropathy, with the intensity of staining following the sequence ICAM-1 > VCAM-1 > ELAM-1. ELAM-1 and VCAM-1 showed a patchy distribution of moderate staining in the tissues, including the mesangium, crescents, adhesions, and tubules. In contrast, there was marked linear ICAM-1 staining throughout the vascular walls. ELAM-1 and VCAM-1 were positive on the basolateral surfaces of a few proximal tubular epithelial cells in association with inflammatory cell infiltration, while ICAM-1 was found on the brush border. ICAM-1 was positive in the glomerular capillary walls and interstitial vessels of the control kidney tissue, while ELAM-1 and VCAM-1 were virtually absent. By electron microscopy, ELAM-1 positivity on the urinary surface of the parietal/visceral epithelial cells was often associated with adherent mononuclear cells in the urinary space. VCAM-1 positivity was increased in the perinuclear space and/or cytoplasm of mesangial cells as well as at the mesangial cell-endothelial cell interface. These findings suggest that ELAM-1 and VCAM-1 may be more closely related than ICAM-1 to the major histopathological changes occurring in IgA nephropathy, including mesangial expansion, formation of crescents and adhesions, and tubulointerstitial injury.

Lack of Epidemiological Evidence for a Role of Resolved Hepatitis B Virus Infection in Hepatocarcinogenesis in Patients Infected with Hepatitis C Virus in Japan

Objective: The role of resolved hepatitis B virus (HBV) infection in promoting hepatocellular carcinoma (HCC) in patients infected with hepatitis C virus (HCV) in Japan was evaluated by epidemiological surveys. Methods: Antibody to hepatitis B core (anti-HBc) was determined in age-matched blood donors, and the frequency was compared with that in patients with HCV-associated HCC in Japan. Results: Anti-HBc was detected significantly more frequently in the blood donors with than without antibody to HCV (anti-HCV; 76/135 or 56.3% vs. 65/255 or 25.5%, p < 0.001). In the patients with HCV-associated HCC, anti-HBc was detected in 109 of 202 (54.0%), which was comparable to the frequency in anti-HCV-positive blood donors (56.3%). Among the blood donors with anti-HCV, the prevalence of anti-HBc was no different between those with and without HCV RNA in serum (40/77 or 51.9% vs. 36/58 or 62.1%). Conclusions: The individuals of an age with high cancer frequency (≧40 years) in Japan would have been exposed to HBV frequently (>50%), whether or not they have developed HCV-associated HCC. Despite repeated assertions in the literature, no epidemiological evidence was obtained for a role of past HBV infection in hepatocarcinogenesis in patients infected with HCV in Japan.

Early Dynamics of Hepatitis C Virus in the Circulation of Chimpanzees with Experimental Infection

Two chimpanzees were inoculated with hepatitis C virus (HCV) and followed on a daily basis for 12 days. HCV RNA became detectable in their sera on day 5 by polymerase chain reaction with the detection limit of 102 copies/ml. Based on an exponential growth observed until 8 or 9 days after inoculation in their sera, the doubling time of HCV in the circulation was estimated at 6.3–8.6 h and log time (time required to grow 10-fold) at 31.3– 42.9 h. The exact doubling time of HCV determined in them would help plan an efficient strategy for screening out blood donors in the window period of infection between the exposure and the development of antibody to HCV in serum.

Ultrastructural localization of vascular cell adhesion molecule-1 in proliferative and crescentic glomerulonephritis

Recent studies have demonstrated an important role of vascular cell adhesion molecule-1 (VCAM-1) in the pathogenesis of nephritis. In the present study, renal biopsy specimens from patients with proliferative and crescentic glomerulonephritis were subjected to immunoelectron microscopy using an anti-VCAM-1 monoclonal antibody. In control normal kidney tissue, VCAM-1 expression was restricted to the free surface of parietal epithelial cells. In diseased glomeruli, VCAM-1 was expressed on the free surface of parietal and visceral epithelial cells, on the luminal surface of capillary endothelial cells, on infiltrating monocyte/macrophage-like cells, on mesangial cells, and in the matrix of the expanded mesangium. There was also VCAM-1 expression on almost all cell types in the crescents, including macrophage-like cells, fibroblast-like cells, and epithelial cells. Some cells also showed VCAM-1 positivity in the rough endoplasmic reticulum and the perinuclear space. Both the glomerular capillary lumen and urinary spaces of Bowmans capsule contained positive reaction products, which were often associated with exocytosis by the surrounding cells. VCAM-1 was predominantly expressed on the basal and lateral surfaces of a few proximal tubules, but it could not be localized ultrastructurally. These findings suggest that production and secretion of VCAM-1 by both infiltrating monocyte/macrophages and resident glomerular cells may be related to the pathogenesis of proliferative and crescentic glomerulonephritis.

Hepatocyte Growth Factor Localization in Primary Glomerulonephritis and Drug-Induced Interstitial Nephritis

Noriaki Yorioka, MD, 2nd Department of Internal Medicine, Hiroshima University School of Medicine, 1-2-3 Kasumi Minami-ku, Hiroshima City 734 (Japan) Table 1. Correlation between the distribution of HGF on tubule epithelial cells and primary glomerulonephritis, drug-induced interstitial nephritis and control tissue. Dear Sir, Hepatocyte growth factor (HGF) was initially identified as a mitogen for hepatocytes by Nakamura et al. [1] in 1984. Recently, HGF has been shown to be a mitogen for a variety of cell types, including renal tubular cells [2, 3]. Nagaike et al. [4] examined changes in HGF mRNA expression, HGF activity and HGF receptor in the rat kidney following unilateral nephrectomy or treatment with carbon tetrachloride, and suggested that HGF might function as a renotropic factor during renal regeneration after injury. In this study, we present the immunohis-tochemical localization of HGF in patients with primary glomerulonephritis and drug-induced interstitial nephritis. We examined 37 patients (18 males and 19 females; age: 15-74 years, mean ± SD; 40.9 ± 17.6) who underwent renal biopsy prior to treatment in the Second Department of Internal Medicine, Hiroshima University School of Medicine, Hiroshima, Japan. These patients were diagnosed as follows: primary glomerulonephritis, 25 patients (IgA nephropathy, 15 patients; membranous glomerulonephritis, 6 patients; focal glomer-ulosclerosis, 4 patients); druginduced interstitial nephritis, 9 patients (NSAIDs, 4 patients; anticancer agents, 2 patients; antibiotics, 2 patients; interferon-γ, 1 patient), and minimal changes, for control tissue, 3 patients. Kidney tissue fixed in formalin and embedded in paraffin was used to localize HGF. The sections were deparaffmized using xylene and alcohol, and the antigen was unmasked in 0.01% trypsin diluted with 0.06 M PBS, pH 7.2, for 30 min at 37 °C. The primary antibody (monoclonal anti-HGF antibody, Otsuka Pharmaceuticals, Japan) was applied overnight at 4 o C, followed by incubation with the secondary antibody for 30 min at room temperature. Immuno-products were visualized by applying DAB. The relationship between the presence of HGF and the histological stages of IgA nephropathy [5] was evaluated.

Direct infusion of ascites into the blood circuit during hemodiafiltration in uremic patients with cirrhosis.

Two chronic dialysis patients with massive ascites caused by cirrhosis were treated by infusion of their ascites directly into the blood circuit. This stabilized their hemodynamics during dialysis, facilitating the control of weight gain and ascites, and thus markedly improving their general condition. Long-term use of this therapy was able to prevent the accumulation of ascitic fluid. Interestingly, fever occurred when this therapy was performed with hemodialysis, but not with hemofiltration or hemodiafiltration, suggesting that a pyrogen in the ascites was removed by filtration.