Ultrastructural study of lymphocytic interaction with hepatocytes and endothelial cells in acute non-A, non-B hepatitis

In the liver biopsy specimens of all six patients with acute non-A, non-B hepatitis, the lymphocytic interaction with hepatocytes and sinusoidal endothelial cells was observed by electron microscopic study. Lymphocytes were in a close contact with damaged hepatocytes and interrupted endothelial cells, and the microvilli on the surface of these damaged hepatocytes were degenerated and lost. These findings pointed out the possibility that the lymphocyte may play one of the important roles in hepatocytic damage and endothelial cell damage in acute non-A, non-B hepatitis.     

Studies on transmission of human non-A, non-B hepatitis to marmosets

Two sera obtained from four healthy blood donors, which caused non-A, non-B post-transfusion hepatitis in two recipients, were experimentally inoculated into nine marmosets. Three of seven marmosets developed acute hepatitis characterized by the elevation of serum concentrations of glutamic pyruvic transaminase (GPT) and/or isocitric dehydrogenase (ICD) 8-11 weeks after inoculation. Four of seven showed histopathological changes of acute hepatitis in liver biopsy specimens during the biochemically acute phase. In electron microscopic examination, attached membrane-like structures, which consisted of two-unit membranes of two neighboring endoplasmic reticula with electron-dense material between them, were noted in cytoplasm of hepatocytes during the acute phase of hepatitis. Furthermore, acute-phase sera obtained from two animals were inoculated into four additional marmosets, and non-A, non-B hepatitis was successfully passaged in two of them. The results of this study indicate that certain species of marmoset monkeys are susceptible to human non-A, non-B hepatitis agents and provide a useful animal model for non-A, non-B hepatitis.     

Occurrence of Tubuloreticular Inclusions in Acute Viral Hepatitis

Light and electron microscopy were used to study the liver needle-biopsy material of 20 patients with acute viral hepatitis. According to clinical and serologic data, 5 cases proved to be acute viral hepatitis type A, 13 were type B, and 2 were type non-A/non-B. In 2 of the hepatitis type A, in 6 of the hepatitis type B, and in one of the type non-A/non-B cases, tubuloreticular inclusions (TRI) were found in the endoplasmic reticulum of the sinusoidal endothelial cells of the liver.

These data support the possible presumption that the inclusions represent a characteristic reaction of the endoplasmic reticulum to different influences, as for example to viral infection, rather than to the virus itself.     

Non-A/non-B hepatitis in experimentally infected chimpanzees: cross-challenge and electron microscopic studies

Inoculation of eight chimpanzees with factor VIII, factor IX, or "H" strain plasma resulted in enzymatic and histopathologic evidence of non-A/non-B hepatitis in all eight animals. Challenge of two chimpanzees convalescent from factor VIII-induced disease with either factor IX or "H" strain plasma resulted in non-A/non-B hepatitis only in the animal inoculated with factor IX materials. Reciprocal cross-challenge of a chimpanzee convalescent from factor IX-induced disease with factor VIII also produced unequivocal enzymatic and histopathologic evidence of non-A/non-B hepatitis. Cross-challenge of a chimpanzee convalescent from "H" strain-induced non-A/non-B hepatitis with factor VII did not cause a second bout of non-A/non-B hepatitis. These findings suggest the factor VIII materials and "H" strain plasma used in these studies share a common etiologic agent (or agents), but that factor VIII and factor IX may contain two distinct agents. Electron microscopic (EM) examination of thin-sectioned, acute-phase liver biopsies from all but one of the chimpanzees receiving the primary inocula revealed the presence of abnormal hepatocyte cytoplasmic structures previously shown to be associated with non-A/non-B hepatitis. Crystalline structure containing 25 to 30 nm particles were visualized by EM in the cytoplasm of endothelial or Kupffer cells in acute-phase liver biopsies obtained from three chimpanzees inoculated with either factor VIII materials or "H" strain plasma.     

Community-acquired non-A, non-B hepatitis: clinical characteristics and chronicity

The characteristics of 86 patients with acute non-A, non-B hepatitis were compared to 23 patients with acute hepatitis A and 76 with acute hepatitis B by medical record reviews of patients seen at 5 hospitals in Baltimore, Maryland, as part of case-control study of viral hepatitis. Results of serum aminotransferase levels, bilirubin, albumin, and prothrombin times alone could not distinguish the type of viral hepatitis because of extensive overlap. The alanine aminotransferase range for non-A, non-B hepatitis was 56 to 1819 IU/liters, for hepatitis A 250 to 1995 IU/liters, and for hepatitis B 203 to 2120 IU/liters. The ranges of aspartate aminotransferase and bilirubin for the types of hepatitis also overlapped. Fewer patients with non-A, non-B hepatitis or hepatitis A had a prolonged prothrombin time compared to patients with hepatitis B. Hepatic encephalopathy was seen only in two patients with hepatitis B. Forty-two percent of non-A, non-B hepatitis patients followed for 6 months or longer continued to have elevated alanine aminotransferase levels. Chronic alanine aminotransferase elevation was independent of the source of infection: transfusion, parenteral drug use, or all other sources. Prolonged follow-up is necessary to evaluate chronicity in patients with non-A, non-B hepatitis.     

Further evidence of cellular changes associated with non-A, non-B hepatitis

Two distinct types of ultrastructural changes were found in the hepatocytes of chimpanzees infected with two forms of non-A, non-B hepatitis. In the type of infection that was of long incubation, there was a marked cytoplasmic derangement of the endoplasmic reticulum, with the formation of tubules, but no pathological changes in the nuclei. In the short-incubation type of non-A, non-B hepatitis, induced experimentally in a chimpanzee that had recovered from the long-incubation type of infection, nuclear alterations were found together with the presence of aggregates of particles measuring 15--27 nm in diameter. Cytoplasmic tubules were not seen in this type of non-A, non-B infection.     

Spumaviruses isolated from sources containing agents of non-A, non-B (NANB) hepatitis do not cause NANB hepatitis

Serum and liver tissue containing infective non-A, non-B hepatitis virus were shown to contain a retrovirus-like agent that replicated when inoculated into chimpanzee liver cell cultures in vitro. The virus appeared to assemble its core particles in association with tubular structures reminiscent of those characteristically seen in non-A, non-B hepatitis virus-infected chimpanzee liver in vivo, and produced syncytial cytopathic effects in a number of continuous and a primary mammalian liver cells. The agents were neutralized by acute and convalescent sera from human and chimpanzee cases of non-A, non-B hepatitis, as well as by antisera against simian spumavirus type 7, but not type 6. Aluminum chloride failed to abolish viral infectivity. There was no evidence of virus replication or hepatitis in chimpanzees inoculated with a seventh passage of one of the isolates. Thus the data suggest that the isolates are not causally related to non-A, non-B hepatitis, as was previously postulated.     

Non-a, non-b hepatitis: identification of hepatitis-B-like virus particles in serum and liver

Hepatitis B virus-like particles including: small spheres and filaments 15--25 nm in diameter together with a 35--40 nm Dane particle-like virion have been identified in sera of patients with non-A, non-B hepatitis. In a coded serological study, such particles were detected transiently in 3/4 acute, and persistently in 7/8 chronic cases of non-A, non-B hepatitis with non-A, non-B antigenemia. Only 2/12 similar cases without non-A, non-B antigens (Ag) in serum had detectable particles but neither patients with drugs, or type A hepatitis, nor cases of obstructive jaundice. The particles did not express hepatitis B surface (HBs) or non-A, non-B Ag at their surface but were associated, in three patients, with significant endogenous DNA polymerase activity. Furthermore, particles similar to hepatitis B cores (BHc) and also associated with DNA polymerase activity were demonstrated by sucrose gradient ultracentrifugation of a liver homogenate obtained from a patient who had died of non-A, non-B hepatitis. The non-A, non-B hepatitis virion described here appears, therefore, as a hepatitis B-like virus. The exact kinship between these two agents is currently being investigated.     

Detection of an antigen-antibody system in serum associated with human non-A, non-B hepatitis.

An antigen was detected by counterelectrophoresis in serum samples from six of seven chimpanzees during the acute phase of experimentally induced non-A, non-B hepatitis using antiserum from a chimpanzee convalescent from human non-A, non-B hepatitis. This antigen could not be detected in 35 preinoculation serum samples from these chimpanzees, or in 94 weekly bleedings from three chimpanzees with hepatitis A and three chimpanzees with hepatitis B. The antigen was detected in serum samples obtained from three humans with chronic non-A, non-B hepatitis whose blood had transmitted non-A, non-B hepatitis to other humans (including a nurse by accidental needlestick) and to chimpanzees by experimental inoculation. In addition, the antigen was detected in serum obtained retrospectively from 11 to 31 former blood donors whose blood had transmitted posttransfusion non-A, non-B hepatitis several years previously to recipients of a single unit of their blood. Antibody to this antigen was detected in convalescent serum samples from all seven chimpanzees studied, in convalescent serum from the nurse infected by accidental needlestick, and in serum from a hemodialysis patient convalescent from non-A, non-B hepatitis.     

Ultrastructural alterations in hepatocytes and sinus endothelia in experimental non-A, non-B hepatitis in chimpanzees with and without immunoglobulin prophylaxis

Three chimpanzees were inoculated with an infectious factor VIII preparation. Two of the chimpanzees received in addition a human immunoglobulin preparation as used for prophylaxis in humans. All three chimpanzees developed an acute limited non-A, non-B hepatitis as judged from light and electron microscopic markers after an incubation period of two weeks. The use of immunoglobulin did not prevent the infection. A prolonged incubation of 15 weeks, however, was observed in one animal when alanine aminotransferase (ALT) elevation was used as criterion of infection. In the electron microscope, non-A, non-B hepatitis was characterized by tubular structures, spongelike inclusions and attaching curved membranes, in the absence of nuclear viruslike particles. An additional finding were viruslike particles in crystalline arrays which were found in the cytoplasm of sinusoidal-lining endothelial cells and tubuloreticular complexes.     

Precipitin system detected in sera from patients with non-A,non-B hepatitis

A precipitin system detected by immunodiffusion was identified using sera from patients with non-A, non-B hepatitis. This system resembled those described by other authors as serological tests for non-A, non-B antigens and antibodies. However, testing of sera from a large number of patients with acute and chronic liver diseases revealed that this precipitin system was not specific for non-A, non-B hepatitis. Further characterization of the precipitin system demonstrated that it did not represent a true antigen-antibody reaction. Precipitin lines detected by immunodiffusion during the course of acute and chronic non-A, non-B hepatitis may not represent reactions specific for this disease.     

Seroepidemiology of hepatitis E virus in patients with non-A, non-B, non-C hepatitis in Hungary

Many cases of acute hepatitis remain undiagnosed and the hepatitis E virus (HEV) is emerging in industrialized countries. The aim of this study was to assess the role HEV as causative agent in acute non-A, non-B, and non-C hepatitis patients in Hungary. 10.5% of the 264 acute non-A, non-B, and non-C hepatitis patients tested had anti-HEV IgG and 1.9% had anti-HEV IgM as tested by ELISA. After confirmation by Western blot 6.1% of the acute non-A, non-B, and non-C hepatitis patients had anti-HEV IgG antibodies only and 1.1% of the patients had both IgG and IgM. All 19 patients that were positive for anti-HEV IgG and/or IgM tested negative for HEV RNA by PCR. Only a small proportion of the acute hepatitis cases in the southwest of Hungary are assumed to be attributed to HEV infection, however, hepatitis E should be considered along with hepatitis A, B, and C in the diagnosis of acute hepatitis.     

Absence of detectable hepatitis B virus DNA in sera and liver of chimpanzees with non-A, non-B hepatitis

The risk of hepatitis B infections has been reduced by screening of blood donors for hepatitis B surface antigen (HBsAg). However, recipients remain at significant risk of developing post-transfusion hepatitis. Studies have shown that non-A, non-B hepatitis virus(es) are responsible for the majority of post-transfusion hepatitis infections. In spite of many efforts, these non-A, non-B hepatitis viruses have not yet been identified. Epidemiological studies, however, suggest that non-A, non-B hepatitis shares many features with hepatitis B. Recently, Wands et al [1982] showed, in chimpanzees infected with non-A, non-B hepatitis agents, the presence of antigenemia or viremia by radioimmunoassay with monoclonal antibodies directed toward distinct determinants of HBsAg and by molecular hybridization analysis. They suggested that non-A, non-B hepatitis agents may be related, but distinct variant(s) of hepatitis B virus (HBV). In this study, five chimpanzees were inoculated with three different agents that have been shown to transmit non-A, non-B hepatitis. The following inocula were used (I) a factor VIII preparation kindly provided by D.W. Bradley, (II) acute phase serum from a chimpanzee infected with the F strain kindly provided by A.J. Zuckerman, and (III) a DS-antigen serum previously shown by us to transmit non-A, non-B hepatitis [Duermeyer et al, 1983]. All chimpanzees developed a rise in transaminase levels between 8 and 10 weeks after inoculation. None of the chimpanzees was positive for any markers of HBV infection. No evidence was obtained of infection with hepatitis A, cytomegalovirus, or Epstein-Barr virus. One chimpanzee developed chronic liver disease.(ABSTRACT TRUNCATED AT 250 WORDS)     

Acute sporadic hepatitis in Sudanese children.

Eighty consecutive cases of acute viral hepatitis and 80 controls selected from a public pediatric clinic were entered into a study of acute sporadic hepatitis in Khartoum, Sudan. Study subjects were 14 years of age or younger and were mainly from a low socioeconomic level. Non-A, non-B hepatitis was diagnosed by exclusion in 35 (43.8%) patients, hepatitis A in 27 (33.8%), acute hepatitis B in 8 (10.0%), possible Epstein-Barr virus (EBV) hepatitis in 1 patient; and dual hepatitis A and B infection in 1 patient. Eight acute cases were positive for HBsAg but negative for anti-HBc IgM and anti-HAV IgM. Delta hepatitis was not identified in any study subject. A household case of jaundice and acquaintance with an individual outside of the household with jaundice during the prior 6 months were associated with non-A, non-B hepatitis. There was no association between parenteral exposure and non-A, non-B hepatitis. These findings suggest that enterically transmitted non-A, non-B hepatitis may be a major cause of acute sporadic hepatitis in children in this area, as well as a cause of epidemic hepatitis.     

Liver histopathology of the hepatitis A virus infection: a comparison with hepatitis type B and non-a, non-b

Liver biopsies from 86 patients with serologically established acute hepatitis A were evaluated for quantitative and qualitative light microscopic features together with biopsies from 78 patients with acute hepatitis type B and 76 patients with acute hepatitis type non-A, non-B. Hepatitis A was characterised by more pronounced portal inflammation than hepatitis non-A, non-B (p less than 0.01) but less conspicuous parenchymal changes (focal necrosis, Kupffer cell proliferation, acidophil bodies, ballooning) than found in hepatitis type B (p less than 0.01). Steatosis occurred in 10% of the hepatitis A biopsies compared with 26% (p less 0.01) and 6% (not significant) in the hepatitis non-A, non-B and B groups, respectively. A comparison between the histological findings in women and men revealed that iron deposits occurred in more than half of the men compared to less than 20% of the women (p less than 0.01) irrespective of hepatitis type. Histological and biochemical follow-up was available in 36 patients with hepatitis A. For the majority of these patients the bilirubin concentration reached normal values within one month of the initial biopsy. The activity of serum transaminases showed good correlation with the degree of histological resolution. Non-specific reactive hepatitis with slightly raised serum transaminases were often seen during recovery from hepatitis A. These patients may be misinterpreted as cases of acute non-A, non-B hepatitis.     

Autoantibodies against liver cell membrane antigens detected by enzyme-linked immunosorbent assay in acute and chronic liver disease

An enzyme-linked immunosorbent assay was developed to detect serum antibody binding to liver membrane antigen derived from human hepatoma cell line SK-Hep-1. When we tested sera from 214 patients with this assay, IgM antibodies were detected in 100% of patients with acute type A, but not with type B or non-A, non-B hepatitis. IgM antibodies were also found in highest frequency (76%) and titer in patients with autoimmune chronic active hepatitis (CAH) among chronic liver disease groups. IgG antibodies occurred in over 50% of patients with acute type A hepatitis, type B chronic active liver disease (CALD), and autoimmune CAH. IgA antibodies were present in 43% of the patients with alcoholic liver disease, but were also seen in other patient groups. When freshly isolated rat hepatocytes were used as target cells, prevalences and titers similar to those obtained with SK-Hep-1 were found. The levels of serum membrane binding antibody were significantly reduced by the addition of human liver-specific membrane lipoprotein in all patient groups. In particular, IgM antibodies became negative in over 50% of patients with CALD (both type B and non-A, non-B) and autoimmune CAH, whereas in acute hepatitis over 50% lost their positivity for IgG antibody. These results indicate that circulating liver membrane binding autoantibodies are heterogeneous, occurring in hepatitis virus-induced acute and chronic liver disease as well as in autoimmune CAH.     

Clinical aspects of non-A, non-B hepatitis infection

Although non-A, non-B hepatitis is usually a mild subclinical illness, 40% of cases of fulminant viral hepatitis are attributed to infection by this agent. The administration of coagulation factor IX concentrates before liver biopsy in 17 patients with chronic liver disease was followed by the development of hepatitis in four, which proved fatal in three cases. The diagnosis was confirmed by transmission in chimpanzees, and further studies demonstrated the existence of two types of non-A, non-B hepatitis with different incubation periods and specific ultrastructural changes in the hepatocytes. The progression of 40% of cases of acute viral hepatitis to chronic liver disease and the development of chronic liver disease in renal and hepatic transplant recipients is very disturbing. It is likely that this type of hepatitis is an aetiological factor in some cases of hepatitis B surface antigen-negative chronic active hepatitis.     

In vitro effect of lymphoblastoid alpha-interferon on subpopulations of effector cells mediating cytotoxicity for autologous hepatocytes in hepatitis B and non-A, non-B

Controlled clinical trials are currently under way to assess the efficacy of interferon (IFN) in hepatitis B virus (HBV) infection. In the present study, T-enriched and non-T-enriched lymphocytes from six patients with acute and six patients with chronic HBV infection were cocultured with autologous liver cells with and without alpha-IFN (lymphoblastoid) at a concentration of 1000 U/ml of culture medium, in an 18 h cytotoxicity assay. IFN produced a significant enhancement of non-T-cell cytotoxicity in patients with acute and chronic HBV infection, from 34.3 +/- 19.6% to 60.0 +/- 11.2%, P less than 0.03, and from 41.2 +/- 17.2% to 65.5 +/- 9.8%, P less than 0.01, respectively. In contrast, no significant effect was observed on T cell-mediated cytotoxicity in either group of patients. The in vitro effect of alpha-IFN was also evaluated in four patients who developed chronic non-A, non-B hepatitis following blood transfusion, but no significant stimulatory effect was noted on either T- or non-T cell cytotoxicity. Similarly, no significant increase was observed in control subjects. The significant enhancement of non-T cell cytotoxicity, but not of T-cell cytotoxicity, for autologous hepatocytes in HBV infection suggests that alpha-IFN produces a selective stimulatory effect on non-T cells. The absence of a similar effect in patients with chronic non-A, non-B hepatitis and control subjects suggests that HBV infection alters the susceptibility of hepatocytes to IFN-stimulated non-T cell damage.     

Acute sporadic hepatitis E in children living in Cairo, Egypt.

Seventy-three pediatric patients with acute hepatitis and 19 control patients without liver disease living in Cairo, Egypt, were evaluated with a newly developed Western blot assay for IgM antibody to hepatitis E virus (IgM anti-HEV). The mean age of acute hepatitis patients was 6.4 years (range, 1-13 years); 56% were male. Among the 73 acute cases, hepatitis A was diagnosed in 30 (41%), possible acute hepatitis B in three (4%), hepatitis E in nine (12%), and by exclusion, non-A, non-B hepatitis in 29 (40%). Two additional acute cases were positive for both IgM anti-HAV and IgM anti-HEV. None of the 19 control subjects had IgM anti-HEV. Parenteral risk factors were associated with cases of non-A, non-B hepatitis but were not associated with acute hepatitis E. Contact with a family member with jaundice was associated with acute hepatitis A. In contrast to prior epidemics of enterically-transmitted non-A, non-B hepatitis, HEV was found to be a common cause of acute hepatitis in a pediatric population. This study provides additional evidence that HEV may be a frequent cause of acute sporadic hepatitis among children living in some developing countries.     

Etiology and outcome of acute viral hepatitis in Korean adults

One hundred and sixteen Korean adults with biopsy-proven acute viral hepatitis were studied to determine the etiology and the outcome of the disease using paired sera obtained during acute and convalescent phases. The prevalence of acute viral hepatitis A, B, D and non-A non-B were 3.4%, 60.3%, 0.9% and 35.3%, respectively: hepatitis B virus infection was the most common cause and the hepatitis D virus superinfection was almost negligible. Only eleven (26.8%) of 41 patients with AVH NANB were negative for all serological markers of HBV. The rest (73.2%) were positive for at least one HBV marker: HBsAg was positive in 31.7%. Therefore, the presence of HBV serologic markers in the sera does not exclude the diagnosis of AVH NANB in Korea. In patients with acute viral hepatitis B, 27% remained positive for HBsAg. Chronic hepatitis developed in 12.8% and 17% patients with acute hepatitis B and non-A non-B, respectively. Progression to chronic hepatitis in patients with acute viral hepatitis B and non-A non-B occurred more commonly, although statistically not significant, in male sex and in patients who did not have clinical jaundice during the acute phase and who showed bridging necrosis in their liver biopsies. Age did not influence the progression to chronic hepatitis.