Behavior of theophylline-sensitive T lymphocytes in viral A, B, non-A, non-B hepatitis. I. Methodological aspects

To evaluate the behaviour of the Theophylline-sensitive T lymphocytes subpopulation some modifications of the standard procedure are proposed. Lymphoprep purified lymphocytes were counted in a Neubauer hemocytometer after Acridine Orange stain, viability was evaluated by Ethidium Bromide counterstain and monocytes contamination was evaluated by the peroxidase stain. Sheep red blood cells were treated with AET, Theophylline was used at 3 mM (final concentration) and the results compared with untreated lymphocytes; the enumeration of the rosetting lymphocytes was facilitated by adding Acridine Orange prior to the resuspension. The modifications described were able to increase the % of rosetting T lymphocytes, to eliminate differences depending by different lots of sheep red blood cells and to decrease differences depending by subjective evaluation of the rosetting T lymphocytes.     

Epidemic focus of non-A, non-B viral hepatitis in a plasmapheresis unit]

Non-A, non-B hepatitis has been diagnosed in 12 blood donors in a plasmapheresis unit. The course of the disease has been symptomatic, accompanied by jaundice, fatigue, and nausea in 8 cases, and subclinical in the remaining 4 patients. Nine patients were followed-up to 2 years and only 2 patients liver biochemical tests were normalized permanently. The biopsies performed, a year after the acute phase of hepatitis period revealed chronic active disease in patients, chronic persistent hepatitis in 2 patients, acute hepatitis in one, and normal liver in one patient. Repeated liver biopsies, performed one year later, have basically shown similar lesions except one patient in whom chronic active hepatitis progressed to incipient liver cirrhosis. No symptoms of the disease have been usually noted in patients with chronic form of the disease, and liver function tests have occasionally been normal.     

Incidence of hepatitis non-A,non-B compared with types A and B in hospital patients.

To establish the relative frequencies of types A, B and non-A, non-B hepatitis, stored samples of blood from all the cases of acute viral hepatitis seen over a period of 9 years in a general hospital for adults were classified according to their type by presently available serologic methods. The study included 456 episodes of hepatitis in 447 patients, distributed as follows: 114 episodes of hepatitis A (25%), 282 of hepatitis B (62%) and 60 of hepatitis non-A, non-B (13%). The episodes of non-A, non-B hepatitis were equally distributed between the sexes, suggesting a mode of transmission different from that of hepatitis A or B, which had male/female ratios of 2.4 and 3.1 respectively. The low proportion of hepatitis non-A, non-B may not reflect its real frequency, since it often escapes clinical recognition.     

A cDNA clone closely associated with non-A, non-B hepatitis.

A lambda gt11 cDNA library was constructed from RNA purified from hepatitis B viral surface antigen-negative human plasma with high alanine aminotransferase activity. A cDNA clone, designated as C8-2, was isolated by immunoscreening with mixed sera from non-A, non-B hepatitis (NANBH) carrier and convalescent chimpanzees. The recombinant protein produced by C8-2 reacted specifically with sera of patients in the chronic phase of NANBH. The sequence of C8-2, 269 bp, did not hybridized with any human or chimpanzee genomic DNA, and had no homology with those of primates and viruses. The existence of this sequence in RNA of possibly infectious plasma was shown by RNA blot hybridization and by Southern blot analysis of products amplified by the polymerase chain reaction. These results strongly suggest that C8-2 is derived from the agent of this viral hepatitis.     

The epidemiological patterns of hepatitis non-A, non-B

The etiological structure of viral hepatitides in three cities of the Volga region was studied. The proportion of hepatitis non A, non B was established: it varied from 3.6% to 18.1% at different territories and depended on the total morbidity level, on the season, as well as on the age of patients. A suggestion was made concerning the role of the parenteral mechanism of the transfer of hepatitis non A, non B at the territory under study.     

Primate animal models and titered inocula for the study of human hepatitis A, hepatitis B, and non-A, non-B hepatitis

Although many primate species have been inoculated with the agents of human hepatitis A, B, and non-A, non-B, only a small number of species have been shown to be susceptible, and only the chimpanzee (Pan troglodytes) has been shown to be reproducibly susceptible to all three types of human hepatitis. Infectious inocula containing each agent have been identified in different laboratories and the end-point titers of infectivity determined, in most cases by inoculation of chimpanzees. These inocula and the nonhuman primate models have permitted investigators to study the inactivation of these agents and to evaluate passive and active immunization against the agents.     

The epidemiological problems of viral non-A, non-B hepatitis with a fecal-oral mechanism of infection transmission

The results of the study of the epidemiological regularities of viral hepatitis non-A, non-B with the fecal-oral mechanism of the transfer of infection in one of the regions of the Turkmenian SSR are presented. These results have been confirmed by serological and virological laboratory methods. The territories, groups of the population (age groups) and the time of the maximum epidemic risk of infection have been determined. The main (primary) role of the water route (polluted ground water) in the spread of this infection has been proved.     

Lymphocyte and neutrophil dysfunction associated with hepatitis B virus and hepatitis non-A, non-B virus infection in the chimpanzee.

Chimpanzees were examined for the effect of viral hepatitis infections on specific and nonspecific immune response mechanisms. The data suggest that infection with either hepatitis B virus or hepatitis non-A, non-B virus may result in suppression of cellular immune response components. Mitogen-induced lymphocyte proliferation was lower in virus-infected chimpanzees than in naive animals. Neutrophils from virus infected animals exhibited decreased or altered chemiluminescence kinetics.     

Cloning and expression of cDNAs from enterically-transmitted non-A, non-B hepatitis virus.

The fragment gene of enterically-transmitted non-A, non-B hepatitis virus (ET-NANBHV) was cloned as a cDNA and inserted into an expression vector pUEX2. The recombinant protein was expressed in Escherichia coli HB101 as a fusion protein with beta-galactosidase (beta-Gal). The fusion protein reacted with the sera of infected cynomolgus monkeys and of patients from Myanmar. This reaction was highly related with ET-NANBHV infection, and obviously demonstrates in that the recombinant protein can be used for the detection of ET-NANBHV infection.     

IgM anti-hepatitis C virus in patients with chronic non-A, non-B hepatitis and their relationship to viral replication

Patients with hepatitis C virus (HCV) infection may have different patterns of antibody response to various structural and non-structural viral antigens. We have correlated the serological patterns to the clinical features of chronic infection and to viral replication in 68 HCV-Ab-positive patients with chronic liver disease at different stages (19 with cirrhosis-hepatocellular carcinoma, 38 with chronic active hepatitis and 11 with chronic persistent hepatitis). Serum samples from each patient were assayed for HCV-IgM by enzyme immunoassay and for HCV-RNA by the polymerase chain reaction using primer sets derived from the 5'-non-coding region. The prevalence of HCV-IgM was high (54 patients (79.4%)) and the study showed a good correlation between high values of anti-HCV-IgM and the presence of HCV-RNA in serum, since HCV-RNA was detected in 35 of the 54 IgM-positive patients (64.8%) and notably in 19 of the 20 subjects with high levels of specific IgM. Conversely, all the 35 sera containing HCV-RNA were also reactive for HCV-IgM, while none of the HCV-IgM-negative sera was HCV-RNA reactive. Positivity rates for both HCV-RNA and IgM anti-HCV were higher in the more advanced stages of disease; thus, the clinical pattern of HCV chronic hepatitis seems to be strictly related to the serological pattern and the presence of HCV-RNA.     

Association of human hepatocellular membrane fusions with non-A, non-B hepatitis

Liver biopsies from patients with alcoholic hepatitis, chemical hepatitis, or viral hepatitis types A, B, or non-A, non-B were examined by electron microscopy. Circular, fused, cytoplasmic membranes were observed in hepatocytes of 17% of patients with hepatitis type B and 92% of patients with hepatitis type non-A, non-B. The membrane alterations were not observed in hepatocytes of patients with the other types of hepatitis. The greater frequency of altered cytoplasmic membranes in hepatocytes of patients with non-A, non-B hepatitis was shown to be statistically significant (p less than 0.05) when compared to that in patients with viral hepatitis type B.     

Transmission of non-A, non-B hepatitis agent to chimpanzees from patients of epidemic hepatitis

Two chimpanzees were inoculated intravenously with acute-phase sera obtained from two patients with epidemic hepatitis. They developed histopathologically confirmed hepatitis. Electron microscopic examination of the liver showed peculiar cytoplasmic tubular structures in the hepatocytes. These ultrastructural findings were similar to those described for the livers of chimpanzees inoculated with the F strain of non-A, non-B hepatitis agent derived from a posttransfusion hepatitis case. The chimpanzee that had recovered from hepatitis caused by the F strain of non-A, non-B hepatitis agent was re-challenged with the serum from one of the patients. The chimpanzee developed neither clinical signs nor histological changes of hepatitis. These results suggested that non-A, non-B hepatitis agent was involved not only in post-transfusion hepatitis but also in epidemic hepatitis.