Replication In Liver Research Articles

Long-term follow-up of hepatitis G virus/GB virus C replication in liver during and after interferon therapy in patients coinfected with hepatitis C and G viruses.

We investigated changes in the titers of positive and negative strands of hepatitis G virus (HGV) RNA and hepatitis C virus (HCV) RNA in serum and liver during and 1 to 3 years after interferon therapy in patients with chronic hepatitis C coinfected with HGV/ GBV-C. Eight (6%) of 134 patients with chronic hepatitis C treated with interferon were positive for HGV RNA and were examined retrospectively. Titers of positive and negative strands of HGV RNA and HCV RNA were determined by strand-specific reverse transcriptase-polymerase chain reaction. Before therapy, HGV RNA titers in liver were lower than those in serum (P = 0.0169), while HCV RNA titers in liver were significantly higher than those in serum (P = 0.0074). No negative strands of HGV RNA were detected in serum, liver, or peripheral blood mononuclear cells in any patients. With interferon therapy, 5 of the 8 patients lost HCV RNA from serum and liver, with sustained normal liver biochemical values and significant histological improvement. HGV RNA disappeared transiently from serum and liver at the end of therapy in 5 patients, but reappeared again after therapy in 4 of them. Two of the 8 patients naturally lost HGV RNA after completion of therapy. These findings suggest that: (1) HGV/GBV-C does not appear to replicate in liver, serum, or peripheral blood mononuclear cells, (2) detection of HGV RNA in liver and peripheral blood mononuclear cells may be a mere reflection of serum HGV RNA, and (3) the long-term clinical outcome of chronic HGV/GBV-C infection appeared to be benign.

Hepatitis G virus infection in American patients with cryptogenic cirrhosis: no evidence for liver replication.

It is unclear whether hepatitis G virus (HGV) can lead to chronic liver disease and cirrhosis. Eighty-nine patients with end-stage liver disease undergoing liver transplantation were studied: 50 were diagnosed as having cryptogenic cirrhosis while 39 had nonviral chronic liver disease. Five (10%) in the former and 1 (2.6%) in the latter group (not significantly different) were positive for HGV RNA in serum. All 6 HGV-infected patients were negative for the presence of the HGV RNA minus strand in the liver when tested with a strand-specific Tth-based reverse transcription-polymerase chain reaction, and 5 were positive for the presence of the plus strand, albeit at low levels. This implies that the liver is not the primary replication site for HGV, at least in a significant proportion of patients. Absence of liver replication explains the reported lack of association between HGV infection and liver pathology encountered in many clinical settings.

Ribavirin conjugated with lactosaminated poly- l-lysine : Selective delivery to the liver and increased antiviral activity in mice with viral hepatitis

Ribavirin (RIBV) is a useful drug in the treatment of chronic type C hepatitis but displays a toxicity for red blood cells (RBC), which limits its dosage and necessitates withdrawal in some patients. Selective concentration of RIBV in liver should improve therapeutic results. Liver targeting can be achieved by coupling the drug to galactosyl-terminating peptides, which specifically enter hepatocytes. In the present work, we conjugated RIBV to lactosaminated poly- l-lysine ( l-Poly(Lys)), a hepatotropic carrier enabling intramuscular (IM) administration of conjugates. The l-Poly(Lys)-RIBV conjugate had a heavy drug load (312–327 μg of RIBV in 1 mg of conjugate) and was very soluble in 0.9% NaCl (200 mg/mL). The conjugate was devoid of acute toxicity in mouse. When incubated with human or mouse blood, it did not release the drug. After IM administration to mice, the conjugate was selectively taken up by the liver, where the drug was released in a pharmacologically active form. This was demonstrated using mice infected with a strain of murine hepatitis virus (MHV) sensitive to RIBV. Coupled RIBV, IM injected, inhibited MHV replication in liver at a daily dose two to three times lower than that of the free drug. In mice IM injected with a conjugate tritiated in the RIBV moiety, the ratios between the levels of radioactivity in liver and RBC were two times higher than in animals injected with free tritiated RIBV. In conclusion, the present results support the possibility that the chemotherapeutic index of RIBV in chronic type C hepatitis can be increased by conjugation with l-Poly(Lys).

Antibodies to tumor necrosis factor alpha prevent increases in cell replication in liver due to the potent peroxisome proliferator, WY-14,643.

Several structurally dissimilar hypolipidemic drugs, plasticizers and halogenated hydrocarbons induce peroxisomes in hepatocytes, and cause hepatocellular adenoma and carcinoma in rats and mice. The mechanism by which these agents act is unknown, although recent studies have suggested a link between increased cell proliferation and hepatic cancer caused by peroxisome proliferators. Here, we demonstrate that neutralizing antibodies to tumor necrosis factor alpha (TNF alpha) block increases in protein kinase C and cell proliferation due to [4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio]acetic acid (WY-14,643), a hypolipidemic drug and potent peroxisome proliferator that causes tumors. WY-14,643 moderately elevated the level of TNF alpha mRNA in the liver. TNF alpha was detected immunohistochemically exclusively in Kupffer cells. These results demonstrate that WY-14,643 acts as an indirect mitogen on hepatocytes via TNF alpha. We propose that the Kupffer cell, a major source of TNF alpha in the liver, is involved in the mechanism of the mitogenic effect of WY-14,643.

Renal activation of trichloroethene and S-(1,2-dichlorovinyl)-L-cysteine and cell proliferative responses in the kidneys of F344 rats and B6C3F1 mice.

Covalent binding of reactive intermediates formed by renal beta-lyase activation of S-(1,2-dichlorovinyl)-L-cysteine (DCVC) has been suggested to be responsible for the greater renal sensitivity of rats than mice to the carcinogenic effects of chronic treatment with trichloroethene (TRI). Previous work demonstrated that the activation of DCVC results in acid-labile adducts to protein that can be distinguished from adducts formed by other pathways of TRI metabolism. By analyzing acid-labile adduct formation, the relationship between DCVC formation and activation from TRI and increases in rates of cell division in the kidneys of male F344 rats and B6C3F1 mice could be investigated. The delivered dose of DCVC from an oral dose of 1000 mg/kg TRI was approximately six times greater in rats than mice. However, renal activation of DCVC in mice was approximately 12 times greater than in rats. Therefore, the overall activation of TRI was about two times greater in mice than rats. Induction of cell replication in liver and kidney following doses of 1, 5, or 25 mg/kg DCVC or 1000 mg/kg TRI was also measured through the use of miniosmotic pumps that delivered BrdU subcutaneously for 3 d. Acid-labile adduct formation from DCVC and TRI displayed a consistent relationship with increased cell replication in mice and between mice and rats. Both cell replication and acid-labile adduct formation in rats given 25 mg/kg DCVC were approximately equal to that observed in mice given 1 mg/kg. Increased cell replication was not observed in rats receiving 1 or 5 mg/kg DCVC or 1000 mg/kg TRI, nor were there histological signs of nephrotoxicity. Thus, net activation of TRI by the cysteine S-conjugate pathway was found to be greater in mice than rats and these findings appeared related to differences in cell proliferative responses of the kidneys of the two species. Based on these data, it would appear that other factors must contribute to the greater sensitivity of the rat to the induction of renal carcinogenesis by TRI.

Hepatitis C virus replication in liver and peripheral blood mononuclear cells of interferon-alpha-treated and untreated patients with chronic hepatitis C.

Serum, liver tissue, and peripheral blood mononuclear cells (PBMC) were obtained from 72 patients with chronic hepatitis C, 19 of whom had received interferon-alpha (IFN-alpha) treatment. Hepatitis C virus (HCV) RNA was detected by polymerase chain reaction in the serum and liver of all patients with relapse after treatment and in 51 of 53 (96%) sera, 28 of 29 (97%) livers, and all of 13 (100%) PBMC from untreated patients but not in 3 patients with completely sustained responses. Minus-stranded HCV RNA was found in 5 of 14 (35%) sera, 16 of 17 (94%) livers, and 7 of 13 (53%) PBMC from untreated patients. The results indicate that HCV RNA is present not only in serum and liver but also in PBMC from the vast majority of untreated anti-HCV-seropositive patients and also in patients who respond to IFN-alpha treatment but then relapse. HCV replicates in the liver of almost all patients, but replication may possibly also occur in PBMC. By contrast, HCV RNA seems to be eradicated in patients with completely sustained responses.

Expression of pre‐S1, pre‐S2, S and X peptides in relation to viral replication in livers with chronic hepatitis B

The expression of large (pre-S1), middle (pre-S2), major S (S) polypeptides of the envelope (HBs) and X peptides of hepatitis B virus (HBV) was investigated in 37 liver specimens with chronic hepatitis B by indirect immunoperoxidase staining. Primary antisera utilized were polyclonal ones against HBs (poly-HBs), core (HBc) and X and monoclonal ones against pre-S1, pre-S2 and S with (particle-S) or without (peptide-S) conformational structure. The localization of HBs proteins in hepatocytes was classified into three types: diffuse, membranous and inclusion. The peptide-S and pre-S2 were expressed at nearly the same frequency as poly-HBs in all types, whereas particle-S was found less frequently (18/29 cases) in the inclusion type, and pre-S1 was recognized relatively rarely (9/33 cases) in the membranous type. As for staining intensity, peptide-S and pre-S2 were almost identical to poly-HBs which stained the most strongly among all three staining types. Particle-S was similar to poly-HBs in the membranous type, but was weak in the inclusion type in the majority. While pre-S1 was stained in a similar intensity to poly-HBs in the diffuse and inclusion types, it was weak or negative in the membranous type. Thus, envelope particles indicated by particle-S staining appeared to be located most frequently in the membranous type, but their assembly might be suppressed in the inclusion type where pre-S1 was well expressed. The X peptide was more frequently detected in the liver with serum HBe antigen and/or HBV DNA. The X peptide was stained exclusively in the cytoplasm of hepatocytes and was correlated with the cytoplasmic HBc antigen. The X peptide was not observed differently between cases with and those without cirrhosis. This suggests that the expression of X peptide tends to occur with virus replication but not with disease progression.

Site of reovirus replication in liver is determined by the type of hepatocellular insult

Reovirus type 1 strain Lang is restricted from replicating in adult murine livers. In noninjured livers, approximately 1% of hepatocytes express reovirus antigen during infection. However, hepatocytes can be induced to express reovirus antigen if challenged with either toxins or trauma. We used selective hepatotoxins or surgical trauma to demonstrate that reovirus antigen localization in liver is determined by the site of hepatocellular insult and the timing of the virus inoculum.

Repair and replication of DNA in rat brain and liver during foetal and post-natal development, in relation to nitroso-alkyl-urea induced carcinogenesis.

The susceptibility of rat brain to induction of cancer by N-nitroso-N-ethyl-urea (NEU) increases dramatically from a very low level in the 12 day foetus to a maximum at the time of birth, and then decreases with age. Liver tumors are rarely induced by a single treatment with NEU at any age. If induction of cancer by nitroso-alkyl-ureas depends on replication of DNA containing the mispairing base O6-alkylguanine, susceptibility would reflect the balance between the protective effect of removal of the base by repair mechanisms and the potentiating effect of cell replication. The capacity of tissues to remove O6-alkyl-guanine from DNA depends on the amount of alkyl acceptor protein (AAP) present. To study the concept that carcinogenesis results from replication of alkylated DNA, the AAP contents and relative rates of DNA replication were studied in brain and liver of rats at various stages of development, from the 12 day foetus to the 48 week old rat. Replication in liver was approximately ten times higher than in intra-cranial contents at each stage of development studies. The AAP content was higher in the 12 day foetal brain than later, decreasing to low levels as susceptibility to NEU increased until the time of birth, and then remaining low in the adult. The peak sensitivity of brain therefore corresponds to the time at which AAP content is low and rate of DNA replication is high. With liver, AAP levels are low in the foetus, although higher than in brain, and increase after birth. The higher level of AAP in the foetal liver compared with that of brain is possibly sufficient to explain why cancer is not induced in liver in spite of the high rate of DNA replication. The results are consistent with the concept that replication of alkylated DNA is an essential event in initiation. There may be no quantitative relationship between replication and repair and susceptibility to cancer on comparison of different tissues, owing to the fact that, at the cellular level, cancer is a rare event. The amount of mispairing at replication necessary to bring it about may depend on the detailed organisation of the genome, and hence on cell type.

Amino acids and control of nucleolar size, the activity of RNA polymerase I, and DNA synthesis in liver.

The volume of nucleolar material per nucleus and the activity of RNA polymerase I (RNA nucleotidyltransferase I) become doubled in the liver cells of rats that are fed for several days a diet that lacks essential amino acids. Omission of methionine from a fully supplemented diet is equivalent to leaving out all the amino acids, and the responses to a deficiency of tryptophan are about 40% as great. Deprivation of one of the remaining essential amino acids gives either small responses or none at all. Supplementation of the methionine-free diet with cystine blocks the nucleolar enlargement and the enhancement of the polymerase activity that would otherwise take place, but the dispensable amino acid does not affect the responses to a deprivation of one of the other essential amino acids. After deprivation of all the essential amino acids or only methionine, hepatocytes make DNA when the rat is fed a meal with protein. A preparatory diet lacking in tryptophan is much less effective; a deficiency in any of the other indispensable compounds tested fails to prepare the liver for DNA synthesis. The results give hope that elucidation of the means by which methionine deprivation affects the nucleolus will also provide information on the regulation of nuclear DNA replication in liver. One attractive possibility is that the amino acid deficiency acts by producing some imbalance in protein metabolism.

Nucleolar hypertrophy and nuclear DNA replication in liver.

Hypertrophy of nucleoli is associated with DNA synthesis in the hepatocytes of untreated rats just as it is in stimulated animals. Nucleolar enlargement is not a sufficient change to ensure that the parenchymal liver cell can make DNA.

A mouse hepatotropic variant of influenza virus

A hepatotropic variant of avian influenza virus A/Turkey/England 63 (Hav 1, Nav 3) was selected by serial passages in mouse liver. Adaptation to this organ was established after 13 in vivo passages and was found to improve during further passages as shown by increasing rates of replication in livers of ICR mice. The mutant virus finally selected was stable and differed from the original virus mainly in lethality upon intraperitoneal injection in mice, in its ability to grow to high titers in livers of susceptible animals and in plaque morphology in chick embryo fibroblasts. No differences were detected in hemagglutination inhibition and neutralization by standard mouse antisera. Pathogenicity for the liver was independent of the route of inoculation, included other laboratory animals sensitive to influenza virus and could be inhibited by amantadine. Fatal hepatitis in 50 per cent of susceptible mice by the intraperitoneal route required from 10 to 20 EID50-. Pathological changes consisted of severe necrosis of liver parenchyma accompanied by release of F antigen into the serum and were apparently due to virus replication in hepatic cells as evidenced by immunofluorescence. The main implications of this animal model for studies on experimental hepatitis and on myxovirus-host interactions in an organ not usually associated with influenza are discussed.