Equilibrium centrifugation studies of hepatitis C virus: evidence for circulating immune complexes

Hijikata, Makoto; Shimizu, Yohko K.; Katō, Hirokazu; Iwamoto, Aikichi; Shih, Jennifer; Alter, Harvey J.; Purcell, Robert H.; Yasuda, Hiroshi

doi:10.1128/jvi.67.4.1953-1958.1993

Cited by 343 publications

(166 citation statements)

References 21 publications

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“…We selected a donor blood sample that had been shown to be infectious to both human and chimpanzees. [20][21][22][23] This plasma was obtained from a healthy blood donor who was implicated in the transmission of HCV. The donor blood was later found to be positive for anti-HCV antibody.…”

Section: Methodsmentioning

confidence: 99%

“…18,19 To construct a complete cDNA clone of the HCV genome, we selected a blood donor sample which was shown to be infectious to both humans and chimpanzees. [20][21][22][23] From that sample we have cloned several fragmental cDNAs and constructed a full-length cDNA containing the 98-base pair (bp)-3ЈX sequence. We also compared the sequence in the hypervariable region (HVR) of HCV in the donor from whom the cDNA was derived with those derived from the recipient and the experimentally infected chimpanzee.…”

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confidence: 99%

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Full-length complementary DNA of hepatitis C virus genome from an infectious blood sample

et al. 1998

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We constructed a full-length complementary DNA (cDNA) clone of hepatitis C virus (HCV) from a blood sample of an HCV carrier. The blood from the carrier was eventually transfused to a patient who later developed typical posttransfusion hepatitis C. It was also shown to be infectious to chimpanzees. We obtained 12 overlapping cDNA fragments altogether, covering the entire HCV genome. By subcloning and sequencing, clones considered to constitute the major population were selected. We could also detect 98 base pairs of extra sequences at the 3Ј end of the genome. After confirming the overlapping sequences, we combined the fragments to make a full-length cDNA. The HCV population in the donor was heterogeneous, as determined by their nucleotide sequences of the hypervariable region in envelope protein, but a few virus clones were selected in the recipient after transmission. The similar convergence of the virus population was previously observed when the same blood sample was injected into a chimpanzee. Interestingly, virus clones isolated during the acute phase in the recipient and the chimpanzee had sequences in the hypervariable region identical to that of the full-length cDNA clone. The full-length cDNA clone of HCV constructed in this study may originate from infectious virus clones.(HEPATOLOGY 1998;27:621-627.)Comparative study of the amino acid sequence of hepatitis C virus (HCV) with those of flaviviruses and pestiviruses and gene expression experiments in bacteria, yeast, and animal cells have revealed that the proteins of HCV are processed by a host-derived signalase and cleaved by virus-coded proteases. [1][2][3][4][5] In vitro culture systems that support partial replication of this virus have been developed from human T-and B-cell lines, 6,7 human fetal hepatocytes, 8 chimpanzee hepatocytes, 9 and human primary hepatocytes. 10 However, fully efficient, long-term viral replication has not yet been established. To study the biology of HCV and its pathogenesis, it is essential to establish an efficient in vitro cell culture system or an infectious complementary DNA (cDNA) clone to support the complete replication of HCV.As a first step to construct an infectious cDNA clone of HCV, it is important to select an adequate sample containing infectious HCV. To date, a considerable number of ''complete'' clones of the HCV genome have been reported. However, it is not certain that those clones have truly originated from infectious HCV, because the materials used usually were pooled plasma samples. Furthermore, the plasma of HCV carriers or patients is generally composed of quasispecies of HCV population. 11 To construct an infectious cDNA clone of RNA viruses, it is crucial to retain both 5Ј and 3Ј ends of the sequence, which are highly conserved and are considered to play essential roles for RNA synthesis, transcription, and translation. 12 Many have reported that the 3Ј untranslated region (UTR) of HCV consisted of poly(U) [13][14][15][16] or poly(A) 17 homopolymer tracts. A novel 98-nucleotide (nt) sequence down...

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Section: Methodsmentioning

confidence: 99%

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Full-length complementary DNA of hepatitis C virus genome from an infectious blood sample

et al. 1998

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“…Viral RNA is detected in fractions ranging from 1.03 to 1.25 gAEmL )1 in a sucrose gradient. Low-density HCV particles are associated with either low-or verylow-density lipoproteins [24][25][26][27][28] and have been shown by assays of chimpanzee model to be the most infectious fraction [29,30]. HCV particles of higher densities correspond to particles associated with immunoglobulin, free particles [28,30,31] or free nucleocapsids [32].…”

Section: Infection Of Primary Cell Cultures and Cell Linesmentioning

confidence: 99%

Recent contributions of in vitro models to our understanding of hepatitis C virus life cycle

et al. 2007

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“…The binding of immunoglobulins to lipoprotein-HCV complexes further affects the density of these particles [19,23,26]. For most HCV-positive sera, the majority of HCV RNA banded at buoyant densities of about 1.03-1.08 gAEmL )1 and 1.17-1.25 gAEmL )1 , which represent densities of VLDL ⁄ LDL and lipoprotein-free particles, respectively [12,[18][19][20][21][22]. Occasionally, a third population of HCV RNA-containing material was observed at a medium density of about 1.13-1.16 gAEmL )1 [15,27].…”

Section: Physicochemical Properties Of Hcvmentioning

confidence: 99%

“…Two studies analyzed the correlation between the buoyant density of HCV RNA-containing material and infectivity in chimpanzees [20,30]. Bradley et al [30] separated infected human serum into five fractions by density centrifugation and determined the infectious titer of each fraction by injecting chimpanzees with 10fold serial dilutions of the fractions.…”

Section: Correlation Of Infectivity and Lipoprotein Association Of Hcvmentioning

confidence: 99%

How does hepatitis C virus enter cells?

Diedrich

2006

The FEBS Journal

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The World Health Organization estimates that 170 million people, 3% of the world population, are infected with hepatitis C virus (HCV) [1]. The majority of those infected (55-85%) fail to clear the virus and become chronic carriers manifested by the persistent presence of detectable virus in the serum [2]. The clinical course of chronic hepatitis C is highly variable ranging from mild hepatitis (inflammation of the liver), fibrosis (scaring of the liver), cirrhosis (end-stage fibrosis) to hepatocellular carcinoma (liver cancer). Liver damage is not directly caused by the virus, but by the interplay between the virus and the immune system that results in the replacement of healthy liver tissue with fibrous scar tissue. About 20% of patients with chronic hepatitis C will develop liver cirrhosis within 20 years. Once cirrhosis is established, the rate of hepatocellular cancer development is 1-4% per year [3]. The standard treatment for chronic HCV infection is pegylated a-interferon in combination with the nucleoside analogue ribavirin. About 55% of patients respond to the therapy and show a sustained reduction in viral titer [4]. Few treatment options exist for nonresponders. Ribavarin and a-interferon have general antiviral properties not specifically related to HCV. Drugs interfering specifically with HCV RNA replication or translation and processing of HCV proteins are not available yet, but a few promising candidates are in clinical testing [5,6].Since the discovery of HCV in 1989, the major bottleneck in HCV research has been the lack of a robust and reliable cell culture system for the propagation of the virus, and the absence of a nonprimate animal model. While cultured liver cells can be infected with clinical HCV isolates, the process has been inefficient, transient and not always reproducible [7]. Our current knowledge about the mechanism of viral cell entry comes from several different approaches including vaccination of chimpanzees, structural studies of Hepatitis C virus (HCV) exists in different forms in the circulation of infected people: lipoprotein bound and lipoprotein free, enveloped and nonenveloped. Viral particles with the highest infectivity are associated with lipoproteins, whereas lipoprotein-free virions are poorly infectious. The detection of HCV's envelope proteins E1 and E2 in lipoprotein-associated virions has been challenging. Because lipoproteins are readily endocytosed, some forms of HCV might utilize their association with lipoproteins rather than E1 and E2 for cell attachment and internalization. However, vaccination of chimpanzees with recombinant envelope proteins protected the animals from hepatitis C infection, suggesting an important role for E1 and E2 in cell entry. It seems possible that different forms of HCV use different receptors to attach to and enter cells. The putative receptors and the assays used for their validation are discussed in this review.

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Equilibrium centrifugation studies of hepatitis C virus: evidence for circulating immune complexes

Cited by 343 publications

References 21 publications

Full-length complementary DNA of hepatitis C virus genome from an infectious blood sample

Full-length complementary DNA of hepatitis C virus genome from an infectious blood sample

Recent contributions of in vitro models to our understanding of hepatitis C virus life cycle

How does hepatitis C virus enter cells?

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