Hepatitis C virus‐RNAs in plasma and in peripheral blood mononuclear cells of hemophiliacs with chronic hepatitis C: Evidence for viral replication in peripheral blood mononuclear cells

Willems, M; Peerlinck, Kathelijne; Moshage, Han; Deleu, I; Eynde, Christel Van den; Vermylen, Jozef; Yap, Sing Hiem

doi:10.1002/jmv.1890420314

Cited by 93 publications

(64 citation statements)

References 26 publications

(7 reference statements)

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“…In the literature it has been suggested that PBMC can function as an important extrahepatic reservoir and a possible site for HCV replication. The evidence for this assumption was based on the demonstration of negative-strand HCV-RNA in association with PBMC from HCV infected patients by a RT/PCR technique [16][17][18][19] .…”

Section: Discussionmentioning

confidence: 99%

HCV-RNA positivity in peripheral blood mononuclear cells of patients with chronic HCV infection: does it really mean viral replication

Meier

Mihm²,

Braun³

et al. 2001

WJG

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Subject headings hepatitis C like viruses; hepatitis C, chronic; RNA, viral/blood; virus replication; monocytes; interferon alpha/therapeutic use; polymerase chain reaction Abstract AIM To analyze the association of HCV-RNA with peripheral blood mononuclear cells (PBMC) and to answer the question whether HCV-RNA positivity in PBMC is due to viral replication. METHODS HCV-RNA was monitored in serum and PBMC preparations from 15 patients with chronic HCV infection before, during and after an IFN-α α α α α therapy using a nested RT/PCR technique. In a second approach, PBMC from healthy donors were incubated in HCV positive plasma. RESULTS In the IFN-α α α α α responding patients, HCV-RNA disappeared first from total RNA preparations of PBMC and then from serum. In contrast, in relapsing patients, HCV-RNA reappeared first in serum and then in PBMC. A quantitative analysis of the HCV-RNA concentration in serum was performed before and after transition from detectable to non detectable HCV-RNA in PBMC-RNA and vice versa. When HCV-RNA was detectable in PBMC preparations, the HCV concentration in serum was significantly higher than the serum HCV-RNA concentration when HCV-RNA in PBMC was not detectable. Furthermore, at no time during the observation period was HCV specific RNA observed in PBMC, if HCV-RNA in serum was under the detection limit. Incubation of PBMC from healthy donors with several dilutions of HCV positive plasma for two hours showed a concentration dependent PCR positivity for HCV-RNA in reisolated PBMC. CONCLUSION The detectability of HCV-RNA in total RNA from PBMC seems to depend on the HCV concentration in serum. Contamination or passive adsorption by circulating virus could be the reason for detection of HCV-RNA in PBMC preparations of chronically infected patients.

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Section: Discussionmentioning

confidence: 99%

HCV-RNA positivity in peripheral blood mononuclear cells of patients with chronic HCV infection: does it really mean viral replication

Meier

Mihm²,

Braun³

et al. 2001

WJG

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“…One of the hybrid cell clones, IMY-N9, had 98 chro- 6 3 0 10 Ϫ1. 6 3 0 *Necessary HCV-RNA content for 50% tissue culture infection in samples (HCV RNA titer/TCID 50 /mL). †Culture media obtained from HCV in vitro culture system (Ito T, et al 1996).…”

Section: Discussionmentioning

confidence: 99%

Acquisition of susceptibility to hepatitis C virus replication in HepG2 cells by fusion with primary human hepatocytes: Establishment of a quantitative assay for hepatitis C virus infectivity in a cell culture system

et al. 2001

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Hepatitis C virus (HCV) replicates in human and chimpanzee hepatocytes. To characterize the nature of HCV and evaluate antiviral agents, the development of an HCV replication system in a cell culture is essential. We developed a cell line derived from human hepatocytes by fusing them with a hepatoblastoma cell line, HepG2, and obtained several clones. When we tested the clones for their ability to support HCV replication by nested RT-PCR, we found 1 clone (IMY-N9) that was more susceptible to HCV replication than HepG2. The negative-strand HCV RNA was detected in IMY-N9 by strand-specific RT-PCR, and viral RNA was identified in culture supernatant during the culture. Hepatitis C virus (HCV) is now recognized as the principal agent in parentally transmitted non-A, non-B hepatitis. HCV is a single-stranded positive-sense RNA virus of about 9,500 nucleotides (nt) that encodes a large polyprotein, 1,2 which is processed into viral structural and nonstructural proteins. Recently, full-length HCV RNA transcripts from HCV cDNAs were shown to be infectious in chimpanzees. 3,4,5 Although our understanding of the molecular biology of HCV has progressed rapidly, little is known about its biological characteristics, because in vivo studies have been limited to the studies performed using chimpanzees that have been infected or transfected with HCV. The development of efficient animal cell culture systems for HCV or readily available animal models are priorities for the study of HCV. HCV replication has been described based on studies of liver tissue and peripheral blood mononuclear cells (PBMC) from infected patients. 6,7 Animal cell culture systems for HCV replication have been developed from human T and B cell lines, 8,9,10 human fetal liver cells, 11,12 and chimpanzee primary hepatocytes. 13 Efficient long-term viral replication, however, has not been reported in any system. HCV replication may be stringently restricted to differentiated hepatocytes, which may have receptors for HCV on their cell surface and specific factors to support viral growth. Recently, a host cellular factor, polypyrimidine-tract-binding protein (PTB), was reported to bind the HCV 3Ј-end, and was thought to be a cis-acting element for HCV replication. 14,15 PTB is also related to the mechanism of HCV translational regulation, 16,17 indicating that several cellular factors may be involved in the mechanism of HCV propagation. We previously reported that HCV replicated efficiently in primary cultured human hepatocytes, 18 and another group also confirmed our findings. 19 However, we still need to develop a more convenient cell-culture system because it is difficult to obtain a constant source of human hepatocytes. In this study, we fused human hepatocytes with a hepatoblastoma cell line, HepG2, and obtained hybrid cell lines. We then tested the ability of the hybrids to support HCV replication by monitoring HCV RNA titers in cultured cells using a real-time detection PCR (RTD-PCR). 20 The results showed that several hybrid cell lines were more suscept...

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“…29,[67][68][69][70][71][72] Initial studies similarly reported extrahepatic replication of HCV in PBMCs (Table 3). 58,[73][74][75][76][77] However, as the detection assays were not able to adequately differentiate between positive-and negative-strand HCV RNAs, theses results should be interpreted with extreme caution. Subsequent studies have now convincingly demonstrated negative-strand HCV RNA -albeit in a minority of PBMCs tested -particularly among immunosuppressed patients, such as HIV/AIDS patients 68,69,71 and liver transplant recipients.…”

Section: Evidence For Extrahepatic Replication Of Hcvmentioning

confidence: 99%

Extrahepatic replication of HCV: Insights into clinical manifestations and biological consequences

2006

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Hepatitis C virus‐RNAs in plasma and in peripheral blood mononuclear cells of hemophiliacs with chronic hepatitis C: Evidence for viral replication in peripheral blood mononuclear cells

Cited by 93 publications

References 26 publications

HCV-RNA positivity in peripheral blood mononuclear cells of patients with chronic HCV infection: does it really mean viral replication

HCV-RNA positivity in peripheral blood mononuclear cells of patients with chronic HCV infection: does it really mean viral replication

Acquisition of susceptibility to hepatitis C virus replication in HepG2 cells by fusion with primary human hepatocytes: Establishment of a quantitative assay for hepatitis C virus infectivity in a cell culture system

Extrahepatic replication of HCV: Insights into clinical manifestations and biological consequences

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