In vitro infection of peripheral blood mononuclear cells by hepatitis C virus

Abstract

To study the in vitro susceptibility of peripheral blood mononuclear cells (PBMC) to hepatitis C virus (HCV), we incubated cells from healthy donors with HCVpositive sera. Using RT-PCR and in situ hybridization, the genomic viral RNA was detected in PBMC and in their supernatants until 25 days post-incubation. The PBMC of the different donors were not all permissive to HCV, but results were more constantly positive when cells from four donors were pooled. Quantification of the genomic viral RNA by the branched-DNA assay showed a decrease in the HCV RNA concentration during the first week of culture followed by a peak during the second or third week, and also an increase in the total amount of viral RNA in the inoculated cells.Although HCV RNA could be detected in the supernatants by RT-PCR, the concentration was very low. Using a sense-specific RT-PCR method, the HCV negative-strand was also detected in the cells but not in the supernatants. In two experiments PBMC were successfully infected using HCV-positive culture supernatants, therefore suggesting that infectious particles can be produced in this system. Our findings demonstrate that PBMC are permissive for HCV replication in vitro but the replication level is very low. The HCV RNA concentration measured in PBMC of 10 chronically infected patients was not significantly higher than the maximal concentration obtained in PBMC infected in vitro.

Nucleotide and Amino Acid Complexity of Hepatitis C Virus Quasispecies in Serum and Liver

“…All these data, the phylogenetic clustering of serum and liver sequences (in 37 of 39 patients; Fig. 1) and the finding that virus level was correlated between both compartments, argue against a significant contribution (in quantitative terms) of extrahepatic HCV replication to the serum (9,43). Alternatively, differences in the clearance rates of some variants might be responsible for the observed differences (19).…”

The genotype of the hepatitis C virus in patients with HCV-related B cell non-Hodgkin’s lymphoma

“…[12][13][14][15][16][17][18][19] HCV might be a causative cofactor in the pathogenesis of B cell NHL: it is found in a significative percentage of some histotypes of NHL, 8,11,33,34 it is known to be able to infect B lymphocytes circulating in the peripheral blood (PB) 35 or infiltrating the bone marrow (BM) 36 and the liver, 37 and in vitro studies have shown its ability to infect a human BM-derived B cell line and the PB MNC from healthy subjects. [38][39][40] If this is the case, it would appear appropriate to assess the prevalence of different HCV genotypes among the patients bearing a B cell NHL, with the aim of finding a possible correlation between a particular genotype and the lympho-proliferative disorder.…”

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

“…Several complementary strategies to investigate extrahepatic replication of HCV include DNA-based expression systems, 41,[49][50][51][52][53] lymphocyte cell lines that constitutively express HCV, 21,23,54 peripheral blood mononuclear cell (PBMC) cultures from HCV-infected persons, 21,55,56 direct viral infection of extrahepatic cell types with HCV-infected sera, 35,[57][58][59][60] in situ hybridization and immunohistochemistry, 38,[61][62][63][64] and laser capture microdissection. 37,65 The relevance of each of these strategies to extrahepatic replication of HCV is not currently known and can only be determined through extensive validation and cross-comparison amongst all available methodologies.…”