Authentic Patient-Derived Hepatitis C Virus Infects and Productively Replicates in Primary CD4 + and CD8 + T Lymphocytes In Vitro

Abstract

ABSTRACT Accumulated evidence indicates that immune cells can support the replication of hepatitis C virus (HCV) in infected patients and in culture. However, there is a scarcity of data on the degree to which individual immune cell types support HCV propagation and on characteristics of virus assembly. We investigated the ability of authentic, patient-derived HCV to infect in vitro two closely related but functionally distinct immune cell types, CD4 + and CD8 + T lymphocytes, and assessed the properties of the virus produced by these cells. The HCV replication system in intermittently mitogen-stimulated T cells was adapted to infect primary human CD4 + or CD8 + T lymphocytes. HCV replicated in both cell types although at significantly higher levels in CD4 + than in CD8 + T cells. Thus, the HCV RNA replicative (negative) strand was detected in CD4 + and CD8 + cells at estimated mean levels ± standard errors of the means of 6.7 × 10 2 ± 3.8 × 10 2 and 1.2 × 10 2 ± 0.8 × 10 2 copies/μg RNA, respectively ( P < 0.0001). Intracellular HCV NS5a and/or core proteins were identified in 0.9% of CD4 + and in 1.2% of CD8 + T cells. Double staining for NS5a and T cell type-specific markers confirmed that transcriptionally competent virus replicated in both cell types. Furthermore, an HCV-specific protease inhibitor, telaprevir, inhibited infection in both CD4 + and CD8 + cells. The emergence of unique HCV variants and the release of HCV RNA-reactive particles with biophysical properties different from those of virions in plasma inocula suggested that distinct viral particles were assembled, and therefore, they may contribute to the pool of circulating virus in infected patients. IMPORTANCE Although the liver is the main site of HCV replication, infection of the immune system is an intrinsic characteristic of this virus independent of whether infection is symptomatic or clinically silent. Many fundamental aspects of HCV lymphotropism remain uncertain, including the degree to which different immune cells support infection and contribute to virus diversity. We show that authentic, patient-derived HCV productively replicates in vitro in two closely related but functionally distinct types of T lymphocytes, CD4 + and CD8 + cells. The display of viral proteins and unique variants, the production of virions with biophysical properties distinct from those in plasma serving as inocula, and inhibition of replication by an antiviral agent led us to ascertain that both T cell subtypes supported virus propagation. Infection of CD4 + and CD8 + T cells, which are central to adaptive antiviral immune responses, can directly affect HCV clearance, favor virus persistence, and decisively influence the development and progression of hepatitis C.