HCV Infection and Innate Immunity

Clinical Center

Investigators

Linked publications, trials & patents

Abstract

Chronic HCV infection remains a global health concern due to its involvement in hepatic and extrahepatic diseases, including B cell non-Hodgkin lymphoma (BNHL). Clinical and epidemiological evidence support a causal role for HCV in BNHL development, although mechanistic insight is lacking. The existence of extrahepatic reservoirs of HCV replication, particularly in PBMCs, remains highly controversial. It is unclear how B cells become dysregulated during chronic HCV infection. 1) We observed that the association of HCV with CD19+ B cells is mediated by the complement system. In addition, using antibodies against cell surface markers, we showed that the binding complex mainly involved CD21 (complement receptor 2), CD19, CD20, and CD81 (Hepatology 2016). 2) Our group has demonstrated that free HCV is opsonized by complement and binds to CR1 on erythrocytes (ECR1); the presence of HCV specific antibody significantly increased this binding (Hepatology 2018). Since B-cells proliferation, in response to antigenic stimulation or polyclonal activation, may predispose to genetic aberrations (mutation, gene translocation, gene fusion, chromosomal amplification or deletion) we used RNA-sequencing, to obtain both mutational and gene expression data from B-cells obtained from patients who are HCV+, HBV+, HDV+ and B-NHL+, and compare to patients who are HCV-, HBV-, HDV-, NHL+; HCV+, HBV+, HDV+, NHL-, and healthy controls. As recipient of an NIAID-NCI grant: "Investigation of HCV and HBV-Mediated B-Cell Malignant Transformation and Prioritization of Treatment for HCV-Related NHL in Mongolia, we have received more than 200 samples from Mongolia, including PBMC, plasma, and matched paraffin embedded lymphatic tissues, and have performed RNA-sequencing on 75 peripheral B cell samples isolated from PBMCs. 1) Peripheral B cells from HCV-only patients demonstrated a gene signature consistent with chronic viral infection that indicated elevated Type I interferon signaling and general immunosuppression. This pattern was not observed in B cells from patients with HCV-associated BNHL. In these patients, we observed enrichment of an anergic-like gene signature mirroring that observed in autoimmune disorders and other HCV-associated lymphoproliferative disorders. Additionally, we observed significant clonal enrichment in HCV-associated BNHL samples of immunoglobulin genes known to be associated with autoimmunity and lymphoproliferative disorders. Clonal enrichment was correlated with expression of multiple epigenetic regulatory genes, which were found to be overexpressed in BNHL patients with and without concurrent HCV infection. Our data support viral-mediated clonal expansion of anergic-like B cells as a potential contributing factor to HCV-associated BNHL development and suggest epigenetic dysregulation may be a common pathway in viral- and non-viral-associated lympho-progression. In HCV-associated BNHL, epigenetic mechanisms may also be involved in regulation of B cell anergy and represent an attractive target for clinical interventions. 2) To help elucidate the role for HBV and HDV infection in the development of BNHL, we completed the RNA-sequencing was performed on peripheral B cells collected from patients with chronic HBV mono-infection, HBV/HDV co-infection with or without concomitant BNHL, BNHL-only patients, and healthy donors. Our results suggest that dysregulated epigenetic and RNA-mediated gene expression may be an early factor of lymphomagenesis common to both non-viral and viral-mediated BNHL. Additionally, chronic HBV/HDV co-infection and, especially, chronic HBV mono-infection results in altered snoRNA expression with potential impacts on ribosome biogenesis and subunit composition, and this may represent a viral-specific mechanism contributing to lymphoma development (Manuscript Submitted). 3) The detection of HCV replication in B cells would support the hypothesis of direct virus-mediated lymphomagenesis, where the virus could trigger oncogenic events via intracellular viral proteins. We conducted a study to further elucidate these mechanisms by visualizing HCV positive and negative strand RNA in B cells using RNAscope fluorescent in situ hybridization. Preliminary data showed successful visualization of CD19 mRNA and HCV positive-strand RNA. Optimization of the negative-strand HCV RNA probe is ongoing Visualization of this RNA would indicate viral proliferation within B cells, confirming the existence of extrahepatic reservoirs for HCV and providing more direct evidence of viral protein expression in HCV associated lymphoma (Manuscript in progress).