In Vitro Hepatitis C Virus Infection and Hepatic Choline Metabolism.

Kaelan Gobeil Odai,Morgan D Fullerton,John P Pezacki,Rodney S Russell,Conor O’Dwyer,Tyler A Shaw,Marc-André Langlois,Shauna Han,Mojgan Rezaaifar,Peyman Ghorbani,Rineke Steenbergen,D Lorne Tyrrell,Tyler M Renner,Angela M Crawley

doi:10.3390/v12010108

Abstract

Choline is an essential nutrient required for normal neuronal and muscular development, as well as homeostatic regulation of hepatic metabolism. In the liver, choline is incorporated into the main eukaryotic phospholipid, phosphatidylcholine (PC), and can enter one-carbon metabolism via mitochondrial oxidation. Hepatitis C virus (HCV) is a hepatotropic positive-strand RNA virus that similar to other positive-strand RNA viruses and can impact phospholipid metabolism. In the current study we sought to interrogate if HCV modulates markers of choline metabolism following in vitro infection, while subsequently assessing if the inhibition of choline uptake and metabolism upon concurrent HCV infection alters viral replication and infectivity. Additionally, we assessed whether these parameters were consistent between cells cultured in fetal bovine serum (FBS) or human serum (HS), conditions known to differentially affect in vitro HCV infection. We observed that choline transport in FBS- and HS-cultured Huh7.5 cells is facilitated by the intermediate affinity transporter, choline transporter-like family (CTL). HCV infection in FBS, but not HS-cultured cells diminished CTL1 transcript and protein expression at 24 h post-infection, which was associated with lower choline uptake and lower incorporation of choline into PC. No changes in other transporters were observed and at 96 h post-infection, all differences were normalized. Reciprocally, limiting the availability of choline for PC synthesis by use of a choline uptake inhibitor resulted in increased HCV replication at this early stage (24 h post-infection) in both FBS- and HS-cultured cells. Finally, in chronic infection (96 h post-infection), inhibiting choline uptake and metabolism significantly impaired the production of infectious virions. These results suggest that in addition to a known role of choline kinase, the transport of choline, potentially via CTL1, might also represent an important and regulated process during HCV infection.

Highlights

Hepatitis C virus (HCV) is a positive-strand (+)RNA virus that is inherently primate-borne and principally targets the liver [1]
To establish a kinetic profile of choline uptake in the Huh7.5 cell line cultured in fetal bovine serum (FBS), we first performed uptake experiments using [3 H]-choline and established that transport was linear over the course of 30 min
We demonstrate that short term HCV infection of FBS, but not human serum (HS)-cultured Huh7.5 cells, results in a substantial impairment of choline uptake and incorporation into PC

Summary

Introduction

Hepatitis C virus (HCV) is a positive-strand (+)RNA virus that is inherently primate-borne and principally targets the liver [1]. 71 million people worldwide are infected with HCV, or approximately 1% of the global population [3]. Some cases only lead to mild illness, most infected individuals (75–85%) develop chronic HCV infection. While recent advancements in direct-acting antiviral therapy have proven to be highly efficacious (~98% cure rates), barriers to therapy access, viral resistance, and low diagnosis rates minimizing efforts towards virus elimination, chronic HCV infection remains a significant medical concern. With no widely available vaccine, and culminating in chronic hepatitis, cirrhosis, as well as one of the world’s leading causes of death, hepatocellular carcinoma (HCC) [4], HCV infection has important implications in global health outcomes. Together with non-alcoholic fatty liver disease, HCV infection are the most common indications for a liver transplant in the Western world [3]

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