Gastroenterology. 2013 Dec;145(6):1404-1413.e10. doi: 10.1053/j.gastro.2013.08.034. Epub 2013 Aug 22.
Department of Medicine, Johns Hopkins University Baltimore, Maryland.
BACKGROUND & AIMS: Hepatitis C virus (HCV) predominantly infects hepatocytes, but many hepatocytes are not infected; studies have shown that HCV antigens cluster within the liver. We investigated spatial distribution and determinants of HCV replication in human liver samples.
METHODS: We analyzed liver samples from 4 patients with chronic HCV infection (genotype 1, Metavir scores 0-1) to estimate the proportion of infected hepatocytes and the amount of HCV viral RNA (vRNA) per cell. Single-cell laser capture microdissection was used to capture more than 1000 hepatocytes in grids, to preserve geometric relationships. HCV vRNA and interferon-induced transmembrane protein 3 (IFITM3) messenger RNA (the transcript of an interferon-stimulated gene) were measured in the same hepatocytes by quantitative polymerase chain reaction and assembled in maps to identify areas of high and low HCV replication.
RESULTS: Patients' serum levels of HCV RNA ranged from 6.87 to 7.40 log10 IU/mL; the proportion of HCV-infected hepatocytes per person ranged from 21% to 45%, and the level of vRNA ranged from 1 to 50 IU/hepatocyte. Infection was not random; we identified clustering of HCV-positive hepatocytes using infected-neighbor analysis (P < .0005) and distance to the kth nearest neighbor compared with random distributions, obtained by bootstrap simulations (P < .02). Hepatocytes that expressed IFITM3 did not appear to cluster and were largely HCV negative.
CONCLUSIONS: We used single-cell laser capture and high-resolution analysis to show that in human liver HCV infects hepatocytes in nonrandom clusters, whereas expression of antiviral molecules is scattered among hepatocytes. These findings show that quantitative single-cell RNA measurements can be used to estimate the abundance of HCV vRNA per infected human hepatocyte and are consistent with cell-cell propagation of infection in the absence of clustered IFITM3.
Copyright © 2013 AGA Institute. Published by Elsevier Inc. All rights reserved.
KEYWORDS: AIDS Linked to Intravenous Experience, ALIVE, HCV, HIV, IFITM3, ISG, Intrahepatic Infection, LCM, PCR, Virology, cDNA, complementary DNA, hepatitis C virus, human immunodeficiency virus, interferon-induced transmembrane protein 3, interferon-stimulated gene, laser-capture microdissection, mRNA, messenger RNA, polymerase chain reaction, qPCR, quantitative polymerase chain reaction, scLCM, single-cell laser-capture microdissection, vRNA, viral RNA
PMID: 23973767 [PubMed - in process]