Mapping HCV Infection in the Liver

Using single-cell laser capture and high-resolution analysis, researchers show that hepatitis C virus (HCV) infects hepatocytes in the human liver in nonrandom clusters, whereas expression of anti-viral molecules is scattered among hepatocytes. The findings are presented in the December issue of Gastroenterology.

HCV predominantly infects hepatocytes, but most hepatocytes in the liver remain uninfected—HCV antigens have been observed to cluster. This suggests a localized mechanism of intra-hepatic propagation and control. Understanding this process could increase our understanding of infection and strategies for treatment.

Abraham J. Kandathil et al. analyzed liver samples from 4 patients with chronic HCV infection to estimate the proportion of infected hepatocytes and the amount of HCV RNA per cell using single-cell laser capture microdissection (LCM).

LCM unites light microscopy with a low-intensity ultraviolet laser, allowing researchers to ensnare enriched cellular material from tissue samples while preserving positional information, because the tissue is not homogenized. Kandathil et al. made improvements to the technique to increase its resolution, developing single-cell LCM, which allowed them to compare host and viral RNAs.

Studying viral replication in liver tissues from patients with chronic HCV infections, Kandathil et al. estimated the amount of HCV RNA per infected hepatocyte, to determine how the virus spreads in vivo and localize host cell expression of antiviral molecules.

The authors used their data to create a map of viral RNA in hepatocytes, which they called the viroscape. The viroscape shows hepatocytes containing narrow HCV replication peaks surrounded by broad regions with minimal or no HCV vRNA that resembled valleys (see video).

HCV viroscape with superimposed IFITM3 landscape. HCV RNA from 1 patient is represented as contoured gray peaks above the xy plane, and IFITM3 mRNA in the same hepatocytes is represented as contoured terrain below the xy plane.

HCV viroscape with superimposed IFITM3 landscape. HCV RNA from 1 patient is represented as contoured gray peaks above the xy plane, and IFITM3 mRNA in the same hepatocytes is represented as contoured terrain below the xy plane.

Kandathil et al. found that the proportion of HCV-infected hepatocytes per person ranged from 21% to 45%, and the level of viral RNA ranged from 1 to 50 IU/hepatocyte. However, infection was not random—the authors saw clusters of HCV-positive hepatocytes. These clusters in the hepatic viroscapes indicate cell-to-cell propagation of infection.

Kandathil et al. characterized the spatial association between intrahepatic HCV replication and innate immune signaling, and found that although expression of interferon-stimulated genes was sporadic, it was not specifically targeted toward or away from HCV-positive hepatocytes.

Clustering of HCV-infected hepatocytes did not appear to be caused by short-range immunologic control. IFITM3, an interferon-λ–induced protein that has direct antiviral effects against HCV in cell culture, did not appear to be directed specifically toward or away from infected hepatocytes (green in video).

In liver tissues from some subjects, the author found an association between the peak of viral RNA in a cluster and the number of cells in the cluster. This might suggest that infected hepatocytes depend on the robustness of viral replication in the hepatocyte most permissive to viral replication. Alternatively, the hepatocyte with the highest viral RNA copy number could have been the earliest infected cell of a cluster.

Cell-to-cell propagation of HCV could have important implications for vaccine design and drug development—strategies to inhibit entry of extracellular virions could be insufficient for HCV control if cell-to-cell spread of infection is rampant.

The authors hope for future studies with expanded viroscapes, so they can analyze expression of other host genes that control or support HCV replication.

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About Kristine Novak, PhD, Science Editor

Dr. Kristine Novak is the science editor for Gastroenterology and Clinical Gastroenterology and Hepatology, both published by the American Gastroenterological Association. She has worked as an editor at biomedical research journals and as a science writer for more than 12 years, covering advances in gastroenterology, hepatology, cancer, immunology, biotechnology, molecular genetics, and clinical trials. She has a PhD in cell biology and an interest in all areas of medical research.
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4 Responses to Mapping HCV Infection in the Liver

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