Visualizing hepatitis C virus infections in human liver by two-photon microscopy

Area of Science:

• Hepatology
• Virology
• Optical Imaging

Background:

• Hepatitis C virus (HCV) infection is a major cause of cirrhosis and liver cancer.
• Understanding HCV pathogenesis is hindered by low viral protein expression in the liver, limiting in situ detection of infected cells.

Purpose of the Study:

• To evaluate advanced optical imaging techniques for determining the extent and distribution of HCV-infected cells in liver tissue.
• To assess the proportion of hepatocytes infected with HCV in patients with chronic infection.

Main Methods:

• Combined 2-photon microscopy with virus-specific, fluorescent quantum dot probes.
• Analyzed frozen liver tissue sections from patients with chronic HCV infection.
• Detected viral core and nonstructural protein 3 antigens, and double-stranded RNA.

Main Results:

• Successfully detected HCV proteins and double-stranded RNA without confounding autofluorescence.
• Confirmed specificity through antibody blocking and antigen colocalization.
• Found 7-20% of hepatocytes infected in patients with high viral RNA loads (≥10^5 IU/mL).
• Observed infected cells in clusters, suggesting cell-to-cell spread.
• Noted abundant double-stranded RNA in central cluster cells, scarce in peripheral cells, indicating recent infection.

Conclusions:

• Two-photon microscopy offers high sensitivity for detecting HCV proteins and replication markers.
• HCV infection involves a limited number of hepatocytes and is a dynamic process.
• Cell-to-cell spread of HCV appears constrained, possibly by host responses.