Failure of the CD8 T cell response in an adenoviral liver infection with high-level antigen expression

Question:

The liver is considered as a lymphoid organ disposing of tolerogenic properties that render it an attractive target site for pathogens like hepatitis viruses. The clinical outcome depends on the kinetics of the virus-host interaction and on the strength of the innate and adaptive immune response. As virus-specific CD8 T cells are known to play a role in viral infections and their dysfunction is observed during infections with high levels of replicating virus combined with high and sustained secretion of viral antigens (i.e. HBV), this project aims to elucidate the mechanisms of the immune system or liver-milieu that lead to the failure in clearing high-level adenoviral liver infection.

Methods:

Our in vivo model system consists of a recombinant, replication-deficient adenovirus encoding for the transgenes eGFP, ovalbumin and CBG99-luciferase under a CMV promotor (Ad-CMV-GOL). Mice have been infected with increasing doses Ad-CMV-GOL and the immune response was characterized by in vivo bioluminescence imaging. Intrahepatic virus-specific CD8 T cells have been characterized by flow cytometry.

Results:

We investigated the outcome of anti-viral immunity after infection with 10⁵ to 10⁹ infectious units (iu) Ad-CMV-GOL and found a dose-dependent outcome of anti-viral immunity. In 10⁵ to 10⁷ iu Ad-CMV-GOL infected mice infection is cleared within 20 days, whereas in 10⁹ iu Ad-CMV-GOL infected mice bioluminescence signal as well as adenoviral genomes are detectable up to 3 months after infection. Although, after high-dose infection, liver damage is detectable by elevated serum ALT levels and virus-specific CD8 T cells expand, we identified them as non-functional. Intrahepatic virus-specific CD8 T cells are characterized by expression of PD-1, Lag-3 and surprisingly by a lack of the T cell receptor. Because of the missing T cell receptor on the cell surface of these CD8 T cells, they could not be re-stimulated with ovalbumin-derived SIINFEKL-peptide. Re-stimulation with PMA/Ionomycin, however, led to production of IFNγ showing the principal ability of these cells for cytotoxicity. Re-isolation of intrahepatic virus-specific CD8 T cells and ex vivo co-cultivation in the absence of antigen restored T cell receptor cell surface expression to normal levels.

Conclusions:

Here, we identify that high-level antigen expression by virus-infected hepatocytes renders anti-viral CD8 T cells non-functional. On the one hand, this non-functional state may be a safety mechanism to prevent liver failure due to overwhelming hepatocyte elimination, on the other hand this leads to persistent liver infection.