The pivotal role of infected hepatocytes in determining the outcome of antiviral CD8 T cell immunity in the liver

• Applicant: Privatdozent Dr. Dirk Wohlleber
• Subject Area: Immunology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 452431680

The priming and effector function of antiviral CD8 T cells are thoroughly characterized and understood. Whereas much is known how effector CD8 T cells migrate into infected organs recognizing virus-infected cells, less is known about the function of the infected target cell during CD8 T cell effector function. We hypothesize that viral infection of the liver sensitizes hepatocytes towards enhanced cell death induction. Sensitization of infected hepatocytes leads to more efficient CD8 T cell effector function with less effector cells preventing excessive immunopathology. To prove this hypothesis, we aim to establish in vitro assays where primary murine and human hepatocytes are virus-infected and challenged with single CTL effector molecules (e.g. TNF, Fas, Perforin/GranzymeB). Thereby, an enhanced sensitivity of virus-infected hepatocytes towards death inducing molecules shall be demonstrated. Furthermore, we will make use of a co-culture system of primary murine hepatocytes and effector CD8 T cells derived from transgenic mice for single T cell receptor specificities. By uncoupling antigen-presentation through peptide loading of hepatocytes and sensitization by viral infection, we will demonstrate that sensitization enables target cell elimination even at low peptide concentrations, which otherwise would lead to escape from CTL effector function. At the same time, we will estimate to what extent sensitization improves antiviral immunity by calculating numbers of CTLs necessary for elimination of given amount of target cells and time required for elimination of target cells with and without sensitization of target cells by viral infection. In parallel, we will identify in vivo if viral infection sensitizes hepatocytes towards TNF receptor superfamily ligands. Therefore, virus-infected mice will be will be treated by death inducing molecules identified before in vitro and monitored for liver damage and antiviral effects of ligands. In addition, a sophisticated in vivo system will be established, where we will uncouple antigen presentation by hepatocytes from sensitization through viral infection in the living organism. The effect of viral sensitization will be quantified by measuring elimination of antigen-presenting hepatocytes in real-time using non-invasive in vivo bioluminescence imaging and titration of antigen-specific effector CD8 T cells. We will demonstrate that target cell elimination requires less effector cells upon sensitization. In addition, we will be able to demonstrate that target cell sensitization is a naturally evolved mechanism to overcome low antigen-presentation and to protect healthy tissue. In summary, this project will identify the target cells decisive for an effective antiviral immunity. This will expand the current knowledge of antiviral immunity and change the view from a strict T cell-centered more towards to a target cell-centered role during elimination of viral infection.

• DFG Programme: Research Grants