Project 4: APOB-specific CD4 and CD8 T cells exacerbate atherosclerosis

Project 4 Summary Project 4 in this PPG with 4 projects and 3 cores is focused on the T cell response to apolipoprotein B (ApoB), an atherosclerosis antigen, in humans and mice. The central hypothesis is that the autoimmune response is initially atheroprotective, but becomes pro-atherogenic (switches) as atherosclerosis progresses. We will address this in humans with and without coronary artery disease (Cardiovascular Assessment Virginia [CAVA] and CT-CAVA cohorts) and in newly generated Apoe-/- FoxP3-ERT2-Cre-GFP ROSA26-fl-STOP-fl- RFP lineage tracker mice. We propose 3 specific aims. Aim 1 is to define the ApoB-specific CD4 and CD8 T cells in PBMCs from CAD cases and controls. This includes T cell transcriptomes, TCRα and β sequences, cell surface phenotypes by Ab-Seq, and finding the immunodominant ApoB epitopes to which they respond. The main methods are single cell (sc) RNA-Seq, antibody (Ab)-Seq and T cell receptor (TCR)-Seq. To find the immunodominant epitopes, of key relevance for cell-based and immunomodulatory therapeutics, we will use a megapool of 208 MHC-binding human ApoB peptides, followed by systematic deconvolution tested by Elispot, intracellular cytokine staining (ICS), cytokine capture assays (CCA) and antigen-induced marker (AIM) assays. Aim 2 is to test how ApoB-specific T cells develop by combining the new mouse model with ApoB-specific tetramers. Our preliminary data support the hypothesis that some ApoB-specific CD4 T cells already exist in very young mice. Some of these cells are effector T cells (non-Tregs). Other FoxP3+ CD4+ Tregs in mice switch their transcriptomes from atheroprotective to pro-atherogenic with progression of atherosclerosis. Mechanistically, we will test the metabolic and epigenetic hypotheses of Treg to exTreg switch. Aim 3 is to test the hypothesis that ApoB-specific T cells in humans with cardiovascular disease include Tregs, exTregs and non-Tregs, using scRNA-Seq and deconvolution (CIBERSORT) methods. When the proposed work is done, we will know the dominant MHC-II-restricted epitopes in human ApoB and the polarization of responding T cells by scRNA-Seq, Ab-Seq and TCR-Seq. We will know the mechanism of Treg phenotype switching and the proportion of Tregs, exTregs and other ApoB-specific T cells in PBMCs (CAVA and CT-CAVA cohorts) and endarteriectomy transcriptomes (BIKE cohort).