Immunosurveillance and Immunopathology Core

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Project Summary The Immunosurveillance and Immunopathology Core (IIC) by performing mechanistic studies on samples from the 3 projects of this application will serve a synergistic scientific role with strong emphasis on transplant tolerance, the central focus of this Program. The IIC will perform assays to analyze the immune alterations in peripheral circulation, graft and spleen induced by the tolerance regimens proposed in Projects 1, 2 and 3. The Transplantation Tolerance Laboratory (TTL) has developed a series of Standard Operating Procedures (SOPs) to analyze the donor-specific immune responses in NHP transplant recipients that will be shared across all the projects to harmonize the analyses proposed across all the projects in the U19 program. The Core has optimized, validated, and adopted several cutting-edge technologies and pipelines for bioinformatic data analysis of this complex dataset across multiple tissue compartments. Comprehensive immune analyses of samples from the 3 projects by the IIC will aid in avoidance of assay and analysis variations resulting in harmonization of results that will aid in insights obtained in one model to be synergistically deployed across other projects. IIC will perform assays outlined in the four Specific Aims presented below and conduct bioinformatics and data analysis pipeline to assess the immune alterations induced by the distinct tolerance regimens utilized in the Projects 1, 2 and 3. Aim 1: Define the alterations in the immune landscape of renal transplant recipients by high-dimensional CyTOF profiling of circulating immune cells during induction and maintenance of tolerance. Aim 2: Generate and validate TCR signatures using single cell TCRseq to track donor-antigen specific CD4+ and CD8+ T cells. Aim 3: Define the fate and function of donor-antigen specific CD4+ T cells with MHC class II tetramers and single cell ATAC and RNA sequencing in serial peripheral blood mononuclear cells of tolerant and rejecting renal allograft recipients. Aim 4: Analyze the spatio-temporal organization of immune cells in renal allografts, spleen, and lymph nodes at the transcription level by Digital Spatial Profiling and at the protein level by MIBI-TOF. Analysis of multiple tissue compartments longitudinally to generate a large, but highly controlled, dataset. We intentionally apply this multi-omics approach to advantage parallel, rather than sequential, discrimination of system level analyses of immune mechanisms that contribute to transplant tolerance. Analysis of samples from the 3 projects utilizing distinct tolerance regimens will provide new immunological insights on induction and maintenance of tolerance. Moreover, the performance of assays across projects supports lateral transfer of knowledge to identify commonality, independent of strategy, revealing principal features underlying immune tolerance to advance towards successful clinical application. IIC will interact regularly with the PIs of the 3 Projects and with the administrative core to design, execute, analyze and trouble shoot the studies proposed in this application.