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[Preprint]. 2023 Feb 13:2023.02.08.524808. [Version 2] doi: 10.1101/2023.02.08.524808

Single cell transcriptomic analysis of renal allograft rejection reveals novel insights into intragraft TCR clonality

Tiffany Shi, Ashley R Burg, J Timothy Caldwell, Krishna Roskin, Cyd M Castro-Rojas, P Chukwunalu Chukwuma, George I Gray, Sara G Foote, Jesus Alonso, Carla M Cuda, David A Allman, James S Rush, Catherine H Regnier, Grazyna Wieczorek, Rita R Alloway, Adele R Shields, Brian M Baker, E Steve Woodle, David A Hildeman
PMCID: PMC9934650  PMID: 36798151

Abstract

Bulk analysis of renal allograft biopsies (rBx) identified RNA transcripts associated with acute cellular rejection (ACR); however, these lacked cellular context critical to mechanistic understanding. We performed combined single cell RNA transcriptomic and TCRα/β sequencing on rBx from patients with ACR under differing immunosuppression (IS): tacrolimus, iscalimab, and belatacept. TCR analysis revealed a highly restricted CD8 + T cell clonal expansion (CD8 EXP ), independent of HLA mismatch or IS type. Subcloning of TCRα/β cDNAs from CD8 EXP into Jurkat76 cells (TCR -/- ) conferred alloreactivity by mixed lymphocyte reaction. scRNAseq analysis of CD8 EXP revealed effector, memory, and exhausted phenotypes that were influenced by IS type. Successful anti-rejection treatment decreased, but did not eliminate, CD8 EXP , while CD8 EXP were maintained during treatment-refractory rejection. Finally, most rBx-derived CD8 EXP were also observed in matching urine samples. Overall, our data define the clonal CD8 + T cell response to ACR, providing novel insights to improve detection, assessment, and treatment of rejection.

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