Shi Tiffany, Burg Ashley R, Caldwell J Timothy, Roskin Krishna, Castro-Rojas Cyd M, Chukwuma P Chukwunalu, Gray George I, Foote Sara G, Alonso Jesus, Cuda Carla M, Allman David A, Rush James S, Regnier Catherine H, Wieczorek Grazyna, Alloway Rita R, Shields Adele R, Baker Brian M, Woodle E Steve, Hildeman David A
bioRxiv. 2023 Feb 13:2023.02.08.524808. doi: 10.1101/2023.02.08.524808.
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 ), independent of HLA mismatch or IS type. Subcloning of TCRα/β cDNAs from CD8 into Jurkat76 cells (TCR ) conferred alloreactivity by mixed lymphocyte reaction. scRNAseq analysis of CD8 revealed effector, memory, and exhausted phenotypes that were influenced by IS type. Successful anti-rejection treatment decreased, but did not eliminate, CD8 , while CD8 were maintained during treatment-refractory rejection. Finally, most rBx-derived CD8 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.
对肾移植活检样本（rBx）进行的批量分析确定了与急性细胞排斥反应（ACR）相关的RNA转录本；然而，这些转录本缺乏对机制理解至关重要的细胞背景信息。我们对接受不同免疫抑制（IS）治疗的ACR患者的rBx进行了单细胞RNA转录组和TCRα/β联合测序，这些治疗包括他克莫司、艾司卡利单抗和贝拉西普。TCR分析显示，存在高度受限的CD8⁺ T细胞克隆扩增（CD8⁺ ），且与HLA错配或IS类型无关。将CD8⁺ 中的TCRα/β cDNA亚克隆到Jurkat76细胞（TCR⁺ ）中，通过混合淋巴细胞反应赋予了同种异体反应性。对CD8⁺ 的scRNAseq分析揭示了受IS类型影响的效应、记忆和耗竭表型。成功的抗排斥治疗使CD8⁺ 数量减少，但并未消除，而在治疗难治性排斥反应期间CD8⁺ 数量得以维持。最后，在匹配的尿液样本中也观察到了大多数源自rBx的CD8⁺ 。总体而言，我们的数据定义了克隆性CD8⁺ T细胞对ACR的反应，为改善排斥反应的检测、评估和治疗提供了新的见解。
