Table2_Immune landscape in rejection of renal transplantation revealed by high-throughput single-cell RNA sequencing.xlsx

Background: The role of the cellular level in kidney transplant rejection is unclear, and single-cell RNA sequencing (scRNA-seq) can reveal the single-cell landscape behind rejection of human kidney allografts at the single-cell level.
Methods: High-quality transcriptomes were generated from scRNA-seq data from five human kidney transplantation biopsy cores. Cluster analysis was performed on the scRNA-seq data by known cell marker genes in order to identify different cell types. In addition, pathways, pseudotime developmental trajectories and transcriptional regulatory networks involved in different cell subpopulations were explored. Next, we systematically analyzed the scoring of gene sets regarding single-cell expression profiles based on biological processes associated with oxidative stress.
Results: We obtained 81,139 single cells by scRNA-seq from kidney transplant tissue biopsies of three antibody-mediated rejection (ABMR) patients and two acute kidney injury (AKI) patients with non-rejection causes and identified 11 cell types, including immune cells, renal cells and several stromal cells. Immune cells such as macrophages showed inflammatory activation and antigen presentation and complement signaling, especially in rejection where some subpopulations of cells specifically expressed in rejection showed specific pro-inflammatory responses. In addition, patients with rejection are characterized by an increased number of fibroblasts, and further analysis of subpopulations of fibroblasts revealed their involvement in inflammatory and fibrosis-related pathways leading to increased renal rejection and fibrosis. Notably, the gene set score for response to oxidative stress was higher in patients with rejection.
Conclusion: Insight into histological differences in kidney transplant patients with or without rejection was gained by assessing differences in cellular levels at single-cell resolution. In conclusion, we applied scRNA-seq to rejection after renal transplantation to deconstruct its heterogeneity and identify new targets for personalized therapeutic approaches.

背景：细胞水平在肾移植排斥反应中的作用尚不明确，而单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）能够在单细胞层面揭示人类肾脏同种异体移植排斥反应背后的单细胞图谱。
方法：本研究从5例人类肾移植活检组织芯中获取了高质量的转录组数据。基于已知的细胞标记基因对单细胞RNA测序数据进行聚类分析，以识别不同的细胞类型。此外，本研究还探究了不同细胞亚群所涉及的通路、拟时间发育轨迹以及转录调控网络。随后，基于与氧化应激相关的生物学过程，本研究系统分析了基于单细胞表达谱的基因集评分。
结果：本研究通过单细胞RNA测序，从3例抗体介导排斥反应（antibody-mediated rejection, ABMR）患者与2例非排斥原因导致的急性肾损伤（acute kidney injury, AKI）患者的肾移植组织活检样本中，共获得81139个单细胞，鉴定出11种细胞类型，包括免疫细胞、肾实质细胞以及多种基质细胞。巨噬细胞等免疫细胞呈现出炎症激活、抗原呈递及补体信号通路活化的特征，尤其在排斥反应患者中，部分特异性表达于排斥反应场景的细胞亚群展现出特定的促炎反应。此外，排斥反应患者的成纤维细胞数量显著增加，对成纤维细胞亚群的进一步分析显示，这些细胞参与了炎症及纤维化相关通路，进而加剧肾移植排斥反应与纤维化进程。值得注意的是，排斥反应患者体内氧化应激应答相关基因集的评分更高。
结论：本研究通过单细胞分辨率的细胞水平差异分析，揭示了伴或不伴排斥反应的肾移植患者之间的组织学差异。综上，本研究将单细胞RNA测序应用于肾移植后排斥反应的研究，解析了其异质性，并为个性化治疗方案找到了新的靶点。