免疫逃避诱导多能干细胞（iPSC）衍生的肾脏类器官，被称为“隐形”类器官，有望用于临床移植。为了解决免疫排斥问题，我们研究了移植前对肾脏类器官进行基因改造的 I 类人类白细胞抗原 (HLA) 的影响。通过使用 CRISPR-Cas9，我们成功敲除 β-2-微球蛋白 (B2M)，从而产生缺乏 HLA I 类表面表达的 iPSC。在体外，B2M 敲除可保护源自这些 iPSC 的肾脏类器官免受 T 细胞排斥。为了评估体内保护作用，将未修饰的（对照）和 B2M-/- 肾类器官移植到移植了人外周血单核细胞 (PBMC) 的人源化小鼠中。证实人类 PBMC 成功植入，4 周后，我们观察到对照和 B2M-/- 类器官之间 CD4 和 CD8 T 细胞的浸润率、增殖或细胞毒性没有明显差异。两组类器官均表现出组织完整性受损，表现为肾小管炎和肾小管完整性丧失。值得注意的是，虽然 B2M-/- 类器官未能在其细胞表面表达 HLA I 类，但在对照和移植到含有人 PBMC 的小鼠体内的 B2M-/- 类器官中，预先存在 HLA II 类表达。 HLA II 类表达不仅限于抗原呈递细胞，在肾类器官的上皮细胞中也很明显，这对其移植带来了额外的免疫挑战。因此，我们得出结论，仅 B2M 敲除不足以保护 iPSC 衍生的肾类器官免受 T 细胞介导的免疫排斥。此外，我们的研究结果表明，调节 HLA II 类信号传导对于预防移植后的排斥反应是必要的。© 作者 2023。由牛津大学出版社出版。

Immune evasive induced pluripotent stem cell (iPSC)-derived kidney organoids, known as "stealth" organoids, hold promise for clinical transplantation. To address immune rejection, we investigated the impact of genetically modifying human leukocyte antigen (HLA) class I in kidney organoids prior to transplantation. By using CRISPR-Cas9, we successfully knocked out beta-2-microglobulin (B2M), resulting in iPSCs devoid of HLA class I surface expression. In vitro, the B2M knockout protected kidney organoids derived from these iPSCs against T-cell rejection. To assess in vivo protection, unmodified (control) and B2M-/- kidney organoids were transplanted into humanized mice engrafted with human peripheral blood mononuclear cells (PBMCs). Successful engraftment of human PBMCs was confirmed, and after 4 weeks, we observed no discernible difference in the infiltration rate, proliferation, or cytotoxicity of CD4+ and CD8+ T cells between control and B2M-/- organoids. Both groups of organoids showed compromised tissue integrity, displaying tubulitis and loss of tubule integrity. Notably, while B2M-/- organoids failed to express HLA class I on their cell surface, there was preexisting expression of HLA class II in both control and B2M-/- organoids transplanted into mice with human PBMCs. HLA class II expression was not limited to antigen-presenting cells but also evident in epithelial cells of the kidney organoid, posing an additional immunological challenge to its transplantation. Consequently, we conclude that B2M knockout alone is insufficient to protect iPSC-derived kidney organoids from T-cell-mediated immune rejection. Additionally, our findings suggest that modulating HLA class II signaling will be necessary to prevent rejection following transplantation.© The Author(s) 2023. Published by Oxford University Press.