Characterization and effective expansion of CD4-CD8- TCRab+ T cells from individuals living with type 1 diabetes

CD4-CD8- TCRab+ (double-negative [DN]) T cells represent a rare T cell population that promotes immunological tolerance through various cytotoxic mechanisms. In mice, autologous transfer of DN T cells has shown protective effects against autoimmune diabetes and graft-versus-host disease. Here, we characterized humanDNT cells from people living with type 1 diabetes (PWT1D) and healthy controls. We found that while DN T cells and CD8+ T cells share many similarities, DN T cells are a unique T cell population, both at the transcriptomic and protein levels. We also show that by using various cytokine combinations, human DN T cells can be expanded in vitro up to 1,000- fold (mean >250-fold) and remain functional post-expansion. In addition, we report that DN T cells from PWT1D display a phenotype comparable to that of healthy controls, efficiently expand, and are highly functional. As DN T cells are immunoregulatory and can prevent T1D in various mouse models, these observations suggest that autologous DN T cells may be amenable to therapy for the prevention or treatment of T1D. Overall design: RNA-seq profiling of human CD4+, CD8+, TCRgd+ and DN T cells from 3 healthy donors, ex vivo and after 14-day expansions in vitro.

CD4-CD8- TCRαβ+（双阴性[DN]）T细胞是一类罕见的T细胞亚群，可通过多种细胞毒性机制介导免疫耐受。在小鼠模型中，自体转输DN T细胞已被证实可对自身免疫性糖尿病及移植物抗宿主病（GVHD）发挥保护性作用。本研究对1型糖尿病患者（people living with type 1 diabetes，简称PWT1D）及健康对照人群的人源DN T细胞进行了系统表征。研究发现，尽管DN T细胞与CD8+ T细胞存在诸多共性，但无论是转录组层面还是蛋白质组层面，DN T细胞均为一类独特的T细胞亚群。此外，本研究证实，通过多种细胞因子组合的培养体系，人源DN T细胞可在体外扩增最高达1000倍（平均扩增倍数＞250倍），且扩增后仍维持功能活性。进一步研究显示，1型糖尿病患者的DN T细胞表型与健康对照人群无显著差异，可实现高效扩增并保持高度功能活性。鉴于DN T细胞具有免疫调节功能，且在多种小鼠模型中可预防1型糖尿病，上述结果提示自体DN T细胞或可用于1型糖尿病的预防或治疗。整体实验设计：对3名健康供者来源的CD4+、CD8+、TCRγδ+及DN T细胞进行RNA测序表征分析，涵盖离体原代样本及体外14天扩增培养后的样本。