METHODS: Global profiling of the tRFs present in pancreatic islets of 4- and 8-week-old NOD mice and in extracellular vesicles released by activated CD4+ T lymphocytes was performed by small RNA-seq. Changes in the level of specific fragments were confirmed by quantitative PCR. The transfer of tRFs from immune cells to beta cells occurring during insulitis was assessed using an RNA-tagging approach. The functional role of tRFs increasing in beta cells during the initial phases of type 1 diabetes was determined by overexpressing them in dissociated islet cells and by determining the impact on gene expression and beta cell apoptosis.
RESULTS: We found that the tRF pool was altered in the islets of NOD mice during the initial phases of type 1 diabetes. Part of these changes were triggered by prolonged exposure of beta cells to proinflammatory cytokines (IL-1β, TNF-α and IFN-γ) while others resulted from the delivery of tRFs produced by CD4+ T lymphocytes infiltrating the islets. Indeed, we identified several tRFs that were enriched in extracellular vesicles from CD4+/CD25- T cells and were transferred to beta cells upon adoptive transfer of these immune cells in NOD.SCID mice. The tRFs delivered to beta cells during the autoimmune reaction triggered gene expression changes that affected the immune regulatory capacity of insulin-secreting cells and rendered the cells more prone to apoptosis.
CONCLUSIONS: Our data point to tRFs as novel players in the crosstalk between the immune system and insulin-secreting cells and suggest a potential involvement of this novel class of non-coding RNAs in type 1 diabetes pathogenesis.
METHODS: Sequences are available from the Gene Expression Omnibus (GEO) with accession numbers GSE242568 and GSE256343.
方法:通过小RNA-seq对存在于4周龄和8周龄NOD小鼠胰岛中以及由活化的CD4+T淋巴细胞释放的细胞外囊泡中的tRF进行了全局分析。通过定量PCR确认特异性片段水平的变化。使用RNA标记方法评估在胰岛炎期间发生的tRF从免疫细胞向β细胞的转移。在1型糖尿病的初始阶段,tRF在β细胞中增加的功能作用是通过在解离的胰岛细胞中过表达它们并通过确定对基因表达和β细胞凋亡的影响来确定的。
结果:我们发现,在1型糖尿病的初始阶段,NOD小鼠胰岛中的tRF池发生了改变。这些变化的一部分是由β细胞长时间暴露于促炎细胞因子(IL-1β,TNF-α和IFN-γ),而其他原因是CD4T淋巴细胞浸润胰岛产生的tRF的传递。的确,我们鉴定了几个tRF,这些tRF在CD4+/CD25-T细胞的胞外囊泡中富集,并在NOD中这些免疫细胞过继转移后转移至β细胞.SCID小鼠。在自身免疫反应期间递送至β细胞的tRF触发了影响胰岛素分泌细胞的免疫调节能力并使细胞更易于凋亡的基因表达变化。
结论:我们的数据表明,tRF是免疫系统和胰岛素分泌细胞之间相互作用的新型参与者,并提示这类新型非编码RNA可能参与1型糖尿病的发病机制。
方法:序列可从基因表达Omnibus(GEO)获得,登录号为GSE242568和GSE256343。