Table4_Circulating hsa-miR-320a and its regulatory network in type 1 diabetes mellitus.xlsx

IntroductionIncreasing evidence from human and animal model studies indicates the significant role of microRNAs (miRNAs) in pancreatic beta cell function, insulin signaling, immune responses, and pathogenesis of type 1 diabetes (T1D).
MethodsWe aimed, using next-generation sequencing, to screen miRNAs from peripheral blood mononuclear cells of eight independent Kuwaiti-Arab families with T1D affected siblings, consisting of 18 T1D patients and 18 unaffected members, characterized by no parent-to-child inheritance pattern.
ResultsOur analysis revealed 20 miRNAs that are differentially expressed in T1D patients compared with healthy controls. Module-based weighted gene co-expression network analysis prioritized key consensus miRNAs in T1D pathogenesis. These included hsa-miR-320a-3p, hsa-miR-139-3p, hsa-miR-200-3p, hsa-miR-99b-5p and hsa-miR-6808-3p. Functional enrichment analysis of differentially expressed miRNAs indicated that PI3K-AKT is one of the key pathways perturbed in T1D. Gene ontology analysis of hub miRNAs also implicated PI3K-AKT, along with mTOR, MAPK, and interleukin signaling pathways, in T1D. Using quantitative RT-PCR, we validated one of the key predicted miRNA-target gene-transcription factor networks in an extended cohort of children with new-onset T1D positive for islet autoantibodies. Our analysis revealed that hsa-miR-320a-3p and its key targets, including PTEN, AKT1, BCL2, FOXO1 and MYC, are dysregulated in T1D, along with their interacting partners namely BLIMP3, GSK3B, CAV1, CXCL3, TGFB, and IL10. Receiver Operating Characteristic analysis highlighted the diagnostic potential of hsa-miR-320a-3p, CAV1, GSK3B and MYC for T1D.
DiscussionOur study presents a novel link between hsa-miR-320a-3p and T1D, and highlights its key regulatory role in the network of mRNA markers and transcription factors involved in T1D pathogenesis.

引言 越来越多基于人类及动物模型的研究证据表明，微小RNA（microRNAs, miRNAs）在胰腺β细胞功能、胰岛素信号传导、免疫应答以及1型糖尿病（type 1 diabetes, T1D）的发病机制中发挥重要作用。
方法 本研究借助下一代测序技术，对8个独立的科威特阿拉伯家族的外周血单个核细胞中的miRNAs进行筛选。这些家族均存在受累同胞，共纳入18名T1D患者与18名健康对照个体，且未观察到亲子代遗传模式。
结果 本研究分析显示，相较于健康对照，T1D患者体内存在20种差异表达的miRNAs。基于模块的加权基因共表达网络分析筛选出了T1D发病机制中的关键共识miRNAs，包括hsa-miR-320a-3p、hsa-miR-139-3p、hsa-miR-200-3p、hsa-miR-99b-5p及hsa-miR-6808-3p。差异表达miRNAs的功能富集分析表明，PI3K-AKT通路是T1D中受扰动的关键通路之一。枢纽miRNAs的基因本体分析进一步显示，除PI3K-AKT通路外，mTOR、MAPK及白细胞介素信号通路也与T1D发病机制相关。本研究通过实时定量逆转录聚合酶链反应，在扩展队列的新发T1D患儿（胰岛自身抗体检测阳性）中验证了关键的预测性miRNA-靶基因-转录因子调控网络。分析发现，hsa-miR-320a-3p及其关键靶基因（包括PTEN、AKT1、BCL2、FOXO1与MYC）在T1D中存在表达失调，其相互作用伙伴BLIMP3、GSK3B、CAV1、CXCL3、TGFB及IL10亦呈现异常表达。受试者工作特征曲线分析表明，hsa-miR-320a-3p、CAV1、GSK3B及MYC具备T1D诊断潜力。
讨论 本研究揭示了hsa-miR-320a-3p与T1D之间的全新关联，并阐明了其在T1D发病机制相关的mRNA标志物及转录因子网络中的关键调控作用。