Cao Minkai, Bu Chaozhi, Zhang Jingjing, Ren Yongwei, Zhou Guanlun, Chen Chao, Han Guorong, Jiang Shi-Wen, Wen Juan
Department of Gynecology and Obstetrics, The Second Hospital of Nanjing, Nanjing University of Chinese Medicine, Nanjing 210003, China.
Department of Obstetrics and Gynecology, The Affiliated Wuxi Matemity and Child Health Care Hospital of Nanjing Medical University, Wuxi 214002, Jiangsu, China.
Int J Endocrinol. 2022 Jun 11;2022:9218113. doi: 10.1155/2022/9218113. eCollection 2022.
Impaired glucose metabolism and insulin sensitivity have been linked to the pathogenesis of gestational diabetes mellitus (GDM). Exosomes secreted by the umbilical cord mesenchymal stromal cells (UMSCs) and circular RNAs (circRNAs) derived from exosomes have been shown to be associated with the progression of GDM-related complications.
UMSCs were isolated from umbilical cords and identified through flow cytometry. Exosomes were isolated from UMSCs and were then characterized. The expression levels of RNA of hsa_circ_0046060, mmu_circ_0002819, and miR-338-3p were determined by quantitative real-time polymerase chain reaction (RT-qPCR). The intracellular glucose intake and glycogen content were measured using a High Sensitivity Glucose Assay Kit and Glycogen Assay Kit, respectively. Bioinformatics analysis and luciferase reporter assay were used to validate interactions among hsa_circ_0046060, miR-338-3p, and G6PC2. The expression of insulin receptor substrate-1 (IRS-1) and its phosphorylated form, (p-IRS-1), as well as G6PC2, was determined through western blotting.
UMSCs and exosomes were successfully isolated and identified. The upregulation of hsa_circ_0046060 decreased the intracellular glucose content in L-02 cells (43.45 vs. 16.87 pM/mg, =0.0002), whereas shRNA-mediated downregulation reversed this effect (16.87 vs. 33.16 pM/mg, =0.0011). Mmu_circ_0002819 in mice aggravated dysregulated glucose metabolism (49.88 vs. 21.69 pM/mg, =0.0031) and insulin sensitivity (0.20 vs. 0.11 mg/mL, =0.03) in GDM mice, which was abrogated by the knockdown of mmu_circ_0002819. The results of luciferase reporter assay revealed that miR-338-3p and G6PC2 were the potential targets of has_circ_0046060. Western blotting results showed that the reduced activation of IRS-1 induced by GDM (1.25 vs. 0.54, =0.0001) could be rescued by the administration of si-circ-G-UMSC-EXOs (0.54 vs. 1.17, =0.0001).
Taken together, the inhibition of hsa_circ_0046060 expression in exosomes from GDM-derived UMSCs can alleviate GDM by reversing abnormal glucose metabolism and insulin resistance and .
葡萄糖代谢受损和胰岛素敏感性与妊娠期糖尿病（GDM）的发病机制有关。脐带间充质基质细胞（UMSCs）分泌的外泌体以及源自外泌体的环状RNA（circRNAs）已被证明与GDM相关并发症的进展有关。
从脐带中分离UMSCs，并通过流式细胞术进行鉴定。从UMSCs中分离外泌体，然后对其进行表征。通过定量实时聚合酶链反应（RT-qPCR）测定hsa_circ_0046060、mmu_circ_0002819和miR-338-3p的RNA表达水平。分别使用高灵敏度葡萄糖检测试剂盒和糖原检测试剂盒测量细胞内葡萄糖摄取和糖原含量。采用生物信息学分析和荧光素酶报告基因检测来验证hsa_circ_0046060、miR-338-3p和G6PC2之间的相互作用。通过蛋白质印迹法测定胰岛素受体底物-1（IRS-1）及其磷酸化形式（p-IRS-1）以及G6PC2的表达。
成功分离并鉴定了UMSCs和外泌体。hsa_circ_0046060的上调降低了L-02细胞中的细胞内葡萄糖含量（43.45对16.87 pM/mg，P = 0.0002），而shRNA介导的下调则逆转了这种作用（16.87对33.16 pM/mg，P = 0.0011）。小鼠中的mmu_circ_0002819加重了GDM小鼠的葡萄糖代谢失调（49.88对21.69 pM/mg，P = 0.0031）和胰岛素敏感性（0.20对0.11 mg/mL，P = 0.03），而mmu_circ_0002819的敲低则消除了这种作用。荧光素酶报告基因检测结果显示，miR-338-3p和G6PC2是has_circ_0046060的潜在靶标。蛋白质印迹结果表明，GDM诱导的IRS-1激活降低（1.25对0.54，P = 0.0001）可通过给予si-circ-G-UMSC-EXOs来挽救（0.54对1.17，P = 0.0001）。
综上所述，抑制GDM来源的UMSCs外泌体中hsa_circ_0046060的表达可通过逆转异常葡萄糖代谢和胰岛素抵抗来缓解GDM。
