Zhang Yan, Du Mingyu, Zhu Baosheng, Ma Runmei, Zhang Jinman
Department of Medical Genetics, The First People's Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, No. 157 Jinbi Road, Xishan District, Kunming, Yunnan, 650000, China.
National Health Commission Key Laboratory of Preconception Health Birth in Western China, Kunming, Yunan, China.
Diabetol Metab Syndr. 2025 Apr 26;17(1):138. doi: 10.1186/s13098-025-01692-x.
We aimed to probe the effect of microRNA (miR)-455-3p derived from human umbilical cord mesenchymal stem cell exosomes (hucMSCs-Exos) in targeting calcium/calmodulin-dependent protein kinase 2 inhibitor 1 (CAMK2N1) to mitigate trophoblast cell injury in gestational diabetes mellitus (GDM).
hucMSCs were cultured, and Exos were isolated and characterized using transmission electron microscopy and Western blot (WB) for exosomal surface markers CD63, TSG101, and Calnexin. An in vitro GDM model was established by exposing human trophoblast cells (HRT-8/SVneo) to high glucose (HG), followed by intervention with Exos or overexpression of CAMK2N1. RT-qPCR or WB was applied to test miR-455-3p and CAMK2N1 expression levels. CCK-8 assay was adopted to assess cell proliferation, Transwell assay was applied to test cell migration, and flow cytometry was implemented to assess cell apoptosis. Bioinformatics websites and a dual-luciferase reporter gene assay were conducted to verify the targeting relationship between miR-455-3p and CAMK2N1.
The successfully isolated Exos expressed the exosomal markers CD63 and TSG101. Treatment with hucMSCs-Exos and hucMSCs-Exos carrying miR-455-3p mimic enhanced the migration and proliferation of HG-induced HRT-8/SVneo cells while reducing cell apoptosis. In contrast, the miR-455-3p inhibitor and overexpression of CAMK2N1 reversed these protective effects, leading to decreased cell migration and proliferation and increased apoptosis. Furthermore, bioinformatics analysis and experimental validation confirmed that miR-455-3p directly targeted and negatively regulated CAMK2N1.
miR-455-3p derived from hucMSCs-Exos exerts a protective effect against trophoblast cell injury in GDM by targeting and downregulating CAMK2N1.
我们旨在探究源自人脐带间充质干细胞外泌体（hucMSCs-Exos）的微小RNA（miR）-455-3p靶向钙/钙调蛋白依赖性蛋白激酶2抑制剂1（CAMK2N1）对减轻妊娠期糖尿病（GDM）中滋养层细胞损伤的作用。
培养hucMSCs，使用透射电子显微镜和蛋白质免疫印迹法（WB）检测外泌体表面标志物CD63、TSG101和钙连接蛋白，对分离出的外泌体进行鉴定。通过将人滋养层细胞（HRT-8/SVneo）暴露于高糖（HG）环境建立体外GDM模型，随后用外泌体或过表达CAMK2N1进行干预。应用逆转录定量聚合酶链反应（RT-qPCR）或WB检测miR-455-3p和CAMK2N1的表达水平。采用细胞计数试剂盒-8（CCK-8）法评估细胞增殖，采用Transwell法检测细胞迁移，采用流式细胞术评估细胞凋亡。利用生物信息学网站和双荧光素酶报告基因检测验证miR-455-3p与CAMK2N1之间的靶向关系。
成功分离出的外泌体表达外泌体标志物CD63和TSG101。用hucMSCs-Exos和携带miR-455-3p模拟物的hucMSCs-Exos处理可增强HG诱导的HRT-8/SVneo细胞的迁移和增殖，同时减少细胞凋亡。相反，miR-455-3p抑制剂和CAMK2N1过表达逆转了这些保护作用，导致细胞迁移和增殖减少，凋亡增加。此外，生物信息学分析和实验验证证实miR-455-3p直接靶向并负向调节CAMK2N1。
源自hucMSCs-Exos的miR-455-3p通过靶向并下调CAMK2N1对GDM中的滋养层细胞损伤发挥保护作用。
