人脐带间充质干细胞来源的细胞外囊泡携带 microRNA-29a 通过靶向 HMG-Box 转录因子/Wnt/-Catenin 信号通路改善原发性卵巢功能不全小鼠的卵巢功能。
Human Umbilical Cord Mesenchymal Stem Cell-Derived Extracellular Vesicles Carrying MicroRNA-29a Improves Ovarian Function of Mice with Primary Ovarian Insufficiency by Targeting HMG-Box Transcription Factor/Wnt/-Catenin Signaling.
机构信息
Department of Gynecology and Obstetrics, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China.
Reproductive Medicine Center, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China.
出版信息
Dis Markers. 2022 Jul 2;2022:5045873. doi: 10.1155/2022/5045873. eCollection 2022.
BACKGROUND
Primary ovarian insufficiency (POI) is a female disease characterized by ovarian function loss under 40 years old. Transplantation of exosomes is an encouraging regenerative medicine method that has the potential for restoring ovarian functions post-POI with high efficiency. Therefore, we investigate the therapeutic efficacy and potential mechanisms of human umbilical cord mesenchymal stem cell- (UCMSC-) derived exosomes on ovarian dysfunction post-POI.
METHODS
The model of POI was established by intraperitoneal injection with 5 mg/kg cisplatin. The mouse ovarian function was detected by measuring the levels of anti-Mullerian hormone, follicle-stimulating hormone, and estradiol and detecting the morphological changes. For in vitro experiments, the characterization and identification of UCMSCs and UCMSC-derived exosomes were done by observation of morphologies and flow cytometry. To exclude the interference effect of nonspecific precipitation substances, UCMSCs were treated with RNase A or RNase A in combination with Triton X-100. Granulosa cell (GC) identification was performed using immunofluorescence. GC proliferation and viability were assessed using 5-ethynyl-2'-deoxyuridine (EdU) assays and Cell Counting Kit-8 (CCK-8), and GC apoptosis was calculated by flow cytometry. Gene expression and protein levels were evaluated using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blotting. The binding relationship between miR-29a and HMG-box transcription factor (HBP1) was verified by luciferase reporter assays.
RESULTS
In vitro, the human UCMSC-derived exosomes carrying miR-29a upregulation promoted the proliferation of GCs and suppressed their apoptosis. In vivo, miR-29a upregulation reserved the mature follicles and restored the ovarian functions. miR-29a targeted HBP1 and negatively regulated its expression. HBP1 upregulation rescued the miR-29a upregulation-induced inhibition in GC apoptosis and inactivated the Wnt/-catenin pathway.
CONCLUSION
The exosomal miR-29a derived from human UCMSCs improves the ovarian function by targeting HBP1 and activating the Wnt/-catenin pathway.
背景
原发性卵巢功能不全（POI）是一种以 40 岁以下卵巢功能丧失为特征的女性疾病。外泌体移植是一种有前途的再生医学方法，有可能高效恢复 POI 后的卵巢功能。因此，我们研究了人脐带间充质干细胞（UCMSC）衍生的外泌体对 POI 后卵巢功能障碍的治疗效果和潜在机制。
方法
通过腹腔注射 5mg/kg 顺铂建立 POI 模型。通过测量抗苗勒管激素、卵泡刺激素和雌二醇的水平以及检测形态变化来检测小鼠的卵巢功能。对于体外实验，通过观察形态和流式细胞术对 UCMSC 和 UCMSC 衍生的外泌体进行特征和鉴定。为了排除非特异性沉淀物质的干扰作用，用 RNase A 或 RNase A 联合 Triton X-100 处理 UCMSC。通过免疫荧光法鉴定颗粒细胞（GC）。使用 5-乙炔基-2'-脱氧尿苷（EdU）测定法和细胞计数试剂盒-8（CCK-8）评估 GC 增殖和活力，并通过流式细胞术计算 GC 凋亡。通过逆转录定量聚合酶链反应（RT-qPCR）和蛋白质印迹法评估基因表达和蛋白水平。通过荧光素酶报告基因实验验证 miR-29a 与 HMG 盒转录因子（HBP1）之间的结合关系。
结果
体外，携带 miR-29a 上调的人 UCMSC 衍生外泌体促进 GC 增殖并抑制其凋亡。在体内，miR-29a 上调保留了成熟卵泡并恢复了卵巢功能。miR-29a 靶向 HBP1 并负调控其表达。HBP1 上调挽救了 miR-29a 上调诱导的 GC 凋亡抑制，并激活了 Wnt/-catenin 通路。
结论
来自人 UCMSC 的外泌体 miR-29a 通过靶向 HBP1 并激活 Wnt/-catenin 通路来改善卵巢功能。
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