人尿源外泌体介导靶向ac4C修饰的NR5A1modRNA诱导自体脂肪间充质干细胞分化为Leydig样细胞的研究

原发性性腺功能低下症严重影响男性身心健康，而具有睾酮分泌功能的Leydig样细胞移植具有一定的治疗前景。传统的方法诱导干细胞分化为Leydig样细胞不仅耗时长、且效率低。前期实验证实，通过慢病毒过表达NR5A1能使hADMSCs快速、高效地分化为Leydig样细胞，但转基因不安全，很难应用于临床。研究显示，modRNA具有安全、快速的表达特点，而外泌体是一种潜在的天然载体。鉴于此，我们推测外泌体加载modNR5A1将是获取Leydig样细胞的一种安全、有效的策略。因此，本项目拟以人尿源性外泌体为载体，采用优化的超声法加载靶向ac4C修饰的modNR5A1，与自体hADMSCs共培养后，分析睾酮生成及诱导分化后的细胞特性，并探讨靶向ac4C修饰的modNR5A1作用特点。同时结合体内实验，探讨诱导分化后的细胞移植对雄性性腺功能低下症模型大鼠的疗效，为男性性腺功能低下症提供一个新的治疗策略。

Primary hypogonadism seriously affects physical and mental health of men, and Leydig-like cell transplantation with testosterone secretion function has a therapeutic potential. It is time-consuming and inefficient to induce stem cells to differentiate into Leydig-like cells by traditional methods. Our previous study showed that lentivirus-mediated overexpression of NR5A1 could convert hADMSCs into Leydig-like cells rapidly and efficiently, but there are safety problems in transgenic technology, which is difficult to be applied in clinical practice. Recent studies have shown that modRNA has the characteristics of high safety and rapid expression, and exosome is a potential natural carrier. Based on this, we speculate that exosomes loading modNR5A1 will be a safe and effective strategy for obtaining Leydig-like cells. Therefore, for this study, urine-derived exosomes will be used as carrier to load ac4C-modified modNR5A1 by optimized ultrasonic method, testosterone generation and differentiated cells are analyzed after co-culture with autologous hADMSCs, and then the role of ac4C modification is investigated. Furthermore, an in-vivo experiment will be carried out in hypogonadism rat model to evaluate the therapeutic effect of Leydig-like cells derived from hADMSCs. Eventually, it will provide a new treatment strategy for male hypogonadism.