从Notch信号通路探讨miR-124a对骨髓间质干细胞内源性胰岛分化潜能的调控机制及对糖尿病肾病的保护作用

Chronic kidney disease is a leading cause of mortality and morbidity in western countries which affects about 11% of the adult population.Among such patients, 50% patients were induced by diabetic nephropathy,which is a complication of diabetes and is defined as progressive kidney disease caused by angiopathy of the capillaries supplying the kidney glomeruli. Bone mesenchymal stem cells (BMSCs) are multipotent, self-renewing cells that can be found in almost all postnatal organs and tissues. The main functional characteristics of BMSCs are their immunomodulatory ability, capacity for self-renewal, and differentiation into mesodermal tissues. BMSCs have been used for the treatment of diabetic nephropathy in nonobese diabetic/severely compromised immunodeficient mice. However,it is not clear whether BMSCs can propagate after engraftment in the kidney. In addition, successful BMSCs treatment of diabetic nephropathy could be explained by BMSCs competence to differentiate into insulin-producing beta cells followed by decrease of glycemia and glycosuria,factors important for damaging renal cells. However, such efficiency of trans- differentiation is low and there is lack of large scale therapeutic applications at present.We hypothesized that miR-124a might motivate the endogenous differentiation of BMSCs into insulin-producing cells through Notch signal and BMSCs transplantation might prevent the pathological changes in the glomeruli and enhances their regeneration resulting in improved kidney function in diabetic animals.Our study may provide a simple, reliable procedure to obtain autologous stem cells that could provide a potentially unlimited source for transplantation. It may help to solve the major treatment problems of diabetic nephropathy.

骨髓间质干细胞（BMSCs）分化为胰岛素分泌细胞在糖尿病肾病修复过程中具有重要作用，如何提高其体外向胰岛素分泌细胞分化的效率是丞待解决的问题。本课题在前期研究基础上，提出"miR-124a可能通过抑制Notch信号通路激发BMSCs 内源性胰岛分化潜能，促进BMSCs体外分化为功能性胰岛"的新观点：筛选出调控胚胎胰腺发育的miRNAs- - miR-124a，通过模拟胚胎发育过程，激发BMSCs内源性胰岛分化潜能，促进BMSCs分化为功能性胰岛；在此基础上，通过Notch信号途径抑制剂构建类似于Notch信号敲除模型，探讨Notch信号通路在miR-124a调控BMSCs内源性胰岛分化潜能中的作用，进而通过动物模型研究BMSCs对糖尿病肾病的治疗作用及可能机制。本课题将miRNAs与干细胞成功结合，将为干细胞在糖尿病肾病细胞治疗中的应用提供一条新的思路。

慢性肾脏病（chronic kidney disease，CKD）的发生率和发病率呈上升趋势，已经成为全球关注的公共卫生问题；发达国家有50%以上的慢性肾脏病由糖尿病引起；预计到2025年，中国将成为世界糖尿病第一大国，糖尿病肾病也将成为我国导致ESRD的首位病因。因此有效控制糖尿病及糖尿病肾病是当前临床医学治疗学的重要课题。开发行之有效的糖尿病的治疗方法是全球范围内医学界面临的严重挑战，也是我国医学科学的重点研究课题。其实无论是1型糖尿病还是2型糖尿病，功能性胰岛细胞不足都是导致高血糖发生和疾病进展的主要决定性因素。干细胞诱导分化为胰岛细胞进行替代治疗无疑是糖尿病患者脱离胰岛素、恢复正常血糖控制最有希望的途径之一。骨髓间质干细胞（BMSCs）分化为胰岛素分泌细胞在糖尿病肾病修复过程中具有重要作用，如何提高其体外向胰岛素分泌细胞分化的效率是丞待解决的问题。本课题首先引入Notch信号通路：发现通过γ-分泌酶特异性抑制剂DAPT阻断Notch信号通路后，BMSCs分化为胰岛素和胰高血糖素阳性细胞显著增高，而且阻断Notch信号通路能通过改变胰腺关键转录因子的表达显著增强BMSCs体外分化能力，为BMSCs在糖尿病肾病细胞治疗中的应用奠定基础。接下来，通过糖尿病肾病动物模型发现，BMSCs移植能通过促进内源性胰岛细胞再生、减轻糖尿病肾病模型的肾小球肥大、减轻肾脏纤维化和足细胞转分化等方面减轻糖尿病大鼠的血糖、蛋白尿，改善糖尿病大鼠的预后；随后，初步建立了BMSCs和胰岛体外共同培养体系，并研究了高血糖对胰腺发育关键基因的影响。我们研究发现，BMSCs移植后在肾脏定植较少，而促进干细胞靶向迁移是亟待解决的问题。因此，本课题通过调控经典Wnt信号通路上调骨髓间质干细胞表面CXCR4表达量从而促进BMSCs的靶向迁移，为BMSCs移植治疗糖尿病肾病奠定基础。