乳凝集素在糖尿病肠道病变中的分子机制及干预研究

Diabetic intestinal disease is a common complication of diabetes and is more common in elderly patients. It is mainly related to diabetic autonomic neuropathy, intestinal hormone abnormalities, intestinal histological changes, intestinal microecological changes, causing pain, nausea, vomiting, diarrhea, constipation and other symptoms, seriously affecting the life of patients. We used proteomics technology to study diabetic mice and found that differential protein MFG-E8 has a promoting effect on the development of diabetic complications. Although in recent years, MFG-E8 has been gradually in the study of various diseases began to attract people's attention, but the molecular mechanism and regulation of intestinal endocrine mechanism in diabetes in intestinal complications is not clear. This project aims to establish the glycosylation of cell injury model and aging animal models with diabetes mellitus induced by STZ in SAMP8 mice, to construct interference or over-expression carrier of MFG-E8 both in vivo and in vitro transfection vector, and the D-pinitol intervention, clear the molecular mechanism of MFG-E8 in intestinal endocrine cell glycosylation injury and the diabetic intestinal diseases, and the protective effect of D-pinitol intervention. Therefore，provide new drug targets for clinical prevention and treatment of diabetic complications and various aging related disease.

糖尿病肠道病变是糖尿病常见的并发症，主要与糖尿病自主神经病变、胃肠道激素异常、胃肠道组织形态学改变、肠道微生态改变等因素相关，引起疼痛、恶心、呕吐、腹泻、便秘等症状，严重影响患者的生活，在老年患者中尤为常见。我们前期应用蛋白质组学技术对糖尿病小鼠进行研究，发现差异蛋白乳凝集素（MFG-E8）对糖尿病多个靶器官病变的发生发展均具有促进作用，但其在糖尿病肠道并发症中作用的分子机制及调控肠道内分泌激素的作用机制尚不明确。本项目拟建立糖基化损伤小肠内分泌细胞模型，并以STZ诱导的SAMP8快速老化小鼠建立衰老糖尿病动物模型，构建MFG-E8的干扰或过表达载体进行体内和体外转染，并应用D-松醇干预，明确MFG-E8在糖基化损伤和糖尿病肠道病变中的分子机制，以及D-松醇的保护作用，为临床预防和治疗糖尿病肠道并发症乃至多种衰老相关性疾病提供新的干预及药物靶点。

糖尿病肠道病变是糖尿病常见的并发症，主要与糖尿病自主神经病变、胃肠道激素异常、胃肠道组织形态学改变、肠道微生态改变等因素相关，引起疼痛、恶心、呕吐、腹泻、便秘等症状。我们前期应用蛋白质组学技术对糖尿病小鼠进行研究，发现了差异蛋白乳凝集素(MFG-E8)，其对糖尿病多个靶器官病变的发生发展均发挥重要的作用，但在糖尿病肠道并发症中及调控肠道内分泌激素的作用机制尚不明确。本项目建立了糖基化损伤小肠内分泌细胞模型，并以STZ诱导的SAMP8快速老化小鼠建立衰老糖尿病动物模型，构建MFG-E8的干扰或过表达载体进行转染，并应用D-松醇干预。研究发现，与正常对照组相比，应用STZ诱导的SAMP8糖尿病小鼠及SAMR1糖尿病小鼠回肠组织在光镜下出现钝变、短缩、绒毛尖脱落等病理改变。Western blot和免疫荧光染色显示糖尿病小鼠的MFG-E8蛋白表达显著降低，p-MLKL和HMGB1表达增加。此外，在晚期糖基化终末产物(advanced glycation end products, AGEs)诱导的STC-1细胞中，发现了促炎介质(TNF-α、IL-1β、IL-6)的释放和HMGB1的表达，以及RIPK3/MLKL信号通路的激活。D松醇预处理可显著减少促炎介质，增加MFG-E8的表达。MFG-E8小干扰RNA (siRNA)干预促进了RIPs通路和炎症因子的活性，而MFG-E8过表达载体干预则抑制了RIPs通路和炎症因子的活性。MFG-E8下调是糖尿病相关肠道炎症损伤发生发展中的重要机制。本项目研究结果表明，MFG-E8过表达和D松醇对糖尿病坏死诱导的肠道炎症起保护作用，并能够维持STC-1细胞功能。本研究明确了MFG-E8在糖基化损伤和糖尿病肠道病变中的分子机制，以及D-松醇的保护作用，为临床预防和治疗糖尿病肠道并发症乃至多种衰老相关性疾病提供了新的干预及药物靶点。