云南道地药材滇黄精多糖结构及降血糖机理研究

Diabetes is now a common and serious global health problem. Although western medicine can effectively control their glucose levels, they also have unwanted and serious side effects. Plant polysaccharides in the prevention and treatment of diabetes mellitus and its complications advantage has been widespread concerned by the pharmaceutical industry. Rhizoma Polygonatum kingianum is used for treatment of diabetes mellitus in Yunnan Yi folk for a long term, but its material basis for efficacy and mechanism of anti-diabetes remains unclear. The total polysaccharide and the polysaccharide that separated from it have remarkable effect on reducing blood sugar in the preliminary research, which has been authorized 2 patents. This study based on the pharmacodynamics and chemical basis, the specific glycosidase cutting technology with GC-MS, IR, NMR, X ray diffraction and other modern analysis method will apply to clarify the molecular structure information of polysaccharides. At the same time, the spontaneous and experimental of type 2 diabetes mellitus animal model, the model of liver cells and fat cells will be used to study the effect and mechanism of hypoglycemic activity of a polysaccharide from the Rhizome Polygonatum kingianum. After that, preliminary analyze the relationship between its structure and hypoglycemic activity. The fruits of the research will expected to be developed into new drug with independent intellectual property rights, which will provide a strong scientific support for the follow-up product development.

糖尿病是严重危害健康的全球公共卫生问题，虽然降糖西药能有效控制高血糖，但不良反应严重，植物活性多糖在防治糖尿病及其并发症的优势已受到医药界普遍关注。滇黄精在云南彝族民间长期用于治疗糖尿病，疗效确切，但其活性物质基础和作用机制尚未明了。项目组前期研究首次发现其总多糖和从中分离得到的均一多糖PPS-I具有显著的降血糖作用，成果已获专利2项。本研究基于已有药效学和化学基础，拟用GC-MS、IR、NMR、X射线衍射等现代分析方法，研究滇黄精中降血糖活性均一多糖的理化性质，阐明其分子结构信息。并采用自发性和实验性2型糖尿病动物模型，以及离体肝细胞和脂肪细胞模型，从体内和体外探讨滇黄精多糖降血糖的作用及机理，并初步分析其结构与降血糖活性之间的关系。研究成果有望开发成具有自主知识产权的新型防治糖尿病天然药物，为后续的产品研发、变资源优势为产业优势提供强有力的科学支撑。

本研究从滇黄精中得到滇黄精总多糖PPS，并从中分离PPS-1 ~ 6 六种均一多糖。采用光谱（紫外、红外光谱）、色谱（PMP-HPLC）、质谱（1H，13C NMR，1H-1H COSY）理化方法，重点对滇黄精中的均一多糖PPS-1、PPS-2的的结构进行了解析。同时，建立了稳定高效的滇黄精多糖PPS实验室制备路线。通过2型糖尿病的小鼠模型对滇黄精总多糖和均一多糖抗糖尿病活性进行了研究，证实了PPS和PPS-1均可使糖尿病小鼠生活状况改善，空腹血糖值（FBG）显著性降低，改善糖尿病小鼠的葡萄糖耐量，降低TC、TG、LDL水平，显著增加肝、肌糖原含量，缓解糖尿病小鼠肾脏病理性结构变化，修复糖尿病所引起的胰腺损害，PPS-2未发现类似作用。此外，通过建立高胰岛素诱导的HepG2细胞胰岛素抵抗模型(IR-HepG2)，研究PPS及其单体(PPS1~4)对IR-HepG2葡萄糖消耗的影响及对胰岛素信号通路中关键酶或蛋白的mRNA表达水平的影响。结果显示滇黄精总多糖PPS及四种均一多糖，除PPS-3外，均可改善胰岛素抵抗，其中PPS、PPS-1和PPS-4作用显著。机制研究表明滇黄精多糖PPS改善胰岛素抵抗的机理为增强胰岛素受体底物1（IRS1）、磷脂酰肌醇3激酶 (PI3K)，蛋白激酶B (AKT)的表达水平，作用于转录因子叉头蛋白 (FoxO1)，正向调控磷酸烯醇式丙酮酸羧激酶 (PEPCK)、葡萄糖-6-磷酸 (G6Pase) 等关键酶的表达，改善胰岛素传导，增加胰岛素敏感性，从而通过多种途径改善胰岛素抵抗。而均一多糖PPS1对AKT的相对表达无明显作用，但FOXO1，G6pase的表达促进影响极显著，PPS-1对IR-HepG2细胞的改善作用可能是直接对该通路下游的FOXO1作用，提高G6pase和PEPCK的表达，从而促进葡萄糖的消耗。研究成果将为后续的产品研发，变资源优势为产业优势提供强有力的科学支撑。