基于肠道菌群的人参、黄连“药对”治疗2型糖尿病的抗炎机制研究

The drug pair of ginseng and rhizoma coptidis is often used to treat type 2 diabetes. The preliminary studies have suggested that the ginseng and rhizoma coptidis drug pair can improve insulin resistance and regulate glucose metabolism, which may be related to its anti-inflammatory effect . But the underlying mechanism is unclear. Moreover, it has been proved that the low-grade inflammation state of type 2 diabetes has close relationship with intestinal flora disorder. Therefore we propose a hypothesis that the anti-inflammation and glucose regulation effect of ginseng and rhizoma coptidis drug pair may act through regulating the intestinal flora, correcting the increased absorption of endotoxin and changing the composition of intestinal short chain fatty acids. In order to test this hypothesis, this study will choose metformin as contrast to observe the impact of ginseng and rhizoma coptidis drug pair on blood glucose, fasting insulin and inflammatory factors. Meanwhile, the whole genome of intestinal flora will be detected by macro genome sequencing technology and mice intestinal fatty acids will be detected using gas chromatography. We also measure levels of serum LPS, Zonulin and LPS by enzyme-linked immunoassay, and detect intestinal tissue Occludin expression level with the protein imprinting method. The db/db mice will be used in this study. In the end, the result is expected to uncover the anti-inflammatory mechanism of ginseng and rhizoma coptidis drug pair based on intestinal flora. This study will provide important scientific basis for prevention and treatment of Traditional Chinese medicine for diabetes mellitus.

人参、黄连“药对”为中医治疗2型糖尿病的常用对药，前期研究证实人参、黄连“药对”能够改善胰岛素抵抗，调节糖代谢，机制可能与抗炎作用有关，但其抗炎机制不明确。目前，2型糖尿病的低度炎症状态与肠道菌群紊乱相关已经得到证实。因此我们提出假说：人参、黄连“药对”通过调节肠道菌群，纠正因菌群紊乱导致的内毒素吸收增加以及肠道短链脂肪酸成分改变，起到抗炎作用，从而能够调节血糖。为验证此假说，本研究将以二甲双胍为对照，观察人参、黄连“药对”对db/db小鼠血糖、空腹胰岛素以及炎症因子的影响，同时采用宏基因组测序技术对小鼠进行全基因组测序，采用气相色谱法测定小鼠肠道短链脂肪酸，采用酶联免疫法测定血清LPS、Zonulin、LPS等的水平，蛋白印迹法测定肠道组织Occludin表达水平。最终，从调节肠道菌群的角度，明确人参、黄连“药对”抗炎、改善胰岛素抵抗的机制，这将为中医药防治糖尿病提供重要的科学依据。

糖尿病（diabetes mellitus,DM）是一组常见的以葡萄糖和脂肪代谢紊乱、血浆葡萄糖水平增高为特征的代谢内分泌疾病，我们认为 2 型糖尿病的基本病机为热伤气阴，治疗强调清热益气养阴法的应用。人参、黄连“药对”为此治法的代表药物，也是中医治疗2型糖尿病的常用对药，前期研究证实人参、黄连“药对”能够改善胰岛素抵抗，调节糖代谢，机制可能与抗炎作用有关，但其抗炎机制不明确。目前，2型糖尿病的低度炎症状态与肠道菌群紊乱相关已经得到证实。为了进一步研究其抗糖尿病的作用机制。本课题以二甲双胍为对照，观察人参、黄连“药对”对db/db小鼠血糖、胰岛素以及炎症因子的影响，同时采用宏基因组测序技术对小鼠进行全基因组测序，采用LAL法测定血清LPS的水平，蛋白印迹法测定肠道组织Occludin表达水平。研究发现，肠道菌群失调在2型糖尿病小鼠中普遍存在。而中药干预显著的改变了糖尿病小鼠的肠道微生物丰富度及多样性。人参、黄连药对可以显著降低dbdb小鼠血糖，降低血清lps水平及胰腺IL1β表达，增加肠道组织 Occludin 的表达。人参、黄连药对对肠道菌群紊乱有调节作用，且可能通过调节肠道菌群，纠正因菌群紊乱导致的内毒素吸收增加、肠道屏障受损起到抗炎作用。宏基因分析发现人参、黄连药对能减少与lps合成、核黄素代谢和过氧化物酶体通路有关的prevotellaceae（普雷沃氏菌科）、rikenellaceae（理研菌科）和helicobacteraceae（螺杆菌科）等菌群相关丰度。还能上调与淀粉和蔗糖的代谢以及戊糖和葡萄糖醛酸相互转化通路相关的bacteroidaceae（拟杆菌科）的丰度，这些菌群可能是黄连、人参药对治疗2型糖尿病所调节的关键菌群。