肠道微生物代谢产物短链脂肪酸（SCFAs）在妊娠期糖尿病发生中的作用及机制探讨

Short chain fatty acid (SCFAs), which is produced by gut microbiota, has been increasingly shown to play a key role in the development of Type 2 diabetes mellitus. During pregnancy, there is a significant remodeling of the gut microbiota. However, the role of SCFAs in the gestational diabetes (GDM) is not clear. Our mouse study has demonstrated that free fatty acid receptor 2 (FFAR2), the specific receptor for SCFAs, contributes to glucose homeostasis during pregnancy. Based on our previous studies and other evidence, we hypothesize that aberrant change in gut microbiota during pregnancy leads to decreased level of SCFAs, resulting in activation of the downstream NF-kB pathway and subsequent modulation of various inflammatory responses via FFAR2 and eventually causing GDM. We aim to initiate a cohort study to enroll pregnant women during their early pregnancy. In both control subjects and women diagnosed with GDM, we propose to assess their metabolic profile for Week 8-12 and Week 24-28 of pregnancy by measuring serum and cecum SCFA levels, index of insulin resistance (IR), and expression of FFAR2, NF-kB pathway, adipokines and cytokines in white fat tissues (obtained from selected subjects who undergo C-sections). Additionally, we will generate an IR model in primary adipocytes derived from these subjects and challenge the primary adipocytes with SCFAs, selective agonist or siRNA for FFAR2 to further investigate the role of adipose SCFAs-FFAR2 pathway in the development of GDM. Finally, we seek to utilize Ffar2 knockout mouse model to further test our hypothesis. The overarching goal of our study is to elucidate the role of SCFAs-FFAR2 pathway in the pathogenesis of GDM and explore potential targets for prevention and treatment of GDM.

研究表明，肠道微生物代谢产物短链脂肪酸（SCFAs）和2型糖尿病发生关系密切。在孕期，肠道微生物组成发生重构，但SCFAs和妊娠期糖尿病（GDM)的关系尚不清楚。课题组前期动物实验发现SCFAs的FFRA2受体贡献了妊娠期糖稳态；基于前期研究及其他证据， 课题组提出如下假说：在孕期，肠道微生物代谢紊乱，产生SCFAs减少，通过其FFAR2受体激活下游NF-kB通路，调控炎症反应，介导了GDM的发生。我们拟采用人群队列研究，招募孕早期女性，动态留取血清及粪便标本，随访妊娠结局；对确定的GDM患者及正常孕妇检测SCFAs、脂肪组织中胰岛素抵抗（IR)、FFAR2、NFkB活化及相关炎症反应因子表达情况；对剖宫产孕妇脂肪细胞原代培养，诱导IR同时给予SCFAs、FFAR2激动或抑制及利用Ffar2基因敲除孕鼠实验等对假说进行验证，以期进一步阐明GDM的发生机制，为其防治寻找关键干预靶点。

本项目探讨了肠道微生物代谢产物短链脂肪酸（SCFAs）在GDM中的作用和机制，研究分为人群研究，细胞实验和动物实验三部分开展。第一部分为人群研究，招募孕早期女性建立人群队列，动态留取血清及粪便标本，随访妊娠结局。通过检测孕妇孕早期血清SCFAs水平发现，GDM组血清丙酸、丁酸、异戊酸的水平均低于对照组（P<0.01）。另外，对孕妇粪便的高通量测序分析显示，GDM组肠道菌群及其动态变化有明显差异。与糖脂代谢指标的相关性分析结果显示，一些产生SCFAs的菌群与血糖、血脂水平及胰岛素抵抗存在相关性。以上结果说明孕早期肠道SCFAs产生菌的相关变化与后期GDM发生风险相关，这从肠道微生态角度揭示孕期糖脂代谢的可能调控机制，为进一步的分子机制研究提供了研究靶点。随后，我们对SCFAs与GDM发生之间可能的机制进行了研究，纳入20例GDM孕妇及20例正常对照孕妇，在剖宫产时留取大网膜及皮下脂肪组织，利用western-blot和RT-PCR的方法检测胰岛素抵抗（IR）、FFAR2-NFkB炎症通路及相关炎症因子的表达情况，结果显示GDM孕妇和正常对照组孕妇之间无差异，考虑研究样本为剖宫产分娩时采集，已经为膳食干预过，GDM组和对照组两者结果的趋同可能与几个月的膳食干预有关。第二部分为细胞实验，利用剖宫产孕妇脂肪间充质干细胞原代培养成熟脂肪细胞，并诱导IR细胞模型，发现添加SCFAs能够提高脂肪细胞IRS1的表达，提示SCFAs对脂肪细胞的胰岛素抵抗有改善作用。第三部分为动物实验，通过向无菌小鼠移植GDM孕妇的肠道菌群，小鼠出现血糖升高和胰岛素抵抗，进一步验证了肠道菌群的异常与GDM的因果关系。另外，我们利用饲喂高脂饮食构建GDM孕鼠模型，通过灌胃SCFAs观察到丁酸能够显著改善孕鼠葡萄糖耐量。以上结果均提示肠道微生物及其产物SCFAs可能通过调节糖脂代谢与GDM相关，有望为将来GDM的预防和治疗提供新的干预靶点。