健脾养胃辅助胃癌化疗的微生态学机制

The spleen-stomach is the foundation of acquired constitution. It is an important theorem and superiority characteristics of traditional Chinese medicine (TCM) in treating malignant tumors. Our previous clinic investigation showed that fortifying the spleen and nourishing the stomach treatment(FSNST) could significantly inhibit the severe adverse reactions of chemotherapy-treatment in gastric cancer patients.However,its biological mechanism remains poorly explored. The composition and characteristics of gut microbiome has been a hot topic in human disease research. Many studies suggest that the gut microbiome play a key role in the function of TCM spleen & stomach and may be the key target of the treatment of fortifying the spleen. And the microbiome in the tongue coating could be a new potential indicator of syndrome differentiation of the stomach diseases. In view of this, the present project will conduct a case-control study. The cases are the patients with gastric cancer (GC),including 1) GC patiens with continuous chemotherapy-treatment (CT) and 2) GC patients with continuous CT plus FSNST, while the controls are those who have the medical exam with no severe organic wrong. Microbiome diversity of the tongue coating and faeces is assessed by high-throughput sequencing of 16S/18S rRNA genes. Based on evaluating the curative effect of FSNST and guiding by the theory of medical statistics and bioinformatics, multi-factors statistical analysis is adopted among the microbiome in the tongue coating and faeces, the function of TCM spleen-stomach, treatment options, demographic characteristics, and so on. The aims of the project are 1) to dynamically characterize the variation of microbial composition among GC patients with CCT, GC patients with CCT & FSNST, and controls, 2) to identify the core microbial flora that related to spleen & stomach and in the tongue coating and faces, 3) to dynamically explore the key microbial flora that related to GC patients with CCT & FSNST, and 4) to explain the microbiota mechanism of FSNST of GC. Findings from this study will provide basic information for our understanding of TCM spleen-stomach, and will promote the normative scientific development of the integration of traditional and western medicine in treatment of GC patients.

脾胃乃后天之本，调治脾胃是中医治疗恶性肿瘤的重要法则和优势特色。临床发现健脾养胃可显著降低胃癌化疗不良反应，然而其生物学机制尚不清楚。肠道菌群的结构和特征已成为人类疾病研究的一个热点。研究提示肠道菌群是中医脾功能的关键执行者和健脾治疗的关键靶点，而舌苔菌群则是胃病证候分类的新潜在指标。鉴于此，本项目拟以胃癌患者（连续接受化疗或健脾养胃辅助化疗）为研究对象，无重大器质性病变的体检者为对照，运用高通量测序技术对舌苔和粪便微生物的16SrRNA和18S rRNA 基因测序。基于健脾养胃辅助胃癌化疗临床评价，动态解析舌苔和肠道菌群结构；运用统计学和生物信息学方法对菌群结构与脾胃功能、治疗方案、人口学特征等进行多因素关联分析，鉴定反映脾胃功能的舌苔与粪便的核心菌群，动态揭示健脾养胃有效辅助胃癌化疗的关键菌群，阐明健脾养胃辅助胃癌化疗的微生态学机制，推动中西医结合治疗胃癌的科学发展。

脾胃是后天之本，调治脾胃是中医治疗恶性肿瘤的重要法则和优势特色。本项目招募90例胃癌化疗患者，健脾养胃辅助胃癌化疗为观察组51例，单纯胃癌化疗为化疗组39例；无重大器质性病变的86例体检者为对照，年龄性别匹配。观察组和化疗组在3个采样点（第一次化疗前，第二次化疗前和第三次化疗前）采集舌苔和粪便，对照组收集1次舌苔和粪便，基于16S/18S rRNA 基因进行高通量测序检测舌苔和粪便菌群。.化疗前，比较胃癌患者与对照组的舌苔和粪便菌群差异。结果显示，与对照组相比，胃癌患者舌苔细菌多样性指数Shannon显著降低，舌苔真菌丰富度Ace显著升高，胃癌患者粪便细菌和真菌的丰富度Ace均显著降低。LEfSe判别分析菌群差异，发现舌苔70个细菌分类单元和131个真菌分类单元，粪便115个细菌分类单元和46个真菌分类单元。进一步ROC分析，发现4个舌苔细菌（OTUs）和17个粪便细菌（OTUs）、5个舌苔真菌（OTUs）和4个粪便真菌（OTUs）对胃癌具有潜在诊断价值。而且，较高丰度的舌苔韦荣菌科（Veillonellaceae）与胃癌无进展期大于12个月相关。.化疗前，观察组和化疗组菌群组成无显著差异；随着胃癌化疗的进行，比较胃癌患者3个采样点的舌苔和粪便菌群，化疗组和观察组均未发现显著差异，提示舌苔和粪便菌群具有可塑性，在化疗间期可以发生了肠道菌群恢复。.化疗前，观察组和化疗组生活质量无显著统计学差异（P > 0.05）；完成三个化疗周期后，观察组治疗后的生活质量显著高于化疗组（P < 0.05）。分析观察组和化疗组第三个采样点的菌群结构差异，结果发现，18个粪便细菌分类单元、74个粪便真菌分类单元、45个舌苔细菌分类单元、28个舌苔真菌分类单元与健脾养胃治法相关，健脾养胃治法可以增加舌苔和粪便的益生菌的相对丰度，降低致病菌的相对丰度，逆转化疗导致的菌群变化。.通过本项目研究，为研究健脾养胃治法的科学内涵提供了潜在的菌群靶点，为理解脾胃功能与肠道菌群的关系提供了新思路，为后续开展动物实验提供了基础。