通过肠道菌群紊乱诱导的炎症级联反应研究补肾化痰方治疗绝经后骨质疏松症的机制

Studies have shown that the key to avoid postmenopausal osteoporosis(PMOP)and its development is to find safe and effective measures from traditional Chinese medicine, which interfere with intestinal flora disorders and the systemic inflammatory response as soon as possible.Based on the understanding of lipid metabolism and the correlation with PMOP and phlegm,we suggest that it should be treated from phlegm on the basis of tonifying kidney. Previous studies in our project group suggest that Bushen Huatan Decoction can improve bone mineral density of PMOP model rats, and regulate the intestinal flora disorders. Therefore, this project puts forward the hypothesis that: Bushen Huatan Decoction play a role in the treatment of PMOP by improving intestinal flora disorders and intestinal barrier functions, reduce the differentiation and proliferation of Th17 cells mediated by IL-6/JAK2/STAT3 signaling pathway and the secretion of IL-17 and TNF-α, and reduce the activation of osteoclast.This research intends to establish a rat model of postmenopausal osteoporosis by ovariectomy，with probiotics, estrogen supplements and different doses of Bushen Huatan Decoction intragastric intervention. By observing the changes of intestinal flora and intestinal barrier function, inflammatory cascade reaction and bone metabolismin in rats of each group, to systematically study the pathological characteristics of PMOP and the intervention targets and mechanisms of Bushen Huatan Decoction, and provide new methods and scientific basis for the treatment of PMOP by traditional Chinese medicine.

研究表明从中医药中发掘安全有效的措施尽早干预肠道菌群紊乱，减轻其诱导的全身炎症反应，是避免绝经后骨质疏松症（PMOP）发生发展的关键。基于脂代谢与PMOP、痰浊相关性的认识，课题组提出在补肾的基础上，还应从“痰”论治。我们的既往研究证明了补肾化痰方能够提高PMOP模型大鼠骨密度，改善肠道菌群紊乱，故本项目提出假说：补肾化痰方通过改善肠道菌群紊乱及肠道屏障功能，减少IL-6/JAK2/STAT3信号通路介导的Th17细胞分化增殖及IL-17、TNF-α分泌，降低破骨细胞活性，起到治疗PMOP的作用。因此，本研究拟以去卵巢建立PMOP模型，用益生菌、雌激素补充剂及不同剂量补肾化痰方灌胃干预，观察各组大鼠肠道菌群及肠道屏障功能变化、机体炎症级联反应改变及骨代谢特点，系统评价PMOP的病理特征及补肾化痰方干预的作用靶点及机制，为中医药治疗PMOP提出新方法和实验科学依据。

从中医药中发掘安全有效的措施尽早干预肠道菌群紊乱，减轻其诱导的全身炎症反应，是避免绝经后骨质疏松症（PMOP）发生发展的关键。基于脂代谢与PMOP、痰浊相关性的认识，课题组提出在补肾的基础上，还应从“痰”论治。根据预实验结果，本课题将90只大鼠随机分为6组，即假手术组（Sham）、模型组（OVX）、雌激素补充剂组（EV组）、补肾化痰方低、中、高剂量组（分别为BL组0.47 g/ml、BM组0.94 g/ml、BH组1.88g/ml中药药液灌胃），每组15只。对各组的骨代谢、肠道菌群及炎症因子等相关指标进行检测，结果显示，与假手术组相比，去卵巢模型大鼠呈现出骨小梁稀疏、变细、断裂，骨髓腔内增加了大量的脂肪细胞，破骨细胞（OC）的TRAP染色阳性细胞数较Sham组增多，成骨细胞（OB）的ALP染色阳性细胞数较Sham组有轻微升高，血清、结肠与骨组织IL-6、TNF-α、IL-17、STAT3/P-STAT3等炎症因子表达明显升高；肠道屏障（Occludin 、Claudin-1、ZO-1）遭到破坏，肠道菌群多样性下降等病理特征；通过与模型组、雌激素补充剂组进行组间比较，结果不同浓度的补肾化痰方均可以改善OVX大鼠的骨小梁结构；能够降低去卵巢后机体IL-6、IL-17、STAT3/P-STAT3等炎症因子的表达；能够增加OVX模型大鼠肠道菌群的物种丰度及多样性，增加有益菌，减低致病菌。从功能基因预测可见补肾化痰方通过影响肠道代谢、细胞的复制与修复等改善肠道菌群失衡状态。综上所述，补肾化痰方通过调节肠道菌群，影响IL-6/JAK2/STAT3信号通路，减轻机体炎症反应，降低破骨细胞活性，最终起到降低PMOP骨吸收的作用，但作用并不存在明显的剂量依赖。本课题系统评价了PMOP的病理特征及补肾化痰方干预的作用靶点及机制，为中医药治疗PMOP提出新方法和实验科学依据。