清肝益肾祛风方对血管紧张素Ⅱ诱导的高血压小鼠肠道病理及微生态改变的干预研究

Effects of Qinggan Yishen Qufeng Formula on Intestinal Pathological, Microbiome and Metabolic Alterations in Angiotensin Ⅱ-induced Hypertensive Mice

目的探讨清肝益肾祛风方(Qinggan Yishen Qufeng formula,QYQ)对血管紧张素Ⅱ(Angiotensin Ⅱ,Ang Ⅱ)诱导的高血压相关肠道病理改变、肠道菌群及粪便代谢物失调的干预作用。方法将C57BL/6J小鼠随机分为假手术对照组(Sham组)、模型组(Vehicle组)及清肝益肾祛风方组(QYQ组)。Sham组皮下植入含生理盐水的渗透泵,Vehicle组及QYQ组皮下植入含Ang Ⅱ(490 ng/kg/min)的渗透泵。QYQ组自造模后第1日起给予QYQ灌胃,Sham组及Vehicle组给予同体积的生理盐水灌胃,连续28天;分别于造模前1天及造模后第7、14、28天(±2天)采用tail-cuff法测量小鼠血压,以评价QYQ降压效果;实验结束时收集小鼠粪便、远端回肠及近端结肠组织,采用苏木素伊红染色及Masson’s三色染色观察肠道组织形态及纤维化改变,采用16s rDNA测序对肠道菌群进行分析,采用超高效液相色谱-串联质谱对粪便代谢组进行分析,以评价QYQ对Ang Ⅱ诱导的高血压小鼠肠道病理改变、肠道菌群及粪便代谢物失调的干预作用。结果 与Vehicle组比较,QYQ显著抑制Ang Ⅱ诱导的小鼠收缩压升高(P <0.05),显著抑制其远端回肠及近端结肠病理改变的发生(P <0.05);QYQ显著纠正Ang Ⅱ诱导的高血压相关肠道菌群α多样性下降,部分纠正其β多样性改变,并对多种菌群的丰度异常具有抑制作用;QYQ可纠正部分粪便代谢物失调(P <0.05)。结论 QYQ能够显著抑制Ang Ⅱ诱导的小鼠高血压,同时对Ang Ⅱ高血压小鼠肠道病理改变、肠道菌群及粪便代谢物失调具有积极的防治作用。

Objective To explore the interventional effects of Qinggan Yishen Qufeng formula(QYQ) on hypertensionassociated intestinal pathological changes, gut dysbiosis and altered fecal metabolic profiles in Ang II-infused mice.Methods C57BL/6J mice were randomly divided into three experimental groups, namely, vehicle-treated shamoperated control group(Sham control), vehicle-treated Ang II-infused group(Vehicle control) and QYQ-treated Ang IIinfused group(QYQ). Mini-osmotic pumps were implanted subcutaneously for Ang II infusion. The QYQ group received daily QYQ treatment staring from the first day post the implantation of the pumps containing Ang II, while the Sham group and Vehicle group were given equal volume of normal saline in the same manner for 28 days. The blood pressure was measured by tail-cuff method one day before Ang II infusion and on the 7th, 14thand 28thday( ± 2 days) post the implantation of the pumps. The anti-hypertensive effect of QYQ was evaluated. At the end of the experiment, the fecal specimens, distal ileum and proximal colon tissues were collected. The intestinal histology and fibrosis were examined by histological methods including hematoxylin-eosin(H&E) staining and Masson’s trichrome staining. Intestinal microbiota was analyzed by 16s rDNA sequencing. The fecal metabolome was analyzed by ultra-high performance liquid chromatography-tandem mass spectrometry to evaluate the interventional effects of QYQ on hypertension-associated intestinal pathological changes, gut dysbiosis and alterations in the fecal metabolites. Results Compared with the Vehicle group, QYQ significantly inhibited the increase in systolic blood pressure induced by Ang II in mice(P < 0.05),and significantly inhibited the pathological changes in the distal ileum and proximal colon(P<0.05);In addition, QYQ treatment significantly corrected the decrease of Ang II-induced hypertension-related intestinal flora α diversity,partially corrected the changes of β diversity and therapeutically modified the abundance of multiple microorganisms.Lastly, QYQ treatment resulted in partial normalization of the fecal metabolites in the Ang II-infused mice(P < 0.05).Conclusion QYQ can significantly inhibit Ang II-induced high blood pressure in mice. Moreover, QYQ exhibits preventive and therapeutic effects on hypertension-associated intestinal pathological changes, gut dysbiosis and altered fecal metabolic profiles.