基于16S rRNA测序技术探讨骨松益骨方对去卵巢大鼠肠道菌群的影响

Effect of GusongYigu decoction on gut microbiota in ovariectomized rats based on 16S rRNA sequencing

目的基于肠道菌群探究骨松益骨方(GusongYigu decoction,GSYG)治疗绝经后骨质疏松症(postmenopausal osteoporosis,PMOP)的机制。方法SPF级SD大鼠60只随机分成假手术组(Control)、模型组(Model)、骨松益骨方低剂量组(GSYG-L,1.25 g·kg^(-1))、骨松益骨方中剂量组(GSYG-M,2.5 g·kg^(-1)),骨松益骨方高剂量组(GSYG-H,5 g·kg^(-1))和阿仑膦酸钠组(Al,0.89 mg·kg^(-1)),每组10只。HE染色观察骨组织形态变化,micro CT观察大鼠的股骨组织结构变化;取Control组、Model组、GSYG-H组大鼠结肠粪便样本,提取DNA,利用Illumina Miseq平台进行高通量测序。结果与Control组比较,Model组大鼠骨小梁稀疏;BV/TV、Tb.Th、Tb.N及BMD均明显降低(P<0.01),SMI、Tb.Sp明显升高(P<0.01);与Model组比较,GSYG-L、GSYG-M、GSYG-H组和Al组骨小梁数目增加,骨微结构得到改善,且BV/TV、Tb.Th、Tb.N及BMD明显增加(P<0.05),SMI、Tb.Sp明显降低(P<0.05)。肠道菌群测序结果显示,与Control组比较,Model组大鼠肠道菌群多样性降低,厚壁菌门丰度降低,而拟杆菌门丰度增加;与Model组比较,GSYG-H组厚壁菌门丰度增加,拟杆菌门丰度减少。结论GSYG能够改善PMOP患者的骨微结构,增加骨密度,其作用机制可能与增加肠道菌群的多样性、调节肠道菌群结构有关。

Aim To explore the mechanism of GusongYigu decoction(GSYG)in the treatment of postmenopausal osteoporosis(PMOP)based on the gut microbiota.Methods Sixty SPF grade SD rats were randomly divided into Control group(Control),Model group(Model),GSYG low-dose group(GSYG-L,1.25 g·kg^(-1)),medium-dose group(GSYG-M,2.5 g·kg^(-1))and high-dose group(GSYG-H,5 g·kg^(-1))and alendronate group(Al,0.89 mg·kg^(-1)),with 10 rats in each group.The changes of bone histomorphology were detected by HE staining and the changes of femur tissue structure were observed by micro-CT.The fecal samples from the colon of Control group,Model group,GSYG-H group were taken to extract the DNA of fecal samples.The Illumina Miseq platform was used to carry out high-throughput sequencing.Results Compared with the Control group,bone trabecula in the Model group was sparse,BV/TV,Tb.Th,Tb.N and BMD decreased(P<0.01),while SMI and Tb.Sp increased(P<0.01);compared with the Model group,the number of bone trabeculae in GSYG-L,GSYG-M,GSYG-H group and Al group increased,the bone microstructure was improved,BV/TV,Tb.Th,Tb.N and BMD increased significantly(P<0.05),and SMI and Tb.Sp increased(P<0.05).Sequencing results of gut microbiota showed that,compared with the Control group,the gut microbiota diversity of Model group decreased,the flora abundance of Firmicutes decreased,while the Bacteroidetes abundance increased.Compared with the Model group,the Firmicutes abundance increased and Bacteroidetes abundance decreased in GSYG-H group.Conclusions GSYG can improve bone microstructure and increase bone mineral density,and its mechanism may be related to increasing the diversity of gut microbiota and regulating the structure of gut microbiota.