摘要
目的探讨积雪草酸(AA)对卵巢切除(OVX)诱导的绝经后骨质疏松症(PMOP)小鼠骨质流失的影响,并基于调节性T细胞(Treg)/T辅助细胞17(Th17)平衡和肠道群菌(GM)探讨其对骨骼的保护机制。方法采用双侧OVX法制备PMOP小鼠模型,实验分为假手术组(Sham组)、OVX组和AA组。Micro-CT检测股骨组织的骨参数,包括骨密度(BMD)、骨体积分数(BV/TV)、骨小梁数量(Tb.N)、骨小梁厚度(Tb.Th)、骨小梁分离度(Tb.Sp);苏木素-伊红染色观察股骨组织和结肠组织病理改变;酶联免疫吸附试验检测血清中骨钙素(BGP)、抗酒石酸性磷酸酶5b(TRACP-5b)、白细胞介素(IL)-6、IL-17、IL-10、转化生长因子-β(TGF-β)水平;蛋白免疫印迹检测股骨组织中骨保护素(OPG)、核转录因子-κB受体活化因子配体(RANKL)蛋白表达及结肠组织中闭合蛋白-3(Claudin-3)、闭锁小带蛋白1(ZO-1)、咬合蛋白(Occludin)表达;流式细胞术检测脾中Treg、Th17细胞比例;16S rRNA测序分析GM多样性和组成。结果相比Sham组,OVX组小鼠股骨组织发生明显病理改变,骨小梁失去其完整的结构形态,数目减少、排列稀疏,间隙变大且存在断裂现象;肠黏膜可见局部损伤,肠腺萎缩变形,有少量炎性细胞浸润;BMD、BV/TV、Tb.N、Tb.Th,血清BGP、IL-10、TGF-β水平,股骨组织中OPG蛋白表达,脾中Treg细胞比例及Treg/Th17比值均降低(P<0.05);Tb.Sp,血清TRACP-5b、IL-6、IL-17水平,股骨组织中RANKL蛋白表达,结肠组织中Claudin-3、ZO-1、Occludin蛋白表达及脾中Th17细胞比例均升高(P<0.05);Chao1、Shannon、Simpson、observed_species均降低,Firmicutes/Bacteroidetes(F/B)比值升高(P<0.05)。经AA治疗后,上述现象均得到改善。结论AA可通过调节Treg/Th17平衡及GM来改善OVX小鼠的骨质流失。本研究揭示了AA治疗PMOP的新药理机制,为其成为治疗PMOP的候选药物提供了重要证据。
Objective To investigate the effect of asiatic acid(AA)on ovariectomy(OVX)-induced bone loss in postmenopausal osteoporosis(PMOP)mice and to explore its protective mechanism on bone based on regulatory T cell(Treg)/T helper cell 17(Th17)balance and gut microbiota(GM).Methods PMOP mouse model was prepared by bilateral OVX method and divided into Sham group,OVX group and AA group.Bone parameters,including bone density(BMD),bone volume fraction(BV/TV),trabecular number(Tb.N),trabecular thickness(Tb.Th)and trabecular separation(Tb.Sp),were measured by Micro-CT.The pathological changes of femur tissue and colon tissue were detected by hematoxylin-eosin stain-ing.Serum bone gla-protein(BGP),tartrate-resistant acid phosphatase 5b(TRACP-5b),interleukin(IL)-6,IL-17,IL-10,and transforming growth factor-β(TGF-β)levels were detected by enzyme-linked immunosorbent assay.The protein expression of osteoprotegerin(OPG)and receptor activator for nuclear factor-kB ligand(RANKL)in femur tissue and the expression of Claudin-3,occlusive zonoll(ZO-1)and Occludin in colon tissue were detected by Western blot.The Treg cells'and Th17 cells'proportion of spleen was detected by flow cytometry.The diversity and composition of GM were analyzed by 16S rRNA sequencing.Results Compared with the Sham group,the femoral tissue of OvX group had obvious pathological changes,the trabeculae of bone lost its complete structure and shape,the number of bone trabeculae was reduced and sparsely arranged,and the gap was enlarged and fractured;the intestinal mucosa showed local damage,intestinal gland atrophy and deformation,and a small number of inflammatory cells infiltrated;BMD,BV/TV,Tb.N,Tb.Th,serum BGP,IL-10 and TGF-βlevels,OPG pro-tein expression in femur tissue,Treg cell proportion and Treg/Th17 ratio in spleen were decreased(P<0.05),Tb.Sp,serum TRACP-5b,IL-6 and IL-17 levels,RANKL protein expression in femur tissue,Claudin-3,ZO-1 and Occludin protein expres-sion in colon tissue and the proportion of Th17 cells in spleen were increased(P<0.05).Chaol,Shannon,Simpson and ob-served_species all decreased,Firmicutes/Bacteroidetes(F/B)ratio increased(P<0.05).After AA treatment,the above phe-nomena were improved.Conclusion AA improved bone loss in OVX mice by regulating Treg/Th17 balance and GM.This study revealed a new pharmacological mechanism of AA in the treatment of PMOP,and provided important evidence for it to be a candidate drug for the treatment of PMOP.
作者
王斌
董洪洋
高松
江小红
丛嫣
武传昇
WANG Bin;DONG Hongyang;GAO Song;JIANG Xiaohong;CONG Yan;WU Chuansheng(Department of Rheumatology,Lianyungang Hospital of TCM,Lianyungang,Jiangsu 222000,China)
出处
《中国优生与遗传杂志》
2025年第2期253-259,共7页
Chinese Journal of Birth Health & Heredity
基金
江苏省优势学科建设工程项目(YSHL2107-971)。
