基于PPAR-γ/SFRP5信号通路探究三期辨证中药复方对骨质疏松大鼠破骨细胞增殖分化的作用机制  

Mechanism of the three-phase syndrome differentiation Chinese herbal compound on the proliferation and differentiation of osteoclasts in osteoporotic rats based on the PPAR-γ/SFRP5 signal pathway

作　　者：王勤俭 李泊泊 WANG Qinjian;LI Bobo(Henan Provincial Hospital of Traditional Chinese Medicine/The Second Affiliated Hospital of Henan University of Traditional Chinese Medicine,Zhengzhou 450000,Henan,China)

机构地区：[1]河南省中医院(河南中医药大学第二附属医院),河南郑州450000

出　　处：《现代中西医结合杂志》2023年第3期293-301,共9页Modern Journal of Integrated Traditional Chinese and Western Medicine

基　　金：河南省中医药科学研究专项课题(20-21ZYZD03)。

摘　　要：目的 基于过氧化物酶体增殖物激活受体-γ/分泌型卷曲相关蛋白5(PPAR-γ/SFRP5)信号通路探究三期辨证中药复方对骨质疏松大鼠破骨细胞增殖分化的影响。方法 制备骨质疏松大鼠模型,原代分离骨质疏松模型大鼠破骨细胞并培养。取细胞分为对照组(1~6周均用不含任何药物的培养基培养)、一期组(1~2周用含增液汤合益骨汤培养基培养,3~6周用不含任何药物的培养基培养)、二期组(1~2周培养同一期组,3~4周用含归脾汤合益骨汤的培养基培养,5~6周用不含任何药物的培养基培养)和三期组(1~4周培养同二期组,5~6周用含独活寄生汤合益骨汤培养基培养),另取细胞分为sh-PPAR-γ组(细胞中转染pSilencer-shPPAR-γ质粒)、sh-NTC组(细胞中转染pSilencer-non-target control质粒)、sh-SFRP5组(细胞中转染SFRP5-shRNA质粒)、sh-NC组(细胞中转染NC-shRNA质粒)、三期+sh-PPAR-γ组(细胞培养于含增液汤合益骨汤、归脾汤合益骨汤、独活寄生汤合益骨汤的培养基中,同时转染pSilencer-non-target control质粒)、三期+sh-SFRP5组(细胞培养同三期+sh-PPAR-γ组,同时转染SFRP5-shRNA质粒)、三期+sh-PPAR-γ+sh-SFRP5组(细胞培养同三期+sh-PPAR-γ组,同时转染pSilencer-shPPAR-γ质粒和SFRP5-shRNA质粒),TRAP染色检测各组破骨细胞数量,MTT法检测细胞增殖能力,流式细胞术检测细胞凋亡率,Western blot法检测细胞中PPAR-γ、SFRP5、果蝇无翅基因(Wnt)、β-连环蛋白(β-catenin)蛋白表达情况。结果 与对照组比较,一期组、二期组和三期组破骨细胞数量、细胞增殖能力(吸光度值)和细胞中PPAR-γ、SFRP5蛋白表达量均明显降低(P均<0.05),细胞凋亡率和细胞中Wnt、β-catenin蛋白表达量均明显增高(P均<0.05);三期组破骨细胞数量、细胞增殖能力(吸光度值)和细胞中PPAR-γ、SFRP5蛋白表达量均明显低于一期组、二期组(P均<0.05),细胞凋亡率和细胞中Wnt、β-catenin蛋白表达量均明显�Objective It is to explore the effect of the three-phase syndrome differentiation Chinese herbal compound on the proliferation and differentiation of osteoclasts in osteoporotic rats based on the peroxisome proliferator-activated receptor-γ/secreted frizzled-related protein 5(PPAR-γ/SFRP5)signaling pathway.Methods The osteoporosis rat models were prepared,and the osteoclasts of the osteoporosis model rats were isolated and cultured.Some cells were divided into control group(cultured in medium without any drugs for 6 weeks)and phaseⅠgroup(cultured in medium containting Zengye Decoction combined with Yigu decoction from 2 weeks,then in medium without any drugs for 4 weeks),phaseⅡgroup(cultured in the same medium as phaseⅠgroup for 2 weeks,then in medium containing Guipi Decoction plus Yigu decoction for 2 weeks,and in medium without any drugs for anther 2 weeks)and phaseⅢgroup(cultured in the same medium as phaseⅠgroup for 4 weeks,then in medium containing Duhuojijie Decoction plus Yigu decoction for another 2 weeks).The other cells were divided into sh-PPAR-γgroup(transfected with pSilencer-shPPAR-γplasmid),sh-NTC group(transfected with pSilencer-non-target control plasmid),sh-SFRP5 group(transfected with SFRP5-shRNA plasmid),sh-NC group(transfected with NC-shRNA plasmid),phaseⅢ+sh-PPAR-γgroup(cultured in medium containing Zengye Decoction plus Yigu decoction,Guipi Decoction plus Yigu decoction,Duhuojisheng Decoction plus Yigu decoction,and transfected with pSilencer-non-target control plasmid),phaseⅢ+sh-SFRP5 group(cultured in the same medium as phaseⅢ+sh-PPAR-γgroup,and transfected with SFRP5-shRNA plasmid),PhaseⅢ+sh-PPAR-γ+sh-SFRP5 group(cultured in the same medium as PhaseⅢ+sh-PPAR-γgroup and transfected with pSilencer-shPPAR-γplasmid and SFRP5-shRNA plasmid).The number of osteoclasts in each group was detected by TRAP staining,the cell proliferation was detected by MTT assay,the apoptosis rate was detected by flow cytometry,and the expression of PPAR-γ,SFRP5,Drosophila wingless g

关 键 词：骨质疏松症 三期辨证中药复方 过氧化物酶体增殖物激活受体-Γ 分泌型卷曲相关蛋白5 破骨细胞 

分 类 号：R-332[医药卫生]