牛膝多糖调控Wnt/β-catenin通路改善骨质疏松性骨折大鼠骨代谢的研究

Study on Achyranthes Bidentata Polysaccharides Improving Bone Metabolism In Osteoporotic Fracture Rats by Regulating Wnt/Β-Catenin Pathway

目的:研究牛膝多糖对骨质疏松性骨折大鼠骨代谢的改善作用。方法:将50只SD雌鼠随机分为假手术组、模型组、阳性对照组(尼尔雌醇,0.21 mg·kg^(-1))及牛膝多糖高、中、低剂量组(8 g·kg^(-1)、4 g·kg^(-1)、2 g·kg^(-1)),每组10只。除假手术组外,其余大鼠切除两侧卵巢10周后,横向锯断右侧股骨复制骨质疏松性骨折模型,模型制备成功后连续给予相应的药物12周。双能X线骨密度仪检测大鼠股骨骨密度(bone mineral density,BMD)水平;ELISA法检测大鼠骨代谢相关指标血清碱性磷酸酶(alkaline phosphatase,ALP)、骨钙素(bone gla protein,BGP)、Ⅰ型前胶原氨基端前肽(type I pro-collagen propeptide,PINP)、Ⅰ型胶原蛋白交联N端端肽(N-terminal typeⅠcollagen telopeptide,NTx)和抗酒石酸酸性磷酸酶(tartrate-resistant acid phospatage,TRAP)水平;HE染色观察骨组织病理学变化;RT-PCR法检测大鼠骨折端骨组织中β-catenin mRNA、Runx2 mRNA、Osterix mRNA水平;Western Blot法检测大鼠骨折端骨组织β-catenin、胞核β-cate-nin、Runx2、Osterixl及p-β-catenin蛋白表达。结果:与模型组比较,牛膝多糖高、中、低剂量组及阳性对照组显著增加大鼠BMD水平(P<0.05);显著升高大鼠血清ALP、BGP和PINP水平(P<0.05),降低NTx、TRAP水平(P<0.05);显著升高大鼠骨折端骨组织β-catenin mRNA、Runx2 mRNA、Osterix mRNA水平(P<0.05);显著上调大鼠骨折端骨组织β-catenin、胞核β-catenin、Runx2、Osterixl蛋白表达(P<0.05),显著降低p-β-catenin/β-catenin水平(P<0.05);同时骨小梁数量增多,形态增粗,连续性增加。结论:牛膝多糖可改善骨质疏松性骨折大鼠骨代谢,其机制可能与激活Wnt/β-catenin通路有关。

Objective:To study the effect of achyranthes bidentata polysaccharide on bone metabolism in rats with osteoporotic fractures.Methods:50 female SD rats were randomly divided into sham operation group,model group,positive control group(nylestriol,0.21 mg·kg^(-1))and achyranthes bidentata polysaccharide high(8 g·kg^(-1)),medium(4 g·kg^(-1)),and low(2 g·kg^(-1))dose groups,with 10 in each group.Except for the sham operation group,the rest of the rats had their ovaries removed and their femurs were sawn off transversely to prepare an osteoporotic fracture model 10 weeks later.After the model was successfully prepared,the model rats were made corresponding drugs were given to them for 12 weeks.Bone mineral density(BMD)levels in rats were detected by the Dual-energy X-ray bone densitometer.Serum alkaline phosphatase(ALP),bone gla protein(BGP),type I procollagen propeptide(PINP),N-terminal type I collagen telopeptide(NTx),and tartrate-resistant acid phosphatage(TRAP)levels were detected by the ELISA method.Whileβ-catenin mRNA,Runx2 mRNA,Osterix mRNA levels in rat fractured bone tissues were detected by the RT-PCR method.And Western Blot method was used to detectβ-catenin,nucleusβ-catenin,Runx2,Osterix,and p-β-catenin Protein expression.Results:Compared with the model group,In achyranthes bidentata polysaccharide high,medium,and low dose groups and the positive control group,the level of rat BMD(P<0.05)and the level of rat serum ALP,BGP,and PINP(P<0.05)were significantly increased.The levels of NTx and TRAP were reduced.The levels ofβ-catenin mRNA,Runx2 mRNA,and Osterix mRNA in the fractured bone tissue of rats(P<0.05)were significantly increased.And the expression ofβ-catenin,β-catenin,Runx2,and Osterix protein in the fractured bone tissue of rats(P<0.05)was also increased.However,the level of p-β-catenin/β-catenin(P<0.05)was significantly reduced.At the same time,the number of bone trabecula was increased with a thicker shape and better continuity.Conclusion:Achyranthes bidentata polysaccharide can improve bone metabolism in osteoporotic fracture rats,which may be related to the activation of the Wnt/β-catenin pathway.