【摘要】 目的基于构建Bcl2沉默和过表达腺病毒观察Bcl2对成骨细胞骨形态发生蛋白2(bone morphogenetic protein-2,BMP-2)、骨保护素(osteoprotegerin,OPG)蛋白的调节作用及补肾健脾活血方对其影响。方法将培养的UMR-106细胞分为空载腺病毒组(沉默Bcl2空载腺病毒NC-bcl2,过表达Bcl2空载腺病毒WT-bcl2)、Bcl2腺病毒组(沉默Bcl2腺病毒sh-bcl2,过表达Bcl2腺病毒Ad-bcl2),空载腺病毒+空白血清组(NC-bcl2,WT-bcl2)、空载腺病毒+补肾健脾活血方含药血清组(NC-bcl2+ME,WT-bcl2+ME)、腺病毒+空白血清组(sh-bcl2,Ad-bcl2)、腺病毒+补肾健脾活血方血清组(sh-bcl2+ME,Ad-bcl2+ME),用RT-q PCR检测Bcl2、OPG mRNA的变化、应用免疫印迹(Western Blot)检测Bcl2、OPG、BMP-2的变化。结果 (1)荧光倒置显微镜观察包装腺病毒转染UMR-106细胞;(2)在正常培养基中,与NC-bcl2比较,sh-bcl2可以降低Bcl2、OPG、BMP-2蛋白的表达(P均<0.05),与WT-bcl2比较,Ad-bcl2可以提高Bcl2、OPG、BMP-2蛋白的表达(P均<0.05);(3)与NC-bcl2比较,RTq PCR显示sh-bcl2可以降低Bcl2 mRNA的表达,但OPG mRNA无统计学意义;(4)在大鼠血清干预中,与空载腺病毒比较,补肾健脾活血方含药血清均可以增加BMP-2、OPG蛋白表达(P均<0.05);在sh-bcl2干预的细胞中,补肾健脾活血方含药血清提高BMP-2、OPG蛋白的表达(P均<0.05);在Ad-bcl2干预的细胞中,中药干预后,BMP-2、OPG未见明显变化。结论 Bcl2可以影响BMP-2、OPG蛋白的表达,补肾健脾活血方可能通过作用于Bcl2影响成骨细胞成骨相关蛋白的表达。更多还原

【Abstract】 Objective By the construction of the silencing and overexpression adenovirus of Bcl2,to obverse the effect Bcl2 on bone morphogenetic protein-2(BMP-2) and osteoprotegerin(OPG),and the influence of nourishing kidney,invigorating spleen,and promoting blood flow recipe(Recipe). Methods UMR-106 cells were divided into vector group(silencing vector Bcl2,NCbcl2,and overexpression vector Bcl2,WT-bcl2),Bcl2 adenovirus group(silencing Bcl2,sh-bcl2,and overexpression Bcl2,Adbcl2),vector adenovirus + blank serum group(NC-bcl2 and WT-bcl2),vector adenovirus + Recipe serum group(NC-bcl2 + ME and WT-bcl2 + ME),adenovirus + blank serum group(sh-bcl2 and Ad-bcl2),adenovirus + Recipe serum group(sh-bcl2 + ME and Ad-bcl2 + ME). Bcl2 and OPG mRNA were detected with RT-qPCR. The portion expressions of Bcl2,OPG,and BMP-2 weredetected with Western blotting. Results 1) Adenovirus fluorescent in UMR-106 cells was observed with fluorescent inverted microscope. 2) In normal medium,compared with NC-bcl2,sh-bcl2 reduced the protein expressions of Bcl2,OPG,and BMP-2(P < 0. 05). Compared with WT-bcl2,the protein expressions of Bcl2,OPG,and BMP-2 were significantly improved in Ad-bcl2(P < 0. 05). 3) Compared with NC-bcl2,the mRNA expression of bcl2 decreased in sh-bcl2. However,mRNA expression of OPG had no significance. 4) Compared with vector adenovirus in rat serum intervention,the protein expressions of BMP-2 and OPG increased in Recipe serum(P < 0. 05). The protein expressions of BMP-2 and OPG increased in the Recipe serum and sh-bcl2 intervention(P < 0. 05). The protein expressions of BMP-2 and OPG had no significance in Ad-bcl2 intervention. Conclusion Bcl2 could affect the protein expressions of BMP-2 and OPG,and Recipe may affect the osteoclast-related protein expressions through Bcl2.更多还原