血红素氧合酶1和骨形态发生蛋白2共表达腺病毒载体构建及其对酒精性股骨头坏死治疗的研究  

Construction of adenovirus vector co-expressing heme oxygenase 1 and bone morphogenetic protein 2 and its treatment for alcoholic femoral head necrosis

作　　者：杜振宁[1] 王冰一 高强 李杰[1] Du Zhenning;Wang Bingyi;Gao Qiang;Li Jie(Second Department of Orthopedics,Henan Provincial People’s Hospital,Zhengzhou 450003,China)

机构地区：[1]河南省人民医院骨二科,郑州450003

出　　处：《中华实验外科杂志》2022年第3期535-537,共3页Chinese Journal of Experimental Surgery

基　　金：河南省医学科技攻关计划项目(162300410110)。

摘　　要：目的探讨血红素氧合酶1(HO-1)和骨形态发生蛋白-2(BMP-2)在酒精作用下对成骨细胞凋亡的影响。方法利用腺病毒载体分别构建HO-1、HO-1+BMP-2过表达成骨细胞株H-FOB1-19、HB-FOB1-19及空载载体对照组NC-FOB1-19,实时荧光定量聚合酶链反应(RT-qPCR)检测成骨细胞中BMP-2和HO-1的表达水平;蛋白质印迹法(Western blot)检测H-FOB1-19、HB-FOB1-19 HO-1、NC-FOB1-19中HO-1及表达水平。90 mmol/L酒精分别处理上述3组成骨细胞,流式细胞术(FCM)检测酒精处理后各组成骨细胞的凋亡率。两组比较采用t检验。结果qRT-PCR表明H-FOB1-19、HB-FOB1-19组HO-1表达量高于NC-FOB1-19组(3.872±0.286、3.940±0.653比1.010±0.139,t=10.220、6.206,P<0.05)];HB-FOB1-19组中BMP-2的表达量明显高于NC-FOB1-19组(3.573±0.744比1.183±0.025,t=4.678,P<0.05);Western blot表明HO-1在HB-FOB1-19、H-FOB1-19中的表达量为NC-FOB1-19中表达量的(2.03±0.24)、(1.75±0.16)倍,BMP-2在HB-FOB1-19、H-FOB1-19中的表达量为NC-FOB1-19中表达量的(1.92±0.21)、(1.18±0.09)倍;流式细胞术检测90 mmol/L乙醇处理NC-FOB1-19、H-FOB1-19、HB-FOB1-19组成骨细胞,NC-FOB1-19组凋亡率明显高于H-FOB1-19、HB-FOB1-19组[(23.12±0.14)%比(18.17±0.19)%、(13.59±1.30)%,t=7.073、12.670,P<0.05],HB-FOB1-19组的凋亡率低于H-FOB1-19组[(18.17±0.19)%比(13.585±1.295)%,t=6.640,P<0.05]。结论成骨细胞中过表达HO-1可降低酒精作用后成骨细胞凋亡率,同时上调HO-1和BMP-2表达上述作用效果更明显。Objective To investigate the effect of heme oxygenase-1(HO-1)and bone morphogenetic protein-2(BMP-2)on osteoblast apoptosis under the effect of alcohol.Methods The HO-1,HO-1+BMP-2 overexpressing osteoblast cell lines H-FOB1-19,HB-FOB1-19 and the empty vector control group NC-FOB1-19 were constructed by adenovirus vector,and real-time fluorescence quantification was performed.The expression levels of BMP-2 and HO-1 in osteoblasts were detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).Western blotting was used to detect HO-1 and its expression level in H-FOB1-19,HB-FOB1-19 HO-1 and NC-FOB1-19.The three groups of osteocytes were treated with 90 mmol/L alcohol,respectively,and the apoptosis rate of osteocytes in each group was detected by flow cytometry after alcohol treatment.Quantitative data between the two groups were compared by t test.Results RT-qPCR showed that the expression levels of HO-1 in H-FOB1-19,HB-FOB1-19 and NC-FOB1-19 were 3.872±0.286,3.940±0.653,1.010±0.139 respectively.The expression of HO-1 in H-FOB1-19 and HB-FOB1-19 was significantly higher than that in NC-FOB1-19(t=10.220,6.206,P<0.05).The expression of BMP-2 in HB-FOB1-19 was 3.573±0.744,which was significantly higher than that in NC-FOB1-19(t=4.678,P<0.05).Western blotting showed that HO-1 was expressed in HB-FOB1-19.The expression level of HO-1 in HB-FOB1-19 amd H-FOB1-19 was(2.03±0.24)times and(1.75±0.16)times that in NC-FOB1-19 respectively,and the expression of BMP-2 in HB-FOB1-19 and H-FOB1-19 was(1.92±0.21)times and(1.18±0.09)times that in NC-FOB1-19 respectively.After treatment of osteoblasts with 90 mmol/L alcohol in NC-FOB1-19,H-FOB1-19,and HB-FOB1-19 groups,the apoptosis rates were(23.12±0.14)%,(18.17±0.19)%and(13.59±1.30)%,respectively.The apoptosis rate in the HB-FOB1-19 group was significantly lower than that in the NC-FOB1-19 group(t=7.073,P<0.05).The apoptosis rate in H-FOB1-19 group was lower than that in NC-FOB1-19(t=12.670,P<0.01).The apoptosis rate in HB-FOB1-19 group was lower t

关 键 词：酒精性股骨头坏死 腺病毒载体 血红素氧合酶1 骨形态发生蛋白-2 凋亡 

分 类 号：R681.8[医药卫生—骨科学]