菟丝子黄酮通过调控ZBED3-AS1影响成骨细胞凋亡

Total flavones from Cuscuta chinensis affect osteoblast apoptosis by regu⁃lating ZBED3-AS1

目的:探讨菟丝子黄酮(TFCC)对长链非编码RNA含BED型锌指蛋白3反义RNA 1(ZBED3-AS1)表达和成骨细胞凋亡的影响,旨在阐明TFCC调控成骨细胞凋亡的分子机制。方法:将成骨样细胞株UMR-106分为对照组、肿瘤坏死因子α(TNF-α)组、TNF-α+TFCC组、TNF-α+pcDNA3.1组、TNF-α+pcDNA3.1-ZBED3-AS1组、TNF-α+TFCC-5+si-NC组、TNF-α+TFCC-5+si-ZBED3-AS1组、TNF-α+TFCC-5+pcDNA3.1组和TNF-α+TFCC-5+pcDNA3.1-ZBED3-AS1组。采用CCK-8法和集落形成实验检测细胞增殖;流式细胞术检测细胞凋亡;Western blot检测caspase-3和p21的表达;RT-qPCR检测ZBED3-AS1的表达。结果:TNF-α处理后UMR-106细胞的活力和集落形成能力降低,凋亡率升高,p21和caspase-3表达升高,ZBED3-AS1表达降低(P<0.05)。TFCC(1、10和100μg/L)干预或过表达ZBED3-AS1后,UMR-106细胞的活力和集落形成能力升高,凋亡率降低,ZBED3-AS1表达增加,从而减轻TNF-α处理对UMR-106细胞增殖和凋亡的影响(P<0.05)。抑制ZBED3-AS1表达减轻了TFCC对TNF-α作用下UMR-106细胞增殖和凋亡的影响(P<0.05)。过表达ZBED3-AS1增强了TFCC对TNF-α作用下UMR-106细胞增殖和凋亡的影响(P<0.05)。结论:TFCC通过上调ZBED3-AS1表达可促进成骨细胞增殖,抑制TNF-α诱导的成骨细胞凋亡。

AIM:To explore the effect of total flavones from Cuscuta chinensis(TFCC)on the expression of long noncoding RNA zinc finger BED-type containing protein 3 antisense RNA 1(ZBED3-AS1)and osteoblast apoptosis,and to elucidate the molecular mechanism of TFCC on osteoblast apoptosis.METHODS:Osteoblast-like cell line UMR-106 was divided into control group,tumor necrosis factor-α(TNF-α)group,TNF-α+TFCC group,TNF-α+pcDNA3.1 group,TNF-α+pcDNA3.1-ZBED3-AS1 group,TNF-α+TFCC-5+si-NC group,TNF-α+TFCC-5+si-ZBED3-AS1 group,TNF-α+TFCC-5+pcDNA3.1 group,and TNF-α+TFCC-5+pcDNA3.1-ZBED3-AS1 group.The cell viability and proliferation were evaluated by CCK-8 assay and colony formation assay,and the apoptosis was analyzed by flow cytometry.The expression of caspase-3 and p21 was determined by Western blot,and the expression of ZBED3-AS1 was examined by RTqPCR.RESULTS:After TNF-αtreatment,the viability and colony-forming ability of the UMR-106 cells were decreased,the apoptotic rate was increased,the expression of p21 and caspase-3 was increased,and the expression of ZBED3-AS1 was decreased(P<0.05).After intervention with TFCC(1,10 and 100μg/L)or over-expression of ZBED3-AS1,the viability and colony-forming ability of the UMR-106 cells were increased,the apoptosis rate was decreased,and ZBED3-AS1 expression was increased,so as to reduce the effects of TNF-αtreatment on UMR-106 cell proliferation and apoptosis(P<0.05).Inhibition of ZBED3-AS1 reduced the effect of TFCC on the proliferation and apoptosis of UMR-106 cells treated with TNF-α(P<0.05).Over-expression of ZBED3-AS1 enhanced the effect of TFCC on the proliferation and apoptosis of UMR-106 cells treated with TNF-α(P<0.05).CONCLUSION:TFCC promotes the osteoblast proliferation and reduces TNF-α-induced apoptosis by up-regulating the expression of ZBED3-AS1.