ADAM28反义核酸对人牙髓干细胞生物学特性的影响

Effects of ADAM28 AS-ODN on biological property of human dental pulp stem cells(HDPSCs)

目的:探讨金属蛋白酶解离素28(ADAM28)反义核酸(AS-ODN)对人牙髓干细胞(HDPSCs)增殖、分化、凋亡特性的影响并分析可能的作用机制。方法:体外分离培养、鉴定HDPSCs,将FITC荧光标记的ADAM28反义核酸(AS-ODN)和正义对照(S-ODN)分别转染HDPSCs,用半定量反转录聚合酶链反应(RT-PCR)和蛋白印迹法(Westernblot)检测ADAM28AS-ODN转染48h后的封闭效率,应用四唑盐(MTT)比色法、酶动力学法和流式细胞术(FCM)分别检测实验各组HDPSCs的增殖活性、碱性磷酸酶(AKP)的分泌活性以及细胞周期分布和凋亡百分比。采用SPSS13.0软件包的SNK检验进行统计学分析。结果:AS-ODN转染组ADAM28mRNA和蛋白的表达水平均明显下调(P<0.05),细胞增殖活性、增殖指数明显降低(P<0.05),碱性磷酸酶(AKP)分泌水平和细胞凋亡率显著增高(P<0.05)。结论:ADAM28AS-ODN可显著抑制HDPSCs的增殖并影响细胞周期的变化,促进AKP的分泌活性并诱导HDPSCs的凋亡。

Objective：To investigate the effects of a disintegrin and metalloproteinase 28（ADAM28） antisense oligodeoxynucleotide（AS-ODN） on proliferation, differentiation and apoptosis of human dental pulp stem cells（HDPSCs） and possible mechanism.Methods：HDPSCs were isolated culture in vitro and identified.ADAM28 AS-ODN and S-ODN with FITC fluorescence labelling were transfected into HDPSCs respectively.Semiquantitative RT-PCR and Western blot were used to detect the blocking efficiency of ADAM28 AS-ODN after transfected into HDPSCs for 48h.MTT chromatometry, enzyme dynamics method and flow cytometry（FCM） were utilized to respectively determine HDPSCs proliferation activity, secretion activity of alkaline phosphatase（AKP）, cell cycle distribution and apoptosis percentage of each group.Statistical significance was assessed by the Student-Newman-Keuls（SNK） test with SPSS 13.0 software package.Results：In ADAM28 AS-ODN group, the expression levels of ADAM28 mRNA and protein appeared obvious down-regulation（P〈0.05）, HDPSCs proliferation activity and index decreased significantly（P〈0.05）, the secretion level of alkaline phosphatase（AKP） and apoptosis ratio notably raised up（P〈0.05） .Conclusions：ADAM28 AS-ODN could inhibit HDPSCs proliferation and influence the changes of cell cycle,promote AKP secretion activity and induce HDPSCs apoptosis significantly.