富血小板纤维蛋白对人牙髓干细胞增殖、凋亡及碱性磷酸酶活性的影响

Effect of PRF on proliferation, apoptosis and alkaline phosphatase activity of human dental pulp stem cells

目的探讨富血小板纤维蛋白(PRF)对人牙髓干细胞增殖、凋亡及碱性磷酸酶活性的影响及机制。方法从人牙髓组织中分离出人牙髓干细胞(h DPSCs),h DPSCs随机分为对照组及实验组,CCK8实验检测PRF刺激h DPSCs第0、1、3、5及7天时细胞增殖情况;RT-PCR检测PRF刺激h DPSCs第0、3、7及14天时碱性磷酸酶(ALP)的m RNA表达情况;流式细胞仪检测PRF刺激h DPSCs 7 d后细胞的凋亡情况,Western blotting检测活化的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved Caspase-3)、p38、p-p38蛋白表达。结果实验组第1天时细胞的增殖与对照组比较,差异无统计学意义(P>0.05),第3、5、7天时细胞的增殖均高于对照组,差异有统计学意义(P <0.05);实验组第3天时ALP mRNA表达与对照组比较,差异无统计学意义(P>0.05),第7和14天时ALP mRNA表达均高于对照组,差异有统计学意义(P <0.05);细胞的凋亡率及Cleaved Caspase-3蛋白、p-p38蛋白表达实验组与对照组比较,差异有统计学意义(P <0.05),实验组细胞的凋亡率及Cleaved Caspase-3蛋白表达均下降,p-p38蛋白表达上调,p38蛋白表达两组间比较差异无统计学意义(P>0.05)。结论 PRF可促进hDPSCs的增殖,上调ALP的mRNA表达,抑制其凋亡,其机制与激活p38信号通路有关。

Objective To investigate the effect of PRF on proliferation,apoptosis and alkaline phosphatase activity of human dental pulp stem cells.Methods Human dental pulp stem cells (hDPSCs) were isolated from human dental pulp tissue,and were randomly divided into control group and experimental group.Cells in experimental group were co-incubated with PRF for 7 days.Cell proliferation and apoptosis were detected by CCK8 assay and flow cytometry on day 0,1,3,5 and 7,respectively.Expression of alkaline phosphatase,Cleaved Caspase-3,p38 and p-p38 were measured by RT-PCR and Western blotting on day 0,3,7,14.Results Cell proliferation was enhanced significantly on day 3,5 and 7 compared with control group (P<0.01).Expression of alkaline phosphatase were increased significantly on day 7 and 14 when compared with that in control group (P<0.01).Cells apoptosis rate and expression of cleaved Caspase-3 were decreased while expression of p-p38 was increased significantly compared with those in control group (P<0.01).No significant difference in p38 protein was observed between the two groups (P>0.05).Conclusion PRF promotes the proliferation and inhibits the apoptosis of hDPSCs cells through mediation of p38 signaling pathway.