牙髓卟啉单胞菌促成骨细胞凋亡的机制初探

摘要

目的：观察牙髓卟啉单胞菌超声提取物对成骨细胞凋亡的促进作用，并初步探讨其相关分子机制。方法：用浓度为1、10、100μg/mL牙髓卟啉单胞菌超声提取物作用于成骨细胞，分别采用Hoechst33258染色法观察细胞的形态变化；比色法检测细胞中的Caspase－3酶活性改变；同时通过Caspase抑制剂阻断实验分析相关成骨细胞的凋亡途径。结果：牙髓卟啉单胞菌超声提取物能使成骨细胞形态出现染色质浓缩、碎裂等典型的凋亡改变，并可显著提高成骨细胞的Caspase－3酶活性，且具有浓度依赖性（P＜0．05）；Caspase－8抑制剂和Caspase－9抑制剂可以部分抑制牙髓卟啉单胞菌提取物对成骨细胞的促凋亡作用，而Caspase－3抑制剂则能完全阻断牙髓卟啉单胞菌对成骨细胞的促凋亡作用。结论：牙髓卟啉单胞菌超声提取物具有促成骨细胞凋亡的作用，细胞内、外凋亡途径都参与了该过程，并且是一种依赖Caspase－3的凋亡通路。%AIM:To investigate the apoptosis of human osteoblastic hFOB 1.19 cells induced by sonicated extract of P.endodontalis and the relevant molecular mechanism.METHODS:After exposure to sonicated extract of P.endodontalis at 0 (the control),1,10 and 100 μg/mL for 48 h,the morphology changes of hFOB 1.19 cells were observed by Hoechst33258 staining.Caspase-3 activity was detected by colorimetric assay .The apoptosis rate was an-alyzed by flow cytometry after the cells had been treated with the extract of P.endodontalis plus Caspase inhibitors.RE-SULTS:The sonicated extract of P.endodontalis induced chromatin concentration,breakup and other typical apoptosis changes in hFOB 1.19 cells.Meanwhile,the extract increased Caspase-3 activity of hFOB 1.19 cells in a dose-de-pendent manner(P<0.05)Caspase-8 inhibitor and Caspase-9 inhibitor partially inhibited the apoptosis of hFOB 1. 19 cells induced by the extract.Caspase-3 inhibitor completely blocked the apoptosis of hFOB 1.19 cells.CONCLU-SION:The extract of P.endodontalis can promote hFOB 1.19 cell apoptosis.Caspase-3-dependent apoptosis pathway is involved in this process,including both internal and external pathway.