二甲双胍对糖基化终末产物诱导的成骨细胞氧化应激和凋亡的影响

目的: 观察二甲双胍对糖基化终末产物(AGEs)诱导的大鼠颅骨成骨细胞内氧化应激产物活性氧簇(ROS)、细胞早期凋亡的影响。方法: 分离培养大鼠颅骨成骨细胞,2′7′-二乙酰二氯荧光素(DCFH-DA)作为荧光探针,流式细胞检测技术检测细胞内ROS水平,膜联蛋白V-异硫氰酸荧光素/碘化丙啶(Annexin V-FITC/ PI)双染色分析细胞早期凋亡率。结果: 与未经葡萄糖处理的牛血清白蛋白(BSA)组对比,500 μg/mL AGEs显著促进成骨细胞内ROS形成和细胞凋亡(均P＜0.01);给予二甲双胍 (100~500 μmol/L)呈浓度依赖性抑制BSA组及AGEs组成骨细胞内ROS形成和细胞凋亡,浓度分别为500、400 μmol/L 时,对细胞内ROS形成和细胞凋亡的抑制作用达到最强。结论: AGEs显著诱导原代成骨细胞内ROS的形成和细胞凋亡,而二甲双胍能够呈浓度依赖性抑制AGEs诱导的成骨细胞内ROS的形成和细胞凋亡,减轻AGEs对成骨细胞的损害。

Effects of metformin on intracellular reactive oxygen species and apoptosis induced by advanced glycation end products on rat cranioaural osteoblasts

AIM: To explore the effects of metformin (MF) on intracellular reactive oxygen species (ROS) and apoptosis in advanced glycation end products (AGEs)-induced rat cranioaural osteoblasts.METHODS: Rat cranioaural osteoblasts were isolated and cultured. The fluorescence of 2′, 7′-dichlorofluorescin (DCF) was measured by flow cytometry as a mean of estimating the formation of ROS, the cell apoptosis was detected by double-staining (Annexin V-FITC/ PI).RESULTS: With 500 μg/mL AGEs, intracellular ROS and the cell apoptosis were markedly increased(both P＜0.01), as compared with that observed with bovine serum albumin(BSA) group; MF(100-500 μmol/L) decreased intracellular ROS in both BSA group and AGEs group in a dose-dependent manner, with maximal inhibition attained at a concentration of 500 μmol/L. Introducing different concentrations of metformin (100-500 μmol/L) into the medium significantly lowered the number of apoptotic cells in a dose-dependent manner in both BSA group and AGEs group and maximal inhibition was reached at a concentration of 400 μmol/L.CONCLUSION: AGEs markedly induced intracellular ROS and the cell apoptosis. However, metformin significantly decreased intracellular ROS and apoptosis, which could ameliorate deleterious effects of AGEs on osteoblasts.