高糖对人牙髓细胞增殖及氧化应激的影响

Effects of high glucose on proliferation and oxidative stress of human dental pulp cells

目的探讨高糖对人牙髓细胞(HDPCs)增殖和氧化应激的影响,以探索高糖环境对牙髓的影响机制。方法分离培养牙髓细胞,鉴定组织来源,实验分为4组:低糖组(葡萄糖5.5 mmol/L)、正常组(葡萄糖25 mmol/L)、高糖组(葡萄糖50 mmol/L)、高渗组(与50 mmol/L组渗透压相等,并与5.5 mmol/L组葡萄糖浓度相等),CCK-8法检测HDPCs 1、3、5 d后的增殖水平。DCFH-DA检测细胞内活性氧(ROS)水平,酶标法检测丙二醛(MDA)含量,四唑盐法检测超氧化物歧化酶(SOD)活力。流式细胞术分析各组细胞线粒体膜电位。结果(1)高糖组与正常组相比较,可知高浓度葡萄糖对HDPCs的增殖有明显的抑制作用,有统计学意义(P<0.05)。(2)ROS、MDA水平在50 mmol/L葡萄糖浓度下暴露72 h后显著升高,SOD活力降低,相比正常组具有统计学差异(P<0.05)。(3)流式细胞术检测线粒体膜电位显示高糖组荧光值高于正常组,具有统计学差异(P<0.05)。结论高浓度葡萄糖抑制牙髓细胞增殖,导致HDPCs线粒体膜电位下降,诱导氧化应激产生,说明高血糖环境降低了牙髓细胞的活力,也可能是糖尿病患者牙髓组织发生改变的一个潜在因素。

Objective To investigate the effect of high glucose on the proliferation and oxidative stress of human dental pulp cells(HDPCs),and to explore the influencing mechanism of high glucose environment on dental pulp.Methods Dental pulp cells were isolated and cultured,and the source of tissue was identified.The experiment was divided into four groups:low-glucose group(glucose 5.5 mmol/L),normal group(25 mmol/L),high-glucose group(50 mmol/L),and hyperosmolar group(osmotic pressure was equal to that of the 50 mmol/L group and glucose concentration was equal to that of the 5.5 mmol/L group).CCK-8 kits were used to detect the proliferation level of HDPCs after 1 day,3,and 5 days.DCFH-DA was used to detect intracellular ROS level after treatment with high glucose,the content of malondialdehyde(MDA)was measured by enzyme labeling method,and the activity of superoxide dismutase(SOD)was measured by Water-soluble tetrazolium method.Flow cytometry was used to analyze mitochondrial membrane potential in each group.Results(1)Compared with the normal group,high glucose had significant inhibitory effect on the proliferation of HDPCs(P<0.05).(2)The levels of reactive oxygen species(ROS)and malondialdehyde(MDA)increased significantly after exposure to 50 mM glucose for 72 h,and activity of superoxide dismutase(SOD)decreased.Compared with the normal group,there was a statistical difference(P<0.05).(3)Flow cytometry detection of mitochondrial membrane potential showed that the fluorescence value of high glucose group was higher than that of normal group(P<0.05).Conclusion High glucose can inhibit the proliferation of pulp cells,and lead to the decrease in mitochondrial membrane potential of HDPCs,induce oxidative stress,indicating that the high-glycemic environment reduces the vitality of dental pulp cells,which may also be a potential factor for the change of dental pulp tissues in diabetic patients.