1,25-二羟维生素D_3协同槲皮素抑制高糖诱导SH-SY5Y细胞凋亡的研究

Inhibitory effect of 1,25-dihydroxyvitamin D_3 combined with quercetin on apoptosis of SH-SY5Y cells induced by high glucose

目的利用高糖环境中的神经母瘤SH-SY5Y细胞产生炎性反应制备糖尿病神经受损细胞模型,观察1,25-二羟维生素D_3协同槲皮素对高糖环境中神经细胞保护作用,探讨其分子机制。方法采用100mmol/L葡萄糖模拟神经瘤细胞高糖环境,噻唑蓝(MTT)观察不同槲皮素浓度(0、25、50、100、200μmol/L)对细胞的增殖影响。蛋白免疫印迹(Western blot)观察Bcl,Bax,MMP-9蛋白表达,流式细胞仪Annexin-FITC/PI方法分析细胞凋亡,酶联免疫吸附测定法检测细胞上清肿瘤坏死因子-α(TNF-α)和白细胞介素-1β(IL-1β)的表达。结果 100mmol/L葡萄糖明显诱导SH-SY5Y细胞凋亡,MMP-9蛋白表达升高,分泌炎症因子,表现为神经细胞受损。加入1,25-二羟维生素D_3(1nmol/L)和25、50μmol/L槲皮素及两者联合用药可以明显逆转这些变化,其中槲皮素剂量依赖性抑制细胞凋亡和炎症因子表达。结论槲皮素和1,25-二羟维生素D_3能有效保护高糖引起的神经细胞凋亡受损,联合用药有明显的协同作用,实验对指导糖尿病患者使用1,25-二羟维生素D_3联合槲皮素预防糖尿病患者神经损伤有积极指导意义。

Objective To study SH-YSY cells under high glucose to produce inflammatory reaction to pre- pare diabetic nerve damage cell model and research 1,25-dihydroxyvitamin D3 and quercetin synergistic effect on nerve cell protection and its molecular mechansms. Methods MTT had been imployed to observe cell pro- liferation in quercetin （0,25,50,100,200 μmol/I.） and 100 mmol/L glucose. Western blot had been used to obzerva,e Bcl,Bax and MMP9 protein expression,flow cytometry,Annexin,FITC/PI had been use to analysis cell apoptosis, HLISA had been imployed to detect TNF-α and IL-1 beta expression. Results 100 mmol/L glu- cose induced apoptosis in SH-SYSY cells,MMP-9 protein expression increased the secretion of inflammatory cytokines,manifested as nerve cells. Adding glucose with different concentrations of quercetin 25,50 /μmol/L and 1,25-dihydroxyvitamin D3 （1 nmol/L） high glucose can significantly reverse the impact of these changes, the qucrcetin dose dcpcndently inhibited the apoptosis and expression of inflammatory cytokincs. Conclusion quercetin can effectively protect nerve cells from damage induced by high glucose, vitamin D3 has obvious effect on synergy,combine with quercetin in patients with diabetes guidelines for diabetes injury prevention, actively guide the nerve with 1,25-dihydroxycholecalciferol.