活性维生素D改善高糖诱导髓核细胞损伤及机制研究

The effect of active vitamin D on degeneration of nucleus pulposus cells in high glucose environment

目的探讨活性维生素D对高糖诱导髓核细胞炎症、氧化应激、自噬和凋亡的影响。方法分别于正常糖(正常糖组)和高糖(高糖组)环境下培养体外人椎间盘髓核细胞(nucleus pulposus cells,NPCs),首次培养及加入活性维生素D后,分别检测髓核细胞炎症、氧化应激、自噬和凋亡情况。其中,采用ELISA试剂盒检测IL-1β、TNF-α及H_(2)O_(2),蛋白免疫印迹Western blotting(WB)检测SOD、Caspase3和Atg5、Atg12、LC3等指标。结果与正常糖组相较,高糖组髓核细胞炎症反应及氧化应激增强、细胞自噬降低、凋亡增多,差异均具有统计学意义(P<0.05)。与加入活性维生素D前相较,正常糖组髓核细胞生物特征变化无明显统计学意义(P>0.05),高糖组髓核细胞氧化应激及炎症反应减弱,自噬增强,细胞凋亡减弱,差异均具有统计学意义(P<0.05)。与高糖+维生素D组相比,加入p38MAPK抑制剂和NF-κB抑制剂可以显著抑制维生素D导致的生化指标变化,且差异具有统计学意义(P<0.05)。结论活性维生素D通过激活p38MAPK和NF-κB信号通路增强高糖环境下髓核细胞自噬并抑制炎症和凋亡来保护髓核细胞,延缓椎间盘退变。

Objective To explore the effect of active vitamin D on autophagy,inflammation and apoptosis of nucleus pulposus cells in high glucose environment.Methods In vitro,human nucleus pulposus cells(NPCs)were cultured in normal glucose(normal glucose group)and high glucose(high glucose group)environment,and the autophagy,inflammation and apoptosis of NPCS were detected for the first time.Active vitamin D was added in two groups,and the autophagy,inflammation and apoptosis of NPCs were detected again after continued culture.The factors of IL-1β,TNF-αand H_(2)O_(2)were detected by ELISA,SOD,Caspase 3,Atg5,Atg12 and LC3 were detected by Western blotting(WB).Results Compared with normal glucose group,autophagy of NPCs decreased,oxidative stress and apoptosis increased in high glucose group,with statistical significance(P<0.05).There was no statistically significant difference in the change of biological characteristics of NPCs in the normal glucose group(P>0.05),while the low autophagy of NPCs was improved and enhanced,the oxidative stress and inflammatory response was weakened,and the apoptosis decreased in the high glucose group after adding active vitamin D,with statistical significance(P<0.05).Conclusions Active vitamin D protects NPCs and delays disc degeneration by activating autophagy and inhibiting inflammation and apoptosis in high glucose environment.