阻断TLR2和TLR4缓解UVB导致的HaCaT细胞损伤作用

Blocking of TLR2 and TLR4 alleviates adverse effects of UVB on HaCaT cells

目的:本文拟探讨阻断TLR2和TLR4能否缓解紫外线(UVB)导致的人角质形成细胞株-HaCaT细胞损伤作用并探讨其可能的作用机制。方法:采用MTT法确定UVB最佳照射剂量以及C_(29)和TAK-242的最佳作用浓度;Western blot检测细胞凋亡、NF-κB和内质网应激相关蛋白的表达,探讨阻断TLR2和TLR4缓解UVB导致的HaCaT细胞损伤的作用机制。结果:90 s后细胞存活率明显低于对照组,确定UVB最佳照射时间为90 s;阻断TLR2、TLR4后,UVB导致的HaCaT细胞损伤降低,且150μmol/L C_(29)和2μmol/L TAK-242效果最好;与TLR2相比,阻断TLR4后凋亡蛋白Caspase-8,氧化应激蛋白NF-κB、pNF-κB以及内质网应激蛋白p-PERK的表达下降显著,二者抑制剂联合使用效果更加显著。结论:阻断TLR2和TLR4可以抑制UVB导致的人角质形成细胞HaCaT的损伤和凋亡,其可能是通过抑制细胞NF-κB通路和内质网应激发挥作用。

Objective:The study was to explore whether the blocking of Toll like receptor-2(TLR2)and TLR4 would alleviate the adverse effects of Ultraviolet radiation B(UVB)on human keratinocyte-HaCaT cells and possible underlying mechanisms.Methods:MTT method was used to determine the most appropriate dose of UVB,appropriate concentration of C_(29) and TAK-242.Western blot was employed to detect the expression of apoptosis related protein,NF-κB and endoplasmic reticulum stress-related proteins to investigate the possible mechanisms underlying the alleviating effects of blocking TLR2 and TLR4 on the damage of HaCaT cells caused by UVB.Results:The irradiation time of UVB was determined at 90 s since the survival rate of the cells irradiated for 90 s was significantly decreased compared with that of the cells from control.Blocking TLR2 and TLR4 reduced the damage caused by UVB to HaCaT cells and 150μmo/l L C_(29) and 2μmo/l L TAK-242 are the best.Compared with blocking TLR2,blocking TLR4 significantly decreased the expression of Caspase-8,NF-κB,p-NF-κB and p-PERK even further.Moreover,the combination of C_(29) and TAK-242 could produce a better effects against UVB damage.Conclusion:Blocking TLR2 and TLR4 could inhibit the HaCaT cell apoptosis and decrease the damage caused by UVB to HaCaT cells;and the possible underlying mechanisms may be that the blocking alleviates the damage through the inhibition of NF-κB pathway and endoplasmic reticulum stress.