晚期糖基化终末产物AGEs通过牙周膜成纤维细胞的TLR4途径促进牙周炎症

Advanced Glycation End Products Increase Inflammation via TLR4 Pathway in Periodontal Ligament Fibroblasts

目的:探讨糖尿病的重要病理产物晚期糖基化终末产物(AGEs)对牙周膜成纤维细胞(PDLC)中Toll样受体4(TLR4)介导的相关炎症因子表达的作用,以阐明AGEs对牙周炎可能的影响。方法:分别使用不同剂量的AGEs(0、100、300、500 mg/L)处理牙周膜细胞。通过免疫荧光染色α-actin,以检测AGEs对细胞骨架的可能作用。通过Western blot检测不同剂量的AGEs对PLDCs表达TLR4及下游炎症因子IL-1β和TNF-α的影响,以及AGEs抑制剂E5564对TLR4激活的阻断作用。通过免疫荧光再次验证AGEs通过TLR4对IL-1β和TNF-α的激活作用。结果:AGEs对PDLCs的培养和形态并未有影响,对细胞结构蛋白α-actin的表达未见明显影响。Western blot结果显示,与对照组相比,300和500 mg/L的AGEs处理明显增加了PDLCs的TLR4的表达,且300和500 mg/L剂量之间未发现统计学差异。AGEs引起的TLR4的激活继续导致了下游IL-1β和TNF-α的表达上调,Western blot和免疫荧光均证明,加入TLR4特异性抑制剂E5564可以逆转TLR4通路诱导的IL-1β和TNF-α激活。结论:AGEs并未对成纤维细胞的形态和骨架有明显影响,AGEs可能通过TLR4通路促进牙周成纤维细胞的炎症因子IL-1β和TNF-α的释放,促进牙周炎的发展。

Objective:To investigate the effect of advanced glycation end products(AGEs),a product of diabetes,on the expression of Toll-like receptor 4(TLR4)-mediated inflammation in periodontal ligament cells(PDLC).Methods:Different doses of AGEs(0,100,300,500 mg/L)were used to treat PDLCs.α-Actin was stained by immunofluorescence staining to detect the cytoskeleton.Western blot was used to detect the effect of AGEs on the expression of TLR4 and the downstream factors IL-1βand TNF-α,as well as the blocking effect of AGEs inhibitor E5564 on TLR4 activation.Immunofluorescence confirmed the activation of IL-1βand TNF-αvia TLR4 pathway by AGEs.Results:AGEs had no effect on the culture and morphology of PDLCs,and no significant effect on the expression of cell structural proteinα-Actin.300 mg/L and 500 mg/L AGEs significantly increased the TLR4 expression compared with the control group,and no statistical difference was found between the 300 mg/L and 500 mg/L doses.The activation of TLR4 caused by AGEs continued to lead to the up-regulation of IL-1βand TNF-α.Western blot and immunofluorescence proved that the addition of TLR4-specific inhibitor E5564 could reverse the activation of IL-1βand TNF-αinduced by TLR4 pathway.Conclusion:AGEs have no obvious effect on the morphology of fibroblasts.AGEs may promote the release of inflammatory factors IL-1βand TNF-αfrom periodontal fibroblasts through the TLR4 pathway.