糖基化终产物对人牙周膜细胞自噬的影响

Effects of advanced glycation end product on autophagy of human periodontal ligament cells

目的研究在体外培养坏境下糖基化终产物(advanced glycation end of products,AGEs)对人牙周膜细胞(hPDLCs)发生自噬现象的影响。方法选取因正畸治疗而拔除的前磨牙,组织块法分离培养人牙周膜细胞,鉴定后传代培养。用不同浓度的AGEs处理hPDLCs,以单纯DMEM完全培养基作为阴性对照组,培养0、1、3、6、9 h,透射电镜观察细胞自噬体数量的变化和形态,Western blot、免疫荧光法检测自噬蛋白LC3表达的改变,检测在体外培养坏境下AGEs对hPDLCs发生自噬现象的影响。结果 AGEs处理hPDLCs0、1、3、6、9 h后,与对照组比较,实验组自噬相关蛋白LC3-Ⅱ的表达显著变化;与0 h比较,1 h时实验组与对照组自噬水平都明显下降,随时间变化对照组的自噬相关蛋白LC3-Ⅱ无明显改变,实验组在3 h时达到高峰,并随时间变化逐渐降低;3 h时电镜观察和免疫荧光检测到实验组细胞胞浆内自噬体数量增加。结论 AGEs能够提高hPDLFs的自噬水平。

Objective To investigate the influence of advanced glycation end products （AGEs） on autophagy of human periodontal ligament cells （hPDLCs） in vitro. Methods The hPDLCs were isolated from premolars extracted for orthodontic purpose and cultured in vitro using tissue culture method,hPDLCs was treated with different concentrations of AGEs and DMEM complete medium was used as the negative control group：AGEs group （200 μg/ml AGEs） , negative control group （bovine serum albumin BSA）. After0, 1,3,6 and 9 h of culture, these results were further substantiated by Western blot analysis, electron microscopy and Immunofluorescence. Re- sults We found that the lysosome activity significantly increased in AEGs group by electron microscopy and Immunofluorescence. Compared with Oh, the LC3-Ⅱ expression was declined in AGEs group and negative control group at lh. But after lh, the LC3-Ⅱ ex- pression elevated and highest at 3h in AGEs group, while it remained unchanged in negative control group. Conclusions AGEs can strongly induce autophagy of the hPDLCs in vitro.