摘要
目的探讨可溶性RAGE(soluble form of receptor for advanced glycation-end products,sRAGE)对晚期糖基化终产物(advanced glycation end products,AGEs)-晚期糖基化终产物受体(receptor for advanced glycation-end products,RAGE)介导的C57BL/6J小鼠SGCs(spiral ganglion cells,SGCs)凋亡及RAGE受体表达的影响。方法不同浓度的sRAGE作用于培养5天后的螺旋神经节细胞和加入AGEs-BSA作用2 h后的SGCs细胞,采用免疫荧光法鉴定SGCs,通过流式细胞术检测细胞凋亡,以及RT-PCR检测RAGE的mRNA的表达,琼脂糖凝胶电泳对扩增结果进行确认,同时采用western blot技术检测凋亡相关蛋白Bax、BCL-2及caspase-3的表达情况。结果原代培养获得足够数量以及活性良好的SGCs,并用小鼠抗神经微丝蛋白抗体染色以鉴定。单纯向SGCs加入不同浓度的sRAGE,细胞凋亡无明显增加,RAGE受体的表达无明显增多;而不同浓度sRAGE加入AGEs作用的SGCs后,可见SGCs的凋亡随sRAGE浓度增加而减少,呈浓度相关性,RAGE受体的mRNA表达也随之减弱。凋亡相关蛋白中,BCL-2表达升高,Bax及cleaved caspase-3表达降低。结论 s RAGE可以减少AGEs诱导的SGCs的凋亡,并减少膜上RAGE受体的表达,可能与s RAGE竞争性结合AGEs而不产生相应生物学效应有关。
Objective To investigate the effect of sRAGE(soluble form of receptor for advanced glycation-end products) on apoptosis of spiral ganglion cells induced by AGEs(advanced glycation end products) – RAGE(receptor for advanced glycation-end products) and expression of RAGE in 57 BL/6 J mice. Method Cultured SGCs(5 days), with or without 2 hours exposure to AGEs, were treated with sRAGE of various concentrations. SGCs were identified by immunofluorescence. Apoptosis was determined by flow cytometry. mRNA expression for RAGE and BCL-2 was checked by RT-PCR and Agarose electrophoresis. Expression of apoptosis-related proteins Bax, BCL-2 and Caspase-3 were assessed by Western blot. Result The quantity and activity of SGCs in primary culture were adequate as examined immunocytochemically by NF200 monoclonal antibody. There was no significant difference in apoptosis of SGCs without AGE exposure regardless of sRAGE concentrations. sRAGE reduced apoptosis of SGCs previously exposed to AGEs in a concentration-dependent manner. mRNA expression for RAGE was enhanced as sRAGE increased. For apoptosis-related proteins,expression of BCL-2 increased while expression of Bax and cleaved caspase-3 decreased(P<0.05). Conclusion AGEs-RAGE mediated SGC apoptosis and RAGE expression can be inhibited by sRAGE, likely caused by competitive binding between sRAGE and AGEs.
作者
吴小波
林昶
WU Xiaobo;LIN Chang(Department of Otolaryngology,Head&Neck Surgery,First Affiliated Hospital of Fujian Medical University,Fuzhou 350005)
出处
《中华耳科学杂志》
CSCD
北大核心
2019年第6期958-962,共5页
Chinese Journal of Otology
基金
福建省科技合作重点项目2014I0003~~
关键词
SRAGE
AGES
RAGE
C57BL/6J小鼠螺旋神经节细胞
凋亡
Soluble form of receptor for advanced glycation end products
Advanced glycation end products
Receptor for advanced glycation-end products
Mouse spiral ganglion cells of C57BL/6J
Apoptosis
