目的:探索成纤维细胞生长因子10(fibroblast growth factor 10,FGF10)对脂多糖(lipopolysaccha-ride,LPS)刺激下小胶质细胞BV2活化的影响.方法:小鼠BV2小胶质细胞用细胞培养基DMEM培养,置于37℃、5%CO2、饱和湿度的培养箱中培养,1~2 d换液,4~5 d传代.实验分为对照组、LPS组和FGF10组,FGF10组的BV2细胞预先给予FGF101 mg/L 30 min后,在LPS组和FGF10组中加入500 mg/L的LPS,在不同时点进行检测.用倒置显微镜观察小胶质细胞的形态学改变,RT-qPCR和ELISA分别检测肿瘤坏死因子α(TNF-α)转录和蛋白表达水平的改变来观测BV2细胞的活化情况.结果:静息状态下BV2细胞形态呈圆形或椭圆形,经过24 h LPS刺激后,BV2细胞形状向多极或纺锤样改变,活化细胞数量比值明显高于对照组;预先给予FGF10能抑制LPS刺激下的BV2细胞向活化形态改变,活化的BV2细胞明显减少.给予LPS刺激6 h后,LPS组TNF-α的mRNA水平相比于对照组显著升高,然而预先给予FGF10会显著抑制TNF-α的转录.LPS作用24 h后,细胞培养上清液内TNF-α的表达水平与对照组相比显著上升,而预先给予FGF10在蛋白水平显著抑制TNF-α的表达.结论:FGF10能够成功抑制LPS刺激下BV2细胞的活化,有望成为治疗经胶质细胞介导的神经系统炎症性疾病的一种有效药物.%AIM: To investigate the effects of fibroblast growth factor 10 ( FGF10 ) on lipopolysaccharide ( LPS)-induced microglial activation .METHODS:Mouse BV2 microglial cells were maintained in DMEM in a humidified incubator with 95%/5%( V/V) mixture of air and CO 2 at 37℃.The medium was changed every 1 or 2 d.The cells were digested and passaged every 4 or 5 d.The BV2 microglial cells were first pretreated with FGF 10 (1 mg/L) for 30 min and then stimulated with LPS (500 μg/L).The medium and the cells were collected at different time points .The morphologi-cal changes of microglia were visualized under microscope .To evaluate the microglial activation , the transcription and pro-duction of proinflammatory factor tumor necrosis factor-α( TNF-α) were examined by real-time quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively.RESULTS:The morphology of control BV2 microglia showed circular or oval shape .After exposure to LPS for 24 h, the microglia revealed spindle shaped or multipolar morphology , and the percentage of activated cells was significantly increased compared with control group.Pretreatment with FGF10 successfully inhibited the morphological change from normal to activated shape .LPS sti-mulation for 6 h significantly increased the transcription of TNF-α, while FGF10 pretreatment remarkably reversed the effect.In addition, the production of TNF-αincreased in the presence of LPS stimulation for 24 h compared with control group.Pretreatment with FGF10 suppressed LPS-induced TNF-αexpression.CONCLUSION: Pretreatment with FGF10 inhibits the morphological change from normal to activated shape , and remarkably suppressed the transcription and produc-tion of TNF-α.FGF10 successfully suppresses LPS-induced BV2 microglial activation , indicating that FGF10 is a therapeu-tic agent for the treatment of glia-mediated neuroinflammatory diseases .
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