摘要目的 探讨无血清处理人甲状腺髓样癌细胞(TT细胞)内S100A13及成纤维细胞生长因子1( FGF-1)蛋白释放的机制,初步阐明Ca2+在S100A13及FGF-1蛋白释放过程中的作用.方法 Western印迹检测无血清处理TT细胞S100A13及FGF-1蛋白表达；ELISA检测培养上清液FGF-1蛋白浓度；激光共聚焦显微镜实时动态检测无血清处理TT细胞1h内Ca2+浓度([Ca2+]i)变化；间接免疫荧光观察TT细胞内S100A13及FGF-1蛋白荧光分布.结果 无血清处理TT细胞4h和6h后S100A13及FGF-1蛋白表达降低(P＜0.05或P＜0.0l),而TT细胞培养上清液中FGF-1蛋白浓度升高(P＜0.05或P＜0.01).激光共聚焦Ca2+荧光显像发现无血清处理TT细胞23 min内[Ca2+]i保持相对稳定,23 min后[Ca2+]i迅速上升达到峰值1.6μmol/L,第40 win后下降至一个较低水平,第40 min至第60 min[ Ca2+]i接近0.3～0.6 μmol/L.1h内TT细胞平均[Ca2+]i明显高于正常培养组、EGTA组和BAPTA-AM组.加入钙离子螯合剂EGTA组和BAPTA-AM组TT细胞内的S100A13、FGF-1蛋白表达无明显下降.结论 无血清处理诱导TT细胞内S100A13及FGF-1蛋白的释放与[Ca2+]i变化有关；Ca2+螯合剂EGTA、BAPTA-AM能有效拮抗无血清处理引起的TT细胞[Ca2+]i升高,并抑制S100A13及FGF-1蛋白的释放.

abstractsObjective To investigate the effect of serum-deprivation on the changes of [ Ca2+ ] i and the protein release of S100A13 and fibroblast growth factor 1 ( FGF-1 ) in thyroid cancer TT cell,and to reveal the role of Ca2+in the protein release of S100Al3 and FGF-1.Methods The protein expressions of FGF-1 and S100A13 in TT cells under serum-deprivation were detected by Western blot.The released FGF-1 protein from TT cells in the supernatant fluid was detected by ELISA.Realtime dynamic examinations on the change of 1 h [ Ca2+ ] i in TT cells under serum-deprivation were detected by confocal laser scanning microscopy.Then,the effect of EGTA( 2.5 mmol/ L),BAPTA-AM (2.5 μmol/L)on distributions of the fluorescence of S100AI 3 and FGF-1 in TT cells under serumdeprivation for6 h were detected by indirect immunofluorescence.Results The expressions of FGF-1 and S100A13 in TT cells after serum-deprivation for4 h and 6 h were reduced( P＜0.05 or P＜0.01 ),but the released FGF-1 protein from TT cells in the supernatant fluid was elevated ( P＜0.05 or P＜0.01).Confocal laser scanning of Ca2+ imaging indicated that [ Ca2+ ] i of serum-deprivation TT cells maintained the relative stabilization within 23 win,but the rapid rise of [ Ca2+ ] i achieved peak value 1.6 μmol/L after 30 min,and remained stable for about 17 win,and thereafter 40 win slowly dropped to a low level From 40 win to 60 win the [ Ca2+ ] i was about 0.3-0.6 μ mol/L.The average [ Ca2+ ] i was higher than that in normal group,EGTA group,and BAPTA-AM group within 1 h.The protein expressions of S100A13 and FGF-1 did not drop obviously in EGTA group and BAPTA-AM group.Conlusion The release of S100A13 and FGF-1 from TT cell under serum-deprivation is possibly related with the change of [ Ca2+ ]i.Both Ca2+-chelating agents EGTA and BA PTA-AM are able to inhibit the rise of [ Ca2+ ] i and release of S100A 13 and FGF-1 from TT cells under serum-deprivation.