不同表面处理钛片对成骨细胞骨架影响的研究

Effects of different titanium surfaces on F-actin cytoskeleton of osteoblast

目的研究喷砂、酸蚀、碱热处理的钛片表面对成骨细胞F-actin骨架的影响。方法将纯钛钛片根据处理方法不同分为6组:机械打磨组(G组)、喷砂组(SB组)、酸蚀组(SLA组)、光滑碱热组(AH1组)、喷砂碱热组(AH2组)、喷砂酸蚀碱热组(AH3组)。在各组钛片表面培养成骨细胞1、2、4、12 h后,用Phalloidin-TRITC染色,在激光共聚焦显微镜下观察6组钛片表面成骨细胞F-actin骨架的变化。结果成骨细胞接种于6组钛片表面1 h后,各组表面肌动蛋白纤维丝不能清晰看见,SB、AH2、AH3、SLA组肌动蛋白呈环状,纤维分布于粗糙表面隆起部分的边缘。接种2 h后,各组肌动蛋白纤维开始铺展,SB组微丝束有方向性的排列;G组细胞的肌动蛋白纤维成束,排列于细胞周围,有沿沟纹方向铺展趋势;AH1组细胞肌动蛋白纤维于核周围的环绕核呈平行环状排列,周边纤维呈放射状分布;AH2、AH3组纤维呈网状排列,周边纤维汇集呈指状突起,伸入周围孔洞或附着于表面隆起部分的边缘。接种4 h后,G、AH1组肌动蛋白纤维开始定向平行排列;与细胞长轴一致。接种12 h后,各组表面肌动蛋白纤维完全铺展,互相平行排列,与细胞长轴一致,并横跨整个细胞,止于细胞周缘。SB、AH3、SLA组细胞肌动蛋白纤维汇集成束,伸入周围孔洞或附着于表面隆起部分的边缘,将细胞悬挂于凹窝上方。结论成骨细胞接种于6组钛片后,在不同的表面上肌动蛋白的重组有一定的顺序和形态。SB表面最利于细胞骨架排列及伸展。

Objective To evaluate the effects of grooved,alkali-and heat-treated,acid-etched and TiO2 blasted surfaces of titanium substrates on F-actin cytoskeleton of osteoblasts in vitro.Methods Osteoblasts derived from fetal rat calvarial were cultured on 6 different commercially pure titanium discs-grooved（G）,sandblasted（SB）,sandblasted and acid-etching（SLA） surfaces and alkali-and heat-treated（AH1,AH2,AH3） surfaces.For F-actin cytoskeleton measurement,osteoblasts whose filamentous actin was stained with phalloidin-TRITC were cultured for 1,2,4,12 h,evaluated by CLSM observation.Results Osteoblasts attached to the different types of surfaces after 1 hour culture were similar.The actin cytoskeleton formed a ring of cortical filaments around the nucleus after 1 hour on SB,AH2,AH3,SLA surfaces.Actin filaments condensed along edges of pits.The actin filaments of seeded cells were spread after 2 h.The actin filaments on G formed bundles around the nucleus.The filaments began to parallel to the grooves.On AH1,the fibres formed a ring of cortical filaments around the nucleus with some cytoplasmic fibres radially oriented.On AH2,AH3,SB,the fibres orignised in a cytoplasmic meshwork with fibres which terminate at the ridge of depressions.The cell were suspending itself over the depressed areas.Actin filaments on SB were distinct and well formed that were oriented paralled to one another and the long axis of cells.After 4 h,actin filaments appeared organised in a parallel to one another and the long axis of cells.After 12 h,the actin filaments on all surfaces were well spread and were oriented paralled to another and to the long axis of the cell.The filaments formed bundles which reached to holes or adhered to the ridge of raised points,suspending cells over depressed areas.Conclusion After 12 h,the actin filaments on all surfaces were well spread and were oriented paralled to another and to the long axis of the cell.It was concluded that F-actin cytoskeleton of osteoblasts were spread best on SB surfaces among all surfaces.