CAJ | 학술논문

目的探讨纤维连接蛋白Hep Ⅱ功能域对体外培养的人眼小梁细胞的作用.方法对照实验研究.在体外培养的人眼小梁细胞中加入纤维连接蛋白HepⅡ功能域片段,孵育18、24 h,通过光镜与电镜观察小梁细胞的形态学变化,同时应用免疫荧光染色观察纤维连接蛋白HepⅡ功能域对体外培养的人眼小梁细胞肌动蛋白、纽蛋白、β-连接蛋白的影响.结果加入纤维连接蛋白HepⅡ功能域片段10～24 h,光镜下观察,可见小梁细胞逐渐发生形态学变化,表现为细胞收缩、变圆、彼此间分离;激光共聚焦显微镜下观察,呈红色细丝状着染的小梁细胞肌动蛋白染色减少;位于肌动蛋白两端绿色点、簇状着染的Vinculin染色明显减少;β-Catenin免疫荧光染色呈绿色点状,在沿细胞相连的胞膜处分布减少.电镜下观察,可见小梁细胞突起间缝隙增宽.加入纤维连接蛋白Hep Ⅱ功能域片段24 h,小梁细胞恢复至加入Hep Ⅱ功能域片段前的状态.结论纤维连接蛋白Hep Ⅱ功能域能够可逆性的阻断人眼小梁细胞肌动蛋白及其细胞连接.通过信号转导并重组细胞骨架肌动蛋白及其细胞连接有可能成为重要的研究方向.
목적 탐토섬유련접단백Hep Ⅱ공능역대체외배양적인안소량세포적작용.방법 대조실험연구.재체외배양적인안소량세포중가입섬유련접단백HepⅡ공능역편단,부육18、24 h,통과광경여전경관찰소량세포적형태학변화,동시응용면역형광염색관찰섬유련접단백HepⅡ공능역대체외배양적인안소량세포기동단백、뉴단백、β-련접단백적영향.결과 가입섬유련접단백HepⅡ공능역편단10～24 h,광경하관찰,가견소량세포축점발생형태학변화,표현위세포수축、변원、피차간분리;격광공취초현미경하관찰,정홍색세사상착염적소량세포기동단백염색감소;위우기동단백량단록색점、족상착염적Vinculin염색명현감소;β-Catenin면역형광염색정록색점상,재연세포상련적포막처분포감소.전경하관찰,가견소량세포돌기간봉극증관.가입섬유련접단백Hep Ⅱ공능역편단24 h,소량세포회복지가입Hep Ⅱ공능역편단전적상태.결론 섬유련접단백Hep Ⅱ공능역능구가역성적조단인안소량세포기동단백급기세포련접.통과신호전도병중조세포골가기동단백급기세포련접유가능성위중요적연구방향.
Objective To determine the effects of Heparin Ⅱ (Hep Ⅱ) domain on cultured human trabecular meshwork (HTM) cells.Methods HTM cells were cultured and treated with Hep Ⅱ domain for 18 and 24 h.The morphological changes in HTM cells were assessed by light and electron microscopy.Changes in cell morphology and the organization of actin cytoskeleton,Vinculin,β-Catenin were assessed by using immunofluorescence.Results Treatment of Hep Ⅱ domains resulted in morphological changes from 10 to 24 h.In light microscopy,cells rounded up,retracted and detached from each other.In high performance liquid chromatography,Hep Ⅱ domains-treated cells showed that actin fibre bundles were highly concentrated at the periphery of the cells with few actin filaments left in this area;decreased vinculin staining was observed toward the cell periphery;decreased β-Catenin staining was also observed around the cell sub-membrane.Transmission electron microscopy showed expended intercellular space.After 24 h,changes of HTM cells were recovered.Conclusions Hep Ⅱ domains reversiblely blocks actin cytoskeleton and cell-junction of HTM cells.Disorganization of actin cytoskeleton and cell-junction in trabecular meshwork through signal transduction may be a useful strategy for the decrease of intraocular pressure.