遗传霉素(G418)纯化许旺细胞过程中成纤维细胞胞浆空泡化的光镜观察  

Cytoplasm vacuolization of fibroblasts during purification of Schwann cells by geneticin(G418):An optical microscope observation and analysis

作　　者：陈宝[1] 史晓远[1] 向宁[1] 王光林[1] 

机构地区：[1]四川大学华西医院骨科,四川省成都市610041

出　　处：《中国组织工程研究》2012年第14期2593-2596,共4页Chinese Journal of Tissue Engineering Research

基　　金：中国国家自然科学基金资助项目(30772204)~~

摘　　要：背景:以往研究应用遗传霉素(G418)联合差速贴壁及差速分离法除去成纤维细胞用于许旺细胞的纯化,对纯化过程中成纤维细胞形态变化的描述及可能其原理的推测很少。目的:利用光学显微镜采集纯化的不同时期成纤维细胞和许旺细胞形态学照片,描述成纤维细胞病理学变化。方法:利用差速贴壁、差速分离法结合遗传霉素抑制来纯化大鼠坐骨神经来源的许旺细胞。将传代培养的细胞分2组,实验组:细胞重复一次差速贴壁后转移到另一六孔培养板,每2d更换新鲜纯化培养液,用差速分离法传代。对照组:差速分离纯化1次后转移到另一六孔培养板,每2d更换新鲜基础培养液,用差速分离法传代。在纯化的不同时期,利用光学显微镜观察成纤维细胞病理学变化。结果与结论:在纯化的早期,光学显微镜下不能观察到成纤维细胞的病理变化,但成纤维细胞的增殖速度减慢,持续纯化经过3代(三四周)之后,光镜下可观察到逐渐加重的细胞病理变化,直至细胞空泡化。结果表明,应用遗传霉素纯化许旺细胞过程中,成纤维细胞增殖受限较早,但是胞浆的空泡化延迟至第3次传代及以后发生。BACKGROUND：Geneticin（G418） combined with differential adhesion method and differential detachment method are used to eliminate the contaminated fibroblasts in the purification of Schwann cells.Most of the researches prefer to describe purity of Schwann cells,rather than give a description of cell morphology change and its probable cause during the purification process.OBJECTIVE：To describe the pathological changes of fibroblasts through the morphological photos of purified fibroblasts and Schwann cells at different time points taken by optical microscope.METHODS：Schwann cells were isolated from sciatic nerves of rats by geneticin（G418） combined with differential adhesion method and differential detachment method in primary cultivation.The cultivated cells were divided into two groups.In the experimental group,the cells were transferred to another 6-well plate after once differential adhesion,and the waste product accumulated medium was changed to fresh basal medium every 2 days,the differential detachment method was used to passage.In the control group,cells were given differential detachment once again and transferred to another 6-well tissue culture plate,and the waste product accumulated medium was changed to fresh purification medium every 2 days,the differential detachment method was used to passage.Cytopathologic changes of fibroblasts in different periods were observed by optical microscope.RESULTS AND CONCLUSION：Cytopathologic changes of fibroblasts could not be observed under optical microscope in early stage of the purification,the proliferation of the fibroblasts was decreased and the cytoplasm vacuolization of fibroblasts was observed obviously after cells were sub-cultured for three times or more（three to four weeks）.Fibroblast proliferation was restricted in early stage of the Schwann cells purification process by geneticin（G418）,but cytoplasm vacuolization of fibroblasts delayed to the third generation or more.

关 键 词：遗传霉素(G418) 成纤维细胞 细胞空泡化 许旺细胞 光学显微镜 

分 类 号：R394.2[医药卫生—医学遗传学]