CAJ | 학술논문

目的：探讨坍塌反应调节蛋白5（ CRMP5）对神经元突起生长的作用。方法构建CRMP5真核表达载体，采用基因转染、实时定量PCR和免疫印迹技术评估CRMP5基因表达；以空载体为对照组，设3个复孔，用时差成像技术和突起提取技术观察和检测原代培养海马神经元突起的生长。结果成功构建携带增强型绿色荧光蛋白（ EGFP）标签蛋白的CRMP5的真核表达载体。脂质体转染技术可成功把CRMP5基因导入细胞，转染的细胞CRMP5表达高于空载体对照组；CRMP5蛋白表达于神经元的胞体和突起，尤其是胞体、突起起始处和突起末端高表达。过表达CRMP5可明显促进突起生长，主要表现为突起的生长，并形成丰富的侧枝；定量结果显示，CRMP5过表达的细胞突起的长度逐渐延长，而且较空载体转染细胞增多，差异显著（ P＜0.01）。导入CRMP5的细胞突起提取液的吸光度较对照细胞明显升高（P＜0.01）。结论 CRMP5能促进神经元突起及其分支的生长。
목적：탐토담탑반응조절단백5（ CRMP5）대신경원돌기생장적작용。방법구건CRMP5진핵표체재체，채용기인전염、실시정량PCR화면역인적기술평고CRMP5기인표체；이공재체위대조조，설3개복공，용시차성상기술화돌기제취기술관찰화검측원대배양해마신경원돌기적생장。결과성공구건휴대증강형록색형광단백（ EGFP）표첨단백적CRMP5적진핵표체재체。지질체전염기술가성공파CRMP5기인도입세포，전염적세포CRMP5표체고우공재체대조조；CRMP5단백표체우신경원적포체화돌기，우기시포체、돌기기시처화돌기말단고표체。과표체CRMP5가명현촉진돌기생장，주요표현위돌기적생장，병형성봉부적측지；정량결과현시，CRMP5과표체적세포돌기적장도축점연장，이차교공재체전염세포증다，차이현저（ P＜0.01）。도입CRMP5적세포돌기제취액적흡광도교대조세포명현승고（P＜0.01）。결론 CRMP5능촉진신경원돌기급기분지적생장。
Objective To investigate function of collapsin response mediator protein 5 ( CRMP5 ) on neurite outgrowth.Methods The CRMP5 eukaryotic expression vector was constructed and transfected into hippocampal neurons . The gene transfection, Real-time quantitative PCR and Western blotting were used to detect expression of CRMP 5 protein. The lapse-time imaging and neurite extraction were utilized to show neurite outgrowth and differentiation and 3 double-pored were performed, compared with the vector without CRMP5 gene.Results It was successful to construct the CRMP5 eukaryotic expression vector with an EGFP tag .The lipofectamine effectually transfected CRMP 5 into cultured neurons , and the CRMP5 protein was expressed successfully more than the control cells .CRMP5 protein was abundant in cell body , initiation and end of neurites .Overexpression of CRMP5 in neuronal cells significantly promoted outgrowth neurites , and led to the formation of longer neurites with more branches .Accompanying rapid outgrowth of neurites , branches from original neurites were contributed to form a network .The results of neurite length and extraction showed that neurons overexpressing CRMP5 were possessed more and longer neurites (P<0.01), compared with control cells .Conclusion The results suggest that CRMP5 accelerates not only axonal growth but also branching .