CAJ | 학술논문

背景：骨骼肌卫星细胞是一种具有多向分化能力的全能干细胞，存在于骨骼肌间质中，对缺血、缺氧有一定的耐受力，是干细胞工程中重要来源细胞。 　　目的：为联合基因工程细胞心肌成形治疗初步探讨较为简便、经济的骨骼肌卫星细胞体外培养方法，建立一种简单、高效的转染骨骼肌卫星细胞的方法及探讨转染后基因表达的特点。 　　方法：分离、培养兔大腿骨骼肌卫星细胞，用CKK-8法测定其生长曲线。根据质粒和脂质体不同比例分组，用脂质体介导增强型绿色荧光蛋白质粒(plasmid enhanced green fluorescent protein，pEGFP)转染骨骼肌卫星细胞。测定各组转染效率及目标基因表达特点。 　　结果与结论：成功分离培养骨骼肌卫星细胞及转染pEFGF。在合适的质粒和脂质体比例下，转染效率可达35%以上。目标蛋白在转染12 h内开始表达，48-72 h表达最强，1周后逐渐减弱，2周后仍可观察到其表达。阳离子脂质体可介导pEGFP高效转染骨骼肌卫星细胞，转染效率与质粒、脂质体比例密切相关，目标基因表达随时间改变。
배경：골격기위성세포시일충구유다향분화능력적전능간세포，존재우골격기간질중，대결혈、결양유일정적내수력，시간세포공정중중요래원세포。 　　목적：위연합기인공정세포심기성형치료초보탐토교위간편、경제적골격기위성세포체외배양방법，건립일충간단、고효적전염골격기위성세포적방법급탐토전염후기인표체적특점。 　　방법：분리、배양토대퇴골격기위성세포，용CKK-8법측정기생장곡선。근거질립화지질체불동비례분조，용지질체개도증강형록색형광단백질립(plasmid enhanced green fluorescent protein，pEGFP)전염골격기위성세포。측정각조전염효솔급목표기인표체특점。 　　결과여결론：성공분리배양골격기위성세포급전염pEFGF。재합괄적질립화지질체비례하，전염효솔가체35%이상。목표단백재전염12 h내개시표체，48-72 h표체최강，1주후축점감약，2주후잉가관찰도기표체。양리자지질체가개도pEGFP고효전염골격기위성세포，전염효솔여질립、지질체비례밀절상관，목표기인표체수시간개변。
BACKGROUND:Skeletal muscle satel ite cells are totipotential stem cells with multi-directional differentiation potential, locate in skeletal muscle interstitium, have a certain tolerance to ischemia and hypoxia, and are important cells in stem cellengineering. OBJECTIVE:To establish a thrifty, convenient culture procedure and create a simple, efficient method to transfect skeletal muscle satel ite cells, and investigate genetic expression after the transfection for cellular cardiomyoplasty. METHODS:Skeletal muscle satel ite cells were isolated from rabbit thigh and cultured. Their growth curves were determined by CKK-8 method. Grouped by different proportions of the plasmid and liposome, skeletal muscle satel ite cells were transfered by the enhanced green fluorescent protein plasmid based on liposome. After transfection, the efficiency and character of target genetic expression was determined. RESULTS AND CONCLUSION:Satel ite cells were isolated, cultured and transfected successful y. In suitable ratio of plasmid and liposomes, the transfection efficiency reached up to above 35%. The target protein was expressed within 12 hours after transfection, reached maximum in 48-72 hours and decreased gradual y after one week. The expression stil could be observed two weeks latter. The enhanced green fluorescent protein plasmid conducted by cationic liposome could be transfered into skeletal muscle satel ite cells efficiently. The transfection efficiency was correlated closely to the ratio of plasmid and lipofectamine. The change of target gene expression depended on time.