大鼠胚胎皮肤成纤维细胞培养鉴定及真皮层内移植存活性的实验研究

发布时间：2018-05-13 14:48

本文选题：成纤维细胞 + 细胞　；参考：《山东大学》2006年硕士论文

【摘要】：目的：利用体外组织细胞培养及胚胎无瘢痕愈合的一般原理，在体外原代培养胚胎大鼠皮肤成纤维细胞，探索和建立大鼠胚胎皮肤成纤维细胞体外分离、培养及鉴定的技术方法。观察大鼠胚胎皮肤成纤维细胞在正常皮肤以及深二度烧伤创面注射移植后的存活情况。方法：分别用组织块法和酶消化分离法原代培养处于无瘢痕愈合期的Wistar大鼠胚胎皮肤成纤维细胞，对两种方法获得细胞的情况进行比较。用胰蛋白酶+EDTA消化加反复贴壁法分离纯化成纤维细胞。倒置相差显微镜下观察原代培养及传代过程中成纤维细胞的形态学特征。制备细胞爬片，对培养的成纤维细胞用SP方法作免疫细胞化学染色(immunocytochemistry，ICC)，第一抗体分别为波形蛋白(vimentin)、结蛋白(desmin)、角蛋白(pan cytokeratin)。选取生长状态良好的细胞，制作标本，用透射电子显微镜(transmission electron microscope，TEM)观察胚胎成纤维细胞的超微结构。用组织块法和酶消化法对成年Wistar大鼠皮肤成纤维细胞进行培养，并对两类细胞生长以及功能活动情况进行简单的比较研究。细胞计数法绘制两种细胞的生长曲线。流式细胞术(flow cytometry，FCM)测定比较处于对数生长期的两种细胞的细胞周期情况，细胞各周期的百分比测定结果以(?)±S表示，作组间t检验进行统计学分析，，P＜0.05即认为有统计学意义。选择成年健康Wistar大鼠12只，每只体重约250g左右，随机分为两组，A组为正常皮肤移植组，B组为烧伤创面移植组。A组每只背
[Abstract]:Objective : To explore and establish a technique for the in vitro isolation , culture and identification of rat embryonic skin fibroblasts by using the general principles of in vitro tissue culture and embryo - free healing , and to observe the survival of rat embryonic skin fibroblasts after injection transplantation in normal skin and deep second degree burn wounds .

Methods : The embryonic skin fibroblasts were cultured in Wistar rats without scar healing stage by tissue block method and enzyme digestion separation method . The cells were isolated and purified by trypsin + EDTA digestion and repeated adherent method . The cells were cultured and passaged by reversed phase contrast microscope . The cells were stained by SP method for immunohistochemical staining ( ICC ) . The first antibody was vimentin , vimentin , pan cytokeratin , respectively . The ultrastructure of embryonic fibroblasts was observed by transmission electron microscope ( TEM ) .

The growth curve of two kinds of cells was studied by using tissue block method and enzymatic digestion method . The growth curve of two kinds of cells was studied by cell counting method . Flow cytometry ( FCM ) was used to measure the cell cycle of two kinds of cells .

12 adult healthy Wistar rats were randomly divided into two groups : group A , normal skin transplantation group , group B as normal skin graft group and group B as burn wound transplantation group .

【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R329.2

【引证文献】