摘要
背景:脂肪来源干细胞在体内储备丰富,体外增殖快速,具有多向分化潜能,是目前组织工程种子细胞的研究热点。近年来越来越多的研究表明脂肪干细胞在一定条件下可被诱导分化为内皮细胞,促进血管生成。目的:观察兔脂肪干细胞体外分离培养及诱导分化为血管内皮细胞的生物学特性。方法:取兔附睾处脂肪,采用胶原酶消化分离获得脂肪干细胞,体外培养至第3代后加入血管内皮细胞生长因子与碱性成纤维细胞生长因子联合诱导分化,对诱导前后细胞进行形态学观察、生长曲线测定、免疫组织化学染色及流式细胞仪表型检测。结果与结论:兔脂肪干细胞生长旺盛,第3代兔脂肪干细胞呈成纤维细胞样,生长曲线呈"S"型,15代以内细胞形态未见明显变化。免疫荧光法检测Vimentin阳性,流式细胞仪检测CD44表达阳性,CD31表达阴性;诱导后细胞CD31表达阳性,CD44表达阴性。第3代兔脂肪干细胞向血管内皮细胞诱导分化21d显微镜下呈铺路石样形态,血管内皮细胞第Ⅷ因子相关抗原染色细胞阳性,透射电镜下可见W-P小体。提示脂肪干细胞在体外可诱导分化为血管内皮细胞,可为组织工程血管提供理想的种子细胞。
BACKGROUND： Adipose-derived stem cells are regarded as the potential seed cells for tissue engineering due to abundance in vivo, rapid proliferation in vitro, and capacity of multi-directional differentiation. Accumulated evidence supports that adipose-derived stem ceils can be induced to differentiate into endothelial cells and to promote angiogenesis. OBJECTIVE： To study the biological characteristics of vascular endothelial cells differentiated from rabbit adipose-derived stem cells cultured in ~,itro. METHODS： Adipose tissues were obtained from the epididymal fat pads of the rabbits. And adipose-derived stem cells were isolated from adipose tissues by collagenase digestion and cultured in vitro to passage 3. Vascular endothelial growth factor and basic fibroblast growth factor within endothelial cell growth medium were used to induce adipose-derived stem cells differentiation into endothelial-like cells. Celt morphology was observed, and growth curves were drawn before and after induction. Flow cytometry and immunohistochemistry were used to analyze the morphology and type of adipose-derived stem cells and the differentiated cells. RESULTS AND CONCLUSION： Rabbit adipose-derived stem cells grew well, and passage 3 adipose-derived stem calls presented fibroblast-like growth. The growth curve was like ＂S＂ shape. No significant change in cell morphology was detected within passage 15. Vimentin was positive on passage 3 adipose-derived stem calls by indirect immunofluorescence methods. The positive CD44 expression and negative CD32 expression were detected in passage 3 adipose-derived stem cells by flow cytometric analysis. After induction, CD31 became positive while CD44 wasnegative. Paving stone-like call appearance was seen under inverted microscope 21 days after induction. The differentiated cells were Factor VIII-related antigen positively stained with immunohistological method, and WeibeI-Palade body was observed under a transmission electron microscope. Experimental findings indicate that, adipose-derived stem cells can be induced to differentiate into vascularendothelial calls in vitro, and it can offer the ideal seed cells for tissue engineered blood vessels.
出处
《中国组织工程研究》
CAS
CSCD
2013年第23期4224-4231,共8页
Chinese Journal of Tissue Engineering Research
基金
苏州市社会发展科技项目(SYS201244)
江苏省临床医学科技专项资助项目(BL2012051)~~
关键词
干细胞
脂肪干细胞
血管内皮细胞
组织工程血管
流式细胞术
内皮分化
体外诱导
鉴定
家兔
省级基金
干细胞图片文章
stem cells
adipose-derived stem cells
vascular endothelial cells
tissue engineered blood vesselsflow cytometry
endothelial differentiation
induction in vitro
identification
rabbits
provincial grants-supportedpaper
stem cell photographs-containing.paper
