小鼠脂肪间充质干细胞的分离培养及肠道归巢

发布时间：2018-07-08 17:58

本文选题：脂肪组织 + 间质干细胞　；参考：《中国组织工程研究》2015年06期

【摘要】：背景:脂肪来源间充质干细胞因其取材安全、创伤小、易纯化、增殖快等优点而备受关注。目的:建立一种有效、快速分离培养较高纯度小鼠脂肪间充质干细胞的方法,荧光标记后探索其是否可在体内向肠道归巢。方法:取小鼠腹股沟及附睾脂肪,用0.1%Ⅰ型胶原酶消化得到单细胞,并与剩余未消化脂肪组织块一起接种,贴壁培养获得脂肪间充质干细胞,通过细胞形态、细胞表型、生长动力学、成骨成脂分化潜能4个方面进行鉴定,并将脂肪间充质干细胞用PKH67标记后经尾静脉注射移植到溃疡性结肠炎模型小鼠体内,观察其向肠道的归巢情况。结果与结论:脂肪间充质干细胞镜下呈梭形,快速增殖呈漩涡状排列,细胞高表达CD29、CD44、CD90,不表达CD45。成骨诱导碱性磷酸酶染色显示有黑色颗粒生成,茜素红S染色呈红色结节,成脂诱导油红O染色显示有大量脂滴。细胞生长曲线显示第3-5天为对数增长期,细胞生长活力良好。结肠冰冻切片在荧光显微镜下可见绿色荧光光点,并随时间推移有增加趋势。以上结果表明采用胶原酶消化加组织块贴壁法体外分离培养的脂肪间充质干细胞增殖快、纯度高,可向成骨成脂细胞分化,在体内可向结肠归巢并增殖。
[Abstract]:Background: adipose derived mesenchymal stem cells (MSCs) have attracted much attention due to their advantages of safety, small trauma, easy purification and rapid proliferation. Aim: to establish an effective and rapid method for isolation and culture of adipose mesenchymal stem cells (MSCs) from high purity mice. Methods: the adipose mesenchymal stem cells were obtained from mouse inguinal and epididymal adipose cells after digested with 0.1% collagenase and inoculated with residual undigested adipose tissue. The adipose mesenchymal stem cells were obtained by cell morphology and phenotype. The growth kinetics and adipogenic differentiation potential of adipose mesenchymal stem cells were identified. The adipose mesenchymal stem cells were labeled with PKH67 and injected into the ulcerative colitis model mice via tail vein to observe the homing of adipose mesenchymal stem cells to the intestine. Results and conclusion: adipose mesenchymal stem cells were fusiform under microscope, and rapid proliferation was whirlpool. Osteoblast-induced alkaline phosphatase staining showed black granulation, alizarin red S staining showed red nodules, and lipogenic oil red O staining showed a large number of lipid droplets. The cell growth curve showed the logarithmic growth period on day 3-5, and the cell growth activity was good. Green fluorescent spots were observed in colonic frozen sections under fluorescence microscope and increased with time. The results showed that the adipose mesenchymal stem cells isolated and cultured by collagenase digestion and tissue mass adherence could proliferate rapidly and have high purity, and could differentiate into osteoblasts and homing and proliferating into colon in vivo.
【作者单位】：北京大学航天临床医学院消化科;北京大学人民医院临床分子生物学研究所;
【基金】：航天中心医院特色临床医学发展科研项目(YN2013TS03)《间充质干细胞联合内皮祖细胞治疗小鼠溃疡性结肠炎的疗效及机制研究》~~
【分类号】：R329.2

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