猴骨髓间充质干细胞体外诱导分化为脂肪细胞的实验探讨

发布时间：2018-04-10 11:13

本文选题：骨髓 + 间充质干细胞　；参考：《临床与实验病理学杂志》2013年06期

【摘要】：目的体外定向诱导猴骨髓间充质干细胞(bone marrow mesenhymal stem cells,BMSCs)分化为脂肪细胞。方法抽取2～3岁龄雄性恒河猴髂骨骨髓,应用密度梯度离心法提取单核细胞层进行贴壁培养,获取原代(P0)猴BMSCs后进行传代培养,并进行多向分化能力鉴定。分别取第3代(P3)、第5代(P5)、第8代(P8)猴BMSCs,实验组加入脂肪细胞诱导分化培养液,对照组加入常规培养液。倒置相差显微镜下观察细胞形态变化,诱导3周后行油红O染色,计算脂肪细胞分化率,并比较不同代次之间脂肪细胞分化率之间的差异。结果实验抽取的猴骨髓单核细胞具有成脂及成骨分化能力,从而表明其为多能性干细胞。成脂诱导分化培养后第5天,实验组即可观察到少数细胞胞质内出现脂滴,随后胞质内脂滴数量、大小及含脂滴细胞数量逐渐增加,约3周时达到高峰,随后诱导细胞迅速出现死亡;油红O染色示脂滴呈橙红色;脂肪细胞分化率:P3(48.3±10.0)%,P5(58.7±5.3)%,P8(68.1±5.5)%,差异有统计学意义(P0.05)。对照组未见脂滴形成,油红O染色呈阴性。结论脂肪细胞诱导分化培养液可以在体外将猴BMSCs定向诱导分化为脂肪细胞,脂肪细胞分化率随着传代次数增加而增加。
[Abstract]:Objective to induce bone marrow mesenchymal stem cells (marrow mesenhymal stem cells) to differentiate into adipocytes in vitro.Methods the bone marrow of the iliac bone of male rhesus monkeys aged 2 to 3 years old was extracted by density gradient centrifugation. The monocyte layer was isolated for adherent culture, the primary BMSCs was obtained, and the ability of multi-directional differentiation was identified.The third generation of P3, the 5th generation of P5, the 8th generation of P8) monkey BMSCs were taken respectively. The experimental group was added adipocyte induced differentiation medium, the control group was added to the conventional culture medium.The morphologic changes of adipocytes were observed under inverted phase contrast microscope. After 3 weeks of induction, oil red O staining was used to calculate the differentiation rate of adipocytes and to compare the difference of adipocyte differentiation rates among different generations.Results Mononuclear cells extracted from monkey bone marrow had the ability of adipogenic and osteogenic differentiation, which indicated that they were pluripotent stem cells.Lipid droplets were observed in the cytoplasm of a few cells in the experimental group on the 5th day after adipogenic differentiation, followed by an increase in the number, size and number of lipid droplets in the cytoplasm, and reached a peak at about 3 weeks.The differentiation rate of adipocytes was 48.3 卤10.0%. The differentiation rate of adipocytes was 58.7 卤5.3%. The difference was statistically significant (P 0.05).No lipid droplets were found in the control group, but oil red O staining was negative.Conclusion the adipocyte differentiation medium can induce the differentiation of monkey BMSCs into adipocytes in vitro, and the adipocyte differentiation rate increases with the increase of passage times.
【作者单位】：北京市垂杨柳医院病理科;中山大学医学院病理学与病理生理学教研室;北京航天总医院眼科;
【基金】：国家自然科学基金(30271327)
【分类号】：R329

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