大鼠脂肪源性干细胞分离培养的实验研究

THE STUDY OF ISOLATING AND CULTURING ADSCS FROM RATS

目的研究分离、培养、鉴定大鼠脂肪源性干细胞的实验方法。方法提取大鼠双侧腹股沟处脂肪组织,进行原代培养及流式细胞检测仪的鉴定。用MTT法检测不同原代细胞接种密度的细胞分裂增殖率的变化,并观察第1-第13代细胞分裂增殖的特点。结果大鼠脂肪源性干细胞的生长呈大量细胞集落,表面标记物CD44、CD105、CD49d表达阳性,CD106表达阴性。不同的原代细胞接种密度会影响细胞分裂增殖,以1×106/ml细胞密度接种时细胞的增殖速率高于其它对照组。该细胞株经多次传代后仍能保持较强的分裂增殖能力。结论大鼠脂肪源性干细胞分离培养方法简便,不同的原代细胞接种密度会影响细胞分裂增殖,细胞经多次传代后仍能保持较强的分裂增殖能力。

Objective establish a simple and effective method to isolate and culture Adipose-derived stem cells（ ADSCs）. Methods get the adipose from groins of the rats,use the collagenase type I to digest and isolate,inoculate Primary cells into the culture flask with different concentrations,assay the cell with the MTT in 1 - 7 days,identify the cell surface marker of CD44、CD105、CD106、CD49d by flow cytometry instrument,obverse the form of the cells with HE staining,use MTT to determin the chang of the cell proliferation rate with different cell seeding concentration,use use MTT to determin the growth curve of the cell of P1- P13. Results The surface marker CD44、CD49d、CD105 are Positive,CD106 is negative. MTT growth curves suggest that the ability of the cell proliferation remain strong after repeated passage. MTT growth curves aslo suggest that the cells were seeding in different concentrations will affect cell proliferation,Proliferation rate is higher than other control group after sedding cell concentration of 1 × 106ml. Conclusion The method of isolating and culturing the ADSCs is simple. Adipose tissue is a good source of stem cells.