过氧化氢-血清饥饿诱导脂肪组织来源干细胞凋亡模型的建立

目的探讨过氧化氢/血清饥饿(H2O2/SD)法建立脂肪组织来源干细胞(ASCs)凋亡模型的可行性。方法酶消化法及贴壁法从小型猪腹股沟脂肪组织中分离、培养ASCs,流式细胞仪检测ASCs表面抗原CD29、CD31、CD34、CD44、CD45、CD90、CD105、HLA-DR的表达。采用不同浓度H2O2(100μM、200μM、400μM、600μM、800μM)联合SD作用于ASCs 6 h,Annexin V-FITC/PI、罗丹明123检测各组细胞凋亡率及线粒体内跨膜电位(ΔΨm)的变化。结果第4代ASCs表面抗原CD29、CD44、CD90、CD105表达呈阳性,CD31、CD34、CD45、HLA-DR表达呈阴性。与对照组比较,不同浓度的H2O2/SD干预6 h均可引起ASCs不同程度的凋亡,而细胞凋亡率最明显的是200μM组(P<0.05),该浓度组也引起ΔΨm明显下降(P<0.01)。结论过氧化氢联合血清饥饿是一种简单有效诱导干细胞凋亡的方法。

Hydrogen peroxide combined serum deprivation-induced apoptosis in adipose tissue-derived stem cells

Objective To establish an apoptosis model in adipose tissue-derived stem cells （ASCs） by hydrogen peroxide and serum deprivation-induced （H2OJSD） in vitro. Methods The isolation and cultivation of miniswine ASCs was conducted by enzyme digestion and adherent from adipose tissue harvested from inguinal regions. The surface antigens （ CD29, CD31, CD34, CD44, CD45, CD90, CD105, HLA-DR ） of ASCs were examined by flow cytometry. After ASCs were incubated for 6 hours in H202/SD（ 100 μM,200 μM,400 μM,600 μM,800 μM） ,apoptosis was then determined by fluorescence-activated cell sorter （FACS） analysis after staining with Annexin V and propidine iodide（H） and Mitochondrial inner transmembrane potential（△ψm） was detected by FACS analysis after staining with Rhodamine 123. Results The ASCs of the forth passage was positive for CD29, CD44, CD90 and CD105, but negative for CD31, CD34,CD45 and HLA-DR. Compared with control group, the apoptosis rate in 200 μM H2 O2/SD group was increased significently （ P 〈 0.05 ） and △ψm in this group was decreased significently （ P 〈 0.01 ）. Conclusion Hydrogen peroxide combined serum deprivation in vitro in stem cells is a simple and effective method of inducing stem cell apotosis.