油酸联合胰岛素和地塞米松诱导hADSCs成脂

Adipogenesis of hADSCs induced by oleic acid combined with insulin and dexamethasone

目的利用油酸联合胰岛素和地塞米松诱导人脂肪间充质干细胞(human adipose derived stromal cells, hADSCs)成脂,探索一种在人体内脂肪细胞生成的诱导条件,更接近人的生物学状态,对疾病预防、治疗具有指导意义。方法从人大腿部吸脂术所抽取的脂肪组织,分离培养hADSCs,并进行鉴定:免疫荧光染色法检测细胞表面分子的表达;进行成脂、成神经诱导。采用25、50和80μmol/L三种浓度的油酸联合胰岛素和地塞米松诱导hADSCs的成脂,油红O染色观察细胞成脂情况。结果分离原代细胞,镜下观察呈长梭形,漩涡状排列,表面标记物CD49d和CD105为阳性表达。细胞成脂诱导后,胞浆出现大量脂滴,油红O染为红色;经神经诱导后,细胞伸出细长突起,突起之间有网状连接。油酸联合胰岛素和地塞米松可成功诱导hADSCs成脂:可见细胞内脂滴形成,油红O染为红色,三种浓度以80μmol/L浓度组油红O阳性细胞率最高。结论成功从人大腿皮下脂肪分离、培养hADSCs,并使用80μmol/L油酸联合胰岛素和地塞米松对其诱导12d,可以获得满意的成脂效果。

Objective To explore an induction condition for the formation of human adipocytes through the adipogenesis of human adipose derived stromal cells(hADSCs) induced by oleic acid combined with insulin and dexamethasone, which is closer to the human biological state, and more instructive for the prevention and treatment of diseases. Methods The adipose tissue extracted from human thigh liposuction was used to isolate and culture hADSCs, and the expression of cell surface molecules was detected by immunofluorescence staining. Adipogenesis and neurogenic induction were performed. The adipogenesis of hADSCs was induced by oleic acid at 25, 50 and 80μmol/L combined with insulin and dexamethasone which was observed by oil red O staining. Results The primary cells were isolated and observed in a long fusiform shape with a swirling arrangement. The surface markers CD49 d and CD105 were positively expressed;a large amount of lipid droplets appeared in the cytoplasm in adipogenesis, and were stained red by oil red O;the cells protruded into elongated protrusions with a network between the protrusions in nerve induction. hADSCs were successfully induced to adipocytes by oleic acid combined with insulin and dexamethasone: intracellular lipid droplet formation was observed and stained by oil red O. The concentration of oil red O positive cells was highest in the three concentrations at 80μmol/L. Conclusion The hADSCs were successfully isolated from human thigh subcutaneous fat, and the adipogenesis of hADSCs was induced successfully by 80μmol/L oleic acid combined with insulin and dexamethasone.