人脂肪组织源性干细胞分化为神经元样细胞的能力及可塑性

Capability and feasibility of human adipose tissue derived stromal cells differentiating into neuron-like cells

目的:脂肪组织源性干细胞分化为神经元样细胞并表达神经元信号已有报道。实验拟观察人脂肪组织源性干细胞定向诱导分化为神经元样细胞的可能性。方法:实验于2003-09/2005-06在沈阳脑科医院试验中心完成。腹部脂肪由沈阳市第一医院普外科提供,患者对实验知情同意,并经医院伦理委员会批准。提取人脂肪组织分离、原代培养、扩增脂肪组织源性干细胞,以免疫荧光化学染色方法鉴定细胞CD44,CD114,CD34的表达。取第二、三代细胞以PBS缓冲液冲洗后,用含有丁酸酯羟基茴香醚、KCL、丙戊酸、地塞米松、胰岛素的培养液诱导向神经细胞分化,采用免疫荧光化学染色方法鉴定神经元特异性烯醇化酶。结果:原代培养的脂肪组织源性干细胞为梭形散在细胞,培养至7d左右细胞接近融合。荧光倒置显微镜下观察经免疫荧光染色,96.7%细胞呈CD44阳性,CD114抗体、CD34抗体呈阴性。神经元特异性烯醇化酶免疫荧光鉴定可见诱导后细胞出现类似轴突和树突样结构,有类神经元间网络形成,表达神经元特异性标志物神经元特异性烯醇化酶。结论:人脂肪组织源性干细胞可在体外扩增,在一定培养条件下,有分化为神经元样细胞的能力。

AIM： It is reported that adipose tissue derived stromal cells can differentiate into neuron-like cells and express neuron signal. The study aimed to explore the feasibility of inducing the differentiation of human adipose tissue derived stromal cells into neuron-like cells in vitro. METHODS： Experiments were performed at the Experimental Center of Shenyang Brain Hospital from September 2003 to June 2005. Abdominal fat was provided by Department of General Surgery of Shenyang First Hospital. Patients signed informed constents. Experimental procedures were approved by Hospital Ethics Committee. Human adipose tissue derived stromal cells were isolated, cultured and proliferated in vitro, then identified by immunofluorescent staining for CD44, CD114, CD34. Second and third generations cells were incubated with culture medium containing butylated hydroxyani sole, KCL, valproic acid, desamethasone and insulin after washing with PBS, and induced differentiating into neural cells. Neuro-specific enolase was identified by immsunofluorescent staining. RESULTS： Primarily cultured adipose tissue derived stromal cells were fusiform, scattered and fused about 7 days later. 96.7% cells were positive for CD44, and negative for CD114 antibody and CD34 antibody under fluorescent invert microscope. Neuro-specific enolase immunofluorescence showed that adipose tissue derived stromal cells appeared axis-cylinder and dendrite-like structure, and differentiate into neuron-like cells. CONCLUSION： Human adipose tissue derived stromal cells can amplify in vitro, and differentiate into neuron-like cells under a certain condition.