IGF-1基因转染对脂肪间充质干细胞活性的影响及机制研究  

Effects of Insulin-like Growth Factor-1 Transfection on the Bioactivity of Adipose Tissue-derived Mesenchymal Stem Cells and Its Mechanism

作　　者：王菲[1] 蔡晓庆[1] 史亮[1] 陈效安[1] 王益[1] 马凌[1] 

机构地区：[1]兰州军区兰州总医院心血管内科,甘肃兰州730050

出　　处：《现代生物医学进展》2016年第12期2241-2244,2253,共5页Progress in Modern Biomedicine

基　　金：甘肃省自然科学基金项目(1208RJZA213)

摘　　要：目的:探讨携带IGF-1基因慢病毒转染脂肪间充质干细胞(ADMSCs)的可行性,对其引起的细胞凋亡机制做出初步研究,并讨论其与PI3K/Akt通路的关系。方法:分离培养ADMSCs,利用脂质体将携带IGF-1基因的慢病毒载体转染入ADMSCs。实验分为Blank组,Lv-non和Lv-IGF-1三组,转染后用MTT法测定各组细胞的生长情况并绘制生长曲线,流式细胞术测定各组细胞凋亡,Western blot测定各组中IGF-1、Akt、p-Akt及凋亡相关蛋白的表达。结果:成功分离、培养了大鼠脂肪间充质干细胞;携带IGF-1慢病毒载体转染ADMSCs后,流式细胞术检测发现Lv-non和Blank组凋亡率明显高于Lv-IGF-1组(P<0.05)。同时发现Lv-IGF-1组中促凋亡蛋白Caspase-3、Caspase-9蛋白和Bax的表达水平显著下调(P<0.01),抗凋亡蛋白Bcl-2显著上调(P<0.01)。MTT法结果显示Lv-IGF-1能促进细胞生长,在5到6天时显著高于其他两组(P<0.05)。通过检测PI3K/Akt通路发现,Lv-IGF-1组PI3K和p-Akt水平显著高于Blank和Lv-non组(P<0.05),通路被激活。结论:携带IGF-1基因慢病毒载体成功转染ADMSCs。在ADMSCs中过表达IGF-1蛋白可以促进细胞生长,同时抑制细胞凋亡,其机制可能与PI3K/Akt通路蛋白磷酸化相关。Objective： To construct a lentiviral vector expressing gene of insulin-like growth factor-1（IGF-1） and transfect it in cultured ADMSCs and observe the impact of IGF-1 transfection on the bioactivity of ADMSCs and investigate the underlying mechanisms, focusing on its possible relationship with PI3K/Akt cell signaling. Methods： Human-derived ADMSCs were isolated and cultured in vitro, dividing into three groups as： control group, Lv-non and Lv-IGF1 group. Then lentivirus expressing IGF-1 was transfected into ADMSCs. MTT assay was used to detect the proliferation and flow cytometry was used to evaluate IGF1-induced anti-apoptosis effects. Furthermore, the expression of IGF1, Akt, p-Akt and apoptosis related proteins were detected by Western blot.Results： ADMSCs were successfully isolated from Adult abdomen greater omentum adipose tissue and cultured. 48 h after lentivirus carrying IGF1 transfected, green fluorescence was detected. Compared with Lv-non and blank groups, the Lv-IGF1 transfected group showed statistically lower apoptotic rate（P〈0.05）. Compared with control group, the expression of Caspase-3, Caspase-9 and Bax was decreased gradually in ADMSCs transfected with Lv-IGF1（P〈0.01）, while Bcl-2 expression was increased. Meanwhile, the proliferation of ADMSCs was substantially promoted in Lv-IGF1 group after cultivating five or six days. Furthermore, PI3 K and p-AKT expression was notably highest in Lv-IGF1 group among three groups（P〈0.05）. Conclusions： ADMSCs was successfully isolated and cultured from Adult abdomen greater omentum adipose tissues. IGF1 overexpression in ADMSCs could promote the cell proliferation and depress the apoptosis dramatically partially through activating the cell signaling of PI3K/Akt.

关 键 词：脂肪间充质干细胞 慢病毒载体 IGF-1基因 PI3K/AKT通路 

分 类 号：Q782[生物学—分子生物学]