低频脉冲电磁场在三维支架下对人脂肪源干细胞增殖和成骨分化的影响  

Effect of low-frequency pulsed electromagnetic fields on proliferation and osteogenic ability of human adipose-derived stem cells in a three-dimensional scaffold

作　　者：陈宇雄 陈贤哲[1] 倪梦杉 郭文杰[1] 田京[2] Chen Yu-xiong Chen Xian-zhe Ni Meng-shan Guo Wen-jie Tian Jing(The Second Clinical Medical College, Southern Medical University, Guangzhou 510282, Guangdong Province, China Orthopedics Center, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, Guangdong Province, China)

机构地区：[1]南方医科大学第二临床医学院,广东省广州市510282 [2]南方医科大学珠江医院骨科中心,广东省广州市510282

出　　处：《中国组织工程研究》2017年第18期2828-2833,共6页Chinese Journal of Tissue Engineering Research

基　　金：广东省大学生科技创新培育专项资金(pdjh2016b0104);广东省科技计划项目(2013B021800312)~~

摘　　要：背景:目前已有较多实验研究证实低频脉冲电磁场在二维支架下促进多种干细胞向成骨细胞分化,但鲜有研究磁场在三维支架下对干细胞的增殖及成骨分化的影响。目的:探讨在聚已内酯3D培养支架环境下,外加低频脉冲电磁场对人脂肪源干细胞增殖及成骨分化的影响。方法:将第3代人脂肪源干细胞接种于聚已内酯3D培养支架上,分2组培养,实验组给予50 Hz、1 m T的低频脉冲电磁场干预,2 h/d,连续干预14 d;对照组不给于低频脉冲电磁场干预。培养7 d后,Live/Dead染色观察细胞存活情况;培养1,3,5,7 d后,MTT法检测细胞增殖;培养7,14 d,碱性磷酸酶染色及qR T-PCR检测细胞成骨分化水平。结果与结论:(1)Live/Dead染色:人脂肪源干细胞已长入到支架材料的空隙结构内部,在材料中可保持较高的活性;(2)细胞增殖:随着培养时间的延长,两组细胞A值均逐渐升高,实验组培养1,3 d的细胞A值高于对照组(P<0.05);(3)碱性磷酸酶染色:实验组培养7,14 d的碱性磷酸酶阳性染色强于对照组;(4)qR T-PCR检测:培养7 d时,实验组碱性磷酸酶、Ⅰ型胶原基因表达量高于对照组(P<0.01);培养14 d时,实验组骨桥蛋白、Runt相关转录因子2、碱性磷酸酶及Ⅰ型胶原的表达量均明显高于对照组(P<0.05);(5)结果表明:在聚已内酯3D培养支架环境下,外加低频脉冲电磁场可促进人脂肪源干细胞的增殖及成骨分化。BACKGROUND： Nowadays increasing experimental findings have proved that the low-frequency pulsed electromagnetic fields（LPEMF） can induce osteogenic differentiation of a variety of stem cells in the two-dimensional scaffold. However, little is reported on the LPEMF effect on the proliferation and osteogenic differentiation of stem cells in the three-dimensional scaffold. OBJECTIVE： To investigate the effect of LPEMF on the proliferation and osteogenic differentiation of human adipose-derived stem cells（h ASCs） in the 3D Insert-PCL scaffold. METHODS： Passage 3 h ASCs were directly cultured in the 3D Insert-PCL scaffolds followed by LPEMF（50 Hz, 1 m T） exposure, 2 hours per day, for continuous 14 days（experimental group） or no intervention（control group）. After 7 days of culture, Live/Dead staining was used to observe cell survival. After 1, 3, 5, 7 days of culture, MTT assay was used to detect cell proliferation. After 7 and 14 days of culture, the osteogenic differentiation of h ASCs was assessed through the alkaline phosphatase（ALP） staining and q RT-PCR. RESULTS AND CONCLUSION： Live/dead cell staining proved that the h ASCs had a good growth in the 3D Insert-PCL scaffolds as well as a high survival rate. The absorbance values of h ASCs in the two groups were increased gradually with time, and the absorbance value in the experimental group was significantly higher than that in the control group at 1 and 3 days after culture（P〈0.05）. The ALP activity in the experimental group was stronger than that in the control group at 7 and 14 days after culture. q RT-PCR findings showed that at 7 days after culture, the m RNA levels of ALP and type I collagen were significantly higher in the experimental group than the control group（P〈0.01）, while at 14 days after culture, the m RNA levels of osteopontin, Runt-related transcription factor, ALP and type I collagen were significantly higher in the experimental group than the control group（P〈0.05）. To conclude, the LPEMF exposure

关 键 词：生物材料 骨生物材料 聚已内酯3D培养支架 低频脉冲电磁场 人脂肪源干细胞 增殖 成骨分化 

分 类 号：R318[医药卫生—生物医学工程]