小鼠诱导多能干细胞与 PHBHHx 二维膜、三维膜体外共培养制作心肌补片  

Study on PHBHHx co-cultured with mouse induced pluripotent stem cells for use as myocardial patches in vitro

作　　者：许士俊 穆军升[1] 张建群[1] 伯平[1] 

机构地区：[1]首都医科大学附属北京安贞医院心外科、北京市心肺血管疾病研究所,100029

出　　处：《中华胸心血管外科杂志》2016年第7期415-418,共4页Chinese Journal of Thoracic and Cardiovascular Surgery

基　　金：2014年度北京市朝阳区科技计划项目（SF1417）,首都医科大学学生科研创新项目（XSKY2015192）,北京安贞医院院长基金（15P11）

摘　　要：目的：通过3-羟基丁酸与3-羟基己酸的共聚酯膜（ PHBHHx膜）与小鼠诱导多能干细胞（miPSCs）体外共培养，探讨其对miPSCs黏附、存活及分化的影响；寻找一种适合制作心肌补片的生物材料。方法复苏miPSCs，传代后，将miPSCs与PHBHHx二维膜和PHBHHx三维膜共培养，72 h后行扫描电镜观察细胞黏附情况。用含和不含维生素C的分化培养基对其进行诱导分化，通过免疫荧光、流式细胞术测定心肌细胞特异性蛋白cTnT的含量。结果 miPSCs可以在PHBHHx膜上良好的黏附和存活。维生素C作为一种良好的体外诱导剂，可以极大地促进miPSCs向心肌细胞分化，诱导分化21天后免疫荧光测定心肌细胞特异性蛋白cTnT表达阳性；流式细胞术测定cTnT的表达量：加维生素C诱导剂组cTnT＞未加任何诱导剂组[（47.54±1.46）％对（7.02±0.95）％，P＜0.05]。在miPSCs和PHBHHx膜共培养向心肌细胞分化过程中，添加维生素C诱导剂时，cTnT的表达量PHBHHx三维膜细胞培养分化＞PHBHHx二维膜细胞培养分化＞传统的细胞培养皿培养[（60.32±1.76）％对（53.31±1.41）对（47.54±1.46）％，P＜0.05]。结论 miPSCs与PHBHHx膜具有良好的组织相容性，可以在PHBHHx膜上黏附、存活和分化；相对于PHBHHx二维膜培养和传统的培养皿培养，三维PHBHHx膜培养在miPSCs向心肌细胞分化过程中具有更大的优势。Objective To study the effect of polymer material on adhesion , proliferation and differentiation of mouse in-duced pluripotent stem cells via co-culturing in vitro, to find a suitable polymeric biomaterials for miPSCs attachment , prolifera-tion and differentiation , forming myocardial patches as a new repair method for myocardial infarction .Methods miPSCs were recovered, passaged and cultured with PHBHHx two-dimension films and PHBHHx three-dimension films together.The mor-phology of miPSCs attached on the films was visualized under scanning electron microscope （ SEM） .The mouse induced pluri-potent stem cells were induced by differentiation medium that containing vitamin C .Control group did not add any inducer.The survival and differentiation of miPSCs were observed through immunofluorescence and flow cytometry .Results MiPSCs can grow, proliferate and differentiate on PHBHHx films both two-dimension and three-dimension.Vitamin C, as a favourable in-ducer, can markedly improve the efficiency of miPSCs differentiation of cardiomyocytes on PHBHHx films .Immunofluorescence results demonstrated positive cTnT expression.Flow cytometry measured cTnT expression： Vitamin C inducer group（47.54 ± 1.46）%〉without any inducer group（7.02 ±0.95）%（P〈0.05）.In the process of miPSCs differentiating into myocardial cell, after adding Vitamin C, the expression quantity of cTnT：PHBHHx three-dimensional films（60.32 ±1.76）%〉PHBHHx two-dimensional films（53.31 ±1.41）%〉traditional cell culture（47.54 ±1.46）%（P〈0.05）.Conclusion iPSCs can ad-here, survive and differentiate on the PHBHHx film.Vitamin C, as a favourable inducer, can markedly improve the efficiency of miPSCs differentiation of cardiomyocytes .Relative to PHBHHx two-dimensional film culture and traditional culture , the＆amp;nbsp;three-dimensional PHBHHx film culture has a great advantage in the process of miPSCs differentiating into myocardial cells .

关 键 词：多能干细胞 抗坏血酸 肌细胞 心脏 PHBHHx膜 

分 类 号：R654[医药卫生—外科学] R318.08[医药卫生—临床医学]