北京生物医学工程

胚胎干细胞与聚3-羟基丁酸酯-co-4-羟基丁酸酯共培养细胞补片的初步研究

Primary study on 3-hydroxybutyrate-co-4-hydroxybutyrate co-cultured with mouse embryonic stem cell of cell patch

作者：马义祥穆军升张健群伯平

单位：首都医科大学附属北京安贞医院 (北京100029)

分类号：R318.04

出版年·卷·期（页码）：2016·35·1（53-58）

摘要：

目的干细胞移植是具有治疗心血管疾病潜力的有效方法，但是细胞移植过程中缺乏有效的载体会大大影响其成活和增殖。聚3-羟基丁酸酯-co-4-羟基丁酸酯P(3HB-co-4HB)是一种能够完全降解的，具有良好物理特性的高分子材料。探讨P(3HB-co-4HB)这种三维材料是否适合胚胎干细胞在上面生长，并生成细胞补片。方法复苏小鼠胚胎干细胞，传代培养，消化离心后重悬细胞，用悬滴法形成拟胚体(embryid bodies,EB)，将拟胚体与P(3HB-co-4HB)膜共培养；72h共培养后固定并进行扫描电镜和共聚焦显微镜观察细胞黏附情况，DAPI荧光染色观察细胞在P(3HB-co-4HB)膜上形态。结果胚胎干细胞在培养皿中贴壁生长良好，形态正常；拟胚体同P(3HB-co-4HB)膜共培养72h后，所有拟胚体都在三维材料表面良好生长，爬膜率为100%；Confocal显微镜和扫描电镜观察到在2mm的三维膜上，拟胚体生长贴附在生物膜上，细胞形态正常。结论 ESC在P(3HB-co-4HB)表面生长、增殖形成细胞补片。P(3HB-co-4HB)可作为干细胞治疗多种疾病的支架材料之一。

Objective Stem cell transplantation is considered promising for treating cardiovascular disorders. However,lack of an effective carrier impacts their survival and proliferation. Poly 3-hydroxybutyrate-co-4-hydroxybutyrate ［P(3HB-co-4HB)］ is a novel polymer biomaterial with excellent physical characteristics. We assessed the suitability of P(3HB-co-4HB) combined with embryonic stem cells (ESC) for cell patch formation. Methods Mouse embryonic stem cells were recovered，passaged，digested，centrifuged，suspended and form embryoid bodies (EB)by hanging drop method.EB were cultured with P(3HB-co-4HB) together for 72 hours. The morphology of EBs attached on the films was observed by confocal and scanning electron microscopy. DAPI fluorescence staining to observe cell morphology. Results Adherent mouse ESC displayed normal growth and morphology.When cultured as EB for 72 hours,100% grew on the biofilm and cells were present in the 3D biofilm to a depth of 2mm,with normal cell morphology. Conclusions Polymer biomaterial patches composed of P(3HB-co-4HB) permit ESCs growth .P(3HB-co-4HB) is a suitable biomaterial for stem cell transplantation in the treatment of cardiovascular disorders.

参考文献：

［1］Teng CJ,Luo J,Chiu RC,et al. Massive mechanical loss of microspheres with direct intramyocardial injection in the beating heart:Implications for cellular cardiomyoplasty［J］. J Thorac Cardiovasc Surg,2006,132(3):628-632.

［2］Al Kindi AH,Asenjo JF,Ge Y,et al. Microen- capsulation to reduce mechanical loss of microspheres:implications in myocardial cell therapy［J］. Eur J Cardiothorac Surg,2011,39(2):241-247.

［3］Saito T,Kuang JQ,Lin CC,et al. Transcoronary implantation of bone marrow stromal cells ameliorates cardiac function after myocardial infarction［J］. J Thorac Cardiovasc Surg,2003,126(1):114-123.

［4］Mandel Y,Weissman A,Schick R,et al. Human embryonic and induced pluripotent sten cell-derived cardiomyoeytes exhibit beat rate variability and power-law behavior［J］. Circulation,2012,125(7):883-893.

［5］Shiba Y,Fernandes S,Zhu WZ,et al.Human ES-cell- derived cardiomyocytes electrically couple and suppress arrhythmias in injured hearts.［J］. Nature,2012,489(7415):322-325.

［6］Gepstein L,Ding C,Rahmutula D,et al. In vivo assessment of the electrophysiological integration and arrhythmogenic risk of myocardial celltransplantation strategies［J］. Stem Cells,2010,28(12):2151-2161.

［7］Verma V,Verma P,Ray P,et al. Preparation of scaffolds from human hair proteins for tissue-engineering applications［J］. Biomed Mater,2008,3(2):025007.

［8］Nazarov R,Jin HJ,Kaplan DL.Porous 3-D scaffolds from regenerated silk fibroin［J］. Biomacromolecules,2004,5(3):718-726.

［9］Kse GT,Korkusuz F,Korkusuz P,et al.Bone generation on PHBV matrices:an in vitro study［J］. Biomaterials,2003,24(27):4999-5007.

［10］Tobella LM,Bunster M,Pooley A,et al. Biosynthesis of poly-beta-hydroxyalkanoates by Sphingopyxis chilensis S37 and Wautersia sp. PZK cultured in cellulose pulp mill effluents containing 2,4,6-trichlorophenol［J］.J Ind Microbiol Biotechnol,2005,32(9):397-401.

［11］Thakor N,Trivedi U,Patel KC. Biosynthesis of medium chain length Poly(3-hydroxyalkanoates) (mcl-PHAs) by Comamonas testosteroni duringcultivation on vegetable oils［J］.Bioresour Technol,2005,96(17):1843-1850.

［12］Blau HM,Brazelton TR,Weimann JM. The evolving concept of a stem cell:entity or function?［J］. Cell,2001,105(7):829-841.

［13］Orlic D,Hill JM,Arai AE. Stem cells for myocardial regeneration［J］.Circ Res,2002,91(12):1092-1102.

［14］Roell W,Lewalter T,Sasse P,el a1.Engraftment of connexin 43-expressing cells prevents post-infarct arrhythmia［J］.Nature,2007,450 (7171):819-824.

服务与反馈：

【文章下载】【加入收藏】

提示：您还未登录，请登录！点此登录