Department of Biotechnology and Medical Engineering, National Institute of Technology, Rourkela, Odisha, India.
Department of Biotechnology, Institute of Engineering and Technology, Bundelkhand University, Jhansi, Uttar Pradesh, India.
J Biomed Mater Res A. 2020 Mar;108(3):496-514. doi: 10.1002/jbm.a.36831. Epub 2019 Nov 22.
In recent years, stem cell-based therapies shown to have promising effects on the clinical management of ischemic heart disease. Moreover, stem cells differentiation into cardiomyocytes (CMs) can overcome the cell source limitations. The current research involves the isolation and expansion of mesenchymal stem cells (MSCs) and induced pluripotent stem cells (iPSCs), their differentiation into CMs and subsequent construction of tissue-engineered myocardium supported by random and aligned polycaprolactone (PCL) nanofibrous matrices (av. dia: 350-850 nm). Umbilical cord matrix (UCM)-derived MSCs were isolated successfully by routine enzymatic digestion and a nonenzymatic explant culture method and characterized by their morphology, differentiation into different lineages, and surface marker expression. Treatment of UCM-derived MSCs with 5-azacytidine (5 μM) induced their differentiation into putative cardiac cells, as revealed by the expression of cardiac-specific troponin T (cTnT), smooth muscles actin, myogenin (MYOG), smoothelin, cardiac α-actin genes and cTnT, α-actinin proteins by RT-PCR and immunocytochemistry, respectively. However, no beating cells were observed in differentiated MSCs. On the other hand, adult human foreskin-derived iPSCs cultured on Matrigel™-coated aligned PCL nanofibrous matrices showed anisotropic behavior along the PCL nanofibers and, upon differentiation, expressed cardiac-specific cTnT (23.34 vs. 32.55%) proteins and showed more synchronized beating than those differentiated on Matrigel™-coated tissue culture coated polystyrene surfaces. Moreover, aligned PCL nanofibers are able to promote cells orientation parallel to the fibers, thus providing an effective way to control anisotropic nature under in vitro condition.
近年来，基于干细胞的疗法在缺血性心脏病的临床治疗中显示出有前景的效果。此外，干细胞向心肌细胞 (CMs) 的分化可以克服细胞来源的限制。目前的研究涉及间充质干细胞 (MSCs) 和诱导多能干细胞 (iPSCs) 的分离和扩增，它们向 CMs 的分化以及随后在随机和定向聚己内酯 (PCL) 纳米纤维基质（平均直径：350-850nm）支持下构建组织工程心肌。通过常规酶消化和非酶外植体培养方法成功分离出脐带基质 (UCM) 衍生的 MSCs，并通过其形态、向不同谱系的分化和表面标志物表达进行了表征。用 5-氮杂胞苷 (5μM) 处理 UCM 衍生的 MSCs 诱导其分化为拟心肌细胞，通过 RT-PCR 和免疫细胞化学分别检测到心脏特异性肌钙蛋白 T (cTnT)、平滑肌肌动蛋白、肌生成素 (MYOG)、 smoothelin、心脏α-肌动蛋白基因和 cTnT、α-辅肌动蛋白蛋白的表达。然而，在分化的 MSC 中没有观察到搏动细胞。另一方面，在 Matrigel™涂覆的定向 PCL 纳米纤维基质上培养的成人包皮来源的 iPSCs 沿 PCL 纳米纤维表现出各向异性行为，并且在分化后表达心脏特异性 cTnT（23.34 对 32.55%）蛋白，并且比在 Matrigel™涂覆的组织培养涂覆聚苯乙烯表面上分化的蛋白表现出更同步的搏动。此外，定向 PCL 纳米纤维能够促进细胞沿纤维平行取向，从而为在体外条件下控制各向异性提供了有效途径。
