胶原支架上体外三维培养成年大鼠心肌细胞

Collagen scaffold facilitates the three-dimensional culture of myocardial cells from adult rats in vitro

背景:前期研究中已经在Atelocollagen胶原支架培养出了具有收缩功能的乳鼠心肌细胞。目的:在Atelocollagen胶原支架上三维培养成年大鼠心肌细胞。方法:将成年SD大鼠心肌细胞经消化培养并纯化后,种植到Atelocollagen胶原支架上进行三维培养。应用倒置显微镜动态观察心肌细胞在Atelocollagen胶原支架上的生长状况;对三维培养的心肌细胞块进行冰冻切片,分别进行肌钙蛋白免疫组织化学和免疫荧光显色,对支架上生长的细胞进行鉴定。结果与结论:成年SD大鼠心肌细胞接种12h开始贴附支架生长,24h与支架汇合、紧密黏附,但心肌细胞并不产生自律性搏动,细胞之间夹杂少量崩解的细胞碎片;48h换液后支架网孔中崩解的细胞被换掉,剩下大部分细胞贴附在支架壁边缘生长。苏木精-伊红染色显示培养48h细胞与支架形成复合体。形态学鉴定证明支架网孔中生长的主要是心肌细胞。免疫组织化学和免疫荧光显色均显示在支架内生长的大部分是心肌细胞,并且紧贴支架,生长良好。

BACKGROUND：Three-dimensional contractible myocardial cel block of neonatal rats in the Atelocol agen scaffold has been cultured in the previous study. OBJECTIVE：To perform three-dimensional culture of myocardial cel s from adult rats on the Atelocol agen scaffold. METHODS：After being cultured and purified, myocardial cel s from adult Sprague-Dawley rats were implanted onto the Atelocol agen scaffold for three-dimensional cultures. The growth of myocardial cel s on the Atelocol agen scaffold was dynamical y observed by inverted microscopy. Myocardial cel blocks were separately tested by hematoxylin-eosin staining, immunohistochemistry and immunofluorecence of troponin after frozen section. RESULTS AND CONCLUSION：After being implanted, myocardial cel s from adult Sprague-Dawley rats began to grow on the Atelocol agen scaffold in 12 hours, showed mutual integration with the scaffold and the integration became more closely 24 hours later. However, the myocardial cel s did not have autorhythmiciting beats at al , and among the cel s there were some cel debris. After replaced with the same fresh medium 48 hours later, the cel debris was washed away and most of the rest cel s grew on the edge of the Atelocol agen scaffold. After 48 hours, the cel s formed a complex with the scaffold tested by hematoxylin-eosin staining, and the cel s which grew wel and were coalesced with the scaffold meshes were mainly myocardial cel s, identified by immunohistochemistry and immunofluorecence.