SD大鼠乳鼠胰岛细胞体外培养方法探讨

In vitro culture of primary islet cells of suckling rats

目的探讨一种获得足量、纯度高和功能良好的大鼠胰岛细胞的体外培养方法.方法采用胶原酶对胰腺组织进行分次短时间消化;细胞接种18 h时,倒置显微镜下观察细胞生长情况;将细胞转入新的培养板培养,定期收集培养液上清.分别检测胰岛素、淀粉酶含量及葡萄糖刺激下的胰岛素分泌水平.结果SD大鼠乳鼠原代培养的胰岛细胞中成纤维细胞明显减少,双硫腙可染率85%～90%,锥虫兰染色细胞活力90%以上,培养7～11d的细胞分泌功能正常.结论用胶原酶反复短时间消化胰腺组织,细胞接种后及时转板的方法可获得纯度较高、生长良好、适合实验研究的胰岛单层细胞.

Objective To find a new method for obtaining abundant, well-purified and functionally active rat islet cells cultured in vitro. Method The pancreatic tissues of suckling rats underwent repeated digestion for short durations with collagenase, and the cell growth was observed under inverted microscope 18 h after cell inoculation. The cultured cells were then transferred to a new culture plate and the supernate was harvested regularly to determine the concentration of insulin, amylase and glucose-stimulated insulin release. Results The fibroblast cells in the primary cultured suckling rat cells were obviously reduced, and the ratio of dithizone-stained cells were 85%-90% with the viability exceeding 90% as assessed by trypan blue staining. The secretion function of the cultured cells remained normal after a 7- to 11-day culture. Conclusion Repeated digestion of the pancreatic tissues for short durations with collagenase and timely cell transfer to new plate may help achieve highly purified viable monolayer islet cells that are well applicable for experimental studies.