小鼠角膜上皮细胞诱导获得小胶质样细胞

Obtaining microglia-like cells after mouse corneal epithelial cell induction

背景:小胶质细胞是中枢神经系统最主要的免疫细胞,对研究多种疾病的发生和治疗有重要意义,但体外个体化细胞的获得受限。目的:建立一种新的小胶质细胞诱导培养体系,将小鼠角膜上皮细胞诱导成为小胶质样细胞并对其进行鉴定。方法:裂解2周龄小鼠角膜上皮组织获得上皮细胞,先后加入含有白细胞介素34(100 ng/mL)、集落刺激因子1(5 ng/mL)和转化生长因子β(1 ng/mL)、白细胞介素6(1 ng/mL)、重组血管内皮生长因子(1 ng/mL)的改良培养基,并且持续处理到16 d。通过转录水平、蛋白表达水平、亚细胞定位、流式分析和形态学观察多角度评估诱导后的细胞性质。结果与结论:诱导后获得表达TMEM119、CXCL11、CD11b、CD68小胶质细胞特征分子的细胞,形态学观察、流式分析和荧光检验后确定其表达产物具有较高丰度,与小胶质细胞极为相似。

BACKGROUND:Microglial cells are the predominant immune cells of the central nervous system and are important for the study of the development and treatment of many diseases.However,the methods for obtaining individualized cells in vitro are still lacking.OBJECTIVE:To establish a new microglia induction culture system to induce mouse corneal epithelial cells into microglia-like cells.METHODS:Corneal epithelial tissues of 2-week-old mice were lysed to obtain epithelial cells,and modified media containing interleukin-34(100 ng/mL),colony stimulating factor 1(5 ng/mL),transforming growth factorβ(1 ng/mL),interleukin-6(1 ng/mL),and recombinant vascular endothelial growth factor(1 ng/mL)were successively added.Treatment lasted up to 16 days.Cell properties were evaluated after induction by transcriptional level,protein expression level,subcellular localization,flow cytometry and morphological observation.RESULTS AND CONCLUSION:After induction,microglia-like cells which express characteristic molecules of TMEM119,CXCL11,CD11b and CD68 were obtained.Morphological observation,flow cytometry and fluorescence analysis confirmed that the expression products of TMEM119,CXCL11,CD11b and CD68 microglia had high abundance,which was very similar to microglia.