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目的建立小鼠脾脏基质细胞培养的方法 ,分离出成纤维网状细胞并对其进行鉴定。方法取健康C57小鼠脾脏,胶原酶消化后,用磁珠分选出CD45-脾脏基质细胞,接种于α-MEM培养基,传代培养待其稳定生长。流式细胞仪进一步分选出成纤维网状细胞(gp38+CD31-),并用PCR、免疫荧光的方法检测其趋化因子表达。结果利用磁珠分离出脾脏的基质细胞培养后可以稳定传代,成功的分选出小鼠脾脏成纤维网状细胞(gp38+CD31-),并检测到其CCL19和CCL21的表达。结论成功分离培养脾脏基质细胞,并分选出有功能的成纤维网状细胞,为实验研究提供稳定的细胞来源。
Objective To establish a method of culturing mouse splenic stromal cells and to isolate and identify fibroblast-like cells. Methods The spleen of healthy C57 mice were digested with collagenase and CD45-splenic stromal cells were isolated by magnetic beads. The cells were inoculated into α-MEM medium and subcultured to be stable growth. Fibroblast cells (gp38 + CD31-) were further sorted by flow cytometry and chemokine expression was detected by PCR and immunofluorescence. Results Spleen stromal cells isolated by magnetic beads could be stably passaged after culture, and the spleen fibroblast reticulocytes (gp38 + CD31-) were successfully sorted and the expression of CCL19 and CCL21 was detected. Conclusion Splenic stromal cells were successfully isolated and sorted into functional fibroblast reticular cells, which provided a stable source of cells for experimental research.