大鼠腹膜间皮细胞的分离培养方法

Establishment of method for primary culture and passage of rat peritoneal mesothelial cells

目的建立简便有效的大鼠腹膜间皮细胞(passage of rat peritoneal mesothelial cells,RPMC)的体外培养方法。方法采用0.25%胰蛋白酶-0.02%EDTA消化大鼠大网膜及脾胃韧带,细胞悬液进行离心培养,沉淀重悬后进行培养。通过形态学、免疫荧光、免疫组化法鉴定细胞。结果分离培养的细胞在光镜下呈铺路鹅卵石状,免疫荧光鉴定显示细胞角蛋白阳性,免疫组化鉴定显示波形蛋白抗原阳性,第Ⅷ因子、白细胞CD45抗原阴性,证实培养的细胞为RPMC。结论胰蛋白酶消化法是一种简便而实用的分离RPMC的方法。

Objective To establish a convenient and effective method for primary culture and passage of rat peritoneal mesothelial cells（RPMCs）.Methods Omental specimens were obtained from SD rats and digested with 0.25% trysin-0.02%EDTA for 10 min.Cell deposit was suspended and was cultured in DMEM/F12 containing 15% fatal calf serum.Cells were verified by morphology,cytoimmunohistochemistry and cytoimmunofluorescence analysis.Results The confluent cells showed cobblestone-like appearance under microscope.Cytoimmunofluorescence analysis showed a positive staining of cytokeratin.Cytoimmunohistochemistry analysis showed a positive staining of vimentin,a negative staining of CD45 and factor Ⅷ.Conclusion The method established in the study is an easy and practical one to obtain RPMCs.