利用Rhodamine 123染色初步分离及纯化肠上皮干细胞

目的 探讨肠上皮干细胞初步的分离、纯化方法. 方法 利用Rhodamine 123对小鼠肠上皮细胞进行染色,流式细胞仪分选Rhodaminel23(Rho)染色阴性(Rholow)和强阳性细胞群(Rhobn);RT-PCR检测Rholow和Rhobn细胞musashi-1和p-糖蛋白1(p-g-p1)mRNA的表达;流式细胞术分析细胞周期和musashi-1蛋白表达情况. 结果 肠上皮细胞Rhodamine123染色可明显分为3个细胞群,分别是Rho阴性、Rho弱阳性和Rho强阳性细胞,分别约占总细胞的12.34%、45.26%和41.40%;成功分选出Rholow和Rhobri细胞群;前者musashi-1和p-g-p1mRNA表达阳性,后者p-g-p1mRNA少量表达;Rholow细胞群大部分细胞处于G0/G1期,约10.37%细胞musashi-1蛋白表达阳性. 结论 利用Rhodamine 123染色特点可实现肠上皮干细胞的初步分离、纯化.

Modest isolation and enrichment of intestinal epithelial stem cells by rhodamine 123 Staining

Objective To discuss the technology of modest isolating and enriching the intestinal epithelial stem cells. Methods Mouse intestinal epithelial cells were stained by Rhodamine 123 (Rho), sorting the Rhodamine 123 low staining cell population ( Rholow ) and Rhodamine 123 strong straining cell population ( Rhobri ) by fluorescence activated cell sorting(FACS) in flow cytometer; Detecting the musaashi-1 and p-glycoprotein 1 (p-g-p1) mRNA expression of two groups by RT-PCR; Analyzing the cell cycle and the percentage of the musashi-1 positive cells by flow cytometry. Results The intestinal epithelial cells were divided into three groups, Rhodamine 123 low staining cells( 12. 34% of total cells), Rhodamine 123 middle staining cells (45.26% of total cells) and Rhodamine 123 strong staining cells ( 41. 40% of total cells). The Rholow cell fraction and Rhobri cell fraction were isolated successfully. Both of musashi-1 and p-g-p1 mRNA were strongly expressed in Rho1ow cell fraction, and Rhobri cell fraction little expressed p-g-p1 mRNA. Most of Rho1ow cells were in G0/G1 phase, and the musashi-1 positive cells were about 10.37% of total cells in this fraction. Conclusion The intestinal epithelial stem cells can be modestly isolated and enriched by Rhodamine 123 staining.