尾加压素Ⅱ对大鼠肾小管上皮细胞的促丝裂作用

Mitogenic effect of urotensin Ⅱ on renal tubular epithelial cells in rats

目的:研究尾加压素Ⅱ(UⅡ)对大鼠肾小管上皮细胞的促丝裂作用。方法:采用机械分离和酶消化法获取肾小管上皮细胞进行培养、鉴定;用免疫细胞化学法定位溴脱氧尿嘧啶核苷(BrdU)掺入到肾小管上皮细胞DNA内;用MTT和BrdU掺入法观察不同浓度UⅡ(0、10-10、10-9、10-8、10-7和10-6mol.L-1)对肾小管上皮细胞增殖和DNA合成的影响,以确定其促丝裂作用。结果:培养的细胞经Cytokeratin-18免疫细胞化学染色后,强度不一和不均匀分布的棕黄色颗粒定位于大部分细胞的胞核周围和胞浆内。培养的肾小管上皮细胞经BrdU掺入和免疫细胞化学显色后,细胞核内有棕黄色颗粒沉着。10-10mol.L-1UⅡ组的MTT值、BrdU掺入量与对照组(0 mol.L-1UⅡ)比较差异无显著性(P>0.05);10-9和10-8mol.L-1UⅡ组的MTT值、BrdU掺入量高于对照组(P<0.01或P<0.05);与对照组比较,10-7和10-6mol.L-1UⅡ组的MTT值明显增加(P<0.05),但增加的幅度较10-9和10-8mol.L-1UⅡ组低,而BrdU掺入量无明显改变。结论:UⅡ对体外培养的大鼠肾小管上皮细胞具有促丝裂作用。

Objective To investigate the mitogenic effects of urotensin Ⅱ （U Ⅱ ） on renal tubular epithelial cells in rats. Methods The renal tubular epithelial cells in rats were collected by mechanical and enzyme digestive methods and were cultivated and characterized; using immunocytochemistry to investigate bromodeoxyuridine （BrdU） incorporation of DNA in renal tubular epithelial cells of rats; the effects of UⅡ with different concentrations （0, 10^-10, 10^-9 , 10^-8 , 10^-7 and 10^-6 mol · L^-1 ） on proliferation in rat renal tubular epithelial cells were observed using methyl thiazolyl tetrazolium assay （MTT assay） and BrdU incorporation was applied to idetect DNA synthesis in the cells. Results Cytokeratin-18 immunocytochemical staining showed that yellow granules with different intensities and uneven distribution were localized in nucleus surrounding and cytoplasm of the most renal tubular epithelial cells. Yellow granules were deposited in nuclei of cultural renal tubular epithelial cells by immunocytochemical staining following BrdU incorporation. The A values of MTT assay and BrdU incorporation in 10^-10mol · L^-1 U Ⅱ group were not significantly different from those in control group. In 10^-9 and 10^-8 mol · L^-1 UI] groups, the A values of MTT assay and BrdU incorporation were significantly higher than those in control group （P〈0. 01 or P〈0. 05）. Compared with control group, the A values of MTT assay in 10^-7 and10^-6 mol · L^-1 U Ⅱ groups were significantly higher （P〈0.05）, which was weaker than that difference in 10^-9 mol · L^-1 and 10^-8 mol · L^-1 groups, BrdU incorporation were not significantly different. Conclusion Urotensin 11 reveals a mitogenic effect on rat renal tubular epithelial cells in vitro.