钾通道与SD大鼠前列腺上皮细胞增殖和凋亡关系的实验研究

The experimental study on the relationship between potassium channel and proliferation and apoptosis of SD rat prostatic epithelial cells

目的观察钾通道在SD大鼠前列腺上皮细胞增殖和凋亡中的作用。方法采用胶原酶消化SD大鼠前列腺组织建立原代SD大鼠前列腺上皮细胞株,角化蛋白免疫组织化学染色鉴定培养细胞,将SD大鼠前列腺上皮细胞分为对照组、钾通道阻断剂(四乙胺,TEA)组、开放剂(尼可地尔,Nicorandil)组,TEA的浓度为1、5、15mmol/L,Nicorandil的浓度为10、25、100μmol/L,各作用于细胞24、48、72h绘制细胞生长曲线,运用MTT方法、Hoechst33258细胞核染色法和流式细胞仪(FCM)观察传代后细胞增殖和凋亡状况。结果培养的细胞具有典型的上皮细胞形态特征,Pan-Keratin染色阳性,与对照组相比,MTT法和Hoechst33258染色法均显示,15mmol/L的钾离子通道阻滞剂TEA显著增加前列腺上皮细胞数(P<0.01);25μmol/L～100μmol/L/Nicorandil显著减少前列腺上皮细胞数(P<0.01),且FCM显示前列腺上皮细胞有明显的凋亡峰。Hoechst33258染色法显示细胞核固缩、裂解,呈现明显的凋亡形态特征。结论钾离子通道对体外培养的SD大鼠前列腺上皮细胞增殖与凋亡状态起重要的调节作用。

Objective To observe the role of potassium channel between potassium channel and proliferation and apoptosis of SD rat prostatic epithelial cells in vitro. Methods The primary culture was prepared by collagenase dissociation of minced prostatic tissue. Cells were cultured in serum -free prostate epithelial cell growth media and identified by immunocytochemical studies. The cells were divided into control, potassium channel blocker TEA and potassium channel opener Nicorandil and cultured in some special media, TEA under the concentration of 1 、 5 、 15mmol/L, Nicorandil under the concentration of 10、 25、 100 μmol/L, After 24、 48, 72h , draw the cells curves by couting the cell number, MTT assay, staining in the nucleus by Hoechst33258 and FCM. Results The cultured cells had the typical morphological features of epithelium and stained positive. Compared with the control group, MTT assay and stained by Hoechst33258 all showed that TEA significantly increased the cell number at 15mmol/L （P〈0.01）. However, Nicorandil at 25 μmol/L-100 μmol/L the cell number decreased significantly（P〈0.01）, FCM indicated a peak of apoptosis obviously, stained by Hoechst33258 showed that the nucleus of the cells have a typical features of apoptosis. Conclusion Potassium channel plays an important role in the relationship between the proliferation and apoptosis of SD rat prostatic epithelial cells in vitro.