| 摘要: |
| 目的:探讨5α-双氢睾酮(DHT)对前列腺癌LNCaP细胞钙离子移动的影响及其机制。方法:应用Fura-2/AM Ca2+ 荧光探针法结合MiraCal荧光成像系统动态检测DHT刺激以及Ca2+通道阻滞剂干预后LNCaP细胞内钙离子浓度(\[Ca2+\]i)的变化。应用MTT法观察细胞活力,流式细胞仪观察早期细胞凋亡率。结果:DHT浓度为1、10、100和1 000 nmol/L时,能快速诱导\[Ca2+\]i升高,在20 s～3 min升至峰值。细胞外液无Ca2+时,1 000 nmol/L DHT未能诱导\[Ca2+\]i升高。细胞膜L-型电压门控Ca2+通道阻滞剂维拉帕米(50 μmol/L)、地尔硫(100 μmol/L)或硝苯地平(5 mmol/L)37℃孵育细胞5 min后,能完全抑制1 000 nmol/L DHT诱导的\[Ca2+\]i升高。磷脂酶C抑制剂新霉素(1 mmol/L)37℃孵育细胞5 min或兰尼定受体阻滞剂普鲁卡因(50 mmol/L)37℃孵育细胞3 min后,对1 000 nmol/L DHT诱导的\[Ca2+\]i升高没有影响。1 000 nmol/L DHT作用细胞48 h后,与1 000 nmol/L DHT作用前用维拉帕米预孵细胞相比,细胞光密度(D)值\[(0.67±0.10) vs (2.13±0.16))和早期细胞凋亡率\[(14.31±2.29)% vs (1.07±0.19)%\]差异有统计学意义(P＜0.01)。结论:DHT可快速地、剂量依赖性地诱导LNCaP细胞\[Ca2+\]i升高;DHT诱导的LNCaP细胞\[Ca2+\]i的升高是通过细胞外Ca2+经细胞膜L-型电压门控Ca2+通道流入细胞内实现的,细胞内贮钙库未释放Ca2+。DHT诱导的LNCaP细胞\[Ca2+\]i升高促进细胞凋亡、抑制细胞生长。 |
| 关键词: 前列腺肿瘤 5α-双氢睾酮 钙；钙通道 |
| DOI：10.3724/SP.J.1008.2008.00 |
| 投稿时间：2008-01-25修订日期：2008-04-09 |
| 基金项目:国家自然科学基金（30225046）. |
|
| Effects of 5α-dihydrostestosterone on calcium mobilization in prostate cancer LNCaP cells |
| TANG Yuan-jie1,2, SUN Ying-hao1*, GAO Xu1, XU Chuan-liang1, WANG Lin-hui1 |
| (1. Department of Urology, Changhai Hospital, Second Military Medical University, Shanghai 200433, China*2. Department of Urology, Hospital of Chinese People’s Armed Police Forces, Jiangsu Regional Headquarters, Yangzhou 225003) |
| Abstract: |
| Objective:To investigate the effects of 5α - dihydrostestosterone (DHT) on calcium mobilization and growth of prostate cancer cell line LNCaP.Methods: Intracellular calcium concentration (\[Ca2+\]i) was assayed by MiraCal Image System using Fura-2/AM as Ca2+ fluorescence probe. Cell viability was observed by MTT assay and apoptosis by flow cytometry. Results: The calcium levels rapidly increased following addition of DHT, with the latency of response only in seconds. DHT at the concentrations of 1, 10, 100 and 1 000 nmol/L increased \[Ca2+\]i from (28±5), (29±5), (28±4) and (28±9) nmol/L to (31±3) ( P>0.05,65±9) (P<0.01), (193±33) (P<0.001) and (208±42) nmol/L (P<0.001), respectively. The response induced by 1 000 nmol/L DHT was similar to that induced by 100 nmol/L DTH. DHT 1 000 nmol/L did not increase \[Ca2+\]i under extracellular Ca2+ -free condition. Blockers of L-type voltage-gated calcium channels, including verapamil (50 μmol/L), diltiazem (100 μmol/L) or nifedipine (5 mmol/L) at 37℃ for 5 min prior to stimulation with 1 000 nmol/L DHT, completely inhibited DHT-induced \[Ca2+\]i rise. Pre-treatment with inhibitor of phospholipase C such as neomycin sulfate (1 mmol/L) at 37℃ for 3 min or inhibitor of ryanodine receptor such as procaine (50 mmol/L) at 37℃ for 3 min had no influence on \[Ca2+\]i rise induced by 1 000 nmol/L DHT. The optical density (D) values and early apoptosis rates of the cells stimulated with 1 000 nmol/L DHT for 48 h were significantly different from those of cells pre-treated with verapamil prior to stimulation with 1 000 nmol/L DHT (\[0.67±0.10\] % vs \[2.13±0.16\] % and \[14.31±2.29\] % vs \[1.07±0.19\] %,P<0.01). |
| Key words: prostatic neoplasms 5α-dihydrostestosterone calcium calcium channels |
