Effects of 5α-dihydrostestosterone on calcium mobilization and growth of prostate cancer cell line LNCaP / 第二军医大学学报

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°C 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°C for 3 min or inhibitor of ryanodine receptor such as procaine (50 mmol/L) at 37°C 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). Conclusion: DHT can induce rapid [Ca2+]i, rise in LNCaP cells in a concentration-dependent manner. The increase of [Ca2+]i, induced by DHT involves L-type voltage-gated calcium channels, but does not involve release of intracellular Ca2+ stores. The increase of [Ca2+]i induced by DHT increases apoptosis and inhibits growth of LNCaP cells.