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Testosterone and prolactin increase carboxypeptidase‐D and nitric oxide levels to promote survival of prostate cancer cells
Author(s) -
Thomas Lynn N.,
Morehouse Timothy J.,
Too Catherine K.L.
Publication year - 2012
Publication title -
the prostate
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.295
H-Index - 123
eISSN - 1097-0045
pISSN - 0270-4137
DOI - 10.1002/pros.21446
Subject(s) - lncap , downregulation and upregulation , endocrinology , medicine , testosterone (patch) , nitric oxide , viability assay , prolactin , apoptosis , transfection , prostate cancer , dihydrotestosterone , chemistry , biology , androgen , cell culture , cancer , biochemistry , hormone , gene , genetics
BACKGROUND Plasma‐membrane carboxypeptidase‐D (CPD) releases arginine from extracellular substrates. Arginine is converted intracellularly to nitric oxide (NO). This study determined the effects of testosterone (T) and prolactin (PRL) on CPD expression, and the role(s) of CPD in NO production and survival of prostate cancer (PCa) cells. METHODS LNCaP cells were treated with T and/or PRL. CPD expression was measured. Regulation by T (low doses) was determined using transfected cells overexpressing 5α‐reductase type‐1 (5αR1), which converts T to the more potent dihydrotestosterone. The effects of siRNAs targeting CPD (siCPDs) on NO production, cell viability, and apoptosis were determined using DAF2‐DA, MTS, and Annexin‐V assays. The effects of PRL/T on CPD/NO levels in PC‐3, MDA‐PCa‐2b, and 22Rv1 cells were also evaluated. RESULTS In LNCaP cells, 10 nM T and 10 ng/ml PRL‐upregulated CPD mRNA/protein levels. In pTRE‐transfectants, 1 nM T‐upregulated CPD mRNA levels by ∼2‐fold over controls, whereas 0.1 nM T caused similar upregulation in pTRE‐5αR1‐transfectants. In LNCaP cells cultured in arginine‐free medium, addition of furylacryloyl‐Ala‐Arg (FAR; CPD substrate) increased NO levels. NO production, with FAR, was enhanced by PRL and/or T. siCPDs decreased NO production and cell viability, but increased apoptosis. QPCR analysis showed T/PRL‐upregulation of CPD in 22Rv1, MDA‐PCa‐2b, and PC‐3 cells. NO production was doubled by T/PRL in 22Rv1 cells, tripled by T in MDA‐PCa‐2b cells, and marginally increased by PRL in MDA‐PCa‐2b and PC‐3 cells. CONCLUSIONS T and PRL upregulate CPD and NO levels in PCa cells. CPD increases NO production to promote PCa cell survival. Prostate 72:450–460, 2012. © 2011 Wiley Periodicals, Inc.