The Role of Serotonin (5-Hydroxytryptamine_1A and _1B) Receptors in Prostate Cancer Cell Proliferation

Purpose

Serotonin (5-hydroxytryptamine), a monoamine neurotransmitter released by prostate neuroendocrine cells, has a fundamental role in tumor growth, differentiation and gene expression. We investigated the effect of 5-hydroxytryptamine and 5-hydroxytryptamine antagonists on the growth of prostate cancer cells and we identified 5-hydroxytryptamine receptor expression in PC3 cells and in human hormone refractory prostate cancer tissue.

Materials and Methods

A total of 12 preparations of hormone refractory PC3 human prostate cancer cells were incubated with 5-hydroxytryptamine, or the 5-hydroxytryptamine receptor antagonists 5-hydroxytryptamine_1A, _1B, _1D, ₂, ₃ or ₄. After 72 hours cell viability was assessed using the crystal violet assay. PC3 cells treated with 5-hydroxytryptamine_1A and _1B antagonists were investigated for apoptosis using flow cytometry. PC3 cells and sections of hormone refractory human prostate cancer tissue were studied by immunohistochemistry and Western blot analysis.

Results

In PC3 cells 5-hydroxytryptamine caused dose dependent proliferation with a maximum increase of 15% in 12 preparations at a concentration of 10⁻⁸ M at 72 hours compared to controls (p <0.0001). At a concentration of 10⁻⁴ M at 72 hours the 5HT_1A antagonist NAN-190 hydrobromide and the 5-hydroxytryptamine_1B antagonist SB224289 HCl (Tocris Laboratories, Bristol, United Kingdom) induced a 20% and 78% inhibitory effect, respectively, on PC3 cell growth compared to that in controls (p <0.0001). In PC3 cells 5-hydroxytryptamine_1A and _1B antagonists demonstrated apoptosis after 24 and 48 hours of incubation. Immunostaining for 5-hydroxytryptamine_1A and _1B receptors was seen in PC3 cells and prostate cancer tissue. Western blot analysis demonstrated 5-hydroxytryptamine_1A and _1B receptor proteins with 46 and 43 kDa bands, respectively.

Conclusions

In PC3 prostate cancer cells 5-hydroxytryptamine_1A and to a greater extent 5-hydroxytryptamine_1B antagonists significantly inhibit growth and induce apoptosis. To our knowledge growth inhibition caused by the 5-hydroxytryptamine_1B antagonist SB224289 HCl is a novel finding, as is apoptosis caused by the 2 antagonists 5-hydroxytryptamine_1A and _1B. This effect is most likely mediated via 5-hydroxytryptamine_1A and _1B receptors. Therefore, our results imply that 5-hydroxytryptamine_1A and in particular 5-hydroxytryptamine_1B receptor antagonists warrant further investigations as potential anti-neoplastic agents.

Key Words:  prostate , prostatic neoplasms , serotonin , gene expression

Abbreviations and Acronyms:  5HT, 5-hydroxytryptamine , 5HTR, 5HT receptor , ABC, avidin-biotin complex , DES, diethylstilbestrol , FITC, fluorescein isothiocyanate , NE, neuroendocrine , PBS, phosphate buffered saline , RT, room temperature , SDS, sodium dodecyl sulfate , TBS, tris buffered saline