Serotonin synthesis and metabolism-related molecules in a human prostate cancer cell line

Abstract

Prostate cancer is one of the most common tumors in males and its incidence is steadily increasing worldwide. Serotonin or 5-hydroxytryptamine (5-HT) is a well-known neurotransmitter that mediates a wide variety of physiological effects. An increase in the number of 5-HT-releasing neuroendocrine (NE) cells has been correlated with tumor progression. However, it is particularly unclear whether released 5-HT or the release of 5-HT has a role in tumor cell growth. We hypothesized that 5-HT synthesis and metabolism in NE cells regulate the growth of prostate cancer cells. In the present study, 5-HT was found to play a role as a cell growth factor in prostate cancer cells. Moreover, the pharmacological inhibition of 5-HT synthesis and metabolism interrupted the growth of prostate cancer cells. To confirm the existence of 5-HT in prostate cancer cells, we performed ELISA, HPLC, RT-PCR and immunohistochemical analyses. A high expression of tryptophan hydroxylase (TPH-1), dopa decarboxylase (DDC) and monoamine oxidase A (MAO-A) was noted in the prostate cancer cells when compared with normal prostate cells. Previous studies showed that 5-HT stimulated the proliferation of prostate cancer cells mediated by 5-HT receptors 5-HTR1A and R1B. However, cell proliferation was significantly inhibited when siRNA for both DDC and TPH-1 was transfected to the cells. Consequently, we propose that the secretion system of prostate NE cells capable of 5-HT synthesis and metabolism plays a significant role in prostate tumor generation and progression. These findings provide crucial clues for the development of potential pharmacotherapeutics to slow prostate tumor progression.

Figures

Figure 1

The serotonin synthesis and metabolism pathway and components affect cell proliferation. (A) Chemical structural pathway of the serotonin synthesis and metabolism system. (B) 5-HT significantly stimulated the growth of LNCaP cells in a dose-dependent manner (Black bar, +FBS; red bar, −FBS). (C) Antagonists of serotonin synthesis enzymes (p-chlorophenylalanine and benserazide) inhibited LNCaP cell growth by 50% at a concentration of 10 μM compared to the control at 72 h. (D) At a concentration of 0.01 μM, monoamine oxidase A (MAO-A)-specific antagonist (clorgyline) induced the cell growth effect by 30%, while treatment with a higher concentration showed 40% inhibition of cell proliferation compared to the control cells. Non-selective MAO antagonist (iproniazid) had no significant inhibitory effect on cell growth.

Figure 2

The serotonin synthesis and metabolism system plays a role in prostate cancer cells. (A) Total RNA was isolated from PrEC, LNCaP and PC-3 cells. Following reverse transcription, PCR was performed using specific primers for the components. (B) ELISA analysis of the serotonin content in both normal prostate and cancer cell lines. PC-3 cells had a high serotonin content, whereas LNCaP cells had a low serotonin content as compared to the normal prostate cell line. Inset, immunohistochemical analysis with both anti-5-HT Ab and DAPI staining in LNCaP cells. (C) HPLC analysis for 5-HTP, 5-HT and 5-HIAA contents in prostate-related cell lines.

Figure 3

Dopa decarboxylase (DDC) and tryptophan hydroxylase (TPH-1) siRNA expression inhibits cell proliferation. LNCaP cells were cultured in RPMI-1640 with 10% FBS. After siRNA was transfected to the cell lines (A, TPH-1 target; B, DDC target), the cells were cultured with 5-HT (10 μM) or without 5-HT for 7 days. (Black box, control; blue box, siRNA only; red box; siRNA + 5-HT)