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CD8+ T cells promote proliferation of benign prostatic hyperplasia epithelial cells under low androgen level via modulation of CCL5/STAT5/CCND1 signaling pathway.

ABSTRACT: Previous studies by our group have shown that low intra-prostatic dihydrotestosterone (DHT) induced BPH epithelial cells (BECs) to recruit CD8+ T cells. However, the influence of the recruited CD8+ T cells on BECs under a low androgen level is still unknown. Here, we found CD8+ T cells have the capacity to promote proliferation of BECs in low androgen condition. Mechanism dissection revealed that interaction between CD8+ T cells and BECs through secretion of CCL5 might promote the phosphorylation of STAT5 and a higher expression of CCND1 in BECs. Suppressed CCL5/STAT5 signals via CCL5 neutralizing antibody or STAT5 inhibitor Pimozide led to reverse CD8+ T cell-enhanced BECs proliferation. IHC analysis from Finasteride treated patients showed PCNA expression in BECs was highly correlated to the level of CD8+ T cell infiltration and the expression of CCL5. Consequently, our data indicated infiltrating CD8+ T cells could promote the proliferation of BECs in low androgen condition via modulation of CCL5/STAT5/CCND1 signaling. The increased secretion of CCL5 from the CD8+ T cells/BECs interaction might help BECs survive in a low DHT environment. Targeting these signals may provide a new potential therapeutic approach to better treat BPH patients who failed the therapy of 5α-reductase inhibitors.

SUBMITTER: Yang Y

PROVIDER: S-EPMC5316951 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundBenign prostatic hyperplasia (BPH) is the most common urological disease in elderly men, but the underlying pathophysiological mechanisms are complex and not fully understood. Phosphodiesterase type 5 inhibitors (PDE5-Is) used to treat BPH could upregulate the cyclic guanosine monophosphate (cGMP)-dependent protein kinase G (PKG) signaling, which was shown to blunt inflammation in the prostate. Our previous findings indicate that CD8+ T cells promote the proliferation of BPH epithelial cells (BECs) in low androgen conditions through secretion of CCL5; however, the role of the cGMP/PKG pathway in the process is unclear.MethodsParaffin-embedded tissues were used for expression quantity of CD8+ T cells, CCL5, cyclin D1, and PDE5 protein by immunohistology in prostate specimens which were/were not treated with finasteride 5 mg daily for at least 6 months before surgery. BPH-1 cells were cocultured with or without CD8 + T cells or PDE5-Is in low androgen conditions for 4 days. The conditioned media, BPH-1 cells, and CD8 + T cells were harvested for the subsequent experiments. The quantitative polymerase chain reaction was used for assaying the level of messenger RNA expression of CCL5. CCL5 in the conditioned media was detected by the enzyme-linked immunosorbent assay. The effect of PDE5-Is on cocultured BPH-1/CD8 + T-cell proliferation was detected by the cell counting kit-8. A high-fat diet (HFD)-induced prostatic hyperplasia rat model was used to investigate the effect of cGMP/PKG activation in CD8 + T cells in vivo.ResultsCD8+ T-cell infiltration into human BPH tissues was positively correlated with the expression of CCL5, cyclin D1, and PDE5, whereas in an HFD-induced prostatic hyperplasia rat model, the activation of the cGMP/PKG signaling by a PDE5-I could suppress the CD8 + T-cell infiltration and the CCL5 and cyclin D1 expression. Furthermore, the activation of the cGMP/PKG pathway inhibited CCL5 secretion by CD8 + T cells by downregulating nuclear factor-κB p65 phosphorylation, which reduced the growth of BPH-1 through CCL5/STAT5/CCND1 signaling.ConclusionsOur results indicate that the upregulation of the cGMP/PKG/p65 signaling reduces CCL5 secretion in CD8 + T cells, which in turn decreases the proliferation of BECs in low androgen conditions, suggesting that the combination of 5α reductase inhibitors lowering androgen levels and PDE5-Is may be a novel, more effective treatment for BPH patients.

Project description:Early studies suggested macrophages might play roles in inflammation-associated benign prostatic hyperplasia (BPH) development, yet the underlying mechanisms remain unclear. Here we first showed that CD68(+) macrophages were identified in both epithelium and the stromal area of human BPH tissues. We then established an in vitro co-culture model with prostate epithelial and macrophage cell lines to study the potential impacts of infiltrating macrophages in the BPH development and found that co-culturing prostate epithelial cells with macrophages promoted migration of macrophages. In a three-dimensional culture system, the sphere diameter of BPH-1 prostate cells was significantly increased during coculture with THP-1 macrophage cells. Mechanism dissection suggested that expression levels of epithelial-mesenchymal transition (EMT) markers, such as N-cadherin, Snail, and TGF-?2, were increased, and administration of anti-TGF-?2 neutralizing antibody during co-culture suppressed the EMT and THP-1-mediated growth of BPH-1 cells, suggesting THP-1 might go through EMT to influence the BPH development and progression. Importantly, we found that modulation of androgen receptor (AR) in BPH-1 and mPrE cells significantly increased THP-1 and RAW264.7 cell migration, respectively, and enhanced expression levels of EMT markers, suggesting that AR in prostate epithelial cells might play a role in promoting macrophage-mediated EMT in prostate epithelial cells. Silencing AR function via an AR degradation enhancer, ASC-J9, decreased the macrophage migration to BPH-1 cells and suppressed EMT marker expression. Together, these results provide the first evidence to demonstrate that prostate epithelial AR function is important for macrophage-mediated EMT and proliferation of prostate epithelial cells, which represents a previously unrecognized role of AR in the cross-talk between macrophages and prostate epithelial cells. These results may provide new insights for a new therapeutic approach to battle BPH via targeting AR and AR-mediated inflammatory signaling pathways.

Project description:BackgroundMale lower urinary tract symptoms (LUTS) occur in more than half of men above 50 years of age. LUTS were traditionally attributed to benign prostatic hyperplasia (BPH) and therefore the clinical terminology often uses LUTS and BPH interchangeably. More recently, LUTS were also linked to fibrogenic and inflammatory processes. We tested whether osteopontin (OPN), a proinflammatory and profibrotic molecule, is increased in symptomatic BPH. We also tested whether prostate epithelial and stromal cells secrete OPN in response to proinflammatory stimuli and identified downstream targets of OPN in prostate stromal cells.MethodsImmunohistochemistry was performed on prostate sections obtained from the transition zone of patients who underwent surgery (Holmium laser enucleation of the prostate) to relieve LUTS (surgical BPH, S-BPH) or patients who underwent radical prostatectomy to remove low-grade prostate cancer (incidental BPH, I-BPH). Images of stained tissue sections were captured with a Nuance Multispectral Imaging System and histoscore, as a measure of OPN staining intensity, was determined with inForm software. OPN protein abundance was determined by Western blot analysis. The ability of prostate cells to secrete osteopontin in response to IL-1β and TGF-β1 was determined in stromal (BHPrS-1) and epithelial (NHPrE-1 and BHPrE-1) cells by enzyme-linked immunosorbent assay. Quantitative polymerase chain reaction was used to measure gene expression changes in these cells in response to OPN.ResultsOPN immunostaining and protein levels were more abundant in S-BPH than I-BPH. Staining was distributed across all cell types with the highest levels in epithelial cells. Multiple OPN protein variants were identified in immortalized prostate stromal and epithelial cells. TGF-β1 stimulated OPN secretion by NHPrE-1 cells and both IL-1β and TGF-β1 stimulated OPN secretion by BHPrS-1 cells. Interestingly, recombinant OPN increased the mRNA expression of CXCL1, CXCL2, CXCL8, PTGS2, and IL6 in BHPrS-1, but not in epithelial cell lines.ConclusionsOPN is more abundant in prostates of men with S-BPH compared to men with I-BPH. OPN secretion is stimulated by proinflammatory cytokines, and OPN acts directly on stromal cells to drive the synthesis of proinflammatory mRNAs. Pharmacological manipulation of prostatic OPN may have the potential to reduce LUTS by inhibiting both inflammatory and fibrotic pathways.

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CD8+ T cells promote proliferation of benign prostatic hyperplasia epithelial cells under low androgen level via modulation of CCL5/STAT5/CCND1 signaling pathway.

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