P2X7R Modulates NEK7-NLRP3 Interaction to Exacerbate Experimental Autoimmune Prostatitis via GSDMD-mediated Prostate Epithelial Cell Pyroptosis.

Chronic prostatitis is one of the most common urologic diseases that troubles young men, with unclear etiology and ineffective treatment approach. Pyroptosis is a novel model of cell death, and its roles in chronic prostatitis are unknown. In this study, P2X7R, NEK7, and GSDMD-NT expression levels were detected in prostate tissues from benign prostate hyperplasia (BPH) patients and experiment autoimmune prostatitis (EAP) mice. P2X7R agonist, antagonist, NLRP3 inhibitor, and disulfiram were used to explore the roles of the P2X7R-NEK7-NLRP3 axis in prostate epithelial cell pyroptosis and chronic prostatitis development. We found that P2X7R, NEK7, and GSDMD-NT were highly expressed in the prostate epithelial cells of BPH patients with prostatic inflammation and EAP mice. Activation of P2X7R exacerbated prostatic inflammation and increased NLRP3 inflammasome component expressions and T helper 17 (Th17) cell proportion. Moreover, P2X7R-mediated potassium efflux promoted NEK7-NLRP3 interaction, and NLRP3 assembly and activation, which caused GSDMD-NT-mediated prostate epithelial cell pyroptosis to exacerbate EAP development. Disulfiram could effectively improve EAP by inhibiting GSDMD-NT-mediated prostate epithelial cell pyroptosis. In conclusion, the P2X7R-NEK7-NLRP3 axis could promote GSDMD-NT-mediated prostate epithelial cell pyroptosis and chronic prostatitis development, and disulfiram may be an effective drug to treat chronic prostatitis.

Botanical preparation HX109 inhibits macrophage-mediated activation of prostate epithelial cells through the CCL4-STAT3 pathway: implication for the mechanism underlying HX109 suppression of prostate hyperplasia.

Benign prostatic hyperplasia (BPH) is one of the most frequently observed diseases in the elderly male population worldwide. A variety of factors such as aging, hormonal imbalance, chronic inflammation, and oxidative stress play an important role in its pathogenesis. We have previously shown that HX109, an ethanol extract prepared from 3 plants (Taraxacum officinale, Cuscuta australis, and Nelumbo nucifera), alleviates prostate hyperplasia in the BPH rat model and suppresses AR signaling by upregulating Ca2+/CAMKKß and ATF3. In this study, we used macrophage cell lines to examine the effects of HX109 on inflammation, which is considered an important causative factor in BPH pathogenesis. In the co-culture system involving macrophage-prostate epithelial cells, HX109 inhibited macrophage-induced cell proliferation, migration and epithelial-mesenchymal transition (EMT) by inhibiting the expression of CCL4 and the phosphorylation of STAT3. Furthermore, HX109 inhibited the expression of inflammatory cytokines and the phosphorylation of p65 NF-κB in a concentration dependent manner. Taken together, our results suggested that HX109 could regulate macrophage activation and its crosstalk with prostate cells, thereby inhibiting BPH.

[Gene expression profiles in normal human prostate epithelial cells exposed to low-dose cadmium: A bioinformatics analysis].

OBJECTIVE: To analyze the biological network regulation and key proteins of gene expression microarray in human normal prostate epithelial cells after treated with low-dose cadmium, and provide some new theoretical evidence for the pathogenesis of cadmium-related prostate cancer. METHODS: We downloaded 19 copies of gene chip data from the Gene Expression Omnibus (GEO), involving 9 samples of prostate epithelial cells exposed to low-dose cadmium and 10 cases of normal control. Using the Gene-Cloud of Biotechnology Informs platform, GenClip2.0 and Sytoscape 3.5.1, we screened differentially expressed genes, explored their protein interaction networks and biological pathways and, from the perspective of transcriptome, analyzed the changes in the genetic network of normal human prostate epithelial cells and their possible molecular biological functions after low-dose cadmium treatment. RESULTS: Totally, 1 050 (1.92%) differentially expressed genes were found in the prostate epithelial cells treated with low-dose cadmium, involved in such biological functions as the cell physiological process, MAPK regulation, regulation of intracellular signal transduction, and immunological effect. The HSP90AB1, BUB3 and PRKAR1A genes were the core nodes of the protein network, which showed statistically significant differences in their expressions and a correlation with the malignant transformation of normal cells. CONCLUSIONS: Low-dose cadmium can cause genetic changes in normal human prostate epithelial cells and the differentially expressed genes are mainly involved in such biological functions as the cell physiological process, MAPK regulation, regulation of intracellular signal transduction, and immunological effect.

Gene expression profiles in normal human prostate epithelial cells exposed to low-dose cadmium: A bioinformatics analysis / 中华男科学杂志

Objective@#To analyze the biological network regulation and key proteins of gene expression microarray in human normal prostate epithelial cells after treated with low-dose cadmium, and provide some new theoretical evidence for the pathogenesis of cadmium-related prostate cancer.@*METHODS@#We downloaded 19 copies of gene chip data from the Gene Expression Omnibus (GEO), involving 9 samples of prostate epithelial cells exposed to low-dose cadmium and 10 cases of normal control. Using the Gene-Cloud of Biotechnology Informs platform, GenClip2.0 and Sytoscape 3.5.1, we screened differentially expressed genes, explored their protein interaction networks and biological pathways and, from the perspective of transcriptome, analyzed the changes in the genetic network of normal human prostate epithelial cells and their possible molecular biological functions after low-dose cadmium treatment.@*RESULTS@#Totally, 1 050 (1.92%) differentially expressed genes were found in the prostate epithelial cells treated with low-dose cadmium, involved in such biological functions as the cell physiological process, MAPK regulation, regulation of intracellular signal transduction, and immunological effect. The HSP90AB1, BUB3 and PRKAR1A genes were the core nodes of the protein network, which showed statistically significant differences in their expressions and a correlation with the malignant transformation of normal cells.@*CONCLUSIONS@#Low-dose cadmium can cause genetic changes in normal human prostate epithelial cells and the differentially expressed genes are mainly involved in such biological functions as the cell physiological process, MAPK regulation, regulation of intracellular signal transduction, and immunological effect.

Role of GPR30 in estrogen-induced prostate epithelial apoptosis and benign prostatic hyperplasia.

Several studies have implicated estrogen and the estrogen receptor (ER) in the pathogenesis of benign prostatic hyperplasia (BPH); however, the mechanism underlying this effect remains elusive. In the present study, we demonstrated that estrogen (17ß-estradiol, or E2)-induced activation of the G protein-coupled receptor 30 (GPR30) triggered Ca2+ release from the endoplasmic reticulum, increased the mitochondrial Ca2+ concentration, and thus induced prostate epithelial cell (PEC) apoptosis. Both E2 and the GPR30-specific agonist G1 induced a transient intracellular Ca2+ release in PECs via the phospholipase C (PLC)-inositol 1, 4, 5-triphosphate (IP3) pathway, and this was abolished by treatment with the GPR30 antagonist G15. The release of cytochrome c and activation of caspase-3 in response to GPR30 activation were observed. Data generated from the analysis of animal models and human clinical samples indicate that treatment with the GPR30 agonist relieves testosterone propionate (TP)-induced prostatic epithelial hyperplasia, and that the abundance of GPR30 is negatively associated with prostate volume. On the basis of these results, we propose a novel regulatory mechanism whereby estrogen induces the apoptosis of PECs via GPR30 activation. Inhibition of this activation is predicted to lead to abnormal PEC accumulation, and to thereby contribute to BPH pathogenesis.

Signalling pathways associated with IL-6 production and epithelial-mesenchymal transition induction in prostate epithelial cells stimulated with Trichomonas vaginalis.

Trichomonas vaginalis (Tv) has been found in patient tissue of benign prostatic hyperplasia (BPH), and suggested to cause chronic prostatitis. IL-6 is known as one of the important factors of chronic inflammation in prostate cancer. Patients with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) had higher levels of IL-6 in seminal plasma. Furthermore, inflammatory conditions induced by pathogen infections have been shown to promote epithelial-mesenchymal transition (EMT). Here, we investigated the signals involved in IL-6 production by human prostate epithelial cells (PECs) stimulated with Tv and examined whether Tv induces EMT in PECs. We found that PECs stimulated with Tv increased the production of IL-6, as well as the expression of TLR2, TLR4, MAPKs (p38, JNK, ERK), NF-κB and JAK2/STAT3, and levels of ROS. Inhibition of TLR2 or TLR4 reduced IL-6 production as well as expression of these other factors, and agents inhibiting ROS, MAPKs, NF-κB and JAK reduced IL-6 production. However, when PECs were stimulated with Tv, transcripts of mesenchymal cell markers increased, and epithelial cell markers decreased. In addition, the induction of EMT was suppressed by inhibitors of JAK or NF-κB. These findings are the first evidence that Tv infection of prostate epithelial cells may induce EMT.

Trichomonas vaginalis induces IL-1ß production in a human prostate epithelial cell line by activating the NLRP3 inflammasome via reactive oxygen species and potassium ion efflux.

BACKGROUND: Trichomonas vaginalis is a sexually transmitted protozoan parasite that causes vaginitis in women, and urethritis and prostatitis in men. IL-1ß is synthesized as immature pro-IL-1ß, which is cleaved by activated caspase-1. Caspase-1 is, in turn, activated by a multi-protein complex known as an inflammasome. In this study, we investigated the inflammatory response of a prostate epithelial cell line (RWPE-1) to T. vaginalis and, specifically, the capacity of T. vaginalis to activate the NLRP3 inflammasome. METHODS: RWPE-1 cells were stimulated by live T. vaginalis, and subsequent expression of pro-IL-1ß, IL-1ß, NLRP3, ASC and caspase-1 was determined by real-time PCR and Western blotting. IL-1ß and caspase-1 production was also measured by ELISA. To evaluate the effects of NLRP3 and caspase-1 on IL-1ß production, the activated RWPE-1 cells were transfected with small interfering RNAs to silence the NLRP3 and caspase-1 genes. Activation of the NLRP3 inflammasome was observed by fluorescence microscopy. Intracellular reactive oxygen species (ROS) were evaluated by spectrofluorometry. RESULTS: When RWPE-1 cells were stimulated with live T. vaginalis, the mRNA and protein expression of IL-1ß, NLRP3, ASC, and caspase-1 increased. Moreover, silencing of NLRP3 and caspase-1 attenuated T. vaginalis-induced IL-1ß secretion. The NADPH oxidase inhibitor DPI and high extracellular potassium ion suppressed the production of IL-1ß, caspase-1, and the expression of NLRP3 and ASC proteins. The specific NF-κB inhibitor, Bay 11-7082, inhibited IL-1ß production, and also inhibited the production of caspase-1, ASC and NLRP3 proteins. CONCLUSIONS: T. vaginalis induces the formation of the NLRP3 inflammasome in human prostate epithelial cells via ROS and potassium ion efflux, and this results in IL-1ß production. This is the first evidence for activation of the NLRP3 inflammasome in the inflammatory response by prostate epithelial cells infected with T. vaginalis. Prostate 76:885-896, 2016. © 2016 Wiley Periodicals, Inc.

EGFR and IGF-1R in regulation of prostate cancer cell phenotype and polarity: opposing functions and modulation by T-cadherin.

T-cadherin is an atypical glycosylphosphatidylinsoitol-anchored member of the cadherin superfamily of adhesion molecules. We found that T-cadherin overexpression in malignant (DU145) and benign (BPH-1) prostatic epithelial cell lines or silencing in the BPH-1 cell line, respectively, promoted or inhibited migration and spheroid invasion in collagen I gel and Matrigel. T-cadherin-dependent effects were associated with changes in cell phenotype: overexpression caused cell dissemination and loss of polarity evaluated by relative positioning of the Golgi/nuclei in cell groups, whereas silencing caused formation of compact polarized epithelial-like clusters. Epidermal growth factor receptor (EGFR) and IGF factor-1 receptor (IGF-1R) were identified as mediators of T-cadherin effects. These receptors per se had opposing influences on cell phenotype. EGFR activation with EGF or IGF-1R inhibition with NVP-AEW541 promoted dissemination, invasion, and polarity loss. Conversely, inhibition of EGFR with gefitinib or activation of IGF-1R with IGF-1 rescued epithelial morphology and decreased invasion. T-cadherin silencing enhanced both EGFR and IGF-1R phosphorylation, yet converted cells to the morphology typical for activated IGF-1R. T-cadherin effects were sensitive to modulation of EGFR or IGF-1R activity, suggesting direct involvement of both receptors. We conclude that T-cadherin regulates prostate cancer cell behavior by tuning the balance in EGFR/IGF-1R activity and enhancing the impact of IGF-1R.

Molecular functions of the iron-regulated metastasis suppressor, NDRG1, and its potential as a molecular target for cancer therapy.

N-myc down-regulated gene 1 (NDRG1) is a known metastasis suppressor in multiple cancers, being also involved in embryogenesis and development, cell growth and differentiation, lipid biosynthesis and myelination, stress responses and immunity. In addition to its primary role as a metastasis suppressor, NDRG1 can also influence other stages of carcinogenesis, namely angiogenesis and primary tumour growth. NDRG1 is regulated by multiple effectors in normal and neoplastic cells, including N-myc, histone acetylation, hypoxia, cellular iron levels and intracellular calcium. Further, studies have found that NDRG1 is up-regulated in neoplastic cells after treatment with novel iron chelators, which are a promising therapy for effective cancer management. Although the pathways by which NDRG1 exerts its functions in cancers have been documented, the relationship between the molecular structure of this protein and its functions remains unclear. In fact, recent studies suggest that, in certain cancers, NDRG1 is post-translationally modified, possibly by the activity of endogenous trypsins, leading to a subsequent alteration in its metastasis suppressor activity. This review describes the role of this important metastasis suppressor and discusses interesting unresolved issues regarding this protein.

Inflammatory response of prostate epithelial cells to stimulation by Trichomonas vaginalis.

BACKGROUND: Trichomonas vaginalis is known as the most common cause of sexually transmitted infection. However, its prevalence may have been underestimated. Trichomonads are detected in prostatic tissue in benign prostatic hyperplasia, prostatitis, and prostate cancer. Our objective was to investigate whether T. vaginalis could induce an inflammatory response in prostate epithelium. METHODS: The cytokine production by human prostate epithelial cell (RWPE-1) activated with T. vaginalis was determined by ELISA and real-time PCR. Intracellular ROS was evaluated by flow cytometry or spectrofluorometry. The protein levels of MAP kinase, NF-κB were analyzed by Western blot. The migration of neutrophil and monocyte were performed in 24-well microplates with filter insert. RESULTS: Incubation of cells of a human prostate epithelial cell line with a live T. vaginalis T016 isolate increased expression of the inflammatory mediators IL-1ß, CCL2, and CXCL8. In addition, ROS, MAPK, and NF-κB activities increased, while inhibitors of ROS, ERK, and NF-κB reduced IL-1ß production. Medium conditioned by incubation of RWPE-1 cells with T. vaginalis contained IL-1ß and stimulated the migration of human neutrophils and monocytes (THP-1 cell line). CONCLUSIONS: We conclude that T. vaginalis may increase IL-1ß expression in human prostate epithelium through activation of ROS, ERK, and NF-κB, and this in turn may induce the migration of neutrophils and monocytes and lead to an inflammatory response. This research is the first attempt to confirm inflammatory reaction caused by T. vaginalis in prostate epithelial cell.

Tumor necrosis factor alpha increases aerobic glycolysis and reduces oxidative metabolism in prostate epithelial cells.

BACKGROUND: Chronic inflammation promotes prostate cancer formation and progression. Furthermore, alterations in energy metabolism are a hallmark of prostate cancer cells. However, the actions of inflammatory factors on the energy metabolism of prostate epithelial cells have not been previously investigated. This is the first study to report on the effect of the inflammatory cytokine tumor necrosis factor alpha (TNFα) on the glycolytic and oxidative metabolism, and the mitochondrial function of widely used prostate epithelial cells. METHODS: Pre-malignant RWPE-1 and cancerous LNCaP and PC-3 cells were treated with low-dose TNFα. Glycolytic and oxidative metabolism was quantified by measuring extracellular acidification and oxygen consumption rates, respectively. ATP content and lactate export were measured by luminescence and fluorescence, respectively. Mitochondrial content and the expression of glucose transporter 1 (GLUT1), peroxisome proliferator-activated receptor co-activator 1 alpha (PGC-1α), and Cytochrome C were measured by flow cytometry. RESULTS: Our data suggest that TNFα increases glycolysis, ATP production, and lactate export, while it reduces oxidative metabolism and mitochondrial function in prostate epithelial cells. The highly aggressive PC-3 cells tend to be less responsive to the actions of TNFα than the pre-malignant RWPE-1 and the non-aggressive LNCaP cells. CONCLUSIONS: Cellular energetics, that is, glycolytic and oxidative metabolism is significantly influenced by low-level inflammation in prostate epithelial cells. In widely used prostate epithelial cell models, the micro-environmental inflammatory cytokine TNFα induces aerobic glycolysis while inhibiting oxidative metabolism. This supports the hypothesis that low-level inflammation can induce Warburg metabolism in prostate epithelial cells, which may promote cancer formation and progression.

Expression of somatomedin-receptor in anoxic prostate epithelial cells / 中国医师进修杂志

Objective To observe the different expression of somatomedin-receptor in cell membrane of prostate epithelial cells at anoxic or normoxic condition.Methods Human prostate epithelial cells line RWPE-1 were cultured in vitro.At 4,8,12,24,48 h after cells had been seeded,the gene and protein expression of epidermal growth factor receptor (EGFR),fibroblast growth factor receptor (FGFR),transforming growth factor β1 receptor (TGF- β 1R),insulin-like growth factor-1 receptor (IGF-1 R) and vascular endothelial growth factor receptor (VEGFR) in prostate epithelial cells were tested by RT-PCR and immunohistochem-istry methods,respectively.Results The expression of mRNA and protein of EGFR,FGFR,IGF-1R,TGF- β1R,VEGFR were significantly increased in anoxic and normoxic prostate epithelial cells (P ＜ 0.01 ).At different time point,the expression of mRNA and protein of EGFR,FGFR,IGF-1R,TGF- β1R,VEGFR significantly higher in anoxic than those in normoxic prostate epithelial cells (P＜ 0.01 )besides 4 h EGFR mRNA,12 h EGFR protein,4 h IGF-1R mRNA,4 and 8 h IGF-1R protein,4 and 8 h TGF-β 1R mRNA,4 and 8 h TGF-β 1R protein,4 h VEGFR mRNA (P ＞ 0.05).Conclusion Anoxic prostate epithelial cell can up-regulate the expression of somatomedin-receptor.