BACKGROUND:Benign prostatic hyperplasia (BPH) is the most common disease in aging males. BPH growth is androgen-dependent, and it appears that androgens preferentially stimulate the growth of BPH as compared to normal prostate. The mechanism of preferential androgen stimulation of BPH growth is not clear. Our previous studies suggest that primary BPH stromal cells, but not the paired normal adjacent prostate (NAP) stromal cells from the same patients, could stimulate prostatic epithelial cell proliferation in 3D co-culture. However, whether androgen regulation of prostatic stromal cells is altered in BPH nodules is not clear. METHODS:Paired BPH and NAP primary stromal cells were prepared from the same patients. Early passage paired BPH and NAP primary cells were treated with androgens and/or antiandrogens for RNA-seq analysis. RNA-seq data were analyzed using principal component analysis (PCA) and functional enrichment analysis. Conditioned medium of the paired BPH and NAP primary cells in the presence or absence of androgens were used in three-dimensional (3D) culture of benign prostatic epithelial cells. RESULTS:RNA-seq data analysis suggests that androgen-induced gene expression in BPH primary stromal cells is heightened compared to paired NAP stromal cells. AR activation in BPH stromal cells appears to enhance cellular motility, vascular and extracellular matrix remodeling, and paracrine signaling, while simultaneously modulating cell cycle and metabolic processes. Compared to regular BPH stromal conditioned medium, conditioned medium of androgen-stimulated BPH stromal cells further enhanced prostatic epithelial cell proliferation in 3D culture. In contrast, medium of NAP stromal cell culture conditioned in the presence of androgen had no significant effect on prostatic epithelial cell proliferation in 3D culture. CONCLUSIONS:Our findings suggest that androgen stimulation of prostatic epithelial cell growth via paracrine prostatic stromal signaling is enhanced in BPH and is associated with heightened androgen-responsiveness of BPH stromal cells.

Lower urinary tract symptoms (LUTS), associated with benign prostatic hyperplasia (BPH), are an aging-related disease, with more than 210 million cases worldwide. Estrogen exposure and estrogen regulation have been implicated in a variety of disease processes, with estrogen receptor (ER)-α pathways associated with disease progression and ERβ pathways considered to be disease-protective through enhanced apoptosis and reduced cellular proliferation. Preclinical models of LUTS/BPH have shown that ERα activation contributes to disease initiation and progression. Self-identified African American (AA) men have a high incidence of LUTS/BPH, with increased incidence of non-surgical treatment failure, larger prostates at time of surgery, and surgery occurring at a younger age compared with self-identified European American (EA) men. While circulating estrogen levels are higher in AA individuals, regulation of ERs, particularly ERβ, in normal and LUTS/BPH human prostate has not been well characterized. In this study, we examined differences in ER expression between peripheral zone (PZ) and transition zone (TZ) prostate tissues using multiplex, multispectral imaging. Additionally, we assessed changes in ERs and steroid metabolism genes involved in ERβ signaling between normal and LUTS/BPH prostate samples. Our study revealed underlying differences in steroid metabolism gene expression between normal AA and EA prostates, which were further altered with LUTS/BPH. Importantly, the contribution of ERα to LUTS/BPH was more pronounced in EA prostate samples, whereas AA prostate samples exhibited an overall increase in the expression of both ER and estrogen metabolism-related genes. Although estrogens have also been implicated in collagen deposition in the prostate of LUTS/BPH patients, we did not observe significant differences in collagen deposition between AA and EA samples. These results suggest that racial differences in steroid hormone signaling pathways within the benign prostate represent a promising area for the development of precision-based therapies to reduce LUTS in aging men. © 2025 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Glucocorticoid receptor (GR) plays important roles in many diseases including prostate cancer. Intracellular shuttling of GR is thought to be an important mechanism regulating its localization to the nucleus required for transactivation of GR target genes. Here, using fluorescent microscopy coupled with pulse-chase and nucleocytoplasmic fractionation coupled with western blot, we provided evidence that GR can be imported and then degraded in the nucleus in the absence of ligand. We also showed that nuclear GR was stabilized by glucocorticoid hormone and that hormone withdrawal caused nuclear GR degradation, but not export. Further analysis showed that GR ubiquitination occurred predominantly in the nucleus compared with cytoplasm and was suppressed by glucocorticoids. Using small interfering RNA knockdown, we showed that loss of E3 ligase CHIP significantly inhibited GR ubiquitination and degradation in the nucleus, while enhancing the expression of GR target gene SGK1. These findings support an updated model that GR nucleocytoplasmic trafficking is a 1-way trip, involving nuclear import but not export. Future studies should focus on defining the mechanisms regulating GR ubiquitination and degradation in the nucleus, which may lead to novel approaches to modulate GR function for disease treatment. SIGNIFICANCE STATEMENT: This study suggests that glucocorticoid receptor (GR) nucleocytoplasmic trafficking is a 1-way trip, involving nuclear import but not export. This will guide future studies on defining the mechanisms regulating GR nuclear localization, which may lead to novel approaches to modulate GR function for disease treatment.

Declining mitochondrial function is an established feature of aging and contributes to most aging-related diseases through its impact on various pathologies such as chronic inflammation, fibrosis and cellular senescence. Our recent work suggests that benign prostatic hyperplasia, which is an aging-related disease frequently associated with inflammation, fibrosis and senescence, is characterized by a decline in mitochondrial function. Here, we utilize glycolytic restriction and pharmacologic inhibition of the mitochondrial electron transfer chain complex I to promote mitochondrial dysfunction and identify the cellular processes impacted by declining mitochondrial function in benign prostate stromal cells. Using this model, we show that mitochondrial dysfunction induced alterations in cell-cell and cell-matrix adhesion, elevated fibronectin expression, resistance to anoikis and stress-induced premature senescence (SIPS). We also showed that ablation of ZC3H4, a transcription termination factor implicated in anoikis-resistance and reduced in BPH relative to normal prostates, phenocopied various phenotypes in the human BHPrS1 prostate stromal cell line that resulted from inhibition of complex I. Furthermore, ZC3H4 ablation resulted in the elevation of mitochondrial superoxide (mtROS) and mitochondrial membrane potential, altered mitochondrial morphology and NAD+/NADH ratio, and reduced CI function in BHPrS1 cells. Thus, ZC3H4 loss promotes mitochondrial dysfunction to drive pathophysiologic changes in the stromal compartment that are features of the aging prostate.

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research & Pathophysiology (PD03)1 May 2024PD03-06 MITOCHONDRIAL COMPLEX I PROTEIN NDUFS3 IS DECREASED IN BENIGN PROSTATIC HYPERPLASIA PATIENTS TREATED WITH CELECOXIB AND/OR FINASTERIDE Chandler N. Hudson, Teresa T. Liu, Taro Igarashi, Nathalie El-Khoury, Nnamdi Ihejirika, Kelsey Paxton, Juliann Jaumotte, Rajiv Dhir, Joel B. Nelson, Donald B. DeFranco, William A. Ricke, Naoki Yoshimura, Zhou Wang, and Laura E. Pascal Chandler N. HudsonChandler N. Hudson , Teresa T. LiuTeresa T. Liu , Taro IgarashiTaro Igarashi , Nathalie El-KhouryNathalie El-Khoury , Nnamdi IhejirikaNnamdi Ihejirika , Kelsey PaxtonKelsey Paxton , Juliann JaumotteJuliann Jaumotte , Rajiv DhirRajiv Dhir , Joel B. NelsonJoel B. Nelson , Donald B. DeFrancoDonald B. DeFranco , William A. RickeWilliam A. Ricke , Naoki YoshimuraNaoki Yoshimura , Zhou WangZhou Wang , and Laura E. PascalLaura E. Pascal View All Author Informationhttps://doi.org/10.1097/01.JU.0001009388.01015.e5.06AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Benign prostatic hyperplasia (BPH) is treated with multiple medical therapies and surgical options with variable outcomes. Some of the heterogeneity in treatment efficacy may come from an incomplete understanding of their molecular and cellular impact. A decline in essential mitochondrial protein expression has been observed in BPH tissues, suggesting mitochondrial dysfunction may be a factor in BPH pathogenesis. Both finasteride and celecoxib have been associated with mitochondrial toxicity in other tissues. The objective of this study was to determine whether the BPH therapies finasteride and celecoxib might impact mitochondrial function in prostate tissues, specifically their effects on NDUFS3. METHODS: NDUFS3, E-cadherin and inflammatory cell immunostaining were performed on simple prostatectomy BPH specimens from patients naïve to androgen manipulation who were treated with celecoxib and/or finasteride for 28 days in a four-arm, phase II randomized, single-blind clinical trial. NDUFS3 and inflammatory cell staining was also conducted in a murine model of celecoxib treatment in male mice. Quantification analyses of immunostaining were performed. RESULTS: NDUFS3 was decreased in BPH compared to normal adjacent prostate. Patients treated with celecoxib and/or finasteride had significantly decreased NDUFS3, and however there was no change in inflammatory cell infiltration compared to untreated patients. Mice treated with celecoxib also displayed a significant decrease in NDUFS3 immunostaining and no change in inflammatory cell infiltration. CONCLUSIONS: These findings suggest that celecoxib and/or finasteride are associated with a decline in NDUFS3 in prostate tissues but no change in inflammatory cell infiltration. Given that BPH has recently been associated with increased prostatic mitochondrial dysfunction, therapeutic targeting of BPH with celecoxib and/or finasteride may exacerbate existing mitochondrial dysfunction in BPH patients. Future development of therapeutic strategies improving mitochondrial function may improve the efficacy of celecoxib and/or finasteride in BPH patients. Source of Funding: This work was funded in part by NIH grants K01 AG059899 from NIA to TL; U54 DK104310, R01 DK131175, and R01 DK127081 from NIDDK to WAR; U54 DK112079 and R56 DK107492 from NIDDK to ZW; and CAIRIBU Convergence Award to LEP from NIDDK. This project also used the Pitt Biospecimen Core and was supported in part by award P30CA047904 © 2024 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 211Issue 5SMay 2024Page: e78 Advertisement Copyright & Permissions© 2024 by American Urological Association Education and Research, Inc.Metrics Author Information Chandler N. Hudson More articles by this author Teresa T. Liu More articles by this author Taro Igarashi More articles by this author Nathalie El-Khoury More articles by this author Nnamdi Ihejirika More articles by this author Kelsey Paxton More articles by this author Juliann Jaumotte More articles by this author Rajiv Dhir More articles by this author Joel B. Nelson More articles by this author Donald B. DeFranco More articles by this author William A. Ricke More articles by this author Naoki Yoshimura More articles by this author Zhou Wang More articles by this author Laura E. Pascal More articles by this author Expand All Advertisement PDF downloadLoading ...

Background: Older men frequently develop lower urinary tract symptoms attributed to benign prostatic hyperplasia (LUTS/BPH). Risk factors for LUTS/BPH include sedentary lifestyle, anxiety/depression, obesity, and frailty, which all increase with age. Although physical exercise may reduce the progression and/or severity of LUTS/BPH, the age-related mechanisms responsible remain unknown. Methods: Voiding symptoms, body mass, and frailty were assessed after 4-weeks of voluntary wheel running in 2-month (n = 10) and 24-month (n = 8) old C57Bl/6J male mice. In addition, various social and individual behaviors were examined in these cohorts. Finally, cellular and molecular markers of inflammation and mitochondrial protein expression were assessed in prostate tissue and systemically. Results: Despite running less (aged vs young X = 12.3 vs 30.6 km/week; p = .04), aged mice had reduced voiding symptoms (X = 67.3 vs 23.7; p < .0001) after 1 week of exercise, which was sustained through week 4 (X = 67.3 vs 21.5; p < .0001). Exercise did not affect voiding symptoms in young mice. Exercise also increased mobility and decreased anxiety in both young and aged mice (p < .05). Exercise decreased expression of a key mitochondrial protein (PINK1; p < .05) and inflammation within the prostate (CD68; p < .05 and plasminogen activator inhibitor-1; p < .05) and in the serum (p < .05). However, a frailty index (X = 0.17 vs 0.15; p = .46) and grip strength (X = 1.10 vs 1.19; p = .24) were unchanged after 4 weeks of exercise in aged mice. Conclusions: Voluntary aerobic exercise improves voiding behavior and mobility, and decreases prostatic mitochondrial protein expression and inflammation in aged mice. This promising model could be used to evaluate molecular mechanisms of aerobic exercise as a novel lifestyle intervention for older men with LUTS/BPH.

BACKGROUND:Our research focused on the assessment of the impact of systemic inhibition of Trk receptors, which bind to nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), on bladder hypersensitivity in two distinct rodent models of prostatic inflammation (PI). METHODS:Male Sprague-Dawley rats were divided into three groups (n = 6 each): the control group (no PI, vehicle administration), the untreated group (PI, vehicle administration), and the treated group (PI, nonselective Trk inhibitor, GNF 5837, administration). PI in rats was induced by a intraprostatic injection of 5% formalin. Posttreatment, we carried out conscious cystometry and a range of histological and molecular analyses. Moreover, the study additionally evaluated the effects of a nonselective Trk inhibitor on bladder overactivity in a mouse model of PI, which was induced by prostate epithelium-specific conditional deletion of E-cadherin. RESULTS:The rat model of PI showed upregulations of NGF and BDNF in both bladder and prostate tissues in association with bladder overactivity and inflammation in the ventral lobes of the prostate. GNF 5837 treatment effectively mitigated these PI-induced changes, along with reductions in TrkA, TrkB, TrkC, and TRPV1 mRNA expressions in L6-S1 dorsal root ganglia. Also, in the mouse PI model, GNF 5837 treatment similarly improved bladder overactivity. CONCLUSIONS:The findings of our study suggest that Trk receptor inhibition, which reduced bladder hypersensitivity and inflammatory responses in the prostate, along with a decrease in overexpression of Trk and TRPV1 receptors in sensory pathways, could be an effective treatment strategy for male lower urinary tract symptoms associated with PI and bladder overactivity.

BACKGROUND:Benign prostatic hyperplasia (BPH) is a condition generally associated with advanced age in men that can be accompanied by bothersome lower urinary tract symptoms (LUTS) including intermittency, weak stream, straining, urgency, frequency, and incomplete bladder voiding. Pharmacotherapies for LUTS/BPH include alpha-blockers, which relax prostatic and urethral smooth muscle and 5ɑ-reductase inhibitors such as finasteride, which can block conversion of testosterone to dihydrotestosterone thereby reducing prostate volume. Celecoxib is a cyclooxygenase-2 inhibitor that reduces inflammation and has shown some promise in reducing prostatic inflammation and alleviating LUTS for some men with histological BPH. However, finasteride and celecoxib can reduce mitochondrial function in some contexts, potentially impacting their efficacy for alleviating BPH-associated LUTS. METHODS:To determine the impact of these pharmacotherapies on mitochondrial function in prostate tissues, we performed immunostaining of mitochondrial Complex I (CI) protein NADH dehydrogenase [ubiquinone] iron-sulfur protein 3 (NDUFS3) and inflammatory cells on BPH specimens from patients naïve to treatment, or who were treated with celecoxib and/or finasteride for 28 days, as well as prostate tissues from male mice treated with celecoxib or vehicle control for 28 days. Quantification and statistical correlation analyses of immunostaining were performed. RESULTS:NDUFS3 immunostaining was decreased in BPH compared to normal adjacent prostate. Patients treated with celecoxib and/or finasteride had significantly decreased NDUFS3 in both BPH and normal tissues, and no change in inflammatory cell infiltration compared to untreated patients. Mice treated with celecoxib also displayed a significant decrease in NDUFS3 immunostaining and no change in inflammatory cell infiltration. CONCLUSIONS:These findings suggest that celecoxib and/or finasteride are associated with an overall decrease in NDUFS3 levels in prostate tissues but do not impact the presence of inflammatory cells, suggesting a decline in mitochondrial CI function in the absence of enhanced inflammation. Given that BPH has recently been associated with increased prostatic mitochondrial dysfunction, celecoxib and/or finasteride may exacerbate existing mitochondrial dysfunction in some BPH patients thereby potentially limiting their overall efficacy in providing metabolic stability and symptom relief.

BACKGROUND Age is the greatest risk factor for lower urinary tract symptoms attributed to benign prostatic hyperplasia (LUTS/BPH). While LUTS/BPH can be managed with pharmacotherapy, treatment failure has been putatively attributed to numerous pathological features of BPH (e.g., prostatic fibrosis, inflammation). Mitochondrial dysfunction is a hallmark of aging, however its impact on the pathological features of BPH remains unknown. METHODS Publicly available gene array data was analyzed. Immunohistochemistry examined mitochondrial proteins in human prostate. The effect of complex I inhibition (rotenone) on a prostatic cell line was examined using qPCR, immunocytochemistry, and Seahorse assays. Oleic acid was tested as a bypass of complex I inhibition. Aged mice were treated with OA to examine its effects on urinary dysfunction. Voiding was assessed longitudinally, and a critical complex I protein measured. RESULTS Mitochondrial function and fibrosis genes were altered in BPH. Essential mitochondrial proteins (i.e., VDAC1/2, PINK1 and NDUFS3) were significantly (P<0.05) decreased in BPH. Complex I inhibition in cultured cells resulted in decreased respiration, altered NDUFS3 expression, increased collagen deposition and gene expression. Oleic acid ameliorated these effects. Oleic acid treated aged mice had significantly (P<0.05) improved voiding function and higher prostatic NDUFS3 expression. CONCLUSION Complex I dysfunction is a potential contributor to fibrosis and lower urinary tract dysfunction in aged mice. Oleic acid partially bypasses complex I inhibition and therefore should be further investigated as a mitochondrial modulator for treatment of LUTS/BPH. Hypotheses generated in this investigation offer a heretofore unexplored cellular target of interest for the management of LUTS/BPH.

Supplemental Figure S2 shows the co-immunoprecipitation of SBD and AR truncated mutations.

Supplementary Figure from (+)-JJ-74–138 is a Novel Noncompetitive Androgen Receptor Antagonist

Supplementary Figures S1-S7

Background: Risk factors for prostate cancer include age, environment, race and ethnicity. Genetic vari-ants in cyclic-adenosine-monophosphate-response-element-binding protein 3 regulatory factor (CREBRF) gene are frequently observed in Pacific Islanders, a population with elevated prostate cancer incidence. CREBRF has been shown to play a role in other cancers, however its function in prostate homeostasis and tumorigenesis has not been previously explored. We determined the incidence of CREBRF alterations in publicly available databases and examined the impact of CREBRF deletion on the murine prostate in order to determine whether CREBRF impacts prostate physiology or pathophysiology. Methods: Alterations in CREBRF were identified in prostate cancer patients via in silico analysis of several publicly available datasets through cBioPortal. Male Crebrf knockout and wild-type littermate mice were generated and examined for prostate defects at 4 months of age. Immunohistochemical stain-ing of murine prostate sections was used to determine the impact of Crebrf knockout on proliferation, apoptosis, inflammation and blood vessel density in the prostate. Serum adipokine levels were measured using a Luminex Multiplex Assay. Results: CREBRF alterations were identified in up to 4.05% of prostate tumors and the mutations identified were categorized as likely damaging. Median survival of prostate cancer patients with genetic alterations in CREBRF was 41.23 months, compared to 131 months for patients without these changes. In the murine model, the prostates of Crebrf knockout mice had reduced epithelial proliferation and increased TUNEL+ apoptotic cells. Circulating adipokines PAI-1 and MCP-1 were also altered in Crebrf knockout mice compared to age-matched con-trols. Conclusions: Prostate cancer patients with genetic alterations in CREBRF had a significantly decreased overall survival suggesting that wild type CREBRF may play a role in limiting prostate tumorigenesis and progression. The murine knockout model demonstrated that CREBRF could modulate proliferation and apoptosis and macrophage density in the prostate. Serum levels of adipokines PAI-1 and MCP-1 were also altered and may contribute to the phenotypic changes observed in the prostates of Crebrf knockout mice. Future studies focused on populations susceptible to CREBRF mutations and mechanistic studies will be required to fully elucidate the potential role of CREBRF in prostate tumorigenesis.

Supplemental Figure S4 shows the effect of 18BBQU on AR protein level in C4-2 cells.

Supplemental Figure S3 shows the effect of Hsp70 knockdown on AR protein level in LNCaP cells.

Supplemental Figure S1 shows the Hsp70 and Hsc70 peptide sequences identified by proteomics.

AbstractIdentification of novel androgen receptor (AR) antagonists may lead to urgently needed new treatments for patients with prostate cancer resistant to current AR antagonists. AR is presently the main target for treating prostate cancer. Clinically approved AR antagonists compete with dihydrotestosterone (DHT) for binding to the ligand-binding domain (LBD) of AR, and patients eventually develop resistance to these treatments. One approach to overcoming resistance is to discover compounds that inhibit AR in alternative ways. Our lab previously identified a small molecule, JJ-450, that is capable of inhibiting AR lacking LBD. To optimize the efficacy of this class of inhibitors, we developed structural analogues of JJ-450 and identified (+)-JJ-74–138 as a promising candidate. Here, we show that (+)-JJ-74–138 is more potent than JJ-450 in the inhibition of androgen-independent AR activity in enzalutamide-resistant LN95 cells. Further studies showed (+)-JJ-74–138 inhibition of castration-resistant PSA expression in all tested castration-resistant prostate cancer (CRPC) cells. (+)-JJ-74–138 inhibited mRNA expression of AR and ARv7 target genes and reduced AR level in the nucleus in the absence of androgens. Also, this analogue noncompetitively inhibited androgen-stimulated AR activity in C4–2, LN95, and 22Rv1 CRPC cells. At low dosages, (+)-JJ-74–138 inhibited the proliferation of enzalutamide-resistant AR-positive LN95 and 22Rv1 cells, but not AR-negative PC3 and DU145 cells. A surface plasmon resonance assay detected (+)-JJ-74–138 binding to AR and a chromatin immunoprecipitation assay indicated (+)-JJ-74–138 inhibited AR binding to androgen response elements. In addition, (+)-JJ-74–138 inhibited 22Rv1 xenograft tumor growth. Our observations suggest that (+)-JJ-74–138 is a novel noncompetitive AR antagonist capable of inhibiting enzalutamide-resistant CRPC.

Background: Prostatic inflammation is closely linked to the development and progression of benign prostatic hyperplasia (BPH). Clinical studies of non-steroidal anti-inflammatory drugs, which inhibit cyclooxygenase-2 (COX-2), targeting prostate inflammation patients with symptomatic BPH have demonstrated conflicting results, with some studies demonstrating symptom improvement and others showing no impact. Thus, understanding the role of the cyclooxygenases in BPH and prostatic inflammation is important. Methods: The expression of COX-1 was analyzed in a cohort of donors and BPH patients by immunohistochemistry and compared to previously determined characteristics for this same cohort. The impact of mitochondrial dysfunction on COX-1 and COX-2 was determined in experiments treating human benign prostate epithelial cell lines BPH-1 and RWPE-1 with rotenone and MitoQ. RWPE-1 cells were transfected with small interfering RNA specific to complex 1 gene NDUFS3. Results: COX-1 expression was increased in the epithelial cells of BPH specimens compared to young healthy organ donor and normal prostate adjacent to BPH and frequently co-occurred with COX-2 alteration in BPH patients. COX-1 immunostaining was associated with the presence of CD8+ cytotoxic T-cells, but was not associated with age, prostate size, COX-2 or the presence of CD4+, CD20+ or CD68+ inflammatory cells. In cell line studies, COX protein levels were elevated following treatment with inhibitors of mitochondrial function. MitoQ significantly decreased mitochondrial membrane potential in RWPE-1 cells. Knockdown of NDUFS3 stimulated COX-1 expression. Conclusion: Our findings suggest COX-1 is elevated in BPH epithelial cells and is associated with increased presence of CD8+ cytotoxic T-cells. COX-1 can be induced in benign prostate epithelial cells in response to mitochondrial complex I inhibition, and knockdown of the complex 1 protein NDUFS3. COX-1 and mitochondrial dysfunction may play more of a role than previously recognized in the development of age-related benign prostatic disease.