ROS-NLRP3 signaling pathway induces sterile inflammation after thulium laser resection of the prostate.

The sterile inflammation (SI) of the urinary tract is a common problem requiring serious consideration after prostatectomy. This study mainly focuses on the role of the reactive oxygen species-NLR family, pyrin domain-containing 3 (ROS-NLRP3) signaling pathway in SI after thulium laser resection of the prostate (TmLRP). Urinary cytokines were determined in patients who received TmLRP, and heat shock protein 70 (HSP70) was detected in the resected tissues. The involvement of ROS signaling in HSP70-induced inflammation was explored in THP-1 cells with or without N-acetyl- l-cysteine (NAC) pretreatment. The function of NLRP3 and Caspase-1 was determined by Western blot analysis, enzyme-linked immunosorbent assay (ELISA), and polymerase chain reaction. These phenomena and mechanisms were verified by the beagle models that received TmLRP. Clinical urine samples after TmLRP showed high expression of inflammatory factors and peaked 3-5 days after surgery. The high expression of HSP70 in the resected tissues was observed. After HSP70 stimulation, the expression of ROS, NLRP3, Caspase-1, and interleukin-18 (IL-18) increased significantly and could be reduced by ROS inhibitor NAC. The expression of IL-1ß and IL-18 could be inhibited by NLRP3 or Caspase-1 inhibitors. In beagle models that received TmLRP, HSP70, NLRP3, Caspase-1, IL-1ß, and IL-18 were highly expressed in the wound tissue or urine, and could also be reduced by NAC pretreatment. Activation of the ROS-NLRP3 signaling pathway induces SI in the wound after prostatectomy. Inhibition of this pathway may be effective for clinical prevention and treatment of SI and related complications after prostatectomy.

QKI-6 inhibits bladder cancer malignant behaviours through down-regulating E2F3 and NF-κB signalling.

Quaking homolog (QKI) is a member of the RNA-binding signal transduction and activator of proteins family. Previous studies showed that QKI possesses the tumour suppressor activity in human cancers by interacting with the 3'-untraslated region (3'-UTR) of various gene transcripts via the STAR domain. This study first assessed the association of QKI-6 expression with clinicopathological and survival data from bladder cancer patients and then investigated the underlying molecular mechanisms. Bladder cancer tissues (n = 223) were subjected to immunohistochemistry, and tumour cell lines and nude mice were used for different in vitro and in vivo assays following QKI-6 overexpression or knockdown. QKI-6 down-regulation was associated with advanced tumour TNM stages and poor patient overall survival. QKI-6 overexpression inhibited bladder cancer cell growth and invasion capacity, but induced tumour cell apoptosis and cell cycle arrest. Furthermore, ectopic expression of QKI-6 reduced tumour xenograft growth and expression of proliferation markers, Ki67 and PCNA. However, knockdown of QKI-6 expression had opposite effects in vitro and in vivo. QKI-6 inhibited expression of E2 transcription factor 3 (E2F3) by directly binding to the E2F3 3'-UTR, whereas E2F3 induced QKI-6 transcription by binding to the QKI-6 promoter in negative feedback mechanism. QKI-6 expression also suppressed activity and expression of nuclear factor-κB (NF-κB) signalling proteins in vitro, implying a novel multilevel regulatory network downstream of QKI-6. In conclusion, QKI-6 down-regulation contributes to bladder cancer development and progression.

Heat injured stromal cells-derived exosomal EGFR enhances prostatic wound healing after thulium laser resection through EMT and NF-κB signaling.

BACKGROUND: This study investigated shallow heat injury to prostate stromal fibroblasts and epithelial cells and their interaction to regulate the wound healing and the underlying molecular events. METHODS: Prostate stromal fibroblasts and epithelial cells were cultured individually or cocultured and subjected to shallow heat injury for assessments of cell proliferation, migration, apoptosis, cell cycle distribution, and gene expression. The supernatant of heat-injured WPMY-1 cells was collected for exosome extraction and assessments. Furthermore, beagle dogs received thulium laser resection of the prostate (TmLRP) and randomly divided into Gefitinib, GW4869, and control treatment for the histological analysis, tissue re-epithelialization, and epidermal growth factor receptor (EGFR) expression on the prostatic wound surface. Immunofluorescence was to evaluate p63-positive basal progenitor cell trans-differentiation and macrophage polarization and ELISA was to detect cytokine levels in beagles' urine. RESULTS: Shallow heat injury caused these cells to enter a stressed state and enhanced their crosstalk. The prostate stromal fibroblasts produced and secreted more exosomal-EGFR and other cytokines and chemokines after shallow heat injury, resulting in increased proliferation and migration of prostate epithelial cells during wound healing. The wound healing of the canine prostatic urethra following the TmLRP procedure was slower in the Gefitinib and GW4869 treatment group than in the control group of animals. Immunofluorescence and ELISA showed that reduced EGFR expression interrupted macrophage polarization but increased the inflammatory response. CONCLUSIONS: Shallow heat injury was able to promote the interaction of prostate stromal cells with prostate epithelial cells to enhance wound healing. Stromal-derived exosomal-EGFR plays a crucial role in the balance of the macrophage polarization and prostatic wound healing.

M1 macrophage mediated increased reactive oxygen species (ROS) influence wound healing via the MAPK signaling in vitro and in vivo.

Thulium laser resection of the prostate (TmLRP), a major treatment for benign prostatic hyperplasia (BPH), has several postoperative complications that affect the patients' quality of life. The aim of this study was to investigate the effect of the M1 macrophage-secreted reactive oxygen species (ROS) on prostatic wound healing, and the role of MAPK signaling in this process. A co-culture model in vitro was established using macrophages and prostate epithelial or stromal cells. Cell proliferation, migration, apoptosis, MAPK pathway-related gene expression levels were evaluated by standard assays. In addition, an in vivo model of prostatectomy was established in beagles by subjecting them to TmLRP, and were either treated with N-acetyl-L-cysteine (NAC) and or placebo. Wound healing and re-epithelialization were analyzed histopathologically in both groups, in addition to macrophage polarization, oxidative stress levels and MAPK pathway-related proteins expressions. Intracellular ROS levels were significantly increased in the prostate epithelial and stromal cells following co-culture with M1-like macrophages and H2O2 exposure via MAPK activation, which affected their proliferation, migration and apoptosis, and delayed the wound healing process. The cellular functions and wound healing capacity of the prostate cells were restored by blocking or clearing the macrophage-secreted ROS. In the beagle model, increased ROS levels impaired cellular functions, and appropriate removing ROS accelerated the wound healing process.

TRAF6 regulates proliferation of stromal cells in the transition and peripheral zones of benign prostatic hyperplasia via Akt/mTOR signaling.

BACKGROUND: Increased prostatic smooth muscle tone and hyperplastic growth contribute to urethral obstruction and voiding symptoms in benign prostatic hyperplasia (BPH). It has been suggested that different proliferative potential of stromal cells between transition zone (TZ) and adjoining regions of the prostate plays a significant role in the development of BPH. However, the molecular mechanisms of this hyperplastic process remain unclear. We found tumor necrosis factor receptor-associated factor 6 (TRAF6) highly expressed in TZ stromal cells compared to peripheral zone (PZ) stromal cells by gene array analyzes. Therefore, we aim to study the potential mechanisms of stromal TRAF6 in promoting BPH progression. METHODS: Stromal cells obtained from BPH-derived primary cultures. The TRAF6-siRNA vector were constructed and transfected into cultured human BPH primary TZ stromal cells, and TRAF6-overexpressing vector were constructed and transfected into cultured human BPH primary PZ stromal cells. Stromal cells were recombined with BPH-1 cells then subcutaneously inoculated into the kidney capsule of male nude mice. Cell proliferation was evaluated by CCK-8 assay. Multiple proteins in the Akt/mTOR pathway were assessed using western blot. RESULTS: TRAF6 levels were increased in TZ stroma compared with PZ stroma of BPH. The in vitro cell culture and in vivo cell recombination revealed that selective downregulation of TRAF6 in TZ stromal cells led to suppression of the proliferation, while upregulation of TRAF6 in PZ stromal cells enhanced the proliferation. We found that the Phosphorylation and Ubiquitination of Akt as well as the Phosphorylation of mTOR, P70S6K were decreased when TRAF6 was downregulated in primary cultured TZ stromal cells of BPH. CONCLUSIONS: TRAF6 can promote the proliferation of stromal cells of BPH via Akt/mTOR signaling. Our results may make stromal TRAF6 responsible for zonal characteristic of BPH and as a promising therapeutic strategy for BPH treatment.

Androgen Deprivation Accelerates the Prostatic Urethra Wound Healing After Thulium Laser Resection of the Prostate by Promoting Re-Epithelialization and Regulating the Macrophage Polarization.

BACKGROUND: Complications after a thulium laser resection of the prostate (TmLRP) are related to re-epithelialization of the prostatic urethra. Since prostate growth and development are induced by androgen, the aim of this study was to determine the role and explore the mechanism of androgen in wound healing of the prostatic urethra. METHODS: Beagles that received TmLRPs were randomly distributed into a castration group, a testosterone undecanoate (TU) group, and a control group. The prostate wound was assessed once a week using a cystoscope. Histological analysis was then carried out to study the re-epithelialization of the prostatic urethra in each group. The inflammatory response in the wound tissue and urine was also investigated. RESULTS: The healing of the prostatic urethra after a TmLRP was more rapid in the castration group and slower in the TU group than that in the control group. Castration accelerated re-epithelialization by promoting basal cell proliferation in the wound surface and beneath the wound and by accelerating the differentiation of basal cells into urothelial cells. Castration reduced the duration of the inflammatory phase and induced the conversion of M1 macrophages to M2 macrophages, thus accelerating the maturation of the wound. By contrast, androgen supplementation enhanced the inflammatory response and prolonged the inflammatory phase. Moreover, the anti-inflammatory phase was delayed and weakened. CONCLUSION: Androgen deprivation promotes re-epithelialization of the wound, regulates the inflammatory response, and accelerates wound healing of the prostatic urethra after a TmLRP. Prostate 77:708-717, 2017. © 2017 Wiley Periodicals, Inc.

[Adjuvant hormonal therapy immediately after radical surgery for high-risk organ-confined or locally advanced prostate cancer].

OBJECTIVE: To evaluate the effect of adjuvant hormonal therapy (AHT) immediately after radical surgery for high- risk organ-confined or locally advanced prostate cancer using the PSA-related biochemical relapse rate within 2 years after surgery. METHODS: We retrospectively analyzed 62 cases of high-risk organ-confined or locally advanced prostate cancer. The patients were treated by laparoscopic radical prostatectomy or radical retropubic prostatectomy after MRI and ECT systemic bone imaging examination, which revealed no regional lymph node or bone metastasis. Thirty-two of the patients (group A) received AHT orally or subcutaneously from 2 weeks to 1 months after operation, and another 30 (group B) were left untreated. We followed up the patients for 2 years, measuring the serum PSA level every 3 months, performing ECT every 6 months, and recording the adverse reactions, medication dura- tion, and the patients'quality of life. RESULTS: All the operations were successfully accomplished. The rate of 2-year biochemical relapse-free survival was 78.13% (25/32) in group A and 53.33% (16/30) in group B. CONCLUSION: AHT immediately after radical surgery can improve the rate of biochemical relapse-free survival of the patients with high-risk organ-confined or locally advanced prostate cancer and check the progression and metastasis of the disease.

Porous Se@SiO2 nanosphere-coated catheter accelerates prostatic urethra wound healing by modulating macrophage polarization through reactive oxygen species-NF-κB pathway inhibition.

Benign prostatic hyperplasia (BPH) patients experience complications after surgery. We studied oxidative stress scavenging by porous Se@SiO2 nanospheres in prostatic urethra wound healing after transurethral resection of the prostate (TURP). Beagle dogs were randomly distributed into two groups after establishing TURP models. Wound recovery and oxidative stress levels were evaluated. Re-epithelialization and the macrophage distribution at the wound site were assessed by histology. The mechanism by which porous Se@SiO2 nanospheres regulated macrophage polarization was investigated by qRT-PCR, western blotting, flow cytometry, immunofluorescence and dual luciferase reporter gene assays. Our results demonstrated that Porous Se@SiO2 nanosphere-coated catheters advance re-epithelization of the prostatic urethra, accelerating wound healing in beagle dogs after TURP, and improve the antioxidant capacity to inhibit oxidative stress and induced an M2 phenotype transition of macrophages at the wound. By restraining the function of reactive oxygen species (ROS), porous Se@SiO2 nanospheres downregulated Ikk, IκB and p65 phosphorylation to block the downstream NF-κB pathway in macrophages in vitro. Since activation of NF-κB signaling cascades drives macrophage polarization, porous Se@SiO2 nanospheres promoted macrophage phenotype conversion from M1 to M2. Our findings suggest that porous Se@SiO2 nanosphere-coated catheters promote postoperative wound recovery in the prostatic urethra by promoting macrophage polarization toward the M2 phenotype through suppression of the ROS-NF-κB pathway, attenuating the inflammatory response. STATEMENT OF SIGNIFICANCE: The inability to effectively control post-operative inflammatory responses after surgical treatment of benign prostatic hyperplasia (BPH) remains a challenge to researchers and surgeons, as it can lead to indirect cell death and ultimately delay wound healing. Macrophages at the wound site work as pivotal regulators of local inflammatory response. Here, we designed and produced a new type of catheter with a coating of porous Se@SiO2 nanosphere and demonstrated its role in promoting prostatic urethra wound repair by shifting macrophage polarization toward the anti-inflammatory M2 phenotype via suppressing ROS-NF-κB pathway. These results indicate that the use of porous Se@SiO2 nanosphere-coated catheter may provide a therapeutic strategy for postoperative complications during prostatic urethra wound healing to improve patient quality of life.

5-ARI induces autophagy of prostate epithelial cells through suppressing IGF-1 expression in prostate fibroblasts.

OBJECTIVES: 5α-reductase inhibitor (5-ARI) is a commonly used medicine in the treatment of lower urinary tract symptoms (LUTS) associated with benign prostatic hyperplasia (BPH). Our study mainly focuses on the mechanism of BPH development after 5ARI treatment. MATERIALS AND METHODS: Prostate specimens from patients were collected. Insulin-like growth factor 1 (IGF-1), Beclin-1, LC3 levels, was analysed by immunohistochemistry. The role IGF-1 on autophagic flux in prostate epithelial cells was studied. Additionally, effect of autophagy on recombinant grafts consisting of prostate stromal and epithelial cells in nude mice was investigated. RESULTS: We demonstrated that IGF-1 expression is down-regulated in prostate fibroblasts after long-term 5-ARI application. A decrease in IGF-1 levels was found to activate autophagic flux through the mTOR pathway in prostate epithelial cells, while the inhibition of IGF-1 receptor function induced autophagy in prostate epithelial cells. In addition, we revealed that blocking autophagic flux initiation can reduce the volume of recombinant grafts in vivo. Finally, our findings suggest that long-term 5-ARI application reduces IGF-1 secretion by prostatic stromal cells, thereby inducing autophagy of prostatic epithelial cells, which is one of the mechanisms underlying BPH pathogenesis and progression. CONCLUSIONS: Focusing on the autophagy induced by low levels of IGF-1 in prostatic epithelial cells, after elucidating AR signalling impairment of prostate stromal cells, might provide a novel strategy for the treatment and prevention of BPH development.

Macrophages are targets of retinoic acid signaling during the wound-healing process after thulium laser resection of the prostate.

BACKGROUND: The wound-healing process is very important for reducing complications after thulium laser resection of the prostate (TmLRP). The retinoic acid (RA) signaling pathway has been well studied in the wound-healing process of the skin and other organs. The goals of this study were to identify the role of RA signaling in the repair of the prostate after TmLRP and to investigate the molecular mechanism of this process. RESULTS: Retinoic acid receptors (RARs) were present in the prostate, and their expression was increased after TmLRP. RARß was expressed in the macrophages and may be related to the role of stromal cells in the wound-healing process. In vitro, RA enhanced the function of anti-inflammatory macrophages and promoted stromal cell activation and angiogenesis. Arg1 was also increased via RARß after treatment with RA. MATERIALS AND METHODS: The expression of RARs was analyzed in vivo by immunohistochemistry (IHC), real time qPCR, and western blot analysis. THP-1 cells were co-treated with or without RA and stimulating factor and then assessed by ELISA and qPCR. The supernatants from these cells were cultured with stromal cells and vascular endothelial cells, and the effects on these cells were analyzed. CONCLUSIONS: We found that RA signaling was involved in the wound-healing process of the prostate after TmLRP. RA treated macrophages activated stromal cells and promoted angiogenesis. RARß was the key isoform in this process.