Rab25 is involved in hypospadias via the ß1 integrin/EGFR pathway.

BACKGROUND: Hypospadias is a common congenital abnormality of the penile. Abnormal regulation of critical genes involved in urethral development leads to hypospadias. We used the Rab25-/- mice and foreskin fibroblasts transfected with lentivirus in vitro and in vivo to investigate the role of Rab25 in hypospadias. METHODS: The expression levels of various molecules in tissue samples and foreskin fibroblasts were confirmed using molecular biology methods (western blotting, PCR, immunohistochemistry, etc.). A scanning electron microscope (SEM) was used to visualize the external morphology of genital tubercles (GTs) of gestation day (GD) 18.5 male wild-type (WT) and Rab25-/- mice. RESULTS: An expanded distal cleft and V-shaped urethral opening were observed in GD 18.5 Rab25-/- mice. We demonstrated that Rab25 mediated hypospadias through the ß1 integrin/EGFR pathway. In addition, silencing Rab25 inhibited cell proliferation and migration and promoted apoptosis in the foreskin fibroblasts; Ki-67- and TUNEL-positive cells were mainly concentrated near the urethral seam. CONCLUSION: These findings suggest that Rab25 plays an essential role in hypospadias by activation of ß1 integrin/EGFR pathway, and Rab25 is a critical mediator of urethral seam formation in GD18.5 male fetal mice.

Birth trauma simulated by vaginal distention induced possible irreversible changes in the composition of the fiber types in the external urethral sphincter of rats.

The external urethral sphincter (EUS) is crucial in urinary continence development. Understanding the morphological features of the EUS in female rats after vaginal distention (VD), using a model of birth trauma, would aid in evaluating its functional and metabolic properties. Our recent study demonstrated that the EUS in female rats expresses one slow (type 1) and two fast (types 2A and 2B) myosin heavy chain (MHC) isoforms. Our preliminary experiment revealed that type 2B isoform expression was markedly reduced in the EUS 4 weeks after VD. Here, we aimed to examine the expression patterns of these three types of MHC isoforms, and an embryonic MHC, a marker of regeneration fibers, in the EUS of rats 3 days and 1, 2, and 8 weeks after VD using immunofluorescence staining. Hence, type 2B fibers were selectively damaged early in post-VD and did not recover fully later. Muscle regeneration in the sphincter peaked 1 week after trauma using a marker of immature fibers, embryonic myosin heavy chain. Electron microscopy revealed that the EUS of female rats was composed of mitochondria-rich muscle fibers. Myoblasts or immature muscle fibers were discovered in the sphincter layer 1 week after trauma. These results suggest that myogenesis after VD may not contribute to restoring normal fiber composition in a female rat's EUS.

"Urethral-Sparing" Robotic Radical Prostatectomy: Critical Appraisal of the Safety of the Technique Based on the Histologic Characteristics of the Prostatic Urethra.

BACKGROUND: The prostatic urethra (PU) is conventionally resected during robot-assisted radical prostatectomy (RALP). Recent studies demonstrated the feasibility of the extended PU preservation (EPUP). AIMS: To describe the histologic features of the PU. METHODS: The PU was evaluated using cystoprostatectomy and RALP specimens. Cases of PU infiltration by prostate cancer or distortion by benign hyperplastic nodules were excluded. The thickness of the chorion and distance between the urothelium and prostate glands were measured. Prostate-specific antigen expression in the PU epithelium was evaluated with immunohistochemistry. Descriptive statistics were used. RESULTS: Six specimens of PU were examined. Histologically, the following layers of the PU were observed: (1) urothelium with basal membrane, (2) chorion, and (3) prostatic peri-urethral fibromuscular tissue. The chorion measures between 0.2 and 0.4 mm. There is not a distinct urethral muscle layer, but rather muscular fibers that originate near the prostatic stroma and are distributed around the PU. This muscular tissue appears to be mainly represented in the basal and apical urethra, but not in the middle urethra. The mean distance between the chorion and prostatic glands is 1.74 mm, with significant differences between base of the prostate, middle urethral portion, and apex (2.5 vs. 1.49 vs. 1.23 mm, respectively). PSA-expressing cells are abundant in the PU epithelium, coexisting with urothelial cells. CONCLUSIONS: The exiguity of thickness of the PU chorion, short distance from glandular tissue, and coexistence of PSA-expressing cells in the epithelium raise important concerns about the oncologic safety of EPUP.

Quantification of the Androgen and Estrogen Receptors in the Penile Tissues of Hypospadias in Comparison with Normal Children.

INTRODUCTION: Hypospadias is a common congenital abnormality typified by a proximally placed ectopic urethral meatus along the ventral surface of the penis. Androgen receptor (AR) and estrogen receptor (ER) expression in the hypospadias tissues may be altered in hypospadias. METHODOLOGY: We evaluated by immunohistochemistry the AR and ER expression in 75 tissues from hypospadias repair, and compared this expression to that of tissue from 75 patients undergoing circumcision. We also compared the intensity of AR and ER expression between different severities of hypospadias. RESULTS: AR quantitative grading score decreased with severity of hypospadias, while the ER score increased as the hypospadias worsened, which was statistically significant (p-value <0.05). CONCLUSION: The penile tissue AR expression is decreased and ER expression is increased with increasing severity of hypospadias.

Purine nucleoside phosphorylase as a target to treat age-associated lower urinary tract dysfunction.

The lower urinary tract (LUT), including the bladder, urethra and external striated muscle, becomes dysfunctional with age; consequently, many older individuals suffer from lower urinary tract disorders (LUTDs). By compromising urine storage and voiding, LUTDs degrade quality of life for millions of individuals worldwide. Treatments for LUTDs have been disappointing, frustrating both patients and their physicians; however, emerging evidence suggests that partial inhibition of the enzyme purine nucleoside phosphorylase (PNPase) with 8-aminoguanine (an endogenous PNPase inhibitor that moderately reduces PNPase activity) reverses age-associated defects in the LUT and restores the LUT to that of a younger state. Thus, 8-aminoguanine improves LUT biochemistry, structure and function by rebalancing the LUT purine metabolome, making 8-aminoguanine a novel potential treatment for LUTDs.

Effects of low-dose insulin or a soluble guanylate cyclase activator on lower urinary tract dysfunction in streptozotocin-induced diabetic rats.

AIMS: To examine the effects of low-dose insulin or a soluble guanylate cyclase activator (sGC) on lower urinary tract dysfunction (LUTD) in rats with diabetes mellitus (DM). MAIN METHODS: Female Sprague-Dawley rats were divided into non-DM control (N), DM induced by streptozotocin (65 mg/kg), with low-dose insulin (DI), DM with vehicle (D), and DM with sGC (GC) groups. In GC group, BAY 60-2770 (1 mg/kg/day) was orally administered in 6-8 weeks after DM. Voiding assay at 2, 4, and 8 weeks after DM, cystometry, and urethral pressure recordings at 8 weeks of DM were performed. mRNA levels of NO-related markers and cGMP protein levels in the urethra, and ischemia and inflammation markers in the bladder were evaluated by RT-PCR. KEY FINDINGS: Moderate levels of high blood glucose were maintained in Group DI versus Group D. The 24-h voided volume was significantly higher in Group D versus Groups N and DI. Non-voiding contractions were significantly greater, and voiding efficiency and urethral pressure reduction were significantly lower in Group D versus Groups N, DI, and GC. Urethral cGMP levels were significantly lower in Group D versus Groups N and GC. mRNA levels of PDE5 in the urethra and ischemia and inflammation markers in the bladder increased in Group D versus Group N or DI was reduced after sGC treatment. SIGNIFICANCE: DI rats with a lesser degree of bladder and urethral dysfunction might be useful as a slow-progressive DM model. sGC activation could be an effective treatment of LUTD in DM.

Distribution, shape, and immunohistochemical characteristics of serotonin-immunoreactive neuroendocrine cells in the urethra and periurethral genital organs in mice.

The aim of this study is to clarify the disibution, shape, and immunohistochemical characteristics of serotonin-immunoreactive neuroendocrine cells (SIR-NECs) in mouse prostate and in the surrounding genital organs by histological and immunohistochemical analysis of the light microscopic serial sections of urethra. We collected lower urinary tracts from 13-week-old mice and observed the distribution pattern and shape of the SIR-NECs by serial light microscopy. The organs on the sections were divided into three anatomical zones to clarify the distribution pattern of SIR-NECs: (1) zone A, the ducts near the prostatic urethra; (2) zone B, the ducts outside the urethral sphincter; and (3) zone C, the acinus areas. Sections were double immune-stained with antibodies against serotonin and one of neuroendocrine-related factors (NRFs), including 10 neural cell markers and eight neurotransmitters, and also 4',6-diamino-2-phenylindole (DAPI). In addition, SIR-NECs were double immune-stained with antibodies against cytokeratin 5 (CK5) and p63, together with DAPI. SIR-NECs were mostly localized in zone A, and no SIR-NECs were observed in zone C. The proportion of flask-shaped SIR-NECs was approximately 15% in zones A and B. No flask-shaped SIR-NECs were observed in urethral epithelia. The NRFs co-localized with SIR-NEC were calcitonin gene-related peptide, CD56, chromogranin A, neuron-specific enolase, neuron cytoplastic protein 9.5, and synaptophysin (72.3%, 73.2%, 88.9%, 92.3%, 91.7%, and 81.9%, respectively). CK5 and p63 were not co-localized with SIR-NECs.　In this study, SIR-NEC of the urethra and the surrounding genital organs was ubiquitous in the urethra and the ducts near the urethra and co-expressed specific nerve-related NRFs.

Age-associated bladder and urethral coordination impairment and changes in urethral oxidative stress in rats.

AIMS: We examined age-associated changes in bladder and urethral coordination involving the nitric oxide (NO)/soluble guanylyl cyclase (sGC) system, which induces urethral smooth muscle relaxation, and urethral ischemic/oxidative stress changes in rats. MAIN METHODS: Sixteen female Sprague-Dawley rats were divided into young (3 months old) and middle-aged (12-15 months old) groups. Urethral activity was evaluated by simultaneously recording intravesical pressure under isovolumetric conditions and urethral perfusion pressure (UPP) under urethane anesthesia. Sodium nitroprusside (SNP, 0.1 mg/kg), an NO donor, and BAY 41-2272, a novel NO-independent stimulator of sGC (0.1 mg/kg), were administered intravenously to both groups. N-nitro-l-arginine methyl ester hydrochloride (l-NAME, 100 mg/kg) was also injected intravenously, to inhibit NO synthase activity in both groups. Staining for the ischemic marker, hypoxia-inducible factor-1α (HIF-1α), and the oxidative stress markers, 8-hydroxy-2'-deoxyguanosine (8-OHdG) and malondialdehyde (MDA), was performed on tissue sections of the urethra, in both groups. KEY FINDINGS: Baseline UPP and UPP changes were significantly lower in middle-aged rats than in young rats. After administration of SNP and BAY 41-2272, baseline UPP and UPP nadir were significantly decreased in both groups. After administration of l-NAME, UPP change/bladder contraction amplitude in young rats was still lower than at baseline but was completely restored to control levels in middle-aged rats. Immunoreactivity of HIF-1α, 8-OHdG, and MDA was higher in middle-aged rats than in young rats. SIGNIFICANCE: Age-associated ischemic and oxidative stress in the urethra might be correlated with impairment of the NO/sGC system and with coordination of the bladder and urethra.

Effects of Anti-Inflammatory Nanofibers on Urethral Healing.

Protracted postsurgical inflammation leading to postoperative complications remains a persistent problem in urethral reconstruction. Nanofibers in the form of peptide amphiphiles expressing anti-inflammatory peptides (AIF-PA) have positively modulated local inflammatory responses. Urethroplasty is performed to repair 5 mm ventral urethral defects with: uncoated small intestinal submucosa (SIS); SIS dip-coated with AIF-PA1 (anti-inflammatory treatment), or SIS dip-coated with AIF-PA6 (control) on 12-week-old male Sprague Dawley rats (n = 6/group/timepoint). Animals are euthanized at 14 and 28 d postsurgery. Hematoxylin-eosin, Masson's Trichrome, and immunohistochemistry with primary antibodies against myeloperoxidase (MPO; neutrophils), CD68, CD86, CD206 (macrophages), and proinflammatory cytokines TNFα and IL-1ß are performed. Complete urethral healing occurs in 3/6 uncoated SIS (50%), 2/6 SIS+AIF-PA6 (33.3%), and 5/6 SIS+AIF-PA1 (83.3%) animals at 14 d and all at 28 d. Application of AIF-PA1 to SIS substitution urethroplasty decreases MPO+ neutrophils, CD86+ M1 proinflammatory macrophages, TNFα, and IL-1ß levels while concurrently increasing levels of CD206+ M2 proregenerative/anti-inflammatory macrophages at the anastomoses and the regenerated tissue at the wound bed (REGEN). AIF-PA1 treatment enhances the healing process, contributing to earlier, complete urethral healing, and increased angiogenesis. Further studies are needed to elucidate the specific mechanism of inflammatory response modulation on angiogenesis and overall urethral healing.

Transient receptor potential channels in sensory mechanisms of the lower urinary tract.

Disruptions to sensory pathways in the lower urinary tract commonly occur and can give rise to lower urinary tract symptoms (LUTS). The unmet clinical need for treatment of LUTS has stimulated research into the molecular mechanisms that underlie neuronal control of the bladder and transient receptor potential (TRP) channels have emerged as key regulators of the sensory processes that regulate bladder function. TRP channels function as molecular sensors in urothelial cells and afferent nerve fibres and can be considered the origin of bladder sensations. TRP channels in the lower urinary tract contribute to the generation of normal and abnormal bladder sensations through a variety of mechanisms, and have demonstrated potential as targets for the treatment of LUTS in functional disorders of the lower urinary tract.

Deletion of the lysyl oxidase-like 1 gene induces impaired elastin fiber synthesis and inefficient urethral closure in rats.

We investigated the bladder and urethral function in a rat model lacking the protein lysyl oxidase-like 1 (Loxl1). Female nulliparous rats of Loxl1-/- or age-matched wild type (WT) rats had leak-point pressure testing, cystometry, histopathological analyses of lower urinary tract, and contractile response of isolated detrusor strips to carbachol and electric field stimulation. The Loxl1-/- rats showed increased looseness and redundancy of the skin, the decreased intercontraction interval and voided volume in cystometry, the lower leak-point pressure, thinner elastic fibers of the mesentery, bladder, urethra and vagina, and smaller contractile response of detrusor strips to carbachol when compared to the WT rats. Thus, the insufficient hydrostatic mechanism of urethra via submucosal impaired elastin synthesis might reduce the resting urethral closure pressure and the diminished cholinergic contractile response of detrusor smooth muscle might be involved in bladder activity in the Loxl1-/- rats.

A Rho kinase inhibitor (Fasudil) suppresses TGF-ß mediated autophagy in urethra fibroblasts to attenuate traumatic urethral stricture (TUS) through re-activating Akt/mTOR pathway: An in vitro study.

AIMS: Transforming growth factor-ß (TGF-ß) mediated super-activation of urethra fibroblasts contributes to the progression of traumatic urethral stricture (TUS), and the Rho-associated kinase inhibitors, Fasudil, might be a novel therapeutic agent for TUS, but the underlying mechanisms had not been studied. MATERIALS AND METHODS: The primary urethral fibroblasts (PUFs) were isolated from rabbit urethral scar tissues and cultured in vitro, and the PUFs were subsequently treated with TGF-ß (10 µg/L) to simulate the realistic conditions of TUS pathogenesis. Next, the PUFs were exposed to Fasudil (50 µM) and autophagy inhibitor 3-methyladenine (3-MA) treatment. Genes expression was examined by Western Blot and immunofluorescence staining, and cellular functions were determined by MTT assay and Transwell assay. KEY FINDINGS: TGF-ß promoted cell proliferation, migration, autophagy, and secretion of extracellular matrix (ECM), including collagen I and collagen III, which were reversed by co-treating cells with both Fasudil and 3-MA. In addition, TGF-ß treatment decreased the expression levels of phosphorylated Akt (p-Akt) and mTOR (p-mTOR) to inactivate the Akt/mTOR pathway in the PUFs, which could be re-activated by Fasudil. Then, the fibroblasts were treated with the Pan-Akt inhibitor (GDC-0068), and we surprisingly found that GDC-0068 abrogated the inhibiting effects of Fasudil on cell autophagy and proliferation in the PUFs treated with TGF-ß. SIGNIFICANCE: Fasudil regulated Akt/mTOR pathway mediated autophagy to hamper TGF-ß-mediated super-activation in PUFs, which supported that Fasudil might be an ideal candidate therapeutic agent for TUS treatment for clinical utilization.

Well-differentiated neuroendocrine tumors of the lower urinary tract: biologic behavior of a rare entity.

The spectrum of neuroendocrine (NE) tumors in the genitourinary tract ranges from the aggressive large and small cell carcinomas to the often benign paraganglioma and well-differentiated neuroendocrine tumor (WD-NET). At least 15 pure lower urinary tract (LUT) WD-NETs have been described. Owing to the rarity of WD-NET in the LUT and the limited number of reported cases, a better definition of their biologic long-term behavior is warranted. Herein, we aim to describe 10 new cases of WD-NET arising in the LUT and expand on follow-up findings. Ten consultation cases were identified and included 6 men and 4 women who ranged from 45 to 73 years of age. Seven cases arose in the bladder with one located in the bladder neck, 1 arose in the prostatic urethra, 1 arose in the female urethra, and 1 arose in the left ureteral orifice. All lesions were confined to the lamina propria, and tumor architecture was pseudoglandular in all cases. Associated cystitis cystica et glandularis was identified in 5 cases; urothelial papilloma and florid von Brunn's nests were found in 2 additional cases. Immunohistochemical staining for synaptophysin and chromogranin was diffusely positive in 9 cases and focal in 1 case, and the Ki-67 proliferation index was 5% or less in all tumors. Follow-up ranged from 37 to 137 months (mean = 82; median = 77), and there was no evidence of residual disease or recurrence in any of the 10 patients during the follow-up period.

Regenerating Urethral Striated Muscle by CRISPRi/dCas9-KRAB-Mediated Myostatin Silencing for Obesity-Associated Stress Urinary Incontinence.

Overweight females are prone to obesity-associated stress urinary incontinence (OA-SUI), and there are no definitive medical therapies for this common urologic condition. This study was designed to test the hypothesis that regenerative therapy to restore urethral striated muscle (stM) and pelvic floor muscles might represent a valuable therapeutic approach. For the in vitro experiment, single-guide RNAs targeting myostatin (MSTN) were used for CRISPRi/dCas9-Kruppel associated box (KRAB)-mediated gene silencing. For the in vivo experiment, a total of 14 female lean ZUC-Leprfa 186 and 14 fatty ZUC-Leprfa 185 rats were used as control and CRISPRi-MSTN treated groups, respectively. The results indicated that lentivirus-mediated expression of MSTN CRISPRi/dCas9-KRAB caused sustained downregulation of MSTN in rat L6 myoblast cells and significantly enhanced myogenesis in vitro. In vivo, the urethral sphincter injection of lentiviral-MSTN sgRNA and lentiviral-dCas9-KRAB significantly increased the leak point pressure, the thickness of the stM layer, the ratio of stM to smooth muscle, and the number of neuromuscular junctions. Downregulation of MSTN with CRISPRi/dCas9-KRAB-mediated gene silencing significantly enhanced myogenesis in vitro and in vivo. It also improved urethral continence in the OA-SUI rat model.

Evaluation of surgical procedures of mouse urethra by visualization and the formation of fistula.

Visualization of the surgically operated tissues is vital to improve surgical model animals including mouse. Urological surgeries for urethra include series of fine manipulations to treat the increasing number of birth defects such as hypospadias. Hence visualization of the urethral status is vital. Inappropriate urethral surgical procedure often leads to the incomplete wound healing and subsequent formation of urethro-cutaneous fistula or urethral stricture. Application of indocyanine green mediated visualization of the urethra was first performed in the current study. Indocyanine green revealed the bladder but not the urethral status in mouse. Antegrade injection of contrast agent into the bladder enabled to detect the urethral status in vivo. The visualization of the leakage of contrast agent from the operated region was shown as the state of urethral fistula in the current hypospadias mouse model and urethral stricture was also revealed. A second trial for contrast agent was performed after the initial operation and a tendency of accelerated urethral stricture was observed. Thus, assessment of post-surgical conditions of urogenital tissues can be improved by the current analyses on the urethral status.

Clinician-collected urethra swab: A good alternative sample type using cobas 4800 system for Chlamydia trachomatis and Neisseria gonorrhoeae in men.

BACKGROUND: Nucleic acid amplification tests (NAATs) are being used increasing to detection of CT (Chlamydia trachomatis) and NG (Neisseria gonorrhoeae) infections for superior sensitivity and specificity than other tests. Male first-void urine (FVU) sample is the optimal sample type for detection of CT and NG by NAATs. Although not being the recommended by NAATs, clinician-collected urethra swab (CCUS) is perhaps a good alternative sample type compared with the FVU sample in men. METHODS: Paired samples (FVU and CCUS) from one hundred male outpatients were simultaneously detected by urine pattern and swab pattern using cobas 4800 CT/NG assay on cobas 4800 system for the detection of CT and NG, respectively. And twenty-one positive controls were also detected on cobas 4800 system. RESULTS: The CT/NG cycle thresholds (Ct) value of urine pattern is lower than that of swab pattern for the same positive samples (clinical samples and positive controls) on the cobas 4800 CT/NG assay. The final CT/NG results of two sample patterns from patients were highly consistent except for four discordant results. CONCLUSION: CCUS is validated for a good alternative sample type for the CT/NG detection on the cobas 4800 system in this study.

A comparison of prostatic development in xenografts of human fetal prostate and human female fetal proximal urethra grown in dihydrotestosterone-treated hosts.

The goal of this paper is to explore the ability of the human female urogenital sinus immediately below the bladder (proximal urethra) to undergo prostatic development in response to dihydrotestosterone (DHT). To establish this idea, xenografts of human fetal female proximal urethra were grown in castrated nude mouse hosts receiving a subcutaneous DHT pellet. To verify the prostatic nature of the resultant glands, DHT-treated human fetal female urethral xenografts were compared with human fetal prostatic xenografts (derived from male specimens) grown in untreated and DHT-treated castrated mouse hosts and human fetal female proximal urethral xenografts grown in untreated castrated hosts. The resultant glands observed in DHT-treated human fetal female proximal urethral xenografts expressed 3 prostate-specific markers, NKX3.1, prostate specific antigen and prostatic acid phosphatase as well as the androgen receptor. Glands induced by DHT exhibited a protein expression profile of additional immunohistochemical markers (seven keratins, RUNX1, ESR2, TP63 and FOXA1) consistent with the unique spatial pattern of these proteins in prostatic ducts. Xenografts of human fetal female proximal urethra grown in DHT-treated hosts also expressed one of the salient features of prostatic development, namely androgen responsiveness. The experimental induction of prostatic differentiation from human fetal female proximal urethra makes possible future in-depth analysis of the molecular pathways directly involved in initiation of human prostatic development and subsequent epithelial differentiation, and more important whether the molecular pathways involved in human prostatic development are similar/identical versus different from that in murine prostatic development.

Bladder outlet obstruction disrupts circadian bladder function in mice.

The circadian clock programs daily rhythms and coordinates multiple behavioural processes, including micturition. Partial bladder outlet obstruction (pBOO) in mice produces hyperactive voiding. However, long-term effects of pBOO on bladder function have not been clarified. In this study, we investigated micturition under conditions of impaired circadian bladder function by inducing long-term pBOO by tying the proximal urethra. Micturition behavior was evaluated at 1, 3, 6 and 12 months after surgery. We used automated voided stain on paper method for a precise micturition recording for mice. And quantitative assessment of gene expression was performed at 24 months after pBOO surgery using qRT-PCR procedure. The micturition frequencies in the pBOO group were significantly decreased at 3, 6, and 12 months compared to those at 1 month after operation in the same group (p < 0.05). Body weight of pBOO mice was significantly increased compared to sham operated mice at 12 months. The expression level of mRNA was exhibited a 3.4-fold nominal increased for a 5-HT2B receptor in the pBOO group compared to the sham group. The current study found that long-term pBOO led to disruption of the circadian bladder function (the day/night cycle) in mice, similar to those observed in human as nocturia. This disruption is possible involvement of the gain of body weight and/or serotonergic alteration after pBOO.

Urethral luminal epithelia are castration-insensitive cells of the proximal prostate.

BACKGROUND: Castration-insensitive epithelial progenitors capable of regenerating the prostate have been proposed to be concentrated in the proximal region based on facultative assays. Functional characterization of prostate epithelial populations isolated with individual cell surface markers has failed to provide a consensus on the anatomical and transcriptional identity of proximal prostate progenitors. METHODS: Here, we use single-cell RNA sequencing to obtain a complete transcriptomic profile of all epithelial cells in the mouse prostate and urethra to objectively identify cellular subtypes. Pan-transcriptomic comparison to human prostate cell types identified a mouse equivalent of human urethral luminal cells, which highly expressed putative prostate progenitor markers. Validation of the urethral luminal cell cluster was performed using immunostaining and flow cytometry. RESULTS: Our data reveal that previously identified facultative progenitors marked by Trop2, Sca-1, KRT4, and PSCA are actually luminal epithelial cells of the urethra that extend into the proximal region of the prostate, and are resistant to castration-induced androgen deprivation. Mouse urethral luminal cells were identified to be the equivalent of previously identified human club and hillock cells that similarly extend into proximal prostate ducts. Benign prostatic hyperplasia (BPH) has long been considered an "embryonic reawakening," but the cellular origin of the hyperplastic growth concentrated in the periurethral region is unclear. We demonstrate an increase in urethral luminal cells within glandular nodules from BPH patients. Urethral luminal cells are further increased in patients treated with a 5-α reductase inhibitor. CONCLUSIONS: Our data demonstrate that cells of the proximal prostate that express putative progenitor markers, and are enriched by castration in the proximal prostate, are urethral luminal cells and that these cells may play an important role in the etiology of human BPH.

Preparation and Characterization of Nano-Laponite/PLGA Composite Scaffolds for Urethra Tissue Engineering.

The purpose of this study was to construct a biomimetic urethral repair substitute. The nano-Laponite/polylactic acid-glycolic acid copolymer (PLGA) fiber scaffolds were produced to replicate the natural human urethra tissue microenvironment. PLGA (molar ratio 50:50) and Laponite were used in this study as raw materials. The nano-Laponite/PLGA scaffolds were fabricated via electrospinning technology. After preparing the material, the microstructural and mechanical properties of the nano-Laponite/PLGA scaffold were tested via scanning electron microscopy and electronic universal testing. The effects of different amounts of Laponite on the degradation of the nano-Laponite/PLGA scaffold were studied. Human umbilical vein endothelial cells (HUVECs) were co-cultured with PLGA and nano-Laponite/PLGA scaffolds for 24, 48, or 72 h. Scanning electron microscopy results illustrated that the microstructure of the scaffold fabricated by electrospinning was similar to that of the natural extracellular matrix. When the electrospinning liquid contained 10% Laponite, the nano-Laponite/PLGA stress-strain curve illustrated that the scaffold has strong elastic deformation ability. HUVECs exhibited good growth on the nano-Laponite/PLGA scaffold. When the scaffold contained 1% Laponite, the cell proliferation rate in the CCK-8 test was significantly better than that for the other three materials, displaying good cell culture characteristics. The 1% nano-Laponite/PLGA composite scaffold can be used as a suitable urethral repair material, but its performance requires further development and research.