Three-dimensional ultrastructural and anatomical analysis of prostatic neuroendocrine cells in mice.

BACKGROUND: A few studies have examined the ultrastructure of prostatic neuroendocrine cells (NECs), and no study has focused on their ultrastructure in three dimensions. In this study, three-dimensional ultrastructural analysis of mouse prostatic NECs was performed to clarify their anatomical characteristics. METHODS: Three 13-week-old male C57BL/6 mice were deeply anesthetized, perfused with physiological saline and 2% paraformaldehyde, and then placed in 2.5% glutaraldehyde in 0.1 M cacodylate (pH 7.3) buffer for electron microscopy. After perfusion, the lower urinary tract, which included the bladder, prostate, coagulation gland, seminal vesicle, upper vas deferens, and urethra, was removed, and the specimen was cut into small cubes and subjected to postfixation and en bloc staining. Three-dimensional ultrastructural analysis was performed on NECs, the surrounding cells, tissues, and nerves using focused ion beam/scanning electron microscope tomography. RESULTS: Twenty-seven serial sections were used in the present study, and 32 mouse prostatic NECs were analyzed. Morphologically, the NECs could be classified into three types: flask, flat, and closed. Closed-shaped NECs were always adjacent to flask-shaped cells. The flask-shaped and flat NECs were in direct contact with the ductal lumen and always had microvilli at their contact points. Many of the NECs had accompanying nerves, some of which terminated on the surface in contact with the NEC. CONCLUSIONS: Three-dimensional ultrastructural analysis of mouse prostatic NECs was performed. These cells can be classified into three types based on shape. Novel findings include the presence of microvilli at their points of contact with the ductal lumen and the presence of accompanying nerves.

Insights on Platelet-Derived Growth Factor Receptor α-Positive Interstitial Cells in the Male Reproductive Tract.

Male infertility is a significant factor in approximately half of all infertility cases and is marked by a decreased sperm count and motility. A decreased sperm count is caused by not only a decreased production of sperm but also decreased numbers successfully passing through the male reproductive tract. Smooth muscle movement may play an important role in sperm transport in the male reproductive tract; thus, understanding the mechanism of this movement is necessary to elucidate the cause of sperm transport disorder. Recent studies have highlighted the presence of platelet-derived growth factor receptor α (PDGFRα)-positive interstitial cells (PICs) in various smooth muscle organs. Although research is ongoing, PICs in the male reproductive tract may be involved in the regulation of smooth muscle movement, as they are in other smooth muscle organs. This review summarizes the findings to date on PICs in male reproductive organs. Further exploration of the structural, functional, and molecular characteristics of PICs could provide valuable insights into the pathogenesis of male infertility and potentially lead to new therapeutic approaches.

Soluble Immune Checkpoint Molecules as Predictors of Efficacy in Immuno-Oncology Combination Therapy in Advanced Renal Cell Carcinoma.

Immuno-oncology (IO) combination therapy is the first-line treatment for advanced renal cell carcinoma (RCC). However, biomarkers for predicting the response to IO combination therapy are lacking. Here, we investigated the association between the expression of soluble immune checkpoint molecules and the therapeutic efficacy of IO combination therapy in advanced RCC. The expression of soluble programmed cell death-1 (sPD-1), soluble programmed cell death ligand-1 (sPD-L1), soluble PD-L2 (sPD-L2), and lymphocyte activation gene-3 (sLAG-3) was assessed in plasma samples from 42 patients with advanced RCC who received first-line IO combination therapy. All IMDC risk classifications were represented among the patients, including 14.3, 57.1, and 28.6% with favorable, intermediate, and poor risk, respectively. Univariate analysis revealed that prior nephrectomy, sPD-L2 levels, and sLAG-3 levels were significant factors affecting progression-free survival (PFS), whereas multivariate analyses suggested that sPD-L2 and sLAG-3 levels were independent prognostic factors for PFS. In a univariate analysis of the overall survival, prior nephrectomy and sPD-L2 levels were significant factors; no significant differences were observed in the multivariate analysis. No significant correlation was observed between the sPD-L2 and sLAG-3 levels and PD-L2 and LAG-3 expression via immunohistochemistry. In conclusion, sPD-L2 and sLAG-3 expression may serve as a potential biomarker for predicting IO combination therapy efficacy.

Sheet-like interstitial cells connect epithelial and smooth muscle cells in the mouse prostate.

Epithelial-stromal relationship in the prostate gland is crucial for maintaining homeostasis, including functional differentiation, proliferation, and quiescence. Pathological stromal changes are believed to cause benign prostatic hyperplasia (BPH). The prostate stromal tissue is known to have several subtypes of interstitial cells that connect the epithelium and smooth muscle. However, the characteristics of their morphology and connection patterns are not fully understood. Therefore, we aimed to investigated the three-dimensional morphology and intercellular interactions of interstitial cells in the prostate ventral lobe of mature wild-type mice using immunohistochemistry and focused ion beam-scanning electron microscopy tomography (FIB-SEM tomography). The prostate interstitial cells exhibited immunohistochemical subtypes, including PDGFRα single-positive, CD34 single-positive, and CD34 and PDGFRα double-positive. PDGFRα single-positive cells were observed as elongated cells just below the epithelium, CD34 single-positive cells were observed as polygonal cells in the area away from the epithelium, and double-positive cells were observed as elongated cells situated slightly deeper than PDGFRα single-positive cells. Furthermore, connexin43-immunoreactive puncta were observed on interstitial cells just beneath the epithelium, suggestive of possible electrical connections among the PDGFRα single-positive interstitial cells. Three-dimensional structural analysis using FIB-SEM tomography revealed sheet-like multilayered interstitial cells that appear to separate the glandular terminal from the deeper interstitial tissue, which includes smooth muscle and capillaries. Further, epithelial cells might be indirectly connected to the smooth muscle and nerve fibers via these sheet-like multilayered interstitial cellular networks. These findings suggest that the cellular network that separates the glandular terminals from the deep interstitial tissue functionally bridges the epithelium and smooth muscle, possibly playing a pivotal role in prostate tissue homeostasis through the epithelial-smooth muscle or epithelial-stromal relationships.

Three-Dimensional Ultrastructural and Volume Analysis of the Redundant Nuclear Envelope of Developing and Matured Sperm in Mice.

The ultrastructure of the nuclear envelope (NE) and redundant NE (RNE) of the spermatozoon cannot be observed in detail using conventional electron microscopy. Thus, this study aimed to employ transmission electron microscopy (TEM) and focused ion beam/scanning electron microscopy (FIB/SEM) tomography to fill this research gap. Male mice aged 13 weeks were deeply anesthetized, and the testes and vas deferens were extracted and processed for electron microscopy. In round spermatids, the acrosomal vesicle compressed the nucleus, and the acrosomal center was depressed. The nucleoli concentrated on the contralateral side of the acrosome formation site. In mature spermatozoa, the RNE accumulated in the neck with the residual bodies. The NE pores exhibited a hexagonal pattern. The body surface area and volume of the nuclei of spermatids and spermatozoa in each maturation phase were analyzed using FIB/SEM tomography. The body surface area and volume of the nuclei decreased during spermatid maturation into spermatozoa. The RNE converged at the sperm neck and possessed a honeycomb structure. The method used revealed that the nuclei of spermatids gradually condense as they mature into spermatozoa. This method may be used to analyze small tissues, such as RNE, and detect morphological abnormalities in microtissues, such as spermatozoa.

Three-Dimensional Ultrastructural Analysis of the Head-Most Mitochondrial Roots of Mice Spermatozoa Using Focused Ion Beam/Scanning Electron Microscopy (FIB/SEM) Tomography.

This study aimed to clarify the three-dimensional ultrastructure of head-side mice spermatozoa mitochondria. Six 13-week-old male C57BL/6 mice were deeply anesthetized, perfused with 2% paraformaldehyde, and placed in 2.5% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.3) for electron microscopy. After perfusion, the vas deferens was removed, and the specimens were cut into small cubes and subjected to postfixation and en bloc staining. Three-dimensional ultrastructural analysis was performed on five mitochondria on the spermatozoa head using conventional transmission electron microscopy (TEM) and focused ion beam/scanning electron microscopy (FIB/SEM) tomography. Conventional TEM analysis showed that head-side mitochondria were not spiral in morphology but clearly horizontal to the sperm axis. However, this was difficult to evaluate further using conventional TEM. In the FIB/SEM analysis, the first and second head-most mitochondria were flat and straight, with no helix, and shaped as an attachment plug with two electrodes, and their tail side contacted the third mitochondrion. The third mitochondrion was shorter than the fourth and fifth and had a semicircular arching structure. The fourth and fifth mitochondria were spiral-shaped and intertwined. The redundant nuclear envelope encircled the head-most mitochondria. This ultrastructural analysis clarified that the head-most mitochondria have a unique morphology.

Sarcopenia and the Therapeutic Effects of Androgen Receptor-axis-targeted Therapies in Patients With Castration-resistant Prostate Cancer.

BACKGROUND/AIM: Sarcopenia is a syndrome characterized by the progressive and generalized loss of skeletal muscle mass and has been reported to be a poor prognostic factor for taxane-treated castration-resistant prostate cancer (CRPC). However, whether sarcopenia affects androgen receptor axis-targeted therapies (ARATs) remains unknown. In the present study, we investigated the association between sarcopenia in CRPC and treatment outcomes of ARATs. PATIENTS AND METHODS: From January 2015 to September 2022, 127 patients who received ARATs as 1st-line treatment for CRPC at our two hospitals were included in the study. We retrospectively evaluated sarcopenia using computed tomography images and investigated whether sarcopenia affects the progression-free survival (PFS) and overall survival (OS) of patients with CRPC treated with ARATs. RESULTS: Out of 127 patients, 99 were diagnosed with sarcopenia. The PFS of the sarcopenic group administered ARATs was significantly better than that of the non-sarcopenic group. Furthermore, in the multivariate analysis of PFS, sarcopenia was an independent favourable prognostic factor. However, there was no significant difference in the OS between the sarcopenic and non-sarcopenia groups. CONCLUSION: ARATs could more effectively treat patients with CRPC and sarcopenia than patients with CRPC without sarcopenia. Sarcopenia may positively influence the therapeutic effects of ARATs.

Prognostic Value of Absolute Lymphocyte Count in Patients with Advanced Renal Cell Carcinoma Treated with Nivolumab Plus Ipilimumab.

Nivolumab and ipilimumab (NIVO + IPI) is standard therapy for patients with advanced renal cell carcinoma (RCC). Absolute lymphocyte count (ALC) is a valuable prognostic factor in patients with various cancers treated with immune checkpoint inhibitors. Herein, we determined the prognostic value of pretreatment ALC in advanced RCC patients treated with NIVO + IPI as first-line therapy. Data from 46 advanced RCC patients treated with NIVO + IPI between September 2018 and August 2022 were retrospectively reviewed and analyzed. Median progression-free survival (PFS) and overall survival (OS) were significantly shorter in patients with low than high ALC (PFS: p = 0.0095; OS: p = 0.0182). Multivariate analysis suggested that prior nephrectomy [hazard ratio (HR) = 3.854, 95% confidence interval (CI) = 1.433-10.359, p = 0.0075] and pretreatment ALC (HR = 2.513, 95% CI = 1.119-5.648, p = 0.0257) were independent factors for PFS. Our new prognostic ALNx model based on ALC and prior nephrectomy suggested that the poor-risk group was a predictor of significantly worse PFS (p < 0.0001) and OS (p = 0.0016). Collectively, the developed ALNx model may be a novel predictor of response in advanced RCC patients treated with NIVO + IPI.

Improved Survival of Real-world Japanese Patients With Advanced Renal Cell Carcinoma Treated With Immuno-oncology Combination Therapy.

BACKGROUND/AIM: Immuno-oncology (IO) combination therapy has become the standard of treatment for advanced renal cell carcinoma (RCC). In this retrospective study, we compared the efficacy of first-line molecular targeted therapy (MTT), administered as monotherapy, and IO combination therapy using real-world data of Japanese patients. PATIENTS AND METHODS: The clinical information of 202 patients with RCC who received MTT (n=144) or IO combination therapy (n=58) at the Kurume University Hospital from May 2008 to May 2022 was collected and retrospectively analyzed. The Cox proportional hazards model was used for univariate and multivariate analyses, with hazard ratios (HRs) and 95% confidence intervals (CIs) calculated. RESULTS: The patients treated with IO combination therapy had a prolonged progression-free survival (PFS) compared with those treated with MTT (p=0.0038). IO combination therapy was significantly associated with a better PFS in patients with intermediate (p=0.0072) and poor risk (p=0.0411) but not in those with favorable risk (p=0.5434). Furthermore, overall survival with IO combination therapy was significantly better in patients at poor risk (p=0.0335). Multivariate analyses suggested that prior nephrectomy (HR=1.501, 95%CI=1.048-2.150, p=0.0268) and first-line therapy (HR=1.962, 95%CI=1.288-2.986, p=0.0017) were independent prognostic factors for PFS. CONCLUSION: IO combination therapy significantly improved the PFS of patients with advanced RCC, especially those with intermediate- and poor-risk disease. Further investigations focusing on the improvement of survival are warranted.

Three-dimensional analysis of interstitial cells in the lamina propria of the murine vas deferens by confocal laser scanning microscopy and FIB/SEM.

The present study aimed to explore the three-dimensional (3D) ultrastructure of interstitial cells (ICs) within the lamina propria of the murine vas deferens and the spatial relationships between epithelial cells and surrounding cells. Focused ion beam scanning electron microscopy and confocal laser scanning microscopy were performed. ICs within the lamina propria had a flat, sheet-like structure of cytoplasm with multiple cellular processes. In addition, two types of 3D structures that comprised cell processes of flat, sheet-like ICs were observed: one was an accordion fold-like structure and the other was a rod-shaped structure. ICs were located parallel to the epithelium and were connected to each other via gap junctions or adherens junctions. Moreover, multiple sphere-shaped extracellular vesicle-like structures were frequently observed around the ICs. The ICs formed a complex 3D network comprising sheet-like cytoplasm and multiple cell processes with different 3D structures. From this morphological study, we noted that ICs within the lamina propria of murine vas deferens may be involved in signal transmission between the epithelium and smooth muscle cells by physical interaction and by exchanging extracellular vesicles.

Immediate Prostate-specific Antigen Decline After Enzalutamide Following Abiraterone Predicts Survival in Castration-resistant Disease.

BACKGROUND/AIM: Although the sequential use of abiraterone and enzalutamide is not recommended because of possible cross-resistance, many patients with metastatic castration-resistant prostate cancer (mCRPC) are receiving sequential abiraterone and enzalutamide in the real world, and a subset of patients can benefit from sequential therapy with these drugs. This study aimed to identify patients who could benefit from the sequential use of enzalutamide after abiraterone use. PATIENTS AND METHODS: We included 70 patients with mCRPC who received enzalutamide sequentially following abiraterone treatment. Decline in the prostatespecific antigen (PSA) levels at 4 weeks after enzalutamide initiation and the association between decline in PSA levels and survival were analyzed. RESULTS: Sixteen men (22.9%) achieved a decline of >50% in PSA levels after 4 weeks of enzalutamide administration. Overall survival (OS) after enzalutamide among men with >50% decline at 4 weeks was significantly better than that among men with a PSA decline <50% (not reached vs. 34 months, p=0.008). OS after first-line abiraterone treatment for men with PSA decline >50% and <50% was not reached and 46 months, respectively (p=0.007). A PSA decline of >50% at 4 weeks of enzalutamide administration was an independent predictor of longer OS. CONCLUSION: A PSA decline of >50% at 4 weeks after the start of sequential enzalutamide treatment following abiraterone treatment predicted long-term survival in patients with mCRPC. Early PSA decline can identify patients who benefit from second-line enzalutamide after abiraterone treatment and can be useful as a decision-making tool regarding treatment.

Survival outcomes of non-definitive therapy for muscle-invasive bladder cancer.

To analyze the risks and survival outcomes of non-definitive therapy (nDT) for muscle-invasive bladder cancer (MIBC), which may provide useful information for future treatment selection, the present study analyzed 124 patients who were diagnosed with MIBC (cT2-4aN1-2M0) and treated at Kurume University Hospital (Kurume, Japan) with definitive therapy (DT; including radical cystectomy and trimodal therapy) or nDT [transurethral resection of bladder tumor (TURBT) monotherapy or TURBT plus chemotherapy]. Differences in survival outcomes between the two groups were estimated using the Kaplan-Meier method and analyzed using the log-rank test. Cox proportional hazards regression models were used for multivariate analysis of each survival outcome. Of the 124 patients, 45% were treated with nDT, and among these, 50% were treated with TURBT monotherapy and 50% were treated with TURBT plus chemotherapy. Of the patients who chose definitive treatment, 69% were treated with radical cystectomy. The median age in the nDT group was 77 years, which was significantly higher than that in the DT group. Additionally, the proportion of patients with poor performance status, high Charlson comorbidity index and high neutrophil-lymphocyte ratio values was significantly higher in the nDT group. nDT was associated with significantly reduced overall survival, cancer-specific survival and progression-free survival rates, and was a poor prognostic factor for all survival outcomes compared with DT. In conclusion, nDT was associated with a high cancer-related mortality risk. The most appropriate treatment method should be discussed with the patients after providing them with sufficient information on the risks and benefits of each treatment method.

Immune-related adverse events are clinical biomarkers to predict favorable outcomes in advanced renal cell carcinoma treated with nivolumab plus ipilimumab.

BACKGROUND: Immune checkpoint inhibitors cause various immune-related adverse events. The present study examined the association between the incidence of immune-related adverse events and survival outcomes in patients treated with nivolumab plus ipilimumab for patients with advanced renal cell carcinoma. In addition, we compared the effect of adverse event profiles on survival for patients receiving nivolumab plus ipilimumab. METHODS: A total of 35 patients with advanced renal cell carcinoma who were treated with nivolumab plus ipilimumab from August 2018 to August 2021 were retrospectively reviewed and analyzed. Cox proportional hazards models were used for univariate and multivariate analyses, and hazard ratio and 95% confidence intervals were calculated. RESULTS: Of the 35 patients, 22 (62.9%) experienced immune-related adverse events. The median progression-free survival (P = 0.0012) and overall survival (P = 0.0147) were significantly longer in patients with immune-related adverse events than in those without immune-related adverse events. Multivariate analysis showed that the incidence of immune-related adverse events was an independent factor for progression-free survival (hazard ratio = 4.940, 95% confidence interval: 1.558-15.664, P = 0.0067). Skin reaction was a positive predictive immune-related adverse events for progression-free survival (hazard ratio = 9.322, 95% confidence interval: 1.954-44.475, P = 0.0051). CONCLUSION: Patients with advanced renal cell carcinoma with immune-related adverse events had superior clinical outcomes of nivolumab plus ipilimumab treatment than those without immune-related adverse events. Skin immune-related adverse events may be effective biomarkers in patients with advanced renal cell carcinoma treated with nivolumab plus ipilimumab.

Three-dimensional ultrastructure and histomorphology of mouse circumvallate papillary taste buds before and after birth using focused ion beam-scanning electron microscope tomography.

Early taste buds are formed from placode cells. Placode cells differentiate into Type I-Ⅲ cells at birth; however, the ultrastructure of these first taste cells remain elusive. Here, we used focused ion beam-scanning electron microscopy (FIB-SEM) to analyze taste buds on the dorsal surface of the circumvallate papilla on embryonic day (E) 18.5 and postnatal day (P) 1.5. The taste buds on E18.5 existed as a mass of immature cells. One of the immature cells extended the cell process to the surface of the epithelium from the taste bud mass. Cytoplasm of this cell contained many mitochondria and vesicles in the apical region. The taste buds at P1.5 had small taste pores and had an onion-shaped structure. Most of the cells in the taste buds extended toward the taste pores. Some of the cells in the taste buds were Type II-like cells with glycogen in their cytoplasm. In this study, it was shown in three dimensions that immature cells extend to the surface of epithelium before the formation of the taste pore. Subsequently, the formation of taste pores and maturation of taste buds progress simultaneously.

Three-Dimensional Analysis of Interstitial Cells in the Smooth Muscle Layer of Murine Vas Deferens Using Confocal Laser Scanning Microscopy and FIB/SEM.

The smooth muscle contraction of the vas deferens has the important function of transporting sperm. Interstitial cells (ICs) play a critical role in the pacing and modulation of various smooth muscle organs by interactions with nerves and smooth muscle. Elucidating the three-dimensional (3D) architecture of ICs is important for understanding their spatial relationship on the mesoscale between ICs, smooth muscle cells (SMCs), and nerves. In this study, the 3D ultrastructure of ICs in the smooth muscle layer of murine vas deferens and the spatial relationships between ICs, nerves, and smooth muscles were observed using confocal laser scanning microscopy and focused ion beam/scanning electron microscopy. ICs have sheet-like structures as demonstrated by 3D observation using modern analytical techniques. Sheet-like ICs have two types of 3D structures, one flattened and the other curled. Multiple extracellular vesicle (EV)-like structures were frequently observed in ICs. Various spatial relations were observed in areas between ICs, nerves, and SMCs, which formed a complex 3D network with each other. These results suggest that ICs in the smooth muscle layer of murine vas deferens may have two subtypes with different sheet-like structures and may be involved in neuromuscular signal transmission via physical interaction and EVs.

Absolute lymphocyte count is an independent predictor of survival in patients with metastatic renal cell carcinoma treated with nivolumab.

OBJECTIVE: Programmed cell death-1 antibody therapy has demonstrated improved progression-free survival and overall survival in patients with metastatic renal cell carcinoma. However, there are limited studies on biomarkers that can predict the efficacy of immune checkpoint inhibitors. We examined the influence of peripheral inflammatory biomarkers on the clinical outcomes of patients with metastatic renal cell carcinoma treated with nivolumab. METHODS: Data of 38 patients with metastatic renal cell carcinoma, who were treated with nivolumab monotherapy after receiving at least one molecular targeted therapy from November 2016 to February 2021, were retrospectively reviewed and analyzed. RESULTS: Median progression-free survival and overall survival were significantly shorter in patients with low absolute lymphocyte count (<1300/µl) versus those with high absolute lymphocyte count (progression-free survival: P = 0.0102; overall survival: P = 0.0026). Median overall survival was shorter in patients with high neutrophil-lymphocyte ratio (≥3.0) versus those with low neutrophil-lymphocyte ratio (P = 0.0344). Multivariate analysis showed that absolute lymphocyte count was an independent factor for progression-free survival (hazard ratio = 2.332, 95% confidence interval = 1.012-5.375, P = 0.0468) and overall survival (hazard ratio = 4.153, 95% confidence interval = 1.108-15.570, P = 0.0347). Increased absolute lymphocyte count, 1 month after nivolumab initiation, was a positive predictive factor for progression-free survival (hazard ratio = 0.419, 95% confidence interval = 0.189-0.926, P = 0.0317) and overall survival (hazard ratio = 0.285, 95% confidence interval = 0.091-0.890, P = 0.0308). CONCLUSION: Our study indicates that peripheral absolute lymphocyte count, before nivolumab initiation, is a predictor of poor response in metastatic renal cell carcinoma. Additionally, increased absolute lymphocyte count, 1 month post-nivolumab initiation, can be a predictor of the effects of nivolumab.

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.

Morphological analysis of interstitial cells in murine epididymis using light microscopy and transmission electron microscopy.

Smooth muscle contraction of the epididymis plays an important role in sperm transport. Although PDGFRα-positive interstitial cells (PDGFRα (+) ICs) are thought to be involved in controlling smooth muscle movement via intercellular signaling, they have not yet been reported to date in the epididymis. Therefore, we aimed to investigate the morphological characteristics of PDGFRα (+) ICs in the interstitial space of the murine epididymis. Immunohistochemistry showed that PDGFRα (+) ICs co-labeled with CD34 (PDGFRα (+) CD34 (+) ICs were distributed in the interstitial space of the murine epididymis from the initial segment (IS) to the cauda of the epididymis. PDGFRα (+) ICs that were not co-labeled with CD34 (PDGFRα (+) CD34 (-) ICs) were observed just beneath the epithelium from the corpus to the cauda but not in the IS. Both types of PDGFRα (+) ICs were in close proximity to each other as well as the surrounding nerves and macrophages. In addition, PDGFRα (+) CD34 (-) ICs beneath the epithelium were also in close proximity to the basal cells. Using transmission electron microscopy, we identified ICs that possessed elongated and woven cellular processes and were in close proximity to each other, surrounding the cells in the interstitial space. In the murine epididymis, it is suggested that there are two subtypes of ICs that show different distribution patterns depending on the segment, which may reflect segmental differences in mechanisms of sperm transport, forming a cellular network by physical interactions in the murine epididymis.

Identification of PDGFRα-positive interstitial cells in the distal segment of the murine vas deferens.

Platelet-derived growth factor receptor-α (PDGFRα)-positive interstitial cells (ICs) are widely distributed in various organs and may be involved in the motility of various tubular organs. We, for the first time, aimed to investigate the distribution, immunohistochemical characteristics, and ultrastructure of PDGFRα-positive ICs in murine vas deferens, using confocal laser scanning microscopy, transmission electron microscopy (TEM), and immuno-electron microscopy (immuno-EM). For immunofluorescence, we used antibodies against PDGFRα and other markers of ICs. PDGFRα-positive ICs were distributed widely in the lamina propria, smooth muscles, and serosal layers. Although most PDGFRα-positive ICs labeled CD34, they did not label CD34 in the subepithelial layers. Additionally, PDGFRα-positive ICs were in close proximity to each other, as also to the surrounding cells. TEM and immuno-EM findings revealed that PDGFRα-positive ICs established close physical interactions with adjacent ICs. Extracellular vesicles were also detected around the PDGFRα-positive ICs. Our morphological findings suggest that PDGFRα-positive ICs may have several subpopulations, which can play an important role in intercellular signaling via direct contact with the IC network and the extracellular vesicles in the murine vas deferens. Further investigation on PDGFRα-positive ICs in the vas deferens may lead to understanding the vas deferens mortility.

Ectopic subcutaneous transplantation of fetal rat urogenital sinus and seminal vesicle promotes the organ growth and formation.

PURPOSE: The fate of subcutaneously transplanted urogenital sinus (UGS) and seminal vesicle (SV) was investigated in the present study. MATERIALS AND METHODS: Fetal UGS and SV extracted from 20-embryonic-day-old male normal and GFP transgenic rats were subcutaneously transplanted into 7-week-old male immunologically inhibited rats. The transplants were then examined at 2, 4, 8, and 16 weeks after transplantation. We analyzed the survival ratio, weight, and histopathology as well as the immunohistochemical characteristics of the transplanted tissues. For control experiments, 2-, 4-, 8-, and 16-week-old normal male rats were used. RESULTS: Almost all of the transplanted tissues survived under the skin, and the tissue weights increased over time after transplantation. The histopathological characteristics and immunohistochemical staining pattern with certain antibodies of the transplanted tissues were similar to those of normal adult rat prostate and seminal vesicle. The transplanted GFP transgenic tissues demonstrated spontaneous growth and organ formation under the skin, showing distribution and movement of transplanted cells and tissues. CONCLUSION: Subcutaneously transplanted fetal UGS and SV were able to develop into mature adult organs.