Effect of prostate volume on f/tPSA value: A cross-sectional study.

Previous studies have suggested that there is a positive correlation between prostate-specific antigen (PSA) levels and prostate volume (PV). A better understanding of the possible influence of PV on a ratio of free to total PSA (f/tPSA) may improve the diagnostic value of the prostate disease. The study group consisted of 342 men with lower urinary tract symptoms (LUTS). All patients underwent urinary tract ultrasonography and had tests carried out on PSA, serum glucose, total cholesterol, triglyceride, HDL, LDL and blood pressure. Univariate and multivariate analyses were used to assess the associations between prostate volume and f/tPSA value. We found no obvious relationship between prostate volume and f/tPSA value when PSA >10 ng/ml but did observe a positive correlation when 4 ng/ml < PSA ï¼œ 10 ng/ml (hazard ratio [HR]: 0.0012; 95% confidence interval [CI]: 0.0009-0.0248). With increasing prostate volume, multivariate analysis showed an obvious increase in f/tPSA value (HR: 0.0011; 95% CI: 0.0007-0.0015) (p ≤ .0001). We confirmed that prostate volume could affect the f/tPSA levels in serum. There was an obvious positive correlation between prostate volume and f/tPSA level when PSA levels were between 4 and 10ng/dl. There was no significant correlation between prostate volume and f/tPSA value when PSA >10 ng/ml.

Effect of high-density lipoprotein on penile erection: A cross-sectional study.

Previous studies have shown that elevated levels of high-density lipoprotein (HDL) could inhibit penile erection, but the relationship between HDL and the erection of the penile tip or base has not been extensively researched. We investigated the effects of HDL on erection of the penile tip and base through a cross-sectional study of 113 patients with erectile dysfunction, using a cut-off score of ≤21 on the International Index of Erectile Function-5. The following patient data were collected: nocturnal penile tumescence; blood pressure; platelet count; platelet distribution width; mean platelet volume; plateletcrit; and levels of serum glucose, total cholesterol, triglyceride, HDL, and low-density lipoprotein. Univariate and multivariate analyses were used to assess the association between HDL levels and the erection of the penile tip and base. We confirmed that HDL had a beneficial effect on penile erectile function. We also found that when the HDL level exceeded the normal range, the change in HDL had a significant effect on the penile base. In addition, our study did not find any relationship between platelet parameters and erection of the penile tip or penile base.

Verbascoside attenuates experimental varicocele-induced damage to testes and sperm levels through up-regulation of the hypothalamus-pituitary-gonadal (HPG) axis.

CONTEXT: Verbascoside (VB), which is found in many medicinal plant families, exhibits biological activities in various diseases. However, its effects on varicocele (VCL)-induced damage remain unknown. OBJECTIVE: To investigate the effects and mechanism of VB on experimental rats with varicocele (VCL)-induced damage. MATERIALS AND METHODS: Sixty sexually mature male Sprague-Dawley (SD) rats were divided into six groups (n = 10): control, control-sham, VCL-vehicle (normal saline), and VCL + VB groups (50, 100, and 200 mg/kg/day, intraperitoneally). After 4 weeks of VB treatment, all animals were sacrificed, and the body and testicular weight, sperm quality parameters, histopathology, antioxidant status, and hormone levels were tested. The levels of gonadotropin-releasing hormone (GnRH) and gonadotropin-inhibitory hormone in the hypothalamus were detected by western blot. RESULTS: Compared with the VCL-vehicle group (41.14%), administration of VB significantly increased the sperm viability (59.29, 65.45, 84.93%). VB groups showed higher Johnson's score (3.57 ± 0.15, 4.71 ± 0.26, 7.93 ± 0.37) than VCL-vehicle group (2.72 ± 0.24). Antioxidant status and hormone levels alterations were also observed. Meanwhile, the mean number of apoptotic tubules (8.15 ± 0.96, 6.61 ± 1.05, 2.17 ± 0.08) and apoptotic index showed a marked decrease. Compared with the VCL-vehicle group (0.21 ± 0.09), the VB groups (0.36 ± 0.07, 0.42 ± 0.06, 0.88 ± 0.10) showed considerable increases in GnRH. DISCUSSION AND CONCLUSIONS: VB has protective effects on reproductive organs and VB may be therapeutically useful in the treatment of varicocele through up-regulation of the HPG axis.

Relationship between penile erection and the ratio of estradiol to testosterone: A retrospective study.

Previous studies have found that the ratio of estradiol to testosterone (E2/T ratio) has a negative effect on sexual function, but the relationship between the E2/T ratio and erection of the penis is not clarified. We conducted a retrospective study of 183 patients with erectile dysfunction and 52 healthy men to investigate the relationship between penis base erection and tip erection. All participants underwent nocturnal penile tumescence tests and medical history checks and had relevant biochemical and endocrine indicators measured. The ratio of estradiol to testosterone was calculated. The relationship between E2/T ratio and erectile time of penile tip and penile base was determined by univariate analysis, multivariate analysis and stratification analysis. After adjusting for mixed factors, the results showed that the E2/T ratio had a more significant negative effect on the base of the penis compared with the tip of the penis (Hazard ratio: -4.34 95% CI: -6.52, -2.16 p = .0001). Moreover, when the effective erection time was ≥10 min, the negative effect of E2/T on penile root erection was more obvious (HR ratio: -4.46 95% CI: -6.50, -2.43 p < .0001). In summary, our study demonstrated a negative relationship between E2/T ratio and penile erection, particularly at the root of the penis.

Microenvironment derived from metanephros transplantation inhibits the progression of acute kidney injury in glycerol-induced rat models.

Background: Embryonic metanephros is the mammalian renal anlagen, which is considered as a potential source for the regeneration of functional whole kidneys. Some studies reported that metanephros implanted into unilateral nephrectomized animals can develop into kidney tissue. However, kidneys are nephrotoxic in renal failure patients, and whether metanephros can grow in nephrotoxic has not been reported. This study aims to investigate the growth of metanephros in acute nephrotoxic environment and analyze the therapeutic effect of metanephros microenvironment on acute kidney injury (AKI).Methods: AKI was induced in 200 g Wistar rats by giving intramuscular injections of 50% glycerol (10 mL/kg) in their hind limbs. 45 rats were divided randomly into three groups (control, glycerin, and metanephros). Metanephros group was transplanted two metanephroi (embryonic day 15) into the renal capsule of AKI rats. Glycerin group was AKI rats without transplantation. Control group was untreated.Results: Mature glomeruli and tubules were detected in the grafts in metanephros group, which means that metanephroi can grow into tissues with mature kidney structure under acute nephrotoxic. Then, we assessed the renal function of host rats and found that there were fewer tubular necrosis in metanephros group than glycerin group, and the serum creatinine and urea nitrogen were significantly lower in metanephros group than glycerin group.Conclusion: These results suggested that embryonic metanephroi can grow into tissues with mature kidney structure under acute nephrotoxic, and the graft microenvironment was effective in inhibiting the progression of AKI, which provides a new approach for the treatment of acute renal injury.

Terpenoids with vasorelaxant effects from the Chinese liverwort Scapania carinthiaca.

Four new diterpenoids scapanacins A-D (1-4) including one kaurane and three clerodane derivatives, along with eleven known compounds (9-15), were isolated from the Chinese liverwort Scapania carinthiaca J.B. Jack ex Lindb. Their structures were determined based on extensive spectroscopic analyses, and electronic circular dichroism (ECD) calculations. Vasorelaxant activity assays of the clerodane-type diterpenoids 2, and 4-8 revealed that they relaxed 3rd-order rat mesenteric arterioles pre-contracted with norepinephrine (NE). Further assays with scapanacin D (4) confirmed that the vasodilatation was mediated through inhibition of Ca2+ influx via voltage-dependent Ca2+ channels (VDCs), and this Ca2+ channel blocking effect was also confirmed by inhibiting the extracellular Ca2+ influx in MOVAS cells. Besides, very little decrease of the relaxant activity caused by 4 on endothelium-denuded mesenteric arterioles with NE also suggested the vasodilatation was mainly produced by inhibiting Ca2+-induced contraction of smooth muscle. In addition, cytotoxicity testing showed that compounds 1 and 9 with α,ß-unsaturated ketone exhibited inhibitory activities against a small panel of human cancer cell lines.

Impact of O6-methylguanine-DNA methyltransferase expression on the drug resistance of clear cell renal cell carcinoma.

OBJECTIVE: The deoxyribonucleic acid-repair protein O(6)-methylguanine-deoxyribonucleic acid methyltransferase is a major determinant of resistance of cells to various alkylating drugs. Its expression profile is different in different cancer types. Here, we studied the expression and function of O(6)-methylguanine-deoxyribonucleic acid methyltransferase in clear cell renal cell carcinoma. METHODS: The expression of O(6)-methylguanine-deoxyribonucleic acid methyltransferase was evaluated in clear cell renal cell carcinoma tissues and cell lines by quantitative real-time polymerase chain reaction and immunohistochemistry. The relationship between O(6)-methylguanine-deoxyribonucleic acid methyltransferase expression and clinicopathological characteristics was analyzed. To further investigate the function of O(6)-methylguanine-deoxyribonucleic acid methyltransferase in clear cell renal cell carcinoma resistance to alkylating agents, siRNA targeting O(6)-methylguanine-deoxyribonucleic acid methyltransferase were used to silence the O(6)-methylguanine-deoxyribonucleic acid methyltransferase expression. RESULTS: We found that O(6)-methylguanine-deoxyribonucleic acid methyltransferase is over-expressed in clear cell renal cell carcinoma tissues and cell lines. O(6)-methylguanine-deoxyribonucleic acid methyltransferase expression is related with tumor progression in clear cell renal cell carcinoma patients. Up-regulation of O(6)-methylguanine-deoxyribonucleic acid methyltransferase plays a critical role in primary resistance to alkylating agents. CONCLUSIONS: The overexpression of O(6)-methylguanine-deoxyribonucleic acid methyltransferase contributes to resistance of clear cell renal cell carcinoma to standard chemotherapy. Our results have significance for understanding a new pathway of the development of drug resistance of clear cell renal cell carcinoma.

The vascular and neurogenic factors associated with erectile dysfunction in patients after pelvic fractures.

Erectile dysfunction (ED) is a common complication of pelvic fractures. To identify the vascular and neurogenic factors associated with ED, 120 patients admitted with ED after traumatic pelvic fracture between January 2009 and June 2013 were enrolled in this study. All patients answered the International Index of Erectile Function (IIEF-5) questionnaire. Nocturnal penile tumescence (NPT) testing confirmed the occurrence of ED in 96 (80%) patients on whom penile duplex ultrasound and neurophysiological testing were further performed. Of these ED patients 29 (30%) were demonstrated only with vascular abnormality, 41 (42.7%) were detected only with neural abnormality, 26 (27.1%) revealed mixed abnormalities. Of the 55 patients (29+26) with vascular problems, 7 patients (12.7%) with abnormal arterial response to intracavernous injection of Bimix (15mg papaverine and 1mg phentolamine), 31 (56.4%) with corporal veno-occlusive dysfunction and 17 (30.9%) had both problems. Of the 67 (41+26) patients with abnormal neurophysiological outcomes, 51 (76.1%) with abnormal bulbocavernosus re?ex (BCR), 20 (29.9%) with pathological pudendal nerve evoked potentials (PDEPs) and 25 (37.3%) with abnormal posterior tibial somatosensory nerve evoked potentials (PTSSEPs). Our observation indicated that neurogenic factors are important for the generation of ED in patients with pelvic fracture; venous impotence is more common than arteriogenic ED.

Laminin 411 acts as a potent inducer of umbilical cord mesenchymal stem cell differentiation into insulin-producing cells.

BACKGROUND: Diabetes mellitus (DM) is an incurable metabolic disease constituting a major threat to human health. Insulin-producing cells (IPCs) differentiated from mesenchymal stem cells (MSCs) hold great promise in the treatment of DM. The development of an efficient IPC induction system is a crucial step for the clinical application of IPCs for DM. Laminin 411 is a key component of the basement membrane and is involved in the regulation of cell differentiation; however, little is known about a role of laminin 411 in the regulation of IPC differentiation from human MSCs. METHODS: MSCs were isolated from human umbilical cord (UC-MSCs) and expanded in an in vitro culture system. UC-MSCs were then cultured in the IPC induction and differentiation medium in the presence of laminin 411. Flow cytometry, Quantitative realtime PCR, immunofluorescence staining, ELISA, Western blotting and other techniques were applied to determine IPC generation, insulin expression and related mechanisms. To evaluate potential therapeutic efficacy of IPCs induced from UC-MSCs, a type-1 diabetes (T1DM) rat model was generated using streptozotocin. Blood glucose, insulin levels, and survival of rats were monitored periodically following intravenous injection of the tested cells. RESULTS: Laminin 411 markedly induced the expression of the genes Foxa2 and Sox17, markers for pancreatic precursor cells, efficiently induced IPC differentiation from MSCs, and up-regulated insulin expression at both mRNA and protein levels. Furthermore, the expression of the genes known to govern insulin expression including Pdx1 and Ngn3 was markedly induced by laminin 411, which suggests that Pdx1 and Ngn3 signaling pathways are involved in laminin 411 induced-insulin expression machinery. More importantly, administration of laminin 411-induced IPCs rapidly and significantly down-regulated fasting blood glucose levels, significantly reduced the HbA1c concentration and markedly improved the symptoms and survival of T1DM rats. CONCLUSIONS: Our results demonstrate that laminin 411 acts as a potent differentiation inducer of IPCs from UC-MSCs via the Pdx1 and Ngn3 signaling pathways. Moreover, transfusion of laminin 411 induced-IPCs more efficiently improves symptoms and survival of T1DM rats. These novel finding highlights a potential clinical application of laminin 411 induced-IPCs in the treatment of T1DM, which calls for further studies.

Maturation of Nephrons by Implanting hPSC-derived Kidney Progenitors Under Kidney Capsules of Unilaterally Nephrectomized Mice.

BACKGROUND: Human pluripotent stem cell (hPSC)-derived kidney organoids may contribute to disease modeling and the generation of kidney replacement tissues. However, the realization of such applications requires the induction of hPSCs into functional mature organoids. One of the key questions for this process is whether a specific vascular system exists for nephrogenesis. Our previous study showed that short-term (2 weeks) implantation of hPSC-derived organoids below the kidney capsules of unilaterally nephrectomized and immunodeficient mice resulted in the enlargement of organoids and production of vascular cells, although signs of maturation were lacking. METHODS: Organoids were induced for 15 days in vitro and then grafted below kidney capsules of the same unilaterally nephrectomized immunodeficient mouse model to examine whether medium-term (4 weeks) implantation could improve organoid maturation and vascularization, as evaluated by immunofluorescence and transmission electron microscopy. RESULTS: We demonstrated that after 2-4 weeks of implantation, renal organoids formed host-derived vascularization and matured without any exogenous vascular endothelial growth factor. Glomerular filtration barrier maturation was evidenced by glomerular basement membrane deposition, perforated glomerular endothelial cell development, and apical, basal podocyte polarization. A polarized monolayer epithelium and extensive brush border were also observed for tubular epithelial cells. CONCLUSIONS: Our results indicate that the in vivo microenvironment is important for the maturation of human kidney organoids. Stromal expansion and a reduction of nephron structures were observed following longer-term (12 weeks) implantation, suggesting effects on off-target cells during the induction process. Accordingly, induction efficiency and transplantation models should be improved in the future.

LSR targets YAP to modulate intestinal Paneth cell differentiation.

Lipolysis-stimulated lipoprotein receptor (LSR) is a multi-functional protein that is best known for its roles in assembly of epithelial tricellular tight junctions and hepatic clearance of lipoproteins. Here, we investigated whether LSR contributes to intestinal epithelium homeostasis and pathogenesis of intestinal disease. By using multiple conditional deletion mouse models and ex vivo cultured organoids, we find that LSR elimination in intestinal stem cells results in the disappearance of Paneth cells without affecting the differentiation of other cell lineages. Mechanistic studies reveal that LSR deficiency increases abundance of YAP by modulating its phosphorylation and proteasomal degradation. Using gain- and loss-of-function studies, we show that LSR protects against necrotizing enterocolitis through enhancement of Paneth cell differentiation in small-intestinal epithelium. Thus, this study identifies LSR as an upstream negative regulator of YAP activity, an essential factor for Paneth cell differentiation, and a potential therapeutic target for necrotizing enterocolitis.

A case with type â ¡ vesicouterine fistula.

Here, we report a 41-year-old female with type II VUF after hysteromyomectomy.We report the diagnosis of VUF by imaging method, and provide a feasible treatment for this complication after pelvic surgery.

Comprehensive analysis of FRAS1/FREM family as potential biomarkers and therapeutic targets in renal clear cell carcinoma.

Background: FRAS1 (Fraser syndrome protein 1), together with FREM1 (the Fras1-related extracellular matrix proteins 1) and FREM2, belonging to the FRAS1/FREM extracellular matrix protein family, are considered to play essential roles in renal organogenesis and cancer progression. However, their roles in kidney renal clear cell carcinoma (KIRC) remain to be elucidated. Methods: FRAS1/FREM RNA expression analysis was performed using TCGA/GTEx databases, and valided using GEO databases and real-time PCR. Protein expression was peformed using CPTAC databases. Herein, we employed an array of bioinformatics methods and online databases to explore the potential oncogenic roles of FRAS1/FREM in KIRC. Results: We found that FRAS1, FREM1 and FREM2 genes and proteins expression levels were significantly decreased in KIRC tissues than in normal tissues. Decreased FRAS1/FREM expression levels were significantly associated with advanced clinicopathological parameters (pathological stage, grade and tumor metastasis status). Notably, the patients with decreased FRAS1/FREM2 expression showed a high propensity for metastasis and poor prognosis. FRAS1/FREM were correlated with various immune infiltrating cells, especially CD4+ T cells and its corresponding subsets (Th1, Th2, Tfh and Tregs). FRAS1 and FREM2 had association with DNA methylation and their single CpG methylation levels were associated with prognosis. Moreover, FRAS1/FREM might exert antitumor effects by functioning in key oncogenic signalling pathways and metabolic pathways. Drug sensitivity analysis indicated that high FRAS1 and FREM2 expression can be a reliable predictor of targeted therapeutic drug response, highlighting the potential as anticancer drug targets. Conclusion: Together, our results indicated that FRAS1/FREM family members could be potential therapeutic targets and valuable prognostic biomarkers of KIRC.

Generation of the integration-free induced pluripotent stem cell line (FHUSTCi001-A) from a patient with glomerulopathy with fibronectin deposits harboring FN1 mutation.

Glomerulopathy with fibronectin deposits (GFND) is an autosomal dominant kidney disease exhibiting microscopic hematuria, proteinuria, and hypertension that may lead to end-stage renal failure. In this study, using non-integrative episomal vectors an induced pluripotent stem cell (iPSC) line, FHUSTCi001-A, was derived from peripheral blood mononuclear cells of an 11-year-old boy with GFND carrying a heterozygous c.5602G > A (p.V1868M) mutation in the FN1 gene. The generated iPSC line has a normal karyotype, expresses pluripotency markers, and has the capacity to form all three germ layers in vivo. This iPSC line offers a useful cellular model to study the pathogenesis of GFND disease.

Loss of CLDN5 in podocytes deregulates WIF1 to activate WNT signaling and contributes to kidney disease.

Although mature podocytes lack tight junctions, tight junction integral membrane protein claudin-5 (CLDN5) is predominantly expressed on plasma membranes of podocytes under normal conditions. Using podocyte-specific Cldn5 knockout mice, we identify CLDN5 as a crucial regulator of podocyte function and reveal that Cldn5 deletion exacerbates podocyte injury and proteinuria in a diabetic nephropathy mouse model. Mechanistically, CLDN5 deletion reduces ZO1 expression and induces nuclear translocation of ZONAB, followed by transcriptional downregulation of WNT inhibitory factor-1 (WIF1) expression, which leads to activation of WNT signaling pathway. Podocyte-derived WIF1 also plays paracrine roles in tubular epithelial cells, as evidenced by the finding that animals with podocyte-specific deletion of Cldn5 or Wif1 have worse kidney fibrosis after unilateral ureteral obstruction than littermate controls. Systemic delivery of WIF1 suppresses the progression of diabetic nephropathy and ureteral obstruction-induced renal fibrosis. These findings establish a function for podocyte CLDN5 in restricting WNT signaling in kidney.

Identification of immune-related genes that predict prognosis and risk of bladder cancer: bioinformatics analysis of TCGA database.

BACKGROUND: Bladder cancer (BLCA) is the major tumor of the urinary system, and immune-related genes (IRGs) contribute significantly to its initiation and prognosis. RESULTS: A total of 51 prognostic IRGs significantly associated with overall survival were identified. Functional enrichment analysis revealed that these genes were actively involved in tumor-related functions and pathways. Using multivariate Cox regression analysis, we detected 11 optimal IRGs (ADIPOQ, PPY, NAMPT, TAP1, AHNAK, OLR1, PDGFRA, IL34, MMP9, RAC3, and SH3BP2). We validated the prognostic value of this signature in two validation cohorts: GSE13507 (n = 165) and GSE32894 (n = 224). Furthermore, we performed a western blot and found that the expression of these IRGs matched their mRNA expression in TCGA. Moreover, correlations between risk score and immune-cell infiltration indicated that the prognostic signature reflected infiltration by several types of immune cells. CONCLUSION: We identified and validated an 11-IRG-based risk signature that may be a reliable tool to evaluate the prognosis of BLCA patients and help to devise individualized immunotherapies. METHODS: Bioinformatics analysis was performed using TCGA and ImmPort databases. Cox regression was used to identify prognostic signatures. Two external GEO cohorts and western blotting of samples were performed to validate the IRG signature.

Generation of an iPSC line (HRMSDUi001-A) from an azoospermic patient with 45,XY,der(13;14)(q10;q10) karyotype.

Robertsonian translocation between chromosomes 13 and 14 is reported to be a cause of male infertility. Here, we established an iPSC line (HRMSDUi001-A) from an azoospermic patient with 45,XY,der(13;14)(q10;q10) karyotype. Peripheral blood mononuclear cells were reprogrammed to generate hiPSCs by non-integrating delivery of OCT4, SOX2, KFL4 and MYC. The iPSC line expressed pluripotency markers, could differentiate into cells of three germ layers in vitro, and carried 45,XY,der(13;14)(q10;q10) karyotype.

Age-related reductions in the excitability of phasic dorsal root ganglion neurons innervating the urinary bladder in female rats.

Previous studies have revealed an impairment in bladder sensory transduction in aged animals. To examine the contributions of electrical property changes of bladder primary afferents to this impairment, we compared the electrical properties of dorsal root ganglion (DRG) neurons innervating the bladder among young (3 months), middle-aged (12 months), and old (24 months) female rats. The DRG neurons were labeled using axonal tracing techniques. Whole-cell current-clamp recordings of small and medium-sized neurons were performed to assess their passive and active properties. Two patterns of firing were identified based on responses to super-threshold stimuli (1.5, 2.0, 2.5, and 3.0 × rheobase): tonic neurons fired more action potentials (APs), whereas phasic neurons fired only one AP at the onset of stimulus. Tonic neurons were smaller and had a slower rate of AP rise, longer AP duration, more depolarized voltage threshold, and greater rheobase than phasic neurons. In phasic neurons, there was an age-associated increase in voltage threshold and an increase of rheobase (P < 0.05), suggesting an age-related decrease in excitability. In addition, both middle-aged and old rats had longer AP durations and slower rates of AP rise than young rats (P < 0.05). In tonic neurons, old rats had a greater AP overshoot and greater rate of AP rise, but no age-associated changes were identified in any other electrical properties. Our results suggest that the electrical properties of tonic and phasic bladder afferents are differentially altered with aging. A decrease in excitability may contribute to age-related reductions in bladder sensory function.

Cell Deformation at the Air-Liquid Interface Evokes Intracellular Ca2+ Increase and ATP Release in Cultured Rat Urothelial Cells.

Urothelial cells have been implicated in bladder mechanosensory transduction, and thus, initiation of the micturition reflex. Cell deformation caused by tension forces at an air-liquid interface (ALI) can induce an increase in intracellular Ca2+ concentration ([Ca2+]i) and ATP release in some epithelial cells. In this study, we aimed to examine the cellular mechanisms underlying ALI-induced [Ca2+]i increase in cultured urothelial cells. The ALI was created by stopping the influx of the perfusion but maintaining efflux. The [Ca2+]i increase was measured using the Ca2+ imaging method. The ALI evoked a reversible [Ca2+]i increase and ATP release in urothelial cells, which was almost abolished by GdCl3. The specific antagonist of the transient receptor potential vanilloid (TRPV4) channel (HC0674) and the antagonist of the pannexin 1 channel (10panx) both diminished the [Ca2+]i increase. The blocker of Ca2+-ATPase pumps on the endoplasmic reticulum (thapsigargin), the IP3 receptor antagonist (Xest-C), and the ryanodine receptor antagonist (ryanodine) all attenuated the [Ca2+]i increase. Degrading extracellular ATP with apyrase or blocking ATP receptors (P2X or P2Y) with pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) significantly attenuated the [Ca2+]i increase. Our results suggest that both Ca2+ influx via TRPV4 or pannexin 1 and Ca2+ release from intracellular Ca2+ stores via IP3 or ryanodine receptors contribute to the mechanical responses of urothelial cells. The release of ATP further enhances the [Ca2+]i increase by activating P2X and P2Y receptors via autocrine or paracrine mechanisms.