Bladder Interstitial Cells Research Articles

Single-cell profiling reveals various types of interstitial cells in the bladder.

Clarifying the locations, molecular markers, functions and roles of bladder interstitial cells is crucial for comprehending the pathophysiology of the bladder. This research utilized human, rat and mouse bladder single-cell sequencing, bioinformatics analysis and experimental validation. The main cell types found in human, rat and mouse bladder tissues include epithelial cells, smooth muscle cells, endothelial cells, fibroblasts, myofibroblasts, neurons and various immune cells. Our study identified two significant types of interstitial cells (PTN+ IGFBP6+ PI16 (CD364)+ CD34+ ) and myofibroblasts (STC1+ PLAT+ TNC+ ). These two types of interstitial cells are mainly located in the subepithelial lamina propria, between muscles and between muscle bundles. In the CYP (cyclophosphamide)-induced bladder injury mouse model, the interaction types and signals (MK, MIF, GDF and CXCL) of fibroblasts and myofibroblasts significantly increased compared with the normal group. However, in the aging mouse model, the signals CD34, LAMININ, GALECTIN, MK, SELPLG, ncWNT, HSPG, ICAM and ITGAL-ITGB2 of fibroblasts and myofibroblasts disappeared, but the signals PTN and SEMA3 significantly increased. Our findings identified two crucial types of interstitial cells in bladder tissue, fibroblasts and myofibroblasts, which play a significant role in normal bladder physiology, CYP-induced bladder injury and aging bladder development.

TrkA inhibition alleviates bladder overactivity in cyclophosphamide-induced cystitis by targeting hyperpolarization-activated cyclic nucleotide-gated channels.

To investigate the potential of Tropomyosin receptor kinase A (TrkA) for the treatment of interstitial cystitis/ bladder pain syndrome (IC/BPS). Sixty-four female rats were randomly assigned to the control and cyclophosphamide (CYP) groups. Quantitative reverse transcription polymerase chain reaction was utilized to detect the mRNA level of TrkA. Western blot analysis was used to measure the protein levels of TNF-α, IL-6, and TrkA. Immunostaining was used to detect the expression of TrkA in bladder sections. Contractility studies and urodynamic measurements were utilized to test the spontaneous contractions of detrusor muscle strips and the global bladder activity, respectively. Rat models of chronic cystitis were successfully established. The mRNA and protein levels of TrkA were significantly increased in the bladders of CYP-treated rats. Also, results of immunohistochemical staining and immunofluorescence staining showed that increased TrkA expression in the CYP group was mainly observed in the urothelium layer and bladder interstitial Cajal-like cells (ICC-LCs) but not in the detrusor smooth muscle cells. The specific inhibitor of TrkA, GW441756 (10 μM), significantly suppressed the robust spontaneous contractions of detrusor muscle strips in the CYP group and alleviated the overall bladder overactivity of CYP-treated rats. However, the inhibitory effects of GW441756 (10 μM) on the spontaneous contractions of detrusor muscle strips and the overall bladder activity were eliminated after pretreatments with the specific blocker of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, ZD7288 (50 μM). Our results suggested that increased TrkA expression during chronic cystitis promotes the development of bladder overactivity by targeting the HCN channels.

Studies of ultrastructure, gene expression, and marker analysis reveal that mouse bladder PDGFRA+ interstitial cells are fibroblasts.

Fibroblasts are crucial to normal and abnormal organ and tissue biology, yet we lack basic insights into the fibroblasts that populate the bladder wall. Candidates may include bladder interstitial cells (also referred to as myofibroblasts, telocytes, and interstitial cells of Cajal-like cells), which express the fibroblast-associated marker PDGFRA along with VIM and CD34 but whose form and function remain enigmatic. By applying the latest insights in fibroblast transcriptomics, coupled with studies of gene expression, ultrastructure, and marker analysis, we observe the following: 1) that mouse bladder PDGFRA+ cells exhibit all of the ultrastructural hallmarks of fibroblasts including spindle shape, lack of basement membrane, abundant endoplasmic reticulum and Golgi, and formation of homotypic cell-cell contacts (but not heterotypic ones); 2) that they express multiple canonical fibroblast markers (including Col1a2, CD34, LY6A, and PDGFRA) along with the universal fibroblast genes Col15a1 and Pi16 but they do not express Kit; and 3) that PDGFRA+ fibroblasts include suburothelial ones (which express ACTA2, CAR3, LY6A, MYH10, TNC, VIM, Col1a2, and Col15a1), outer lamina propria ones (which express CD34, LY6A, PI16, VIM, Col1a2, Col15a1, and Pi16), intermuscular ones (which express CD34, VIM, Col1a2, Col15a1, and Pi16), and serosal ones (which express CD34, PI16, VIM, Col1a2, Col15a1, and Pi16). Collectively, our study revealed that the ultrastructure of PDFRA+ interstitial cells combined with their expression of multiple canonical and universal fibroblast-associated gene products indicates that they are fibroblasts. We further propose that there are four regionally distinct populations of fibroblasts in the bladder wall, which likely contribute to bladder function and dysfunction.NEW & NOTEWORTHY We currently lack basic insights into the fibroblasts that populate the bladder wall. By exploring the ultrastructure of mouse bladder connective tissue cells, combined with analyses of their gene and protein expression, our study revealed that PDGRA+ interstitial cells (also referred to as myofibroblasts, telocytes, and interstitial cells of Cajal-like cells) are fibroblasts and that the bladder wall contains multiple, regionally distinct populations of these cells.

Inhibition of U-II/UT signaling ameliorates cystitis-associated bladder hyperactivity by targeting the RhoA/Rho-kinase pathway.

Urotensin II (U-II) and its receptor (UT) are involved in the pathogenesis of various diseases; however, their association with the development of cystitis has not been elucidated. The present study was designed to investigate the functional role of U-II/UT signaling in cyclophosphamide (CYP)-induced cystitis. A total of 60 female rats were randomly divided into the control and CYP-treated groups. Intraperitoneal injection of CYP successfully induced cystitis in rats of the CYP-treated group. The protein and mRNA expression levels of U-II and UT were significantly enhanced in rat bladder tissues of the CYP-treated group. Furthermore, the results of the immunofluorescence staining analysis demonstrated that CYP treatment apparently increased the expression levels of UT in the urothelium layer, detrusor smooth muscle, and bladder interstitial Cajal-like cells. The selective antagonist of UT, SB657510 (10μm), significantly suppressed the CYP-induced increase in the spontaneous contractions of muscle strips and ameliorated the bladder hyperactivity of CYP-treated rats. Moreover, CYP treatment significantly increased the protein expression levels of Ras homolog family member (Rho) A and Rho-associated protein kinase 2 in rat bladder tissues. Following pretreatment with the Rho-kinase inhibitor Y-27632 (10μm), the inhibitory effects of SB657510 (10μm) on the spontaneous contractions of muscle strips were eliminated. In conclusion, the results of the present study suggested that activation of U-II/UT signaling promoted the development of cystitis-associated-bladder hyperactivity by targeting the RhoA/Rho-kinase pathway, indicating that the U-II/UT signaling could serve as a novel target for the treatment of interstitial cystitis/bladder pain syndrome.

The hyperpolarization-activated, cyclic nucleotide-gated channel resides on myocytes in mouse bladders and contributes to adrenergic-induced detrusor relaxation.

Control of urinary continence is predicated on sensory signaling about bladder volume. Bladder sensory nerve activity is dependent on tension, implicating autonomic control over detrusor myocyte activity during bladder filling. Hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channels are known contributors to bladder control, but their mechanism of action is not well understood. The lack of a definitive identification of cell type(s) expressing HCN in the bladder presents a significant knowledge gap. We recently reported a complete transcriptomic atlas of the C57BL/6 mouse bladder showing the dominant HCN paralog in mouse bladder, Hcn1, is limited to a subpopulation of detrusor smooth myocytes (DSMs). Here, we report details of these findings, along with results of patch-clamp experiments, immunohistochemistry, and functional myobath/tension experiments in bladder strips. With the use of a transgenic mouse expressing fluorescence-tagged α-smooth muscle actin, our data confirmed location and function of DSM HCN channels. Despite previous associations of HCN with postulated bladder interstitial cells, neither evidence of specific interstitial cell types nor an association of nonmyocytes with HCN was discovered. We confirm that HCN activation participates in reducing sustained (tonic) detrusor tension via cAMP, with no effect on intermittent (phasic) detrusor activity. In contrast, blockade of HCN increases phasic activity induced by a protein kinase A (PKA) blocker or a large-conductance Ca2+-activated K+ (BK) channel opener. Our findings, therefore, suggest a central role for detrusor myocyte HCN in regulating and constraining detrusor myocyte activity during bladder filling.

MP49-12 A NOVEL BLADDER FIBROSIS PATHWAY THROUGH PDGFRα + CELLS ASSOCIATED WITH ESTROGEN DEFICIENCY AND REPLACEMENT

You have accessJournal of UrologyCME1 May 2022MP49-12 A NOVEL BLADDER FIBROSIS PATHWAY THROUGH PDGFRα+ CELLS ASSOCIATED WITH ESTROGEN DEFICIENCY AND REPLACEMENT Ken Lee, Masatoshi Eto, Kenton Sanders, and Sang don Koh Ken LeeKen Lee More articles by this author , Masatoshi EtoMasatoshi Eto More articles by this author , Kenton SandersKenton Sanders More articles by this author , and Sang don KohSang don Koh More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002624.12AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: E2 deficiency may induce fibrosis that leads to bladder dysfunction. We aimed at studying the effects of E2 on bladder function and assessing changes of bladder PDGFRα+ cells associated with inflammation and fibrosis. METHODS: C57BL/6 female mice were bilaterally ovariectomized (OVX) or sham surgery was performed at 7 weeks old and then E2 was applied subcutaneously in OVX mice for two weeks before experiment. Each protocol was done 1, 3 and 7 months after OVX. In vivo cystometrogram (CMG), mice were restrained with specific holder under awake condition. qPCR, western blotting (WB) and immunohistochemistry (IHC) was carried out to investigate the relationship between E2 deficiency/replacement and fibrosis marker. To further explore this, bladder was divided into urothelium/suburothelial layer (UT) and detrusor smooth muscle layer (SM) and then cultured with control media, E2 (10 ng/ml) or TNFα (100 ng/ml) for 7 days. Cultured tissues were frozen at -80°C to study fibrosis marker. RESULTS: Bladder compliance was decreased in 1, 3 and 7 month OVX groups (20.3±6.5, 10.2±1.5 and 14.2±2.7 μl/mmHg) compared to age-matched sham groups (35.1±6.1, 24.6±3.2 and 24.2±4.5 μl/mmHg), while E2 recovered bladder compliance. Bladder capacity was reduced in 3 and 7 month OVX groups compared to sham, but was not fully recovered by E2. Intercontraction interval (ICI) was reduced in 3 and 7 month OVX groups (10.3±1.8 and 10.7±1.5 min) compared to age-matched sham groups (17.0±1.1 and 17.0±3.0 min) but was not rescued by E2 treatment. Unexpectedly, E2 markedly increased the number of non-voiding contractions (NVCs). Estrogen receptor (ERα) was dominantly immunoreactive against detrusor, however bladder interstitial cells were immunopositive upon ERβ. Colocalizations of PDGFRα+ and ERβ positive interstitial cells were observed both in UT and SM. Expression of PDGFRα and collagen 1a1 was upregulated in 3 month OVX group compared to sham, while E2 replacement further increased the expression of PDGFRα and fibrosis markers compared to age-matched sham or OVX groups. In qPCR of cultured bladder tissue, TNFα dramatically upregulated the expression of Acta2, TIMP2, Tgfβ, Col1a1, Col1a2, cadherin and IL6 in UT but had marginal effects on SM. CONCLUSIONS: Urothelium/suburothelial layer may be common site for the initiation of fibrosis. Inflammation or E2 deficiency/replacement may cause fibrosis through the activation of PDGFRα+ cells. These novel findings might point to a novel approach toward the treatment of bladder fibrosis. Source of Funding: Supported by NIDDK, RO1 DK098388 © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e858 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ken Lee More articles by this author Masatoshi Eto More articles by this author Kenton Sanders More articles by this author Sang don Koh More articles by this author Expand All Advertisement PDF DownloadLoading ...

Expression and activity of pyruvate kinase in bladder interstitial cells of cajal in rats with diabetic cystopathy

Objective To investigate the expression and activity of pyruvate kinase in interstitial cells of Cajal (ICCs) in the bladder of rats with diabetic cystopathy (DCP). Methods Forty 8-week-old female rats were divided into two groups randomly, 31 as the experiment group and the rest 9 as the control group. The experiment group was intraperitoneally injected with streptozotocin (STZ) to establish diabetic models and 18 rats were succeeded to be induced. Nine diabetic rats were further raised for 12 weeks getting DCP rats. ICCs from bladder muscle were cultured. The expression of pyruvate kinase protein and mRNA was detected by Western blotting and real-time quantitative polymerase chain reaction (Real-time PCR) respectively, and pyruvate kinase activity was measured. Results The expression levels of pyruvate kinase protein and mRNA and activity in DCP rats were reduced significantly as compared with those of the diabetic rats and control rats [(0.537±0.015), (0.624±0.026), (0.098±0.010) U/g·prot respectively, vs. (0.722±0.026), (0.739±0.045), (0.178±0.017) U/g·prot, vs. (0.923±0.031), (1.000±0.022), (0.277±0.030) U/g·prot; P<0.01]. Conclusion The reduction of pyruvate kinase activity in ICCs may play an important role in the development of DCP. Key words: Diabetic cystopathy; Pyruvate kinase; Interstitial cells of cajal

Increased Piezo1 channel activity in interstitial Cajal-like cells induces bladder hyperactivity by functionally interacting with NCX1 in rats with cyclophosphamide-induced cystitis

The Piezo1 channel is a mechanotransduction mediator, and Piezo1 abnormalities have been linked to several clinical disorders. However, the role of the Piezo1 channel in cystitis-associated bladder dysfunction has not been documented. The current study aimed to discover the functional role of this channel in regulating bladder activity during cyclophosphamide (CYP)-induced cystitis. One hundred four female rats were randomly assigned to the control, CYP-4h, CYP-48h and CYP-8d groups. CYP successfully induced acute or chronic cystitis in these rats. CYP treatment for 48h or 8d significantly increased Piezo1 channel expression in bladder interstitial Cajal-like cells (ICC-LCs), and the increase in CYP-8d rats was more prominent. In addition, 2.5 μM Grammostola spatulata mechanotoxin 4 (GsMTx4) significantly attenuated bladder hyperactivity in CYP-8d rats by inhibiting the Piezo1 channel in bladder ICC-LCs. Furthermore, by using GsMTx4 and siRNA targeting the Piezo1 channel, we demonstrated that hypotonic stress-induced Piezo1 channel activation significantly triggered Ca2+ and Na+ influx into bladder ICC-LCs during CYP-induced chronic cystitis. In addition, the Piezo1 channel functionally interacted with the relatively activated reverse mode of Na+/Ca2+ exchanger 1 (NCX1) in bladder ICC-LCs from CYP-8d rats. In conclusion, we suggest that the functional role of the Piezo1 channel in CYP-induced chronic cystitis is based on its synergistic effects with NCX1, which can significantly enhance [Ca2+]i and result in Ca2+ overload in bladder ICC-LCs, indicating that the Piezo1 channel and NCX1 are potential novel therapeutic targets for chronic cystitis-associated bladder hyperactivity.

Decreased hyperpolarization-activated cyclic nucleotide-gated channels are involved in bladder dysfunction associated with spinal cord injury

Spinal cord injury (SCI) leads to bereft voluntary control of bladder, but the possible role of spontaneous excited system in bladder of SCI patients is poorly understood. Hyper polarization-activated cyclic nucleotide-gated (HCN) channels are deemed to regulate the spontaneous contraction of bladder, our study explored the functional role of HCN channels in SCI induced neurogenic bladder. Sixty female Sprague-Dawley rats were randomized into control, sham and SCI groups. Rat models subjected to SCI at S2 levels were successfully established and were assessed using hematoxylin and eosin staining and cystometry. In SCI rats, the mRNA and protein expression levels of HCN channels and the Ih density were significantly reduced, and expression levels of several bladder HCN1 channel regulatory proteins were also significantly changed. The effects of 50 µM forskolin and 50 µM 8-bromoadenosine 3′,5′-cyclic monophosphate on [Ca2+]i of isolated bladder interstitial cells of Cajal-like cells were significantly decreased in SCI rats. The spontaneous contractions in detrusor strips from SCI rats were significantly weakened. Furthermore, detrusor strips from SCI rats exhibited decreased tolerance to two doses of ZD7288 (10 and 50 µM). Taken together, our results indicate that the decreased bladder HCN channel expression and function induced by altered regulatory proteins are involved in the pathological process of SCI induced neurogenic bladder, which present HCN channels as valid therapeutic targets for treating this disease.

Relationship between the change of bladder interstitial cells of Cajal and bladder compliance post-spinal cord transection at different time points in rats

Objective To observe the change of bladder compliance and bladder interstitial cells of Cajal (ICCs) post-suprasacral spinal cord transection at different time points in rats and the effect of Imatinib mesylate on the complicance of neurogenic bladder caused by long-term suprasacral spinal cord transaction in rats. Methods One hundred and two female Wistar rats were randomly divided into 6 groups: group A (sham group of 6 weeks); group B (suprasacral spinal cord transaction group of 6 weeks); group C and group E (sham group of 12 weeks); group D and group F (suprasacral spinal cord transaction group of 12 weeks). The rats in groups A to D were used to study the change of bladder compliance and bladder ICCs post-suprasacral spinal cord transection at different time points. The rats in group E and group F were used to study the effect of Imatinib mesylate on the complicance of neurogenic bladder caused by long-term suprasacral spinal cord transaction. Spinal cord transection at T10 level was built in spinal cord transaction groups, while group A, group C and group F underwent sham operaion. Urodynamics was detected at 6th or 12th week after surgery. We observed the changes of ICCs in the bladder wall by immunohistochemistry. Urodynamics was also detected in group E and group F post-surgery 12 weeks before and after intravenous administration of Imatinib mesylate (5 μmol and 20 μmol respectively). Results The bladder compliance was significantly increased [(0.146±0.107) ml/cmH2O vs. (0.066±0.019) ml/cmH2O, P=0.017] and the number of ICCs was significantly decreased [(3.180±1.976) vs. (4.920±1.564), P=0.014] in group B as compared with group A. The bladder compliance was significantly lower [(0.046±0.023) ml/cmH2O vs. (0.063±0.013) ml/cmH2O] and the number of ICCs was significantly higher [(7.710±2.024) vs. (4.750±1.138), P=0.000] in group D than in group C. The bladder compliance in group E was increased by intravenous administration of 20 μmol Imatinib mesylate [from (0.070±0.029) ml/cmH2O to (0.096±0.036) ml/cmH2O]. It increased the bladder compliance of group F in a dose-dependent manner. The bladder compliance in group F was increased by 5 μmol and 20 μmol Imatinib mesylate from (0.047±0.021) ml/cmH2O to (0.065±0.019) ml/cmH2O and (0.077±0.019) ml/cmH2O respectively, P=0.000. Conclusion The bladder compliance was increased first, then decreased and the number of ICCs changed adversely in the T10 spinal cord transection rats. The bladder compliance of the rats at 12th week post-suprasacral spinal cord transection could be improved by inhibiting the ICCs function. Not only the fibrosis, but also the number of ICCs of the bladder wall increased might be the pathological basis of the reduced bladder compliance in suprasacral spinal cord transaction rats. Key words: Spinal cord injury; Neurogenic bladder; Bladder compliance; Interstitial cells of Cajal; Imatinib mesylate

Cyclophosphamide-induced HCN1 channel upregulation in interstitial Cajal-like cells leads to bladder hyperactivity in mice

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are confirmed to be expressed in bladder interstitial Cajal-like cells (ICC-LCs), but little is known about their possible role in cystitis-associated bladder dysfunction. The present study aimed to determine the functional role of HCN channels in regulating bladder function under inflammatory conditions. Sixty female wild-type C57BL/6J mice and sixty female HCN1-knockout mice were randomly assigned to experimental and control groups, respectively. Cyclophosphamide (CYP)-induced cystitis models were successfully established in these mice. CYP treatment significantly enhanced HCN channel protein expression and Ih density and significantly altered bladder HCN1 channel regulatory proteins. Carbachol (CCH) and forskolin (FSK) exerted significant effects on bladder ICC-LC [Ca2+]i in CYP-treated wild-type (WT) mice, and HCN1 channel ablation significantly decreased the effects of CCH and FSK on bladder ICC-LC [Ca2+]i in both naive and CYP-treated mice. CYP treatment significantly potentiated the spontaneous contractions and CCH (0.001–10 μM)-induced phasic contractions of detrusor strips, and HCN1 channel deletion significantly abated such effects. Finally, we demonstrated that the development of CYP-induced bladder overactivity was reversed in HCN1−/− mice. Taken together, our results suggest that CYP-induced enhancements of HCN1 channel expression and function in bladder ICC-LCs are essential for cystitis-associated bladder hyperactivity development, indicating that the HCN1 channel may be a novel therapeutic target for managing bladder hyperactivity.

205 - The stem cell growth factor receptor KIT is not expressed on interstitial cells in bladder

205 - The stem cell growth factor receptor KIT is not expressed on interstitial cells in bladder

The stem cell growth factor receptor KIT is not expressed on interstitial cells in bladder.

The mast/stem cell growth factor receptor KIT has long been assumed to be a specific marker for interstitial cells of Cajal (ICC) in the bladder, with possible druggable perspectives. However, several authors have challenged the presence of KIT + ICC in recent years. The aim of this study was therefore to attempt to clarify the conflicting reports on KIT expression in the bladder of human beings, rat, mouse and guinea pig and to elucidate the possible role of antibody‐related issues and interspecies differences in this matter. Fresh samples were obtained from human, rat, mouse and guinea pig cystectomies and processed for single/double immunohistochemistry/immunofluorescence. Specific antibodies against KIT, mast cell tryptase (MCT), anoctamin‐1 (ANO1) and vimentin were used to characterize the cell types expressing KIT. Gut (jejunum) tissue was used as an external antibody control. Our results revealed KIT expression on mast cells but not on ICC in human, rat, mouse and guinea pig bladder. Parallel immunohistochemistry showed KIT expression on ICC in human, rat, mouse and guinea pig gut, which confirmed the selectivity of the KIT antibody clones. In conclusion, we have shown that KIT + cells in human, rat, mouse and guinea pig bladder are mast cells and not ICC. The present report is important as it opposes the idea that KIT + ICC are present in bladder. In this perspective, functional concepts of KIT + ICC being involved in sensory and/or motor aspects of bladder physiology should be revised.

Waterworks.

Having taught for some years in medical school, I still vividly remember the A+ student who collapsed in the library hyperventilating while studying pathological ventilation patterns, definitely a case of too much focussing all of one's attention on the physiology text in front of herself. I have always wondered what happens when they progress to the urine production chapter. Jokes aside, the human urinary tract is a fascinating, tightly regulated organ system. Understanding its function is of increasing relevance for medical caregivers, especially those dealing with ever-ageing patient populations in the Western world. Demographic change forces us to refocus our attention to diseases of the elderly with ever-increasing life expectancies, such as non-neurogenic male lower urinary tract symptoms (Gratzke et al. 2015), overactive bladder symptoms of the elderly (Wolff et al. 2015) and bladder cancer. Age and male gender are the risk factors for bladder cancer, with a median age at diagnosis of 70 years and a male-to-female ratio of 3 : 1. Incidence rates are on the rise, with approx. 380 000 new cases and approx. 150 000 deaths per year worldwide (Knowles & Hurst 2015). Traditionally, the kidney had been one of the hot topics of physiological research, which has increasingly been extending its focus from basic mechanisms and molecular kidney physiology (Svenningsen et al. 2014) to disease mechanisms underlying and/or associated with Western lifestyles and ageing populations, such as essential hypertension (Damkjaer et al. 2014), metabolic syndrome and acute and chronic kidney injury (Seibert et al. 2013, Singer et al. 2013, Svenningsen et al. 2013). Acta Physiologica has recently added to our understanding of lower urinary tract structure and function and its pathophysiology. In 2013, Nevéus & Sillén shared a closer look on the early development of bladder function and disturbances in childhood. Embryonic bladder development starts at an early gestational age. At birth, higher CNS centres control micturition, while control of urinary function develops as the child grows and matures, a process which, for many parents, resembles a rather bumpy, curvy ride. Often, however, simple measures such as parent education and non-medical bladder training help significantly. Continued incontinence or nocturnal enuresis may become major issue for an astonishingly high percentage of parents, as approx. 5–10% of 7-year-old children suffer from daytime incontinence and/or nocturnal enuresis, with some never outgrowing it (Nevéus & Sillén 2013). Bladder problems in childhood increase the risk of developing according dysfunctions in adulthood (Nevéus & Sillén 2013) such as nocturia, which is, due to its multi-factorial aetiopathogenesis, now viewed as a distinct clinical condition rather than a mere symptom (Osman et al. 2013). The excellent articles of both Griffiths & Fowler (2013) and de Groat & Wickens (2013) have shed light on recent insights into the mechanisms and circuits by which the CNS controls micturition. Urothelial signalling cascades give rise to the sensation that initiates the perception of ‘needing to go’ (Birder & Wyndaele 2013), while continence and voiding are controlled in close cooperation of the autonomous and somatic systems (Ochodnicky et al. 2013). When we take a closer look at signalling cascades and events in the lower urinary tract, purinergic signalling pathways are of major importance (Avelino et al. 2013, Birder et al. 2013, Burnstock 2013). Current research on bladder interstitial cells, several different subgroups of which have been recently identified within the bladder wall to be interacting with nerves and smooth muscle and probably critically involved in intercellular communication and key physiological functions (McCloskey 2013), is but one example of an emerging field in research on lower urinary tract structure and function. Both neurodegenerative disease (Feinstein et al. 2015) and traumatic damage to the spinal cord (Thomas et al. 2014, Gomez-Amaya et al. 2015) are often accompanied by urinary tract dysfunction. Improved overall survival of patients suffering from according pathologies are accompanied by an increasing relevance of these dysfunctions, which may severely affect the quality of life and ability to participate in day-to-day activities. Thus, a better understanding of the molecular and (patho-) physiological mechanisms will help provide novel opportunities for prognostic patient evaluation, monitoring and treatment. Time for a comfort break. None.

Changes in hyperpolarization-activated cyclic nucleotide-gated channel expression and activity in bladder interstitial cells of Cajal from rats with detrusor overactivity.

To investigate changes in the expression and function of the hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, interstitial cells of Cajal (ICCs) were isolated from detrusor overactivity (DO) rats induced with partial bladder outlet obstruction (PBOO). We hypothesized that increased expression of HCN channels in ICCs would enhance the excitability of ICCs and bladder in DO rats. Forty adult female Sprague-Dawley rats were randomly assigned to control and DO groups. The expression of HCN isoforms in the rat bladders was detected using reverse transcription-polymerase chain reaction (RT-PCR). Whole-cell patch-clamp techniques and laser confocal microscopy were used to explore the effects of the HCN channel blocker ZD7288 on Ih current and intracellular calcium levels ([Ca(2+)]i) in freshly isolated ICCs. The effect of ZD7288 on bladder contraction was evaluated using a bladder smooth muscle strip test. The results of RT-PCR showed that HCN1-4 isoforms increased expression in the DO group compared with the control group. The current density of Ih in the bladder ICCs was increased. A moderate concentration of ZD7288 (50 μmol/L) significantly decreased the [Ca(2+)]i levels in freshly isolated ICCs from sham and DO bladders. The smooth strip tests indicated that ZD7288 (50 μmol/L) suppressed the amplitude of smooth muscle strip contractions in the sham and DO bladders. All HCN channel isoforms were highly expressed in bladders from the DO group, which was correlated with increased Ih currents in DO ICCs, suggesting that the HCN channels might play an important role in the pathological processes of DO.

Expression and function of muscarinic subtype receptors in bladder interstitial cells of cajal in rats.

To locate the muscarinic (M) M2 and M3 receptors in bladder interstitial cells of Cajal (ICCs) and to determine the effects of M2 and M3 agonists on bladder ICCs. A total of 30 adult male Sprague-Dawley rats weighing 225-250 g were used in this study. Double-labeled fluorescence of muscarinic receptors and c-kit was performed for co-localization. To evaluate the effect of muscarinic agents on the excitation of bladder ICCs, we analyzed the inward current of bladder ICCs using the whole-cell patch clamp. The effect of muscarinic agents on the carbachol-induced inward currents was evaluated with the whole-cell patch clamp. M2 and M3 receptors were confirmed in the stroma ICCs in rats' bladders with double-labeled immunofluorescence. Spontaneous action potential was observed in freshly isolated bladder ICCs. The carbachol-induced inward Ca2+ current in ICCs can be blocked by atropine. The M2 receptor antagonist methoctramine (1 μM) showed a weak inhibitory capability on the inward Ca2+ current [from 74.8 ± 9.6 to 63.3 ± 13.8 Pascal (pA), n = 12, P = .03]. While the M3 receptor antagonist 4-diphenyl-acetoxy-N-methyl-piperidine methiodide (4-DAMP) (1 μM) significantly inhibited the inward Ca2+ current (from 78.4 ± 11.2 to 17.3 ± 7.9 pA, n = 12, P < .001). Bladder ICCs express M2 and M3 cholinergic receptors. Most muscarinic cholinergic receptor antagonists, especially the M3 antagonists, can effectively inhibit the carbamylcholine- induced inward current of bladder ICCs.

Bladder interstitial cells and pathophysiology

Bladder interstitial cell (IC) is a cell, which lacks thick filaments and dense bodies but with incomplete basement membrane, rough endoplasmic reticulum and golgi apparatus. IC is divided into 4 subtypes: lamina propria IC, intramuscular IC, IC between the detrusor bundles and perivascular IC. There are different ion currents and related activation pathways in the lamina propria IC and intramuscular IC. Ca2+ signaling pathways play an important role in the communication between IC and detrusor. Any bladder lesions affecting the ion current and Ca2+ signaling pathways can lead to bladder dysfunction. The bladder lesions include bladder outlet obstruction, bladder pain syndrome, interstitial cystitis, neurogenic bladder and diabetes. Imatinib mesylate is currently an available treatment target in IC, and electrical stimulation of acupuncture therapy is a new direction.

Do we understand any more about bladder interstitial cells?-ICI-RS 2013

To present a brief review on discussions from "Do we understand any more about lower urinary tract interstitial cells?" session at the 2013 International Consultation on Incontinence-Research Society (ICI-RS) meeting in Bristol, UK. Discussion focused on bladder interstitial cell (IC) subtypes, their localization and characterization, and communication between themselves, the urothelium, and detrusor smooth muscle. The role of ICs in bladder pathologies and new methods for studying ICs were also addressed. ICs have been studied extensively in the lower urinary tract and have been characterized based on comparisons with ICs of Cajal in the gastro-intestinal tract. In fetal bladders it is believed that ICs drive intrinsic contractions to expel urine through the urachus. These contractions diminish postpartum as bladder innervation develops. Voiding in human neonates occurs when filling triggers a spinal cord reflex that contracts the detrusor; in rodents, maternal stimulation of the perineum triggers voiding. Following spinal cord injury, intrinsic contractions, and spinal micturition reflexes develop, similar to those seen during neonatal development. These enhanced contractions may stimulate nociceptive and mechanosensitive afferents contributing to neurogenic detrusor overactivity and incontinence. The IC-mediated activity is believed to be initiated in the lamina propria by responding to urothelial factors. These IC may act syncytially through gap junction coupling and modulate detrusor activity through unknown mechanisms. There has been a great deal of information discovered regarding bladder ICs, however, many of their (patho)physiological functions and mechanisms are still unclear and necessitates further research. Neurourol. Urodynam. 33:573-576, 2014. © 2014 Wiley Periodicals, Inc.

Change of the interstitial cells of Cajal in the bladder of guinea pig with diabetic cystopathy

Objective To investigate the bladder interstitial cells of Cajal (ICC) function in the pathogenesis of the diabetic cystopathy.Methods Fifty short hair Britain female guinea pigs were divided into two groups randomly,42 as the experiment group and the other 8 as the control group (group C).The experiment group was intraperitoneally injected with streptozotocin (STZ) to establish diabetic models.Diabetic guinea pigs were further divided into the decompensated bladder function group (namely DCP,group A) and compensated group (group B) by urodynamic study.Immunofluorescence staining,confocal laser microscope and transmission electron microscopy (TEM) technology were used to detect the morphology and ultrastructure changes of the ICC.Results The percentage of the ICC in group A was significantly lower than that of the group B and group C [(2.18 ± 0.04) ％ vs.(2.80 ± 0.05,2.81 ± 0.04) ％ respectively,P ＜0.01].Under the electron microscopic observation,the mitochondria in the ICC were reduced in the DCP group.The substrate also became less and shallow,and crest got shorter or even disappeared.Conclusion ICC may play an important role in the development of the DCP. Key words: Guinea pig; Diabetic Cystopathy; Interstitial cell of Cajal

Functional innervation of bladder interstitial cells

Interstitial cells (IC) in the urinary bladder have been studied for more than a decade. It is now known that IC represent a heterogeneous group of cells with sub-populations in the mucosal and detrusor layers; however, there is continued debate on their role in normal and diseased bladder.