Effects of fatty acid metabolites on nocturia.

Dysregulation of circadian rhythm can cause nocturia. Levels of fatty acid metabolites, such as palmitoylethanolamide (PEA), 9-hydroxy-10E,12Z-octadecadienoic acid (9-HODE), and 4-hydroxy-5E,7Z,10Z,13Z,16Z,19Z-docosahexaenoic acid (4-HDoHE), are higher in the serum of patients with nocturia; however, the reason remains unknown. Here, we investigated the circadian rhythm of fatty acid metabolites and their effect on voiding in mice. WT and Clock mutant (ClockΔ19/Δ19) mice, a model for nocturia with circadian rhythm disorder, were used. Levels of serum PEA, 9-HODE, and 4-HDoHEl were measured every 8 h using LC/MS. Voiding pattern was recorded using metabolic cages after administration of PEA, 9-HODE, and 4-HDoHE to WT mice. Levels of serum PEA and 9-HODE fluctuated with circadian rhythm in WT mice, which were lower during the light phase. In contrast, circadian PEA and 9-HODE level deteriorated or retreated in ClockΔ19/Δ19 mice. Levels of serum PEA, 9-HODE, and 4-HDoHE were higher in ClockΔ19/Δ19 than in WT mice. Voiding frequency increased in PEA- and 4-HDoHE-administered mice. Bladder capacity decreased in PEA-administered mice. The changes of these bladder functions in mice were similar to those in elderly humans with nocturia. These findings highlighted the novel effect of lipids on the pathology of nocturia. These may be used for development of biomarkers and better therapies for nocturia.

Different effects of GsMTx4 on nocturia associated with the circadian clock and Piezo1 expression in mice.

OBJECTIVES: Nocturia is a major problem in geriatric patients. Clock genes regulate circadian bladder function and Piezo type mechanosensitive ion channel component 1 (Piezo1) that senses bladder fullness. We utilized WT and Clock mutant (ClockΔ19/Δ19: nocturia phenotype) mice to determine if the effects of GsMTx4, a Piezo1 inhibitor, is dependent on circadian Piezo1 expression in the bladder. METHODS: We compared voiding behavior in mice after the administration of vehicle, low dose, or high dose of GsMTx4. Intraperitoneal injections (IP) were performed at Zeitgeber time (ZT) 0, lower Piezo1 expression phase (ZT0-IP) and ZT12, higher Piezo1 expression phase (ZT12-IP). Urine volume (Uvol), voiding frequency (VF), and urine volume per void (Uvol/v) were measured using metabolic cages. RESULTS: VF decreased at ZT12-IP in WT mice only with high dose of GsMTx4 but showed no effects in ClockΔ19/Δ19 mice. VF decreased significantly at ZT0-IP in WT mice after both doses, but only decreased after high dose in ClockΔ19/Δ19 mice. Uvol/v increased in WT mice at ZT0-IP after both doses and at ZT12-IP after high dose. Uvol/v increased in ClockΔ19/Δ19 mice only at ZT0-IP after high dose. GsMTx4 did not affect Uvol in both mice at ZT12-IP. A decrease in Uvol was observed in both mice at ZT0-IP; however, it was unrelated to GsMTx4-IP. CONCLUSIONS: The effects of GsMTx4 changed associated with the circadian clock and Piezo1 expression level. The maximum effect occurred during sleep phase in WT. These results may lead to new therapeutic strategies against nocturia.

Phenotyping the Association between Nocturia and Hypertension: A Systematic Review and Meta-Analysis.

PURPOSE: The association between nocturia and hypertension has been widely reported yet remains poorly characterized, precluding a more refined understanding of blood pressure as it relates to the clinical urology setting. We synthesized current evidence on the relationship between nocturia and hypertension as a function of nocturia severity, age, gender, race, body mass index and diuretic use. MATERIALS AND METHODS: We searched PubMed®, EMBASE® and Cochrane databases for studies published up to May 2020. Random effects meta-analyses were performed to identify pooled odds ratios for nocturia given the presence of hypertension. Meta-regression and subgroup analyses were performed to identify differences across study samples. RESULTS: Of 1,193 identified studies, 25 met the criteria for inclusion. The overall pooled OR for the association of nocturia with hypertension was 1.25 (95% CI 1.21-1.28, p <0.001). Pooled estimates were 1.20 (1.16-1.25, p <0.001) and 1.30 (1.25-1.36, p <0.001) using a 1-void and 2-void cutoff for nocturia, respectively (p <0.001 between cutoffs). The association was more robust in patient-based (1.74 [1.54-1.98], p <0.001) vs community-based (1.24 [1.24-1.29], p <0.001) study samples (p <0.001). The association was stronger in females compared to males (1.45 [1.32-1.58] vs 1.28 [1.22-1.35], p <0.001), and Black (1.56 [1.25-1.94]) and Asian (1.28 [1.23-1.33]) vs White subgroups (1.16 [1.08-1.24]; p <0.05 for both). No effect was observed for age or body mass index. Evidence on diuretics was limited. CONCLUSIONS: Hypertension is associated with a 1.2-fold to 1.3-fold higher risk of nocturia. This association is more robust at a higher nocturia cutoff, in patient-based study samples, among females and in Black and Asian patients, but unrelated to age or body mass index.

Neurologic, clinical, and immunologic features in a cohort of HTLV-1 carriers with high proviral loads.

A high proviral load (PVL) is recognized as a risk factor for human T cell leukemia virus-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), but there is a lack of prospective studies evaluating whether or not HTLV-1 carriers with high PVL are at risk of developing HAM/TSP or other HTLV-1-related diseases. Here, we compare the incidence of clinical manifestations and the cytokine levels in 30 HTLV-1 carriers with high (> 50,000 copies/106 PBMC) and an equal number of subjects with low proviral load. Participants were followed for 3 to 16 years (median of 11 years). The PVL, IFN-γ, TNF, and IL-10 levels were quantified at entry and at the end of the follow-up. Among the self-reported symptoms in the initial evaluation, only the presence of paresthesia on the hands was more frequent in the group with high PVL (p < 0.04). The production of IFN-γ was higher in the group with high PVL group (median of 1308 versus 686 pg/ml, p < 0.011) when compared with the control group in the first assessment. There was no difference in the occurrence of urinary symptoms or erectile dysfunction, periodontal disease, Sicca syndrome, and neurologic signs between the two groups during the follow-up. The observation that none of the HTLV-1 carriers with high PVL and with exaggerated inflammatory response progressed to HAM/TSP indicates that other factors in addition to the PVL and an exaggerated immune response are involved in the pathogenesis of HAM/TSP.

Intermittent restraint stress induces circadian misalignment in the mouse bladder, leading to nocturia.

Intermittent stress disrupts the circadian rhythm in clock genes such as Per2 only in peripheral organs without any effect on the central circadian clock in the suprachiasmatic nucleus. Here, the effect of restraint stress (RS) on circadian bladder function was investigated based on urination behavior and gene expression rhythms. Furthermore, PF670462 (PF), a Per2 phosphorylation enzyme inhibitor, was administered to investigate the effects on circadian bladder re-alignment after RS. Two-hour RS during the light (sleep) phase was applied to mice (RS mice) for 5 days. The following parameters were then examined: urination behaviors; clock gene expression rhythms and urinary sensory-related molecules such as piezo type mechanosensitive ion channel component 1 (Piezo1), transient receptor potential cation channel subfamily V member 4 (TRPV4), and Connexin26 (Cx26) in the bladder mucosa; Per2 expression in the excised bladder of Per2luciferase knock-in mice (Per2::luc); in vivo Per2 expression rhythms in the bladder of Per2::luc mice. Control mice did not show altered urination behavior in the light phase, whereas RS mice exhibited a higher voiding frequency and lower bladder capacity. In the bladder mucosa, RS mice also showed abrogated or misaligned Piezo1, TRPV4, Connexin26, and clock gene expression. The rhythmic expression of Per2 was also altered in RS mice both in excised- and in vivo bladder, compared with control mice. After PF administration, voiding frequency was reduced and bladder capacity was increased during the light phase in RS mice; the in vivo Per2 expression rhythm was also fully restored. Therefore, RS can alter circadian gene expression in the bladder during the light phase and might cause nocturia via changes in circadian bladder function due the dysregulation of clock genes. Amending the circadian rhythm therapeutically could be applied for nocturia.

The time-dependent variation of ATP release in mouse primary-cultured urothelial cells is regulated by the clock gene.

AIMS: The sensation of bladder fullness (SBF) is triggered by the release of ATP. Therefore, the aim of this study was to investigate whether time-dependent changes in the levels of stretch-released ATP in mouse primary-cultured urothelial cells (MPCUCs) is regulated by circadian rhythm via clock genes. METHODS: MPCUCs were derived from wild-type and Clock mutant mice (ClockΔ19/Δ19 ), presenting a nocturia phenotype. They were cultured in elastic silicone chambers. Stretch-released ATP was quantified every 4 h by ATP photon count. An experiment was also performed to determine whether ATP release correlated with the rhythm of the expression of Piezo1, TRPV4, VNUT, and Connexin26 (Cx26) in MPCUCs regulated by clock genes with circadian rhythms. MPCUCs were treated with carbenoxolone, an inhibitor of gap junction protein; were derived from VNUT-KO mice; or treated with Piezo1-siRNA, TRPV4-siRNA, and Cx26-siRNA. RESULTS: Stretch-released ATP showed time-dependent changes in wild-type mice and correlated with the rhythm of the expression of Piezo1, TRPV4, VNUT, and Cx26. However, these rhythms were disrupted in ClockΔ19/Δ19 mice. Carbenoxolone eliminated the rhythmicity of ATP release in wild-type mice. However, time-dependent ATP release changes were maintained when a single gene was deficient such as VNUT-KO, Piezo1-, TRPV4-, and Cx26-siRNA. CONCLUSIONS: ATP release in the bladder urothelium induces SBF and may have a circadian rhythm regulated by the clock genes. In the bladder urothelium, clock gene abnormalities may disrupt circadian ATP release by inducing Piezo1, TRPV4, VNUT, and Cx26. All these genes can trigger nocturia.

The oscillation of intracellular Ca2+ influx associated with the circadian expression of Piezo1 and TRPV4 in the bladder urothelium.

We previously showed that bladder functions are controlled by clock genes with circadian rhythm. The sensation of bladder fullness (SBF) is sensed by mechano-sensor such as Piezo1 and TRPV4 in the mouse bladder urothelium. However, functional circadian rhythms of such mechano-sensors remain unknown. To investigate functional circadian changes of these mechano-sensors, we measured circadian changes in stretch-evoked intracellular Ca2+ influx ([Ca2+] i ) using mouse primary cultured urothelial cells (MPCUCs). Using Ca2+ imaging, stretch-evoked [Ca2+] i was quantified every 4 h in MPCUCs derived from wild-type (WT) and Clock Δ19/Δ19 mice, which showed a nocturia phenotype. Furthermore, a Piezo1 inhibitor GsMTx4 and a TRPV4 inhibitor Ruthenium Red were applied and stretch-evoked [Ca2+] i in MPCUCs was measured to investigate their contribution to SBF. Stretch-evoked [Ca2+] i showed a circadian rhythm in the WT mice. In contrast, Clock Δ19/Δ19 mice showed disrupted circadian rhythm. The administration of both GsMTx4 and Ruthenium Red eliminated the circadian rhythm of stretch-evoked [Ca2+] i in WT mice. We conclude that SBF may have a circadian rhythm, which is created by functional circadian changes of Piezo1 and TRPV4 being controlled by clock genes to be active during wakefulness and inactive during sleep. Abnormalities of clock genes disrupt SBF, and induce nocturia.

Machine Learning on a Genome-wide Association Study to Predict Late Genitourinary Toxicity After Prostate Radiation Therapy.

PURPOSE: Late genitourinary (GU) toxicity after radiation therapy limits the quality of life of prostate cancer survivors; however, efforts to explain GU toxicity using patient and dose information have remained unsuccessful. We identified patients with a greater congenital GU toxicity risk by identifying and integrating patterns in genome-wide single nucleotide polymorphisms (SNPs). METHODS AND MATERIALS: We applied a preconditioned random forest regression method for predicting risk from the genome-wide data to combine the effects of multiple SNPs and overcome the statistical power limitations of single-SNP analysis. We studied a cohort of 324 prostate cancer patients who were self-assessed for 4 urinary symptoms at 2 years after radiation therapy using the International Prostate Symptom Score. RESULTS: The predictive accuracy of the method varied across the symptoms. Only for the weak stream endpoint did it achieve a significant area under the curve of 0.70 (95% confidence interval 0.54-0.86; P = .01) on hold-out validation data that outperformed competing methods. Gene ontology analysis highlighted key biological processes, such as neurogenesis and ion transport, from the genes known to be important for urinary tract functions. CONCLUSIONS: We applied machine learning methods and bioinformatics tools to genome-wide data to predict and explain GU toxicity. Our approach enabled the design of a more powerful predictive model and the determination of plausible biomarkers and biological processes associated with GU toxicity.

Clock Genes Regulate the Circadian Expression of Piezo1, TRPV4, Connexin26, and VNUT in an Ex Vivo Mouse Bladder Mucosa.

OBJECTIVES: ClockΔ19/Δ19 mice is an experimental model mouse for nocturia (NOC). Using the bladder mucosa obtained from ClockΔ19/Δ19 mice, we investigated the gene expression rhythms of mechanosensory cation channels such as transient receptor potential cation channel subfamily V member 4 (TRPV4) and Piezo1, and main ATP release pathways including vesicular nucleotide transporter (VNUT) and Connexin26(Cx26), in addition to clock genes. MATERIALS AND METHODS: Eight- to twelve-week-old male C57BL/6 mice (WT) and age- and sex-matched C57BL/6 ClockΔ19/Δ19 mice, which were bred under 12-h light/dark conditions for 2 weeks, were used. Gene expression rhythms and transcriptional regulation mechanisms in clock genes, mechanosensor, Cx26 and VNUT were measured in the mouse bladder mucosa, collected every 4 hours from WT and ClockΔ19/Δ19 mice using quantitative RT-PCR, a Western blot analysis, and ChIP assays. RESULTS: WT mice showed circadian rhythms in clock genes as well as mechanosensor, Cx26 and VNUT. Their expression was low during the sleep phase. The results of ChIP assays showed Clock protein binding to the promotor regions and the transcriptional regulation of mechanosensor, Cx26 and VNUT. In contrast, all of these circadian expressions were disrupted in ClockΔ19/Δ19 mice. The gene expression of mechanosensor, Cx26 and VNUT was maintained at a higher level in spite of the sleep phase. CONCLUSIONS: Mechanosensor, Cx26 and VNUT expressed with circadian rhythm in the mouse bladder mucosa. The disruption of circadian rhythms in these genes, induced by the abnormalities in clock genes, may be factors contributing to NOC because of hypersensitivity to bladder wall extension.

The Clock mutant mouse is a novel experimental model for nocturia and nocturnal polyuria.

AIMS: The pathophysiologies of nocturia (NOC) and nocturnal polyuria (NP) are multifactorial and their etiologies remain unclear in a large number of patients. Clock genes exist in most cells and organs, and the products of Clock regulate circadian rhythms as representative clock genes. Clock genes regulate lower urinary tract function, and a newly suggested concept is that abnormalities in clock genes cause lower urinary tract symptoms. In the present study, we investigated the voiding behavior of Clock mutant (ClockΔ19/Δ19 ) mice in order to determine the effects of clock genes on NOC/NP. METHODS: Male C57BL/6 mice aged 8-12 weeks (WT) and male C57BL/6 ClockΔ19/Δ19 mice aged 8 weeks were used. They were bred under 12 hr light/dark conditions for 2 weeks and voiding behavior was investigated by measuring water intake volume, urine volume, urine volume/void, and voiding frequency in metabolic cages in the dark and light periods. RESULTS: No significant differences were observed in behavior patterns between ClockΔ19/Δ19 and WT mice. ClockΔ19/Δ19 mice showed greater voiding frequencies and urine volumes during the sleep phase than WT mice. The diurnal change in urine volume/void between the dark and light periods in WT mice was absent in ClockΔ19/Δ19 mice. Additionally, functional bladder capacity was significantly lower in ClockΔ19/Δ19 mice than in WT mice. CONCLUSIONS: We demonstrated that ClockΔ19/Δ19 mice showed the phenotype of NOC/NP. The ClockΔ19/Δ19 mouse may be used as an animal model of NOC and NP. Neurourol. Urodynam. 36:1034-1038, 2017. © 2016 Wiley Periodicals, Inc.

Integrated models for the prediction of late genitourinary complaints after high-dose intensity modulated radiotherapy for prostate cancer: making informed decisions.

BACKGROUND AND PURPOSE: To develop predictive models for late radiation-induced hematuria and nocturia allowing a patient individualized estimation of pre-treatment risk. MATERIALS AND METHODS: We studied 262 PCa patients treated with curative intensity modulated radiotherapy to the intact prostate or prostate bed. A total of 372 variables were used for prediction modeling, among which 343 genetic variations. Toxicity was scored using an in-house developed toxicity scale. Predictor selection is achieved by the EMLasso procedure, a penalized logistic regression method with an EM algorithm handling missing data and crossvalidation avoiding overfit. Model performance was expressed by the area under the curve (AUC) and by sensitivity and specificity. RESULTS: Variables of the model predicting late hematuria (36/262) are bladder volume receiving ⩾75 Gy, prostatic transurethral resection and four polymorphisms. (AUC = 0.80, sensitivity = 83.3%, specificity = 61.5%). The AUC drops to 0.67 when the genetic markers are left out. The model that predicts for late nocturia (29/262) contains the minimal clinical target volume (CTV) dose, the CTV volume and three polymorphisms (AUC = 0.76, sensitivity = 75.9%, specify = 67.4%). This model is a better predictor for nocturia compared to the nongenetic model (AUC of 0.60). CONCLUSIONS: We were able to develop models that predict for the occurrence of late radiation-induced hematuria and nocturia, including genetic factors which might improve the prediction of late urinary toxicity.

Acute radiation-induced nocturia in prostate cancer patients is associated with pretreatment symptoms, radical prostatectomy, and genetic markers in the TGFß1 gene.

PURPOSE: After radiation therapy for prostate cancer, approximately 50% of the patients experience acute genitourinary symptoms, mostly nocturia. This may be highly bothersome with a major impact on the patient's quality of life. In the past, nocturia is seldom reported as a single, physiologically distinct endpoint, and little is known about its etiology. It is assumed that in addition to dose-volume parameters and patient- and therapy-related factors, a genetic component contributes to the development of radiation-induced damage. In this study, we investigated the association among dosimetric, clinical, and TGFß1 polymorphisms and the development of acute radiation-induced nocturia in prostate cancer patients. METHODS AND MATERIALS: Data were available for 322 prostate cancer patients treated with primary or postoperative intensity modulated radiation therapy (IMRT). Five genetic markers in the TGFß1 gene (-800 G>A, -509 C>T, codon 10 T>C, codon 25 G>C, g.10780 T>G), and a high number of clinical and dosimetric parameters were considered. Toxicity was scored using an symptom scale developed in-house. RESULTS: Radical prostatectomy (P<.001) and the presence of pretreatment nocturia (P<.001) are significantly associated with the occurrence of radiation-induced acute toxicity. The -509 CT/TT (P=.010) and codon 10 TC/CC (P=.005) genotypes are significantly associated with an increased risk for radiation-induced acute nocturia. CONCLUSIONS: Radical prostatectomy, the presence of pretreatment nocturia symptoms, and the variant alleles of TGFß1 -509 C>T and codon 10 T>C are identified as factors involved in the development of acute radiation-induced nocturia. These findings may contribute to the research on prediction of late nocturia after IMRT for prostate cancer.

Involvement of urinary bladder Connexin43 and the circadian clock in coordination of diurnal micturition rhythm.

Nocturnal enuresis in children and nocturia in the elderly are two highly prevalent clinical conditions characterized by a mismatch between urine production rate in the kidneys and storage in the urinary bladder during the sleep phase. Here we demonstrate, using a novel method for automated recording of mouse micturition, that connexin43, a bladder gap junction protein, is a negative regulator of functional bladder capacity. Bladder connexin43 levels and functional capacity show circadian oscillations in wild-type mice, but such rhythms are completely lost in Cry-null mice having a dysfunctional biological clock. Bladder muscle cells have an internal clock, and show oscillations of connexin43 and gap junction function. A clock regulator, Rev-erbα, upregulates connexin43 transcription as a cofactor of Sp1, using Sp1 cis-elements of the promoter. Therefore, circadian oscillation of connexin43 is associated with the biological clock and contributes to diurnal changes in bladder capacity, which avoids disturbance of sleep by micturition.

Genetic influences are important for most but not all lower urinary tract symptoms: a population-based survey in a cohort of adult Swedish twins.

BACKGROUND: The relative importance of genetic and environmental factors for the occurrence of lower urinary tract symptoms (LUTS) is poorly understood. OBJECTIVE: To (1) estimate the prevalence of urinary incontinence (UI), overactive bladder (OAB), and other LUTS and (2) to assess the heritability of these symptoms. DESIGN, SETTING, AND PARTICIPANTS: Cross-sectional survey of LUTS in a national population-based cohort of Swedish twins 20-46 yr of age (n=42 582) from the Swedish Twin Registry. MEASUREMENTS: Prevalence rates were determined and heritability of LUTS (in female twins) was assessed using indicators of twin similarity. RESULTS AND LIMITATIONS: A total of 25 364 twins completed the questionnaire (response rate: 59.6%). LUTS were more common in women (UI: 7%; OAB: 9%; nocturia: 61%; micturition frequency: 18%) than in men (UI: 1%; OAB: 5%; nocturia: 40%; micturition frequency: 11%), and prevalence increased with age. The strongest genetic effects were observed for UI, frequency, and nocturia. The lowest estimate for genetic effects was observed for OAB where environmental effects dominated, and more specifically shared family environment accounted for a third or more of the total variation. For stress UI, a fifth of the total variation in susceptibility to the disorder could be attributed to shared environment. Nonshared environmental effects were seen in the range of 45-65% for the various LUTS. The prevalence of LUTS was low in the men, and there were too few male cases to compute measures of similarity or heritability estimates. CONCLUSIONS: This study provides robust evidence of a genetic influence for susceptibility to UI, frequency, and nocturia in women. In contrast, shared environmental factors seem more important for the predisposition to develop OAB, which may reflect familial patterns such as learning from parental behaviours.

[Familial clustering of symptoms typical for OSAS]. / Rodzinne wystepowanie dolegliwosci charakterystycznych dla obturacyjnego bezdechu podczas snu.

The aim of the study was to compare the incidence of obstructive sleep apnoea syndrome (OSAS) symptoms in relatives of subjects with OSAS and in relatives without OSAS but with clinical symptoms of this disease. The study group consisted of 186 relatives of patients with OSAS and 117 relatives of patients with symptoms of OSAS in whom the disease was not confirmed by polysomnography. They were all mailed a questionnaire with questions concerning anthropometric data, the presence of symptoms typical for OSAS and the presence of concomitant diseases. Analysis of the obtained data revealed an increased frequency of snoring, sleep apnea and nycturia in the relatives of patients with OSAS when compared to relatives of patients without OSAS, but the difference was not statistically significant. The incidence of daytime OSAS symptoms was significantly higher in the group of relatives of patients with OSAS. No differences in the incidence of arterial hypertension, ischaemic heart disease and diabetes mellitus were found.