Age is associated with reduced urethral pressure and afferent activity in rat.

Age-related changes in the lower urinary tract (LUT) can affect the coordination of reflexes and increase the incidence of bladder disorders in elderly. This study examines the age-related loss of urethral signaling capability by measuring the afferent activity directly. We find that less urethral pressure develops in response to fluid flow in old rats compared to young rats and that pressure and flow evoke less urethral afferent activation. These findings are consistent with our previous study demonstrating that the urethra-to-bladder reflex, which is required for efficient voiding, becomes weaker with age. We measured the pudendal afferent response in young (4-7 months) and old (18-24 months) rats to fluid flow in the urethra across a range of flow rates. We used paraffin embedding and hematoxylin and eosin staining to quantify age-related changes in the sensory branch of the pudendal nerve. Urethral afferent signaling in response to the same urethral flow rates was weaker in older animals. That is, the sensitivity of urethra afferents to flow decreased with age, and higher flow rates were required in older animals to recruit urethra afferents. There was also a reduction in the myelin thickness of pudendal afferents in old rats, which is a possible contributing factor to the sensory activity. Furthermore, the same flow rates evoked less pressure in the urethras of old animals, indicating there is an age-related change of the urethral tissue that reduces the pressure stimulus to which these afferents respond. These results help characterize the underlying changes in LUT system with age.

Changes in urethral smooth muscle and external urethral sphincter function with age in rats.

To confirm changes in urethral activity with age, both intravesical pressure and urethral perfusion pressure (UPP) were recorded and external urethral sphincter electromyography (EUS-EMG) was performed. A total of 33 female Sprague Dawley rats aged 3 months (young rats), 12 months (middle-aged rats), and 24 months (aged rats) were used. Bladder activity was evaluated using continuous cystometry. Urethral activity was evaluated by simultaneously recording intravesical pressure and UPP in isovolumetric conditions under urethane anesthesia in each group. Additionally, EUS-EMG activity was monitored under the same conditions. In continuous cystometry, the amplitude of bladder contractions was not different among the three groups; nevertheless, residual urine volume was significantly increased in middle-aged and aged rats, as compared in young rats. With respect to UPP, the change in UPP was significantly smaller in aged rats (60%) and middle-aged rats (64%) than in young rats. Furthermore, the mean amplitude of high-frequency oscillations of the EUS was significantly lower in aged (61%) and middle-aged rats (70%) than in young rats. EUS-EMG revealed EUS bursting activity during voiding with clear active and silent phases in young rats but unclear active and silent phases in aged rats. Masson's trichrome staining of the urethra showed EUS atrophy in aged rats compared to young and middle-aged rats. The results indicate that aging induces two urethral dysfunctions in the urethral smooth muscle and EUS, which may lead to dyscoordination between the urinary bladder and urethra.

Current and Future Directions of Stem Cell Therapy for Bladder Dysfunction.

Stem cells are capable of self-renewal and differentiation into a range of cell types and promote the release of chemokines and progenitor cells necessary for tissue regeneration. Mesenchymal stem cells are multipotent progenitor cells with enhanced proliferation and differentiation capabilities and less tumorigenicity than conventional adult stem cells; these cells are also easier to acquire. Bladder dysfunction is often chronic in nature with limited treatment modalities due to its undetermined pathophysiology. Most treatments focus on symptom alleviation rather than pathognomonic changes repair. The potential of stem cell therapy for bladder dysfunction has been reported in preclinical models for stress urinary incontinence, overactive bladder, detrusor underactivity, and interstitial cystitis/bladder pain syndrome. Despite these findings, however, stem cell therapy is not yet available for clinical use. Only one pilot study on detrusor underactivity and a handful of clinical trials on stress urinary incontinence have reported the effects of stem cell treatment. This limitation may be due to stem cell function loss following ex vivo expansion, poor in vivo engraftment or survival after transplantation, or a lack of understanding of the precise mechanisms of action underlying therapeutic outcomes and in vivo behavior of stem cells administered to target organs. Efficacy comparisons with existing treatment modalities are also needed for the successful clinical application of stem cell therapies. This review describes the current status of stem cell research on treating bladder dysfunction and suggests future directions to facilitate clinical applications of this promising treatment modality, particularly for bladder dysfunction.

Exposure to water-accommodated fractions of two different crude oils alters morphology, cardiac function and swim bladder development in early-life stages of zebrafish.

The present study investigated the developmental toxicity of water-accommodated fractions (WAFs) of Oman crude oil (OCO) and Merey crude oil (MCO) on zebrafish (Danio rerio) in early-life stages (ELS). Based on total petroleum hydrocarbons (TPH), LC50 values manifested that OCO WAF was 1.2-fold more lethal to zebrafish embryos than MCO WAF. As for hatching rate, EC50 value for OCO WAF was 5.7-fold lower than that for MCO WAF. To evaluate the sublethal morphological effects, semi-quantitative extended general morphological score (GMS) and general teratogenic score (GTS) systems were adopted. The GMS and GTS scores indicated that the WAFs caused remarkable developmental delay and high frequencies of malformation in a dose-dependent manner. Additionally, OCO and MCO WAFs exposure exhibited severe bradycardia (reduced heart rate) and overt reduction of stroke volume, with a concomitant decrease in the cardiac output. Meanwhile, the WAFs also induced dose-dependent down-regulated expressions of several key functional genes of excitation-contraction coupling in cardiomyocytes, including ryr2, atp2a2a, atp2a2b, ncx1h, and kcnh2. For key gene markers of swim bladder development, results showed that high dose of TPH induced significant down-regulation of hb9 and anxa5 with no obvious change of acta2, suggesting that the WAFs could affect the specification and development of epithelium and outer mesothelium of swim bladder in zebrafish ELS. A strong negative relationship between the failure of swim bladder inflation and cardiac dysfunction via cardiac output was found. All these findings provide novel insights into the complicated mechanisms of the developmental toxicity of crude oil on fish in ELS.

Mesenchymal stromal cells modulate the molecular pattern of healing process in tissue-engineered urinary bladder: the microarray data.

BACKGROUND: Molecular mechanisms underlying the regenerative process induced by stem cells in tissue-engineered urinary bladder are poorly explained. The study was performed to explore the pathways associated with regeneration process in the urinary bladder reconstructed with adipose tissue-derived mesenchymal stromal cells (ASCs). METHODS: Rat urinary bladders were reconstructed with bladder acellular matrix (BAM) (n = 52) or BAM seeded with adipose tissue-derived mesenchymal stromal cells (ASCs) (n = 52). The process of bladder healing was analyzed at 7, 30, 90, and 180 days postoperatively using macroscopic histologic and molecular techniques. Gene expression was analyzed by microarrays and confirmed by real-time PCR. RESULTS: Numerous differentially expressed genes (DEGs) were identified between the bladders augmented with BAM seeded with ASCs or BAM only. Pathway analysis of DEGs allows to discover numerous pathways among them Hedgehog, TGF-ß, Jak-STAT, PI3-Akt, and Hippo modulated by ASCs during the healing process of tissue-engineered urinary bladder. Real-time PCR analysis confirmed upregulation of genes involved in the Hedgehog signaling pathway including Shh, Gli1, Smo, Bmp2, Bmp4, Wnt2, Wnt2b, Wnt4, Wnt5a, and Wnt10 in urinary bladders reconstructed with ASC-seeded grafts. CONCLUSION: The study provided the unequivocal evidence that ASCs change the molecular pattern of healing in tissue-engineered urinary bladder and indicated which signaling pathways triggered by ASCs can be associated with the regenerative process. These pathways can be used as targets in the future studies on induced urinary bladder regeneration. Of particular interest is the Hedgehog signaling pathway that has been upregulated by ASCs during healing of tissue-engineered urinary bladder.

Development of the human bladder and ureterovesical junction.

The urinary bladder collects urine from the kidneys and stores it until the appropriate moment for voiding. The trigone and ureterovesical junctions are key to bladder function, by allowing one-way passage of urine into the bladder without obstruction. Embryological development of these structures has been studied in multiple animal models as well as humans. In this report we review the existing literature on bladder development and cellular signalling with particular focus on bladder development in humans. The bladder and ureterovesical junction form primarily during the fourth to eighth weeks of gestation, and arise from the primitive urogenital sinus following subdivision of the cloaca. The bladder develops through mesenchymal-epithelial interactions between the endoderm of the urogenital sinus and mesodermal mesenchyme. Key signalling factors in bladder development include shh, TGF-ß, Bmp4, and Fgfr2. A concentration gradient of shh is particularly important in development of bladder musculature, which is vital to bladder function. The ureterovesical junction forms from the interaction between the Wolffian duct and the bladder. The ureteric bud arises from the Wolffian duct and is incorporated into the developing bladder at the trigone. It was previously thought that the trigonal musculature developed primarily from the Wolffian duct, but it has been shown to develop primarily from bladder mesenchyme. Following emergence of the ureters from the Wolffian ducts, extensive epithelial remodelling brings the ureters to their final trigonal positions via vitamin A-induced apoptosis. Perturbation of this process is implicated in clinical obstruction or urine reflux. Congenital malformations include ureteric duplication and bladder exstrophy.

Age-related changes in function and gene expression of the male and female mouse bladder.

We investigated age-related changes in in vivo and in vitro functions and gene expression of the bladder of male and female mice. Mature and aged (12 and 27-30 month old) C57BL/6 mice of both sexes were used. Frequency volume, conscious free-moving cystometry and detrusor contractile and relaxant properties in in vitro organ bath were evaluated. mRNA expression level of muscarinic, purinergic, and ß-adrenergic receptors and gene expression changes by cDNA microarray analysis of the bladder were determined. Cystometry demonstrated storage and voiding dysfunctions with ageing in both sexes. Detrusor strips from aged mice showed weaker contractile responses particularly in the cholinergic component and weaker relaxant responses to isoproterenol. These age-related impairments were generally severer in males. mRNA expression of bladder tissue was decreased for M3 muscarinic receptors in aged males and ß2-adrenoceptors in aged females. cDNA microarray analysis results, albeit substantial sex difference, indicated "cell-to-cell signaling and interaction" as the most common feature of age-related gene expression. In summary, aged mice demonstrated voiding and storage dysfunctions resembling to detrusor hyperactivity with impaired contractility (DHIC), which were more pronounced in males. Genomic changes associated with aging may contribute to the age-related bladder functional deterioration in mice.

Mode of Surgical Injury Influences the Source of Urothelial Progenitors during Bladder Defect Repair.

The bladder urothelium functions as a urine-blood barrier and consists of basal, intermediate, and superficial cell populations. Reconstructive procedures such as augmentation cystoplasty and focal mucosal resection involve localized surgical damage to the bladder wall whereby focal segments of the urothelium and underlying submucosa are respectively removed or replaced and regeneration ensues. We demonstrate using lineage-tracing systems that urothelial regeneration following augmentation cystoplasty with acellular grafts exclusively depends on host keratin 5-expressing basal cells to repopulate all lineages of the de novo urothelium at implant sites. Conversely, repair of focal mucosal defects not only employs this mechanism, but in parallel host intermediate cell daughters expressing uroplakin 2 give rise to themselves and are also contributors to superficial cells in neotissues. These results highlight the diversity of urothelial regenerative responses to surgical injury and may lead to advancements in bladder tissue engineering approaches.

Bladder regeneration in a canine model using a bladder acellular matrix loaded with a collagen-binding bFGF.

Bladder reconstruction remains challenging for urological surgery due to lack of suitable regenerative scaffolds. In a previous study, we had used a collagen-binding basic fibroblast growth factor (CBD-bFGF) to bind bFGF to the collagen scaffold, which could promote bladder regeneration in rats. However, the limited graft size in rodent models cannot provide enough evidence to demonstrate the repair capabilities of this method for severely damaged bladders in humans or large animals. In this study, the CBD-bFGF was used to activate a bladder acellular matrix (BAM) scaffold, and the CBD-bFGF/BAM functional scaffold was assessed in a canine model with a large segment defect (half of the entire bladder was resected). The results demonstrated that the functional biomaterials could promote bladder smooth muscle, vascular, and nerve regeneration and improve the function of neobladders. Thus, the CBD-bFGF/BAM functional scaffold may be a promising biomaterial for bladder reconstruction.

[PATHOGENICALLY INDUCED APOPTOSIS CAUSED BY HYPOXIC EFFECTS IN THE URINARY SYSTEM ORGANS OF FETUSES AND NEWBORNS (EXPERIMENTAL STUDY)].

The purpose of the study was to identify the characteristics of apoptosis in the kidneys, ureters and bladder of fetuses and newborns in the modeling of chronic intrauterine hypoxia, acute postnatal hypoxia and mixed hypoxia. An experiment was conducted on WAG rats for modeling high altitude hypoxia. Experimental animals were divided into four groups: I - control - fetuses and newborns from healthy rats; II - modeling of chronic intrauterine hypoxia; III - modeling of acute postnatal hypoxia; IV - modeling of mixed hypoxia. The material of the study was the tissue of the kidneys, ureters and bladder of fetuses and newborns. In group I in the kidneys of fetuses the mean value of the number of p53-positive cells was 7.83±0.31, newborns - 5.40±0.28; in the ureters and bladder of fetuses - 5.77±0.29 and 6.97±0.32, newborns - 3.58±0.21 and 5.36±0.28. In the kidneys in group II the mean value of the number of p53-expressing cells in fetuses was 1.43±0.50, in newborns - 21.72±0.58; in group III in newborns - 15.03±0.63; in group IV in newborns - 33.33±0.72. The mean value of the number of p53-expressing cells in the ureters and bladder in group II in fetuses was 13.17±0.49 and 11.83±0.43, in newborns - 16.24±0.37 and 15.38±0.37; in group III in newborns - 7.25±0.27 and 8.68±0.32; in group IV in newborns - 19.63±0.31and 21.03±0.40. As the result of the study it was found that experimental hypoxia induced apoptotic processes in the kidneys, ureters and bladder of fetuses and newborns, the severity of which was moderate in the modeling of acute postnatal hypoxia, expressed in the modeling of chronic intrauterine hypoxia and strongly expressed in the modeling of mixed hypoxia. Under the influence of acute postnatal hypoxia, chronic intrauterine hypoxia and mixed hypoxia in the ureters and bladder of fetuses and newborns p53-positive cells were located evenly in all layers of the wall of these organs, whereas in the kidneys p53-positive cells prevailed in the tubular component. In the modeling of chronic intrauterine hypoxia apoptotic processes in the kidneys, ureters and bladder increased in newborns in comparison with fetuses.

Developmental changes in contractile responses to cholinergic stimuli: role of calcium sensitization and related pathways.

This study was performed to analyze the developmental changes in bladder response to cholinergic stimulation in detail, highlighting calcium sensitization (CS) and its related pathways. Rats were divided into three groups in accordance with reported time of developmental milestones (newborns, days 1-4; youngsters, days 5-14; and grown-ups, days 15-28). Following cholinergic stimulation (carbachol, 5 µM), the contractile response to detrusor was analyzed with respect to three phases (initial phasic, tonic, and superimposed phasic contractions). Contractile responses were analyzed by their dynamic and kinetic aspects. The responses were further compared in varying external calcium concentrations and in the presence of inhibitors of protein kinase C (PKC) and Rho kinase (ROCK), which are involved in CS. The responses of newborns contrasted with the others by their short and brisk initial phasic contractions, prominent tonic contractions, and delayed participation of irregular superimposed phasic contractions. With development, phasic contractions became prominent, and tonic contractions diminished. These developmental changes in phasic contractions were reproduced when exposed to increasing calcium concentrations. Application of specific inhibitors and molecular phasic analysis revealed that PKC was functional in tonic contractions of the newborns, whereas ROCK took over its role with development. Within a few days of birth, rats' bladders experienced drastic changes in contractile mechanisms. This included dominance of phasic contractions over tonic contractions due to increased calcium dependence and the maturational shift of the calcium sensitivity mechanism from PKC to ROCK.

Stage- and subunit-specific functions of polycomb repressive complex 2 in bladder urothelial formation and regeneration.

Urothelium is the protective lining of the urinary tract. The mechanisms underlying urothelial formation and maintenance are largely unknown. Here, we report the stage-specific roles of PRC2 epigenetic regulators in embryonic and adult urothelial progenitors. Without Eed, the obligatory subunit of PRC2, embryonic urothelial progenitors demonstrate reduced proliferation with concomitant dysregulation of genes including Cdkn2a (p16), Cdkn2b (p15) and Shh. These mutants display premature differentiation of keratin 5-positive (Krt5+) basal cells and ectopic expression of squamous-like differentiation markers. Deletion of Ezh2, the major enzymatic component of PRC2, causes upregulation of Upk3a+ superficial cells. Unexpectedly, Eed and Eed/Ezh2 double mutants exhibit delayed superficial cell differentiation. Furthermore, Eed regulates the proliferative and regenerative capacity of adult urothelial progenitors and prevents precocious differentiation. Collectively, these findings uncover the epigenetic mechanism by which PRC2 controls urothelial progenitor cell fate and the timing of differentiation, and further suggest an epigenetic basis of urothelial maintenance and regeneration.

Embryonic kidney function in a chronic renal failure model in rodents.

BACKGROUND: Rapid advancements have been made in alternative treatments for renal diseases. Our goal for renal regeneration is to establish a kidney graft derived from human embryonic tissues. In this study, we investigated the effects of host renal failure on the structure and activity of transplanted embryonic kidney and bladder, and found that diuretics effectively induced urine production in the transplanted kidney. METHODS: Uremic conditions were reproduced using a 5/6 renal infarction rat model. An embryonic kidney plus bladder (embryonic day 15) was isolated from a pregnant Lewis rat and transplanted into the para-aortic area of a 5/6 renal-infarcted Lewis rat. Following growth, the embryonic bladder was successfully anastomosed to the host ureter. RESULTS: We assessed graft function in terms of survival rates and found no differences between normal (n = 5) and renal failure (n = 8) groups (median survival: 70.5 vs 74.5 h; p = 0.331) in terms of survival, indicating that the grafts prolonged rat survival, even under renal failure conditions. Furosemide (n = 9) significantly increased urine volume compared with saline-treated controls (n = 7; p < 0.05), confirming that the grafts were functional. We also demonstrated the possibilities of an in vivo imaging system for determining the viability of transplanted embryonic kidney with bladder. CONCLUSION: The results of this study demonstrate that transplanted embryonic kidney and bladder can grow and function effectively, even under uremic conditions.

Hydronephrosis in the Wnt5a-ablated kidney is caused by an abnormal ureter-bladder connection.

The Wnt5a null mouse is a complex developmental model which, among its several posterior-localized axis defects, exhibits multiple kidney phenotypes, including duplex kidney and loss of the medullary zone. We previously reported that ablation of Wnt5a in nascent mesoderm causes duplex kidney formation as a result of aberrant development of the nephric duct and abnormal extension of intermediate mesoderm. However, these mice also display a loss of the medullary region late in gestation. We have now genetically isolated duplex kidney formation from the medullary defect by specifically targeting the progenitors for both the ureteric bud and metanephric mesenchyme. The conditional mutants fail to form a normal renal medulla but no longer exhibit duplex kidney formation. Approximately 1/3 of the mutants develop hydronephrosis in the kidneys either uni- or bilaterally when using Dll1Cre. The abnormal kidney phenotype becomes prominent at E16.5, which approximates the time when urine production begins in the mouse embryonic kidney, and is associated with a dramatic increase in apoptosis only in mutant kidneys with hydronephrosis. Methylene blue dye injection and histologic examination reveal that aberrant cell death likely results from urine toxicity due to an abnormal ureter-bladder connection. This study shows that Wnt5a is not required for development of the renal medulla and that loss of the renal medullary region in the Wnt5a-deleted kidney is caused by an abnormal ureter-bladder connection.

Evidence of vagus nerve sprouting to innervate the urinary bladder and clitoris in a canine model of lower motoneuron lesioned bladder.

AIMS: Complete spinal cord injury does not block perceptual responses or inferior solitary nucleus activation after genital self-stimulation, even though the vagus is not thought to innervate pelvic structures. We tested if vagus nerve endings sprout after bladder decentralization to innervate genitourinary structures in canines with decentralized bladders. METHODS: Four reinnervation surgeries were performed in female hounds: bilateral genitofemoral nerve transfer to pelvic nerve with vesicostomy (GNF-V) or without (GFN-NV); and left femoral nerve transfer (FNT-V and FNT-NV). After 8 months, retrograde dyes were injected into genitourinary structures. Three weeks later, at euthanasia, reinnervation was evaluated as increased detrusor pressure induced by functional electrical stimulation (FES). Controls included un-operated, sham-operated, and decentralized animals. RESULTS: Increased detrusor pressure was seen in 8/12 GFNT-V, 4/5 GFNT-NV, 5/5 FNT-V, and 4/5 FNT-NV animals after FES, but not decentralized controls. Lumbar cord segments contained cells labeled from the bladder in all nerve transfer animals with FES-induced increased detrusor pressure. Nodose ganglia cells labeled from the bladder were observed in 5/7 nerve transfer animals (1/2 GNT-NV; 4/5 FNT-V), and from the clitoris were in 6/7 nerve transfer animals (2/2 GFNT-NV; 4/5 FNT-V). Dorsal motor nucleus vagus cells labeled from the bladder were observed in 3/5 nerve transfer animals (1/2 GFNT-NV; 2/3 FNT-V), and from the clitoris in 4/5 nerve transfer animals (1/2 GFNT-NV; 3/3 FNT-V). Controls lacked this labeling. CONCLUSIONS: Evidence of vagal nerve sprouting to the bladder and clitoris was observed in canines with lower motoneuron lesioned bladders. Neurourol. Urodynam. 36:91-97, 2017. © 2015 Wiley Periodicals, Inc.

Fibroblast growth factor receptor signaling in kidney and lower urinary tract development.

Fibroblast growth factor receptors (FGFRs) and FGF ligands are highly expressed in the developing kidney and lower urinary tract. Several classic studies showed many effects of exogenous FGF ligands on embryonic renal tissues in vitro and in vivo. Another older landmark publication showed that mice with a dominant negative Fgfr fragment had severe renal dysplasia. Together, these studies revealed the importance of FGFR signaling in kidney and lower urinary tract development. With the advent of modern gene targeting techniques, including conditional knockout approaches, several publications have revealed critical roles for FGFR signaling in many lineages of the kidney and lower urinary tract at different stages of development. FGFR signaling has been shown to be critical for early metanephric mesenchymal patterning, Wolffian duct patterning including induction of the ureteric bud, ureteric bud branching morphogenesis, nephron progenitor survival and nephrogenesis, and bladder mesenchyme patterning. FGFRs pattern these tissues by interacting with many other growth factor signaling pathways. Moreover, the many genetic Fgfr and Fgf animal models have structural defects mimicking numerous congenital anomalies of the kidney and urinary tract seen in humans. Finally, many studies have shown how FGFR signaling is critical for kidney and lower urinary tract patterning in humans.

Age-related changes in bladder function with altered angiotensin II receptor mechanisms in rats.

AIMS: To examine alterations in expression of angiotensin II type 1 receptors (AT1R) which induce organ tissue remodeling, angiotensin II type 2 receptors (AT2R) which protect against it, and related molecules in the bladder of matured rats with bladder dysfunction. METHODS: Female SD rats of three different ages were used: 8 weeks old (8W; n = 5), 9 months old (9M; n = 5), and 15 months old (15M; n = 5). After cystometry, the expression levels of AT1R, connexin43 (Cx43), MAP kinase (MAPK), collagen1, AT2R, PPAR-γ, adiponectin (Adipo), and adiponectin receptor (Adipo-R) were investigated in the bladder. RESULTS: Pressure threshold, post-void residual volume and the number of non-voiding contractions were significantly increased in 15M versus 8W rats (P < 0.01). Maximum voiding pressure was significantly decreased in 15M versus 8W rats (P < 0.05). There was no significant difference in CMG parameters between 8W and 9M rats. In the bladder, the mRNA expression of AT1R, Cx43, MAPK, collagen 1, AT2R, PPAR-γ, Adipo, and Adipo-R were significantly higher in 15M than in 8W rats. The relative expression ratio of AT1R protein against AT2R protein in the mucosa and detrusor was significantly increased in 15M versus 8W rats. CONCLUSIONS: These results indicate that matured rats exhibit not only bladder overactivity but also impaired voiding, which are associated with upregulation of AT1R. The upregulation of AT2R also may play a significant role in the suppressing of AT1R induced remodelling. However, because AT1R upregulation is more dominant than AT2R increases, AT2R activation may not be sufficient to suppress AT1R stimulation in matured rats. Neurourol. Urodynam. 35:908-913, 2016. © 2015 Wiley Periodicals, Inc.

Time-dependent bladder tissue regeneration using bilayer bladder acellular matrix graft-silk fibroin scaffolds in a rat bladder augmentation model.

With advances in tissue engineering, various synthetic and natural biomaterials have been widely used in tissue regeneration of the urinary bladder in rat models. However, reconstructive procedures remain insufficient due to the lack of appropriate scaffolding, which should provide a waterproof barrier function and support the needs of various cell types. To address these problems, we have developed a bilayer scaffold comprising a porous network (silk fibroin [SF]) and an underlying natural acellular matrix (bladder acellular matrix graft [BAMG]) and evaluated its feasibility and potential for bladder regeneration in a rat bladder augmentation model. Histological (hematoxylin and eosin and Masson's trichrome staining) and immunohistochemical analyses demonstrated that the bilayer BAMG-SF scaffold promoted smooth muscle, blood vessel, and nerve regeneration in a time-dependent manner. At 12weeks after implantation, bladders reconstructed with the BAMG-SF matrix displayed superior structural and functional properties without significant local tissue responses or systemic toxicity. These results demonstrated that the bilayer BAMG-SF scaffold may be a promising scaffold with good biocompatibility for bladder regeneration in the rat bladder augmentation model.

Development of seahorse (Hippocampus reidi, Ginsburg 1933): histological and histochemical study.

Biological aspects and global demand for aquarium promote seahorses as new species with high potential for commercial purposes; however, the low newborn survival rate represents the main bottleneck of seahorses farming. In this study, the organogenesis of the Hippocampus reidi was analysed from release until the 30th day after birth, using histological and histochemical approaches. To study the stages of their early life, 360 individuals were killed, sectioned, and stained with haematoxylin and eosin, periodic acid-Schiff, and Sudan Black B techniques. At birth, mouth and anus were open, the swim bladder inflated, and the visual system highly developed. Among the results, it was emphasized the presence of the yolk sac until the 2nd day after birth, the loops of the intestine to accommodate its elongation, and the ability of the larvae to absorb lipids in the anterior and posterior tract of the intestine. A short time (7/8 days) between reabsorption of yolk sac and formation of gonads was registered, with primordial follicles visible from the 10th day after birth. For the first time, organogenesis in H. reidi was described in detail; seahorses underwent a marked metamorphosis, and the indirect development observed in this species lead up to reconsider the term "juvenile" used for H. reidi during this period.

Bladder function development and its urodynamic evaluation in neonates and infants less than 2 years old.

AIMS: To understand the function development of bladder and its evaluation in neonates and infants less than 2 years old. METHODS: Literature on neonatal and infant bladder function development and urodynamic evaluation were collected and reviewed. RESULTS: Normal range of bladder volume, pressure during voiding and other parameters in neonates and infants less than 2 years old is far from set up, making interpretation of UDS findings difficult. This review provides insight into the bladder development process and problems of the lower urinary tract in this age group with special emphasis on the urodynamic evaluation. CONCLUSIONS: Further animal and human studies will increase our understanding of bladder development leading toward mature function. UDS are still important in providing information for early bladder dysfunction in newborns and infants.