Exploring the Role of miR-132 in Rat Bladders and Human Urothelial Cells during Wound Healing.

Urinary bladder wound healing shares many features with skin healing, involving several molecular players, including microRNAs (miRs). This study investigated the role of miR-132 in urothelial cells. We analyzed miR-132 expression in rat bladder using in situ hybridization and conducted gain and loss of miR-132 function assays in primary human urothelial cells (HUCs). These assays included cell proliferation and migration studies. To explore the regulation of miR-132 expression, cells were treated with wound-healing-related factors such as interleukin 6 (IL-6), interleukin 10 (IL-10), and transforming growth factor beta-1 (TGF-ß1). Predictive bioinformatics and a literature review identified potential miR-132 targets, which were validated through real-time polymerase chain reaction (RT-PCR) and Western blot analysis. miR-132 was found to promote cellular proliferation and migration during the early stages of urothelial wound repair. Its expression was modulated by key cytokines such as IL-6, IL-10, and TGF-ß1. miR-132 played a crucial role in urothelial wound healing by enhancing cell proliferation and migration, regulated by cytokines, suggesting its action within a complex regulatory network. These findings highlight the therapeutic potential of targeting miR-132 in bladder injury repair, offering new insights into bladder repair mechanisms.

Copy number variants suggest different molecular pathways for the pathogenesis of bladder exstrophy.

Bladder exstrophy is a rare congenital malformation leaving the urinary bladder open in the midline of the abdomen at birth. There is a clear genetic background with chromosome aberrations, but so far, no consistent findings apart from 22q11-duplications detected in about 2%-3% of all patients. Some genes are implicated like the LZTR1, ISL1, CELSR3, and the WNT3 genes, but most are not explained molecularly. We have performed chromosomal microarray analysis on a cohort of 140 persons born with bladder exstrophy to look for submicroscopic chromosomal deletions and duplications. Pathogenic or possibly pathogenic microdeletions or duplications were found in 16 patients (11.4%) and further 9 with unknown significance. Two findings were in regions linked to known syndromes, two findings involved the same gene (MCC), and all other findings were unique. A closer analysis suggests a few gene networks that are involved in the pathogenesis of bladder exstrophy; the WNT-signaling pathway, the chromosome 22q11 region, the RIT2 and POU families, and involvement of the Golgi apparatus. Bladder exstrophy is a rare malformation and is reported to be associated with several chromosome aberrations. Our data suggest involvement of some specific molecular pathways.

Exploring the Concept of In Vivo Guided Tissue Engineering by a Single-Stage Surgical Procedure in a Rodent Model.

In severe malformations with a lack of native tissues, treatment options are limited. We aimed at expanding tissue in vivo using the body as a bioreactor and developing a sustainable single-staged procedure for autologous tissue reconstruction in malformation surgery. Autologous micro-epithelium from skin was integrated with plastically compressed collagen and a degradable knitted fabric mesh. Sixty-three scaffolds were implanted in nine rats for histological and mechanical analyses, up to 4 weeks after transplantation. Tissue integration, cell expansion, proliferation, inflammation, strength, and elasticity were evaluated over time in vivo and validated in vitro in a bladder wound healing model. After 5 days in vivo, we observed keratinocyte proliferation on top of the transplant, remodeling of the collagen, and neovascularization within the transplant. At 4 weeks, all transplants were fully integrated with the surrounding tissue. Tensile strength and elasticity were retained during the whole study period. In the in vitro models, a multilayered epithelium covered the defect after 4 weeks. Autologous micro-epithelial transplants allowed for cell expansion and reorganization in vivo without conventional pre-operative in vitro cell propagation. The method was easy to perform and did not require handling outside the operating theater.

Fertility Potential is Impaired in Boys with Bilateral Ascending Testes.

PURPOSE: Ascending testes have been documented to be descended in the scrotum within the first year of life and then reascended. The aim of this study was to investigate to what extent the fertility potential was impaired in boys with such testes compared to the fertility potential of boys with late referral congenital cryptorchidism. MATERIALS AND METHODS: A total of 153 consecutive boys underwent bilateral orchiopexy at age 2 to 7 years (median 3.9) between 2011 and 2018. Of the patients 67 were diagnosed with bilateral ascended testes and 86 with late referral bilateral congenital cryptorchidism. We assessed serum levels of inhibin B and gonadotropins and histological parameters, number of germ cells per tubule cross-section and number of type A dark (Ad) spermatogonia per tubule cross-section. All values were compared to our normal material. RESULTS: Number of germ cells per tubule cross-section of boys with ascended testes (median 0.50, range 0 to 2.29) was not significantly higher compared to boys with congenital cryptorchidism (median 0.37, range 0 to 2.57; p=0.11). Mean number of germ cells per tubule cross-section was below normal range in 40 boys with ascending testes (60%) vs 57 boys with late referral congenital cryptorchidism (66%, p=0.40). Biopsies absent of Ad spermatogonia were noted in 31% of boys with ascending testes (21 of 67) vs 34% of boys with congenital cryptorchidism (29 of 86, p=0.76). Serum levels of inhibin B and gonadotropins did not differ between the 2 groups. CONCLUSIONS: The fertility potential of boys with bilateral ascended testes was impaired to almost the same level as that of boys with bilateral congenital cryptorchidism and should therefore be surgically corrected as soon as the diagnosis of ascended testes is settled.

The impact of early and successful orchidopexy on hormonal follow-up for 208 boys with bilateral non-syndromic cryptorchidism.

PURPOSE: Inhibin-B is produced by Sertoli cells and decreased values might be associated with impaired fertility potential. The aim of the study was to evaluate the impact of bilateral orchidopexy on serum inhibin-B and follicle-stimulating hormone (FSH). METHODS: A cohort study including 208 bilateral cryptorchid boys (median age: 1.7 year) was evaluated with serum inhibin-B and FSH in relation to histological parameters. Based on the fertility potential, the boys were divided into three subgroups. At follow-up (median age: 2.7 years) the boys were evaluated with FSH and in case of inhibin-B using multiple of the median (MoM). RESULTS: Inhibin-B MoM improved significantly at follow-up. In 32 boys with high FSH at orchidopexy 63% normalized FSH and 59% increased MoM inhibin-B, but 31% had impaired inhibin-B at follow-up. In 105 boys with transient hypogonadotropic hypogonadism, 52% increased inhibin-B MoM but 31% had impaired inhibin-B at follow-up. In 71 boys with normal FSH, inhibin-B, and G/T, 54% increased inhibin-B MoM and 15% had impaired inhibin-B at follow-up. The effect of the surgery was best in patients younger than 1 year. CONCLUSION: Orchidopexy, especially before 1 year of age, improves the fertility potential in bilateral cryptorchidism. At follow-up, 26% (54/208) had a risk of infertility based on inhibin-B.

Fertility Potential is Compromised in 20% to 25% of Boys with Nonsyndromic Cryptorchidism despite Orchiopexy within the First Year of Life.

PURPOSE: One of the concerns surrounding cryptorchidism is the risk of impaired fertility. Current guidelines recommend orchiopexy at age 6 to 12 months to optimize fertility outcome. We evaluated the fertility potential of boys with nonsyndromic cryptorchidism who underwent orchiopexy within the recommended age range to clarify the need for eventual supplemental treatment modalities. MATERIALS AND METHODS: We retrospectively evaluated mini-puberty hormones (follicle-stimulating hormone, luteinizing hormone and inhibin B) and testicular biopsies from boys with cryptorchidism who underwent orchiopexy within the first year of life between 2010 and 2019. We histologically analyzed germ cell number and type A dark spermatogonia number per seminiferous tubule cross-section in relation to normal values. RESULTS: Of the 333 boys with nonsyndromic cryptorchidism 83 (25%, 21% with bilateral cryptorchidism) had a reduced number of germ cells. A total of 70 boys (21%) had low serum inhibin B, of whom 32 (46%) had a decreased number of germ cells and 23 (33%) had a decreased number of type A dark spermatogonia (p <0.01). Overall, 75 boys (23%) had no type A dark spermatogonia present. CONCLUSIONS: Despite early and successful orchiopexy, 20% to 25% of boys with cryptorchidism may be at risk for infertility based on hormonal and histological data. Blood test and testicular biopsy are mandatory to identify boys at high risk for infertility, in whom additional treatment modalities and followup may be needed.

Molecular and histological studies of bladder wound healing in a rodent model.

The field of regenerative medicine encounters different challenges. The success of tissue-engineered implants is dependent on proper wound healing. Today, the process of normal urinary bladder wound healing is poorly characterized. We aspired to explore and elucidate the natural response to injury in an in vivo model in order to further optimize tissue regeneration in future studies. In this study, we aimed to characterize histological and molecular changes during normal healing in a rat model by performing a standardized incisional wound followed by surgical closure. We used a rodent model (n = 40) to follow the healing process in the urinary bladder for 28 days. Surgical exposure of the bladder without incision (n = 40) was performed in controls. Histological characterization and western blot analyses of proteins was carried out using specific staining and markers for inflammation, proliferation, angiogenesis, and tissue maturation. For the molecular characterization of gene expression total RNA was collected for RT2 -PCR in wound healing pathway arrays. Analysis of histology revealed distinct, but overlapping, phases of healing with a local inflammatory response (days 1-8) simultaneous with a rapid formation of granulation tissue and proliferation (days 2-8). We also identified significant changes in gene expression related to inflammation, proliferation, and extracellular matrix formation. Healing of an incisional wound in a rodent urinary bladder demonstrated that all the classical phases of wound healing: hemostasis, inflammation, proliferation followed by tissue maturation were present. Our data suggest that the bladder and the skin share similar molecular signaling during wound healing, although we noted differences in the duration of each phase compared to previous studies in rat skin. Further studies will address whether our findings can be extrapolated to the human bladder.

Changes in arterial pressure and markers of nitric oxide homeostasis and oxidative stress following surgical correction of hydronephrosis in children.

OBJECTIVE: Recent clinical studies have suggested an increased risk of elevated arterial pressure in patients with hydronephrosis. Animals with experimentally induced hydronephrosis develop hypertension, which is correlated to the degree of obstruction and increased oxidative stress. In this prospective study we investigated changes in arterial pressure, oxidative stress, and nitric oxide (NO) homeostasis following correction of hydronephrosis. METHODS: Ambulatory arterial pressure (24 h) was monitored in pediatric patients with hydronephrosis (n = 15) before and after surgical correction, and the measurements were compared with arterial pressure measurements in two control groups, i.e. healthy controls (n = 8) and operated controls (n = 8). Markers of oxidative stress and NO homeostasis were analyzed in matched urine and plasma samples. RESULTS: The preoperative mean arterial pressure was significantly higher in hydronephrotic patients [83 mmHg; 95% confidence interval (CI) 80-88 mmHg] than in healthy controls (74 mmHg; 95% CI 68-80 mmHg; p < 0.05), and surgical correction of ureteral obstruction reduced arterial pressure (76 mmHg; 95% CI 74-79 mmHg; p < 0.05). Markers of oxidative stress (i.e., 11-dehydroTXB2, PGF2α, 8-iso-PGF2α, 8,12-iso-iPF2α-VI) were significantly increased (p < 0.05) in patients with hydronephrosis compared with both control groups, and these were reduced following surgery (p < 0.05). Interestingly, there was a trend for increased NO synthase activity and signaling in hydronephrosis, which may indicate compensatory mechanism(s). CONCLUSION: This study demonstrates increased arterial pressure and oxidative stress in children with hydronephrosis compared with healthy controls, which can be restored to normal levels by surgical correction of the obstruction. Once reference data on ambulatory blood pressure in this young age group become available, we hope cut-off values can be defined for deciding whether or not to correct hydronephrosis surgically.

Advantages of Reduced Prophylaxis after Tubularized Incised Plate Repair of Hypospadias.

PURPOSE: Concerns about antibiotic resistance, adverse drug reactions and questionable medical benefits have led to changes in prophylactic antibiotic management in hypospadias repair at our clinic. In March 2010 our guidelines were changed from continuous prophylaxis for 14 days to 1 dose preoperatively and another at removal of the stent. We analyze the effects of this new regimen. MATERIALS AND METHODS: We performed a prospective journal cohort study of all our hypospadias operations from June 2008 to December 2011. We collected data from consecutive patients undergoing primary tubularized incised plate repair and postoperative stent. Patients operated on before March 2010 were compared to those operated on later. End points were postoperative infection requiring antibiotics and any complication that required redo surgery. RESULTS: The study included 113 primary tubularized incised plate repairs with postoperative stents. Patient distribution was the same in both groups. Of 58 patients in the group receiving continuous antibiotic prophylaxis 17 had a complication and/or infection, compared to 9 of 55 patients receiving 2-dose prophylaxis. The infection rate was 5% in the continuous prophylaxis group and 4% in the 2-dose group. In contrast to our expectations, a lower complication rate was observed in the group with lower antibiotic dose without an increased risk of infection. CONCLUSIONS: There is little documented evidence concerning benefits of antibiotic prophylaxis for postoperative complications, which gives rise to large variations in clinical practice. In our study lower antibiotic dose did not increase the number of infections, but rather decreased complication rates. We advocate antibiotic prophylaxis with only a 2-dose regimen.

Long-term followup of men born with hypospadias: urological and cosmetic results.

PURPOSE: We present data on long-term functional and cosmetic results after hypospadias surgery. MATERIALS AND METHODS: Males older than 18 years with hypospadias treated in Sweden were asked to participate in the study, as well as age matched controls and circumcised men. All participants answered questionnaires, and a subgroup was examined during an outpatient visit. Relationships with outcome were analyzed using analysis of variance and regression analysis. RESULTS: A total of 167 patients with a mean age of 34 years and 169 controls with a mean age of 33 years answered the questionnaire. Of the patients 63% had distal, 24% mid and 13% proximal hypospadias. A total of 46 patients and 49 controls presented for physical examination. Patients were significantly less satisfied with the penile cosmetic outcome regarding all parameters of the Penile Perception Score. There was a difference in penile length between patients and controls (mean 9.7 vs 11.6 cm, p <0.001). More patients than controls reported voiding dysfunction symptoms (p = 0.003). Patients had a lower maximum urinary flow rate than controls (p = 0.001). These differences were most prominent between patients with proximal hypospadias and controls. CONCLUSIONS: Men operated on for hypospadias were less satisfied with the cosmetic result than controls, and had a shorter penile length. Patients presented with more symptoms of voiding dysfunction and displayed a lower maximum urinary flow rate. Patients with proximal hypospadias were more affected than those with milder hypospadias. Our results indicate that patients with hypospadias can be subgrouped and that those with severe phenotypes should be followed more closely during childhood as well as later in adulthood.

Autologous micrografting improves regeneration of tissue-engineered urinary conduits in vivo.

Urogenital reconstructive malformation surgery is sometimes hampered by lack of tissue for the repair. We have previously shown that autologous micrografting allows for single-staged scaffold cellularization after surgical implantation. Here, a collagen-based scaffold reinforced with biodegradable mesh and a stent was implanted as a bladder conduit in ten full-grown female minipigs. We aimed to assess short-term regenerative outcomes, safety, and feasibility of implanting tubular urinary micrografted scaffolds versus acellular controls. Five scaffolds were embedded with autologous urothelial micrografts harvested perioperatively. After six weeks, all animals were assessed by cystoscopy, CT-urography, and microanatomical assessment of the urinary conduits. The procedure proved technically feasible within the confines of a regular surgical theater, with duration-times comparable to corresponding conventional procedures. No animals experienced postoperative complications, and all implanted conduits were patent at follow-up. Improved tissue regeneration was observed in the micrografted conduits compared with the acellular controls, including increased luminal epithelialization, increased cell proliferation, decreased cell apoptosis, and increased conduit vascularization. We concluded that single-staged on-site construction and implantation of tissue engineered urinary conduits proved feasible and safe, with improved regenerative potentials in micrografted conduits. This study presents a new approach to urinary conduits, and merits further investigations for advancement towards clinical translation.

Surgical Implantation of Single-Staged Tissue-Engineered Urothelial Tubes in a Minipig Model.

Reconstructive surgeries are often challenged by a lack of grafting tissue. In the treatment of urogenital malformations, the conventional solution has been harvesting gastrointestinal tissue for non-orthotopic reconstruction due to its abundance to reestablish normal function in the patient. The clinical outcomes after rearranging native tissues within the body are often associated with significant morbidity; thus, tissue engineering holds specific potential within this field of surgery. Despite substantial advances, tissue-engineered scaffolds have not yet been established as a valid surgical treatment alternative, mainly due to the costly and complex requirements of materials, production, and implantation. In this protocol, we present a simple and accessible collagen-based tubular scaffold embedded with autologous organ-specific tissue particles, designed as a conduit for urinary diversion. The scaffold is constructed during the primary surgical procedure, comprises commonly available surgical materials, and requires conventional surgical skills. Secondly, the protocol describes an animal model designed to evaluate the short-term in vivo outcomes post-implantation, with the possibility of additional variations to the procedure. This publication aims to demonstrate the procedure step-by-step, with special attention to the use of autologous tissue and a tubular form.

A qualitative content analysis of the experience of living with hypospadias: varying impact on identity and interpersonal relationships.

Objectives: There is a lack of in-depth studies on men's personal experiences of having hypospadias across different aspects of their lives. We therefore aimed to explore the experience of having hypospadias in relation to identity and interpersonal relationships. Subjects and methods: Using purposive sampling, we included 17 adult men aged 20-49 with variation in hypospadias phenotype. The informants further represented variation in sexuality, relationship status, parental status, and familial cultural context. In-depth interviews were conducted with each informant and the data was analysed using qualitative content analysis. Results: We identified four categories. Firstly, The internal experience of hypospadias in relation to being different, being impacted, and being masculine. The remaining three categories related to interpersonal spaces: Intimate spaces, comprising personal relationship with sex, having sex, and being in a relationship; Familial spaces, comprising being a son, and becoming a father; and Public spaces, comprising being hidden, being naked, and peeing. We identified the latent theme varying impact and coping, highlighting differences in experiences relating to both the internal and interpersonal. Discussion: Issues related to hypospadias included struggles with identity and confidence, as well as recurring patterns of social and sexual avoidance. While informants generally related to certain shared experiences, there is large variation in how much hypospadias impacts life, ranging from hardly at all to extensively. This could also fluctuate over time, with puberty and adolescence being an especially sensitive period. Functional and aesthetic outcomes are potentially important for well-being, especially in the case of more severe complications, while personal and interpersonal circumstances play a role in coping and the overall experience of the individual. Conclusion: Healthcare, research, and other channels such as patient groups may be able to offer support to those who need it to help more boys and men with hypospadias live unhindered lives.

Fetal biomarkers for lower urinary tract obstruction secondary to posterior urethral valves.

Today, prenatal diagnosis of congenital urogenital malformations is mostly dependent on anatomical variations found on imaging. However, these findings can mislead us in telling us when to intervene, and about post-natal prognosis. Since many findings are dependent on multiple assessments, delayed diagnosis can occur, leading to less optimal outcomes compared to early intervention. Analyses of fetal urinary biomarkers have been proposed as a method of finding biological changes that are predictive for diagnosis and prognosis in fetuses at risk of kidney disease. We interviewed a group of researchers that have demonstrated that by combining multiple omics traits extracted from fetal urine, the biological variability found in single omics data can be circumvented. By analyzing multiple fetal urine peptides and metabolites at single time point, the prognostic power of postnatal renal outcome in fetuses with lower urinary tract obstruction is significantly increased. In this interview, we inquired about the technical aspects of the tests, challenges, and limitations the research group have come across, and how they envision the future for multi-omics fetal analysis in the clinic.

A perioperative layered autologous tissue expansion graft for hollow organ repair.

Tissue engineering has not been widely adopted in clinical settings for several reasons, including technical challenges, high costs, and regulatory complexity. Here, we introduce the Perioperative Layered Autologous Tissue Expansion graft (PLATE graft), a composite biomaterial and collagen-reinforced construct with autologous epithelium on one side and smooth muscle tissue on the other. Designed to mimic the structure and function of natural hollow organs, the PLATE graft is unique in that it can be produced in a standard operating theatre and is cost-effective. In this proof-of-principle study, we test its regenerative performance in eight different organs, present biomechanical and permeability tests, and finally explore its in vivo performance in live rabbits.

The importance of animal specificity in animal experimentation, part I: Anatomy in relation to pediatric urology.

Why and when is animal experimentation relevant? The answer to this question depends on the research question. In this short educational article we aim to raise awareness of the importance of formulating a very specific research question before choosing an animal species. An awareness of anatomical and physiological differences vis-a-vis similarities between species, will increase the potential for obtaining data that is relevant for translation to human conditions.

The importance of animal specificity in animal experimentation, part II: Physiological challenges and opportunities in relation to pediatric urology.

When performing animal experimentation in Pediatric Urology studies, it is important to be aware of physiological differences between species and to understand when relevant disease models are available. Diseased animal models may be more relevant in many cases, rather than performing studies in healthy and normally developed animals. For example, they may be more appropriate for the study of congenital malformations, to investigate the secondary effects of prenatal urinary obstruction, to study the effect of prenatal exposure to endogenous or exogenous factors which may lead to disease, or in testing bioengineered structures. In this short educational article, we aim to describe some disease models that have been used to simulate human pathologies and how, if properly designed, these studies can lead to important new knowledge for human translation. In addition, we also highlight the importance of formulating a research question(s) before deciding on the animal experimental model and species to choose.

Precision medicine and rare diseases in pediatric urology.

Precision Medicine holds promise for helping us manage specific phenotypes of common diseases. For rare diseases such as hypospadias, DSD, and pediatric solid tumors, it can also reveal underlying risk factors and pathogenesis. Professors Ann Nordgren and Anna Lindstrand share their experiences in the development and ongoing initiatives of the Swedish national project on Precision Medicine and how it could change the care of pediatric urology patients.