Urothelial Injury Research Articles

Vitamin A affects urothelial injury and regeneration in cyclophosphamide-induced cystitis

Vitamin A affects urothelial injury and regeneration in cyclophosphamide-induced cystitis

Prolonged urinary retention in COVID-19 survivors: observational study.

Initially diagnosed as a respiratory disease, SARS-CoV-2 revealed numerous extrapulmonary implications. The aim of this study was to investigate prolonged urinary retention in survivors post-COVID-19 infection that led to hospitalization. A retrospective cohort analysis included male and female patients hospitalized during the COVID-19 pandemic in a reference center hospital. Exclusions were patients with a history of lower urinary tract surgeries or symptoms, including urinary incontinence, those using medication affecting detrusor contractile activity, and those with established neurological diseases. Clinical, laboratory, and radiographic data were obtained from medical records and analyzed using chi-square, Fisher's exact, Mann-Whitney, Kruskal-Wallis, and Dunn tests. The study included 834 patients, with 471 (56.5%) male and 363 (43.5%) female. Of these, 300 patients used a urinary catheter, and 12.6% were unable to remove it due to sustained urinary retention. Orotracheal intubation, thrombocytopenia, urinary tract infections, and higher Sequential Organ Failure Assessment scores were associated with urinary retention. Correlation analysis showed that the highest percentage of pulmonary involvement on computed tomography was related to longer catheterization time and failed attempts to remove the catheter, affecting men and women equally. Urinary tract involvement in COVID-19 infection is increasingly evident. The correlation between COVID-19 severity and failure to remove the urinary catheter in a similar percentage of men and women reinforces the hypothesis that sex-independent urothelial injury and bladder dysfunction might be caused by COVID-19.

Beyond the urothelium: Interplay between autonomic nervous system and bladder inflammation in urinary tract infection, bladder pain syndrome with interstitial cystitis and neurogenic lower urinary tract dysfunction in spinal cord injury-ICI-RS 2023.

Inflammation and neuronal hypersensitivity are reactive protective mechanisms after urothelial injury. In lower urinary tract dysfunctions (LUTD), such as urinary tract infection (UTI), bladder pain syndrome with interstitial cystitis (BPS/IC) and neurogenic LUTD after spinal cord injury (SCI), chronic inflammation can develop. It is unclear how the protective reactionary inflammation escalates into chronic disease in some patients. During its 2023 meeting in Bristol, the International Consultation on Incontinence-Research Society (ICI-RS) reviewed the urothelial and inflammatory changes after UTI, BPS/IC and SCI. Potential factors contributing to the evolution into chronic disease were explored in a think-tank. Five topics were discussed. (1) Visceral fat metabolism participates in the systemic pro-inflammatory effect of noradrenalin in BPS/IC and SCI. Sympathetic nervous system-adipocyte-bladder crosstalk needs further investigation. (2) Sympathetic hyperactivity also potentiates immune depression in SCI and needs to be investigated in BPS/IC. Gabapentin and tumor necrosis factor-α are promising research targets. (3) The exact peripheral neurons involved in the integrative protective unit formed by nervous and immune systems need to be further identified. (4) Neurotransmitter changes in SCI and BPS/IC: Neurotransmitter crosstalk needs to be considered in identifying new therapeutic targets. (5) The change from eubiosis to dysbiosis in SCI can contribute to UTI susceptibility and needs to be unraveled. The think-tank discussed whether visceral fat metabolism, immune depression through sympathetic hyperactivity, peripheral nerves and neurotransmitter crosstalk, and the change in microbiome could provide explanations in the heterogenic development of chronic inflammation in LUTD. High-priority research questions were identified.

P16 Immunohistochemical Expression in Nephrogenic Adenoma.

Nephrogenic adenoma (NA) is a rare metaplastic entitycommonly associated with a prior urothelial injury. Most are seen in the urinary bladder and a minority involve the urethra. In this study, we evaluated the expression of p16 as a surrogate marker of this entity and correlated it with clinical pathological parameters. A total of 17 cases ofNA were retrospectively studied to assess the immunohistochemical expression of p16 and its value for the diagnosis of this entity.

Update on the Effect of the Urinary Microbiome on Urolithiasis.

Microbiota are ecological communities of commensal, symbiotic, and pathogenic microorganisms. The microbiome could be involved in kidney stone formation through hyperoxaluria and calcium oxalate supersaturation, biofilm formation and aggregation, and urothelial injury. Bacteria bind to calcium oxalate crystals, which causes pyelonephritis and leads to changes in nephrons to form Randall's plaque. The urinary tract microbiome, but not the gut microbiome, can be distinguished between cohorts with urinary stone disease (USD) and those without a history of the disease. In the urine microbiome, the role is known of urease-producing bacteria (Proteus mirabilis, Klebsiella pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Providencia stuartii, Serratia marcescens, and Morganella morganii) in stone formation. Calcium oxalate crystals were generated in the presence of two uropathogenic bacteria (Escherichia coli and K. pneumoniae). Non-uropathogenic bacteria (S. aureus and Streptococcus pneumoniae) exhibit calcium oxalate lithogenic effects. The taxa Lactobacilli and Enterobacteriaceae best distinguished the healthy cohort from the USD cohort, respectively. Standardization is needed in urine microbiome research for urolithiasis. Inadequate standardization and design of urinary microbiome research on urolithiasis have hampered the generalizability of results and diminished their impact on clinical practice.

Evaluating the Potential of a New Low-Profile Urinary Catheter in Preventing Catheter-Associated Urinary Tract Infections: A Prospective Randomized Blinded Clinical Trial

Aim To investigate the efficacy of a new low-profile catheter on incidence of the catheter-associated urinary tract infections (CAUTI) in comatose patients admitted to the intensive care unit. Background Catheter-induced urothelial injury is a key component in the development of urinary tract infections in catheterized patients. Methods In this prospective randomized blinded clinical trial, 80 patients requiring indwelling urinary catheterization were equally randomized to either the standard Foley catheter (control) or the low-profile catheter (experimental) group. The signs of urinary tract infection for comatose patients were considered (ie, ≥105 of colony-forming unit/milliliter of urine, hematuria, serum leukocytes, and body temperature) and recorded at baseline and on days 3 and 5 after catheterization. The analysis of covariance was applied by the SPSS-20 software at a 95% confidence level. Results An increasing proportion of patients with elevated urinary colony counts were seen in the Foley catheter group compared with the low-profile catheter group (12.5% vs 5%). However, there were no between-group differences in the urinary colony counts and body temperature after controlling for antibiotic doses and fluid intake. Patients in the low-profile catheter group had significantly lower rates of hematuria and serum leukocytes than those in the Foley catheter group. Conclusion A newly designed low-profile urinary catheter has demonstrated a trend toward reducing the incidence of CAUTI in patients with indwelling urinary catheters. Further studies with larger sample sizes and follow-up are needed to confirm the benefits.

Fibromyxoid Nephrogenic Adenoma: A Series of 43 Cases Reassessing Predisposing Conditions, Clinical Presentation, and Morphology.

Nephrogenic adenoma is a benign epithelial lesion of the genitourinary tract that arises from the reimplantation and proliferation of shed renal tubular cells in areas of urothelial injury and denudation. Fibromyxoid nephrogenic adenoma is a rare variant that consists of compressed spindle-shaped renal epithelial cells in a fibromyxoid background. Only 14 observations of this variant are reported in the literature. We performed a retrospective analysis of fibromyxoid nephrogenic adenomas from 3 large reference centers. We identified 43 lesions in 6 women and 36 men (2 in 1 man) with a median age of 72 years (range, 31 to 94y). Median lesion size was 0.7cm (range, 0.2 to 5cm). Nephrogenic adenomas were in the bladder (n=15), prostate/prostatic urethra (n=14), kidney (n=7), ureter (n=3), penile urethra (n=3), and urethral diverticulum (n=1). One of the kidney lesions developed in an end-stage kidney and radiologically mimicked cancer. Of 37 patients with information, 36 had predisposing conditions including prior biopsy, transurethral resection of bladder tumor, resection, Foley catheter, BCG treatment, urinary stones, (chemo)radiation, or diverticulum. Only 4/37 (10.8%) had a history of prior irradiation. Fifteen lesions had pure fibromyxoid morphology and 28 were admixed classic and fibromyxoid patterns. Three nephrogenic adenomas involved prostatic stroma, 3 renal sinus fat, 2 muscularis propria (1 bladder, 1 renal pelvis), 1 perinephric fat, and 1 corpus spongiosum. Ten fibromyxoid nephrogenic adenomas were intermixed with urothelial carcinoma, 1 with prostate adenocarcinoma, and 1 with malignant melanoma. By immunohistochemistry, PAX8 was positive in all the examined lesions (n=31). Napsin A was negative in all examined fibromyxoid nephrogenic adenomas (n=30). Twenty of them had classic nephrogenic adenoma component which was positive for napsin A. Similar to classic nephrogenic adenoma, fibromyxoid nephrogenic adenoma can occur anywhere along the urinary tract and is associated with a prior history that causes urothelial injury. In nearly a quarter of the cases, fibromyxoid nephrogenic adenoma extended beyond the lamina propria. Unlike previously suggested, fibromyxoid nephrogenic adenoma is not specifically related to prior radiation therapy. Awareness of this variant is important to avoid misdiagnosis and overtreatment.

Urine-Derived Stem Cells for Epithelial Tissues Reconstruction and Wound Healing.

Epithelial tissue injury can occur on any surface site of the body, particularly in the skin or urethral mucosa tissue, due to trauma, infection, inflammation, and toxic compounds. Both internal and external body epithelial tissue injuries can significantly affect patients’ quality of life, increase healthcare spending, and increase the global economic burden. Transplantation of epithelial tissue grafts is an effective treatment strategy in clinical settings. Autologous bio-engineered epithelia are common clinical skin substitutes that have the specific advantages of avoiding tissue rejection, obviating ethical concerns, reducing the risk of infection, and decreasing scarring compared to donor grafts. However, epithelial cells are often obtained from the individual’s skin and mucosa through invasive methods, which cause further injury or damage. Urine-derived stem cells (USC) of kidney origin, obtained via non-invasive acquisition, possess high stemness properties, self-renewal ability, trophic effects, multipotent differentiation potential, and immunomodulatory ability. These cells show versatile potential for tissue regeneration, with extensive evidence supporting their use in the repair of epidermal and urothelial injuries. We discuss the collection, isolation, culture, characterization, and differentiation of USC. We also discuss the use of USC for cellular therapies as well as the administration of USC-derived paracrine factors for epidermal and urothelial tissue repair. Specifically, we will discuss 3D constructions involving multiple types of USC-loaded hydrogels and USC-seeded scaffolds for use in cosmetic production testing, drug development, and disease modeling. In conclusion, urine-derived stem cells are a readily accessible autologous stem cell source well-suited for developing personalized medical treatments in epithelial tissue regeneration and drug testing.

The Uroprotective Efficacy of Total Ginsenosides in Chinese Ginseng on Chemotherapy with Cyclophosphamide

Hemorrhagic cystitis is a recognizable complication of cyclophosphamide (CYP) attributable to its lively metabolite acrolein, which produces urothelial injury. The study intended to examine the uroprotective efficacy of total ginsenosides in Chinese ginseng (TGCG) in CYP-induced hemorrhagic cystitis. In total, 24 virgin female rats were randomized into four groups as follows: group 1 (control group; injected with normal saline), group 2 (injected with CYP plus a placebo with normal saline), group 3 (given CYP and TGCG (200 mg/kg)), and group 4 (given CYP and 2-mercaptoethane sulfonate sodium (Mesna, 30 mg/kg)). An evaluation by cystometry was conducted. Values of the voiding interval were assessed in anesthetized rats and histological examinations of the bladders were measured. In the cystometry analysis, the voiding interval was significantly reduced in the CYP group. TGCG and Mesna significantly increased in the voiding interval values, individually. Bladder edema and urothelial injury were examined after contact with CYP. Contrasted to the group given CYP, CYP-induced hemorrhagic cystitis, TGCG significantly increased the urothelial thickness, and significantly reduced scores of mucosal break and submucosal edema in the bladder. In conclusion, these findings mean that the treatment with TGCG in CYP rats can avoid hemorrhagic cystitis. TGCG decreases urothelial injury. TGCG may participate as the chief character of uroprotection in CYP-induced hemorrhagic cystitis.

Role of ERK signaling in bladder urothelium in response to cyclophosphamide injury.

Mice with inducible urothelial deletion of fibroblast growth factor receptor 2 (ShhCreERT2;Fgfr2 Fl/Fl ) injured with cyclophosphamide had aberrant basal cell endoreplication and poor regeneration. The endoreplication correlated with an absence of phosphorylated (activated) ERK expression in urothelium. We assessed whether inhibiting ERK activity phenocopied the urothelial defects in injured Fgfr2 mutant mice. We co‐administered cyclophosphamide and an ERK inhibitor (ERKi) systemically in mice and assessed general histology and immunofluorescence for various markers post injury. Since AKT also signals downstream of FGFR2, we assessed effects of an AKT inhibitor (AKTi) on cyclophosphamide injury. ERK knockdown did not affect urothelial injury or proliferation 24 h after cyclophosphamide. Conversely, ERK inhibition led to larger basal cell nuclei, more submucosal hemorrhage and attenuated uroplakin staining 3 days after injury versus vehicle‐treated mice. Compared to vehicle‐treated mice, ERKi‐treated mice had a trend for more Ki67+ urothelial cells and had statistically fewer phospho‐Histone H3+ cells normalized to Ki67 and higher basal cell DNA content, consistent with endoreplication 3 days after injury. Ten days after injury, ERKi‐treated mice still had signs of poor urothelial regeneration with absent or aberrant expression of differentiation markers and ectopic lumenal expression of keratin 14 (basal progenitor marker). Co‐administration of the AKTi led to no apparent urothelial defects 3 days after cyclophosphamide. Thus, ERK knockdown (but not AKT knockdown) leads to urothelial regenerative responses after cyclophosphamide reminiscent of Fgfr2 mutant mice. Together, it appears that FGFR2 acts through ERK to prevent aberrant urothelial basal cell endoreplication and ensure normal regeneration after cyclophosphamide.

Abstract 5431: FGF7 peptide (FGF7p) mimetic mitigates urothelial injury from cyclophosphamide or radiation

Abstract Background: Hemorrhagic cystitis from chemical or radiation bladder injury can cause life-threatening bleeding and/or urothelial cancer. Fibroblast growth factor 7 (FGF7) blocks urothelial apoptosis induced by cyclophosphamide (CPP) or radiation, but there are limitations to its use in patients, including an inability to apply it by direct bladder infusion and the high cost due to the large size of the protein (18.8 kDa). Purpose: We previously identified a small peptide that equaled or exceeded the mitigation effect of full length FGF2 on acute radiation syndrome. Benefits of developing a similar FGF7 biomimetic peptide include lower likelihood of inflammation, longer shelf life, higher purity, and lower cost than the full-length protein. Methods: Based on the 3-dimensional structure of FGF2 peptide, a corresponding FGF7 peptide (FGF7p) consisting of 19 amino acids was identified and synthesized. FGF7p or vehicle was given subcutaneously (SQ) to female mice subjected to sham injury, intraperitoneal (IP) CPP or external beam radiation over the bladder. One day after injury, bladders were harvested. Slides with paraffin embedded tissues underwent H&E staining, TUNEL and immunofluorescence (IF) assays. Results: In uninjured control mice, a 20 mg/kg threshold dose of FGF7p induced expression of phosphorylated (activated) FRS2a (pFRS2a) and pAKT in urothelium (consistent with cytoprotective effects of FGF7, albeit at a 4x higher dose than full length FGF7). Unexpectedly, FGF7p activation of urothelial FRS2a and AKT was delayed 24 hours compared to FGF7. FGF7p (20 mg/kg) or vehicle was given at 72 and 48 hours prior to CPP (150 mg/kg). One day after CPP, TUNEL staining revealed an increase in apoptotic and sloughing urothelial cells in vehicle-treated mice compared to in FGF7p-treated mice. IF for pAKT and its targets, pS6K and pBAD, revealed minimal staining in vehicle-treated mice, but strong urothelial staining for all protective markers in FGF7p-treated mice. Using the same dosing strategy, we subjected anesthetized mice to 10 Gy radiation over the bladder. One day after injury, TUNEL staining revealed many more apoptotic urothelial nuclei in control mice, than in FGF7p-treated mice. Conclusions: FGF7p appears to block bladder urothelial apoptosis via AKT and its targets, in a similar manner to FGF7. FGF7p is 200x less expensive to make vs. FGF7 and is likely to work via direct bladder infusion (avoiding systemic side effects) due to its small size. Our future studies will clarify long term benefits of FGF7p and assess effectiveness of direct bladder infusion. Citation Format: Sridhar Narla, Lori P. Rice, David Ostrov, Steven G. Swarts, Dietmar W. Siemann, Carlton M. Bates. FGF7 peptide (FGF7p) mimetic mitigates urothelial injury from cyclophosphamide or radiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5431.

FGF7 peptide (FGF7p) mimetic mitigates bladder urothelial injury from cyclophosphamide.

Although full‐length fibroblast growth factor 7 (FGF7) blocks cyclophosphamide‐induced urothelial apoptosis in mice, limitations include high production costs because of its large size. We previously identified a small peptide derived from FGF2 that mitigated acute radiation syndrome as well as full‐length FGF2. Based on the sequence of the FGF2 peptide, we synthesized a corresponding 19 amino acid FGF7 peptide (FGF7p). Our objectives were to determine if systemic FGF7p triggered the downstream targets and protected against cyclophosphamide bladder injury similar to full‐length FGF7. We administered FGF7p or vehicle subcutaneously (SQ) to mice subjected to no injury or intraperitoneal (IP) cyclophosphamide and harvested bladders 1 day after injury. We then performed hematoxylin and eosin, TUNEL and immunofluorescence (IF) staining. In uninjured mice, a 20 mg/kg threshold FGF7p dose induced expression of phosphorylated (activated) FRS2α (pFRS2α), and pAKT in urothelium (consistent with cytoprotective effects of FGF7). We then gave FGF7p (20 mg/kg) or vehicle at 72 and 48 h prior to cyclophosphamide. One day after injury, TUNEL staining revealed many more apoptotic urothelial cells with vehicle treatment versus FGF7p treatment. IF for pAKT and readouts of two anti‐apoptotic AKT targets (BAD and mTORC1) revealed minimal staining with vehicle treatment, but strong urothelial expression for all markers with FGF7p treatment. In conclusion, FGF7p appears to block bladder urothelial apoptosis via AKT and its targets, similar to FGF7. FGF7p is much more inexpensive to make and has a longer shelf life and higher purity than FGF7.

Urothelial Injury Triggers Adaptive Remodeling That Limits Congenital and Acquired Obstructive Nephropathy

Urothelial Injury Triggers Adaptive Remodeling That Limits Congenital and Acquired Obstructive Nephropathy

Urothelial progenitors in development and repair.

Urothelium is a specialized multilayer epithelium that lines the urinary tract from the proximal urethra to the kidney. In addition to proliferation and differentiation during development, urothelial injury postnatally triggers a robust regenerative capacity to restore the protective barrier between the urine and tissue. Mounting evidence supports the existence of dedicated progenitor cell populations that give rise to urothelium during development and in response to injury. Understanding the cellular and molecular basis for urothelial patterning and repair will inform tissue regeneration therapies designed to ameliorate a number of structural and functional defects of the urinary tract. Here, we review the current understanding of urothelial progenitors and the signaling pathways that govern urothelial development and repair. While most published studies have focused on bladder urothelium, we also discuss literature on upper tract urothelial progenitors. Furthermore, we discuss evidence supporting existence of context-specific progenitors. This knowledge is fundamental to the development of strategies to regenerate or engineer damaged or diseased urothelium.

A randomized study comparing conventional percutaneous nephrolithotomy versus check pyeloscopic percutaneous nephrolithotomy.

Objective:The objective of this study is to compare the advantages of check-pyeloscopic percutaneous nephrolithotomy (CP-PCNL) over conventional PCNL (C-PCNL) in the management of renal calculi.Materials and Methods:All patients with renal calculi requiring PCNL who attended the department of urology in a tertiary care center from December 2016 to October 2018 were included in the study. The patients were randomized into two groups of 50 each. Each group underwent PCNL in a conventional or check pyeloscopic method, respectively.Results:A total number of complications were more in C-PCNL group comprising of two pelvic tears, 8 infundibular tears, 1 minor urothelial injury, 2 cases of bleeding lasting for more than 24 hours, while in CP-PCNL group, complications were less (comprising of 1 minor urothelial injury and 1 case of infundibular tear). Statistical analysis was used to compare both the groups, and the results were statistically significant (P = 0.027).Conclusion:Check pyeloscopy at the time of PCNL is a safe, effective, and economical technique. This study shows that the complications such as blood loss, urothelial injury, and postoperative pain are less when check pyeloscopy is done during PCNL.

Removal of proximally migrated double J ureteric stent through a percutaneous nephrostomy route in an infant

Proximal migration of double J-stent (DJ stent) can be a reason for failure of its cystoscopic removal. Antegrade percutaneous nephrostomic removal of the stent can be done in such cases. Although the technique is described in adults and older children, the technique requires some modification when used in infants because of smaller calibre of pelvicalyceal system. Here we describe a case of a 6 month old male child who was treated by robotic assisted pyeloplasty for left sided ureteropelvic junction obstruction followed by placement of DJ stent. Cystoscopic removal of the stent 6 weeks after surgery was not possible due to proximal migration of the stent. Hence, antegrade percutaneous nephrostomic removal of the stent was performed using micro-snare. Certain novel technical modifications, like advancement of a diagnostic angiographic catheter in pelvicalyceal system, has been used in this case to reduce the risk of urothelial injury. Thus, antegrade removal of DJ stent with few technical modifications can be used safely in infants for removal of proximally migrated stents.

Platelet-Rich Plasma Ameliorates Cyclophosphamide-Induced Acute Interstitial Cystitis/Painful Bladder Syndrome in a Rat Model.

Background: Interstitial cystitis/painful bladder syndrome (IC/PBS) could be treated to ameliorate urothelial injury. Here, we investigated the efficacy of intravesical instillation with platelet-rich plasma (PRP) and hyaluronic acid for acute IC/PBS. Methods: The effects of PRP and hyaluronic acid on the proliferation of normal human fibroblast cells (HFCs) were assessed. Additionally, thirty virgin female rats were randomized into five groups: group 1, saline-injected control; group 2, cyclophosphamide (CYP) plus intravesical instillation with normal saline; group 3, CYP plus intravesical instillation with hyaluronic acid (1 mg/mL); group 4, CYP plus intravesical instillation with PRP; and group 5, CYP plus intravesical instillation with PRP plus hyaluronic acid. A cystometry and histological assessments were performed. The expression of cell junction-associated protein zonula occludens-2 (ZO-2) and inflammatory cytokine interleukin 6 (IL-6) was also measured. Results: Low dose PRP increased proliferation in HFCs. The acute IC/PBS rats showed significantly lower voiding interval values. Voiding interval values were significantly higher in the CYP plus intravesical instillation with PRP group than in the CYP-induced acute IC/PBS group. Additionally, the expression of ZO-2 was increased and IL-6 was decreased in the CYP plus intravesical instillation with PRP group compared with the CYP-induced acute IC/PBS group. Conclusion: These findings suggest that PRP modulate urothelial repair, which ameliorate the increase in urination frequency in rats treated with CYP. Overall, PRP may confer potential benefits by acting as urothelial repair modulators.

Astragalus polysaccharides and astragaloside IV ameliorates cyclophosphamide-induced mouse model of overactive bladder

ObjectivePrevious studies have shown that Astragalus polysaccharides (APS) and Astragaloside IV (AS-IV) protect against inflammation-related cell damage and exhibit immune enhancement. Since urothelial injury may result in an overactive bladder (OAB), the aim of this study was to investigate the efficacy of APS and AS-IV on urothelial injury in an experimental animal model. Materials and methodsThe effects of APS and AS-IV on the proliferation and migration of primary human urothelial cells (HUCs) or primary human fibroblast cells (HFCs) were assessed using an in vitro wounding model and colorimetric thiazolyl blue assays. Sixty virgin female mice were randomized into five groups: group 1–saline-injected plus treatment with H2O, group 2–cyclophosphamide (CYP) plus treatment with H2O, group 3–CYP plus treatment with solifenacin succinate (SS; 10 mg/kg), group 4–CYP plus treatment with AS-IV (100 mg/kg), and group 5–CYP plus treatment with APS (100 mg/kg). Cystometry assessment was conducted and cell junction-associated protein zonula occludens-2 (ZO-2) expression was measured. Voiding interval values (time between voids) were assessed in mice under anesthesia. Lastly, immunohistochemistry analysis was used to confirm the location and level, respectively, of ZO-2 expression. ResultsAPS and AS-IV did not influence the cell viability but increased migration in HFCs compared with the controls. The OAB mice showed significantly lower voiding interval values. Voiding interval values were significantly higher in the CYP plus treatment with APS (100 mg/kg) and AS-IV (100 mg/kg) groups than in the CYP-induced OAB group. Additionally, the expression of ZO-2, a tight junction protein, was increased in the CYP plus treatment APS (100 mg/kg) and AS-IV (100 mg/kg) groups compared with the CYP-induced OAB group. ConclusionThese findings suggest that APS and AS-IV modulate urothelial wound healing, which ameliorates urinary frequency of mice treated with CYP. APS or AS-IV may have the potential benefit of acting as urothelial wound healing modulators.

Human Amniotic Membrane Enriched with Urinary Bladder Fibroblasts Promote the Re-Epithelization of Urothelial Injury

Culturing cells in three-dimensional systems that include extracellular matrix components and different cell types mimic the native tissue and as such provide much more representative results than conventional two-dimensional cell cultures. In order to develop biomimetic bladder tissue in vitro, we used human amniotic membrane (AM) extracellular matrix as a scaffold for bladder fibroblasts (BFs) and urothelial cells. Our aims were to evaluate the integration of BFs into the AM stroma, to assess the differentiation of the urothelium on BFs-enriched AM scaffolds, and to evaluate the AM as a urothelial wound dressing. First, to achieve the optimal integration of BFs into AM stroma, different intact and de- epithelialized AM (dAM) scaffolds were tested. BFs secreted matrix metalloproteinase (MMP)-1 and MMP-2 and integrated into the stroma of all types of AM scaffolds. Second, to establish urothelial tissue equivalent, urothelial cells were seeded on dAM scaffolds enriched with BFs. The BFs in the stroma of the AM scaffolds promoted (1) the proliferation of urothelial cells, (2) the attachment of urothelial cells on AM basal lamina with hemidesmosomes, and (3) development of multilayered urothelium with expressed uroplakins and well-developed cell junctions. Third, we established an ex vivo model of the injured bladder to evaluate the dAM as a wound dressing for urothelial full-thickness injury. dAM acted as a promising wound dressing since it enabled rapid re-epithelization of urothelial injury and integrated into the bladder tissue. Herein, the developed urothelial tissue equivalents enable further mechanistic studies of bladder epithelial–mesenchymal interactions, and they could be applied as biomimetic models for preclinical testing of newly developed drugs. Moreover, we could hypothesize that AM may be suitable as a dressing of the wound that occurs during transurethral resection of bladder tumor, since it could diminish the possibility of tumor recurrence, by promoting the rapid re-epithelization of the urothelium.

Intravesical Administration of Xenogeneic Porcine Urothelial Cells Attenuates Cyclophosphamide-Induced Cystitis in Mice

The urothelium of the bladder, renal pelvis, ureter and urethra is maintained through the regulated proliferation and differentiation of urothelial stem and progenitor cells. These cells provide a rich source of a novel urothelial cell therapy approach that could be used to protect, regenerate, repair and restore a damaged urothelium. Urothelial injury caused by physical, chemical and microbial stress is the pathological basis of cystitis (bladder inflammation). The loss of urothelial integrity triggers a series of inflammatory events, resulting in pain and hematuria such as hemorrhage cystitis and interstitial cystitis. Here we investigate a novel cell therapy strategy to treat cystitis by protecting the urothelium from detrimental stresses through intravesically instilling porcine urothelial cells (PUCs) into the bladder. Using a chemical-induced urothelial injury mouse model of cyclophosphamide (CPP)-induced hemorrhagic cystitis, we determined how the intravesical instillation of PUCs could protect the urothelium from toxic attack from CPP metabolites. We show that intravesical PUC instillation protected the bladder from toxic chemical attack in mice receiving CPP with reduced inflammation and edema. Compared with the vehicle control mice, the proliferative response to chemical injury and apoptotic cells within the bladder tissues were reduced by intravesical PUC treatment. Furthermore, the urothelium integrity was maintained in the intravesical PUC-treated group. After xenogeneic PUCs were introduced and adhered to the mouse urothelium, immunological rejection responses were observed with increased neutrophil infiltration in the lamina propria and higher immune-related gene expression. Our findings provide an innovative and promising intravesical PUC cell therapy for cystitis with urothelial injury by protecting the urothelium from noxious agents.