Characterizing relaxin receptor expression and exploring relaxin's effect on tissue remodeling/fibrosis in the human bladder.

BACKGROUND: Relaxin is an endogenous protein that has been shown to have antifibrotic properties in various organ systems. There has been no characterization of relaxin's role in the human bladder. Our objective was to characterize relaxin receptor expression in the human bladder and assess relaxin's effect on tissue remodeling/fibrosis pathways in bladder smooth muscle cells. METHODS: Relaxin family peptide receptor 1 (RXFP1) and RXFP2 expression was assessed using quantitative reverse transcriptase-PCR (qRT-PCR) and immunohistochemistry (IHC) on primary bladder tissue. Primary human smooth muscle bladder cells were cultured and stimulated with various concentrations of relaxin. Western blot, qRTPCR, ELISA, and zymogram assays were used to analyze fibrosis/tissue remodeling pathway proteins. RESULTS: There was universal mRNA transcript detection and protein expression of relaxin receptors in primary bladder specimens. Immunohistochemistry demonstrated RXFP1 and RXFP2 localizing to both urothelial and smooth muscle cell layers of the bladder. 24 h of in vitro relaxin stimulation did not affect mRNA expression of selected proteins in human bladder smooth muscle cells. However, 48 h of in vitro relaxin stimulation resulted in upregulation of active (p = 0.004) and latent (p = 0.027) MMP-2 in cell lysate, and upregulation of active MMP-2 in supernatant (p = 0.04). There was a dose dependent relationship with increasing expression of MMP-2 with increasing relaxin concentration. Relaxin stimulation resulted in decreased levels of active and total TGF-ß1 in supernatant and extracellular matrix (p < 0.005 with 100 ng/mL relaxin stimulation). CONCLUSIONS: In the human bladder, relaxin receptors are expressed at the dome and trigone and localize to the urothelium and smooth muscle cell layers. Stimulation of human bladder SMCs with relaxin in vitro affects expression of MMP-2 and TGF-ß1.

Laparoscopic-assisted vesicocalicostomy for severe pelvi-ureteral stricture disease.

A 39-year-old female previously treated with shock wave lithotripsy developed extensive ureteral stricture disease. After 2 unsuccessful attempts at retrograde balloon dilatation, she was evaluated at our center for further management. Successful reconstruction was performed with laparoscopic-assisted vesicocalicostomy.

Integrated Ultrasound With Urodynamics Illustrates Effect of Bladder Volume on Upper Tract Dilation: Should we Trust Surveillance Ultrasounds?

OBJECTIVE: To evaluate if ultrasound during urodynamics (uUS) will show that traditional ultrasound (tUS) routinely underestimates the potential magnitude of upper tract dilation (UTD). METHODS: Prospective pilot study of 10 consecutive patients ≥ 5 years of age undergoing same day uUS and tUS. Using randomized images, the study pediatric radiologist determined anterior-posterior renal pelvic diameter (APD), bladder volume, vesicoureteral reflux (VUR) and UTD grades. A single pediatric urologist determined urodynamic bladder capacity and assigned either hostile, intermediate, abnormal but safe, or normal national spina bifida patient registry classification (NSBPR). RESULTS: Bladder volume on tUS was significantly smaller than final bladder volume on uUS (180 vs 363 ml: P<.001). On average, patient reported maximum catheterized/voided volumes were also 82 ml greater than final bladder capacity on uUS. UTD was upgraded in 25% of kidneys and APD increased by 0.6 cm on uUS over that seen on tUS (P=.001). Units with VUR had greater increases in APD (1.2 P=.007 vs. 0.3 cm P=0.06). Changes in APD stratified by NSBPR revealed average increases of up to 1.3 cm. CONCLUSION: Despite instructions to the contrary, patients come for tUS with a relatively empty bladder as compared to either their urodynamic or patient-reported capacity. This translates to a significant underestimation of UTD with tUS, most notably in those with VUR. Alternatives to traditional protocols include insisting patients wait until their bladder is truly full for tUS, retrograde filling their bladder, or performing uUS. Accurate assessment of UTD severity may help guide long term management.

2 year outcome for 8 year old female managed with partial cystectomy for primary bladder clear cell carcinoma.

Bladder cancer is rare in the pediatric population, and clear cell carcinoma is extremely rare with one other pediatric case reported. Here we report the clinical outcome for a medically complicated pediatric patient with muscle invasive clear cell carcinoma treated with partial cystectomy without neoadjuvant or adjuvant therapy. Final pathology was stage T2bN0M0 with negative margins. At 2 years, there is no disease recurrence by cystoscopy, chest and abdominal imaging. Postoperative issues have been related to reduced bladder capacity and compliance and the patient is currently managed with continuous urinary diversion and will require future definitive lower tract reconstruction.

The Role of Genetically Modified Mesenchymal Stem Cells in Urinary Bladder Regeneration.

Recent studies have demonstrated that mesenchymal stem cells (MSCs) combined with CD34+ hematopoietic/stem progenitor cells (HSPCs) can function as surrogate urinary bladder cells to synergistically promote multi-faceted bladder tissue regeneration. However, the molecular pathways governing these events are unknown. The pleiotropic effects of Wnt5a and Cyr61 are known to affect aspects of hematopoiesis, angiogenesis, and muscle and nerve regeneration. Within this study, the effects of Cyr61 and Wnt5a on bladder tissue regeneration were evaluated by grafting scaffolds containing modified human bone marrow derived MSCs. These cell lines were engineered to independently over-express Wnt5a or Cyr61, or to exhibit reduced expression of Cyr61 within the context of a nude rat bladder augmentation model. At 4 weeks post-surgery, data demonstrated increased vessel number (~250 vs ~109 vessels/mm2) and bladder smooth muscle content (~42% vs ~36%) in Cyr61OX (over-expressing) vs Cyr61KD (knock-down) groups. Muscle content decreased to ~25% at 10 weeks in Cyr61KD groups. Wnt5aOX resulted in high numbers of vessels and muscle content (~206 vessels/mm2 and ~51%, respectively) at 4 weeks. Over-expressing cell constructs resulted in peripheral nerve regeneration while Cyr61KD animals were devoid of peripheral nerve regeneration at 4 weeks. At 10 weeks post-grafting, peripheral nerve regeneration was at a minimal level for both Cyr61OX and Wnt5aOX cell lines. Blood vessel and bladder functionality were evident at both time-points in all animals. Results from this study indicate that MSC-based Cyr61OX and Wnt5aOX cell lines play pivotal roles with regards to increasing the levels of functional vasculature, influencing muscle regeneration, and the regeneration of peripheral nerves in a model of bladder augmentation. Wnt5aOX constructs closely approximated the outcomes previously observed with the co-transplantation of MSCs with CD34+ HSPCs and may be specifically targeted as an alternate means to achieve functional bladder regeneration.

The promotion of functional urinary bladder regeneration using anti-inflammatory nanofibers.

Current attempts at tissue regeneration utilizing synthetic and decellularized biologic-based materials have typically been met in part by innate immune responses in the form of a robust inflammatory reaction at the site of implantation or grafting. This can ultimately lead to tissue fibrosis with direct negative impact on tissue growth, development, and function. In order to temper the innate inflammatory response, anti-inflammatory signals were incorporated through display on self-assembling peptide nanofibers to promote tissue healing and subsequent graft compliance throughout the regenerative process. Utilizing an established urinary bladder augmentation model, the highly pro-inflammatory biologic scaffold (decellularized small intestinal submucosa) was treated with anti-inflammatory peptide amphiphiles (AIF-PAs) or control peptide amphiphiles and used for augmentation. Significant regenerative advantages of the AIF-PAs were observed including potent angiogenic responses, limited tissue collagen accumulation, and the modulation of macrophage and neutrophil responses in regenerated bladder tissue. Upon further characterization, a reduction in the levels of M2 macrophages was observed, but not in M1 macrophages in control groups, while treatment groups exhibited decreased levels of M1 macrophages and stabilized levels of M2 macrophages. Pro-inflammatory cytokine production was decreased while anti-inflammatory cytokines were up-regulated in treatment groups. This resulted in far fewer incidences of tissue granuloma and bladder stone formation. Finally, functional urinary bladder testing revealed greater bladder compliance and similar capacities in groups treated with AIF-PAs. Data demonstrate that AIF-PAs can alleviate galvanic innate immune responses and provide a highly conducive regenerative milieu that may be applicable in a variety of clinical settings.

Laparoscopic-assisted vesicocalicostomy for severe pelvi-ureteral stricture disease.

A 39-year-old female previously treated with shock wave lithotripsy developed extensive ureteral stricture disease. After 2 unsuccessful attempts at retrograde balloon dilatation, she was evaluated at our center for further management. Successful reconstruction was performed with laparoscopic-assisted vesicocalicostomy.