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Introduction: Acellular matrices have historically been applied as biologic scaffolds in surgery, wound care, and tissue engineering, albeit with inconsistent outcomes. One aspect that varies widely between products is the selection of decellularization protocol, yet few studies assess comparative effectiveness of these protocols in preserving mechanics, and protein content. This study characterizes bladder acellular matrix (BAM) using two different detergent and enzymatic protocols, evaluating effects on nuclei and DNA removal (≥90%), structure, tensile properties, and maintenance of extracellular matrix proteins., Methods: Porcine bladders were decellularized with 0.5% Sodium Dodecyl Sulfate (SDS) or 0.25% Trypsin-hypotonic-Triton X-100 hypertonic (TT)-based agitation protocols, followed by DNase/RNase agents. Characterization of BAM included decellularization efficacy (DAPI, DNA quantification), structure (histology and scanning electron microscopy), tensile testing (Instron 345C-1 mechanical tester), and protein presence and diversity (mass spectrometry). SDS and TT data was directly compared to the same native bladder using two-tailed paired t-tests. Native, TT, and SDS cohorts for tensile testing were compared using one-way ANOVA; Tukey's post-hoc tests for among group differences., Results: Effective nuclei removal was achieved by SDS- and TT-based protocols. However, target DNA removal was achieved with SDS but not TT. SDS more effectively maintained qualitative tissue architecture compared to TT. The tensile modulus of the TT cohort increased, and stretchability decreased after decellularization in both SDS and TT. UTS was unaffected by either protocol. Higher overall diversity and quantity of core matrisome and matrisome-associated proteins was maintained in the SDS vs TT cohort post-decellularization., Conclusion: The results indicated that detergent selection affects multiple aspects of the resultant BAM biologic product. In the selected protocols, SDS was superior to TT efficacy, and maintenance of gross tissue architecture as well as maintenance of ECM proteins. Decellularization increased scaffold resistance to deformation in both cohorts. Future studies applying biologic scaffolds must consider the processing method and agents used to ensure that materials selected are optimized for characteristics that will facilitate effective translational use., Competing Interests: Conflict of interest Co-founder, board member of Surgi-Zipper (Sturm)., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: to evaluate a membrane based on type B gelatin (G) and porcine urinary bladder extracellular matrix (PUB-EM), highlighting the potential effect of the combination evaluated by biocompatibility and regulation of the expression of transcription factors involved in tissue regeneration. G-PUB-EM membranes were prepared at 12.5, 25, and 50% w/v, and evaluated for biocompatibility with Fibroblast. Chemical characterization by FTIR-ATR showed complex spectra during crosslinking process with glutaraldehyde. Physical tests were performed in deionized water and PBS for 48 h. A significant increase in swelling was observed during the first 2 h. Biocompatibility testing (MTS) and evaluation of the expression profile of genes involved in the cell cycle (Cyclin-D1 VEGF, TNF and NF-κ-B) by PCR showed an increase in viability in a PUB-EM content-dependent way, except for 50% PUB-EM membrane which showed cytotoxic effects with a decrease in cell viability below 70%. The membranes showed an increase in the expression of some factors of cell cycle, as well as inflammatory processes that could promote tissue repair. 12.5 and 25% gelatin type B/porcine urinary bladder extracellular matrix (G/PUB-EM) based membranes have potential for tissue regeneration applications., Impact Statement: The use of membranes based on type B gelatin and porcine urinary bladder for tissue engineering represents a novel strategy. Biocompatibility and signaling pathways play a primary role in tissue repair and wound recovery. Transcription factors that mediate signaling, cell division and vascularization are part of molecules that intervene in the regenerative potential of cells. These techniques will have a significant impact on tissue repair and regeneration and thus stop depending on tissue donors or other surgical sites from the same patient, as is the case with burn patients., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Previous studies showed that the bladder extracellular matrix (B-ECM) could increase the differentiation efficiency of mesenchymal cells into smooth muscle cells (SMC). This study investigates the potential of human amniotic membrane-derived hydrogel (HAM-hydrogel) as an alternative to xenogeneic B-ECM for the myogenic differentiation of the rabbit adipose tissue-derived MSC (AD-MSC). Decellularized human amniotic membrane (HAM) and sheep urinary bladder (SUB) were utilized to create pre-gel solutions for hydrogel formation. Rabbit AD-MSCs were cultured on SUB-hydrogel or HAM-hydrogel-coated plates supplemented with differentiation media containing myogenic growth factors (PDGF-BB and TGF-β1). An uncoated plate served as the control. After 2 weeks, real-time qPCR, immunocytochemistry, flow cytometry, and western blot were employed to assess the expression of SMC-specific markers (MHC and α-SMA) at both protein and mRNA levels. Our decellularization protocol efficiently removed cell nuclei from the bladder and amniotic tissues, preserving key ECM components (collagen, mucopolysaccharides, and elastin) within the hydrogels. Compared to the control, the hydrogel-coated groups exhibited significantly upregulated expression of SMC markers (p ≤ .05). These findings suggest HAM-hydrogel as a promising xenogeneic-free alternative for bladder tissue engineering, potentially overcoming limitations associated with ethical concerns and contamination risks of xenogeneic materials., (© 2024 Federation of American Societies for Experimental Biology.)

Previous studies have shown that growth arrest, dedifferentiation, and loss of original function occur in cells after multiple generations of culture, which are attributed to the lack of stress stimulation. To investigate the effects of multi-modal biomimetic stress (MMBS) on the biological function of human bladder smooth muscle cells (HBSMCs), a MMBS culture system was established to simulate the stress environment suffered by the bladder, and HBSMCs were loaded with different biomimetic stress for 24 h. Then, cell growth, proliferation and functional differentiation were detected. The results showed that MMBS promoted the growth and proliferation of HBSMCs, and 80 cm H 2 O pressure with 4% stretch stress were the most effective in promoting the growth and proliferation of HBSMCs and the expression level of α-smooth muscle actin and smooth muscle protein 22-α. These results suggest that the MMBS culture system will be beneficial in regulating the growth and functional differentiation of HBSMCs in the construction of tissue engineered bladder.

Urinary tract infection (UTI), one of the most common bacterial infections worldwide, is a typical example of an infection that is often polymicrobial in nature. While the overall infection course is known on a macroscale, bacterial behavior is not fully understood at the cellular level and bacterial pathophysiology during multispecies infection is not well characterized. Here, using clinically relevant bacteria, human epithelial bladder cells and human urine, we establish co-infection models combined with high resolution imaging to compare single- and multi-species bladder cell invasion events in three common uropathogens: uropathogenic Escherichia coli (UPEC), Klebsiella pneumoniae and Enterococcus faecalis. While all three species invaded the bladder cells, under flow conditions the Gram-positive E. faecalis was significantly less invasive compared to the Gram-negative UPEC and K. pneumoniae. When introduced simultaneously during an infection experiment, all three bacterial species sometimes invaded the same bladder cell, at differing frequencies suggesting complex interactions between bacterial species and bladder cells. Inside host cells, we observed encasement of E. faecalis colonies specifically by UPEC. During subsequent dispersal from the host cells, only the Gram-negative bacteria underwent infection-related filamentation (IRF). Taken together, our data suggest that bacterial multispecies invasions of single bladder cells are frequent and support earlier studies showing intraspecies cooperation on a biochemical level during UTI., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Ethnopharmacological Relevance: Arctostaphylos uva-ursi (L.) Spreng. (bearberry) is a well-known traditional herbal plant used as a urinary tract disinfectant. Its antiseptic and diuretic properties can be attributed to hydroquinone, obtained by hydrolysis of arbutin., Aim of the Study: This study aimed to determine the toxic profile of free hydroquinone on urinary bladder cells (T24) as a target of therapeutic action., Materials and Methods: Quantitative and qualitative analysis of the extract and the digestive stability and bioavailability of arbutin and hydroquinone were performed by HPLC assay and simulated in vitro digestion, respectively. Cytotoxic effect, reactive oxygen species induction and proteome changes in T24 cells after hydroquinone treatment were determined using Neutral red assay, 2',7'-dichlorofluorescein-diacetate (DCFH-DA) assay and mass spectrometry, respectively., Results: Through in vitro digestion, arbutin was stable, but hydroquinone increased after pepsin treatment (109.6%) and then decreased after the small intestine phase (65.38%). The recommended doses of Uva-ursi had a cytotoxic effect on T24 cells only when all hydroquinone conjugates were converted to free hydroquinone (320 and 900 μg/mL) and the toxic effect was enhanced by recovery. One cup of the therapeutic dose had a prooxidative effect after 4 h of incubation. Shorter time of cell exposure (2 h) to hydroquinone did not have any impact on reactive oxygen species induction. Proteomic analysis found 17 significantly up-regulated proteins compared to control. Hydroquinone activated proteins related to oxidative stress response, stress-adaptive signalling, heat shock response and initiation of translation., Conclusions: Despite the therapeutic properties of bearberry, up-regulated T24 cell proteins are evidence that plant compounds, although from a natural source, may exhibit negative properties., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Single-cell RNA sequencing (scRNA-seq) data has been widely used to profile cellular heterogeneities with a high-resolution picture. Clustering analysis is a crucial step of scRNA-seq data analysis because it provides a chance to identify and uncover undiscovered cell types. Most methods for clustering scRNA-seq data use an unsupervised learning strategy. Since the clustering step is separated from the cell annotation and labeling step, it is not uncommon for a totally exotic clustering with poor biological interpretability to be generated-a result generally undesired by biologists. To solve this problem, we proposed an active learning (AL) framework for clustering scRNA-seq data. The AL model employed a learning algorithm that can actively query biologists for labels, and this manual labeling is expected to be applied to only a subset of cells. To develop an optimal active learning approach, we explored several key parameters of the AL model in the experiments with four real scRNA-seq datasets. We demonstrate that the proposed AL model outperformed state-of-the-art unsupervised clustering methods with less than 1000 labeled cells. Therefore, we conclude that AL model is a promising tool for clustering scRNA-seq data that allows us to achieve a superior performance effectively and efficiently., (© 2021. The Author(s), under exclusive licence to United States and Canadian Academy of Pathology.)

The inflammasome-associated proteins caspase-1, caspase-4 and NLRP3 have been emphasised to be essential in the host cell response during urinary tract infection (UTI) by regulating IL-1β release. Our aim was to investigate how the inflammasome-associated proteins regulate the cell response of bladder epithelial cells during infection with uropathogenic Escherichia coli (UPEC). Human bladder epithelial cells (5637) and CRISPR/Cas9 generated caspase-1, caspase-4 and NLRP3 knockdown cells were stimulated with the UPEC strain CFT073. Using Olink proteomics and real time RT-PCR, we showed that caspase-1, caspase-4 and NLRP3 are vital for the expression of many inflammatory genes and proteins from bladder epithelial cells. When investigating the effect of inflammasome-associated proteins on neutrophils, we found that conditioned medium from UPEC-infected caspase-4 knockdown cells significantly increased phagocytosis of CFT073 and significantly decreased ROS production from neutrophils. In contrast, conditioned medium from UPEC-infected NLRP3 knockdown cells significantly decreased the phagocytosis of CFT073 and significantly increased the ROS production from neutrophils. In conclusion, we showed that the inflammasome-associated proteins contribute to the host cell response during UPEC infection., (© 2022. The Author(s).)

Bladder cancer represents a highly heterogeneous disease characterized by distinct histological, molecular and clinical phenotypes, and a detailed analysis of tumor cell invasion and crosstalks within bladder tumor cells has not been determined. Here, we applied droplet-based single-cell RNA sequencing (scRNA-seq) to acquire transcriptional profiles of 36 619 single cells isolated from seven patients. Single cell transcriptional profiles matched well with the pathological basal/luminal subtypes. Notably, in T1 tumors diagnosed as luminal subtype, basal cells displayed characteristics of epithelial-mesenchymal transition (EMT) and mainly located at the tumor-stromal interface as well as micrometastases in the lamina propria. In one T3 tumor, muscle-invasive tumor showed significantly higher expression of cancer stem cell markers SOX9 and SOX2 than the primary tumor. We additionally analyzed communications between tumor cells and demonstrated its relevance to basal/luminal phenotypes. Overall, our single-cell study provides a deeper insight into the tumor cell heterogeneity associated with bladder cancer progression., (© 2021 UICC.)

Cell-based tissue engineering (TE) has been proposed to improve treatment outcomes in end-stage bladder disease, but TE approaches with 2D smooth muscle cell (SMC) culture have so far been unsuccessful. Here, we report the development of primary bladder-derived 3D SMC spheroids that outperform 2D SMC cultures in differentiation, maturation, and extracellular matrix (ECM) production. Bladder SMC spheroids were compared with 2D cultures using live-dead staining, qRT-PCR, immunofluorescence, and immunoblotting to investigate culture conditions, contractile phenotype, and ECM deposition. The SMC spheroids were viable for up to 14 days and differentiated rather than proliferating. Spheroids predominantly expressed the late myogenic differentiation marker MyH11, whereas 2D SMC expressed more of the general SMC differentiation marker α -SMA and less MyH11. Furthermore, the expression of bladder wall-specific ECM proteins in SMC spheroids was markedly higher. This first establishment and analysis of primary bladder SMC spheroids are particularly promising for TE because differentiated SMCs and ECM deposition are a prerequisite to building a functional bladder wall substitute. We were able to confirm that SMC spheroids are promising building blocks for studying detrusor regeneration in detail and may provide improved function and regenerative potential, contributing to taking bladder TE a significant step forward., Competing Interests: The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (Copyright © 2021 Tim Gerwinn et al.)

The urinary bladder is a highly dynamic organ that undergoes large deformations several times per day. Mechanical characteristics of the tissue are crucial in determining the function and dysfunction of the organ. Yet, literature reporting on the mechanical properties of human bladder tissue is scarce and, at times, contradictory. In this study, we focused on mechanically testing tissue from both human and pig bladders using identical protocols to validate the use of pigs as a model for the human bladder. Furthermore, we tested the effect of two treatments on tissue mechanical properties. Namely, elastase to digest elastin fibers, and oxybutynin to reduce smooth muscle cell spasticity. Additionally, mechanical properties based on the anatomical direction of testing were evaluated. We implemented two different material models to aid in the interpretation of the experimental results. We found that human tissue behaves similarly to pig tissue at high deformations (collagen-dominated behavior) while we detected differences between the species at low deformations (amorphous matrix-dominated behavior). Our results also suggest that elastin could play a role in determining the behavior of the fiber network. Finally, we confirmed the anisotropy of the tissue, which reached higher stresses in the transverse direction when compared to the longitudinal direction., (Copyright © 2021 by ASME.)

Postmenopausal women with ovary hormone deficiency (OHD) are subject to overactive bladder (OAB) symptoms. The present study attempted to elucidate whether low-intensity extracorporeal shock wave therapy (LiESWT) alters bladder angiogenesis, decreases inflammatory response, and ameliorates bladder hyperactivity to influence bladder function in OHD-induced OAB in human clinical trial and rat model. The ovariectomized (OVX) for 12 months Sprague-Dawley rat model mimicking the physiological condition of menopause was utilized to induce OAB and assess the potential therapeutic mechanism of LiESWT (0.12 mJ/mm 2 , 300 pulses, and 3 pulses/second). The randomized, single-blinded clinical trial was enrolled 58 participants to investigate the therapeutic efficacy of LiESWT (0.25 mJ/mm 2 , 3000 pulses, 3 pulses/second) on postmenopausal women with OAB. The results revealed that 8 weeks' LiESWT inhibited interstitial fibrosis, promoted cell proliferation, enhanced angiogenesis protein expression, and elevated the protein phosphorylation of ErK1/2, P38, and Akt, leading to decreased urinary frequency, nocturia, urgency, urgency incontinence, and post-voided residual urine volume, but increased voided urine volume and the maximal flow rate of postmenopausal participants. In conclusion, LiESWT attenuated inflammatory responses, increased angiogenesis, and promoted proliferation and differentiation, thereby improved OAB symptoms, thereafter promoting social activity and the quality of life of postmenopausal participants.

Volume accommodation occurs via a novel mechanism involving interstitial cells in detrusor muscles. The interstitial cells in the bladder are PDGFRα + , and they restrain the excitability of smooth muscle at low levels and prevents the development of transient contractions (TCs). A common clinical manifestation of spinal cord injury (SCI)-induced bladder dysfunction is detrusor overactivity (DO). Although a myogenic origin of DO after SCI has been suggested, a mechanism for development of SCI-induced DO has not been determined. In this study we hypothesized that SCI-induced DO is related to loss of function in the regulatory mechanism provided by PDGFRα + cells. Our results showed that transcriptional expression of Pdgfra and Kcnn3 was decreased after SCI. Proteins encoded by these genes also decreased after SCI, and a reduction in PDGFRα + cell density was also documented. Loss of PDGFRα + cells was due to apoptosis. TCs in ex vivo bladders during filling increased dramatically after SCI, and this was related to the loss of regulation provided by SK channels, as we observed decreased sensitivity to apamin. These findings show that damage to the mechanism restraining muscle contraction during bladder filling that is provided by PDGFRα + cells is causative in the development of DO after SCI., (© 2021. The Author(s).)

The male urogenital system is composed of the reproductive system and the urinary tract; they have an interconnected embryonic development and share one of their anatomical components, the urethra. This system has a highly complex physiology deeply interconnected with the circulatory and nervous systems, as well as being capable of adapting to environmental variations; it also undergoes changes with aging and, in the case of the reproductive system, with seasonality. The stroma is an essential component in this physiological plasticity and its complexity has increased with the description in the last decade of a new cell type, the telocyte. Several studies have demonstrated the presence of telocytes in the organs of the male urogenital system and other systems; however, their exact function is not yet known. The present review addresses current knowledge about telocytes in the urogenital system in terms of their locations, interrelationships, possible functions and pathological implications. It has been found that telocytes in the urogenital system possibly have a leading role in stromal tissue organization/maintenance, in addition to participation in stem cell niches and an association with the immune system, as well as specific functions in the urogenital system, lipid synthesis in the testes, erythropoiesis in the kidneys and the micturition reflex in the bladder. There is also evidence that telocytes are involved in pathologies in the kidneys, urethra, bladder, prostate, and testes., (© 2021 International Federation for Cell Biology.)

Background: Extracellular matrix (ECM) bioscaffolds produced by decellularization of source tissue have been effectively used for numerous clinical applications. However, decellularized tracheal constructs have been unsuccessful due to the immediate requirement of a functional airway epithelium on surgical implantation. ECM can be solubilized to form hydrogels that have been shown to support growth of many different cell types. The purpose of the present study is to compare the ability of airway epithelial cells to attach, form a confluent monolayer, and differentiate on homologous (trachea) and heterologous (urinary bladder) ECM substrates for potential application in full tracheal replacement., Materials and Methods: Porcine tracheas and urinary bladders were decellularized. Human bronchial epithelial cells (HBECs) were cultured under differentiation conditions on acellular tracheal ECM and urinary bladder matrix (UBM) bioscaffolds and hydrogels and were assessed by histology and immunolabeling for markers of ciliation, goblet cell formation, and basement membrane deposition., Results: Both trachea and urinary bladder tissues were successfully decellularized. HBEC formed a confluent layer on both trachea and UBM scaffolds and on hydrogels created from these bioscaffolds. Cells grown on tracheal and UBM hydrogels, but not on bioscaffolds, showed positive-acetylated tubulin staining and the presence of mucus-producing goblet cells. Collagen IV immunolabeling showed basement membrane deposition by these cells on the surface of the hydrogels., Conclusions: ECM hydrogels supported growth and differentiation of HBEC better than decellularized ECM bioscaffolds and show potential utility as substrates for promotion of a mature respiratory epithelium for regenerative medicine applications in the trachea., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Objectives: Much of the information to date in terms of subtypes and function of bladder urothelial cells were derived from anatomical location or by the expression of a small number of marker genes. To have a comprehensive map of the cellular anatomy of bladder urothelial cells, we performed single-cell RNA sequencing to thoroughly characterize mouse bladder urothelium., Materials and Methods: A total of 18,917 single cells from mouse bladder urothelium were analysed by unbiased single-cell RNA sequencing. The expression of the novel cell marker was confirmed by immunofluorescence using urinary tract infection models., Results: Unsupervised clustering analysis identified 8 transcriptionally distinct cell subpopulations from mouse bladder urothelial cells. We discovered a novel type of bladder urothelial cells marked by Plxna4 that may be involved with host response and wound healing. We also found a group of basal-like cells labelled by ASPM that could be the progenitor cells of adult bladder urothelium. ASPM + urothelial cells are significantly increased after injury by UPEC. In addition, specific transcription factors were found to be associated with urothelial cell differentiation. At the last, a number of interstitial cystitis/bladder pain syndrome-regulating genes were found differentially expressed among different urothelial cell subpopulations., Conclusions: Our study provides a comprehensive characterization of bladder urothelial cells, which is fundamental to understanding the biology of bladder urothelium and associated bladder disease., (© 2021 The Authors. Cell Proliferation Published by John Wiley & Sons Ltd.)

Basal and luminal subtypes of invasive bladder tumors have significant prognostic and predictive impacts for patients. However, it remains unclear whether tumor subtype commitment occurs in noninvasive urothelial lesions or in carcinoma in situ (CIS) and which gene pathways are important for bladder tumor progression. To understand the timing of this commitment, we used gene expression and protein analysis to create a global overview of 36 separate tissues excised from a whole bladder encompassing urothelium, noninvasive urothelial lesions, CIS, and invasive carcinomas. Additionally investigated were matched CIS, noninvasive urothelial lesions, and muscle-invasive bladder cancers (MIBC) from 22 patients. The final stage of subtype commitment to either a luminal or basal MIBC occurred at the CIS transition. For all tissues combined, hierarchical clustering of subtype gene expression revealed three subtypes: "luminal," "basal," and a "luminal p53-/extracellular matrix (ECM)-like" phenotype of ECM-related genes enriched in tumor-associated urothelium, noninvasive urothelial lesions, and CIS, but rarely invasive, carcinomas. A separate cohort of normal urothelium from noncancer patients showed significantly lower expression of ECM-related genes compared with tumor-associated urothelium, noninvasive urothelial lesions, and CIS. A PanCancer Progression Panel of 681 genes unveiled pathways specific for the luminal p53-/ECM-like cluster, for example, ECM remodeling, angiogenesis, epithelial-to-mesenchymal transition, cellular discohesion, cell motility involved in tumor progression, and cell proliferation and oncogenic ERBB2/ERBB3 signaling for invasive carcinomas. In conclusion, this study provides insights into bladder cancer subtype commitment and associated signaling pathways, which could help predict therapy response and enhance our understanding of therapy resistance. SIGNIFICANCE: This study demonstrates that CIS is the stage of commitment for determining MIBC tumor subtype, which is relevant for patient prognosis and therapy response., (©2021 American Association for Cancer Research.)

Background: Microscopic analysis of urine sediment is probably the most commonly used diagnostic procedure in nephrology. The urinary cells, however, have not yet undergone careful unbiased characterization., Methods: Single-cell transcriptomic analysis was performed on 17 urine samples obtained from five subjects at two different occasions, using both spot and 24-hour urine collection. A pooled urine sample from multiple healthy individuals served as a reference control. In total 23,082 cells were analyzed. Urinary cells were compared with human kidney and human bladder datasets to understand similarities and differences among the observed cell types., Results: Almost all kidney cell types can be identified in urine, such as podocyte, proximal tubule, loop of Henle, and collecting duct, in addition to macrophages, lymphocytes, and bladder cells. The urinary cell-type composition was subject specific and reasonably stable using different collection methods and over time. Urinary cells clustered with kidney and bladder cells, such as urinary podocytes with kidney podocytes, and principal cells of the kidney and urine, indicating their similarities in gene expression., Conclusions: A reference dataset for cells in human urine was generated. Single-cell transcriptomics enables detection and quantification of almost all types of cells in the kidney and urinary tract., (Copyright © 2021 by the American Society of Nephrology.)

Arsenic is a natural chemical element that is strongly associated with bladder cancer. Understanding the underlying mechanisms behind the association between arsenic and bladder cancer as well as identifying effective preventive interventions will help reduce the incidence and mortality of this disease. The epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) properties play key roles in cancer development and progression. Here, we reported that chronic exposure to arsenic resulted in EMT and increased levels of the CSC marker CD44 in human uroepithelial cells. Furthermore, IL-8 promoted a mesenchymal phenotype and upregulated CD44 by activating the ERK, AKT and STAT3 signaling. Phosphorylation of the human epidermal growth factor receptor 2 (HER2) was key for arsenic-induced IL-8 overexpression and depended on the simultaneous activation of the MAPK, JNK, PI3K/AKT and GSK3β signaling pathways. We also found that genistein inhibited arsenic-induced HER2 phosphorylation and downregulated its downstream signaling pathways, thereby inhibiting progression of EMT, and reducing CD44 expression levels. These results demonstrate that the HER2/IL-8 axis is related to the acquisition of an EMT phenotype and CSCs in arsenic-treated cells. The inhibitory effects of genistein on EMT and CSCs provide a new perspective for the intervention and potential chemotherapy against arsenic-induced bladder cancer., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Current organoid models are limited by their inability to mimic mature organ architecture and associated tissue microenvironments 1,2 . Here we create multilayer bladder 'assembloids' by reconstituting tissue stem cells with stromal components to represent an organized architecture with an epithelium surrounding stroma and an outer muscle layer. These assembloids exhibit characteristics of mature adult bladders in cell composition and gene expression at the single-cell transcriptome level, and recapitulate in vivo tissue dynamics of regenerative responses to injury. We also develop malignant counterpart tumour assembloids to recapitulate the in vivo pathophysiological features of urothelial carcinoma. Using the genetically manipulated tumour-assembloid platform, we identify tumoural FOXA1, induced by stromal bone morphogenetic protein (BMP), as a master pioneer factor that drives enhancer reprogramming for the determination of tumour phenotype, suggesting the importance of the FOXA1-BMP-hedgehog signalling feedback axis between tumour and stroma in the control of tumour plasticity.

Henry Miller stated that "to relieve a full bladder is one of the great human joys". Urination is critically important in health and ailments of the lower urinary tract cause high pathological burden. Although there have been advances in understanding the central circuitry in the brain that facilitates urination 1-3 , there is a lack of in-depth mechanistic insight into the process. In addition to central control, micturition reflexes that govern urination are all initiated by peripheral mechanical stimuli such as bladder stretch and urethral flow 4 . The mechanotransduction molecules and cell types that function as the primary stretch and pressure detectors in the urinary tract mostly remain unknown. Here we identify expression of the mechanosensitive ion channel PIEZO2 in lower urinary tract tissues, where it is required for low-threshold bladder-stretch sensing and urethral micturition reflexes. We show that PIEZO2 acts as a sensor in both the bladder urothelium and innervating sensory neurons. Humans and mice lacking functional PIEZO2 have impaired bladder control, and humans lacking functional PIEZO2 report deficient bladder-filling sensation. This study identifies PIEZO2 as a key mechanosensor in urinary function. These findings set the foundation for future work to identify the interactions between urothelial cells and sensory neurons that control urination.

Introduction: Bladder dysfunction has been considered as one of the most critical health conditions with no proper treatment. Current therapeutic approaches including enterocystoplasty have several limitations. Hence, biofabrication of cell-laden biological allografts using decellularized Goat urinary bladder scaffolds for organ reconstruction/regeneration was major objective of this study., Materials and Methods: An efficient method for decellularization of Goat urinary bladder (N = 3) was developed by perfusion of gradient change of detergents through ureter. The retention of organ architecture, extracellular matrix composition, mechanical properties and removal of cellular components was characterized using histological, cellular and molecular analysis. Further, mesenchymal stem cells (MSCs) from human umbilical cord blood (UCB) were used for preparing biological construct of decellularized urinary bladder (DUB) scaffolds to augment the urinary bladder reconstruction/regeneration., Results: The decellularization method adopted in this study generated completely DUB scaffolds within 10 h at 100 mm Hg pressure and constant flow rate of 1 mL/min. The DUB scaffold retains organ architecture, ECM composition, and mechanical strength. No significant amount of residual nucleic acid was observed post-decellularization. Furthermore, MSCs derived from human UCB engrafted and proliferated well on DUB scaffolds in highly aligned manner under xeno-free condition., Conclusion: Biofabricated humanized urinary bladder constructs provides xeno-free allografts for future application in augmenting urinary bladder reconstruction/regeneration with further development., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Aims: RacGTPase-mediated proliferation and smooth muscle contraction in the lower urinary tract has been recently suggested and may offer putative targets for treamtment of lower urinary tract symptoms. However, RacGTPase function for proliferation of detrusor smooth muscle cells is unknown and the specificity of Rac inhibitors has been questioned. Here, we examined effects of Rac1 knockdown and of the Rac inhibitors NSC23766 and EHT1864 in human bladder smooth muscle cells (hBSMCs)., Main Methods: Rac1 expression was silenced by shRNA expression. Effects of silencing and Rac inhibitors were assessed by CCK-8 assay, EdU staining, RT-PCR, colony formation assay, flow cytometry, and phalloidin staining., Key Findings: Silencing of Rac1 expression reduced the viability (up to 83% compared to scramble shRNA) and proliferation (virtually completely in proliferation assay), increased apoptosis (124%) and the number of dead cells (51%), and caused breakdown of actin organization (56% reduction of polymerized actin compared to scramble shRNA). Effects on proliferation, viability, and actin organization were mimicked by NSC23766 and EHT1864, while both compounds showed divergent effects on cell death (32-fold increase of dead cells by EHT1864, but not NSC23766). Effects of NSC23766 and EHT1864 on viability of hBSMCs were not altered by Rac1 knockdown., Significance: Rac1 promotes proliferation, viability, and cytoskeletal organization, and suppresses apoptosis in bladder smooth muscle cells, which may be relevant in overactive bladder or diabetes-related bladder dysfunction. NSC23766 and EHT1864 mimick these effects, but may act Rac1-independently, by shared and divergent effects., (Copyright © 2020. Published by Elsevier Inc.)

Extracellular matrix (ECM) accumulation plays a key role in the progression of bladder outlet obstruction (BOO). Muscarinic receptors have been widely reported to serve as pivotal regulators in lung tissue remodeling. However, the influence of them on human bladder smooth muscle cells (HBSMCs) and the underlying molecular mechanisms have not yet been evaluated. The purposes of the present study are to investigate the effect of muscarinic receptors on the synthesis of ECM in HBSMCs and the involvement of intracellular signal transducers. The results indicated that M 1 -M 5 muscarinic receptors were all encoded in HBSMCs. The expression rank order was M 2  > M 1  > M 5  > M 3  > M 4 . The gene and protein expression of collagen I (COL1), TIMP-1, and TIMP-2 was carbachol (CCH) concentration-dependently enhanced. The synthesis of COL1 in the supernatant of cell culture medium was significantly elevated by exposure to CCH. The CCH-induced protein expression of COL1, TIMP-1, and TIMP-2, however, was obviously reduced by the pretreatment of muscarinic receptor antagonists, atropine, and M 3 -preferring antagonist (1,1-dimethyl-4-diphenyl-acetoxypiperidinium iodide [4-DAMP]). Furthermore, ERK1/2 was activated by 100 µM CCH when compared with the control group and the pretreatment of ERK1/2 inhibitor significantly suppressed the synthesis of COL1 induced by 100 µM CCH. Besides, CCH-induced phosphorylation of ERK1/2 was remarkably restrained by the pretreatment of 4-DAMP. All in all, these findings demonstrated that M 3 receptor can modulate extracellular matrix synthesis via the ERK1/2 signaling pathway, which may provide potential novel therapeutic targets for BOO., (© 2020 Wiley Periodicals, Inc.)

Aster tataricus L.f. is a traditional Eastern Asian herbal medicine used for the relief of uroschesis-related illnesses and has been demonstrated clinically to exert satisfied effects. However, the mechanism of its therapeutic action remains unclear. The present study aimed to evaluate the protective mechanism of Aster tataricus extract (ATE) on CYP or LPS + ATP-induced interstitial cystitis (IC), we successfully constructed the induced IC Sprague-Dawley (SD) rat model and IC human urothelium cell (SV-HUC-1) model. The main compounds of ATE were determined by LC-MS. After intervention, the changes on the bladder wall morphology and inflammation were observed in each group. SV-HUC1 cell viability was measured by MTT and double stained with Hoechst 33342 and propidium iodide (PI). The expression levels of NLRP3, Pro-caspase-1, Caspsae-1 p20, GSDMD, GSDMD-N and Cleave-IL-1β in vivo and in vitro in different groups were detected by Western blotting. ATE significantly alleviated oedema and haemorrhage and reduced the inflammation index and histopathological score in SD rat bladder. The results of cell revealed that ATE could improve cell viability and decrease pyroptosis ratio. The expression of NLRP3 and other pyroptosis-related protein was remarkably decreased by ATE both in vivo and in vitro. ATE may be used as an inhibitor of NLRP3 in treating IC. The discovery of NLRP3/Caspase-1/GSDMD-N as a new protective pathway provides a new direction for protecting cell against IC., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Urinary tract infections (UTI) frequently progress to chronicity in infected individuals but the mechanisms of pathogenesis underlying chronic UTI are not well understood. We examined the role of interleukin (IL)-17A in UTI because this cytokine promotes innate defense against uropathogenic Escherichia coli (UPEC). Analysis of UPEC persistence and pyelonephritis in mice deficient in IL-17A revealed that UPEC CFT073 caused infection at a rate higher than the multidrug resistant strain EC958. Il17a -/- mice exhibited pyelonephritis with kidney bacterial burdens higher than those of wild-type (WT) mice. Synthesis of IL-17A in the bladder reflected a combination of γδ-T and T H 17 cell responses. Analysis of circulating inflammatory mediators at 24h postinoculation identified predictors of progression to chronicity, including IL-6 and monocyte chemoattractant protein-1 (MCP-1). Histological analysis identified infiltrating populations of neutrophils, NK cells, and γδ T cells in the bladder, whereas neutrophils predominated in the kidney. Analysis of the contribution of flagella to chronicity using hyper-flagellated and fliC-deficient UPEC in WT and Il17a -/- mice revealed that, in a host that is deficient for the production of IL-17A, flagella contribute to bacterial persistence. These findings show a role for IL-17A in defense against chronic UTI and a contribution of flagella to the pathogenesis of infection., (© 2020 Federation of American Societies for Experimental Biology.)

Objective: To determine histological aspects of decellularized bladder graft to achieve a double-sized bladder by novel hourglass technique; using rabbit models., Methods: Sixteen rabbit bladders were decellularized and underwent laboratory investigations. After making a laparotomy incision and exposure of bladders in another 16 rabbits (partial detrusor myomectomy), they were separated into two groups. The fundus of the decellularized scaffold was anastomosed to the fundus of the native bladder via the serosal layer, and the omentum and a double-J stent were placed in the decellularized bladder by no direct contact with the urine (Group A, n=8). In group B (n=8), the bladder was augmented applying the decellularized bladder that was in contact with the urine. After 6 months, the omentum was brought out of the neck of the engineered bladder and the anastomosis was opened. Biopsies were taken at 1, 3, and 9 months postoperatively., Results: Cell removal with preservation of extracellular matrix structure was confirmed in decellularized bladders. Histological examination after 1 month demonstrated few cells at the border of the grafts. After 3 months, the region of the graft was indistinguishable from the natural bladder with continuity of transitional epithelium of natural bladder on the decellularized grafted scaffolds. The organization of muscle layers was similar to native bladder muscle layers after 9 months. IHC staining markers were highly expressed after 9 months. Interestingly, bladders had a high fibrosis grade in group B compared with hourglass technique., Conclusion: We confirmed that decellularized bladder may be a reliable scaffold and viable material for bladder augmentation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: We assessed whether D2-40/podoplanin (PDPN) could be used to identify bladder cancer patients with a higher probability of benefiting from cisplatin-based combination chemotherapy., Patients and Methods: We investigated PDPN expression by immunohistochemical analysis of cystectomy specimens from 96 bladder cancer patients who had undergone radical cystectomy without neoadjuvant or adjuvant cisplatin-based combination chemotherapy until recurrence. We classified the cases into 2 groups according to the achievement of 2-year recurrence-free survival (RFS) and evaluated whether PDPN expression was associated with patient prognosis. We also classified the 96 cases into 3 groups according to the possible need for perioperative chemotherapy based on the response to chemotherapy after recurrence as "unnecessary" (achieving 2-year RFS), "responder" (recurring within 2 years and responding to chemotherapy after recurrence), and "non-responder" (not responding chemotherapy following recurrence) and compared PDPN expression between these groups., Results: Among 13 cases diagnosed with clinically

Recent applications of decellularized tissue have included the use of hydrogels for injectable materials and three-dimensional (3D) bioprinting bioink for tissue regeneration. Microvascular formation is required for the delivery of oxygen and nutrients to support cell growth and regeneration in tissues and organs. The aim of the present study was to evaluate the formation of capillary networks in decellularized extracellular matrix (d-ECM) hydrogels. The d-ECM hydrogels were obtained from the small intestine submucosa (SIS) and the urinary bladder matrix (UBM) after decellularizing with sodium deoxycholate (SDC) and high hydrostatic pressure (HHP). The SDC d-ECM hydrogel gradually gelated, while the HHP d-ECM hydrogel immediately gelated. All d-ECM hydrogels had low matrix stiffness compared to that of the collagen hydrogel, according to a compression test. D-ECM hydrogels with various elastic moduli were obtained, irrespective of the decellularization method or tissue source. Microvascular-derived endothelial cells were seeded on d-ECM hydrogels. Few cells attached to the SDC d-ECM hydrogel with no network formation, while on the HHP d-ECM hydrogel, a capillary network structure formed between elongated cells. Long, branched networks formed on d-ECM hydrogels with lower matrix stiffness. This suggests that the capillary network structure that forms on d-ECM hydrogels is closely related to the matrix stiffness of the hydrogel.

A new, rare trinor-dolabellane diterpenoid, sangiangol A ( 1 ), and one new dolabellane diterpenoid, sangiangol B ( 2 ), together with known cembranes and dolabellanes ( 3 - 8 ), were isolated from the ethyl acetate layer of an extract of an Indonesian marine soft coral, Anthelia sp. Compounds 1 - 8 exhibited moderate cytotoxicity against an NBT-T2 cell line (0.5-10 µg/mL). The structures of the new compounds were determined by analyzing their spectra and a molecular modelling study. A possible biosynthetic pathway for sangiangols A ( 1 ) and B ( 2 ) is presented. Cytotoxicity requires two epoxide rings or a chlorine atom, as in 4 (stolonidiol) and 5 (clavinflol B).

Rationale: Construction of functional vascularized three-dimensional tissues has been a longstanding objective in the field of tissue engineering. The efficacy of using a tissue expander capsule as an induced vascular bed to prefabricate functional vascularized smooth muscle tissue flaps for bladder reconstruction in a rabbit model was tested. Methods: Skin tissue expanders were inserted into the groin to induce vascularized capsule pouch formation. Smooth muscle cells and endothelial progenitor cells were harvested and cocultured to form pre-vascularized smooth muscle cell sheet. Then repeated transplantation of triple-layer cell sheet grafts onto the vascularized capsular tissue was performed at 2-day intervals to prefabricate functional vascularized smooth muscle tissue flaps. Bladder muscular wall defects were created and repaired by six-layer cell sheet graft (sheet only), capsule flap (capsule only) and vascularized capsule prelaminated with smooth muscle cell sheet (sheet plus capsule). The animals were followed for 3 months after implantation and their bladders were explanted serially. Results: Bladder capacity and compliance were maintained in sheet plus capsule group throughout the 3 months. Tissue bath stimulation demonstrated that contractile responses to carbachol and KCl among the three groups revealed a significant difference ( p < 0.05). Histologically, inflammation was evident in the capsule only group at 1 month and fibrosis was observed in sheet only group at 3 months. The vessel density in capsule only and sheet plus capsule group were significantly higher than in the sheet only group at each time point ( p < 0.05). Comparison of the smooth muscle content among the three groups revealed a significant difference ( p < 0.05). Conclusion: These results proved that the capsule may serve as an induced vascular bed for vascularized smooth muscle tissue flap prefabrication. The prefabricated functional vascularized smooth muscle tissue flap has the potential for reliable bladder reconstruction and may create new opportunities for vascularization in 3-D tissue engineering., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Congenital anomalies of the urinary tract are a significant cause of morbidity in infancy, and many congenital anomalies are linked to ureter development; however, the mechanism by which congenital anomalies control ureter development remains unknown. The loss of Robo2 can cause ureter defects and vesicoureteral reflux. However, how Robo2 impacts ureter development is unclear. We found that ROBO2 is expressed in the common nephric duct (CND) and primitive bladder, and impacts CND migration and fusion with the primitive bladder via its novel binding partner retinaldehyde dehydrogenase-2 (RALDH2). Delayed apoptosis that is due to the failure of CND fusion with the primitive bladder in the Robo2 -/- embryo results in an abnormal ureter connection to the CND, which is required for ureter development. We define a novel pathway in which the CND is remodeled by ROBO2 and retinoic acid rescued the ureter anomalies in the Robo2 -/- embryo. These findings may be relevant to diverse disease conditions that are associated with altered signaling in the primitive bladder., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Many important cell types in adult vertebrates have a mesenchymal origin, including fibroblasts and vascular mural cells. Although their biological importance is undisputed, the level of mesenchymal cell heterogeneity within and between organs, while appreciated, has not been analyzed in detail. Here, we compare single-cell transcriptional profiles of fibroblasts and vascular mural cells across four murine muscular organs: heart, skeletal muscle, intestine and bladder. We reveal gene expression signatures that demarcate fibroblasts from mural cells and provide molecular signatures for cell subtype identification. We observe striking inter- and intra-organ heterogeneity amongst the fibroblasts, primarily reflecting differences in the expression of extracellular matrix components. Fibroblast subtypes localize to discrete anatomical positions offering novel predictions about physiological function(s) and regulatory signaling circuits. Our data shed new light on the diversity of poorly defined classes of cells and provide a foundation for improved understanding of their roles in physiological and pathological processes.

Bladder dysfunction is characterized by urgency, frequency (pollakisuria, nocturia), and dysuria and may lead to urinary incontinence. Most of these symptoms can be attributed to disturbed bladder sensitivity. There is growing evidence that, besides the urothelium, suburothelial interstitial cells (suICs) are involved in bladder afferent signal processing. The massive expansion of the bladder during the filling phase implicates mechanical stress delivered to the whole bladder wall. Little is known about the reaction of suICs upon mechanical stress. Therefore, we investigated the effects of mechanical stimulation in cultured human suICs. We used fura-2 calcium imaging as a major physiological readout. We found spontaneous intracellular calcium activity in 75 % of the cultured suICs. Defined local pressure application via a glass micropipette led to local increased calcium activity in all stimulated suICs, spreading over the whole cell. A total of 51% of the neighboring cells in a radius of up to 100 µm from the stimulated cell showed an increased activity. Hypotonic ringer and shear stress also induced calcium transients. We found an 18-times increase in syncytial activity compared to unstimulated controls, resulting in an amplification of the primary calcium signal elicited in single cells by 50%. Our results speak in favor of a high sensitivity of suICs for mechanical stress and support the view of a functional syncytium between suICs, which can amplify and distribute local stimuli. Previous studies of connexin expression in the human bladder suggest that this mechanism could also be relevant in normal and pathological function of the bladder in vivo.

Bladder outlet obstruction (BOO), which is primarily caused by benign prostatic hyperplasia, is a common chronic disease. However, previous studies have most commonly investigated BOO using the acute obstruction model. In the present study, a chronic obstruction model was established to investigate the different pathological alterations in the bladder between acute and chronic obstruction. Compared with chronic obstruction, acute obstruction led to increased expression of proliferating cell nuclear antigen and interleukin‑1β, which are markers of proliferation and inflammation, respectively. Furthermore, increased fibrosis in the bladder at week 2 was observed. Low pressure promoted mice bladder smooth muscle cell (MBSMC) proliferation, and pressure overload inhibited cell proliferation and increased the proportion of dead MBSMCs. Further investigation using serum/glucocorticoid regulated kinase 1 (SGK1) small interfering RNAs indicated that low pressure may promote MBSMC proliferation by upregulating SGK1 and nuclear factor of activated T‑cell expression levels. Therefore, the present study suggested that acute obstruction led to faster decompensation of bladder function and chronic bladder obstruction displayed an enhanced ability to progress to BOO.

Purpose: To evaluate the effect of a new mesenchymal stem cell type derived from the neonatal bladder (nMSC-B) on diabetic bladder dysfunction (DBD)., Materials and Methods: nMSC-B were harvested from neonatal male Sprague-Dawley rat's bladder and expanded in culture. nMSC-B were transferred to Type-1 diabetic rats which were induced by a single dose 45 mg/kg Streptozocin (STZ). Stem cells were transferred via intraperitoneally (IP) (DM-IP group, n:6) and by direct injection to the detrusor (DM-D group, n:6) at 12th week following diabetes and compared with Phosphate Buffered Saline (PBS) injected diabetic rats (DM-PBS group, n:6) and age-matched PBS injected non-diabetic normal rats (NR-PBS group, n:6). All rats were evaluated histopathologically and functionally four weeks after the stem cell treatment., Results: nMSC-B showed improvement in both voiding function and bladder structure. The maximum voiding pressure (MVP) values in the DM-PBS group were lower compare to DM-IP, DM-D and NR-PBS groups (13.27 ± 0.78 vs 16.27 ± 0.61, 28.59 ± 2.09, 21.54 ± 1.00, respectively, P < .001). There was a significant improvement for MVP values in stem cell-treated groups. Immunohistochemical examination revealed decreased bladder smooth muscle (SM), increased fibrosis and desquamation in urothelia in diabetic groups compared to normal group(P < .001). We detected recovery in the stem cell groups. This recovery was more evident in DM-D group.  No statistical difference was observed in SM and fibrosis between DM-D and NR-PBS groups (P = .9)., Conclusion: It was shown that nMSCBs provided amelioration of DBD. We think that nMSC-B constitutes an effective treatment method in DBD.

The complex environment in the bladder weakens the efficacy of intravesical therapy. Hydrogel-based drug delivery systems are poised to revolutionize the delivery of therapeutic agents to bladder lesion sites. This forum article highlights the prospective applications of hydrogels as drug reservoirs in treating chronic bladder diseases., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The lower urinary tract has two main functions, namely, periodic urine storage and micturition; these functions are mediated through central and peripheral neuroregulation. Although extensive research on the lower urinary tract nervous system has been conducted, most studies have focused on primary culture. This protocol introduces a method for the isolation and culture of bladder neurons and glia from Sprague-Dawley rats. In this method, the neurons and glia were incubated in a 37 °C, 5% CO2 incubator for 5-7 days. As a result, they grew into mature shapes suitable for related subsequent immunofluorescence experiments. Cells were morphologically observed using an optical microscope. Neurons, synaptic vesicles, and glia were identified by β-III-tubulin and MAP-2, Synapsin-1, and GFAP staining, respectively. Meanwhile, immunocytochemistry was performed on several neurotransmitter-related proteins, such as choline acetyltransferase, DYNLL2, and SLC17A9.

Cancer cell detection and its stages recognition of life cycle are an important step to analyze cellular dynamics in the automation of cell based-experiments. In this work, a two-stage hierarchical method is proposed to detect and recognize different life stages of bladder cells by using two cascade Convolutional Neural Networks (CNNs). Initially, a hybrid object proposal algorithm (called EdgeSelective) by combining EdgeBoxes and Selective Search is proposed to generate candidate object proposals instead of a single Selective Search method in Region-CNN (R-CNN), and it can exploit the advantages of different mechanisms for generating proposals so that each cell in the image can be fully contained by at least one proposed region during the detection process. Then, the obtained cells from the previous step are used to train and extract features by employing CNNs for the purpose of cell life stage recognition. Finally, a series of comparison experiments are implemented. The results show that the proposed method can obtain better performance than traditional methods either in the stage of cell detection or cell life stage recognition, and it encourages and suggests the application in the development of new anticancer drug and cytopathology analysis of cancer patients in the near future.

Pericardium closure after cardiac surgery is recommended to prevent postoperative adhesions to the sternum. Synthetic materials have been used as substitutes, with limited results because of impaired remodeling and fibrotic tissue formation. Urinary bladder matrix (UBM) scaffolds promote constructive remodeling that more closely resemble the native tissue. The aim of the study is to evaluate the host response to UBM scaffolds in a porcine model of partial pericardial resection. Twelve Landrace pigs were subjected to a median sternotomy. A 5 × 7 cm pericardial defect was created and then closed with a 5 × 7 cm multilayer UBM patch (UBM group) or left as an open defect (control group). Animals were survived for 8 wk. End points included gross morphology, biomechanical testing, histology with semiquantitative score, and cardiac function. The UBM group showed mild adhesions, whereas the control group showed fibrosis at the repair site, with robust adhesions and injury to the coronary bed. Load at failure (gr) and stiffness (gr/mm) were lower in the UBM group compared with the native pericardium (199.9 ± 59.2 versus 405.3 ± 99.89 g, P = 0.0536 and 44.23 ± 15.01 versus 146.5 ± 24.38 g/mm, P = 0.0025, respectively). In the UBM group, the histology resembled native pericardial tissue, with neovascularization, neofibroblasts, and little inflammatory signs. In contrast, control group showed fibrotic tissue with mononuclear infiltrates and a lack of organized collagen fibers validated with a histologic score. Both groups had normal ultrasonography results without cardiac motility disorders. In this setting, UBM scaffolds showed appropriate features for pericardial repair, restoring tissue properties that could help reduce postsurgical adhesions and prevent its associated complications., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Adeno-associated virus (AAV) vectors have been recognized as promising tools for gene delivery. The bladder is a seemingly ideal organ for virus transfer, with easy access through the urethra enabling organ-specific delivery. However, achieving adequate transduction efficiency in the urothelium has been a major challenge because of the barrier function of the glycosaminoglycan (GAG) layer. We investigated optimal pretreatments of the bladder urothelium to maximize transduction efficiency by AAV vectors in vivo. Murine bladders were pretreated with five different chemical agents followed by transurethral instillation with an AAV2 vector encoding a tdTOMATO reporter. After 7 days, transduction efficiency of the urothelium was evaluated. Bladder urothelia pretreated with HCl showed clear evidence of AAV infection and gene delivery. Mice treated with 0.1 N HCl for 4 min showed significantly higher survival rates (nearly 80 %) compared with mice receiving other pretreatment regimens. AAV vector transduction in the urothelium was observed in seven of 20 mice (35 %), and the mean transduction efficiency in these mice was 14.5 %. Thus, HCl pretreatment enhanced transduction efficiency of the mice bladder urothelium by an AAV vector in vivo. Pretreatment with 0.1 N HCl for 4 min was the optimal condition to maximize survival and transduction efficiency of the urothelium., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

TagB, TagC ( t andem a utotransporter g enes B and C ), and Sha ( S erine-protease h emagglutinin a utotransporter) are recently described members of the SPATE (serine protease autotransporters of Enterobacteriaceae ) family. These SPATEs can cause cytopathic effects on bladder cells and contribute to urinary tract infection in a mouse model. Bladder epithelial cells form an important barrier in the urinary tract. Some SPATEs produced by pathogenic E. coli are known to breach the bladder epithelium. The capacity of these newly described SPATEs to alter bladder epithelial cells and the role of the serine protease active site were investigated. All three SPATE proteins were internalized by bladder epithelial cells and altered the distribution of actin cytoskeleton. Sha and TagC were also shown to degrade mucin and gelatin respectively. Inactivation of the serine catalytic site in each of these SPATEs did not affect secretion of the SPATEs from bacterial cells, but abrogated entry into epithelial cells, cytotoxicity, and proteolytic activity. Thus, our results show that the serine catalytic triad of these proteins is required for internalization in host cells, actin disruption, and degradation of host substrates such as mucin and gelatin.

Previous studies have shown that alterations in the mechanical properties of cells may be associated with the onset and progression of some forms of pathology. In this paper, an experimental study of two types of cells, renal (cancer) and bladder (cancer) cells, is described which used acoustic radiation force (ARF) generated by a high-frequency ultrasound focusing transducer and performed on the operating platform of an inverted light microscope. Comparing images of cancer cells with those of normal cells of the same kind, we find that the cancer cells are more prone to deform than normal cells of the same kind under the same ARF. In addition, cancer cells with higher malignancy are more deformable than those with lower malignancy. This means that the deformability of cells may be used to distinguish diseased cells from normal ones, and more aggressive cells from less aggressive ones, which may provide a more rapid and accurate method for clinical diagnosis of urological disease in the future.

Iatrogenic ureteral injury is a dreaded complication of abdominal and pelvic surgeries, and thus, intraoperative identification of ureters is of paramount importance but lacks efficient methods and probes. Herein, we used near-infrared II (NIR-II, 1000-1700 nm) fluorescence imaging with advantages of higher spatial resolution, deeper tissue penetration, lower light scattering, and less tissue autofluorescence to identify ureters by aggregation-induced emission luminogen dots (AIE dots). The intraoperative ureteral injuries and common ureteral diseases can be visualized timely and precisely. Due to the longer emission wavelength and higher quantum yield of the AIE dots, it largely outperforms the commercial indocyanine green dye in brightness and penetration depth. It was the first time to realize the intraoperative identification of ureters in vivo using NIR-II imaging. Thus, our work provides a new platform for intraoperative monitoring during clinical operation.