Urinary Bladder Cancer Induced by N-Butyl-N-(4-Hydroxybutyl)-Nitrosamine.

Urinary bladder cancer is the tenth most common cancer worldwide with high morbidity and mortality. The majority of bladder cancers are urothelial carcinomas. More than half are papillomas or the papillary urothelial carcinomas (stages Ta and T1), which have a relatively good prognosis. Squamous cell carcinomas have a variable survival rate, while carcinomas in situ (Tis) can progress to muscle-invasive urothelial carcinomas (T2) with a poor prognosis. The most challenging feature of bladder cancer is its high recurrence rate, ranging from 50% to 90% of cases. The N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN) model is an invaluable experimental tool for bladder cancer research, as BBN-induced bladder cancer in rodents resembles human bladder cancer in its morphological, biological, and molecular features. We present here a detailed protocol for the treatment of mice and the main expected results.

Selective targeting of lectins and their macropinocytosis in urothelial tumours: translation from in vitro to ex vivo.

Urinary bladder cancer can be treated by intravesical application of therapeutic agents, but the specific targeting of cancer urothelial cells and the endocytotic pathways of the agents are not known. During carcinogenesis, the superficial urothelial cells exhibit changes in sugar residues on the apical plasma membranes. This can be exploited for selective targeting from the luminal side of the bladder. Here we show that the plant lectins Jacalin (from Artocarpus integrifolia), ACA (from Amaranthus caudatus) and DSA (from Datura stramonium) selectively bind to the apical plasma membrane of low- (RT4) and high-grade (T24) cancer urothelial cells in vitro and urothelial tumours ex vivo. The amount of lectin binding was significantly different between RT4 and T24 cells. Endocytosis of lectins was observed only in cancer urothelial cells and not in normal urothelial cells. Transmission electron microscopy analysis showed macropinosomes, endosome-like vesicles and multivesicular bodies filled with lectins in RT4 and T24 cells and also in cells of urothelial tumours ex vivo. Endocytosis of Jacalin and ACA in cancer cells was decreased in vitro after addition of inhibitor of macropinocytosis 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and increased after stimulation of macropinocytosis with epidermal growth factor (EGF). Clathrin, caveolin and flotillin did not colocalise with lectins. These results confirm that the predominant mechanism of lectin endocytosis in cancer urothelial cells is macropinocytosis. Therefore, we propose that lectins in combination with conjugated therapeutic agents are promising tools for improved intravesical therapy by targeting cancer cells.

Combined Lectin- and Immuno-histochemistry (CLIH) for Fluorescence Microscopy.

The function of glycoproteins depends both on their polypeptide chain and sugar residues. For detection and localization of glycoproteins in tissue sections, methods of immunohistochemistry (IHC) and lectin histochemistry (LHC) are usually used separately. For a better understanding of the expression and distribution of variants of glycoproteins, tissue sections can be analyzed by combined lectin- and immuno-histochemistry (CLIH). CLIH exploits the advantages of both IHC and LHC and can therefore contribute to research in glycobiology and other fields of cell biology. Since cancer transformation is accompanied by alterations in the glycosylation of some glycoproteins, CLIH could also be exploited for improved classification of cancers. The chapter considers how CLIH could be employed on paraffin sections and semithin cryosections for fluorescence microscopy. Five different protocols of CLIH are described in detail as well as appropriate negative controls.

Nanodiamonds as Possible Tools for Improved Management of Bladder Cancer and Bacterial Cystitis.

Nanodiamonds (NDs) are a class of carbon nanomaterials with sizes ranging from a few nm to micrometres. Due to their excellent physical, chemical and optical properties, they have recently attracted much attention in biomedicine. In addition, their exceptional biocompatibility and the possibility of precise surface functionalisation offer promising opportunities for biological applications such as cell labelling and imaging, as well as targeted drug delivery. However, using NDs for selective targeting of desired biomolecules within a complex biological system remains challenging. Urinary bladder cancer and bacterial cystitis are major diseases of the bladder with high incidence and poor treatment options. In this review, we present: (i) the synthesis, properties and functionalisation of NDs; (ii) recent advances in the study of various NDs used for better treatment of bladder cancer and (iii) bacterial cystitis; and (iv) the use of NDs in theranostics of these diseases.

Transmission electron microscopy demonstration of reduced endothelial glycocalyx in severe preeclampsia.

INTRODUCTION: Endothelial glycocalyx is a carbohydrate-rich layer lining the luminal side of blood vessels. Its damage was demonstrated in different groups of critically ill patients. Indirect evidence showed that endothelial glycocalyx degradation might be an important factor in pathophysiology of preeclampsia. The aim of our study was to demonstrate endothelial glycocalyx by transmission electron microscopy and to compare its amount in the omentum vessels of pregnant patients with severe preeclampsia and two control groups. METHODS: Patients with severe preeclampsia who had a cesarean section were included in the study. Controls were healthy pregnant people at term with an elective cesarean section and non-pregnant patients of reproductive age who underwent laparoscopy for benign conditions. We performed omentum biopsies in all three groups. Samples were prepared for transmission electron microscopy using perfusion with ruthenium red. We measured the amount of endothelial glycocalyx attached to apical plasma membrane of endothelial cells as the area of glycocalyx observed with transmission electron microscope. RESULTS: In the analysis we included nine patients from each group and demonstrated statistically significant difference in the amount of endothelial glycocalyx among the three groups (p = 0.018). Glycocalyx was significantly reduced in severe preeclampsia (median 1.90 µm2, interquartile range 0.80-4.1 µm2) compared to non-pregnant controls (median 14.34 µm2, interquartile range 3.80-73.32 µm2); p = 0.021. A trend towards reduced glycocalyx amount in preeclampsia vs. pregnant controls and pregnant controls vs. non-pregnant controls was observed but without statistical significance. DISCUSSION: Compared to non-pregnant controls the endothelial glycocalyx was significantly reduced in pregnant patients with severe preeclampsia.

Vitamin A and Retinoids in Bladder Cancer Chemoprevention and Treatment: A Narrative Review of Current Evidence, Challenges and Future Prospects.

Bladder cancer (BC) is the tenth most common cancer worldwide with a high recurrence rate, morbidity and mortality. Therefore, chemoprevention and improved treatment of BC are of paramount importance. Epidemiological studies suggest that adequate vitamin A intake may be associated with reduced BC risk. In addition, retinoids, natural and synthetic derivatives of vitamin A, are intensively studied in cancer research due to their antioxidant properties and their ability to regulate cell growth, differentiation, and apoptosis. Findings from in vivo and in vitro models of BC show great potential for the use of retinoids in the chemoprevention and treatment of BC. However, translation to the clinical practice is limited. In this narrative review we discuss: (i) vitamin A and retinoid metabolism and retinoic acid signalling, (ii) the pathobiology of BC and the need for chemoprevention, (iii) the epidemiological evidence for the role of dietary vitamin A in BC, (iv) mechanistic insights obtained from in vivo and in vitro models, (v) clinical trials of retinoids and the limitations of retinoid use, (vi) novel systems of retinoid delivery, and (vii) components of retinoid signalling pathways as potential novel therapeutic targets.

Immunohistochemistry as a paramount tool in research of normal urothelium, bladder cancer and bladder pain syndrome.

The urothelium, an epithelium of the urinary bladder, primarily functions as blood-urine permeability barrier. The urothelium has a very slow turn-over under normal conditions but is capable of extremely fast response to injury. During regeneration urothelium either restores normal function or undergoes altered differentiation pathways, the latter being the cause of several bladder diseases. In this review, we describe the structure of the apical plasma membrane that enables barrier function, the role of urothelium specific proteins uroplakins and the machinery for polarized membrane transports in terminally differentiated superficial umbrella cells. We address key markers, such as keratins, cancer stem cell markers, retinoic acid signalling pathway proteins and transient receptor potential channels and purinergic receptors that drive normal and altered differentiation in bladder cancer and bladder pain syndrome. Finally, we discuss uncertainties regarding research, diagnosis and treatment of bladder pain syndrome. Throughout the review, we emphasise the contribution of immunohistochemistry in advancing our understanding of processes in normal and diseased bladder as well as the most promising possibilities for improved bladder cancer and bladder pain syndrome management.

Combined lectin- and immuno-histochemistry (CLIH) for applications in cell biology and cancer diagnosis: Analysis of human urothelial carcinomas.

Lectin histochemistry (LHC) and immunohistochemistry (IHC), which demonstrate the composition and localisation of sugar residues and proteins in cell membranes, respectively, are generally used separately. Using these two methods, we previously demonstrated that malignant transformation of urothelial cells results in the alterations of protein glycosylation and reduced expression of urothelium-specific integral membrane proteins uroplakins (UPs). However, the correlation between these changes was not studied yet. To evaluate this correlation, we developed innovative method, which we named combined lectin- and immuno- histochemistry (CLIH). We used human biopsies of 6 normal urothelia and 9 papillary urothelial carcinomas, i.e. 3 papillary urothelial neoplasms of low malignant potential (PUNLMP), 3 non-invasive papillary urothelial carcinomas of low grade (pTa, l.g.), and 3 invasive papillary urothelial carcinomas of high grade (pT1, h.g.). We tested five different protocols (numbered 1-5) of CLIH on paraffin and cryo-semithin sections and compared them with LHC and IHC performed separately. Additionally, we carried out western and lectin blotting with antibodies against UPs and lectins Amaranthus caudatus agglutinin (ACA), Datura stramonium agglutinin (DSA), and jacalin, respectively. We showed that incubation with primary antibodies first, followed by the mixture of secondary antibodies and lectins is the most efficient CLIH method (protocol number 5). Additionally, 300 nm thick cryo-semithin sections enabled better resolution of co-localisation between sugar residues and proteins than 5 µm thick paraffin sections. In the normal urothelium, CLIH showed co-localisation of lectins ACA and jacalin with UPs in the apical plasma membrane (PM) of superficial umbrella cells. In papillary urothelial carcinomas, all three lectins (ACA, DSA and jacalin) labelled regions of apical PM, where they occasionally co-localised with UPs. Western and lectin blotting confirmed the differences between normal urothelium and papillary urothelial carcinomas. Our results show that CLIH, when used with various sets of lectins and antigens, is a useful, quick, and reliable method that could be applied for basic cell biology research as well as detailed subtyping of human urothelial carcinomas.

Vitamin A Rich Diet Diminishes Early Urothelial Carcinogenesis by Altering Retinoic Acid Signaling.

Urinary bladder cancer is one of the leading malignancies worldwide, with the highest recurrence rates. A diet rich in vitamin A has proven to lower the risk of cancer, yet the molecular mechanisms underlying this effect are unknown. We found that vitamin A decreased urothelial atypia and apoptosis during early bladder carcinogenesis induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Vitamin A did not alter urothelial cell desquamation, differentiation, or proliferation rate. Genes like Wnt5a, involved in retinoic acid signaling, and transcription factors Pparg, Ppara, Rxra, and Hoxa5 were downregulated, while Sox9 and Stra6 were upregulated in early urothelial carcinogenesis. When a vitamin A rich diet was provided during BBN treatment, none of these genes was up- or downregulated; only Lrat and Neurod1 were upregulated. The lecithin retinol acyltransferase (LRAT) enzyme that produces all-trans retinyl esters was translocated from the cytoplasm to the nuclei in urothelial cells as a consequence of BBN treatment regardless of vitamin A rich diet. A vitamin A-rich diet altered retinoic acid signaling, decreased atypia and apoptosis of urothelial cells, and consequently diminished early urothelial carcinogenesis.

Lectins as possible tools for improved urinary bladder cancer management.

Urinary bladder cancer is the ninth most common cancer in developed countries with poor prognosis and outcome for the patient due to the challenging diagnosis and limited treatment possibilities. Bladder cancer arises mainly from urothelial cells lining the lumen. Urothelial cells form a three- to five-layered urothelium, which maintains the blood-urine barrier. The carbohydrates that cover the apical surface of superficial urothelial cells, i.e. umbrella cells, are crucial for this function. The composition of the carbohydrate covering is altered during urothelial cancer transformation. These bladder cancer-associated carbohydrate changes are a promising field for diagnosis, therapy and management. Lectins, which are carbohydrate-binding proteins, can be used to detect subtle alterations in carbohydrate composition during urothelial cancer transformation. Extensive research into various lectin applications has already been conducted, but the results are often contradictory and confusing. None of these applications have reached clinical trials. We review the literature and discuss (i) current bladder cancer management, (ii) lectin-based assays for detection of various cancer subtypes, (iii) lectin-based strategies for innovative bladder cancer treatment and finally (iv) lectins in nanotheranostics for personalized bladder cancer management.

Detonation nanodiamonds are promising nontoxic delivery system for urothelial cells.

Detonation nanodiamonds (DNDs) are carbon-based nanomaterials that are among the most promising nanoparticles available for biomedical applications so far. This is due to their biocompatibility, which could be contributed to their inert core and conformable surface nature. However, DNDs cytotoxicity for urothelial cells and the routes of their internalization remains an open question in the aspect of nanodiamond surface. We therefore analyzed four types of DNDs for cytotoxicity and internalization with normal urothelial cells and two types of cancer urothelial cell lines in vitro. Viability of any of the cell types we used was not compromised with any of four DNDs we evaluated after 24-, 48- and 72-h incubation in three different concentrations of DNDs. Transmission electron microscopy revealed that all four types of DNDs were endocytosed into all three types of urothelial cells tested here. We observed DNDs in endosomes, as well as in multivesicular bodies and multilamellar bodies. These results propose using of DNDs as a delivery system for urological applications in human nanomedicine.

Co-culturing porcine normal urothelial cells, urinary bladder fibroblasts and smooth muscle cells for tissue engineering research.

New strategies for culturing and co-culturing of the main types of urinary bladder cells are essential for successful establishment of biomimetic in vitro models, which could be applied for research into, and management of, diverse urological disorders. Porcine normal urothelial cells are available in nearly unlimited amounts and have many properties equivalent to human urothelial cells. In the present study, we established normal differentiated porcine urothelial cells in co-cultures with porcine urinary bladder normal fibroblasts and/or smooth muscle cells. The optimal culture medium for establishment of differentiated urothelial cells, demonstrated by positive immunofluorescence of uroplakins, cytokeratins (CK 7, CK 20), zonula occludens 1 (ZO-1), claudin 4, claudin 8, and E-cadherin, was the medium composed of equal parts of Advanced Dulbecco's modified Eagle's medium (A-DMEM) and MCDB 153 medium with physiological calcium concentration of 2.5 mM and without fetal bovine serum, named UroM (+Ca2+ - S). This medium was also proven to be suitable for culturing of bladder fibroblasts and smooth muscle cells and co-culturing of urothelial cells with these mesenchymal cells. Urothelial cell differentiation was optimal in UroM (+Ca2+ - S) medium in all co-culture conditions and when compared to all conditioned-media combinations. To summarize, these strategies for culturing and co-culturing of urinary bladder urothelial cells with mesenchymal cells could be used as new in vitro models for future basic and applicable research of the urinary bladder and thus potentially also for translational tissue engineering studies.

How to obtain good morphology and antigen detection in the same tissue section?

Most human and animal biopsy samples are routinely embedded in paraffin since this enables the pathologist or researcher to obtain excellent morphology and simplifies storage. Nevertheless, in many cases, the antigen of interest cannot be detected in paraffin section. The alternative available for good immunohistochemistry is preparation of cryosections, which usually provide decent antigen preservation and are frequently used for immunofluorescence. However, cryosections often do not provide efficient morphological details of tissues and cells for pathologic evaluation. In order to obtain good antigen preservation and improve tissue and cell morphology after freezing, we tested three different fixations and freezing methodologies and compared them to routine formaldehyde fixation and paraffin embedding. As a model system, we selected the epithelium of the rat urinary bladder and trachea. On all samples, haematoxylin and eosin staining was performed as well as immunofluorescence with antibodies against tight junction protein ZO-1 and against intermediate filament cytokeratin 7. The best compromise between morphology and immunofluorescence was obtained with "sucrose impregnation prior to freezing" method. Moreover, this procedure is also quicker in comparison to standard paraffin section preparation. To check the clinical relevance of our study, this method was used for human biopsy samples of neoplastic urothelial and bronchial mucosa lesions. Besides good immunofluorescence results, the morphology of these samples was well preserved. We therefore propose that cryosection preparation with sucrose impregnation prior to freezing should be further exploited in other clinical and veterinary applications, since it enables good morphology and antigen preservation.

Biocompatibility of different nanostructured TiO2 scaffolds and their potential for urologic applications.

Despite great efforts in tissue engineering of the ureter, urinary bladder, and urethra, further research is needed in order to improve the patient's quality of life and minimize the economic burden of different lower urinary tract disorders. The nanostructured titanium dioxide (TiO2) scaffolds have a wide range of clinical applications and are already widely used in orthopedic or dental medicine. The current study was conducted to synthesize TiO2 nanotubes by the anodization method and TiO2 nanowires and nanospheres by the chemical vapor deposition method. These scaffolds were characterized with scanning electron microscopy (SEM) and X-ray diffraction (XRD) methods. In order to test the urologic applicability of generated TiO2 scaffolds, we seeded the normal porcine urothelial (NPU) cells on TiO2 nanotubes, TiO2 nanowires, TiO2 nanospheres, and on the standard porous membrane. The viability and growth of the cells were monitored everyday, and after 3 weeks of culturing, the analysis with scanning electron microscope (SEM) was performed. Our results showed that the NPU cells were attached on all scaffolds; they were viable and formed a multilayered epithelium, i.e., urothelium. The apical plasma membrane of the majority of superficial NPU cells, grown on all three different TiO2 scaffolds and on the porous membrane, exhibited microvilli; thus, indicating that they were at a similar differentiation stage. The maximal caliper diameter measurements of superficial NPU cells revealed significant alterations, with the largest cells being observed on nanowires and the smallest ones on the porous membrane. Our findings indicate that different nanostructured TiO2 scaffolds, especially nanowires, have a great potential for tissue engineering and should be further investigated for various urologic applications.

The effect of LDL particles on the behaviour of epithelial noncancer and cancer cell lines after in vitro induced injury.

Cell spreading capability and cell proliferation are the major processes in wound healing of injured epithelia as well as in tumour progression. The effect of low-density lipoprotein (LDL) particles as a major extracellular source of cholesterol was evaluated in the re-epithelisation assay of in vitro induced injury. We selected two noncancer cell lines with different dependence on LDL concentrations, the kidney epithelial cells (MDCK) with higher dependence and keratinocytes (HaCaT) with lower dependence on LDL, and three cancer cell lines originating from epithelial cells: A549 (alveolar), CaCo-2 (intestinal) and RT4 (urothelial). All cells were incubated in a control medium, in an LDL-enriched medium or in an LDL-deficient medium. The LDL-enriched medium stimulated cell spreading of MDCK cells which, together with increased proliferation of these cells, resulted in an enhanced re-epithelisation of in vitro induced injury. LDL deficiency caused lower cell spreading which resulted in a decreased re-epithelisation despite the higher proliferation of MDCK cells in this medium. The re-epithelisation of keratinocytes (HaCaT) was not affected by altered LDL concentrations. In cancer cell lines A549, CaCo-2 and RT4, wide heterogeneity regarding cell proliferation and spreading capability was observed after treatment with different LDL concentrations. LDL had no influence on actin filament and tight junction distribution in any of the tested cell lines. The cholesterol content of all cell types, except for CaCo-2 cells, proved to be independent of the LDL level. Further research of the beneficial effects of LDL is needed to prove LDL as a safe enhancer of epithelial wound healing.

Signalling molecules in the urothelium.

The urothelium was long considered to be a silent barrier protecting the body from the toxic effects of urine. However, today a number of dynamic abilities of the urothelium are well recognized, including its ability to act as a sensor of the intravesical environment. During recent years several pathways of these urothelial abilities have been proposed and a major part of these pathways includes release of signalling molecules. It is now evident that the urothelium represents only one part of the sensory web. Urinary bladder signalling is finely tuned machinery of signalling molecules, acting in autocrine and paracrine manner, and their receptors are specifically distributed among different types of cells in the urinary bladder. In the present review the current knowledge of the formation, release, and signalling effects of urothelial acetylcholine, ATP, adenosine, and nitric oxide in health and disease is discussed.

Molecular ultrastructure of the urothelial surface: insights from a combination of various microscopic techniques.

The urothelium forms the blood-urine barrier, which depends on the complex organization of transmembrane proteins, uroplakins, in the apical plasma membrane of umbrella cells. Uroplakins compose 16 nm intramembrane particles, which are assembled into urothelial plaques. Here we present an integrated survey on the molecular ultrastructure of urothelial plaques in normal umbrella cells with advanced microscopic techniques. We analyzed the ultrastructure and performed measurements of urothelial plaques in the normal mouse urothelium. We used field emission scanning electron microscopy (FESEM), atomic force microscopy (AFM), transmission electron microscopy (TEM) on immunolabeled ultrathin sections (immuno-TEM), and freeze-fracture replicas (FRIL). We performed immunolabeling of uroplakins for scanning electron microscopy (immuno-FESEM). All microscopic techniques revealed a variability of urothelial plaque diameters ranging from 332 to 1179 nm. All immunolabeling techniques confirmed the presence of uroplakins in urothelial plaques. FRIL showed the association of uroplakins with 16 nm intramembrane particles and their organization into plaques. Using different microscopic techniques and applied qualitative and quantitative evaluation, new insights into the urothelial apical surface molecular ultrastructure have emerged and may hopefully provide a timely impulse for many ongoing studies. The combination of various microscopic techniques used in this study shows how these techniques complement one another. The described advantages and disadvantages of each technique should be considered for future studies of molecular and structural membrane specializations in other cells and tissues.

Cystitis: from urothelial cell biology to clinical applications.

Cystitis is a urinary bladder disease with many causes and symptoms. The severity of cystitis ranges from mild lower abdominal discomfort to life-threatening haemorrhagic cystitis. The course of disease is often chronic or recurrent. Although cystitis represents huge economical and medical burden throughout the world and in many cases treatments are ineffective, the mechanisms of its origin and development as well as measures for effective treatment are still poorly understood. However, many studies have demonstrated that urothelial dysfunction plays a crucial role. In the present review we first discuss fundamental issues of urothelial cell biology, which is the core for comprehension of cystitis. Then we focus on many forms of cystitis, its current treatments, and advances in its research. Additionally we review haemorrhagic cystitis with one of the leading causative agents being chemotherapeutic drug cyclophosphamide and summarise its management strategies. At the end we describe an excellent and widely used animal model of cyclophosphamide induced cystitis, which gives researches the opportunity to get a better insight into the mechanisms involved and possibility to develop new therapy approaches.

Correlation between urothelial differentiation and sensory proteins P2X3, P2X5, TRPV1, and TRPV4 in normal urothelium and papillary carcinoma of human bladder.

Terminal differentiation of urothelium is a prerequisite for blood-urine barrier formation and enables normal sensory function of the urinary bladder. In this study, urothelial differentiation of normal human urothelium and of low and high grade papillary urothelial carcinomas was correlated with the expression and localization of purinergic receptors (P2X3, and P2X5) and transient receptor potential vanilloid channels (TRPV1, and TRPV4). Western blotting and immunofluorescence of uroplakins together with scanning electron microscopy of urothelial apical surface demonstrated terminal differentiation of normal urothelium, partial differentiation of low grade carcinoma, and poor differentiation of high grade carcinoma. P2X3 was expressed in normal urothelium as well as in low grade carcinoma and in both cases immunolabeling was stronger in the superficial cells. P2X3 expression decreased in high grade carcinoma. P2X5 expression was detected in normal urothelium and in high grade carcinoma, while in low grade carcinoma its expression was diminished. The expression of TRPV1 decreased in low grade and even more in high grade carcinoma when compared with normal urothelium, while TRPV4 expression was unchanged in all samples. Our results suggest that sensory proteins P2X3 and TRPV1 are in correlation with urothelial differentiation, while P2X5 and TRPV4 have unique expression patterns.

Selective binding of lectins to normal and neoplastic urothelium in rat and mouse bladder carcinogenesis models.

Bladder cancer adjuvant intravesical therapy could be optimized by more selective targeting of neoplastic tissue via specific binding of lectins to plasma membrane carbohydrates. Our aim was to establish rat and mouse models of bladder carcinogenesis to investigate in vivo and ex vivo binding of selected lectins to the luminal surface of normal and neoplastic urothelium. Male rats and mice were treated with 0.05 % N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in drinking water and used for ex vivo and in vivo lectin binding experiments. Urinary bladder samples were also used for paraffin embedding, scanning electron microscopy and immunofluorescence labelling of uroplakins. During carcinogenesis, the structure of the urinary bladder luminal surface changed from microridges to microvilli and ropy ridges and the expression of urothelial-specific glycoproteins uroplakins was decreased. Ex vivo and in vivo lectin binding experiments gave comparable results. Jacalin (lectin from Artocarpus integrifolia) exhibited the highest selectivity for neoplastic compared to normal urothelium of rats and mice. The binding of lectin from Amaranthus caudatus decreased in rat model and increased in mouse carcinogenesis model, indicating interspecies variations of plasma membrane glycosylation. Lectin from Datura stramonium showed higher affinity for neoplastic urothelium compared to the normal in rat and mouse model. The BBN-induced animal models of bladder carcinogenesis offer a promising approach for lectin binding experiments and further lectin-mediated targeted drug delivery research. Moreover, in vivo lectin binding experiments are comparable to ex vivo experiments, which should be considered when planning and optimizing future research.