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1.
Int J Mol Sci ; 22(12)2021 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-34199232

RESUMO

Non-muscle-invasive bladder cancer is the most common form of bladder cancer. The main problem in managing bladder tumors is the high recurrence after the transurethral resection of bladder tumors (TURBT). Our study aimed to examine the fate of intravesically applied cancer cells as the implantation of cancer cells after TURBT is thought to be a cause of tumor recurrence. We established an orthotopic mouse bladder tumor model with MB49-GFP cancer cells and traced them during the first three days to define their location and contacts with normal urothelial cells. Data were obtained by Western blot, immunolabeling, and light and electron microscopy. We showed that within the first two hours, applied cancer cells adhered to the traumatized epithelium by cell projections containing α3ß1 integrin on their tips. Cancer cells then migrated through the epithelium and on day 3, they reached the basal lamina or even penetrated it. In established bladder tumors, E-cadherin and desmoplakin 1/2 were shown as feasible immunohistochemical markers of tumor margins based on the immunolabeling of various junctional proteins. Altogether, these results for the first time illustrate cancer cell implantation in vivo mimicking cellular events of tumor recurrence in bladder cancer patients.


Assuntos
Epitélio/patologia , Recidiva Local de Neoplasia/patologia , Neoplasias da Bexiga Urinária/patologia , Bexiga Urinária/patologia , Animais , Caderinas/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular , Modelos Animais de Doenças , Feminino , Integrina alfa3beta1/metabolismo , Junções Intercelulares/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos Endogâmicos C57BL , Invasividade Neoplásica , Bexiga Urinária/ultraestrutura , Neoplasias da Bexiga Urinária/ultraestrutura , Urotélio/patologia , Urotélio/ultraestrutura
2.
Mol Cell Biochem ; 471(1-2): 113-127, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32519230

RESUMO

N-cadherin is a transmembrane glycoprotein expressed by mesenchymal origin cells and is located at the adherens junctions. It regulates also cell motility and contributes to cell signaling. In previous studies, we identified that its anomalous expression in bladder carcinoma was a tumor progression marker. A pharmacological approach to inhibit N-cadherin expression or to block its function could be relevant to prevent disease progression and metastasis development. The morphological exploration of T24 invasive bladder cancer cells by atomic force microscopy (AFM) revealed a spindle-like shape with fibrous structures. By engaging force spectroscopy with AFM tip functionalized with anti-E or anti-N-cadherin antibodies, results showed that T24 cells expressed only N-cadherin as also demonstrated by Western blotting and confocal microscopy. For the first time, we demonstrated by RTqPCR and Western blotting analyses that the peroxisome proliferator-activated receptor ß/δ (PPARß/δ) agonist GW501516 significantly decreased N-cadherin expression in T24 cells. Moreover, high non-cytotoxic doses of GW501516 inhibited confluent T24 cell wound healing closure. By using AFM, a more sensitive nanoanalytical method, we showed that the treatment modified the cellular morphology and diminished N-cadherin cell surface coverage through the decreasing of these adhesion molecule-mediated interaction forces. We observed a greater decrease of N-cadherin upon GW501516 exposure with AFM than that detected with molecular biology techniques. AFM was a complementary tool to biochemical techniques to perform measurements on living cells at the nanometer resolution level. Taken together, our data suggest that GW501516 could be an interesting therapeutic strategy to avoid bladder cancer cell spreading through N-cadherin decrease.


Assuntos
Antígenos CD/metabolismo , Caderinas/metabolismo , Transição Epitelial-Mesenquimal , Microscopia de Força Atômica/métodos , PPAR delta/agonistas , PPAR beta/agonistas , Tiazóis/farmacologia , Neoplasias da Bexiga Urinária/metabolismo , Antígenos CD/ultraestrutura , Caderinas/ultraestrutura , Linhagem Celular Tumoral , Movimento Celular , Humanos , Transdução de Sinais , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/ultraestrutura
3.
J Cell Biochem ; 117(4): 1027-32, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26515240

RESUMO

The cellular basis of metastasis is poorly understood. An important step to understanding this process is to be able to visualize the routes by which cancer cells migrate from the primary tumor to various distant sites to eventually form metastasis. Our laboratory previously developed single-cell in vivo imaging using fluorescent proteins to label cancer cells. In the present study, using PC-3 human prostate cancer cells labeled with green fluorescent protein (GFP) and orthotopic tumor transplantation, we have imaged in live mice various highly diverse routes by which PC-3 cells metastasize superiorly and inferiorly to distant sites, including in the portal area, stomach area, and urogenital system. Imaging began at day 9, at which time distant metastasis had already occurred, and increased at each imaging point at days 10, 13, 14, and 16. Metastatic cells were observed migrating superiorly and inferiorly from the primary tumor as well as in lymphatic channels and trafficking in various organ systems demonstrating that PC-3 has multiple metastatic routes similar to hormone-independent advanced-stage prostate cancer in the clinic.


Assuntos
Rastreamento de Células/métodos , Diagnóstico por Imagem/métodos , Neoplasias Pancreáticas/diagnóstico , Neoplasias da Próstata/diagnóstico , Neoplasias Gástricas/diagnóstico , Neoplasias Testiculares/diagnóstico , Neoplasias da Bexiga Urinária/diagnóstico , Animais , Linhagem Celular Tumoral , Movimento Celular , Expressão Gênica , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Metástase Linfática , Masculino , Camundongos , Camundongos Nus , Camundongos Transgênicos , Invasividade Neoplásica , Transplante de Neoplasias , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/secundário , Neoplasias Pancreáticas/ultraestrutura , Próstata/metabolismo , Próstata/patologia , Próstata/ultraestrutura , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/ultraestrutura , Neoplasias Gástricas/genética , Neoplasias Gástricas/secundário , Neoplasias Gástricas/ultraestrutura , Neoplasias Testiculares/genética , Neoplasias Testiculares/secundário , Neoplasias Testiculares/ultraestrutura , Transplante Heterólogo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/secundário , Neoplasias da Bexiga Urinária/ultraestrutura
4.
Cancer ; 121(2): 169-78, 2015 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-25132313

RESUMO

Advances in endoscopic imaging technology may improve sensitivity for the detection of bladder cancer and provide a more complete understanding of the urothelial landscape, and it also may lead to improved short-term and long-term cancer control. Fluorescence cystoscopy requires intravesical administration of a photosensitizing agent (5-aminolevulinic acid or hexaminolevulinate), and imaging with a blue-light endoscopy system demonstrably improves the detection of papillary and flat bladder lesions compared with conventional white-light cystoscopy. Prospective phase 3 clinical trials have demonstrated improved diagnostic ability, enhanced tumor resection, and a small but significant reduction in recurrence-free survival. Optical coherence tomography delineates subsurface microarchitecture information about bladder lesions in real time and has the ability to discriminate between noninvasive and invasive cancers. Narrow-band imaging may augment white-light cystoscopy by providing increased contrast between normal and abnormal tissue on the basis of neovascularity. Confocal laser endoscopy has been applied to the urinary tract using thinner probes adapted from use in gastrointestinal malignancies and provides exquisite images at microscopic resolution. More technology is on the horizon that may further enhance our ability to detect and accurately stage bladder tumors and distinguish benign from malignant or dysplastic lesions.


Assuntos
Carcinoma/diagnóstico , Cistoscopia/métodos , Microscopia Confocal , Imagem Multimodal/métodos , Tomografia de Coerência Óptica , Neoplasias da Bexiga Urinária/diagnóstico , Ácido Aminolevulínico , Carcinoma/patologia , Carcinoma Papilar/diagnóstico , Ensaios Clínicos Fase III como Assunto , Fluorescência , Humanos , Fármacos Fotossensibilizantes , Valor Preditivo dos Testes , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/ultraestrutura
5.
Ann Diagn Pathol ; 19(5): 314-9, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26235883

RESUMO

We discuss the histological and immunohistochemical features of 6 cases of urothelial carcinomas of lipid cell variant and 4 cases with shadow cell differentiation, one of which showed additionally sebaceous differentiation, one of which shows additional sebaceous differentiation, from our archive cases from the last 15 years. Conventional urothelial carcinoma (UC) was seen in all lipid cell variant cases, and micropapillary carcinoma was seen in 3. The ratio of the lipid cell component was between 10% and 40% in these 6 cases. Typical histologic features of the lipid cell variant include lipoblast-like cells with a notched nuclear appearance, abundant vacuoles, an eccentric nucleus, and pagetoid spread in some areas. GATA3 and pancytokeratin AE1/AE3 immunohistochemical staining were positive in all cases. Adipophilin was positive in various degrees in 5 of the 6 lipid cell variant cases but was also positive in the case with sebaceous differentiation. α-methylacyl-CoA racemase was positive in the lipid cell areas and negative or focal weakly positive in the conventional UC areas in 4 of the 6 cases. Vimentin, S-100 protein, and PAX8 were negative in the lipid cell component. Follow-up information was available for all cases with follow-up ranging from 6 to 84 months (mean, 34 months). Four patients died of the disease. One pT4 patient who had been followed up for 6 months lives with the disease, whereas another is disease free. In conclusion, the lipid cell variant is a rare UC variant that usually presents at an advanced stage, and tumor cells are histologically similar to lipoblasts, resemble sebaceous differentiation, and show positive immunohistochemical staining with adipophilin.


Assuntos
Carcinoma de Células de Transição/metabolismo , Neoplasias Epiteliais e Glandulares/patologia , Glândulas Sebáceas/patologia , Neoplasias da Bexiga Urinária/patologia , Idoso , Biomarcadores Tumorais , Carcinoma de Células de Transição/patologia , Carcinoma de Células de Transição/ultraestrutura , Diferenciação Celular/fisiologia , Feminino , Fator de Transcrição GATA3/metabolismo , Humanos , Imuno-Histoquímica , Queratinas/metabolismo , Metabolismo dos Lipídeos , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Neoplasias Epiteliais e Glandulares/metabolismo , Neoplasias Epiteliais e Glandulares/ultraestrutura , Perilipina-2 , Racemases e Epimerases/metabolismo , Proteínas S100/metabolismo , Glândulas Sebáceas/metabolismo , Glândulas Sebáceas/ultraestrutura , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/ultraestrutura
6.
Pediatr Surg Int ; 30(1): 123-6, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24030813

RESUMO

Gastrocystoplasty is a surgical form of bladder augmentation which improves bladder capacity and compliance. Patients who undergo bladder augmentation with a gastric remnant are at increased risk for malignancy. The most common types of tumors in this situation were adenocarcinoma and urothelial carcinoma. Most of the adenocarcinomas arise in the gastric remnant or anastomotic site, and adenocarcinomas arising in the residual native bladder are extremely rare. We report on a patient who received gastrocystoplasty 16 years ago. She suffered from recurrent urinary tract infections for a year and cystoscopy showed a tumor in the bladder trigone. Pathologic examination showed tubulovillous adenoma with malignant transformation to adenocarcinoma. The tumor consisted of intact adenomatous architecture from low-grade dysplastic gland to adenocarcinoma, which suggested that the pathogenesis might be related to intestinal metaplasia and dysplasia. The unique location and immunohistologic findings of the tumor suggested that it originated in the bladder mucosa.


Assuntos
Adenocarcinoma/cirurgia , Adenoma Viloso/cirurgia , Estômago/cirurgia , Neoplasias da Bexiga Urinária/cirurgia , Bexiga Urinária/patologia , Procedimentos Cirúrgicos Urológicos/efeitos adversos , Adenocarcinoma/diagnóstico , Adenocarcinoma/ultraestrutura , Adenoma Viloso/diagnóstico , Adenoma Viloso/ultraestrutura , Criança , Feminino , Humanos , Metaplasia , Bexiga Urinária/cirurgia , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/ultraestrutura , Procedimentos Cirúrgicos Urológicos/métodos
7.
BJU Int ; 109(2): 300-5, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21854534

RESUMO

OBJECTIVE: To develop a diagnostic method relying on the preferential accumulation of a dye in non-muscle-invasive bladder cancer (NMIBC) that is visible in conjunction with white-light cystoscopy (WLC). MATERIALS AND METHODS: We investigated in detail the permeation of Evans blue in urothelial cell carcinoma (UCC) spheroids prepared from T24, J82 and RT-112 human cell lines and spheroids composed of normal human urothelial (NHU) cells. To gain more insight into the differential accumulation, all spheroids were investigated ultrastructurally using transmission electron microscopy (TEM). RESULTS: We found that, after exposure to Evans blue for 2 h, UCC spheroids accumulated dramatically more dye than spheroids composed of NHU cells. Using TEM it was found that the malignant spheroids contain similar ultrastructural characteristics, i.e. a wide intercellular space and a decreased number of desmosome-like cell attachments, to those from clinical samples of non-papillary carcinoma in situ of the bladder. CONCLUSION: We believe the present findings could be important for future developments in clinical diagnostics for early bladder cancer detection, staging and grading involving WLC.


Assuntos
Carcinoma de Células de Transição/diagnóstico , Corantes , Cistoscopia/métodos , Detecção Precoce de Câncer/métodos , Azul Evans , Neoplasias da Bexiga Urinária/diagnóstico , Carcinoma de Células de Transição/ultraestrutura , Linhagem Celular Tumoral , Humanos , Luz , Microscopia Eletrônica de Transmissão , Esferoides Celulares/ultraestrutura , Neoplasias da Bexiga Urinária/ultraestrutura , Urotélio/ultraestrutura
8.
Mol Cell Proteomics ; 9(6): 1324-38, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20224111

RESUMO

Exosomes are nanometer-sized vesicles, secreted by various cell types, present in biological fluids that are particularly rich in membrane proteins. Ex vivo analysis of exosomes may provide biomarker discovery platforms and form non-invasive tools for disease diagnosis and monitoring. These vesicles have never before been studied in the context of bladder cancer, a major malignancy of the urological tract. We present the first proteomics analysis of bladder cancer cell exosomes. Using ultracentrifugation on a sucrose cushion, exosomes were highly purified from cultured HT1376 bladder cancer cells and verified as low in contaminants by Western blotting and flow cytometry of exosome-coated beads. Solubilization in a buffer containing SDS and DTT was essential for achieving proteomics analysis using an LC-MALDI-TOF/TOF MS approach. We report 353 high quality identifications with 72 proteins not previously identified by other human exosome proteomics studies. Overrepresentation analysis to compare this data set with previous exosome proteomics studies (using the ExoCarta database) revealed that the proteome was consistent with that of various exosomes with particular overlap with exosomes of carcinoma origin. Interrogating the Gene Ontology database highlighted a strong association of this proteome with carcinoma of bladder and other sites. The data also highlighted how homology among human leukocyte antigen haplotypes may confound MASCOT designation of major histocompatability complex Class I nomenclature, requiring data from PCR-based human leukocyte antigen haplotyping to clarify anomalous identifications. Validation of 18 MS protein identifications (including basigin, galectin-3, trophoblast glycoprotein (5T4), and others) was performed by a combination of Western blotting, flotation on linear sucrose gradients, and flow cytometry, confirming their exosomal expression. Some were confirmed positive on urinary exosomes from a bladder cancer patient. In summary, the exosome proteomics data set presented is of unrivaled quality. The data will aid in the development of urine exosome-based clinical tools for monitoring disease and will inform follow-up studies into varied aspects of exosome manufacture and function.


Assuntos
Exossomos/metabolismo , Proteômica/métodos , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia , Sequência de Aminoácidos , Western Blotting , Linhagem Celular Tumoral , Cromatografia Líquida , Bases de Dados Genéticas , Eletroforese em Gel Bidimensional , Exossomos/química , Exossomos/ultraestrutura , Citometria de Fluxo , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Dados de Sequência Molecular , Nanotecnologia , Proteínas de Neoplasias/química , Proteínas de Neoplasias/urina , Reprodutibilidade dos Testes , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Neoplasias da Bexiga Urinária/ultraestrutura , Neoplasias da Bexiga Urinária/urina
9.
Autophagy ; 17(4): 840-854, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-32116109

RESUMO

Although MIR516A has been reported to be downregulated and act as a tumor suppressor in multiple cancers, its expression and potential contribution to human bladder cancer (BC) remain unexplored. Unexpectedly, we showed here that MIR516A was markedly upregulated in human BC tissues and cell lines, while inhibition of MIR516A expression attenuated BC cell monolayer growth in vitro and xenograft tumor growth in vivo, accompanied with increased expression of PHLPP2. Further studies showed that MIR516A was able to directly bind to the 3'-untranslated region of PHLPP2 mRNA, which was essential for its attenuating PHLPP2 expression. The knockdown of PHLPP2 expression in MIR516A-inhibited cells could reverse BC cell growth, suggesting that PHLPP2 is a MIR516A downstream mediator responsible for MIR516A oncogenic effect. PHLPP2 was able to mediate BECN1/Beclin1 stabilization indirectly, therefore promoting BECN1-dependent macroautophagy/autophagy, and inhibiting BC tumor cell growth. In addition, our results indicated that the increased autophagy by attenuating MIR516A resulted in a dramatic inhibition of xenograft tumor formation in vivo. Collectively, our results reveal that MIR516A has a novel oncogenic function in BC growth by directing binding to PHLPP2 3'-UTR and inhibiting PHLPP2 expression, in turn at least partly promoting CUL4A-mediated BECN1 protein degradation, thereby attenuating autophagy and promoting BC growth, which is a distinct function of MIR516A identified in other cancers.Abbreviation: ATG3: autophagy related 3; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG12: autophagy related 12; BAF: bafilomycin A1; BC: bladder cancer; CHX: cycloheximide; Co-IP: co-immunoprecipitation; CUL3: cullin 3; CUL4A: cullin 4A; CUL4B: cullin 4B; IF: immunofluorescence: IHC-p: immunohistochemistry-paraffin; MIR516A: microRNA 516a (microRNA 516a1 and microRNA 516a2); MS: mass spectrometry; PHLPP2: PH domain and leucine rich repeat protein phosphatase.


Assuntos
Autofagia/genética , Proteína Beclina-1/metabolismo , Carcinogênese/genética , Regulação Neoplásica da Expressão Gênica , MicroRNAs/metabolismo , Fosfoproteínas Fosfatases/genética , Neoplasias da Bexiga Urinária/genética , Regiões 3' não Traduzidas/genética , Sequência de Bases , Carcinogênese/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Proteínas Culina/metabolismo , Regulação para Baixo/genética , Humanos , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , MicroRNAs/genética , Fosfoproteínas Fosfatases/metabolismo , Ligação Proteica , Proteólise , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/ultraestrutura , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Toxicol Lett ; 336: 32-38, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33176187

RESUMO

Tobacco smoking is a major risk factor for human cancers including urinary bladder carcinoma. In a previous study, nicotine enhanced rat urinary bladder carcinogenesis in a two-stage carcinogenesis model. Nicotine also induced cytotoxicity in the bladder urothelium in a short-term study. In the present study, male rats were treated with nicotine (40 ppm) in drinking water co-administered with the NADPH oxidase inhibitor, apocynin (0, 250 or 750 mg/kg) in diet for 4 weeks. The apocynin treatment induced no clinical toxic effects. Reduction of reactive oxygen species (ROS) by apocynin was confirmed by immunohistochemistry of 8-OHdG in the bladder urothelium. Incidences of simple hyperplasia, cell proliferation and apoptosis were reduced by apocynin treatment in the bladder urothelium. However, despite reduction of cell proliferation (labeling index), apocynin did not affect the incidence of simple hyperplasia, apoptosis, or ROS generation in the kidney pelvis urothelium, in addition to 8-OHdG positivity induced by nicotine being lower. In vitro, apocynin (500 µM) reduced ROS generation, but induced cell proliferation in bladder cancer cell lines (T24 and UMUC3 cells). These data suggest that oxidative stress may play a role in the cell proliferation of the bladder urothelium induced by nicotine.


Assuntos
Acetofenonas/farmacologia , Proliferação de Células/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , NADPH Oxidases/antagonistas & inibidores , Nicotina , Neoplasias da Bexiga Urinária/prevenção & controle , Bexiga Urinária/efeitos dos fármacos , Urotélio/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Modelos Animais de Doenças , Humanos , Hiperplasia , Masculino , NADPH Oxidases/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos Endogâmicos F344 , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Bexiga Urinária/enzimologia , Bexiga Urinária/ultraestrutura , Neoplasias da Bexiga Urinária/induzido quimicamente , Neoplasias da Bexiga Urinária/enzimologia , Neoplasias da Bexiga Urinária/ultraestrutura , Urotélio/enzimologia , Urotélio/ultraestrutura
11.
Med Mol Morphol ; 43(2): 96-101, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20683697

RESUMO

Characterization of the signet-ring cell carcinoma (sig) component of a urothelial carcinoma (UC) in the urinary bladder of a 64-year-old man, obtained by transurethral resection of bladder tumor (TUR-BT), is reported. In the present case, a characteristic sig component was detected in approximately 20% of UC, G2 tissues. The sig cells were morphologically similar to those found in gastric cancers and were positively stained with periodic acid-Schiff reaction and Alcian blue and mucicarmine stains. Immunohistochemically, the sig cells were selectively positive for carcinoembryonic antigen (CEA), MUC2, and MUC5AC. These immunohistochemical characteristics were similar to those of sig cells in the stomach, except for the positivity with MUC2. It is interesting to note that CAM5.2-positive sig cells were surrounded by CAM5.2-positive UC cells in a solid nest with no apparent associated adenocarcinoma element. In addition, the ultrastructure of sig cells showed multivacuolar cytoplasmic mucin, which proved to be similar to the ultrastructure of gastric cancers. In the present case of UC, G2 was associated with a sig component. Regarding the origin of the sig component in the bladder, it has been suggested that MUC2-positive sig cells in the bladder might be derived directly from metaplasia of UC, without an associated adenocarcinoma component. From this perspective, it may be noteworthy that sig cells in the bladder were selectively positive for MUC2, exhibiting common antigenicity with mucous cells of the gastric intestinal metaplasia. Because UC associated with a sig component carries a worse prognosis than ordinary UC, the presence of the sig component in any UC should be evaluated even within TUR-BT tissues, as in the present case.


Assuntos
Carcinoma de Células em Anel de Sinete/patologia , Carcinoma de Células em Anel de Sinete/ultraestrutura , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/ultraestrutura , Bexiga Urinária/patologia , Urotélio/patologia , Urotélio/ultraestrutura , Endoscopia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Bexiga Urinária/ultraestrutura
12.
Cells ; 9(6)2020 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-32517366

RESUMO

Exosomes are essential for several tumor progression-related processes, including the epithelial-mesenchymal transition (EMT). Long non-coding RNAs (lncRNAs) comprise a major group of exosomal components and regulate the neoplastic development of several cancer types; however, the progressive role of exosomal lncRNAs in bladder cancer have rarely been addressed. In this study, we identified two potential aggressiveness-promoting exosomal lncRNAs, LINC00960 and LINC02470. Exosomes derived from high-grade bladder cancer cells enhanced the viability, migration, invasion and clonogenicity of recipient low-grade bladder cancer cells and activated major EMT-upstream signaling pathways, including ß-catenin signaling, Notch signaling, and Smad2/3 signaling pathways. Nevertheless, LINC00960 and LINC02470 were expressed at significantly higher levels in T24 and J82 cells and their secreted exosomes than in TSGH-8301 cells. Moreover, exosomes derived from LINC00960 knockdown or LINC02470 knockdown T24 cells significantly attenuated the ability of exosomes to promote cell aggressiveness and activate EMT-related signaling pathways in recipient TSGH-8301 cells. Our findings indicate that exosome-derived LINC00960 and LINC02470 from high-grade bladder cancer cells promote the malignant behaviors of recipient low-grade bladder cancer cells and induce EMT by upregulating ß-catenin signaling, Notch signaling, and Smad2/3 signaling. Both lncRNAs may serve as potential liquid biomarkers for the prognostic surveillance of bladder cancer progression.


Assuntos
Transição Epitelial-Mesenquimal/genética , Exossomos/metabolismo , RNA Longo não Codificante/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Comunicação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , Modelos Biológicos , Gradação de Tumores , Invasividade Neoplásica , RNA Longo não Codificante/genética , Reprodutibilidade dos Testes , Neoplasias da Bexiga Urinária/ultraestrutura
13.
J Cell Biol ; 109(4 Pt 1): 1495-509, 1989 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-2677020

RESUMO

Changes of cell morphology and the state of differentiation are known to play important roles in embryogenesis as well as in carcinogenesis. Examples of particularly profound changes are the conversions of epithelial to mesenchymal cells; i.e., the dissociation of some or all polygonal, polar epithelial cells and their transformation into elongate, fibroblastoid cells of high motility. As an in vitro model system for such changes in cell morphology, we have used cell cultures of the rat bladder carcinoma-derived cell line NBT-II which, on exposure to inducing medium containing a commercial serum substitute (Ultroser G), show an extensive change in their organization (epithelial-mesenchymal transition): the junctions between the epithelial cells are split, the epithelial cell organization is lost, and the resulting individual cells become motile and assume a spindle-like fibroblastoid appearance. Using immunofluorescence microscopy and biochemical protein characterization techniques, we show that this change is accompanied by a redistribution of desmosomal plaque proteins (desmoplakins, desmoglein, plakoglobin) and by a reorganization of the cytokeratin and the actin-fodrin filament systems. Moreover, intermediate-sized filaments of the vimentin type are formed in the fibroblastoid cells. We demonstrate that the modulation of desmosomal proteins, specifically an increase in soluble desmoplakins, is a relatively early event in cell dissociation and in epithelial-mesenchymal transition. In this process, a latent period of 5 h upon addition of inducing medium precedes the removal of these desmosomal components from the plasma membrane. The transition, which is reversible, is dependent on continued protein synthesis and phosphorylation but not on the presence of the inducing medium beyond the initial 2-h period. We discuss the value of this experimental system as a physiologically relevant approach for studying the regulation of the assembly and disassembly of desmosomes and other intercellular adhesion structures, and as a model of the conversion of cells from one state of differentiation into another.


Assuntos
Proteínas do Citoesqueleto/ultraestrutura , Desmossomos/ultraestrutura , Células Tumorais Cultivadas/citologia , Neoplasias da Bexiga Urinária/patologia , Animais , Linhagem Celular , Movimento Celular , Citocalasina D/farmacologia , Proteínas do Citoesqueleto/biossíntese , Desmogleínas , Desmoplaquinas , Células Epiteliais , Fibroblastos/citologia , Imunofluorescência , Immunoblotting , Cinética , Fosforilação , Ratos , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/ultraestrutura , Neoplasias da Bexiga Urinária/ultraestrutura , gama Catenina
14.
J Cell Biol ; 90(3): 769-77, 1981 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-7287822

RESUMO

Intranuclear sodium, potassium, and chloride contents were measured by energy-dispersive x-ray microanalysis in freeze-fractured, freeze-dried, bulk-tumor samples taken from 10 patients suffering from invasive urogenital cancers. Human biopsies were carried out during the first diagnostic interventions before any cytostatic treatment had been applied. Pathohistological diagnosis established the malignancy in each case. The cancers were classified in three types: keratinizing, transitional cell, and hypernephroid carcinoma. More than 250 cell nuclei were measured from each type of cancer. The results were compared with those obtained in intact human urothelium taken from patients having no malignant processes. Proximal and distal tubular epithelial cell nuclei representing the origin of human hypernephroid cancer were also measured in rat kidney because corresponding healthy human material cannot be obtained. The analyses revealed, in all three types of cancer cells, that the average intranuclear sodium content increased more than three-fold, the potassium content decreased 32, 16, and 13%, respectively; meanwhile the chloride content increased, but to a lesser extent than did the sodium. The intranuclear Na+:K+ ratios were more than five-fold higher in the cancer cells on the average, and their distribution histograms were much broader than in the normal human urothelium and in the tubular cell nuclei of the rat kidney. The results obtained fit well with the theory of Cone, C. D., Jr. 1971. J. Theor. Biol. 30: 151-181 according to which the sustained depolarization of the cell membrane may be of mitogenic effect.


Assuntos
Neoplasias Renais/análise , Neoplasias Penianas/análise , Potássio/análise , Sódio/análise , Neoplasias da Bexiga Urinária/análise , Adulto , Idoso , Núcleo Celular/análise , Cloretos/análise , Microanálise por Sonda Eletrônica , Feminino , Humanos , Neoplasias Renais/ultraestrutura , Túbulos Renais/análise , Masculino , Pessoa de Meia-Idade , Neoplasias Penianas/ultraestrutura , Bexiga Urinária/análise , Neoplasias da Bexiga Urinária/ultraestrutura
15.
Int J Nanomedicine ; 14: 9731-9743, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31849465

RESUMO

OBJECTIVE: To investigate differential microRNAs' expression in heterogeneous bladder cancer cells, as well as to investigate the mechanism of changes in invasive and proliferative capacity induced by tunneling nanotubes (TNTs) mediated transport of microRNA between bladder cancer cells of varying histological grade. MATERIALS AND METHODS: Differences in microRNA expression between bladder cancer cells of different grade were identified from a literature review. The identified heterogeneous microRNAs were analyzed by qPCR in T24 (high grade) and RT4 (low grade) bladder cancer cells. Scanning electron microscopy (SEM) and laser confocal fluorescence microscopy (LCM) were used to observe tunneling nanotubes (TNTs) between RT4 and T24 cells. Differentially expressed microRNA was labeled and traced by Fluorescent In Situ Hybridization (FISH) following co-culture of T24 and RT4 cells. MicroRNA mimic and inhibition technologies were applied to investigate how TNTs-mediated intercellular transport of microRNA affects the invasive and proliferative behavior of bladder cancer cells. RESULTS: MicroRNA-155 (miR-155) levels were highly expressed in T24 cells, whereas the same was not true in RT4 cells. MiR-155 was confirmed to be a crucial factor sustaining T24 bladder cancer cell proliferation, migration and cell cycle progression by CCK8, Matrigel test and cell cycle analysis, respectively. After T24 and RT4 co-culture, TNTs were assessed by SEM and LCM between T24 and RT4 cells. In addition, we observed TNTs mediated transport of miR-155 from T24 cells to RT4 cells, which thereby acquired a higher proliferative rate, an increased frequency of cells in the S phase, and increased invasive ability in Matrigel test. At the same time, Deptor, the target protein of miR-155 in RT4 cells, was downregulated, followed by mTOR/4EBP1/p70S6K- eIF4e/S6RP signaling activation. CONCLUSION: MiR-155 was differentially expressed between RT4 and T24 bladder cancer cells. Intercellular transport of miR-155 via TNTs can promote bladder cancer cell reprogramming by Deptor-mTOR signal pathway activation.


Assuntos
Proliferação de Células , MicroRNAs/metabolismo , Nanotubos , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Transporte Biológico , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , Peptídeos e Proteínas de Sinalização Intracelular/genética , MicroRNAs/genética , Microscopia Eletrônica de Varredura , Invasividade Neoplásica , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Neoplasias da Bexiga Urinária/ultraestrutura
16.
Vet Microbiol ; 236: 108396, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31500722

RESUMO

Autophagy is a powerful tool that host cells use to defend against viral infection. Mitophagy, the selective autophagic removal of dysfunctional mitochondria was upregulated in urothelial cancer cells harbouring bovine papillomavirus (BPV) infection, as detected by the expression of BPV E5 protein, the major oncoprotein of bovine Deltapapillomavirus genus. HIF-1α-induced mitophagy receptors, BNIP3 and BNIP3L/Nix, were found to be overexpressed in these cells. The BNIP3 and BNIP3L/Nix receptors were amplified, and amplicon sequencing showed homology between bovine BNPI3 and BNIP3L/Nix sequences deposited in GenBank (accession number: NM_001076366.1 and NM_001034614.2, respectively). The transcripts and protein levels of BNIP3 and BNIP3L/Nix were significantly overexpressed in hypoxic neoplastic cells relative to healthy, non-neoplastic cells. BNIP3 and BNIP3L/Nix interacted with the LC3 protein, a marker of autophagosome (mitophagosome) membrane, ERAS, a small GTPase, and p62, known to be a specific autophagy receptor protein, that plays a role in mitochondrial priming for mitophagy and subsequent elimination. ERAS also interacted with the BPV E5 oncoprotein at mitochondrial level. Furthermore, in anti-Bag3 mitochondrial immunoprecipitates, a complex composed of the Hsc70/Hsp70 chaperone, CHIP co-chaperone, Synpo2, ERAS, LC3, p62, BNPI3, and BNIP3L/Nix was also detected. Bag3 may play a role in mitophagosome formation together with the Synpo2 protein and may be involved in the degradation of Hsc70/Hsp70-bound CHIP-ubiquitinated cargo, in association with its chaperone. ERAS may be involved in mitophagosome maturation via the PI3K signalling pathway. Ultrastructural findings revealed the presence of mitochondria exhibiting severe fragmentation and loss of cristae, as well as numerous mitochondria-containing autophagosomes.


Assuntos
Papillomavirus Bovino 1 , Papillomavirus Bovino 4 , Doenças dos Bovinos/virologia , Infecções por Papillomavirus/veterinária , Proteínas Proto-Oncogênicas/metabolismo , Urotélio/citologia , Animais , Bovinos , Doenças dos Bovinos/patologia , Masculino , Proteínas de Membrana , Proteína Oncogênica p21(ras)/genética , Proteína Oncogênica p21(ras)/metabolismo , Proteínas Oncogênicas Virais , Infecções por Papillomavirus/virologia , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/ultraestrutura , Neoplasias da Bexiga Urinária/veterinária , Neoplasias da Bexiga Urinária/virologia , Urotélio/metabolismo
17.
Peptides ; 29(6): 963-8, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18328599

RESUMO

A novel antimicrobial peptide, polybia-MPI, was purified from the venom of the social wasp Polybia paulista. It has potent antimicrobial activity against both Gram-positive and Gram-negative bacteria, but causing no hemolysis to rat erythrocytes. To date, there is no report about its antitumor effects on any tumor cell lines. In this study we synthesized polybia-MPI and studied its antitumor efficacy and cell selectivity. Our results revealed that polybia-MPI exerts cytotoxic and antiproliferative efficacy by pore formation. It can selectively inhibit the proliferation of prostate and bladder cancer cells, but has lower cytotoxicity to normal murine fibroblasts. In addition, to investigate the structure-activity relationship of polybia-MPI, three analogs in which Leu7, Ala8 or Asp9 replaced by L-Pro were designed and synthesized. L-Pro substitution of Leu7 or Asp9 significantly reduces the content of alpha-helix conformation, and L-Pro substitution of Ala8 can disrupt the alpha-helix conformation thoroughly. The L-Pro substitution induces a significant reduction of antitumor activity, indicating that the alpha-helix conformation of polybia-MPI is important for its antitumor activity. In summary, polybia-MPI may offer a novel therapeutic strategy in the treatment of prostate cancer and bladder cancer, considering its relatively lower cytotoxicity to normal cells.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Substituição de Aminoácidos , Animais , Antibacterianos/síntese química , Antibacterianos/isolamento & purificação , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/ultraestrutura , Proliferação de Células/efeitos dos fármacos , Meios de Cultura Livres de Soro , Relação Dose-Resposta a Droga , Desenho de Fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/ultraestrutura , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/fisiologia , Bactérias Gram-Positivas/efeitos dos fármacos , Bactérias Gram-Positivas/fisiologia , Hemólise/efeitos dos fármacos , Hemólise/fisiologia , Humanos , L-Lactato Desidrogenase/análise , Masculino , Camundongos , Células NIH 3T3 , Prolina/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Neoplasias da Próstata/ultraestrutura , Conformação Proteica , Estrutura Secundária de Proteína , Ratos , Relação Estrutura-Atividade , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/ultraestrutura , Venenos de Vespas/química , Venenos de Vespas/toxicidade , Vespas/química
18.
BMC Urol ; 8: 5, 2008 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-18315881

RESUMO

BACKGROUND: This study evaluated the cytotoxic and antiproliferative efficacy of two well-characterized members of the Cecropin-family of antimicrobial peptides against bladder tumor cells and benign fibroblasts. METHODS: The antiproliferative and cytotoxic potential of the Cecropins A and B was quantified by colorimetric WST-1-, BrdU- and LDH-assays in four bladder cancer cell lines as well as in murine and human fibroblast cell lines. IC50 values were assessed by logarithmic extrapolation, representing the concentration at which cell viability was reduced by 50%. Scanning electron microscopy (SEM) was performed to visualize the morphological changes induced by Cecropin A and B in bladder tumor cells and fibroblasts. RESULTS: Cecropin A and B inhibit bladder cancer cell proliferation and viability in a dose-dependent fashion. The average IC50 values of Cecropin A and B against all bladder cancer cell lines ranged between 73.29 mug/ml and 220.05 mug/ml. In contrast, benign fibroblasts were significantly less or not at all susceptible to Cecropin A and B. Both Cecropins induced an increase in LDH release from bladder tumor cells whereas benign fibroblasts were not affected. SEM demonstrated lethal membrane disruption in bladder cancer cells as opposed to fibroblasts. CONCLUSION: Cecropin A and B exert selective cytotoxic and antiproliferative efficacy in bladder cancer cells while sparing targets of benign murine or human fibroblast origin. Both peptides may offer novel therapeutic strategies for the treatment of bladder cancer with limited cytotoxic effects on benign cells.


Assuntos
Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Antineoplásicos/farmacologia , Proteínas de Insetos/farmacologia , Neoplasias da Bexiga Urinária/patologia , Anti-Infecciosos/uso terapêutico , Peptídeos Catiônicos Antimicrobianos/uso terapêutico , Antineoplásicos/uso terapêutico , Bromodesoxiuridina/análise , Linhagem Celular Tumoral , Membrana Celular/patologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Proteínas de Insetos/uso terapêutico , Microscopia Eletrônica de Varredura , Células Tumorais Cultivadas , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/ultraestrutura
20.
Zhongguo Zhong Yao Za Zhi ; 33(15): 1869-73, 2008 Aug.
Artigo em Zh | MEDLINE | ID: mdl-19007019

RESUMO

OBJECTIVE: To investigate the proliferation, apoptosis and mechanisms on T24 cell of transitional cell carcinoma of bladder (TCCB) by crocin. METHOD: MTT assay was used to evaluate the proliferation of T24 cells. The changes of cell cycle and cell apoptotic percentage were measured by flow cytometry. T24 cells were inoculated into BALB/c nude mice to establish model of carcinoma of bladder. The mice were randomly divided into control group and experimental group. After treatment with 50 mmol x L(-1) crocin, the inhibited growth of tumor was observed. Electronic microscope was used to observe the morphological changes. The expressions of Bcl-2, Bax, Survivin and Cyclin D1 were detected by immunohistochemistry. RESULT: The growth of T24 cells was remarkably inhibited after treatment of crocin. Flow cytometric profiles revealed that crocin led to the increase of the cells in G0/G1 phase, the percentage of cell apoptosis was also increased. Crocin could inhibit the growth of BALB/c xenograft tumor. The morphology changes of cell apoptosis were observed. Bcl-2, Cyclin D1 and survivin expressions determined by immunohistochemical staining were down-regulated after treatment with Bax expression up-regulated. CONCLUSION: Crocin exerts both in vitro and in vivo anti-cancer effect on TCCB T24 cell line. The mechanisms may change tumour cell cycle and induce tumour cell apoptosis by down-regulating the expression of Bcl-2, Survivin, Cyclin D1 and up-regulating the expression of Bax.


Assuntos
Apoptose/efeitos dos fármacos , Carotenoides/farmacologia , Carotenoides/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/patologia , Animais , Carcinoma de Células de Transição/tratamento farmacológico , Carcinoma de Células de Transição/patologia , Carcinoma de Células de Transição/ultraestrutura , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Ciclina D1/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Proteínas Inibidoras de Apoptose , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microscopia Eletrônica de Transmissão , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Repressoras , Survivina , Transplante Heterólogo , Neoplasias da Bexiga Urinária/ultraestrutura
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