Dose-dependent effect of Resveratrol on bladder cancer cells: Chemoprevention and oxidative stress

Background: Over 6 million people die annually in the world because of cancer. Several groups are focused on studying cancer chemoprevention approaches. Resveratrol, a polyphenol, at high dosages, has been reported as antitumor and chemopreventive. However, it has a dose-dependent effect on cell death, even on some cancer cells. Objectives: Our aim was to investigate this dose-dependent effect on human bladder carcinoma ECV304 cells during oxidative stress condition. Methods: For this purpose. ECV304 cells incubated with different Resveratrol concentrations were analyzed as for their metabolic rate, membrane permeability, DNA fragmentation, anti/proapoptotic protein levels and phosphatidylserine exposure after oxidative stress. Results: Resveratrol induced cell death at high concentrations (>20 mu M), but not at low ones (0.1-20 mu M). Pretreatment with 2.5 mu M protected the cells from oxidative damage, whereas 50 mu M intensified the cell death and significantly increased Bad/Bcl-2 ratio (proapoptotic/antiapoptotic proteins). Resveratrol was able to modulate NO and PGE(2) secretion and performed an anti-adhesion activity of neutrophils on PMA-activated ECV304 cells. Conclusions: Resveratrol at high doses induces cell death of ECV304 cells whereas low doses induce protection. Modulation of Bcl-2 protein induced by Resveratrol could be mediating this effect. This information about the role of Resveratrol on cancer alerts us about its dose-dependent effects and could lead the design of future chemoprevention strategies. Published by Elsevier Ireland Ltd.

Increased expression of MMP-9 and IL-8 are correlated with poor prognosis of Bladder Cancer

Background: Extracellular matrix homeostasis is strictly maintained by a coordinated balance between the expression of metalloproteinases (MMPs) and their inhibitors. The purpose of this study was to investigate whether the expression of MMP-9, MMP-2 and its specific inhibitors, are expressed in a reproducible, specific pattern and if the profiles are related to prognosis in Bladder Cancer (BC). Methods: MMP-9, MMP-2 and its specific inhibitors expression levels were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) in fresh-frozen malignant tissue collected from 40 patients with BC submitted to transurethral resection of bladder. The control group consisted of normal bladder tissue from five patients who had undergone retropubic prostatectomy to treat benign prostatic hyperplasia. Results: MMP-9 was overexpressed in 59.0 % of patients, and MMP-2, TIMP-1, TIMP-2, MMP-14, RECK and IL-8 was underexpressed in most of the patients. Regarding prognostic parameters we observed that high-grade tumors exhibited significantly higher levels of MMP-9 and IL-8 (p = 0.012, p = 0.003). Invasive tumors (pT1-pT2) had higher expression levels of MMP-9 than superficial tumors (pTa) (p = 0.026). The same was noted for IL-8 that was more expressed by invasive tumors (p = 0.015, p = 0.048). Most importantly tumor recurrence was related with higher levels of both MMP-9 (p = 0.003) and IL-8 (p = 0.005). Conclusion: We have demonstrated that the overexpression of MMP-9 and higher expression of IL-8 are related to unfavorable prognostic factors of urothelial bladder cancer and tumor recurrence and may be useful in the follow up of the patients.

Bladder expression of CD cell surface antigens and cell-type-specific transcriptomes

Many cell types have no known functional attributes. In the bladder and prostate, basal epithelial and stromal cells appear similar in cytomorphology and share several cell surface markers. Their total gene expression (transcriptome) should provide a clear measure of the extent to which they are alike functionally. Since urologic stromal cells are known to mediate organ-specific tissue formation, these cells in cancers might exhibit aberrant gene expression affecting their function. For transcriptomes, cluster designation (CD) antigens have been identified for cell sorting. The sorted cell populations can be analyzed by DNA microarrays. Various bladder cell types have unique complements of CD molecules. CD9(+) urothelial, CD104(+) basal and CD13(+) stromal cells of the lamina propria were therefore analyzed, as were CD9(+) cancer and CD13(+) cancer-associated stromal cells. The transcriptome datasets were compared by principal components analysis for relatedness between cell types; those with similarity in gene expression indicated similar function. Although bladder and prostate basal cells shared CD markers such as CD104, CD44 and CD49f, they differed in overall gene expression. Basal cells also lacked stem cell gene expression. The bladder luminal and stromal transcriptomes were distinct from their prostate counterparts. In bladder cancer, not only the urothelial but also the stromal cells showed gene expression alteration. The cancer process in both might thus involve defective stromal signaling. These cell-type transcriptomes provide a means to monitor in vitro models in which various CD-isolated cell types can be combined to study bladder differentiation and bladder tumor development based on cell-cell interaction.

Underexpression of MMP-2 and its Regulators, TIMP2, MT1-MMP and IL-8, is Associated with Prostate Cancer

Objective: Extracellular matrix homeostasis is strictly maintained by a coordinated balance between the expression of metalloproteinases (MMPs) and their regulators. The purpose of this study was to investigate whether MMP-2 and its specific regulators, TIMP-2, MT1-MMP and IL-8, are expressed in a reproducible, specific pattern and if the profiles are related to prognosis and clinical outcome of prostate cancer (PCa). Materials and Methods: MMP-2, TIMP-2, MT1-MMP and IL-8 expression levels were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) in freshly frozen malignant and benign tissue specimens collected from 79 patients with clinically localized PCa who underwent radical prostatectomies. The control group consisted of 11 patients with benign prostate hyperplasia (BPH). The expression profile of the MMP-2 and its regulators were compared using Gleason scores, pathological stage, pre-operative PSA levels and the final outcome of the PCa. Results: The analysis of 79 specimens of PCa revealed that MMP-2, TIMP-2, MT1-MMP and IL-8 were underexpressed at 60.0%, 72.2%, 62.0% and 65.8%, respectively, in malignant prostatic tissue in relation to BPH samples. Considering the prognostic parameters, we demonstrated that high Gleason score tumors (>= 7) over-expressed MMP-2 (p = 0.048) and TIMP-2 (p = 0.021), compared to low Gleason score tumors (< 7). Conclusion: We have demonstrated that MMP-2 and its regulators are underexpressed in PCa. Alternatively, overexpression of MMP-2 and TIMP-2 was related to higher Gleason score tumors. We postulate that alterations in metalloproteinase expression may be important in the control of tissue homeostasis related to prostate carcinogenesis and tumor behavior.

Cytostatic in vitro Effects of DTCM-Glutarimide on Bladder Carcinoma Cells

Bladder cancer is a common malignancy worldwide. Despite the increased use of cisplatin-based combination therapy, the outcomes for patients with advanced disease remain poor. Recently, altered activation of the PI3K/Akt/mTOR pathway has been associated with reduced patient survival and advanced stage of bladder cancer, making its upstream or downstream components attractive targets for therapeutic intervention. In the present study, we showed that treatment with DTCM-glutaramide, a piperidine that targets PDK1, results in reduced proliferation, diminished cell migration and G1 arrest in 5637 and T24 bladder carcinoma cells. Conversely, no apoptosis, necrosis or autophagy were detected after treatment, suggesting that reduced cell numbers in vitro are a result of diminished proliferation rather than cell death. Furthermore previous exposure to 10 mu g/ml DTCM-glutarimide sensitized both cell lines to ionizing radiation. Although more studies are needed to corroborate our findings, our results indicate that PDK1 may be useful as a therapeutic target to prevent progression and abnormal tissue dissemination of urothelial carcinomas.

TWIST and p-Akt immunoexpression in normal oral epithelium, oral dysplasia and in oral squamous cell carcinoma

Objectives: The aim of this study was to evaluate the immunoexpression of TWIST and p-Akt proteins in oral leukoplakia (OL) and oral squamous cell carcinoma (OSCC), correlating their expressions with the histological features of the lesions. Study design: Immunohistochemical studies were carried out on 10 normal oral epithelium, 30 OL and 20 OSCC formalin-fixed, paraffin-embedded tissue samples. Immunoperoxidase reactions for TWIST and p-Akt proteins were applied on the specimens and the positivity of the reactions was calculated for 1000 epithelial cells. Results: Kruskal-Wallis and Dunn's post tests revealed a significant difference in TWIST and p-Akt immunoexpression among normal oral mucosa, OL and OSCC. In addition, a significant positive correlation was found between TWIST and p-Akt expressions according to the Pearson's correlation test. Conclusions: The results obtained in the current study suggest that TWIST and p-Akt may participate of the multi-step process of oral carcinogenesis since its early stages.

Immune expression of E-cadherin and alpha, beta and gamma-Catenin adhesion molecules and prognosis for upper urinary tract urothelial carcinomas

Introduction: Cell adhesion molecules (CAM) are required for maintaining a normal epithelial phenotype, and abnormalities in CAM expression have been related to cancer progression, including bladder urothelial carcinomas. There is only one study that correlates E-cadherin and alpha-, beta- and gamma-catenin expression with prognosis of upper tract urothelial carcinomas. Our aim is to study the pattern of immune expression of these CAMs in urothelial carcinomas from the renal pelvis and ureter in patients who have been treated surgically. Our goal is to correlate these expression levels and characteristics with well-known prognostic parameters for disease-free survival. Materials and Methods: We evaluated specimens from 20 patients with urothelial carcinomas of the renal pelvis and ureter who were treated with nephroureterectomy or ureterectomy between June 1997 and January 2007. CAM expression was evaluated by immunohistochemistry in a tissue microarray and correlated with histopathological characteristics and patient outcomes after a mean follow-up of 55 months. Results: We observed a relationship between E-cadherin expression and disease recurrence. Disease recurrence occurred in 87.5% of patients with strong E-cadherin expression. Only 50.0% of patients with moderate expression and 0% of patients with weak or no expression of E-cadherin had disease recurrence (p = 0.014). There was also a difference in disease-free survival. Patients with strong E-cadherin expression had a mean disease-free survival rate of 49.1 months, compared to 83.9 months for patients with moderate expression (p = 0.011). Additionally, an absence of a-catenin expression was associated with tumors that were larger than 3 cm (p = 0.003). Conclusions: We demonstrated for the first time that immune expression of E-cadherin is related to tumor recurrence and disease-free survival rates, and the absence of a-catenin expression is related to tumor size in upper tract urothelial carcinomas.

Standard surgical treatment for benign prostatic hyperplasia is safe for patients over 75 years: Analysis of 100 cases from a high-volume urologic center

OBJECTIVES: In this study, we aimed to determine the complications of standard surgical treatments among patients over 75 years in a high-volume urologic center. METHODS: We analyzed 100 consecutive patients older than 75 years who had undergone transurethral prostatic resection of the prostate or open prostatectomy for treatment of benign prostatic hyperplasia from January 2008 to March 2010. We analyzed patient age, prostate volume, prostate-specific antigen level, international prostatic symptom score, quality of life score, urinary retention, co-morbidities, surgical technique and satisfaction with treatment. RESULTS: Median age was 79 years. Forty-eight patients had undergone transurethral prostatic resection of the prostate, and 52 had undergone open prostatectomy. The median International Prostatic Symptom Score was 20, the median prostate volume was 83 g, 51% were using an indwelling bladder catheter, and the median prostate-specific antigen level was 5.0 ng/ml. The most common comorbidities were hypertension, diabetes and coronary disease. After a median follow-up period of 17 months, most patients were satisfied. Complications were present in 20% of cases. The most common urological complication was urethral stenosis, followed by bladder neck sclerosis, urinary fistula, late macroscopic hematuria and persistent urinary incontinence. The most common clinical complication was myocardial infarction, followed by acute renal failure requiring dialysis. Incidental carcinoma of the prostate was present in 6% of cases. One case had urothelial bladder cancer. CONCLUSIONS: Standard surgical treatments for benign prostatic hyperplasia are safe and satisfactory among the elderly. Complications are infrequent, and urethral stenosis is the most common. No clinical variable is associated with the occurrence of complications.

Standard surgical treatment for benign prostatic hyperplasia is safe for patients over 75 years: analysis of 100 cases from a high-volume urologic center

OBJECTIVES: In this study, we aimed to determine the complications of standard surgical treatments among patients over 75 years in a high-volume urologic center. METHODS: We analyzed 100 consecutive patients older than 75 years who had undergone transurethral prostatic resection of the prostate or open prostatectomy for treatment of benign prostatic hyperplasia from January 2008 to March 2010. We analyzed patient age, prostate volume, prostate-specific antigen level, international prostatic symptom score, quality of life score, urinary retention, co-morbidities, surgical technique and satisfaction with treatment. RESULTS: Median age was 79 years. Forty-eight patients had undergone transurethral prostatic resection of the prostate, and 52 had undergone open prostatectomy. The median International Prostatic Symptom Score was 20, the median prostate volume was 83 g, 51% were using an indwelling bladder catheter, and the median prostatespecific antigen level was 5.0 ng/ml. The most common comorbidities were hypertension, diabetes and coronary disease. After a median follow-up period of 17 months, most patients were satisfied. Complications were present in 20% of cases. The most common urological complication was urethral stenosis, followed by bladder neck sclerosis, urinary fistula, late macroscopic hematuria and persistent urinary incontinence. The most common clinical complication was myocardial infarction, followed by acute renal failure requiring dialysis. Incidental carcinoma of the prostate was present in 6% of cases. One case had urothelial bladder cancer. CONCLUSIONS: Standard surgical treatments for benign prostatic hyperplasia are safe and satisfactory among the elderly. Complications are infrequent, and urethral stenosis is the most common. No clinical variable is associated with the occurrence of complications.

The impact of atmospheric particulate matter on cancer incidence and mortality in the city of Sao Paulo, Brazil

This study aimed to verify the impact of inhalable particulate matter (PM10) on cancer incidence and mortality in the city of Sao Paulo, Brazil. Statistical techniques were used to investigate the relationship between PM10 on cancer incidence and mortality in selected districts. For some types of cancer (skin, lung, thyroid, larynx, and bladder) and some periods, the correlation coefficients ranged from 0.60 to 0.80 for incidence. Lung cancer mortality showed more correlations during the overall period. Spatial analysis showed that districts distant from the city center showed higher than expected relative risk, depending on the type of cancer According to the study, urban PM10 can contribute to increased incidence of some cancers and may also contribute to increased cancer mortality. The results highlight the need to adopt measures to reduce atmospheric PM10 levels and the importance of their continuous monitoring.

Prostate biopsy in patients with long-term use of indwelling bladder catheter: What is the rationale?

Objective: Acute urinary retention (AUR) is expected to occur in 2% to 39% men with benign prostatic hyperplasia. To date, no study has elucidated the effect of long-term use of indwelling bladder catheter on serum prostate specific antigen (PSA) levels and on the incidence of prostate cancer (CaP). The aim of the present study is to analyze the incidence of CaP in patients with long-term use of indwelling bladder catheter and determine some practice patterns on this issue. Materials and methods: The study comprised a retrospective analysis of data from 1,651 patients who had undergone transrectal ultrasound (TRUS)-guided prostate biopsy from July 2004 to June 2009. Among these patients, 198 (12%) were using an indwelling bladder catheter during the biopsy for at least 1 month. The incidence of CaP was recorded according to total PSA levels. Other variables such patient age, free/total PSA rate, PSA density, prostate volume, and duration of catheter use was also analyzed. Men with a digital rectal examination suspicious for cancer were not considered for analysis. Results: Median patient age was 71 years (37 to 89 years). Overall, 25% of patients presented a CaP diagnosis. CaP incidence according to the PSA levels was 0%, 18.9%, 24.5%, and 40.6% for patients with PSA <= 4.0, 4.1-10.0, 10.1-20.0, and >20.0 ng/ml, respectively. When prostate volume was analyzed together, we demonstrated that only 1 (2.4%) patient with PSA below 10.0 ng/ml and prostate volume >60 g had CaP. Median total PSA, PSA density, and prostate volume were statistically different between patients with and without CaP. Conclusions: Prostate biopsy should not be indicated for all patients with diagnosis of BPH and AUR who present an elevated PSA level. Patients with PSA below 10.0 ng/ml, and prostate volume >60 g should only undergo biopsy in selected cases. Patients with PSA >20.0 ng/ml and a prostate volume <= 60 g are at higher risk of CaP diagnosis. (C) 2012 Elsevier Inc. All rights reserved.

Stage, Grade and Behavior of Bladder Urothelial Carcinoma Defined by the MicroRNA Expression Profile

Purpose: We identified miRNA expression profiles in urothelial carcinoma that are associated with grade, stage, and recurrence-free and disease specific survival. Materials and Methods: The expression of 14 miRNAs was evaluated by quantitative reverse transcriptase-polymerase chain reaction in surgical specimens from 30 patients with low grade, noninvasive (pTa) and 30 with high grade, invasive (pT2-3) urothelial carcinoma. Controls were normal bladder tissue from 5 patients who underwent surgical treatment for benign prostatic hyperplasia. Endogenous controls were RNU-43 and RNU-48. miRNA profiles were compared and Kaplan-Meier curves were constructed to analyze disease-free and disease specific survival. Results: miR-100 was under expressed in 100% of low grade pTa specimens (p <0.001) and miR-10a was over expressed in 73.3% (p <0.001). miR-21 and miR-205 were over expressed in high grade pT2-3 disease (p = 0.02 and <0.001, respectively). The other miRNAs were present at levels similar to those of normal bladder tissue or under expressed in each tumor group. miR-21 over expression (greater than 1.08) was related to shorter disease-free survival in patients with low grade pTa urothelial carcinoma. Higher miR-10a levels (greater than 2.30) were associated with shorter disease-free and disease specific survival in patients with high grade pT2-3 urothelial carcinoma. Conclusions: Four miRNAs were differentially expressed in the 2 urothelial carcinoma groups. miR-100 and miR-10a showed under expression and over expression, respectively, in low grade pTa tumors. miR-21 and miR-205 were over expressed in pT2-3 disease. In addition, miR-10a and miR-21 over expression was associated with shorter disease-free and disease specific survival. miRNAs could be incorporated into the urothelial carcinoma molecular pathway. These miRNAs could also serve as new diagnostic or prognostic markers and new target drugs.

The impact of atmospheric particulate matter on cancer incidence and mortality in the city of São Paulo, Brazil

This study aimed to verify the impact of inhalable particulate matter (PM10) on cancer incidence and mortality in the city of São Paulo, Brazil. Statistical techniques were used to investigate the relationship between PM10 on cancer incidence and mortality in selected districts. For some types of cancer (skin, lung, thyroid, larynx, and bladder) and some periods, the correlation coefficients ranged from 0.60 to 0.80 for incidence. Lung cancer mortality showed more correlations during the overall period. Spatial analysis showed that districts distant from the city center showed higher than expected relative risk, depending on the type of cancer. According to the study, urban PM10 can contribute to increased incidence of some cancers and may also contribute to increased cancer mortality. The results highlight the need to adopt measures to reduce atmospheric PM10 levels and the importance of their continuous monitoring.

Morphological alterations and gene and protein expression profiling of bladder tumor cells after treatment with gemcitabine.

Chemical agents used in cancer therapy are associated with cell cycle arrest, activation or deactivation of mechanisms associated to DNA repair and apoptosis. However, due to the complexity of biological systems, the molecular mechanisms responsible for these activities are not fully understood. Thus, studies about gene and protein expression have shown promising results for understanding the mechanisms related to cellular responses and regression of cancer after chemotherapy. This study aimed to evaluate the gene and protein expression profiling in bladder transitional cell carcinoma (TCC) with different TP53 status after gemcitabine (1.56 μM) treatment. The RT4 (grade 1, TP53 wild type), 5637 (grade 2, TP53 mutated) and T24 (grade 3, TP53 mutated) cell lines were used. PCR arrays and mass spectrometry were used to analyze gene and protein expression, respectively. Morphological alterations were observed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The results of PCR array showed that gemcitabine activity was mainly related to CDKN1A, GADD45A and SERTDA1 overexpression, and BAX overexpression only in the wild type TP53 cells. Mass spectrometry demonstrated that gemcitabine modulated the protein expression, especially those from genes related to apoptosis, transport of vesicles and stress response. Analyses using SEM and TEM showed changes in cell morphology independently on the cell line studied. The observed decreased number of microvillus suggests low contact among the cells and between cell and extracellular matrix; irregular forms might indicate actin cytoskeleton deregulation; and the reduction in the amount of organelles and core size might indicate reduced cellular metabolism. In conclusion, independently on TP53 status or grade of bladder tumor, gemcitabine modulated genes related to the cell cycle and apoptosis, that reflected in morphological changes indicative of future cell death.