Enhancing RECK Expression Through miR-21 Inhibition: A Promising Strategy for Bladder Carcinoma Control.

Bladder carcinoma (BC) is the tenth most frequent malignancy worldwide, with high morbidity and mortality rates. Despite recent treatment advances, high-grade BC and muscle-invasive BC present with significant progression and recurrence rates, urging the need for alternative treatments. The microRNA-21 (miR-21) has superexpression in many malignancies and is associated with cellular invasion and progression. One of its mechanisms of action is the regulation of RECK, a tumor suppressor gene responsible for inhibiting metalloproteinases, including MMP9. In a high-grade urothelial cancer cell line, we aimed to assess if miR-21 downregulation would promote RECK expression and decrease MMP9 expression. We also evaluated cellular migration and proliferation potential by inhibition of this pathway. In a T24 cell line, we inhibited miR-21 expression by transfection of a specific microRNA inhibitor (anti-miR-21). There were also control and scramble groups, the last with a negative microRNA transfected. After the procedure, we performed a genetic expression analysis of miR-21, RECK, and MMP9 through qPCR. Migration, proliferation, and protein expression were evaluated via wound healing assay, colony formation assay, flow cytometry, and immunofluorescence.After anti-miR-21 transfection, miR-21 expression decreased with RECK upregulation and MMP9 downregulation. The immunofluorescence assay showed a significant increase in RECK protein expression (p < 0.0001) and a decrease in MMP9 protein expression (p = 0.0101). The anti-miR-21 transfection significantly reduced cellular migration in the wound healing assay (p < 0.0001). Furthermore, in the colony formation assay, the anti-miR-21 group demonstrated reduced cellular proliferation (p = 0.0008), also revealed in the cell cycle analysis by flow cytometry (p = 0.0038). Our results corroborate the hypothesis that miR-21 is associated with BC cellular migration and proliferation, revealing its potential as a new effective treatment for this pathology.

Downregulation of miR-29b is associated with Peyronie's disease.

BACKGROUND: Peyronie's disease (PD) is characterized by the formation of fibrous plaque in tunica albuginea, causing several problems in patients. The etiology of this disease is not fully understood, and there are few effective treatments. To better understand the molecular pathways of PD, we studied miR-29b, a microRNA that could be involved with this illness. MicroRNAs are endogenous molecules that act by inhibiting messenger RNA. MiR-29b regulates 11 of 20 collagen genes and the TGF-ß1 gene, which are related to PD progression. METHODS: We compared miR-29b expression in 11 patients with PD and 14 patients without PD (control group). For the patients with PD, we utilized samples from the fibrous plaque (n = 9), from the tunica albuginea (n = 11), and from the corpus cavernosum (n = 8). For the control group, we utilized samples from the tunica albuginea (n = 14) and from the corpus cavernosum (n = 10). MiR-29b expression was determined by q-PCR. RESULTS: We found a downregulation of miR-29b in the fibrous plaque, tunica albuginea and corpus cavernosum of patients with PD in comparison with the control group (p = 0.0484, p = 0.0025, and p = 0.0016, respectively). CONCLUSION: Although our study has a small sample, we showed for the first time an evidence that the downregulation of miR-29b is associated with PD.

Can increased expression of miR-Let-7c reduce the transition potential of high-grade urothelial carcinoma?

BACKGROUND: Bladder cancer is the leading transitional cell carcinoma affecting men and women with high morbidity and mortality rates, justifying the need to develop new molecular target therapies using microRNAs. This study aimed to evaluate the behavior of the T24 cell line after transfection with miR-Let-7c precursor mimic through invasion, migration, apoptosis, and cell cycle assays. METHODS AND RESULTS: T24 cell was transfected with the Let-7c mimic and its respective control and evaluated after 24 h. The expression levels of miR-Let-7c were analyzed by qPCR. We performed wound healing, Matrigel and flow cytometry, apoptosis, and cell cycle assays to determine its effect on cellular processes. Cells transfected with miR-Let-7c showed increased apoptosis rates (p = 0.019), decreased migration 24 h (p = 0.031) and 48 h (p = 0.0006), invasion potential (p = 0.0007), and cell proliferation (p = 0.002). CONCLUSIONS: Our results demonstrate that miR-Let-7c can act in different pathways of the carcinogenic cellular processes of muscle-invasive urothelial carcinoma cells, inhibiting cell proliferation and increasing apoptosis levels, consequently limiting their invasion potential. However, further studies should be carried out better to elucidate this microRNA's role in high-grade urothelial carcinomas and unveil which targets this microRNA may present, which are intrinsically related to the cancer survival pathways.

Prognostic value of TERF1 expression in prostate cancer.

BACKGROUND: Telomere dysfunction is one of the hallmarks of cancer and is crucial to prostate carcinogenesis. TERF1 is a gene essential to telomere maintenance, and its dysfunction has already been associates with several cancers. TERF1 is a target of miR-155, and this microRNA can inhibit its expression and promotes carcinogenesis in breast cancer. We aim to analyze TERF1, in gene and mRNA level, involvement in prostate cancer progression. RESULTS: Alterations in TERF1 DNA were evaluated using datasets of primary tumor and castration-resistant tumors (CRPC) deposited in cBioportal. The expression of TERF1 mRNA levels was assessed utilizing TCGA datasets, clinical specimens, and metastatic prostate cancer cell lines (LNCaP, DU145, and PC3). Six percent of localized prostate cancer presents alterations in TERF1 (the majority of that was amplifications). In the CRPC cohort, 26% of samples had TERF1 amplification. Patients with TERF1 alterations had the worst overall survival only on localized cancer cohort (p = 0.0027). In the TCGA cohort, mRNA levels of TERF1 were downregulated in comparison with normal tissue (p = 0.0013) and upregulated in tumors that invade lymph nodes (p = 0.0059). The upregulation of TERF1 is also associated with worst overall survival (p = 0.0028) and disease-free survival (p = 0.0023). There is a positive correlation between TERF1 and androgen receptor expression in cancer tissue (r = 0.53, p < 0.00001) but not on normal tissue (r = - 0.16, p = 0.12). In the clinical specimens, there is no detectable expression of TERF1 and upregulation of miR-155 (p = 0.0348). In cell lines, TERF1 expression was higher in LNCaP and was progressively lower in DU145 and PC3 (p = 0.0327) with no differences in miR-155 expression. CONCLUSION: Amplification/upregulation of TERF1 was associated with the worst prognostic in localized prostate cancer. Our results corroborate that miR-155 regulates TERF1 expression in prostate cancer. TERF1 has the potential to become a biomarker in prostate cancer.

Telomeric zinc-finger associated protein (TZAP) in cancer biology: friend or foe?

The new identified protein telomeric zinc-finger associated protein (TZAP) is a negative regulator of telomere length. Since telomere length and telomere maintenance mechanisms are essential to cancer progression, TZAP is considered a new player in cancer biology. Here we aimed to analyze TZAP using the Cancer Genome Atlas data in a Pan-Cancer approach. We gathering data from TCGA Pan-Cancer studies utilizing cBioPortal, GEPIA and UALCAN. In total we analyzed 33 types of cancer (n=9664) and their respective controls (n=711). TZAP is transcribed in all cancers but less than 5% of all tumors show any somatic changes. TZAP was downregulated in kidney chromophobe carcinoma, and upregulated in esophageal cancer, head and neck squamous cell carcinomas, kidney renal clear cell carcinoma and in liver hepatocellular carcinoma. Globally, TZAP expression is related to favorable prognosis, associated to better overall and disease-free survival. Looking to specific tumors, TZAP expression has a dual behavior. Its downregulation is associated with poor prognosis in cervical squamous cell carcinoma, in kidney renal clear cell carcinoma, kidney papillary cell carcinoma, lung adenocarcinoma and pancreas adenocarcinoma. On the contrary, in adrenocortical carcinoma, colon and rectal cancer, brain lower grade glioma and prostate adenocarcinoma the upregulation of TZAP is related with poor prognosis. TZAP expression has a positive correlation with TRF1 and TRF2 in normal tissue but not in cancer. Our analyses indicate that TZAP has an important role in oncology and may be considered as a potential biomarker.

A novel supplement with yeast ß-glucan, prebiotic, minerals and Silybum marianum synergistically modulates metabolic and inflammatory pathways and improves steatosis in obese mice.

OBJECTIVE: To evaluate the synergic effects of a novel oral supplement formulation, containing prebiotics, yeast ß-glucans, minerals and silymarin (Silybum marianum), on lipid and glycidic metabolism, inflammatory and mitochondrial proteins of the liver, in control and high-fat diet-induced obese mice. METHODS: After an acclimation period, 32 male C57BL/6 mice were divided into the following groups: nonfat diet (NFD) vehicle, NFD supplemented, high-fat diet (HFD) vehicle and HFD supplemented. The vehicle and experimental formulation were administered orally by gavage once a day during the last four weeks of the diet (28 consecutive days). We then evaluated energy homeostasis, inflammation, and mitochondrial protein expression in these groups of mice. RESULTS: After four weeks of supplementation, study groups experienced reduced glycemia, dyslipidemia, fat, and hepatic fibrosis levels. Additionally, proliferator-activated receptor-α, AMP-activated protein kinase-1α, peroxisome proliferator-activated receptor γ co-activator-1α, and mitochondrial transcription factor A expression levels were augmented; however, levels of inhibitor of nuclear factor-κB kinase subunit α and p65 nuclear factor-κB expression, and oxidative markers were reduced. Notably, the cortisol/C-reactive protein ratio, a well-characterized marker of the hypothalamic-pituitary-adrenal axis immune interface status, was found to be modulated by the supplement. CONCLUSION: We discovered that the novel supplement was able to modify different antioxidant, metabolic and inflammatory pathways, improving the energy homeostasis and inflammatory status, and consequently alleviated hepatic steatosis.

The role of single nucleotide polymorphisms of miRNAs 100 and 146a as prognostic factors for prostate cancer.

INTRODUCTION: Prostate cancer has a high incidence in men and is the second cause of cancer death among americans male. microRNA (miR) is becoming a potential new prognostic factor for prostate cancer. Single nucleotide polymorphisms (SNPs) are common polymorphisms, characterized by a single exchange of nitrogen based in the DNA. This polymorphism is present in the miRs, altering their function. OBJECTIVE: To evaluate the role of SNP rs1834306 of miR100 and rs2910164 of miR146a in the development and prognosis of prostate cancer. METHODS: One hundred patients diagnosed with prostate cancer and 68 controls were selected. The identification of SNP was rated by quantitative polymerase chain reaction from blood samples, and the analysis was performed within the presence of SNP and the prognostic variables. RESULTS: In the SNP rs1834306 (miR100), a smaller presence of the polymorphic homozygous genotype was identified in patients with PSA >10 ng/mL, (P=0.03); when evaluating only the presence of the polymorphic allele G (P=0.09) it was compared to the presence of the wild type allele A. Among the patients with prostate cancer, SNP rs2910164 (miR146A), the polymorphic allele was more frequent in patients with a Gleason score ⩾7 than in patients with a Gleason score <7, (P=0.043). In patients with prostate cancer, miR100 was overexpressed in those with pT3 staging compared to pT2 and among those who had biochemical recurrence (P = 0.004 and P = 0.011, respectively). CONCLUSIONS: SNP of miR146a acts as a poor prognostic factor (Gleason ⩾7), and the SNP of miR100 is linked to better prognostic data (PSA <10). MiR100 was overexpressed in prostate cancer with worse prognostic factors.

Hypothesis: The triad androgen receptor, zinc finger proteins and telomeres modulates the global gene expression pattern during prostate cancer progression.

Currently, the biggest challenge for prostate cancer (PCa) is to understand the mechanism by which the disease acquires the castration-resistant phenotype and progresses to a fatal disease. PCa has a high genetic heterogeneity, and cannot be separated into well-defined molecular subtypes. Despite this, there is consensus about the role of the androgen receptor (AR) in all stages of the disease, including the transition to the castration-resistant phenotype. Since AR is a transcription factor, we investigated the possibility of PCa presenting a pattern of global gene expression during disease progression. By analyzing the TCGA and CCLE datasets, we were able to find a pattern of waves of genes being expressed during each stage of disease progression. This phenomenon suggests the existence of a mechanism that globally regulates gene expression, being AR, telomeres, and zinc finger proteins (ZNF), three important players. The AR modulates the telomere biology, and its transcription is regulated by ZNF. Recently, a study suggested that the telomere length might influence the expression of ZNF. Thus, we hypothesized that changes in the triad AR, telomeres, and ZNF control gene expression during the progression of PCa.

Physical activity effects on bladder dysfunction in an obese and insulin-resistant murine model.

OBJECTIVE: To investigate the role of physical activity in functional and molecular bladder alterations in an obese and insulin-resistant murine model. METHODS: Wistar rats were randomized into 1. physical activity and standard diet; 2. physical activity and high-fat diet; 3. no physical activity and standard diet; and 4. no physical activity and high-fat diet. Groups 1 and 2 were subjected to a 10-week swimming protocol. Urodynamic study (UDS) was performed, and the expression of genes in the bladder tissue related to the insulin pathway (IRS1/IRS2/PI3K/AKT/eNOS) was assessed using quantitative real-time polymerase chain reaction. RESULTS: Groups 1 and 2 presented lower body weight gains than groups 3 (213.89 ± 13.77 vs 261.63 ± 34.20 grams (g), p = 0.04) and 4 (209.84 ± 27.40 vs 257.57 ± 32.95 g, p = 0.04), respectively. Group 4 had higher insulin level (6.05 ± 1.79 vs 4.14 ± 1.14 ng/ml, p = 0.038) and higher homeostasis model assessment of insulin resistance (HOMA-IR) index (1.95 ± 0.73 vs 1.09 ± 0.37, p = 0.006) than group 1. On UDS, group 4 had greater number of micturition (13.6 ± 4.21 vs 6.0 ± 1.82, p = 0.04), higher postvoid pressure (8.06 ± 2.24 vs 5.08 ± 1.23, p = 0.04), lower capacity (0.29 ± 0.18 vs 0.91 ± 0.41 ml, p = 0.008), and lower bladder compliance (0.027 ± 0.014 vs 0.091 ± 0.034 ml/mmHg, p = 0.016) versus group 1. High-fat diet was related to an underexpression throughout insulin signaling pathway, and physical activity was related to an overexpression of the pathway. CONCLUSIONS: The insulin signaling pathway may be involved in the pathogenesis of bladder dysfunction related to a high-fat diet. Physical activity may help to prevent bladder disfunction induced by a high-fat diet through the insulin pathway.

Tissue expression of MMP-9, TIMP-1, RECK, and miR338-3p in prostate gland: can it predict cancer?

Prostate cancer is the most frequent malignancy affecting men worldwide. Due to the low sensitivity and specificity of the prostate-specific antigen test and the digital rectal exam as screening modalities, several alternatives are being studied. This study aimed to evaluate the application of MMP-9 and its regulators (TIMP-1, RECK, and miR-338-3p) as diagnostic and prognostic indicators of prostate cancer. A total of 134 randomly selected patients under investigation for prostate cancer submitted to a transrectal ultrasound-guided prostate biopsy were enrolled in the study; of these, 61 were positive for the disease (cases), and 73 were negative (control group). The tissue samples were further analyzed by gene and miR-338-3p expression analysis using qRT-PCR (one randomly selected fragment of each patient). Approximately 58% of the patients with prostate cancer presented MMP9 upregulation, while 73%, 65%, and 69% downregulated IMP-1, RECK, and miR-338-3p, respectively. MiR-338-3p was expressed at lower levels in patients with PSA concentrations exceeding 20 ng/mL (p=0.045) and abnormal DRE (p=0.006), while the RECK was more expressed in patients with abnormal DRE (p=0.01). We found that most patients with prostate cancer overexpressed MMP-9; on the other hand, most of them underexpressed TIMP-1, RECK, and miR-338-3p. MiR-338-3p presented as a possible predictor of poor prognosis. Further studies are warranted to evaluate these biomarkers as prognosis factors better.

Treating metastatic prostate cancer with microRNA-145.

Prostate cancer (PCa) is an incurable disease at the metastatic stage. Although there are different options for treatment, the results are limited. MicroRNAs (miRNAs) are a group of small, noncoding, regulatory RNAs with important roles in regulating gene expression. miR-145 is reported to be a key tumor suppressor miRNA (tsmiR) that controls important oncogenes, such as MYC and RAS. In this study, in vitro studies were performed to show the control of MYC and RAS by miR-145. Flow cytometry was used to analyze cell proliferation and apoptosis. The efficacy of miR-145 in treating metastatic PCa was tested in nude mice using a model of bone metastasis promoted by intraventricular injection of PC-3MLuc-C6 cells. Tumor growth was evaluated by an in vivo bioluminescence system. After the full establishment of metastases on day 21, six animals were treated with three intravenous doses of miR-145 (on days 21, 24 and 27), and six were injected with scramble miRNA as controls. Compared to the controls, tumor growth was significantly reduced in animals receiving miR-145, most importantly on day 7 after the third and last dose of miRNA. After discontinuing the treatment, tumor growth resumed, becoming similar to the group of non-treated animals. A decrease in MYC and RAS expression was observed in all cell lines after treatment with miR-145, although statistical significance was achieved only in experiments with LNCaP and PC3 cell lines, with a decrease in 56% (p = 0.012) and 31% (p = 0.013) of RAS expression, respectively. Our results suggest that miR-145 is a potential molecule to be tested for treatment of metastatic, castration-resistant PCa.

Expression of micro-RNAs and genes related to angiogenesis in ccRCC and associations with tumor characteristics.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the third most common urological cancer in adults. Our aim is to evaluate genes and miRNAs expression profiles involved with angiogenesis and tumor characteristics in ccRCC. METHODS: The expression levels of miRNAs miR-99a, 99b, 100; 199a; 106a; 106b; 29a; 29b; 29c; 126; 200a, 200b and their respective target genes: mTOR, HIF1-α, VHL, PDGF, VEGF, VEGFR1 and VEGFR2 were analyzed using qRT-PCR in tumor tissue samples from 56 patients with ccRCC. Five samples of benign renal tissue were utilized as control. The expression levels of miRNAs and genes were related to tumor size, Fuhrman nuclear grade and microvascular invasion. RESULTS: miR99a was overexpressed in most samples and its target gene mTOR was underexpressed, this also occurs for miRNAs 106a, 106b, and their target gene VHL. An increase in miR-200b was correlated with high-risk tumors (p = 0.01) while miR-126 overexpression was associated with Fuhrman's low grade (p = 0.03). CONCLUSIONS: Our results show that in ccRCC there are changes in miRNAs expression affecting gene expression that could be important in determining the aggressiveness of this lethal neoplasia.

Correlation between chromosome 9p21 locus deletion and prognosis in clinically localized prostate cancer.

BACKGROUND: Some studies have reported that deletions at chromosome arm 9p occur frequently and represent a critical step in carcinogenesis of some neoplasms. Our aim was to evaluate the deletion of locus 9p21 and chromosomes 3, 7 and 17 in localized prostate cancer (PC) and correlate these alterations with prognostic factors and biochemical recurrence after surgery. METHODS: We retrospectively evaluated surgical specimens from 111 patients with localized PC who underwent radical prostatectomy. Biochemical recurrence was defined as a prostate-specific antigen (PSA) >0.2 ng/mL and the mean postoperative follow-up was 123 months. The deletions were evaluated using fluorescence in situ hybridization with centromeric and locus-specific probes in a tissue microarray containing 2 samples from each patient. We correlated the occurrence of any deletion with pathological stage, Gleason score, ISUP grade group, PSA and biochemical recurrence. RESULTS: We observed a loss of any probe in only 8 patients (7.2%). The most common deletion was the loss of locus 9p21, which occurred in 6.4% of cases. Deletions of chromosomes 3, 7 and 17 were observed in 2.3%, 1.2% and 1.8% patients, respectively. There was no correlation between chromosome loss and Gleason score, ISUP, PSA or stage. Biochemical recurrence occurred in 83% cases involving 9p21 deletions. Loss of 9p21 locus was significantly associated with time to recurrence (p = 0.038). CONCLUSIONS: We found low rates of deletion in chromosomes 3, 7 and 17 and 9p21 locus. We observed that 9p21 locus deletion was associated with worse prognosis in localized PC treated by radical prostatectomy.

MMP9 overexpression is associated with good surgical outcome in children with UPJO: Preliminary results.

BACKGROUND: Ureteropelvic junction obstruction (UPJO) diagnosed prenatally occurs in 1:150 - 1:1200 pregnancies. Although many studies investigating the molecular changes of this obstructed segment have been performed, the underlying mechanisms are still unclear. The role of extracellular matrix (ECM) components remains controversial, and the investigations in the field of ECM changes, might help the better understanding of the pathogenesis of this common condition. The aim of the present study was to investigate for the first time in the literature whether MMP9 and its specific inhibitors, TIMP1 and RECK, are expressed in a reproducible, specific pattern in UPJ. METHODS: UPJO specimens were obtained from 16 children at the time of dismembered pyeloplasty due to intrinsic UPJ stenosis. Expression levels of the three genes (MMP9, TIMP1 and RECK) were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). Then correlated the expression levels of the genes according to grade study population that was divided in 2 categories according to Society of Fetal Urology classification, grade 3 (moderate) and 4 (severe). For DTPA we subdivided the childrens in 2 groups, obstructive (T 1/2 more than 20 min) and partial obstructive (T 1/2 between 10 and 20 min) and success in a surgery was defined as decrease in T 1/2 to less than 20 min, absence of symptoms, improving renal function and decreasing dilatation on successive exams. RESULTS: MMP9 was underexpressed and TIMP1 and RECK were overexpressed in children with obstructive DTPA but the differences were not statistically significant. Overexpression of MMP9 was higher among patients with severe grade of UPJ compared to those with moderate grade. Surprisingly expression levels of MMP-9 was three times higher in children who were successfully treated by surgery (n = 10) (p = 0.072), so those who were followed for at least 1 year after surgery and remained with improvement in renal function and decreasing dilation on intravenous urogram and TIMP-1 was underexpressed in 100 % of this cases (p = 0.00). CONCLUSIONS: We showed an increase in expression of MMP9 and a decrease in expression of TIMP1 in children who improving renal function and decreasing dilation after surgery. We believe that the higher expression of MMP9 in these cases can reflect an increase in degradation and remodeling process that could be used as a marker for surgical outcome.

Loss of TIMP-1 immune expression and tumor recurrence in localized prostate cancer

Introduction and objective: Overexpression of MMPs has been related to biochemical recurrence after radical prostatectomy. TIMP1 and TIMP2 are controllers of MMPs and the aim of this study is to evaluate the expression levels of MMPs and their regulators using immunohistochemistry in tissue microarray of localized prostate cancer (PC). Materials and Methods: Immune-expression of MMP-9, MMP-2, TIMP1, TIMP-2, MMP-14 and IL8, were analyzed by immunohistochemistry in radical prostatectomy specimens of 40 patients with localized PC who underwent surgery between September 1997 and February 2000. Protein expression was considered as categorical variables, negative or positive. The results of the immune-expression were correlated to Gleason score (GS), pathological stage (TNM), pre-operatory PSA serum levels and biochemical recurrence in a mean follow up period of 92.5 months. Results: The loss of TIMP1 immune-expression was related to biochemical recurrence. When TIMP1 was negative, 56.3% patients recurred versus 22.2% of those whose TIMP1 was positive (p=0.042). MMP-9, MMP-2, IL8 and MMP-14 were positive in the majority of PC. TIMP-2 was negative in all cases. Conclusion: Negative immune-expression of TIMP1 is correlated with biochemical recurrence in patients with PC possibly by failing to control MMP-9, an important MMP related to cancer progression.

Loss of TIMP-1 immune expression and tumor recurrence in localized prostate cancer.

INTRODUCTION AND OBJECTIVE: Overexpression of MMPs has been related to biochemical recurrence after radical prostatectomy. TIMP1 and TIMP2 are controllers of MMPs and the aim of this study is to evaluate the expression levels of MMPs and their regulators using immunohistochemistry in tissue microarray of localized prostate cancer (PC). MATERIALS AND METHODS: Immune-expression of MMP-9, MMP-2, TIMP1, TIMP-2, MMP-14 and IL8, were analyzed by immunohistochemistry in radical prostatectomy specimens of 40 patients with localized PC who underwent surgery between September 1997 and February 2000. Protein expression was considered as categorical variables, negative or positive. The results of the immune-expression were correlated to Gleason score (GS), pathological stage (TNM), pre-operatory PSA serum levels and biochemical recurrence in a mean follow up period of 92.5 months. RESULTS: The loss of TIMP1 immune-expression was related to biochemical recurrence. When TIMP1 was negative, 56.3% patients recurred versus 22.2% of those whose TIMP1 was positive (p=0.042). MMP-9, MMP-2, IL8 and MMP-14 were positive in the majority of PC. TIMP-2 was negative in all cases. CONCLUSION: Negative immune-expression of TIMP1 is correlated with biochemical recurrence in patients with PC possibly by failing to control MMP-9, an important MMP related to cancer progression.

Immune expression of E-cadherin and α, ß and γ-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 Α-, Β- and y-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 Α-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 Α-catenin expression is related to tumor size in upper tract urothelial carcinomas.

Assessment of the in vitro and in vivo genotoxic and antigenotoxic effects of pimaradienoic acid in mammalian cells.

The pimarane-type diterpene, pimaradienoic acid (PA), is known for its diverse biological properties such as antimicrobial, anti-inflammatory and trypanocidal. A preliminary study was undertaken to investigate in vitro the free radical-scavenging potential of PA. In addition, the genotoxic potential of PA and its ability to modulate genotoxicity induced by doxorubicin (DXR) and methyl methanesulfonate (MMS) were studied in Chinese hamster lung fibroblasts (V79 cells) and in male Swiss mice using the comet and micronucleus assays. The DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate) assay showed that PA exerted no antioxidant activity when compared to quercetin. The colony-forming assay using V79 cells showed that PA was cytotoxic at concentrations >5.0µg/mL. Therefore, concentrations of 0.625, 1.25, 2.5, and 5.0µg/mL were used for evaluation of the genotoxic and antigenotoxic potential of PA in V79 cells. For genotoxic and antigenotoxic assessment in Swiss mice, three PA doses were tested (20, 40, and 80mg/kg body weight) based on the solubility limit of the diterpene in dimethylsulfoxide and water. The in vitro results demonstrated that PA induced DNA damage at concentrations of 2.5 and 5.0µg/mL in the comet assay. However, no genotoxic effect was observed in the micronucleus test using V79 cells. In the in vivo evaluation of genotoxicity, a significant increase in the frequency of DNA damage was observed in hepatocytes of animals treated with the highest PA dose (80mg/kg) when compared to the control group, but this difference was not seen in the micronucleus test. Furthermore, PA significantly reduced the frequency of DXR- and MMS-induced micronuclei and extent of DNA damage in in vitro and in vivo test systems.

Immune expression of E-cadherin and α, β and γ-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 α-, β- and γ-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 α-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 α-catenin expression is related to tumor size in upper tract urothelial carcinomas.

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.