The expression of matrix metalloproteinase-9 (MMP-9) has been implicated in progression of atherosclerotic lesions. The role and importance of the signaling pathway in the transcriptional regulation of MMP-9 in human aortic smooth muscle cells (HASMC) was examined. Tumor necrosis factor-alpha (TNF-alpha) stimulated the secretion of MMP-9 in HASMC, as shown by zymography and immunoblot analysis. At the transcriptional levels, TNF-alpha also stimulated the 5'-flanking 710-bp promoter activity of MMP-9. Transcription factors NF-kappaB binding site (-601) and AP-1 binding site (-82) were identified as the cis-elements for TNF-alpha activation, as determined by gel shift assay and mutation analysis. Treatment with U0126, an inhibitor of the extracellular signal-regulated kinase (ERK), significantly downregulated TNF-alpha-induced MMP-9 expression and promoter activity, whereas the inactive analog U0124 had no effect. Furthermore, the transactivation of TNF-alpha-stimulated NF-kappaB and AP-1 was inhibited by U0126 treatment. Finally, the transient transfection of HASMC with dominant negative Ras (RasN17) suppressed TNF-alpha-induced ERK activity, MMP-9 production, and promoter activity. Overexpression of RasN17 also abolished the TNF-alpha-stimulated NF-kappaB and AP-1 activity. In conclusion, the findings herein indicate the activation of the Ras/ERK pathway contributes to the induction of MMP-9 expression in HASMC. In addition, the transcription factors NF-kappaB and AP-1 that are involved in the Ras/ERK-mediated control of MMP-9 regulation on HASMC in response to TNF-alpha have now been identified.
BackgroundWhile several molecular markers of bladder cancer prognosis have been identified, the limited value of current prognostic markers has created the need for new molecular indicators of bladder cancer outcomes. The aim of this study was to identify genetic signatures associated with disease prognosis in bladder cancer.ResultsWe used 272 primary bladder cancer specimens for microarray analysis and real-time reverse transcriptase polymerase chain reaction (RT-PCR) analysis. Microarray gene expression analysis of randomly selected 165 primary bladder cancer specimens as an original cohort was carried out. Risk scores were applied to stratify prognosis-related gene classifiers. Prognosis-related gene classifiers were individually analyzed with tumor invasiveness (non-muscle invasive bladder cancer [NMIBC] and muscle invasive bladder cancer [MIBC]) and prognosis. We validated selected gene classifiers using RT-PCR in the original (165) and independent (107) cohorts. Ninety-seven genes related to disease progression among NMIBC patients were identified by microarray data analysis. Eight genes, a progression-related gene classifier in NMIBC, were selected for RT-PCR. The progression-related gene classifier in patients with NMIBC was closely correlated with progression in both original and independent cohorts. Furthermore, no patient with NMIBC in the good-prognosis signature group experienced cancer progression.ConclusionsWe identified progression-related gene classifier that has strong predictive value for determining disease outcome in NMIBC. This gene classifier could assist in selecting NMIBC patients who might benefit from more aggressive therapeutic intervention or surveillance.
Abstract. miRNAs are small, non-coding RNAs that play important roles in various biological processes. The aims of our study were to investigate whether cell-free miRNAs can be measured in urine samples and might be an accurate biomarker of bladder cancer. Datasets of GSE20418 and GSE19717 were used for analysis, and two miRNAs, miR-145 and miR-200a, were selected for study. A total of 207 patients with primary transitional cell carcinoma of the urinary bladder and 144 healthy normal controls were enrolled. Using quantitative PCR, the levels of miR-145 and miR-200a in urine were measured and compared with the clinicopathological features of bladder cancer. According to our experiments, cell-free miRNAs were present in urine and were stable. Assessment of miR-145 levels was able to distinguish bladder cancer patients from non-cancer controls (77.8% sensitivity and 61.1% specificity for NMIBC, AUC 0.729; 84.1 and 61.1% for MIBC, respectively, AUC 0.790) and showed good correlation with grade (p=0.048). In addition, miR-200a was shown to be an independent predictor of NMIBC recurrence by multivariate analysis (OR 0.449, p=0.013). A higher risk of recurrence was observed among patients with a lower miR-200a level compared to patients with higher miR-200a levels (log-rank test, p=0.040). Urinary cell-free miRNAs show promise as noninvasive biomarkers for diagnosis and recurrence of bladder cancer. IntroductionSurveillance strategies for bladder cancer recurrence have historically relied on the diagnostic combination of cystoscopy and urinary cytology. However, the cystoscopic approach is costly, invasive and uncomfortable. Urinary cytology is a preferable technique for the diagnosis of bladder tumors because of its high specificity; however, it has low sensitivity. For these reasons, many new urine-based tests for urinary bladder cancer have been developed, and screens for bladder tumor antigen (BTA), nuclear matrix protein 22 (NMP22), urine fibrin fibrinogen degradation products (FDP), ImmunoCyt and FISH (UroVysion) have all been approved for clinical use (1,2). However, the specificities of these new urine markers are low in comparison with urinary cytology, although they have higher sensitivities. Thus, none of the currently identified urine markers can replace cystoscopy or urinary cytology (3). miRNAs are small, nonprotein-coding RNA regulators involved in numerous biological and developmental processes (4,5). Cumulative evidence suggests that the dysregulation of miRNA plays an important role in many human disorders, including cancer. Approximately 50% of human miRNAs are encoded in genomic regions that are frequently altered in cancer (6-8). Recently, miRNAs have emerged as highly tissue-specific biomarkers with potential clinical applicability, not only as diagnostic markers but also as prognostic predictors for numerous cancers.Numerous recent studies have explored circulating cell-free miRNAs and provided evidence that miRNAs exist in a stable form in various body fluids, such as blood, urine, saliva, and p...
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