Xiangyi Zheng

Resveratrol induces apoptosis and cell cycle arrest of human T24 bladder cancer cells in vitro and inhibits tumor growth in vivo

Resveratrol, a naturally occurring polyphenolic antioxidant compound present in grapes and red wine, has been reported to hold various biochemical responses. In this preliminary study, we evaluate the chemopreventive potential of resveratrol against bladder cancer and its mechanism of action. Treatment of bladder cancer cells with resveratrol resulted in a significant decrease in cell viability. Resveratrol induced apoptosis through the modulation of Bcl‐2 family proteins and activation of caspase 9 and caspase 3 followed by poly(ADP‐ribose) polymerase degradation. Treatment with resveratrol led to G1 phase cell cycle arrest in T24 cells by activation of p21 and downregulation of cyclin D1, cyclin‐dependent kinase 4, and phosphorylated Rb. Resveratrol also inhibited the phosphorylation of Akt, whereas the phosphorylation of p38 MAPK was enhanced. In addition, resveratrol treatment decreased the expression of vascular endothelial growth factor and fibroblast growth factor‐2, which might contribute to the inhibition of tumor growth on the bladder cancer xenograft model. These findings suggest that reveratrol could be an important chemoprevention agent for bladder cancer. (Cancer Sci 2009)

Cyclin-dependent kinase 4 is a novel target in micoRNA-195-mediated cell cycle arrest in bladder cancer cells

miRNAs are a class of small‐noncoding RNAs capable of negatively regulating gene expression. Here, we found that miR‐195 is down‐regulated in human bladder cancer tissue versus normal adjacent tissue. To better characterize the role of miR‐195 in bladder cancer, we conducted gain of function analysis by transfecting bladder cancer cell line T24 with chemically synthesized miR‐195 mimic. We identified CDK4, an early G1 cell cycle regulator, as a novel target of miR‐195. Selective over‐expression of miR‐195 could induce G1‐phase arrest in T24 cells, and subsequently inhibit T24 cell growth. These findings indicate that miR‐195 could be a potential tumor suppressor in bladder cancer.

MicroRNA-124-3p inhibits cell migration and invasion in bladder cancer cells by targeting ROCK1

BackgroundIncreasing evidence has suggested that dysregulation of certain microRNAs (miRNAs) may contribute to human disease including carcinogenesis and tumor metastasis in human. miR-124-3p is down-regulated in various cancers, and modulates proliferation and aggressiveness of cancer cells. However, the roles of miR-124-3p in human bladder cancer are elusive. Thus, this study was conducted to investigate the biological functions and its molecular mechanisms of miR-124-3p in human bladder cancer cell lines, discussing whether it has a potential to be a therapeutic biomarker of bladder cancer.MethodsThree human bladder cancer cell lines and samples from ten patients with bladder cancer were analyzed for the expression of miR-124-3p by quantitative RT--PCR. Exogenetic overexpression of miR-124-3p was established by transfecting mimics into T24, UM-UC-3 and J82 cells, after that cell proliferation and cell cycle were assessed by MTT assay, flow cytometry and Colony-forming assay. Cell motility and invasion ability were evaluated by wound healing assay and transwell assay. Tissue microarray, and immunohistochemistry with antibodies against ROCK1, MMP2 and MMP9 was performed using the peroxidase and DAB methods. The target gene of miR-124-3p was determined by luciferase assays, quantitative RT--PCR and western blot. The regulation of epithelial-to-mesenchymal transition by miR-124-3p was analyzed by western blot.ResultsmiR-124-3p is frequently down-regulated in bladder cancer both in three bladder cancer cell lines, T24, UM-UC-3, J82 and clinical samples. Overexpression of miR-124-3p induced G1-phase arrest in T24, UM-UC-3 and J82 cell lines and suppressed cell growth in colony-forming assay. miR-124-3p significantly repressed the capability of migration and invasion of bladder cancer cells. In addition, ROCK1 was identified as a new target of miR-124-3p. ROCK1, MMP2, MMP9 were up-regulated in bladder cancer tissues. Furthermore, we demonstrated miR-124-3p could inhibit bladder cancer cell epithelial mesenchymal transfer, and regulated the expression of c-Met, MMP2, MMP9.ConclusionsmiR-124-3p can repress the migration and invasion of bladder cancer cells via regulating ROCK1. Our data indicate that miR-124-3p could be a tumor suppressor and may have a potential to be a diagnostics or predictive biomarker in bladder cancer.

MicroRNA-449a acts as a tumor suppressor in human bladder cancer through the regulation of pocket proteins

Frequent downregulation of microRNA-449a (miR-449a) was detected in 14 human bladder cancer tissues. The restoration of miR-449a inhibited cell growth and induced G1-phase arrest in T24 and 5637 human bladder cancer cells. CDK6 and CDC25a were downregulated after miR-449a treatment, resulting in the functional accumulation of the pocket proteins Rb and p130. The growth of T24 tumor xenografts was suppressed by exogenous miR-449a, and the nuclear proliferation antigen Ki-67 was downregulated in miR-449a-treated tumors. These results suggest a tumor-suppressive role for miR-449a in human bladder cancer.

Up-regulation of p21WAF1/Cip1 by saRNA induces G1-phase arrest and apoptosis in T24 human bladder cancer cells

Very recent studies have reported that chemically synthesized small duplex RNAs complementary to the promoters of target genes can activate gene expression in different cancer cell lines. Such dsRNA have been referred to as saRNA for small activating RNA. The present study was conducted to evaluate the potential of p21(WAF1/Cip1) (p21) induction by small activating RNA targeting the p21 promoter in the treatment of bladder cancer. Using T24 human bladder cancer cells, we found that p21 saRNA caused dose- and time-dependent inhibition of cell proliferation and viability which was associated with induced G1-phase cell cycle arrest and apoptosis. The decreased anti-apoptotic protein Bcl-xL and activation of caspase-3 and PARP also supported the efficacy of the treatment. These data suggest that up-regulation of p21 by saRNA may be an effective way for treating human bladder and other types of cancers.

Up-regulation of E-cadherin by small activating RNA inhibits cell invasion and migration in 5637 human bladder cancer cells

Recent studies have reported that chemically synthesized small duplex RNAs complementary to promoters of target genes can specifically induce gene expression in several cancer cell lines. Such dsRNA, referred to as small activating RNA (saRNA), are involved in the recently described phenomenon called RNA activation (RNAa). Recent findings show that saRNA can inhibit cell proliferation and viability via up-regulation of p21(WAF1/CIP1) (p21) in human bladder cancer cells. In the present study, we demonstrate that induction of E-cadherin expression by saRNA leads to suppression of migration and invasion of 5637 human bladder cancer cells in vitro. The elevated E-cadherin expression was confirmed at transcriptional and protein levels after transfection of a 21-nucleotide dsRNA targeting the E-cadherin promoter (dsEcad). Furthermore, this inhibitory effect was associated with relocalization of beta-catenin from the nucleus to the plasma membrane and decreased beta-catenin-mediated transactivation. These data suggest that activation of E-cadherin by saRNA may have a therapeutic benefit for bladder and other types of cancer.

MicroRNA-409-3p Inhibits Migration and Invasion of Bladder Cancer Cells via Targeting c-Met

There is increasing evidence suggesting that dysregulation of certain microRNAs (miRNAs) may contribute to tumor progression and metastasis. Previous studies have shown that miR-409-3p is dysregulated in some malignancies, but its role in bladder cancer is still unknown. Here, we find that miR-409-3p is down-regulated in human bladder cancer tissues and cell lines. Enforced expression of miR-409-3p in bladder cancer cells significantly reduced their migration and invasion without affecting cell viability. Bioinformatics analysis identified the pro-metastatic gene c-Met as a potential miR-409-3p target. Further studies indicated that miR-409-3p suppressed the expression of c-Met by binding to its 3′-untranslated region. Silencing of c-Met by small interfering RNAs phenocopied the effects of miR-409-3p overexpression, whereas restoration of c-Met in bladder cancer cells bladder cancer cells overexpressing miR-409-3p, partially reversed the suppressive effects of miR-409-3p. We further showed that MMP2 and MMP9 may be downstream effector proteins of miR-409-3p. These findings indicate that miR-409-3p could be a potential tumor suppressor in bladder cancer.

MicroRNA-490-5p inhibits proliferation of bladder cancer by targeting c-Fos.

MicroRNAs (miRNAs) are non-protein-coding sequences that play a crucial role in tumorigenesis by negatively regulating gene expression. Here, we found that miR-490-5p is down-regulated in human bladder cancer tissue and cell lines compared to normal adjacent tissue and a non-malignant cell line. To better characterize the function of miR-490-5p in bladder cancer, we over-expressed miR-490-5p in bladder cancer cell lines with chemically synthesized mimics. Enforced expression of miR-490-5p in bladder cancer cells significantly inhibited the cell proliferation via G1-phase arrest. Further studies found the decreased c-Fos expression at both mRNA and protein levels and Luciferase reporter assays demonstrated that c-Fos is a direct target of miR-490-5p in bladder cancer. These findings indicate miR-490-5p to be a novel tumor suppressor of bladder cancer cell proliferation through targeting c-Fos.

Obesity and Risk of Bladder Cancer: A Meta-analysis of Cohort Studies

OBJECTIVE Previous epidemiologic studies demonstrated that obesity might associated with the risk of bladder cancer. However, many of the actual association findings remained conflicting. To better clarify and provide a comprehensive summary of the correlation between obesity and bladder cancer risk, we conducted a meta-analysis to summarize results of studies on the issue. Stratified analyses were also performed on potential variables and characteristics. METHODS Studies were identified by searching in PubMed and Wanfang databases, covering all the papers published from their inception to March 10, 2013. Summary relative risks (SRRs) with their corresponding 95% confidence intervals (CIs) were calculated by either random-effect or fixed-effect models. RESULTS A total of 11 cohort studies were included in our meta-analysis, which showed that obesity was associated with an increased risk for bladder cancer in all subjects (RR=1.10, 95% CI=1.06-1.16; p=0.215 for heterogeneity; I2=24.0%). Among the 9 studies that controlled for cigarette smoking, the pooled RR was 1.09 (95% CI 1.01-1.17; p=0.131 for heterogeneity; I2=35.9%). No significant publication bias was detected (p = 0.244 for Eggers regression asymmetry test). CONCLUSIONS Our results support the conclusion that obesity is associated with the increased risk of bladder cancer. Further research is needed to generate a better understanding of the correlation and to provide more convincing evidence for clinical intervention in the prevention of bladder cancer.

miR-26a inhibits proliferation and motility in bladder cancer by targeting HMGA1.

It is increasingly clear that microRNAs play a crucial role in tumorigenesis. Recently, emerging evidence suggested that miR‐26a is aberrantly expressed in tumor tissues. In our study, frequent down‐regulation of miR‐26a was observed in 10 human bladder cancer tissues. Forced expression of miR‐26a in the bladder cancer cell line T24 inhibited cell proliferation and impaired cell motility. High mobility group AT‐hook 1 (HMGA1), a gene that modulates cell cycle transition and cell motility, was verified as a novel target of miR‐26a in bladder cancer. These findings indicate an important role for miR‐26a in the molecular etiology of bladder cancer and implicate the potential application of miR‐26a in bladder cancer therapy.