[Frontiers in Bioscience 11, 2275-2285, September 1, 2006]

Growth inhibitory effects of quercetin on bladder cancer cell

Li Ma 1,2, Jean Magloire Feugang 3, Patricia Konarski 3, Jian Wang 3, Jianzhong Lu 1, Shengjun Fu 1, Baoliang Ma 1, Binqiang Tian 1, Changping Zou 3 and Zhingping Wang 1

1 The second hospital of Lanzhou University, Lanzhou, Gansu, China, 2 Life Science School of Lanzhou University, Lanzhou, Gansu, China, 3 Department of OB/GYN, Gynecologic Oncology, University of Arizona, Tucson, Arizona

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and Methods
3.1. Reagents
3.2. Cell culture and treatment
3.3. Cytotoxicity assay
3.4. Cell growth assay
3.5. Colony formation assay
3.6. Detection of apoptosis by two different methods
3.6.1. Hoechst 33258 staining
3.6.2. TUNEL staining
3.7. Cell cycle analysis
3.8. Methylation-specific polymerase chain reaction (MSP-PCR)
3.9. Immunocytochemical staining
4. Results
4.1. Quercetin affects the cell viability
4.2. Quercetin inhibits cell growth and colony formation
4.3. Apoptosis induced by quercetin
4.4. Quercetin affects the cell cycle
4.5. Quercetin decreases the DNA methylation levels of P16 INK4a, Er beta, and RASSF1A genes
4.6. Quercetin inhibits expression of mutant P53 and surviving proteins
4.7. Quercetin does not affect the expression of tumor suppressor gene PTEN
5. Discussion
6. Conclusions
7. Acknowledgments
8. References

1. ABSTRACT

Quercetin, a flavonoid found in many fruits and vegetables, belongs to an extensive class of polyphenolic compounds. Previous studies reported that quercetin inhibits the proliferation of various cancer cells and tumor growth in animal models. We investigated the growth inhibition and colony formation of quercetin on three bladder cancer cells (EJ, J82 and T24). The expression of tumor suppressor genes and oncogenes such as P53, Survivin, PTEN, as well as the methylation status of these genes was also evaluated. We observed that quercetin induced apoptosis in bladder cancer cells in a time- and dose-dependent manner. Quercetin (100 micromolars) significantly inhibited EJ, T24 and J82 cell growth accompanied by an increase in the G0/G1 phase. In all cell lines, quercetin decreased the expression of mutant P53 and Survivin proteins. However, there was no change in the level of PTEN protein. Moreover, the DNA methylation levels of the estrogen receptor (Er-beta), P16INK4a and RASSF1A were strongly decreased (from 35 to 70%) in the quercetin-treated group compared to the control. In conclusion, our study suggested that quercetin inhibits growth, colony formation and hypermethylation of bladder cancer cell lines. Quercetin-induced apoptosis might be associated with a decrease in mutant P53 and Survivin proteins.