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Differential effects of Resveratrol on androgen-responsive LNCaP human prostate cancer cells in vitro and in vivo. P-5

Microarray analysis of the effect of resveratrol on global gene expression in LNCaP cells

Microarray gene expression profiling experiments were conducted as described previously. Briefly, LNCaP cells (2 × 107 cells per T-175 flask) were exposed to 0, 1, 5 or 25 μM resveratrol for 48 h, total RNA was isolated and microarray analysis was performed as described previously using the Affymetrix (Affymetrix, Santa Clara, CA) platform.

The Affymetrix U133 chip was used for gene expression analysis. Resveratrol-responsive genes were identified using the MAS 5.0 suite (Affymetrix) following criteria of a 1.5-fold increase and P < 0.01 as a cutoff as described previously. Triplicate treatments were performed for each concentration. Microarray data are available upon request and also will be deposited in public database.

Determination of the effects of resveratrol on gene expression in LNCaP cells using reverse transcription–polymerase chain reaction

LNCaP cells were plated in six-well plates (1 × 106 cells per well) in Media A and switched to Media B containing 10% CDS 24 h after plating to minimize the effect of serum hormones. Twenty-four hours later, the medium was replaced with fresh medium containing 1 nM R1881 or 17β-estradiol with or without 25 μM resveratrol. Fresh medium containing the test compounds was changed daily and cells were harvested for total RNA isolation using the Trizol method (Invitrogen) after 48 h as described previously.

Real-time reverse transcription–polymerase chain reaction (RT–PCR) was used to quantify expression of the following genes as described previously: PSA; beta-2-microglobulin (B2M); selenoprotein P, plasma, 1 (SEPP1); serine threonine kinase 39 (STK39); cyclin-dependent kinase inhibitor 1A/p21WAF1/CIP1 (CDKN1A) and insulin-like growth factor-1 receptor (IGF-1R).

Article Produced By

Diet, Genomics and Immunology Laboratory, Beltsville Human Nutrition Research Center, Agriculture Research Service, United States Department of Agriculture, 10300 Baltimore Avenue, Building 307C, Room 132, Beltsville, MD. Laboratory of Cellular Regulation and Carcinogenesis, National Cancer Institutes, National Institutes of Health, Department of Nutrition and Food Science, University of Maryland, College Park, Department of Pharmaceutical Sciences and Pharmacology and Toxicology Graduate Program College of Pharmacy, Washington State University, Pullman, National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan. Nutritional Sciences Research Group, Division of Cancer Prevention, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. Biometrical Consulting Service, Beltsville Area, Agriculture Research Service, United States Department of Agriculture, Beltsville, MD 20705, USA. Division of Nutritional Sciences, University of Texas at Austin, Austin, TX 78712, USA. Department of Carcinogenesis, MD Anderson Cancer Center, Smithville, TX 78957, USA
 

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