|Support type:||Scholarships in Brazil - Doctorate|
|Effective date (Start):||July 01, 2007|
|Effective date (End):||June 30, 2011|
|Field of knowledge:||Health Sciences - Pharmacy|
|Principal Investigator:||Rosario Dominguez Crespo Hirata|
|Grantee:||Fabiana Dalla Vecchia Genvigir|
|Home Institution:||Faculdade de Ciências Farmacêuticas (FCF). Universidade de São Paulo (USP). São Paulo , SP, Brazil|
ATP-binding cassette transporter A1 (ABCA1) is a transmembrane protein involved on cholesterol and phospholipids cellular efflux, which is an essential step for the cholesterol reverse transport. Polymorphisms in the ABCA1 gene have been associated with increased risk of coronary heart disease, differences on serum lipid profile and response to lowering-cholesterol drugs. Recently we have shown that atorvastatin, a potent HMGCoA reductase inhibitor, has a significant effect on ABCA1 mRNA expression in mononuclear cells in vivo and this effect is associated with R219K polymorphism. In this study, effects of atorvastatin on ABCA1 mRNA and protein expression will be evaluated in HepG2 and Caco-2 cells and in lymphocytes from hypercholesterolemic individuals (n=30). Total RNA and proteins will be extracted from cell culture after incubation with atorvastatin (0 to 20 µM for 24 h). The percentage of viable cells treated with atorvastatin will be determined by flow cytometry using propidium iodide solution (50 mg/mL in phosphate buffer saline) to detect membrane integrity of the cells. Expression of ABCA1 mRNA and protein will be evaluated by real-time PCR and Western blot and flow cytometry, respectively. Cholesterol and phospholipids efflux will be analyzed by beta counting. ABCA1 mRNA stability will be analyzed by Northern Blotting. Effects of atorvastatin on the activation of transcription factors (AP-1, SP-1, HNF3beta; e LXRalpha/beta) will be evaluated by electrophoretic mobility shift assays (EMSA). The possible link between -105C>T polymorphism and other variables such as ABCA1 mRNA expression and stability and transcription factors activity, as well as R219K polymorphism and cholesterol and phospholipids efflux will be studied in human lymphocytes. The results of this study will contribute for a better understanding of the ABCA1 transporter role on lowering-cholesterol response to atorvastatin.