Abstract 15688: Expression of Antithrombotic Ecto-enzyme ENTPD1 Driven by PDE3 Inhibition
Ectonucleoside triphosphate diphosphohydrolase 1 (or CD39), an enzyme expressed on the plasmalemma of leukocytes and endothelial cells, serves as a potent antithrombotic agent by sequentially phosphohydrolyzing ATP and ADP, to generate AMP. CD39 is thus able to locally deplete the levels of extracellular ATP and ADP, which would otherwise promote inflammation and platelet aggregation by binding P2X and P2Y classes of purinergic receptors found on leukocytes and platelets. The only known pathway for the transcriptional regulation of CD39 was described in recent work by this lab, which demonstrated that increased cAMP levels led to CD39 upregulation in RAW macrophages through the cAMP response elements (CRE) in the CD39 promoter. Cilostazol, a drug currently used in the treatment of claudication in association with peripheral arterial disease, is a specific inhibitor of phosphodiesterase 3 (PDE3), an enzyme responsible for the intracellular depletion of cAMP. Therefore, we hypothesize that cilostazol treatment will upregulate CD39 expression via the blockade of PDE3. Treatment of RAW cells with 30μM cilostazol led to a 3.2-fold increase in CD39 mRNA (18hrs, p<0.01), a 42% increase in CD39 protein (21hrs, p<0.03), and an 8-fold increase in CD39 enzymatic activity (p<0.05), compared to control cells treated with DMSO. To demonstrate that the upregulation of CD39 was indeed specific to PDE3 inhibition the drug milrinone, another PDE3 inhibitor, was used as a treatment on RAW macrophages. Milrinone treatment led to a 74% increase in CD39 protein (50μM, 21hrs, p<0.05) compared to control cells treated with DMSO. Together, these data demonstrate that the expression of functional CD39, a potent antithrombotic and anti-inflammatory enzyme, can be upregulated by the specific blockade of PDE3. This is especially significant since our data shows that CD39 levels can be increased with the administration of a drug that has already approved and tested over time.
- © 2011 by American Heart Association, Inc.