DDAH1 effects on the development of congestive heart failure Abstract Nitric oxide (NO) produced by endothelial nitric oxide synthase (eNOS) is important in maintaining vascular endothelial function and in protecting the heart from adverse ventricular remodeling. Accumulation of the endogenous NOS inhibitors asymmetric dimethyl arginine (ADMA) and Ng-monomethyl-L-arginine (L-NMMA) is a major independent risk factor for cardiovascular diseases including hypertension, congestive heart failure and atherosclerosis. These endogenous NOS inhibitors compete with L-arginine to inhibit NO production by eNOS. ADMA and L-NMMA are eliminated principally by metabolism to L-citrulline by dimethylarginine dimethylaminohydrolase (DDAH). DDAH1 and DDAH2 are encoded by two different genes. We have generated preliminary data indicating that DDAH1 rather than DDAH2 plays the essential role in degrading the NOS inhibitors in tissues such as kidney and brain and thereby regulating NO bioavailability in these tissues. In the heart we find DDAH1 expressed both in coronary endothelium and under the sarcolemma of cardiac myocytes. To address the cell specific role of DDAH1 in regulating endogenous NOS inhibitors, NO bioavailability and cardiovascular function, we have generated three novel tissue specific DDAH1 KO mouse strains. Using these new strains, we propose studies to determine whether the cardioprotective effect of DDAH1 resides in DDAH1 expressed in the cardiac myocytes or in the coronary endothelium.
Specific aims will be addressed: (i) Determine the role of total-DDAH1, and DDAH1 expressed in the vascular endothelium in degrading myocardial ADMA and L-NMMA using endothelial specific and global DDAH1 gene deficient mice;(ii) Determine the cardiac protective effect from DDAH1 expressed in the cardiomyocytes on the development of heart failure produced by chronic systolic overload;and (iii) elucidate the protective effect from DDAH1 expressed in the vascular endothelium on the development of heart failure after chronic systolic overload. We will examine both changes in NO production and ROS generation in the different KO mouse strains. These unique tissue specific DDAH1 KO mice generated in our laboratory will allow us to elucidate cell type specific actions of DDAH1, as well as molecular mechanisms by which DDAH1 protects the overloaded heart.
Cardiovascular disease ranks as America's No. 1 killer that accounts for nearly one million deaths each year. Nitric oxide (NO) is known to exert protective effects on the heart. Accumulation of the endogenous nitric oxide synthase (NOS) inhibitors ADMA and L-NMMA is associated with increased cardiac death and the development of various cardiovascular diseases such as hypertension, coronary disease, atherosclerosis and congestive heart failure (CHF). ADMA and L-NMMA are degraded by the enzyme dimethylarginine dimethylaminohydrolase (DDAH). However, our understanding of the physiology and pathology of this NOS inhibitor system is very limited. For example, although DDAH is reported to increase NO bioavailability by degradation of ADMA and L-NMMA, the in vivo role of DDAH1 vs. DDAH2 in regulating NO bioavailability is not clear. In addition, it is not known whether chronic accumulation of these endogenous NOS inhibitors can directly cause or exacerbate cardiovascular disease. The studies proposed in this application will use unique tissue specific KO mice generated in our laboratory to elucidate both isoform- and cell type specific actions of DDAH. The central hypotheses to be tested are that (i) DDAH1 (not DDAH2) is the essential or sole enzyme responsible for degradation of ADMA and LNMMA in cardiovascular system, (ii) deletion of endothelial DDAH1 will cause accumulation of the endogenous NOS inhibitors and systemic hypertension, and (iii) deletion of DDAH1 will exacerbate the development of CHF in the overloaded heart by decreasing NO bioavailability. We have concrete preliminary data to support these hypotheses. We will examine the influence of chronic accumulation of ADMA and L-NMMA on myocardial NO-bioavailability and on NOS-derived ROS generation. Finally, we will determine whether deletion of DDAH1 in endothelium or in cardiac myocytes impairs the ability of the heart to adapt to chronic pressure overload produced by transverse aortic constriction in mice. These studies will provide new knowledge regarding how DDAH1 acts to regulate production of NO and NOS-derived ROS, and demonstrate whether dysregulation of DDAH1 in either cardiac myocytes or endothelium can contribute to the development of CHF.
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