This is a proposal for a Competing Supplement to NHLBI grant #HL53318, Intercellular Communication in Microvessels. The overarching hypotheses or themes are that smooth muscle and endothelium function as an integrated unit in the regulation of cardiovascular function, and 2.) that the coordinating molecule uniting the two cell types is the connexin. The parent R-01 proposes to study three of the connexins; 43, 40, and 37. Knockout animals for connexin 40 and 37 have been obtained from a collaborator. Because deletion of the connexin 43 is embryologically lethal, we have created cell-specific knockouts in which connexin 43 expression is selectively eliminated in either smooth muscle or endothelium. At the time of submission of the parent R-01 grant we indicated that fuller understanding of the biology of the connexins in the vessel wall would necessitate the production of double knockouts through appropriate cross breeding. We selected connexin 43 and 40 as the first double knockout to be produced based on the facts that we could restrict deletion of 43 to the endothelium, and that our immuno-cytochemistry shows connexin 40 in the mouse to be abundantly expresses in the endothelium compared to the other connexins. We have now found that the endothelial cell-specific Cx43 KO crossed with the Cx40 KO produces rapidly advancing hypertension, profound cardiac hypertrophy and failure at about 3 months of age. The phenotype appears to be a model for overload cardiac hypertrophy and failure. Importantly, animal heterozygous for the endothelial cell deletion show a similar phenotype attesting to the penetrance of the phenotype. We have established a collaboration with the cardiac MRI group here and are now able to measure the progression of cardiac adaptation and failure in this model. These data support the need for an accelerated breeding program to designed to generate sufficient double knockouts, and to apply high throughput screening to the assessment of function.
The specific aims of the Competing Supplement are: 1. to expand our breeding and animal maintenance to allow us to explore the vascular function in animals with double connexin knockouts. 2. to conduct longitudinal measurements on changes in cardiovascular physiology caused by the elimination of one or more of the connexin genes. Specifically, we will follow blood pressure, cardiac output, heart rate, EKG, and peripheral resistance, over several months of the development of hypertension, cardiac hypertrophy and ultimately failure. These data will be correlated with our ongoing measurements of microvascular function. ? ?
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