Anales RANF

P.90 PURINE METABOLISM IN CARDIAC FIBROBLASTS AND EPICARDIUM-DERIVED CELLS, SIMULTANEOUSLY ISOLATED FROM THE INFARCTED MOUSE HEART C. Owenier 1 , C. Alter 1 , J. Hesse 1 , Z. Ding 1 , J. Schrader 1 1 . Heinrich-Heine-University Düsseldorf, Düsseldorf, Germany. Release of nucleotides from the infarcted heart and their extracellular conversion to adenosine play an important role in the process of post-infarct tissue remodeling. Purines are known to have impact on the differentiation of cardiac fibroblasts (CF) into myofibroblasts (= activated cardiac fibroblasts (aCF)). However, the CF/aCF population is most likely heterogeneous and hard to capture by conventionally used markers. Another issue is the contamination of the CF/aCF population with epicardium-derived cells (EPDC), which are formed on the epicardial surface above the infarcted tissue. To discriminate between aCF and EPDC, we established a fast (8 min) and efficient protocol for the simultaneous isolation of aCF and EPDC from the infarcted mouse heart. Quantitative PCR analysis of genes involved in the extracellular purine metabolism revealed that in all cell fractions Enpp1 and Enpp3 is more highly expressed than CD39. In functional assays, however, ATP breakdown was largely inhibited when cells lacked CD39, suggesting an only minor role of Enpp1/3 in ATP breakdown. Concerning the four adenosine receptors, EPDC highly expressed Adora2b, while CF/aCF showed lower expression levels. Analysis of the kinetics of extracellular ATP metabolism in short term cultured aCF and EPDC showed that the main degradation product is AMP with only negligible adenosine formation. This finding supports the hypothesis that AMP generated by aCF and EPDC diffuses to surrounding immune cells to serve as a substrate for CD73 mainly localized on T cells, thereby producing anti- inflammatory adenosine in the infarcted heart.