Abstract 16963: Ankyrin-b-Based Protein Complex Regulates the Ischemic Response in Heart
Background: In heart, ankyrin-B is critical for targeting cardiac membrane proteins. Specially, ankyrin-B dysfunction has been identified following myocardial infarction (MI), suggesting an important role for ankyrin-B in ischemic heart disease. Thus, we hypothesize that ankyrin-B may interact with the ischemia-related membrane-binding partners: KATp channels and the cardiac Na/K ATPase.
Methods: Langendorff-perfused hearts and simulated ischemia in myocytes were employed to check the ischemia response in WT and ankyrin-B+/−mice. Macropatch IKATp and single-channel IKATp were recorded. Co-immunoprecipitation and pulldown analysis were performed in both myocytes and cultured HEK-293. The expression level of related components was examined by immunoblot.
Results: Ankyrin-B+/− mice displayed 38.8% increase in MI size after global ischemia (38.28%±3.84 vs 27.57%±2.78, p<0.05), and a reduced preconditioning response (26.56%±2.19 vs 20.28%±1.71, p< 0.05) when compared with WT mice. Furthermore, unlike WT myocytes, ankyrin-B+/− myocytes displayed no difference in post ischemia viability in response to preconditioning. Notably, ankyrin-B+/− ventricle showed decreased expression of both SUR1 and SUR2A (51% & 47%, n=4, p<0.01) as well as decreased Kir6.2 (∼46%, n=4, p<0.01) and ankyrin-B (n=4, p<0.001). Kir6.1 was not altered by ankyrin-B deficiency. Na/Ca exchanger (NCX) and Na/K ATPase expression levels were also reduced in ankyrin-B+/− ventricle (n=4, p<0.05). Next, we observed 50% decrease of I KATp in ankyrin-B+/− myocytes. Interestingly, a nkyrin-B+/− myocytes were less sensitive to ATP when compared with WT myocytes (K1/2: 24.38 μM vs 41.47 μM, p<0.01). The slope index was equivalent between myocyte genotypes. In the absence of ATP, we observed a significant difference between Po in ankyrin-B+/− myocytes compared with WT myocytes (0.62±0.03 vs 0.48±0.05, n=10; p<0.05). We further demonstrated that KATp channel directly associate with ankyrin-B in heart via Kir6.2. Finally, we demonstrate that ankyrin-B forms a membrane complex with KATp channels and the cardiac Na/K ATPase.
Conclusions: Our new findings define a new cardioprotective role for ankyrin polypeptides through targeting key membrane ion channel and transporters.
- © 2010 by American Heart Association, Inc.