Abstract 10255: Cardiac Fibroblasts Regulate Adult Cardiomyocyte Excitation Contraction Coupling by Soluble Mediators in Co-Culture
Cardiac fibroblasts (CF) are essential constituents of the normal heart and important players in the pathogenesis of heart disease. Using a system which only allows paracrine communication, we tested whether cultured CFs affect adult cardiomyocyte excitation-contraction coupling (ECC) and structure. LV CFs were isolated from Sprague-Dawley adult rats (150-250 g) and after 2 weeks of culture, were cultured on Transwells® above isolated adult Sprague-Dawley LV myocytes (CM+CF) for 24 hours. As a control cardiomyocytes were plated with other cardiomyocytes on the transwell (CM+CM). Di-8-ANEPPS was used to assess cell volume and T-tubule structure, Ca2+ handling was assessed using Indo-1 and Fluo-4 and stimulated sarcomere shortening was recorded. The culture medium was analysed for soluble mediators using cytokine arrays and ELISA kits. Data was analysed using t-tests and presented as mean±SEM. Cardiomyocytes from CM+CF are larger (21040 ± 1234μm3 (n=33) vs. 25910 ± 1604μm3 (n=28) p<0.05) with increased T-tubule density (40.15±1.2 (n=28) vs. 36.27±1.0 (n=29) p<0.05) but unaltered T-tubule regularity. Sarcomere shortening shows a lower amplitude (0.063±0.005μm (n=78) vs. 0.075±0.004μm (n=143) p<0.05) and greater time to 90% peak (0.100±0.004ms (n=92) vs. 0.088±0.003ms (n=141) p<0.05) in the CM+CF group. Ca2+ transients amplitude is depressed (2.74±0.19 f/f0 (n=34) vs. 3.25±0.17f/f0 (n=49) p<0.05) and sarcoplasmic reticulum Ca2+ content reduced (0.2691±0.01676 (n=34) vs. 0.3426±0.01997 (n=32) p<0.01). Ca2+ spark frequency is unaltered but spark duration, peak and width are all reduced in the CM+CF group (p<0.05). TIMP-1 (673.5±41.11 vs. 0.014±0.002 n=4 p<0.01), CINC-1 (254.8±50.05 vs. 0.014 ± 0.003 n=4 p<0.01), VEGF (136.4±17.13 vs. 61.30±19.90 n=4 p<0.05), IGF-1 (0.0300±0.00058 vs. 0.02603±0.000033 n=3 p<0.01) and endothelin (3.225±1.232 vs. 0.4706±0.1841 n=6 p<0.05) are all increased in the CM+CF group. This study shows that cultured CFs can influence adult cardiomyocyte ECC via paracrine communication. These findings are important in understanding the potential of CFs to modulate the activity of cardiomyocytes, and stimulate the need to examine the role of fibroblasts in physiological conditions and during cardiac disease.
- © 2011 by American Heart Association, Inc.