Abstract 19960: Human Cardiac Fibroblasts Increase SR-Dependency of Induced Pluripotent Stem Cell-Derived Cardiomyocyte Calcium Handling by Increasing SR Calcium Uptake and SERCA2a Expression via Direct Physical Contact
Introduction: Cardiac fibroblasts can influence cardiomyocyte structure and function through direct physical interaction and/or by the secretion of soluble factors. Here we assess the relative importance of these different modalities of interaction in terms of their effects on cardiomyocyte electrophysiology.
Methods: Ventricular fibroblasts from the explanted hearts of dilated cardiomyopathy (DCM) patients (n=4) were seeded directly onto a confluent monolayer of human induced pluripotent stem cell-derived cardiomyocytes (iPS-CMs) at a ratio of 2:1 and cultured for 24 hours. iPS-CMs alone were used as control. Ca2+ transients and action potentials were recorded optically using fluo-4 AM and di-8-ANEPPS. Data are presented as percentage change from control where appropriate.
Results: iPS-CM Ca2+ transient duration was significantly reduced by contact with fibroblasts (-8.4±2.2% n=12 p<0.01). Sarcoplasmic reticulum (SR) Ca2+ uptake assessed by caffeine application was highly upregulated (2.07±0.12s-1 vs 0.63±0.07s-1 in control n=26, 23 p<0.001), associated with a significant increase in SR Ca2+ content (F/F0: 3.74±0.11 vs 2.97±0.11 in control n=37, 32 p<0.001) and SERCA2a protein expression (1.02±0.08 vs 0.55±0.01 n=3 p<0.01). iPS-CMs in contact with fibroblasts displayed a greater dependence on SR uptake for Ca2+ extrusion than other mechanisms (SR/NCX/Slow: 60.7/31.9/7.4% vs 23.3/70.9/5.8% in control).
Action potential duration (90%) was significantly prolonged in contact with fibroblasts (+18.8±2.5% n=22 p<0.001 vs control). Screening of culture supernatants from the co-culture has identified the presence of a number of cytokines known to affect cardiomyocyte function including IL-6 (680pg/ml), MIF (1.16ng/ml), MCP-1 (1.25ng/ml) and TGF-β. (110pg/ml).
Conclusions: DCM fibroblasts influence iPS-CM electrophysiology through different modalities; however physical contact with fibroblasts has effects not seen with soluble mediators alone. This contact-dependent shift towards greater SR-dependency represents a novel effect of fibroblasts on cardiomyocyte Ca2+ handling. Determining whether these effects are mediated through mechanical, electrical or paracrine signalling will be the focus of future work.
Author Disclosures: C. Kane: None. P. Dias: None. J. Gorelik: None. C.M. Terracciano: None.
- © 2015 by American Heart Association, Inc.