Introduction: Cardiac fibroblasts are key regulators of the extracellular matrix (ECM), which forms the mechanical environment for cardiomyocytes through extensive integrin binding. Arginine-Glycine-Aspartic Acid (RGD)-motif peptides are integrin binding motifs contained in fibronectin that have been implicated in the regulation of several structural and functional cellular properties. However, the effects of RGD-motif peptides and integrin binding on human myocytes calcium cycling - a fundamental mechanism in normal cardiac function universally altered during disease - are unknown.
Methods: Human induced pluripotent stem-cell derived cardiomyocytes (hiPSC-CMs) were incubated for 24 hours with (1) the RGD peptides GRGDS and GRGDSP (2mM), (2) anti β1 and β3 integrin antibodies (50μg/ml) or (3) both concurrently. Cytoplasmic calcium transients were measured with Fluo4-AM (n/N = total number of areas/total number of dishes).
Results: GRGDS and GRGDSP abbreviate hiPSC-CM calcium transients by reducing time to peak (Fig 1B, ### = p<0.001) and time to 50% decay (Fig 1C, ## = p<0.01). β1 integrin antibodies cause similar changes in calcium transients (Fig 1F, p<0.01). The effects of RGD motif-containing peptides are potentiated by anti-β1 or anti-β3-integrin antibodies (not shown).
Conclusions: Integrin binding ECM proteins cause abbreviation of human myocyte calcium transients, and may have a functional role in physiology and during cardiac disease.