Extracellular Vesicles from Human Cardiac Fibroblasts Modulate Calcium Cycling in Human Stem Cell-Derived Cardiomyocytes

Abstract

Cardiac fibroblasts regulate the development of the adult cardiomyocyte phenotype and cardiac remodeling in disease. We investigate the role that cardiac fibroblasts-secreted extracellular vesicles (EVs) have in the modulation of cardiomyocyte Ca2+ cycling–a fundamental mechanism in cardiomyocyte function universally altered during disease. EVs collected from cultured human cardiac ventricular fibroblasts were purified by centrifugation, ultrafiltration and size-exclusion chromatography. The presence of EVs and EV markers were identified by dot blot analysis and electron microscopy. Fibroblast-conditioned media contains liposomal particles with a characteristic EV phenotype. EV markers CD9, CD63 and CD81 were highly expressed in chromatography fractions that elute earlier (Fractions 1–15), with most soluble contaminating proteins in the later fractions collected (Fractions 16–30). Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were treated with fibroblast-secreted EVs and intracellular Ca2+ transients were analyzed. Fibroblast-secreted EVs abbreviate the Ca2+ transient time to peak and time to 50% decay versus serum-free controls. Thus, EVs from human cardiac fibroblasts represent a novel mediator of human fibroblast-cardiomyocyte interaction, increasing the efficiency of hiPSC-CM Ca2+ handling.

Stem Cell-Derived Extracellular Vesicles in the Treatment of Cardiovascular Diseases

“…Cardiac fibroblasts, which constitute 20% of the heart’s non-myocyte component, might present a solution to the problem of the optimal delivery of therapeutic EVs to the damaged myocardium [ 174 ]. Cardiac fibroblasts (CFs), cells typically of mesenchymal origin, are found in myocardial connective tissue and maintain structure and function by producing and secreting extracellular matrix components, such as collagen, elastin, and glycoproteins [ 175 , 176 , 177 ]. They also play a role in regulating myocardium remodeling during disease by their locally secreted EVs that may carry bioactive molecules, like receptor tyrosine kinases (e.g., DDR2) or proteases (e.g., MMP2), that help regulate various processes, including inflammation, fibrosis, angiogenesis, and cardiomyocyte function [ 174 , 178 , 179 , 180 , 181 , 182 ].…”

“…In contrast, CFEVs have been shown to enhance the efficiency of intracellular Ca 2+ cycling, an important aspect of excitation–contraction coupling, in human iPSC progenitor cardiomyocytes, suggesting an intimate relationship between cardiac stem cell species and the tissue-resident fibroblast population [ 175 ]. This raises the question of whether CFEVs can be reprogrammed to be more cardioprotective, an idea that, apparently, is supported by promising results of the use of haemopoietic stem cell- and induced pluripotent stem cell-derived fibroblasts for the repair of the myocardium after myocardial infarction [ 185 , 186 ].…”