Paracrine activity of cardiac progenitor cells is exerted through the release of exosomes that affect both cellular and extracellular compartment

Exosomes are nano-vesicles released in body fluids and culture media by several types of cells, including stem cells. These small extracellular vesicles are loaded with a cargo of proteins, lipids, mRNA, miRNA, DNA and specific factors that they convey from one cell to another. Therefore, exosomes are involved in cellular intercommunication and signaling and can influence numerous biological mechanisms like cell death, proliferation and differentiation. We hypothesize that cardiac progenitor cells (CPCs) exert the paracrine effects that direct cardiac regeneration and remodeling by delivering exosomes to the injured myocardium. To test our hypothesis, we isolated CPCs (CPC-N) from the heart of donors, and CPCs (CPC-P) and fibroblasts (CF-P) from the heart of patients undergoing heart transplantation. First, we cultured CPC-N and collected the exosomes they released in the culture medium (Exo-CPC-N). Then, we characterized Exo-CPC-N cargo, in terms of mRNA and growth factors, and analyzed their effects on proliferation, migration and apoptosis of CPC-P as well as on extracellular matrix synthesis and deposition by CF-P, by administering Exo-CPC-N to CPC-P and CF-P in vitro. Real-Time PCR analysis revealed that Exo-CPC-N carry specific transcripts for proteins involved in cardiac (Nkx2.5, Mef2C, ACTC1, TBX, GATA6, ACTA2) and mesenchymal differentiation (TM4SF1, Sox9, CD90, CD105), while growth factor array showed the presence of specific factors as HGF, IGF, TGF that are known to enhance cell proliferation, growth and migration. When administrated to CPC-P, Exo-CPC-N not only increased CPC-P proliferation rate and resistance to apoptosis induced by oxidative stress, but also induced the differentiation towards cardiac myocytes as testified by the expression of a well-structured alpha-sarcomeric actin resulting at the immunofluorescence analysis. Additionally, Exo-CPC-N modulated in vitro the CF-P deposition of ECM proteins typical of the cardiac matrix, like collagen IV, fibronectin and laminin. Our study strengthens the hypothesis that exosomes are mediators of CPCs regenerative effects with the potential to empower the regenerative potential of resident cells through the spread of tailored molecules, and are, then, to be considered as a promising alternative to cardiac cell therapy.