Tumor Necrosis Factor-Alpha Induces Proangiogenic Profiling of Cardiosphere-Derived Cell Secretome and Increases Its Ability to Stimulate Angiogenic Properties of Endothelial Cells.

Ischemic heart disease and its complications, such as myocardial infarction and heart failure, are the leading causes of death in modern society. The adult heart innately lacks the capacity to regenerate the damaged myocardium after ischemic injury. Multiple lines of evidence indicated that stem-cell-based transplantation is one of the most promising treatments for damaged myocardial tissue. Different kinds of stem cells have their advantages for treating ischemic heart disease. One facet of their mechanism is the paracrine effect of the transplanted cells. Particularly promising are stem cells derived from cardiac tissue per se, referred to as cardiosphere-derived cells (CDCs), whose therapeutic effect is mediated by the paracrine mechanism through secretion of multiple bioactive molecules providing immunomodulatory, angiogenic, anti-fibrotic, and anti-inflammatory effects. Although secretome-based therapies are increasingly being used to treat various cardiac pathologies, many obstacles remain because of population heterogeneity, insufficient understanding of potential modulating compounds, and the principles of secretome regulation, which greatly limit the feasibility of this technology. In addition, components of the inflammatory microenvironment in ischemic myocardium may influence the secretome content of transplanted CDCs, thus altering the efficacy of cell therapy. In this work, we studied how Tumor necrosis factor alpha (TNFa), as a key component of the pro-inflammatory microenvironment in damaged myocardium from ischemic injury and heart failure, may affect the secretome content of CDCs and their angiogenic properties. We have shown for the first time that TNFa may act as a promising compound modulating the CDC secretome, which induces its profiling to enhance proangiogenic effects on endothelial cells. These results allow us to elucidate the underlying mechanisms of the impact of the inflammatory microenvironment on transplanted CDCs and may contribute to the optimization of CDC efficiency and the development of the technology for producing the CDC secretome with enhanced proangiogenic properties for cell-free therapy.

Urokinase-Type Plasminogen Activator Receptor Regulates Prosurvival and Angiogenic Properties of Cardiac Mesenchymal Stromal Cells.

One of the largest challenges to the implementation of cardiac cell therapy is identifying selective reparative targets to enhance stem/progenitor cell therapeutic efficacy. In this work, we hypothesized that such a target could be an urokinase-type plasminogen activator receptor (uPAR)-a glycosyl-phosphatidyl-inositol-anchored membrane protein, interacting with urokinase. uPAR is able to form complexes with various transmembrane proteins such as integrins, activating intracellular signaling pathway and thus regulating multiple cell functions. We focused on studying the CD117+ population of cardiac mesenchymal progenitor cells (MPCs), expressing uPAR on their surface. It was found that the number of CD117+ MPCs in the heart of the uPAR-/- mice is lower, as well as their ability to proliferate in vitro compared with cells from wild-type animals. Knockdown of uPAR in CD117+ MPCs of wild-type animals was accompanied by a decrease in survival rate and Akt signaling pathway activity and by an increase in the level of caspase activity in these cells. That suggests the role of uPAR in supporting cell survival. After intramyocardial transplantation of uPAR(-) MPCs, reduced cell retention and angiogenesis stimulation were observed in mice with myocardial infarction model compared to uPAR(+) cells transplantation. Taken together, the present results appear to prove a novel mechanism of uPAR action in maintaining the survival and angiogenic properties of CD117+ MPCs. These results emphasize the importance of the uPAR as a potential pharmacological target for the regulation of reparative properties of myocardial mesenchymal progenitor cells.

New Insight on 2D In Vitro Angiogenesis Models: All That Stretches Is Not a Tube.

A Matrigel-based tube formation assay is a simple and widely accepted 2D angiogenesis model in vitro. Extracellular matrix (EM) proteins and growth factors (GFs) from MatrigelTM exclusively trigger endothelial cell (EC) tubular network (ETN) formation. Co-culture of ECs with mesenchymal stromal cells (MSCs) is another and more reliable in vitro angiogenesis assay. MSCs modulate ETN formation through intercellular interactions and as a supplier of EM and GFs. The aim of the present study was to compare the expression profile of ECs in both models. We revealed upregulation of the uPA, uPAR, Jagged1, and Notch2 genes in dividing/migrating ECs and for ECs in both experimental models at 19 h. The expression of endothelial-mesenchymal transition genes largely increased in co-cultured ECs whereas Notch and Hippo signaling pathway genes were upregulated in ECs on MatrigelTM. We showed that in the co-culture model, basement membrane (BM) deposition is limited only to cell-to-cell contacts in contrast to MatrigelTM, which represents by itself fully pre-assembled BM matrix. We suggest that ETN in a co-culture model is still in a dynamic process due to immature BM whereas ECs in the MatrigelTM assay seem to be at the final stage of ETN formation.