Cardiomyocyte differentiation induced in cardiac progenitor cells by cardiac fibroblast-conditioned medium

Abstract

Our previous study showed that after being treated with 5-azacytidine, Nkx2.5(+) human cardiac progenitor cells (CPCs) derived from embryonic heart tubes could differentiate into cardiomyocytes. Although 5-azacytidine is a classical agent that induces myogenic differentiation in various types of cells, the drug is toxic and unspecific for myogenic differentiation. To investigate the possibility of inducing CPCs to differentiate into cardiomyocytes by a specific and non-toxic method, CPCs of passage 15 and mesenchymal stem cells (MSCs) were treated with cardiac ventricular fibroblast-conditioned medium (CVF-conditioned medium). Following this treatment, the Nkx2.5(+) CPCs underwent cardiomyogenic differentiation. Phase-contrast microscopy showed that the morphology of the treated CPCs gradually changed. Ultrastructural observation confirmed that the cells contained typical sarcomeres. The expression of cardiomyocyte-associated genes, such as alpha-cardiac actin, cardiac troponin T, and beta-myosin heavy chain (MHC), was increased in the CPCs that had undergone cardiomyogenic differentiation compared with untreated cells. In contrast, the MSCs did not exhibit changes in morphology or molecular expression after being treated with CVF-conditioned medium. The results indicated that Nkx2.5(+) CPCs treated with CVF-conditioned medium were capable of differentiating into a cardiac phenotype, whereas treated MSCs did not appear to undergo cardiomyogenic differentiation. Subsequently, following the addition of Dkk1 and the blocking of Wnt signaling pathway, CVF-conditioned medium-induced morphological changes and expression of cardiomyocyte-associated genes of Nkx2.5(+) CPCs were inhibited, which indicates that CVF-conditioned medium-induced cardiomyogenic differentiation of Nkx2.5(+) CPCs is associated with Wnt signaling pathway. In addition, we also found that the activation of Wnt signaling pathway was accompanied by higher expression of GATA-4 and the blocking of the pathway inhibited the expression of GATA-4 in CVF-conditioned medium-incubated Nkx2.5(+) CPCs. This finding suggests that Wnt signaling pathway may alter GATA-4 expression and activate the cardiogenic program in the regulation of differentiation. In conclusion, Nkx2.5(+) CPCs have enormous potential for cardiomyogenic differentiation and the CVF-conditioned medium specifically induces CPCs to differentiate into a cardiac phenotype. Wnt signaling pathway is involved in CVF-conditioned medium-induced cardiomyogenic differentiation of Nkx2.5(+) CPCs.

Myocyte-fibroblast communication in cardiac fibrosis and arrhythmias: Mechanisms and model systems

“…However, myocyte-fibroblast interactions also play a key role in humans. Evidence from human embryonic stem cell derived cardiomyocytes highlights that fibroblast interactions (via co-culture or conditioned media) are key drivers of human cardiomyocyte differentiation [125–128]. These interactions may also contribute to human cardiac disease settings, where defects in myocyte-fibroblast coupling may underlie cardiac fibrosis and arrhythmias, which are highly prevalent.…”

Cardiac Progenitor Cells and the Interplay with Their Microenvironment

“…Not only do fibroblasts maintain the supporting matrix of the CPC niche [ 115 ] (the importance of the ECM-cell interaction will be discussed later on in a dedicated paragraph), but they also might influence the differentiation potential of CPCs. It has recently been shown that fibroblast-conditioned medium can induce differentiation via the Wnt signaling pathway [ 136 ] and that fibroblasts produce angiogenic and antiangiogenic factors [ 115 ]. Fibroblasts originate mainly from the epicardium [ 137 ]; therefore, interactions between epicardium-derived cells (EPDCs) and CPCs need to be described.…”

Intramyocardial implantation of differentiated rat bone marrow mesenchymal stem cells enhanced by TGF-β1 improves cardiac function in heart failure rats

“…MSCs are multipotent progenitor cells which can easily be purified and amplified ( 3 , 4 ). Recent studies have shown that MSCs can differentiate into cardiomyocytes (CMCs) or cardiomyocyte-like cells (CLCs) in vivo and in vitro ( 5 , 6 ). 5-azacytidine is a classic inducer that enhances differentiation of MSCs into CMCs by random demethylation.…”