Combination of Cardiac Progenitor Cells from the Right Atrium and Left Ventricle provide Synergistic Paracrine Effects for In Vitro Cardiac Repair

Abstract

Cardiovascular diseases, such as ischaemic heart disease, remain the most common cause of death worldwide. With no effective treatment to replace the loss of cardiac tissue following a myocardial infarction, ischaemic heart disease places a significant socioeconomic burden on medicine. Advances in medical research have introduced new potential treatments, of which regenerative medicine such as stem cell therapy seem to be promising for cardiac repair. Various stem cell types have been investigated, which have different effects in cardiac repair. Combination of different cell types have been shown to improve the therapeutic potential which is thought to be due to synergistic or complimentary reparative effects. Recently, cardiac progenitor cells (CPC) isolated from the right atrial appendage (RAA) and left ventricle have been shown to have different functional effects in vitro. Importantly, conditioned media (CM) from CPCs from the right atrial appendage (RAA CPC) have superior cardioprotective effects, while CM from the left ventricle (LV CPC) have superior angiogenic effects. Due to these distinct functional paracrine effects, this study investigated if the combination of RAA CPCs and LV CPCs from the same patient exert synergistic or complimentary paracrine effects for cardioprotection and angiogenesis in an in vitro model. Both RAA CPCs and LV CPCs expressed the mesenchymal cell markers CD90 and CD105, and were predominantly negative for the haematopoietic cell marker, CD34. CPCs were cultured either alone or in combination and exposed to serum deprivation and hypoxic conditions (1% O2) to stimulate ischaemia. Normoxic conditions (20% O2) was used as a control. Gene expression of HIF1A, AKT1, FGF2 and PDGFA were measured to investigate the cellular responses to stimulated ischaemia. HIF1A mRNA expression was significantly decreased in the combination group of RAA CPCs and LV CPCs (RAA + LV) cultured in hypoxia and in hypoxia irrespective of groups. There were no significant differences in the expression of AKT1, FGF2 and PDGFA. To measure the paracrine effects of CPCs, CM was collected following stimulated ischaemia. IGF-1 (pro-survival factor) and VEGF-A (pro-angiogenic factor) concentrations in CM were measured. IGF-1 concentration was comparable across all CPC groups. However, VEGF-A concentration was increased in CM collected from hypoxia, especially in the RAA and RAA + LV hypoxic CM. CM from all CPC groups reduced apoptosis in AC16 cardiomyocyte cell line exposed to stimulated ischaemia. Interestingly, RAA + LV normoxic CM was the only group that significantly increased the expression of HIF-1α protein in AC16 cardiomyocytes exposed to stimulated ischaemia. CM from all CPC groups induced angiogenesis in human umbilical vein endothelial cells and was the highest in the RAA + LV hypoxic CM. Overall, this study provides evidence that combination of RAA CPCs and LV CPCs may have greater therapeutic effects and have synergistic paracrine effects for cardiac repair. Therefore, in vivo studies are warranted to determine if combination of different stem cell types have greater therapeutic potential than single cell therapies.