Extended characterization of hypoimmunogenic mesenchymal stem cells isolated from the subendocardial layer of human hearts from chronic heart failure patients

Abstract

Purpose: Mesenchymal stem cells (MSC) are pluripotent cells which are known to reside in a number of adult organs. In human heart, various populations of stem/progenitor cells have been isolated by different groups. Several efforts still need to be made to better characterize resident or migrating MSC populations in terms of markers expression and immunogenic potential prior to their use for regenerative medicine applications in heart diseases. Methods: MSC were isolated from the sub-endocardial layer of left ventricle sections of hearts explanted from patients affected by chronic heart failure (CHF). The expression of several markers characteristic of the MSC lineage was assessed at both the protein and RNA level by western blotting, immunocytochemistry and RT-PCR. The differentiative ability was confirmed by exposing cells to differentiation media towards the adicpocyte and the osteoblast lineage. Self-Renewal of cells was evaluated by clonogenicity experiments by the limiting dilution method. Results: MSC from human hearts of patients affected by CHF were successfully extracted and expanded in vitro. These cells spontaneously formed “cardiospheres” with abundant extracellular matrix, and showed for the first time the expression of a number of new and predictive markers: in fact, these cells expressed key markers of the embryonic stem cell phenotype (Oct3/4 and Nanog transcription factors), together with cardio-specific GATAs (-4 and -6). MSC were further positive for the expression of a panel of transcription factors involved in heart development (Nkx2.5, Mef2C, Myocardin, ISL-1), but lacked, when kept undifferentiated, the expression of mature myocyte markers (as MYL-2). Moreover, we assessed for the first time in heart MSC the expression of a favourable panel of markers (CD40-, CD80+, CD86-, HLA-A+, HLA-DR-) which suggests hypoimmunogenic properties for these cells, potentially important in preventing adverse host immune processes after transplantation. Conclusions: The diseased human heart is still a source of MSC populations, even following the development of chronic pathologies as CHF. These cells are capable of self-renewal, express several stem cell markers, showed the expression of promising markers of the cardiac differentiation program, and have a hypoimmunogenic profile of surface markers. Further experiments are underway to differentiate these cells in vitro towards the cardiomyocyte lineage, and to better characterize the expression of other factors which should favour a successful in vivo engraftment of cells preventing host immune rejection processes.