OPLA scaffold, collagen I, and horse serum induce an higher degree of myogenic differentiation of adult rat cardiac stem cells.

In the last few years, a major goal of cardiac research has been to drive stem cell differentiation to replace damaged myocardium. Several research groups have attempted to differentiate potential cardiac stem cells (CSCs) using bi- or three-dimensional systems supplemented with growth factors or molecules acting as differentiating substances. We hypothesize that these systems failed to induce a complete differentiation because they lacked an architectural space. In the present study, we isolated a pool of small proliferating and fibroblast-like cells from adult rat myocardium. The phenotype of these cells was assessed and the characterized cells were cultured in a collagen I/OPLA scaffold with horse serum to obtain fine myocardial differentiation. C-Kit(POS)/Sca-1(POS) CSCs fully differentiated in vitro when an environment more similar to the CSC niche was created. These experiments demonstrated an important model for the study of the biology of CSCs and the biochemical pathways that lead to myocardial differentiation. The results pave the way for a new surgical approach.

HSP90 and eNOS partially co-localize and change cellular localization in relation to different ECM components in 2D and 3D cultures of adult rat cardiomyocytes.

BACKGROUND INFORMATION: Cultivation techniques promoting three-dimensional organization of mammalian cells are of increasing interest, since they confer key functionalities of the native ECM (extracellular matrix) with a power for regenerative medicine applications. Since ECM compliance influences a number of cell functions, Matrigel-based gels have become attractive tools, because of the ease with which their mechanical properties can be controlled. In the present study, we took advantage of the chemical and mechanical tunability of commonly used cell culture substrates, and co-cultures to evaluate, on both two- and three-dimensional cultivated adult rat cardiomyocytes, the impact of ECM chemistry and mechanics on the cellular localization of two interacting signalling proteins: HSP90 (heat-shock protein of 90 kDa) and eNOS (endothelial nitric oxide synthase). RESULTS: Freshly isolated rat cardiomyocytes were cultured on fibronectin, Matrigel gel or laminin, or in co-culture with cardiac fibroblasts, and tested for both integrity and viability. As validation criteria, integrity of both plasma membrane and mitochondria was evaluated by transmission electron microscopy. Cell sensitivity to microenvironmental stimuli was monitored by immunofluorescence and confocal microscopy. We found that HSP90 and eNOS expression and localization are affected by changes in ECM composition. Elaboration of the images revealed, on Matrigel-cultured cardiomyocytes, areas of high co-localization between HSP90 and eNOS and co-localization coefficients, which indicated the highest correlation with respect to the other substrates. CONCLUSIONS: Our three-dimensional adult cardiomyocyte cultures are suitable for both analysing cell-ECM interactions at electron and confocal microscopy levels and monitoring micro-environment impact on cardiomyocyte phenotype.

Cardiac stem cell research: an elephant in the room?

Heart disease is the leading cause of death in the industrialized world, and stem cell therapy seems to be a promising treatment for injured cardiac tissue. To reach this goal, the scientific community needs to find a good source of stem cells that can be used to obtain new myocardium in a very period range of time. Since there are many ethical and technical problems with using embryonic stem cells as a source of cells with cardiogenic potential, many laboratories have attempted to isolate potential cardiac stem cells from several tissues. The best candidates seem to be cardiac "progenitor" and/or "stem" cells, which can be isolated from subendocardial biopsies from the same patient or from embryonic and/or fetal myocardium. Regardless of the technique used to isolate and characterize these cells, it appears that the different cells isolated from adult myocardium to date are all phenotypic variations of a unique cell type that expresses several markers, such as c-Kit, CD34, MDR-1, Sca-1, CD45, nestin, or Isl-1, in various combinations.