ABSTRACT
BACKGROUND: Bone marrow mesenchymal stem cells can be induced into myocardial tissue-like structure in vitro. Myocardial tissue lysates from different parts of the myocardium can be used to construct differential microenvironments. Few studies have investigated the targeted induction efficiency of bone marrow mesenchymal stem cells and the expression of related genes. OBJECTIVE: To investigate the effects of lysates from different parts of the myocardium on the differentiation of bone marrow mesenchymal stem cells into myocardial tissue-like structures and to assess the correlation between the expressions of related genes during induction and myocardial tissue engineering. METHODS: Passage 3 bone marrow mesenchymal stem cells from Sprague-Dawley rats were cultured. Whole heart tissue lysate, atrial muscle tissue lysate, and ventricular muscle tissue lysate were used to construct different microenvironments to induce bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells. The morphological changes and ultrastructure of cells were observed with an inverted phase contrast microscope and transmission electron microscopy. Expression of α-actin and cTnI was detected by immunofluorescence staining. qRT-PCR was used to detect the expression of upstream molecules ANP, HCN4 and downstream molecules MLC-2v in the cAMP/PKA signaling pathway after induction. RESULTS AND CONCLUSION: (1) Each induction cell group followed similar morphological changes: Cardiomyocyte-like cells were capable of autonomic beats, rhythmic contractions and relaxations; under the transmission electron microscope, there were a large number of arranged myofilaments; immunofluorescent staining showed positive expression of α-actin and cTnI. (2) The whole heart tissue lysate was able to induce the differentiation of bone marrow mesenchymal stem cells into rice grain-sized myocardial tissue-like structures with collagen fibers. (3) Atrial muscle tissue lysate could induce bone marrow mesenchymal stem cells to differentiate into atrial muscle-like cells. Autonomic beats appeared earlier, but the beat frequency and duration were shorter. ANP and HCN4 were highly expressed in atrial myocytes. (4) Ventricular muscle tissue lysate induced bone marrow mesenchymal stem cells to differentiate into ventricular muscle-like cells with no secretory granules, and MLC-2v was highly expressed in ventricular muscle-like cells. (5) To conclude, the whole heart tissue lysate can induce bone marrow mesenchymal stem cells to form myocardial tissue-like structures. Atrial muscle tissue lysate can induce bone marrow mesenchymal stem cells to differentiate into atrial muscle-like cells. Ventricular muscle tissue lysate can induce bone marrow mesenchymal stem cells to differentiate into ventricular muscle-like cells. By constructing a specific microenvironment, bone marrow mesenchymal stem cells can be differentiated to cardiomyocytes in different parts, providing laboratory data for the construction of myocardial tissue engineering.
ABSTRACT
BACKGROUND: Owing to the advantages of low sensitization and natural three-dimensional structure, good biocompatibility and cell affinity, acellular heart scaffold materials are of great current interest in cardiac tissue engineering. OBJECTIVE: To investigate the cytocompatibility of an acellular heart scaffold of neonatal rats. METHODS: In order to construct the seed cell-scaffold complex, passage 3 bone marrow mesenchymal stem cells (BMSCs) of Sprague-Dawley neonatal rats were cultured with an acellular heart scaffold of Sprague-Dawley neonatal rats for 7 and 14 days. Hematoxylin eosin staining and scanning electron microscopy were used to observe the growth of BMSCs in the scaffold. The cell-scaffold complex was induced in myocardial tissue lysate for 14 days. BMSCs with planar orientation differentiation for 14 and 20 days were used as control group. RT-PCR was used to detect the expression of myosin heavy chain α-MHC and zinc finger transcription factor GATA-4 in BMSCs. RESULTS AND CONCLUSION: (1) Hematoxylin-eosin staining showed the acellular heart scaffold contained a large number of eosinophilic fibrous structures, and the cell number of cell-scaffold complex after co-culture for 14 days was higher than that after co-culture for 7 days. Under the scanning electron microscope, a large amount of cells adhered to the fiber surface of the acellular scaffold at 14 days of co-culture. (2) BMSCs with planar orientation differentiation for 14 and 20 days had the bamboo-like and myotube-like structures. In the cell-scaffold complex with planar orientation differentiation for 14 days, the expression of α-MHC and GATA-4 could be detected, and their expression levels fulfilled the requirement for the presence of bamboo-like cells and myotube-like structure. These results indicate that the acellular heart scaffold exhibits good cytocompatibility.