Investigating the biological activities of mesenchymal stem ‎cells ‎cultured on skeletal muscle-derived coating substrate

Bone is the hardest and one of the most important tissues in the body. In case of bone damage, the current treatments do not completely repair and regenerate the bone. For this reason, cell-based tissue engineering strategies, especially Mesenchymal Stem Cells (MSCs), have received attention. MSCs have the ability to self-renew and differentiate into different cell types, including bone cells, cartilage cells, and fat cells, among others. They are found in various tissues throughout the body, including bone marrow, adipose tissue, and umbilical cord tissue. Today, MSCs are a valuable resource for regenerative medicine and tissue engineering applications. In addition to the cells, scaffolds are another essential element of tissue engineering. One of these scaffolds is decellularized tissue-derived hydrogels, which are three-dimensional network of hydrophilic polymer chains that can absorb and retain a significant amount of water. In tissue engineering, they mimic the natural extracellular matrix of tissues, providing a suitable environment for cells to attach, proliferate, and differentiate. In the current study, we aimed to investigate the effects of decellularized skeletal muscle-derived hydrogel, known as Myogel, on bone marrow-derived MSCs biological behaviors, including proliferation, viability and migration. In this study, MSCs were isolated from tibia and femur of adult Wistar rats. MSCs were cultured in a complete medium (a-MEM containing 15% fetal bovine serum (FBS) and 1% penicillin/streptomycin (Pen/Strep)). The identity of cells was determined by morphology (using inverted microscope) and expression of specific CD markers (using Flowcytometry). Skeletal muscle was decellularized and accuracy of decellularization was evaluated using special staining. Then Myogel was prepared from digested decellularized skeletal muscle. Here, Myogel substrate was used as the control group, gelatin substrate as the positive control, and un-coated plates as the negative control. The effect of Myogel on survival (MTT method), proliferation (drawing the growth curve and calculating the doubling time of the cell population), cell cycle profile (flow cytometry method), and cell migration (scratch method) were investigated. The MTT test showed that the survival of MSCs in Myogel substrate with a concentration of 0.2 mg/ml was higher than the survival of MSCs in gelatin substrate with a concentration of 0.1 mg/ml and the survival of MSCs in gelatin was higher than the survival of control MSCs. Myogel substrate increased the proliferation and migration of cells and decreased the doubling time of MSCs population. Examining the cell cycle profile showed that a high percentage of cells cultured on Myogel were in the G1 and S phase of the cell cycle, indicating an increase in cell division speed by gelatin and, in the next degree, by Myogel. Therefore, Myogel can be used as a suitable substrate to increase the proliferative and migratory potential of MSCs, which are an important factor in tissue engineering.