Characterization of the in vitro growth and differentiation capabilities of human adipose-derived mesenchymal progenitor cells
Skritakis, Pantos Angelo
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BACKGROUND: Human mesenchymal progenitor cells are multipotent cells that can be harvested from various adult and fetal tissues. They exhibit the potential to differentiate into several cell lineages, most notably osteogenic, chondrogenic, and adipogenic lineages. Conditions such as osteoporosis, metabolic disease, and arthritis are examples of dysfunction of tissues formed by the mesenchyme. The inability of these conditions to be healed by the body’s own mechanisms has raised considerable interest in the potential of using mesenchymal progenitor cells as a therapeutic intervention. This concept opens the possibility of harvesting mesenchymal progenitor cells from an individual, growing them into the desired tissue, and implanting them back into the individual. Treatment of this nature is much less invasive than current methods, overcomes rejection by the immune system, and could potentially demonstrate better outcomes in individuals suffering from degenerative disease of the mesenchyme. AIMS/OBJECTIVES: The aims of this study were to determine and to characterize the differentiation of human adipose-derived mesenchymal progenitor cells into osteocytes, chondrocytes, and adipocytes. The differentiation capacity of the mesenchymal progenitor cells was evaluated through cell staining, immunofluorescence, and RNA sequencing. METHODS: Subcutaneous adipose tissue was collected from patients undergoing elective panniculectomies. The abdominal panniculus was liposuctioned, and small explants of fat were embedded in Matrigel. Mesenchymal progenitor cells were extracted from the explants and plated for differentiation into osteogenic, chondrogenic, and adipogenic lineages. Control cells were grown in parallel in basal media to confirm differentiation. Dye staining for differentiation was performed with Alizarin Red S, Alcian Blue, and Oil Red O, and immunofluorescence staining was performed to indicate lineage-specific markers for differentiation. RNA sequencing was also completed on the different cell lineages. RESULTS: Human adipose-derived mesenchymal progenitor cells displayed the capacity to differentiate into osteogenic, chondrogenic, and adipogenic lineages as evidenced by dye staining. Osteogenic differentiation was confirmed with Alizarin Red S staining of calcium deposits in the differentiated cells, whereas staining in the control resulted in no calcium deposits. Alcian Blue staining confirmed chondrogenic differentiation as glycoproteins secreted by the differentiated cells were evident and different in morphology compared with the control cells. Oil Red O staining indicated adipogenic differentiation by showing lipid droplets in the differentiated cells and no lipid droplets in the control. RNA sequencing provided support that lineage differentiation was successful. Immunofluorescence staining further proved that differentiated cells expressed lineage-specific proteins and demonstrated morphological differences. CONCLUSIONS: This study demonstrates that mesenchymal progenitor cells harvested from human adipose tissue have the potential to differentiate into adipogenic, chondrogenic, and osteogenic cell lineages when induced with differentiation media. The differentiation of these cells can be assessed with dye staining, RNA sequencing, and immunofluorescence staining methods. Further studies should be done to investigate the potential of mesenchymal progenitor cells for therapeutic interventions in the treatment of various illnesses related to the mesenchyme.