The dynamics of changes in myogenic potential of stromal cells from alveolar oral mucosa in 2D and 3D culture

Abstract

The availability of autologic cell sources that would be able to differentiate towards the myogenic direction still remains one of the key issues of substitutive cell therapy for musculoskeletal system dysfunctions and muscular dystrophies. Cells obtained from alveolar oral mucosa are one of the most promising autologic cell sources due to their ontogenetic origin and the ability to repair most injuries of the oral cavity without scar formation. The aim of the present research was to compare the myogenic potential of alveolar mucosa stromal cells in 2D and 3D cell culture. Under the standard conditions of monolayer 2D culture, myogenic differentiation occurred in a differentiation medium (DMEM low glucose + 2% Horse serum) and spontaneously in 40% of cultures in a growth medium; we observed the formation of myotubes as well as the expression of MyoD, a key regulator of initial myogenesis and differentiation of myogenic progenitor cells. In a 3D culture, stromal cells from alveolar oral mucosa formed compact spheroid structures. In 7 days, we observed spontaneous differentiation of cells in spheroids towards the myogenic direction and emergence of not single myotubes but more differentiated structures — myofibrils with characteristic nuclei arrangement and cross-striation determined by staining with sarcomeric a-actinin antibodies. Therefore, we showed that the 3D culture conditions stimulated effective, spontaneous myogenic differentiation of stromal cells from alveolar oral mucosa with the formation of myofibrils, which is characteristic of well-organized muscular tissue.