Induction of chondrogenic differentiation of mouse embryonic mesenchymal stem cells through an in vitro pellet model

Abstract This study employed transforming growth factor beta 3 (TGF-β3) induction of the C3H10T1/2 mesenchymal stem cell (MSC) line to construct an in vitro chondrogenic differentiation model for MSCs. A C3H10T1/2 MSC cell line was cultured, amplified, and the seventh generation of cells was centrifuged to construct pellets, which were divided into a non-induced group and an induced group (treated with TGF-β3, vitamin C, dexamethasone, and ITS). Specimens were taken after 7, 14, 21, and 28 days under non-induced and induced culture to compare these two groups by Alcian Blue staining, collagen type II immunohistochemical staining, and transmission election microscopy (TEM) on days 21 and 42. Cell pellets in the non-induced group were smaller than those in the induced group on days 7, 14, 21, and 28 with the pellet morphology of the induced group being more regular and nearly spherical. Alcian blue staining in the induced group was consistently stronger than that in non-induced group across all time points, and type II collagen immunohistochemical IOD values were significantly higher in the induced group over the non-induced group across all time points. On days 21 and 42, TEM revealed that the induced group displayed greater karyokinesis and a higher euchromatin ratio compared to the non-induced group. This specially constructed pellet model treated with TGF-β3-containing chondrogenic medium can effectively promote chondrogenic differentiation of C3H10T1/2 MSC cells in vitro. This in vitro pellet model should be of value in providing a preliminary cell model reference for further studies of the mechanism of chondroblast differentiation of stem cells.