FOXO1 transcription factor regulates chondrogenic differentiation through transforming growth factor β1 signaling

Abstract

The forkhead box O (FOXO) proteins are transcription factors involved in the differentiation of many cell types. Type II collagen (Col2) Cre-Foxo1-knockout and Col2-Cre-Foxo1,3,4 triple-knockout mice exhibit growth plate malformation. Moreover, recent studies have reported that in some cells, the expressions and activities of FOXOs are promoted by transforming growth factor β1 (TGFβ1), a growth factor playing a key role in chondrogenic differentiation. Here, using a murine chondrogenic cell line (ATDC5), mouse embryos, and human mesenchymal stem cells, we report the mechanisms by which FOXOs affect chondrogenic differentiation. FOXO1 expression increased along with chondrogenic differentiation, and FOXO1 inhibition suppressed chondrogenic differentiation. TGFβ1/SMAD signaling promoted expression and activity of FOXO1. In ATDC5, FOXO1 knockdown suppressed expression of sex-determining region Y box 9 (Sox9), a master regulator of chondrogenic differentiation, resulting in decreased collagen type II α1 (Col2a1) and aggrecan (Acan) expression after TGFβ1 treatment. On the other hand, chemical FOXO1 inhibition suppressed Col2a1 and Acan expression without suppressing Sox9. To investigate the effects of FOXO1 on chondrogenic differentiation independently of SOX9, we examined FOXO1's effects on the cell cycle. FOXO1 inhibition suppressed expression of p21 and cell-cycle arrest in G0/G1 phase. Conversely, FOXO1 overexpression promoted expression of p21 and cell-cycle arrest. FOXO1 inhibition suppressed expression of nascent p21 RNA by TGFβ1, and FOXO1 bound the p21 promoter. p21 inhibition suppressed expression of Col2a1 and Acan during chondrogenic differentiation. These results suggest that FOXO1 is necessary for not only SOX9 expression, but also cell-cycle arrest during chondrogenic differentiation via TGFβ1 signaling.

Ethanol Inhibits Mesenchymal Stem Cell Osteochondral Lineage Differentiation Due in Part to an Activation of Forkhead Box Protein O‐Specific Signaling

“…In our experiments investigating whether FoxO1/3 inhibitor AS1842856 would restore SOX9 levels in the presence of EtOH, we found 1 μ M AS1842856 completely inhibited chondrogenic differentiation. Indeed, a recent study showed that FoxO1 is required for SOX9 expression (Kurakazu et al, 2019). We found that a 10‐fold decrease in AS1842856 permitted chondrogenic differentiation, but did not fully restore SOX9 expression in EtOH‐exposed MSCs.…”

“…More likely, FoxO1/3 activity may be required, in at least at some basal level of activity, for osteogenic and chondrogenic differentiation. Indeed, several studies show that FoxO1/3, as well as endogenous ROS, is required for both osteogenic and chondrogenic differentiation and normal bone turnover (Alund et al, 2016; Kurakazu et al, 2019; Teixeira et al, 2010). Presently, our findings suggest the overexpression of FoxO1/3 is responsible for the inhibition of osteogenesis and chondrogenesis in EtOH‐exposed MSCs.…”

Direct reprogramming of fibroblasts into diverse lineage cells by DNA demethylation followed by differentiating cultures

“…3A and B ). Finally, in chondrogenic media, BMP2 induced chondrogenic differentiation of 5-aza-CN-treated NIH-3T3 fibroblasts, which significantly increased the expression of chondrogenic markers such as aggrecan (ACAN) and type II collagen (COL2) which are major matrix proteins of cartilaginous tissue, and SRY-box 9 (SOX9) which is essential transcription factor for differentiation of precursors cells into chondrocyte [ 25 ], as well as proteoglycan stained by Alcian blue was significantly increased ( Fig. 3C and D ).…”

Potential and recent advances of microcarriers in repairing cartilage defects

“…Some cytokines, such as TGF-β1, can induce MSC chondrogenic differentiation [ 58 ]. Research by Agata Zykwinska confirms that microcarriers based on marine exopolysaccharide (EPS) can encapsulate TGF-β1, which has potential in future applications for repairing cartilage defects [ 59 ].…”