Effect of complete abroagation of hyaluronan synthesis on osteogenic differentiation of mesenchymal stem cells (MSc) in vitro

Mesenchymal stem cells (MSc) show diminished osteogenic differentiation upon complete knockout of all known Hyaluronan (HA) synthases (Has). We demonstrated that Has-knockout completely abrogated the release of HA into the culture medium by MSc and that these MSc showed an impaired osteogenic differentiation detected by Alizarin red staining at days 7 and 12 of differentiation compared to wildtype MSc. To gain insight intor the transcriptomic changes due to Has knockout we compared differentiating MSc at day 7 of differentiation. MSc from mouse femurs and tibiae were cultured in vitro and treated twice with 4-hydroxytamoxifen in order to induce cre recombination for Has2-knockout. Two groups were compared: MSc from wildtype mice (C57Bl6/J) and MSc from Has-total knockout mice (constitutive knockout of Has1 and Has3 plus induced knockout of Has2). MSc were isolated from 8 mice (n=4 per genotype) and the RNA from MSc at day 7 of osteogenic differentiation was analyzed.