All trans retinoic acid inhibits osteogenic differentiation of rat bone marrow mesenchymal stem cells

Abstract:

Objective：To explore the effects of all trans retinoic acid（at-RA） on rat bone marrow mesenchymal stem cells（rBMSCs） osteogenic differentiation and mechanisms. Methods：The rBMSCs were isolated and induced to differentiate into osteoblast. At-RA at concentrations of 0.1，1.0，5.0 μmol/L was added to the osteoblast induced medium. Morphology of the cultured cells was observed us－ing inverted microscope respectively. On the 7th d，histochemical staining and alkaline phosphatase detection kit were used to test the alkaline phosphatase（ALP） activity. On the 21st d，alizarin red staining was employed to detect calcium salt deposition. During osteo－genetic induction，mRNA expressions of both osteocalcin（OCN） and osteopontin（OPN），biomarkers of osteoblast and related differen－tiation regulatory genes，i.e. Runx2，osterix of the osteogenesis regulation pathways and retinoic acid receptors（RARs，?琢，?茁） of the retinoic acid signaling pathways were measured using real-time PCR. Results：On the 7th d of differentiation，the activity of ALP of os－teogenic induced rBMSCs with at-RA cells increased markedly compared with that without at-RA cells（F=107.177，P=0.000）. How－ever，on the 21st d，the calcium salt deposition of at-RA treated rBMSCs significantly reduced compared with that of non-treated rBMSCs（F=57.554，P=0.000）. During osteogenic induction process，OCN，OPN，Runx2，Osterix and RAR?琢 expressions of at-RA treated（5.0 μmol/L） rBMSCs decreased compared with those of non-treated rBMSCs（211.145，0.000；30.835，0.005；106.536，0.000；452.546，0.000；253.159，0.000，respectively for F and P values） on the 21st d. Meanwhile，significantly increased RARβ expression was observed（F=2 294.377，P=0.000） during the whole differentiation process. Conclusion：at-RA can inhibit osteogenic differentiation of rBMSCs. The mechanism may be through up-regulating the expression of RARβ，down-regulating the expression of RARα，and inhibiting the expressions of Runx2 and Osterix.