The Mechanisms Of All Trans Retinoic Acid Inhibits Osteogenic Differentiation Of Rat Bone Marrow Mesenchymal Stem Cells

Objective To explore its effects of all trans retinoic acid (at-RA) onrat bone marrow mesenchymal stem cells (rBMSCs) osteogenicdifferentiation and mechanisms.Materials and Methods In vitro, bone marrow mesenchymal stemcells from3-4weeks SD rats were isolated and cultured. The fourth genera-tion cells were used to induce osteogenesis, and identification by alizarinred staining. At-RA at concentrations (0.1,0.5,1.0,5.0μmol/L) was addedto the osteogenetic differentiation media.rBMSCs were cultured under thefollowing conditions： blank control （no osteogenetic differentiationmedia），control（no at-RA）,and at-RA at0.1,0.5,1.0,5.0μmol/L.Morphology of the cultured cells was observed using inverted microscoperespectively. On day7, histochemical staining and alkaline phosphatasedetection kit was used to test the alkaline phosphatase (ALP) activity.During osteogenetic induction, mRNA expressions of both Runx2，Osterix，OPN and OCN and retinoic acid receptors (RARα, β) of the retinoic acid signaling pathways were measured using real-time quantitative polymerasechain reaction（real-time PCR）.Results （1）rBMSCs can be induced to differentiate into osteoblastin osteogenetic differentiation media.（2）On differentiation day7, theactivity of ALP of osteogenic induced rBMSCs increased markedly withadding at-RA compared with no at-RA addition cells (F=107.177，P=0.000，Pblank vs。0。5μmol/L at-RA=0.011，Pblank vs。1.0μmol/L at-RA=0.000，Pblank vs。5.0μmol/L at-RA=0.000）. On day21, however, the calcium salt deposition wassignificantly reduced for the at-RA treated rBMSCs compared with thenon-treated rBMSCs (F=57.554，P=0.000，Pblank vs。0。5μmol/L at-RA=0.025，Pblank vs。1.0μmol/L at-RA=0.002，Pblank vs。5.0μmol/L at-RA=0.000). During osteogenicinduction process, OCN, OPN, Runx2, Osterix and RARα, β expressionswere decreased for at-RA treated (5.0μmol/L) rBMSCs compared with thenon-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 day21.Meanwhile, significant increased RARβ expression was observed(F=2294.377, P=0.000) during the whole differentiation process.Conclusions (1) rBMSCs could be induced into osteoblast.(2)at-RA can inhibit osteogenic differentiation of rBMSCs.(3) The at-RAinhibits rBMSCs osteogenetic differentiation by inhibits the expression ofRunx2and Osterix of osteogenesis signaling pathways.(4) The mechanismmay be through up-regulating the expression of RARβ and down-regulating the expression of RARα.