Equol reverses the inhibition of cyclosporin A on the proliferation and osteoblastic differentiation of bone marrow mesenchymal stem cells through estrogen receptor/nitric oxide/cyclic guanosine monophosphate signal pathway / 中国组织工程研究

ABSTRACT

BACKGROUND: Long term taking cyclosporin A(CsA)can inhibit osteoblastic differentiation and induce osteoporosis.Equol,which has greater binding affinity to estrogen receptors,can stimulate the proliferation and osteoblastic differentiation.OBJECTIVE: To investigate whether equol may protect against the proliferation and osteoblastic differentiation inhibited by CsA in mouse bone marrow mesenchymal stem cells(BMSCs)cultures and analyze signal pathway of protection.METHODS: Primary mouse BMSCs were cultured by using attachment method and assigned to five groups,which respectively treated with equol or/and CsA in the presence or absence of ICI182780,an estrogen receptor antagonist,and Nω-nitro-L-arginine methyl ester.Under an inverted microscope,morphological changes and mineralization ability of BMSCs were observed.The cell proliferation was measured by[3H]-thymidine incorporation.The osteoblastic differentiation and mineralization of extracellular matrix in BMSCs was assessed by measuring alkaline phosphatase activity and calcium deposition,respectively.Nitric oxide production in the conditioned media and cyclic guanosine monophosphata(cGMP)content in BMSCs were determined by using commercial nitric oxide and cGMP kit,respectively.RESULTS AND CONCLUSION: Equol reversed the decreased[3H]thymidine incorporation(P < 0.05),alkaline phosphatase activity(P < 0.05)and calcium deposition(P < 0.01)of CsA,which was accompanied with the changes of nitric oxide production(P < 0.01)and cGMP content(P < 0.01).The group by co-treatment with equol and CsA possessed higher cells growth density and small mineralized nodes than CsA group on day 12 under an inverted microscope.Moreover,the equol-reversed effect was abolished by ICI182780 and Nω-nitro-L-arginine methyl ester.These indicated that equol can reverse the inhibition of CsA on the proliferation and osteoblastic differentiation of mouse BMSCs through estrogen receptor/nitric oxide/cGMP signal pathway.