title: 24R,25-dihydroxyvitamin D3 promotes the osteoblastic differentiation of human mesenchymal stem cells.
abstract: Although 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3] is considered the most biologically active vitamin D3 metabolite, the vitamin D3 prohormone, 25-hydroxyvitamin D3 [25(OH)D3], is metabolized into other forms, including 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3]. Herein we show that 24R,25(OH)2D3 is fundamental for osteoblastic differentiation of human mesenchymal stem cells (hMSCs). Our approach involved analyses of cell proliferation, alkaline phosphatase activity, and pro-osteogenic genes (collagen 1A1, osteocalcin, vitamin D receptor [VDR], vitamin D3-hydroxylating enzymes [cytochrome P450 hydroxylases: CYP2R1, CYP27A1, CYP27B1 and CYP24A1]) and assessment of Ca(2+) mineralization of extracellular matrix. 24R,25(OH)2D3 inhibited hMSC proliferation, decreased 1α-hydroxylase (CYP27B) expression, thereby reducing the ability of hMSCs to convert 25(OH)D3 to 1α,25(OH)2D3, and promoted osteoblastic differentiation through increased alkaline phosphatase activity and Ca(2+) mineralization. 24R,25(OH)2D3 decreased expression of the 1α,25(OH)2D3 receptor, VDR. 24R,25(OH)2D3 but not 1α,25(OH)2D3 induced Ca(2+) mineralization dependent on the absence of the glucocorticoid analog, dexamethasone. To elucidate the mechanism(s) for dexamethasone-independent 1α,25(OH)2D3 inhibition/24R,25(OH)2D3 induction of Ca(2+) mineralization, we demonstrated that 1α,25(OH)2D3 increased whereas 24R,25(OH)2D3 decreased reactive oxygen species (ROS) production. 25(OH)D3 also decreased ROS production, potentially by conversion to 24R,25(OH)2D3. Upon inhibition of the vitamin D3-metabolizing enzymes (cytochrome P450s), 25(OH)D3 increased ROS production, potentially due to its known (low) affinity for VDR. We hypothesize that vitamin D3 actions on osteoblastic differentiation involve a regulatory relationship between 24R,25(OH)2D3 and 1α,25(OH)2D3. These results implicate 24R,25(OH)2D3 as a key player during hMSC maturation and bone development and support the concept that 24R,25(OH)2D3 has a bioactive role in the vitamin D3 endocrine system.
authors.affiliation: Geriatric Research, Education, and Clinical Center and Research Service (K.M.C., K.K.A., B.A.R., G.A.H.), Bruce W. Carter Veterans Affairs Medical Center, Miami, Florida 33125; and Departments of Biochemistry and Molecular Biology (K.M.C., G.A.H.), Medicine (B.A.R., G.A.H.), and Neurology (B.A.R.), University of Miami Miller School of Medicine, Miami, Florida 33101.
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Curtis
authors.forename: Kristina K
authors.initials: KK
authors.lastname: Aenlle
authors.forename: Bernard A
authors.initials: BA
authors.lastname: Roos
authors.forename: Guy A
authors.initials: GA
authors.lastname: Howard
mesh_terms: D015650:24,25-Dihydroxyvitamin D 3 / Q000494:pharmacology / Q000502:physiology*; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000378:metabolism; D002113:Calcification, Physiologic; D002117:Calcitriol / Q000494:pharmacology / Q000502:physiology; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D002648:Child; D003907:Dexamethasone / Q000494:pharmacology; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000187:drug effects / Q000502:physiology*; D010006:Osteoblasts / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D055815:Young Adult
chemical_list: D005938:Glucocorticoids; D017382:Reactive Oxygen Species; D015650:24,25-Dihydroxyvitamin D 3; D003907:Dexamethasone; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D002117:Calcitriol