1α,25-Dihydroxyvitamin D3 inhibits the differentiation and bone resorption by osteoclasts generated from Wistar rat bone marrow-derived macrophages

Abstract

Abstract. The steroid hormone 1α, 2 D 3 ] plays an important role in maintaining a balance in calcium and bone metabolism. To study the effects of 1α,25-(OH) 2 D 3 on osteoclast (OC) formation and bone resorption, OC differentiation was induced in bone marrow-derived mononuclear cells from Wistar rats with the addition of macrophage colony stimulating factor and receptor activator for nuclear factor-κB ligand in vitro. Cells were then treated with 1α,25-(OH) 2 D 3 at 10 -9 , 10 -8 or 10 -7 mol/l. OCs were identified using tartrate-resistant acid phosphatase staining and activity was monitored in the absorption lacunae by scanning electron microscopy. Expression levels of functional proteins associated with bone absorption, namely carbonic anhydrase II, cathepsin K and matrix metalloproteinase-9 were evaluated by western blot analysis. The results showed that 1α,25-(OH) 2 D 3 inhibited the formation and activation of OCs in a dose-dependent manner and downregulated the expression levels of bone absorption-associated proteins.

Copper-Modified Ti6Al4 V Suppresses Inflammatory Response and Osteoclastogenesis while Enhancing Extracellular Matrix Formation for Osteoporotic Bone Regeneration

“…In brief, bone marrow was washed from the femur with DMEM and cultured in 75 cm 2 flasks (Thermo Fisher Scientific Inc., Pittsburgh, USA) in a medium consisting of 10% fetal bovine serum (FBS) (HyClone, USA), 90% DMEM and 10 ng/mL of rat recombinant macrophage-colony stimulating factor (M-CSF) (Peprotech, Rocky Hill, NJ, USA). The bone marrow-derived macrophages were also used as osteoclast precursor cells and treated with 25 μg/L M-CSF and 45 μg/L receptor activator for nuclear factor-κ B ligand (RANKL) (Peprotech, Rocky Hill, NJ, USA) to induce osteoclasts, as described in the literature . Osteoblasts were isolated from the control and OVX groups, as described in the literature .…”

“…The bone marrow-derived macrophages were also used as osteoclast precursor cells and treated with 25 μg/L M-CSF and 45 μg/L receptor activator for nuclear factor-κ B ligand (RANKL) (Peprotech, Rocky Hill, NJ, USA) to induce osteoclasts, as described in the literature. 26 Osteoblasts were isolated from the control and OVX groups, as described in the literature. 27 In brief, the mandibles of osteoporotic rats were removed under sterile conditions and cut into bone slices.…”

Vitamin D Inhibition of TRPV5 Expression During Osteoclast Differentiation

“…Floating cells, which contained BMMs, were collected and resuspended at the density of rated amount per mL. The cells (plated at concentration of 2 × 10 5 cells/well and 6 × 10 5 cells/well in 24-well and 12-well plates respectively) were cultured with α-MEM supplement with 10% FBS and were treated by M-CSF (10 ng/mL) and RANKL (50 ng/mL) (Peprotech, USA)-induced form OC as previously reported (19). After OC formation, the cells were treated by different concentrations of ruthenium red (RuR, 500 nmol/L) and ECO (5 nmol/L) with or without 1,25(OH) 2 D 3 treatments., A well-known marker of OC, TRAP, was stained by leukocyte acid phosphatase kit (Sigma, USA).…”