Effects of 1,25-(OH)_2-vitamin D_3 on proliferation, differentiation, and secretion of osteoprotegerin and RANKL in cultured marrow mesenchymal cells of rhesus monkey / 中华内分泌代谢杂志

ABSTRACT

Objective To study the effects of 1,25-(OH)_2-vitamin D_3[1,25-(OH)_2D_3] on proliferation,differentiation, and secretion of osteoprotegerin and RANK ligand (RANKL)in cultured marrow mesenchymal cells of rhesus monkey (RhBMSCs). Methods Three different concentrations of 1,25-(OH)_2D_3 (10~(-12) ,10~(-10), and 10~(-8)mol/L)were added to the cultured RhBMSCs in vitro. MTT was used to observe cell proliferation and ELISA technique was used to measure the concentration of alkaline phosphatase (ALP), osteocalcin, osteoprotegerin, and RANKL. Mineralization nodus was identified via alizarin red staining. Results Under different concentration of 1,25-(OH)_2D_3, RhBMSCs proliferation were promoted within 7 days but were suppressed beyond 7 days. No significant dose-dependent manner was found. Differentiation of RhBMSCs into osteoblast was promoted by 1,25-(OH)_2D_3. The levels of ALP and osteocalcin in groups with various concentrations of 1,25-(OH)_2D_3 were higher than those of control group [ALP(ng/ml) ,7 d31.40±1.25,26.50±0.50,28.47± 0.25 vs 13.48±0.26;10 d33.37±0.68,35.30±1.57,33.27±0.67 vs 17.14±0.55;13 d35.37±0.12,30.47± 0. 25 , 30. 27±1.25 vs 16.55 ± 1.13 ; osteocalcin (ng/ml), 7 d4.47±0. 29,4.00 ±0. 60,3.73±0.78 vs 1.63± 0.55;10 d5.63±0.57,5.17±0.15,4.30±0. 10 vs 2.17±0. 15;13 d7.03±0.15,5.53±0.25,5.27±0.31 vs 2.23±0. 55 ; all P < 0. 05], but no typical mineralization nodus was found in either group. Secretions of osteoprotegerin and RANKL from RhBMSCs were stimulated by 1,25-(OH)_2D_3 [osteoprotegerin (pg/ml), 7 d 72.57±0.67,68.00±1.75,64.23±0.87 vs 30. 13±1.72; 10 d62.03±1.62,51.80±1.30,28.93±0.95 vs 18.13±1.40;13 d65.13±0.71,62.43±2.11,44.93+1.63 vs 36.70±0.95 ;RANKL(pg/ml) ,7 d74.33+0.61, 82.37±2.15,85.23±0.45 vs 70.83±1.71 ;10 d83.30±0.46,86.70±0.56,88.23±0.91 vs 74.20±1.83;13 d 81.70±1.81,81.07±0.95,84.70±1.41 vs 72.73±0.97 ;all P<0.05]. The secretion of RANKL was increased at first and then decreased, whereas the secretion of osteoprotegerin had the opposite tendency. The secretions of RANKL and osteoprotegerin were both in a dose-dependent manner. Conclusions 1,25-(OH)_2D_3 promotes differentiation of RhBMSCs into osteoblasts,resulting in increased secretions of osteoprotegerin and RANKL