Gu Jianhong, Tong Xi-Shuai, Chen Guo-Hong, Wang Dong, Chen Yang, Yuan Yan, Liu Xue-Zhong, Bian Jian-Chun, Liu Zong-Ping
College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China; College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009, China.
College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009, China.
J Steroid Biochem Mol Biol. 2015 Aug;152:25-33. doi: 10.1016/j.jsbmb.2015.04.003. Epub 2015 Apr 10.
The hormonally active form of vitamin D3, 1α,25-(OH)2D3, has an important role in bone metabolism. This study examined the effects of 1α,25-(OH)2D3 on the ability of two cytokines, receptor activator of nuclear factor-κB ligand (RANKL) and macrophage-colony stimulating factor (M-CSF), to induce RAW 264.7 cells to form osteoclasts. A TRAP histochemical staining assay and bone resorption analysis were used to identify the rate of formation and activity of osteoclasts. The numbers of osteoclasts formed, and their bone resorption activity, was enhanced by the addition of 1α,25-(OH)2D3. The expression levels of osteoclast-specific proteins that are essential for bone resorption, integrin β3, V-ATPase, CAII, CTSK, TRAP and MMP-9, were detected by western blotting. During 48 h, the expression levels of all these proteins significantly increased. Quantitative real-time polymerase chain reaction was used to determine the expression levels of the transcription factors, c-Fos and NFATcl. The expression levels of c-Fos and NFATc1 also increased 24h after treatment with 1α,25-(OH)2D3. These results suggest that 1α,25-(OH)2D3 can regulate bone metabolism by directly enhancing the formation and maturation of osteoclasts.
维生素D3的激素活性形式1α,25 - (OH)2D3在骨代谢中起重要作用。本研究检测了1α,25 - (OH)2D3对两种细胞因子——核因子κB受体激活剂配体（RANKL）和巨噬细胞集落刺激因子（M - CSF）诱导RAW 264.7细胞形成破骨细胞能力的影响。采用抗酒石酸酸性磷酸酶（TRAP）组织化学染色法和骨吸收分析来鉴定破骨细胞的形成速率和活性。添加1α,25 - (OH)2D3可增强破骨细胞的形成数量及其骨吸收活性。通过蛋白质免疫印迹法检测骨吸收所必需的破骨细胞特异性蛋白——整合素β3、V - ATP酶、碳酸酐酶II（CAII）、组织蛋白酶K（CTSK）、抗酒石酸酸性磷酸酶（TRAP）和基质金属蛋白酶9（MMP - 9）的表达水平。在48小时内，所有这些蛋白的表达水平均显著增加。采用定量实时聚合酶链反应来测定转录因子c - Fos和活化T细胞核因子c1（NFATc1）的表达水平。用1α,25 - (OH)2D3处理24小时后，c - Fos和NFATc1的表达水平也升高。这些结果表明，1α,25 - (OH)2D3可通过直接增强破骨细胞的形成和成熟来调节骨代谢。
