维生素D介导鸡BMSCs和BMMs共培养体系中破骨细胞的形成

Vitamin D Regulates OC Formation in the Co-culture System of Chicken BMSCs and BMMs

为探究1α, 25-(OH)2D3对鸡骨髓间充质干细胞(BMSCs)和骨髓单核巨噬细胞(BMMs)体外共培养体系中破骨细胞(OC)形成的影响,笔者对1α, 25-(OH)2D3处理后的细胞进行抗酒石酸酸性磷酸酶染色(TRAP)、骨吸收功能鉴定、OC标志基因mRNA和蛋白表达的检测。结果表明:1α, 25-(OH)2D3能够上调BMSCs中核因子κB受体活化因子配体(RANKL)的表达,下调骨保护素(OPG)的表达。1α, 25-(OH)2D3处理共培养细胞5 d,OC数目较对照组明显增多,骨吸收活性较对照组极显著增强,基质金属蛋白酶9(MMP-9)、Ⅱ型碳酸酐酶(CAⅡ)、组织蛋白酶K(CtsK)及TRAP等标志性蛋白的mRNA和蛋白表达极显著高于对照组(P<0.01),其中10-8 mol·L^-1 1α, 25-(OH)2D3效果最明显。结果表明,10-8 mol·L^-1 1α, 25-(OH)2D3能够介导鸡BMSCs和BMMs共培养体系中OC的形成,该OC具有骨吸收活性,为进一步研究OC在家禽骨骼及钙磷代谢紊乱性疾病中的作用机制奠定基础。

This study aimed to investigate the effect of 1α, 25-(OH)2D3 on the formation of osteoclast(OC) in the co-culture system of chicken bone marrow mesenchymal stem cells(BMSCs) and bone marrow mononuclear macrophage cells(BMMs). In the study, tartrate-resistant acid phosphatase(TRAP), bone absorption function identification, and OC marker mRNA and protein expression detection were performed on cells treated with 1α, 25-(OH)2D3. The results showed that, 1α, 25-(OH)2D3 up-regulated the expression of receptor activator for nuclear factor-κB ligand(RANKL) and down-regulated the expression of osteoprotegerin(OPG) of BMSCs. Compared to the control group, the number of OC was increased in the co-culture system at day 5 treated by 1α, 25-(OH)2D3. Meanwhile, the bone resorption activity, the mRNA and protein expression of matrix metalloproteinase 9(MMP-9), carbonic anhydrase Ⅱ(CAⅡ), cathepsin K and TRAP were significantly enhanced(P<0.01) by 1α, 25-(OH)2D3. Among them, 10-8 mol· L^-11α, 25-(OH)2D3 had the most obvious effect. The results confirmed that 10-8 mol·L^-1 1α, 25-(OH)2D3 could mediate the formation of OC, which showed bone resorption activity, in the co-culture system of BMSCs and BMMs from chicken. Our study lays a foundation for further research on the mechanism of poultry OC in bone, calcium and phosphorus metabolic diseases.