维生素K_(2)单独及与钙剂联合使用对斑马鱼骨骼健康的改善作用及机制研究

Effect of Vitamin K_(2) Alone and in Combination with Calcium on the Bone Health Improvement and Mechanism in Zebrafish Model

目的:研究维生素K_(2)单独使用及与钙剂联合使用对斑马鱼骨骼健康改善作用及机制。方法:采用转基因Tg系健康斑马鱼作为实验动物,利用泼尼松龙诱导骨质疏松斑马鱼模型,显微镜下观察并拍照,以斑马鱼成骨细胞荧光面积和荧光密度为指标,评价药物处理后斑马鱼骨骼发育情况,并采用Real-time qPCR检测相关基因表达变化。结果:在5~20μg/mL的实验安全剂量范围内,维生素K_(2)及与D_(3)和钙剂联合使用,均能显著改善由泼尼松龙诱导的斑马鱼成骨细胞荧光面积和荧光密度的降低(P<0.05);维生素K_(2)通过促进成骨细胞生成相关的alp、bmp2a和bmp4基因的表达,抑制破骨细胞生成相关的opg、rank1、trap、ctsk基因的表达以及抑制细胞外基质中骨钙化相关基因vdrb、sparc、colla2表达从而发挥功效。结论:维生素K_(2)通过促进成骨细胞生成相关基因的表达,抑制破骨细胞和细胞外基质中骨钙化相关基因的表达,发挥改善斑马鱼骨骼健康的作用,其与钙剂联合给药,可提升其改善骨骼健康能力。

Objective:To investigate the bone health improvement effects and mechanism of vitamin K_(2) alone and in combination with calcium on osteoporotic zebrafish model.Methods:Healthy zebrafish with transgenic Tg line were used as experimental animals.The model of osteoporosis zebrafish was induced by prednisone.The fluorescence area and density of zebrafish osteoblasts were used as indicators to evaluate the bone development of zebrafish which were observed and photographed under microscope.Furthermore,the expression changes of related genes were detected by Real-time qPCR.Results:Vitamin K_(2) alone and in combination with D_(3) and calcium chloride could significantly improve the reduction of fluorescence area and fluorescence density of zebrafish osteoblasts induced by prednisolone in the experimental safe dose range of 5~20μg/mL(P<0.05).Vitamin K_(2) played anti-osteoporosis activity by promoting the expression of alp,bmp2a and bmp4 genes related to osteoblast,inhibiting the expression of opg,rank1,trap and ctsk genes related to osteoclast,and inhibiting the expression of vdrb,sparc and colla2 genes related to bone extracellular matrix.Conclusion:Vitamin K_(2) could improve the bone health of zebrafish by promoting the expression of genes related to osteoblast formation and inhibit the expression of genes related to osteoclasts and bone calcification in extracellular matrix.The combined administration of vitamin K_(2) with D_(3) and calcium chloride could enhance the ability to improve bone health.