鸡zp1基因对成骨细胞矿化的影响

Effect of the chicken zp1 gene on osteoblast mineralization

摘要:

本研究旨在克隆编码鸡透明带1（zona pellucida 1，Zp1）蛋白的zp1基因，并研究其组织表达谱及在成骨细胞矿化中的作用。利用实时荧光定量聚合酶链式反应（real-time fluorescence quantitative polymerase chain reaction，RT-qPCR）检测蛋鸡不同组织和性成熟启动前后胫骨中的zp1表达水平；将Zp1过表达载体转染至诱导分化8 d的鸡颅骨成骨细胞内，检测细胞外矿化基质和矿化相关基因表达。结果表明，鸡zp1基因全长3 045 bp，编码958个氨基酸残基，具有2个N-糖基化位点。zp1基因在产蛋期鸡肝脏中高水平表达，其次为胫骨和卵黄膜，在蛋壳腺和心脏中不表达。鸡性成熟启动后血浆雌激素（estrogen，E2）、胫骨糖胺聚糖（glycosaminoglycan，GAG）含量和zp1表达量均极显著高于性成熟启动前。在过表达Zp1的鸡成骨细胞中添加雌激素后，细胞外矿化基质和矿化相关基因表达水平均显著升高。因此，zp1基因表达具有组织特异性，在雌激素作用下正向调控鸡成骨细胞矿化，为阐明Zp1在鸡产蛋期骨骼中的功能特性奠定了理论基础。

Abstract:

The aim of this study was to clone the chicken zp1 gene encoding zona pellucida 1 (Zp1) and investigate its tissues expression profile and its effect on osteoblast mineralization. The expression level of zp1 was quantified in various tissues of laying hens and in the tibia of the pre- and post-sexual maturity by RT-qPCR. Zp1 overexpressed vector was transfected into chicken calvarial osteoblasts which were induced differentiation for 8 days, and the extracellular mineral and the expression of mineralization-related genes were detected. The full-length chicken zp1 gene is 3 045 bp, encoding 958 amino acids residuals, and has two N-glycosylation sites. The highest expression level of the zp1 gene was found in the liver, followed by the tibia and yolk membrane, while no expression was detected in the heart and eggshell gland. Compared with the pre-sexual maturity hens, the concentration of estrogen (E2) in plasma, the content of glycosaminoglycan (GAG) and the expression level of the zp1 gene in the tibia with post-sexual maturity were higher. The extracellular matrix and the level of osteoblast mineralization-related genes showed a significantly upregulated expression in chicken calvarial osteoblasts with Zp1 overexpressed and addition of estrogen. The expression of the zp1 gene is tissue-specific and positively regulated osteoblast mineralization under the action of estrogen, laying the foundation for elucidating the functional properties of Zp1 in chicken bones during the egg production period.