Li Tingting, Geng Yanqiang, Hu Yun, Zhang Liyang, Cui Xiaoyan, Zhang Weiyun, Gao Feiyu, Liu Zongping, Luo Xugang
Poultry Mineral Nutrition Laboratory, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.
Mineral Nutrition Research Division, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing, China.
Front Vet Sci. 2022 Apr 26;9:875140. doi: 10.3389/fvets.2022.875140. eCollection 2022.
Three experiments were carried out in the present study to investigate whether dentin matrix protein 1 (DMP1) was involved in regulating phosphorus (P) metabolic utilization in primary cultured tibial osteoblasts of broiler chicks. Experiment 1 was conducted to select the optimal osteogenic inductive culture medium and the optimal induction time in primary cultured tibial osteoblasts of broiler chicks. In experiment 2, the siRNAs against DMP1 were designed, synthesized and transfected into primary cultured tibial osteoblasts of broiler chicks, and then the inhibitory efficiencies of siRNAs against DMP1 were determined, and the most efficacious siRNA was selected to be used for the DMP1 silencing. In experiment 3, with or without siRNA against DMP1, primary cultured tibial osteoblasts of broiler chicks were treated with the medium supplemented with 0.0, 1.0 or 2.0 mmol/L of P as NaHPO for 12 days. The P metabolic utilization-related parameters were measured. The results showed that the osteogenic induced medium 2 and 12 days of the optimal induction time were selected; Among the designed siRNAs, the si340 was the most effective ( < 0.05) in inhibiting the DMP1 expression; DMP1 silencing decreased ( < 0.05) the expressions of mRNA and protein, P retention rate, mineralization formation, alkaline phosphatase activity and bone gla-protein content in tibial osteoblasts at all of added P levels. It is concluded that DMP1 silencing inhibited P utilization, and thus DMP1 was involved in regulating P metabolic utilization in primary cultured tibial osteoblasts of broiler chicks, which provides a novel insight into the regulation of the P utilization in the bone of broilers, and will contribute to develop feasible strategies to improve the bone P utilization efficiency of broilers so as to decrease its excretion.
本研究进行了三项实验，以探究牙本质基质蛋白1（DMP1）是否参与调节肉仔鸡原代培养胫骨成骨细胞中磷（P）的代谢利用。实验1旨在选择肉仔鸡原代培养胫骨成骨细胞的最佳成骨诱导培养基和最佳诱导时间。在实验2中，设计、合成针对DMP1的小干扰RNA（siRNAs）并转染至肉仔鸡原代培养胫骨成骨细胞，然后测定siRNAs对DMP1的抑制效率，选择最有效的siRNA用于DMP1沉默。在实验3中，在添加或不添加针对DMP1的siRNA的情况下，用补充有0.0、1.0或2.0 mmol/L磷（以NaHPO形式）的培养基处理肉仔鸡原代培养胫骨成骨细胞12天。测定与磷代谢利用相关的参数。结果表明，选择了成骨诱导培养基2和最佳诱导时间12天；在所设计的siRNAs中，si340在抑制DMP1表达方面最有效（<0.05）；在所有添加磷水平下，DMP1沉默均降低了（<0.05）胫骨成骨细胞中mRNA和蛋白的表达、磷保留率、矿化形成、碱性磷酸酶活性和骨钙素含量。结论是，DMP1沉默抑制了磷的利用，因此DMP1参与调节肉仔鸡原代培养胫骨成骨细胞中磷的代谢利用，这为肉仔鸡骨骼中磷利用的调节提供了新的见解，并将有助于制定可行的策略来提高肉仔鸡骨骼磷利用效率，从而减少其排泄。
