IGF-1对鸽嗉囊上皮细胞脂质形成调控机制的研究

In the preliminary study, concentrations of nutrient components and cell factors in crop milk from male and female parent pigeons were examined, and the lipid meatabolism mechanism in pigeon squabs was also studied. This program will be conducted to investigate the mechanism of lipid formation in pigeon crop epithelial cell under the regulation of insulin-like growth factor 1 (IGF-1). The gene and protein expresions of fatty acid tanslocase (FAT/CD36), keratinocyte lipid-binding protein gene (KLBP), acyl-CoA-binding protein (ACBP), fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC) will be determined to clarify the approaches of lipid formation in crop milk of male and female pigeons, and their sexual differences will also be explained. IGF-1 receptor (IGF-1R) antagonist (BMS-536924) will be used to prove that IGF-1 has an effect on lipid formation in crop in vivo. A primary cell culture method of crop epithelial cell will be established. Depending on this model, the effect of IGF-1 on cell fatty acid transportation and synthesis can be investigated in vitro. And then,the dynamic changing pattern of IGF-1R in crop tissues under different stages will be studied. Thus, the mechanism of IGF-1’s regulation of lipid formation in pigeon crop would be revealed. The whole study could supply the new theoretical evidences for physological reseraches on altricial birds, and a theoretical basis for future studies on nutrient requirements of parent pigeons and producing artificial crop milk would also be provided.

本项目在前期分析比较了雌雄亲鸽分泌鸽乳中营养组分和细胞因子的含量，并对雏鸽的脂肪代谢进行研究的基础上，开展胰岛素样生长因子IGF-1对鸽嗉囊上皮细胞脂质形成机制的研究。通过检测繁殖期雌雄鸽嗉囊组织脂肪酸转运载体（脂肪酸转位酶FAT/CD36、脂肪酸结合蛋白KLBP和酰基CoA结合蛋白ACBP）和脂肪酸从头合成关键酶（脂肪酸合成酶FAS和乙酰CoA羧化酶ACC）基因和蛋白的表达规律，旨在揭示雌雄亲鸽鸽乳脂质形成的途径和性别差异机制。分析IGF-1受体拮抗剂BMS-536924对鸽乳脂质形成的调控作用，并以鸽嗉囊上皮细胞原代培养体系为基础，离体研究IGF-1对鸽嗉囊上皮细胞脂肪酸转运和合成的影响，同时，结合繁殖期雌雄鸽嗉囊组织IGF-1R的表达规律，共同阐明IGF-1调控鸽乳脂质形成的作用机制，丰富晚成雏鸟类生理学的理论，并为肉种鸽营养需求研究，开发符合雏鸽生长需求的新型人工鸽乳提供依据。

人们对鸽乳形成机制的研究甚少，这在很大程度上制约了肉种鸽营养需求和人工鸽乳的开发研究。因鸽乳中碳水化合物的极度缺乏，雏鸽发育前期将脂肪作为维持机体生命活动的最主要能量来源。脂肪酸不仅为机体供能，参与细胞膜的构建，而且在免疫炎症反应和心血管及神经系统的发育和维持正常生理功能方面具有重要的作用。因此，本项目以美国白羽王鸽为研究对象，探讨繁殖期鸽乳脂质形成的调控机制。分别利用组织切片、离体培养、荧光定量PCR等技术研究了繁殖期鸽嗉囊组织形态学、生长因子含量、嗉囊组织相关因子受体和脂肪酸合成及转运相关基因的表达，并基于体外组织培养法探讨了胰岛素样生长因子IGF-1对鸽嗉囊组织脂质形成的影响。结果表明：（1）鸽嗉囊上皮组织在孵化期末至哺育期初期迅速增生，脂滴显著增大且增多，哺育期后期出现退化，且上皮细胞中脂滴数量明显减少，细胞桥粒增多；（2）鸽乳形成高峰期内，种鸽肝脏中糖酵解、糖异生、脂肪代谢关键基因均显著上调；（3）鸽血清和嗉囊组织中IGF-1、EGF、PRL含量呈规律性变化，均在鸽乳形成高峰期出现最大值；（4）孵化期末至哺育期前期，鸽嗉囊脂肪酸转位酶（FAT/CD36）、脂肪酸结合蛋白5（EFABP）和乙酰辅酶A结合蛋白（ACBP）以及脂肪酸合成酶（FAS）和乙酰辅酶A羧化酶（ACC）基因均存在显著上调；（5）孵化期第10天和哺育期第8天鸽嗉囊组织于体外培养0-3天内可维持组织结构的完整性，并于2-3天内细胞活性及相关基因表达保持稳定，离体条件下IGF-1能够诱导鸽嗉囊上皮细胞形成脂质；（6）离体和在体试验证明IGF-1受体拮抗剂BMS-536924能够通过影响脂肪酸的转运和合成有效抑制鸽嗉囊组织脂质的形成。综上所述，繁殖期亲鸽嗉囊和肝脏组织的代谢适应为鸽乳脂质的形成提供生理学和营养学基础。IGF-1通过受体介导参与调控鸽乳的形成，且鸽乳脂质中的脂肪酸来源于血液运输和自身组织合成两种途径。本项目的开展不仅弥补了晚成雏鸟类相关生理学研究的空白，也为后续人工鸽乳和肉种鸽营养需求的研究奠定理论基础。