炎症状态下GR和NF-κB介导的猪脂肪组织ZAG转录调控研究

Zinc alpha 2-glycoprotein (ZAG) is a newly identified adipocytokine. Now it received much attention in human obesity research and early cancer diagnosis, however until now the mechanism of its expression regulation under disease is still unclear. In pigs, there are even no reports on ZAG. We cloned porcine ZAG sequence, and detected its gene expression significantly increased after LPS injection, ZAG content in plasma was also significantly increased in our previously study. However, why inflammation induces ZAG expression increasing is largely unknown. Glucocorticoid (GC) is important hormone involved in stress response, and increased after inflection. GC exerted its function by binding to glucocorticoid receptor (GR). Nuclear factor-κB (NF-κB) participated in many inflammation regulation. Many GR and NF-κB binding sites in the ZAG promoter have been predicted by bioinformatics. Therefore, in the present study, we investigated the ZAG transcript regulation mediated by GR and NF-κB under inflammation state in pigs. The results will provide basis for targeting regulation ZAG expression, and the clinic utilization of ZAG, and give new clue for the research of disease and growth in pigs.

锌α 2糖蛋白（Zinc alpha 2-glycoprotein，ZAG）是一种近年来确认的脂肪因子，目前在人类肥胖、糖尿病等疾病的治疗和癌症诊断中备受关注，具有重要的研究价值，在猪上尚未见报道。本课题组研究发现脂多糖诱导的炎症状态下猪脂肪组织中ZAG表达显著上升，但其机理尚不清楚。糖皮质激素受体( glucocorticoid receptor, GR) 和核因子（nuclear factor-κB，NF-κB）都是炎症诱导的重要转录因子。GR介导糖皮质激素的功能，NF-κB是调控转录多种炎性因子的中心环节。我们用生物信息学预测到猪ZAG基因的启动子存在多个GR以及NF-κB的结合位点。因此本课题拟分析炎症状态下GR 和NF-κB是否参与猪脂肪组织ZAG基因转录调控以及探讨其作用发生的机制。研究结果将为猪ZAG表达的靶向调控提供重要依据，并为猪ZAG功能的进一步认识及应用奠定良好基础。

锌α2糖蛋白（Zinc alpha 2-glycoprotein，ZAG）是一种近年来确认的脂肪因子，目前在人类肥胖、糖尿病等疾病的治疗和癌症诊断中备受关注，具有重要的研究价值，在猪上尚未见报道。本课题组克隆了猪ZAG序列，确认该片段定位于猪3号染色体上。分析了不同品种猪ZAG表达的差异，发现大白猪血液、脂肪和肝脏等组织中ZAG基因和蛋白均显著高于二花脸猪，与猪品种间能量代谢差异相关。研究表明炎症感染情况下猪血浆ZAG含量显著上升，猪肝脏和脂肪中ZAG的蛋白表达显著升高，并且肝脏糖脂代谢发生紊乱。进一步分别以ZAG过表达和敲除小鼠为模型，揭示ZAG在炎症和应激情况下的表达调控及其作用机制。研究结果发现过表达ZAG使肝脏线粒体数量增加，线粒体相关蛋白COX1和UCP3以及线粒体复合物Ⅲ和Ⅴ的活性增加。LPS处理会激活TLR4/NFκB炎症信号通路，促进NFκBp65入核与CBP结合，进而诱导炎症反应。ZAG过表达后通过激活β3-AR/PKA/CREB信号通路，促使CREB发生磷酸化，使CREB与CBP结合，竞争性抑制NFκBp65与CBP结合，从而缓解炎症。在地塞米松模拟应激模型下敲除ZAG显著降低血浆TG和NEFA含量，这一变化是由于ZAG使内脏脂肪组织中脂质动员能力的下降所引起的，GR可能参与了调节。冷应激下过表达ZAG通过β3-AR上调P-P38，从而增加PGC1α表达，激活PPARγ，最终促进白脂转分化为米色脂肪，增加产热。本研究结果揭示了猪ZAG不同品种表达差异和炎症状态下ZAG的表达调控机制，并进一步探索了ZAG在炎症和应激状态下可能的功能。项目以ZAG为纽带，揭示了动物机体代谢和炎症两个要素系统之间的关系，从整合生理学的角度，为猪生长和免疫调控提供新的靶点，为猪健康养殖提供理论依据。本课题资助发表SCI文章14篇，培养博士生2名，硕士生4名。