基于AMPK信号通路探讨胍基乙酸对运输应激黔东南小香鸡肌肉糖酵解的调控作用

The transportation process is considered the most stressful environment challenge experienced by broilers before slaughtering. Transport stress has been reported to induce the acute stress response of broilers and result in the poor postmortem meat quality, which brings great economic losses to poultry industry and meat processing enterprises. Transport stress in Guizhou province, which is a mountain area, maybe more serious. Guanidine acetic acid (GAA), a natural precursor of creatine, could regulate the muscle energy metabolism, muscle glycolysis and improve broiler meat quality. However, the regulation mechanism of GAA on meat glycolysis of broilers in transportation stress is still unknown. Therefore, Southeastern Guizhou xiaoxiang chicken, an excellent native breed of Guizhou province, were chosen to investigate the effects of GAA on meat quality of broilers through analyzing the variation of energy pool, meat glycolytic potential and the key enzymes activities in meat glycolysis. Moreover, AMPK signaling and total protein acetylation protein expression were also investigated to reveal the regulatory mechanism of GAA on energy metabolism and meat glycolysis. This program can provide technical reference and theoretical basis for nutrition regulation and high broiler meat production, and play an important role in the practice of broiler production.

宰前运输应激会引起肉鸡产生强烈的应激反应，导致宰后肉品质变差，给养殖业和肉禽加工企业带来重大经济损失。贵州是多山地区，运输应激问题更加突出。胍基乙酸（GAA）是动物体内肌酸的天然前体，能够调控肌肉细胞能量代谢及肌肉糖酵解进程，改善肉品质。但GAA调控运输应激肌肉糖酵解代谢的机理尚不明确。因此，本研究拟选用贵州省优良地方品种—黔东南小香鸡为研究对象，通过宰前日粮添加GAA，分析肌肉能量代谢池的变化、肌肉糖酵解潜力及糖酵解代谢关键酶活性，揭示GAA对运输应激肌肉糖酵解的调控作用；并深入分析AMPK信号及蛋白总乙酰化表达水平，阐明GAA调控运输应激肉鸡能量代谢及肌肉糖酵解的作用机制。本项目的实施可为宰前应激肉鸡营养调控和生产优质鸡肉提供技术参考和理论依据，在肉鸡生产实践中具有重要的指导意义。

运输应激是宰前应激中最主要的应激之一，能够引起肉鸡强烈的应激反应及肌肉糖原和乳酸含量变化，致使宰后肉质变差。宰前能量调控是一种增加肌肉能量贮备，调控宰后肌肉能量代谢进程，进而改善肉品质的有效技术手段。本项目选择贵州省著名品种—黔东南小香鸡为研究对象，通过在其日粮中添加胍基乙酸（guanidino acetic acid, GAA）,探讨其对宰前运输应激黔东南小香鸡肌肉糖酵解的调控作用。并进一步利用RT-PCR、蛋白免疫印迹及代谢组学，深入探讨了宰前添加GAA对运输应激黔东南小香鸡肌肉糖酵解的调控机制。结果显示：（1）GAA能够缓解运输应激（血液皮质酮水平降低），改善肉品质（增加宰后pH24h值，降低肌肉L*值、蒸煮损失及滴水损失），初步探讨了日粮添加GAA对运输应激黔东南小香鸡肉品质的影响；（2）GAA增加了肌肉能量贮备（增加肌酸及磷酸肌酸含量）、降低了肌肉糖酵解潜力及糖酵解关键酶活性，以上两项研究内容提示，日粮添加GAA能够通过增加能量贮备，缓解宰后肌肉糖酵解从而改善宰后肌肉肉品质；（3）利用C57bl小鼠为模型，通过灌服GAA及腹腔注射激活剂和抑制剂，在宰前进行应激处理。结果发现，GAA增加了肌肉pH24h，降低了肌肉乳酸含量及肌肉LKB1和AMPKα2蛋白表达水平，降低了肌肉总蛋白乙酰化水平。这表明GAA通过介导AMPK信号及蛋白乙酰化水平调控了应激对宰后肌肉的糖酵解进程；（4）根据研究内容（3）确定的AMPK信号关键蛋白表达，进一步测定了运输应激黔东南小香鸡肌肉LKB1、AMPKα1及AMPKα2 mRNA表达、LKB1及AMPKα2 蛋白表达及蛋白总乙酰化水平，揭示了GAA对运输应激黔东南小香鸡肌肉糖酵解的调控机制；（5）进一步利用代谢组学研究了GAA对运输应激黔东南小香鸡肌肉代谢的影响，发现GAA添加影响了宰后肌肉碳水化合物代谢、能量代谢、氧化应激代谢、氨基酸代谢及肌酸代谢，进而影响了肉品质。本项目的研究结果可为宰前应激肉鸡能量调控提供技术参考，在高效优质肉鸡生产实践中具有重要的指导意义。