薯蓣皂苷对蛋鸡脂肪肝综合征肝损伤和氧化应激的缓解作用及其机制研究

Mitigative effect and mechanism of dioscin on liver injury and oxidative stress in laying hens with fatty liver syndrome

旨在探究薯蓣皂苷(dioscin, Dio)对高能低蛋白饲喂诱导蛋鸡脂肪肝综合征脂代谢紊乱和氧化损伤的缓解作用及其机制的影响。以105只150日龄的蛋鸡作为试验对象,分为5组:对照组,HELP(高能低蛋白饲料)组,低剂量组(添加100 mg/kg Dio),中剂量组(添加500 mg/kg Dio),高剂量组(添加1 000 mg/kg Dio),饲喂3个月后剖检观察,测定肝组织AMP-依赖性蛋白激酶(AMPK)和核因子红细胞系2相关因子2(Nrf2)转录水平。体外以棕榈酸钠(PA)诱导鸡肝癌(LMH)细胞代谢紊乱,CCK-8和吖啶橙/碘化丙啶检测细胞死亡,尼罗红染色观察脂滴,荧光探针法检测活性氧(ROS)产生水平,Western blot检测脂代谢、抗氧化相关蛋白表达水平,免疫荧光检测Nrf2入核。结果:与对照组相比,HELP组蛋鸡肝脏发生明显的脂肪变性和出血,中剂量组明显缓解肝脏脂肪变性和出血,提高AMPKα和Nrf2转录水平,Dio缓解PA诱导的LMH细胞死亡,减少LMH细胞中脂滴沉积,并显著上调AMPK通路(P<0.05),同时减少ROS产生并增加Nrf2的表达及入核。综上,Dio能够缓解高能低蛋白饲喂诱导蛋鸡脂肪肝综合征脂代谢紊乱和氧化损伤,激活LMH细胞AMPK和Nrf2信号通路缓解PA诱导的LMH细胞脂代谢紊乱及氧化应激的发生,为其作为一种生理调节剂用于家禽脂肪肝及其相关的营养代谢症的防控提供理论依据。

This study aimed to explore the effect of dioscin(Dio)on lipid metabolism disorder and oxidative damage in laying hens induced by high-energy and low-protein feeding and fatty liver syndrome,and to study the mechanism of the effect.105150-day-old laying hens were used as experimental subjects.And they were divided into five groups:the control group;the HELP group(high-energy and low-protein feed group);the HELP+Dio group(added with 100 mg/kg Dio,500 mg/kg Dio,and 1000 mg/kg Dio,respectively).After three months of feeding,necropsy was performed and the transcription levels of AMP-dependent protein kinase(AMPK)and nuclear factor erythroid 2-related factor 2(Nrf2)in the liver tissue were measured.Then,metabolic disorders of chicken liver cancer cells(Leghorn Male Hepatoma cell line,LMH)were induced by sodium palmitate(PA)in vitro,cell death was detected by CCK-8 and acridine orange/propidium iodide,and lipid droplets were observed by NeilRed staining.Next,the fluorescent probe method was used to detect the production level of the reactive oxygen species(ROS).Finally,Western blot was used to detect the expression levels of lipid metabolism and antioxidantrelated proteins,and immunofluorescence was used to detect the nuclear entry of Nrf2.The results showed that,compared with the control group,the laying hens in the HELP group experienced significant steatosis and hemorrhage in the liver.Adding 500 mg/kg Dio to their diet significantly alleviated liver steatosis and hemorrhage in them,and raised their AMPKαand Nrf2 transcription levels.Dio alleviated PA induced by LMH cell death,reduced lipid droplet deposition in the LMH cells,and significantly increased the AMPK pathway(P<0.05),while reducing ROS production and increasing Nrf2 expression and nuclear import.In conclusion,Dioscin alleviated lipid metabolism disorder and oxidative damage in fatty liver syndrome in laying hens induced by high-energy and low-protein feeding.Diosgenin also activated AMPK and Nrf2 signaling pathways in the LMH cells and alleviated lipid metabolism disorder and oxidative stress induced by PA in these cells.The present results provide a theoretical basis for the use of Diosgenin as a physiological regulator in the prevention and control of poultry fatty liver and related nutritional metabolic diseases.