不同氮源对瘤胃上皮细胞凋亡的影响  

Effect of different nitrogen sources on apoptosis of rumen epithelial cells

作　　者：王永浪 叶恒博 曹应东 沈宇龙 黄俊武 农谨祯 孔志伟 WANG Yonglang;YE Hengbo;CAO Yingdong;SHEN Yulong;HUANG Junwu;NONG Jinzhen;KONG Zhiwei(College of Animal Science and Technology,Guangxi University,Nanning Guangxi 530004,China)

机构地区：[1]广西大学动物科学与技术学院,广西南宁530004

出　　处：《中国饲料》2024年第9期53-59,共7页China Feed

基　　金：国家自然科学基金青年基金(32102576);广西大学高层次人才引进项目(A3340051019)。

摘　　要：本试验采用细胞增殖试验、流式细胞术检测试验、Western blot试验、Hochest33342染色荧光显微镜检测等探究不同氮源(蛋氨酸与氯化铵)对瘤胃上皮细胞凋亡的影响及其可能的调控机制。结果表明:对瘤胃上皮细胞来说,处理36 h时,添加0.12 mmol NH4Cl组和0 mmol Met缺失组的OD值均达到对照组的一半,且均显著低于0.048 mmol Met组、0.12 mmol Met组(对照组)和0.048 mmol NH4Cl组(P <0.05),0 mmol Met组比0.12 mmol NH4Cl组细胞活性显著降低(P <0.05);添加0 mmol Met组的总凋亡率和晚期凋亡率最高,显著高于0.12 mmol Met组(对照组)和0.12 mmol NH4Cl组(P <0.05),而0 mmol Met组和0.12 mmol NH4Cl组的早期细胞凋亡率均显著高于0.12 mmol Met组(对照组)(P <0.05);与对照组相比,0.12 mmol NH4Cl组和0 mmol Met组的线粒体膜电势分别显著下降到71.99%和67.67%;与对照组相比,0 mmol Met组和0.12 mmol NH4Cl组的促调亡蛋白Caspase-3、Bax和p53的表达均显著上调(P <0.05),抑调亡蛋白Bcl-2和Bcl-xl的表达均显著下调(P <0.05);与对照组相比,0 mmol Met组和0.12 mmol NH4Cl组的p-mTOR和p-AMPK蛋白的表达均显著上调(P <0.05),而PI3K和p-AKT的蛋白表达量显著下调(P <0.05)。综上,蛋氨酸缺乏会诱导瘤胃上皮细胞发生凋亡,NH4Cl替代可以缓解凋亡的发生,其诱导过程可能通过PI3K-Akt-mTOR信号通路和AMPK-mTOR信号通路完成,该结果说明了蛋氨酸不可或缺的重要性,可为反刍动物的配合饲料发展研究提供参考。In this experiment,cell proliferation assay,flow cytometry assay,Western blot assay,Hochest33342 staining fluorescence microscopy assay were used to explore the effects of different nitrogen sources(methionine and ammonium chloride)on the apoptosis of rumen epithelial cells and their possible regulatory mechanisms.The results showed that for the rumen epithelial cells,the OD values of both the 0.12 mmol NH4Cl-added group and the 0 mmol Met-deficient group reached half of those of the control group at 36 h of treatment,which were significantly lower than those of the 0.048 mmol Met group,the 0.12 mmol Met group(control group)and the 0.048 mmol NH4Cl group(P<0.05);the total apoptosis rate and late apoptosis rate were the highest in the 0 mmol Met group,which were significantly higher than those in the 0.12 mmol Met group(control group)and 0.12 mmol NH4Cl group(P<0.05),while the early apoptosis rate in the 0 mmol Met group and 0.12 mmol NH4Cl group were significantly higher than those in the 0.12 mmol Met group(control group)(P<0.05);the mitochondrial membrane potential was significantly decreased to 71.99%and 67.67%in the 0.12 mmol NH4Cl group and 0 mmol Met group,respectively,compared with the control group.The expression of the pro-regulatory proteins Caspase-3,Bax and p53 were significantly up-regulated in the 0 mmol Met group and 0.12 mmol NH4Cl group(P<0.05)and the expression of the inhibitory proteins Bcl-2 and Bcl-xl were significantly down-regulated(P<0.05)in the 0 mmol Met and 0.12 mmol NH4Cl groups compared with the control group;the expression of p-mTOR and p-AMPK proteins were significantly up-regulated(P<0.05)in the 0 mmol Met and 0.12 mmol NH4Cl groups compared with the control group,while the expression of PI3K and p-AKT proteins were significantly down-regulated(P<0.05).The above results indicated that methionine deficiency induced apoptosis in rumen epithelial cells,and NH4Cl substitution could alleviate the occurrence of apoptosis,and the induction process could be accomplished through PI3K-Akt-mT

关 键 词：瘤胃上皮细胞 细胞凋亡 蛋氨酸 氯化铵 

分 类 号：S811.2[农业科学—畜牧学]