N-乙酰半胱氨酸对脂多糖刺激仔猪免疫反应和小肠能量状态的影响

doi:10.16431/j.cnki.1671-7236.2021.11.009

摘要/Abstract

摘要： 试验旨在研究N-乙酰半胱氨酸（NAC）对脂多糖（LPS）刺激的仔猪免疫反应和小肠能量状态的影响。按照随机性原则将24头32日龄健康仔猪（11.58 kg±0.26 kg）分为3组（空白组、LPS组和NAC＋LPS组），每组设置8个重复。经过3 d适应期后，各组仔猪饲喂玉米-豆粕型基础饲粮（NAC＋LPS组在此基础上添加500 mg/kg NAC）。试验第21天，仔猪空腹称重后腹腔注射LPS （LPS组和NAC＋LPS组，注射剂量为100 μg/kg BW）或等量的生理盐水（空白组），3 h后前腔静脉采血，然后屠宰取肠黏膜、肝脏和淋巴结等样品，进行血细胞计数分析、小肠黏膜腺苷酸水平测定以及肝脏和淋巴结相关基因检测。结果表明，与空白组相比，LPS组仔猪血液中白细胞和中性粒细胞的数目显著提高（P<0.05），淋巴细胞数目显著降低（P<0.05）；小肠黏膜一磷酸腺苷（AMP）水平和一磷酸腺苷/三磷酸腺苷（AMP/ATP）值显著提高（P<0.05），ATP水平显著降低（P<0.05）；肝脏和淋巴结双链RNA依赖的蛋白质激酶（PKR）、干扰素诱导的四肽重复蛋白1（IFIT1）和黏病毒抗性蛋白1（MX1）基因的相对表达量均显著上调（P<0.05）。与LPS组相比，NAC＋LPS组仔猪血液中白细胞数、中性粒细胞数、嗜酸性粒细胞和淋巴细胞百分比均显著降低（P<0.05）；回肠黏膜AMP水平和AMP/ATP值均显著降低（P<0.05）；肝脏IFIT1、PKR和MX1基因的相对表达量均显著上调（P<0.05），淋巴结IFIT1基因的相对表达量显著下调（P<0.05）。综上所述，饲粮中添加500 mg/kg NAC可缓解LPS刺激引起的仔猪免疫反应，并且能有效改善肠黏膜能量状态。

关键词: N-乙酰半胱氨酸; 脂多糖; 仔猪; 小肠能量状态; 免疫反应

Abstract: This experiment was conducted to determine the effects of N-acetylcysteine (NAC) on the immune response and intestinal mucosal energy status in piglets challenged with lipopolysaccharide (LPS). Twenty-four healthy weaned piglets (11.58 kg±0.26 kg) were randomly allocated into 3 groups (control, LPS and NAC+LPS groups), with 8 replicates in each group. After a 3-day adaptation period, the piglets in each group were fed with basic diet (the NAC+LPS group was supplemented with 500 mg/kg NAC). On the 21st day of the trial, all the piglets were weighed and injected intraperitoneally with LPS (for LPS and NAC+LPS groups, injected with 100 μg/kg BW LPS) or the same amount of normal saline (for control group). 3 hours later, blood samples were collected from the anterior vena cava. Then all piglets were sacrificed, and samples from the small intestinal mucosal, liver and lymph node were collected. Blood cell counts, the level of adenylate in small intestinal mucosal and the relative expression levels of genes were detected. The results showed that:Compared with the control group, the number of white blood cells and neutrophils in the blood was significantly increased (P<0.05), the number of lymphocytes was significantly reduced (P<0.05). AMP level and AMP/ATP ratio in the small intestinal mucosal were increased (P<0.05), and the ATP level was decreased (P<0.05). The relative expression of dsRNA dependent protein kinase R (PKR), interferon-induced protein with tetratricopeptide repeats 1 (IFIT1) and myxovirus resistance protein 1 (MX1) genes in liver and lymph nodes were up-regulated in the LPS group (P<0.05). Compared with the LPS group, the number of white blood cell and neutrophils, the percentage of eosinophils and lymphocytes were decreased (P<0.05), the ileal mucosal AMP level and the AMP/ATP ratio decreased (P<0.05), the relative expression of IFIT1, PKR and MX1 genes in liver were up-regulated (P<0.05), and the relative expression of IFIT1 gene in lymph nodes was down-regulated (P<0.05) in the NAC+LPS group. These results suggested that NAC could alleviate the immune response of piglets and improve the energy state of the intestinal mucosa in LPS-challenged piglets.

Key words: N-acetylcysteine; lipopolysaccharide; piglets; intestinal energy status; immune response

中图分类号:

S816.7

王倩, 汪曼丽, 宁楠, 王帅杰, 谭子涵, 王蕾, 赵迪, 张倩, 侯永清. N-乙酰半胱氨酸对脂多糖刺激仔猪免疫反应和小肠能量状态的影响[J]. 中国畜牧兽医, 2021, 48(11): 3996-4003.

WANG Qian, WANG Manli, NING Nan, WANG Shuaijie, TAN Zihan, WANG Lei, ZHAO Di, ZHANG Qian, HOU Yongqing. Effect of N-acetylcysteine on Immune Response and Intestinal Mucosal Energy Status in Piglets Challenged with Lipopolysaccharide[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(11): 3996-4003.

参考文献

[1] WU Q J, ZHOU Y M, WU Y N, et al. The effects of natural and modified clinoptilolite on intestinal barrier function and immune response to LPS in broiler chickens[J]. Veterinary Immunology and Immuno-pathology, 2013, 153(1-2):70-76.
[2] ZHU H, PI D, LENG W, et al. Asparagine preserves intestinal barrier function from LPS-induced injury and regulates CRF/CRFR signaling pathway[J]. Innate Immunity, 2017, 23(6):546-556.
[3] WANG L, HOU Y Q, YI D, et al. Dietary supplementation with glutamate precursor α-ketoglutarate attenuates lipopolysaccharide-induced liver injury in young pigs[J]. Amino Acids, 2015, 47(7):1309-1318.
[4] 刘坚, 侯永清, 丁斌鹰, 等.α-酮戊二酸对脂多糖应激仔猪肠黏膜能量代谢的影响[J]. 动物营养学报, 2009, 21(6):892-896. LIU J, HOU Y Q, DING B Y, et al. Effect of α-ketoglutaric acid on energy metabolism in intestinal mucosa of weaened pigs chronically challenged with lipopolysaccharide[J]. Chinese Journal of Animal Nutrition, 2009, 21(6):892-896.(in Chinese)
[5] BAVARSAD S R, HARRIGAN M R, ALEXANDROV A V.N-acetylcysteine (NAC) in neurological disorders:Mechanisms of action and therapeutic opportunities[J]. Brain and Behavior, 2014, 4(2):108-122.
[6] YI D, HOU Y Q, XIAO H, et al. N-acetylcysteine improves intestinal function in lipopolysaccharides-challenged piglets through multiple signaling pathways[J]. Amino Acids, 2017, 49(12):1915-1929.
[7] YI D, HOU Y Q, TAN L, et al. N-acetylcysteine improves the growth performance and intestinal function in the heat-stressed broilers[J]. Animal Feed Science and Technology, 2016, 220:83-92.
[8] ZALEWSKA A, SZARMACH I, ENDZIAN P M, et al. The effect of N-acetylcysteine on respiratory enzymes, ADP/ATP ratio, glutathione metabolism, and nitrosative stress in the salivary gland mitochondria of insulin resistant rats[J]. Nutrients, 2020, 12(2):458.
[9] HOU Y Q, WANG L, YI D, et al. N-acetylcysteine reduces inflammation in the small intestine by regulating redox, EGF and TLR4 signaling[J]. Amino Acids, 2013, 45(3):513-522.
[10] HAN J, LIU Y L, FAN W, et al. Dietary L-arginine supplementation alleviates immunosuppression induced by cyclophosphamide in weaned pigs[J]. Amino Acids, 2009, 37(4):643-651.
[11] 孙嵩.血常规以及超敏C反应蛋白检测的临床意义观察[J]. 继续医学教育, 2018, 32(3):144-146. SUN S.Clinical significanceof routine blood and hypersensitive C-reactive protein detection[J]. Continuing Medical Education, 2018, 32(3):144-146.(in Chinese)
[12] BOSKABADI J, MOKHTARI-ZAER A, ABARESHI A, et al. The effect of captopril on lipopolysaccharide-induced lung inflammation[J]. Experimental Lung Research, 2018, 44(4-5):191-200.
[13] 吴天佑.精氨酸对脂多糖诱发泌乳奶牛炎性反应的缓解作用及其分子机制[D].扬州:扬州大学, 2016. WU T Y.Mechanism of relieve effect of arginine on the inflammatory responses induced by lipopolysaccharide in lactating cows[D].Yangzhou:Yangzhou University, 2016.(in Chinese)
[14] 陈庆梓, 扶招弟, 周玉波, 等.N-乙酰半胱氨酸减轻臭氧所致的小鼠肺部炎症反应[J]. 生理学报, 2016, 68(6):767-774. CHEN Q Z, FU Z D, ZHOU Y B, et al. N-acetyllcysteine reduces the ozone-induced lung inflammation response in mice[J]. Acta Physiologica Sinica, 2016, 68(6):767-774.(in Chinese)
[15] LIU Y L, HUANG J J, HOU Y Q, et al. Dietary arginine supplementation alleviates intestinal mucosal disruption induced by Escherichia coli lipopolysaccharide in weaned pigs[J]. British Journal of Nutrition, 2008, 100(3):552-560.
[16] PSARRA A M, SOLAKIDI S, SEKERIS C E.The mitochondrion as a primary site of action of steroid and thyroid hormones:Presence and action of steroid and thyroid hormone receptors in mitochondria of animal cells[J]. Molecular and Cellular Endocrinology, 2006, 246(1-2):21-33.
[17] PLOTNIKOV E Y, PEVZNER I B, ZOROVA L D, et al. Mitochondrial damage and mitochondria-targeted antioxidant protection in LPS-induced acute kidney injury[J]. Antioxidants (Basel, Switzerland), 2019, 8(6):176.
[18] 王惠云, 廖满, 王蕾, 等.N-乙酰半胱氨酸和冷应激对肉鸡肠道组织形态、能量代谢和抗氧化能力的影响[J]. 中国畜牧杂志, 2018, 54(2):52-59. WANG H Y, LIAO M, WANG L, et al. Effects of N-acetylcysteine and cold stress on intestinal morphology, energy status and antioxidant capacity in broilers[J]. Chinese Journal of Animal Science, 2018, 54(2):52-59.(in Chinese)
[19] RUDERMAN N B, CARLING D, PRENTKI M, et al. AMPK, insulin resistance, and the metabolic syndrome[J]. The Journal of Clinical Investigation, 2013, 123:2764-2772.
[20] HOU Y Q, YAO K, WANG L, et al. Effects of α-ketoglutarate on energy status in the intestinal mucosa of weaned piglets chronically challenged with lipopolysaccharide[J]. The British Journal of Nutrition, 2011, 3(106):357-363.
[21] PAN S, WANG N, BISETTO S, et al. Downregulation of adenine nucleotide translocator 1 exacerbates tumor necrosisfactor-α-mediated cardiac inflammatory responses[J]. American Journal of Physiology Heart and Circulatory Physiology, 2015, 308(1):39-48.
[22] DING Y, ZHENG Y, HUANG J, et al. UCP2 ameliorates mitochondrial dysfunction, inflammation, andoxidative stress in lipopolysaccharide-induced acute kidney injury[J]. International Immunopharmacology, 2019, 71:336-349.
[23] ZHONG X, HE J, ZHANG X, et al. UCP2 alleviates tubular epithelial cell apoptosis in lipopolysaccharide-induced acute kidney injury by decreasing ROS production[J]. Biomed Pharmacotherapy, 2019, 115:108914.
[24] DEHART D N, FANG D, HESLOP K, et al. Opening of voltage dependent anion channels promotes reactive oxygen species generation, mitochondrial dysfunction and cell death in cancer cells[J]. Biochemical Pharmacology, 2018, 148:155-162.
[25] FOX L E, LOCKE M C, LENSCHOW D J.Context is key:Delineating the unique functions of IFNα and IFNβ in disease[J]. Frontiers in Immunology, 2020, 11:606874.
[26] HALLER O, STAEHELI P, SCHWEMMLE M, et al. Mx GTPases:Dynamin-like antiviral machines of innate immunity[J]. Trends in Microbiology, 2015, 23(3):154-163.
[27] 刘存.BVDV感染MDBK细胞转录组学分析及牛MX1蛋白对BVDV复制的影响[D].杨凌:西北农林科技大学, 2020. LIU C.Transcriptome analysis of MDBK cells infected with BVDV and the effect of bovine MX1 protein on BVDV replication[D].Yangling:Northwest A&F University, 2020.(in Chinese)
[28] CHANG A, CHEN Y, SHEN W, et al. IFIT1 protects against lipopolysaccharide and D-galactosamine-induced fatal hepatitis by inhibiting activation of the JNK pathway[J]. The Journal of Infectious Diseases, 2015, 212(9):1509-1520.
[29] 晏利琼, 侯永清, 王蕾, 等.N-乙酰半胱氨酸对病毒感染IPEC-1细胞相关基因mRNA水平的影响[A].中国畜牧兽医学会动物营养学分会第十二次动物营养学术研讨会[C].2016. YAN L Q, HOU Y Q, WANG L, et al. Effect of N-acetylcysteine on mRNA levels of related genes in IPEC-1 cells infected by virus[A].The Twelfth Symposium on Animal Nutrition of the Animal Nutrition Branch of the Chinese Society of Animal Husbandry and Veterinary Medicine[C].2016.(in Chinese)