饲粮精氨酸水平对肉仔鸡免疫功能及其抗FAdV-4影响的研究

doi:10.11843/j.issn.0366-6964.2020.04.013

摘要/Abstract

摘要： 本试验旨在研究饲粮精氨酸水平对肉仔鸡免疫功能及其抗禽腺病毒I群4型（fowl adenovirus serotype 4，FAdV-4）的影响。选取1日龄罗斯（Ross）308肉仔鸡300只，随机分为5组（每组5个重复，每个重复12只），设置5个饲粮精氨酸水平组，分别为0.96%、1.20%、1.44%、1.68%和1.92%，试验鸡饲养至21日龄，各重复随机选取4只测定血清免疫指标和免疫相对器官指数。随后各重复再次随机选取6只，并分为感染组和对照组，感染组肌肉注射FAdV-4 NP毒株0.2 mL（TCID₅₀=10^-6.23/0.01 mL），对照组肌肉注射等量灭菌生理盐水，感染2 d后，统计死亡率、发病率，ELISA测定肝组织iNOS水平、qPCR测定肝iNOS mRNA表达量及病毒载量。结果表明：1）饲粮精氨酸水平为1.44%、1.68%和1.92%时，均能显著提高脾脏指数（P<0.05）且精氨酸水平为1.68%能显著提高胸腺指数（P<0.05），而精氨酸水平为0.96%会显著降低脾脏、胸腺和法氏囊指数（P<0.05）。2）1.68%精氨酸水平组能显著提高血清球蛋白（GLO）、IgG水平和球蛋白/白蛋白（G/A）比值（P<0.05）；精氨酸水平为0.96%显著降低血清球蛋白和IgG水平（P<0.05）。3）精氨酸水平为0.96%可显著降低肉仔鸡血清中IL-1β、TNF-α、iNOS和NO水平（P<0.05），而1.68%和1.92%精氨酸水平组均能显著提高肉仔鸡血清中IL-1β、TNF-α、IFN-γ和iNOS水平（P<0.05）。4）肉鸡感染FAdV-4后，0.96%精氨酸水平组的死亡率会显著高于1.20%、1.44%、1.68%和1.92%组（P<0.05）。5）ELISA测定结果显示，1.44%、1.68%和1.92%精氨酸水平组能显著提高感染后的肝组织iNOS水平。6）1.44%、1.68%和1.92%精氨酸水平组均能显著提高肝iNOS mRNA表达水平（P<0.05）；肉鸡感染FAdV-4，iNOS mRNA水平均会显著降低（P<0.05），1.44%、1.68%和1.92%精氨酸水平组能提高感染FAdV-4后肝iNOS mRNA水平（P<0.05）并显著降低肝的病毒载量（P<0.05）。由此可见，肉仔鸡饲粮中精氨酸的不足会降低其机体的免疫功能及抗FAdV-4病毒能力。提高肉仔鸡饲粮中精氨酸水平不仅能提高免疫器官指数、血清免疫因子水平及肝iNOS mRNA表达水平，降低肝的病毒载量，从而提高机体免疫功能和抗病毒能力。

关键词: 精氨酸, 免疫功能, iNOS, FAdV-4

Abstract: The aim of this study was to investigate the effects of dietary arginine levels on immune function in broilers and explore the mechanism of arginine anti-fowl adenovirus serotype 4 (FAdV-4). Three hundred one-day-old broilers(Ross 308) were randomly allocated to 5 groups with 5 replicates per group and 12 chickens per replicate. The experimental diets were set to 5 arginine levels of 0.96%, 1.20%, 1.44%, 1.68%, and 1.92%, respectively. The feeding trial lasted for 21 days. Four broilers were randomly selected from each replicate to determine serum immune index and immune relative organ index. Then, six broilers were randomly selected from each replicate and randomly divided into infected group and control group. The chickens in the infected group were individually infected with 0.2 mL of FAdV-4 NP strain (TCID₅₀=10^-6.23/0.01 mL) by intramuscular injection; the chickens of control group were individually injected with the same volume of sterile saline. Two days after infection, mortality rate and disease incidence were calculated. iNOS levels in livers were determined by ELISA. iNOS mRNA expression and viral load in livers were detected by qPCR. The results showed that:1) Three arginine levels of 1.44%, 1.68%, and 1.92% significantly increased spleen index (P<0.05) and arginine level of 1.68% significantly increased thymus index (P<0.05), while arginine level of 0.96% significantly reduced spleen index, thymus index and bursal index (P<0.05). 2) The arginine level of 1.68% significantly increased serum GLO, IgG levels and G/A value (P<0.05). Arginine level of 0.96% significantly decreased serum GLO and IgG levels (P<0.05). 3) The arginine level of 0.96% significantly decreased serum IL-1β, TNF-α, iNOS and NO levels (P<0.05), while the arginine levels of 1.68% and 1.92% significantly increased IL-1β, TNF-α, IFN-γ and iNOS (P<0.05). 4) After broilers infected with FAdV-4, the mortality rate in the 0.96% group was significantly higher than those of 1.20%, 1.44%, 1.68%, and 1.92% groups (P<0.05). 5) ELISA results showed that 1.44%, 1.68%, and 1.92% arginine levels could significantly increase iNOS levels in liver after infection. 6) The arginine levels of 1.44%, 1.68%, and 1.92% significantly increased the expression of iNOS mRNA in livers (P<0.05). After broiler were infected with FAdV-4, the level of iNOS mRNA in the livers were significantly decreased (P<0.05). High levels of arginine (1.44%, 1.68%, and 1.92%) increased the level of iNOS mRNA expression in the livers after broilers infected with FAdV-4 (P<0.05) and significantly reduced the viral load of the livers (P<0.05). It could be seen that deficiency of arginine in the diet could reduce the immune function and anti-FAdV-4 virus ability of the chicken. Increasing arginine levels in broiler diets not only improved immune organ index, serum immune factor levels, and iNOS mRNA expression levels in the livers, but also reduced viral load of livers, thereby improving immune function and antiviral capacity.

Key words: arginine, immune function, iNOS, FAdV-4

中图分类号:

S852.65

林智鑫, 周嘉鑫, 陈立圳, 高玉云, 王全溪, 王长康. 饲粮精氨酸水平对肉仔鸡免疫功能及其抗FAdV-4影响的研究[J]. 畜牧兽医学报, 2020, 51(4): 772-782.

LIN Zhixin, ZHOU Jiaxin, CHEN Lizhen, GAO Yuyun, WANG Quanxi, WANG Changkang. Effects of Dietary Arginine Levels on Immune Function and Anti-FAdV-4 Capacity of Broilers[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(4): 772-782.

参考文献 44

[1]	LUIKING Y C,POEZE M,RAMSAY G,et al.The role of arginine in infection and sepsis[J].J Parenter Enteral Nutr,2005,29(1S):S70-S74.
[2]	STECHMILLER J K,CHILDRESS B,COWAN L.Arginine supplementation and wound healing[J].Nutr Clin Pract,2005,20(1):52-61.
[3]	方勇军.精氨酸对肉鸭生长性能、免疫机能、胴体品质和血液脂质的影响[D].武汉:武汉工业学院,2009.FANG Y J.Influence of arginine on growth performance,immune action,carcass characteristics and blood lipid of meat duck[D].Wuhan:Wuhan Polytechnic University,2009.(in Chinese)
[4]	刘凤菊,呙于明,王磊.1~3周龄雌性肉仔鸡精氨酸需要量[J].动物营养学报,2011,23(4):571-577.LIU F J,GUO Y M,WANG L.Arginine requirement for female broiler chicks aged from 1 to 3 weeks[J].Chinese Journal of Animal Nutrition,2011,23(4):571-577.(in Chinese)
[5]	BOGDAN C.Nitric oxide and the immune response[J].Nat Immunol,2001,2(10):907-916.
[6]	BOGDAN C.The function of nitric oxide in the immune system[M]//MAYER B.Nitric Oxide.Berlin:Springer,2000:443-492.
[7]	CAPUANO G,RIGAMONTI N,GRIONI M,et al.Modulators of arginine metabolism support cancer immunosurveillance[J].BMC Immunol,2009,10(1):1.
[8]	OCHOA J B,STRANGE J,KEARNEY P,et al.Effects of L-arginine on the proliferation of T lymphocyte subpopulations[J].J Parenter Enteral Nutr,2001,25(1):23-29.
[9]	WU G Y,BAZER F W,DAVIS T A,et al.Arginine metabolism and nutrition in growth,health and disease[J].Amino Acids,2009,37(1):153-168.
[10]	JAHANIAN R.Immunological responses as affected by dietary protein and arginine concentrations in starting broiler chicks[J].Poult Sci,2009,88(9):1818-1824.
[11]	谭玲芳,王安,李越,等.精氨酸对笼养生长期蛋鸭生长性能及免疫功能的影响[J].中国饲料,2013(7):15-18.TAN L F,WANG A,LI Y,et al.Effects of dietary arginine level on growth performance and immunity of caged growing laying ducks[J].China Feed,2013(7):15-18.(in Chinese)
[12]	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.
[13]	XIA J,YAO K C,LIU Y Y,et al.Isolation and molecular characterization of prevalent fowl adenovirus strains in southwestern China during 2015-2016 for the development of a control strategy[J].Emerg Microbes Infect,2017,6(11):e103.
[14]	ZHAO J,RUAN S F,GUO Y,et al.Serological and phylogenetic analysis indicating prevalence of fowl adenovirus in China[J].Vet Rec,2018,182(13):381.
[15]	SCHACHNER A,MATOS M,GRAFL B,et al.Fowl adenovirus-induced diseases and strategies for their control-a review on the current global situation[J].Avian Pathol,2018,47(2):111-126.
[16]	GRIMBLE G K.Adverse gastrointestinal effects of arginine and related amino acids[J].J Nutr,2007,137(6):1693S-1701S.
[17]	WANG Q X,WU Y J,CAI Y L,et al.Spleen transcriptome profile of Muscovy ducklings in response to infection with Muscovy duck reovirus[J].Avian Dis,2015,59(2):282-290.
[18]	WANG Q X,LIU M X,XU L H,et al.Transcriptome analysis reveals the molecular mechanism of hepatic fat metabolism disorder caused by Muscovy duck reovirus infection[J].Avian Pathol,2018,47(2):127-139.
[19]	KWAK H,AUSTIC R,DIETERT R.Influence of dietary arginine concentration on lymphoid organ growth in chickens[J].Poult Sci,1999,78(11):1536-1541.
[20]	王恬.精氨酸对免疫机能的影响及调节作用[J].上海体育学院学报,2006,30(2):76-79.WANG T.Effect and regulatory action of arginine on immune function[J].Journal of Shanghai University of Sport,2006,30(2):76-79.(in Chinese)
[21]	NIRGIOTIS J G,HENNESSEY P J,ANDRASSY R J.The effects of an arginine-free enteral diet on wound healing and immune function in the postsurgical rat[J].J Pediatr Surg,1991,26(8):936-941.
[22]	MA Q Y,HOPER M,ANDERSON N,et al.Effect of supplemental L-arginine in a chemical-induced model of colorectal cancer[J].World J Surg,1996,20(8):1087-1091.
[23]	LEBIEN T W.Arginine:an unusual dietary requirement of pre-B lymphocytes?[J].J Clin Invest,2002,110(10):1411-1413.
[24]	李春松,戴晋军,李彪.日粮中添加禽免疫增强剂对肉鸡免疫功能的影响[J].国外畜牧学-猪与禽,2012,32(9):73-75.LI C S,DAI J J,LI B.Effects of dietary addition of avian immune enhancer on immune function of broilers[J].Pigs and Poultry,2012,32(9):73-75.(in Chinese)
[25]	翟明,李妍,李岩.多肽菌素对肉鸡血清生化指标、血清抗体水平及免疫器官指数的影响[J].中国家禽,2018,40(7):23-26.ZHAI M,LI Y,LI Y.Effects of polypeptide on serum biochemical index,serum antibody level and immune organ index of broilers[J].China Poultry,2018,40(7):23-26.(in Chinese)
[26]	柏美娟,孔祥峰,印遇龙,等.日粮添加精氨酸对肥育猪免疫功能的调节作用[J].扬州大学学报:农业与生命科学版,2009,30(3):45-49.BAI M J,KONG X F,YIN Y L,et al.Regulation of dietary supplemental arginine on immune response in fattening pigs[J].Journal of Yangzhou University:Agricultural and Life Science Edition,2009,30(3):45-49.(in Chinese)
[27]	陈亚军,齐玉梅.精氨酸免疫营养作用的研究进展[J].中国临床营养杂志,2007,15(5):310-314.CHEN Y J,QI Y M.Research advancement of arginine in immunonutrition[J].Chinese Journal of Clinical Nutrition,2007,15(5):310-314.(in Chinese)
[28]	李玉庆,张小平,程爱国.精氨酸的免疫调理作用[J].中国煤炭工业医学杂志,2007,10(2):121-122.LI Y Q,ZHANG X P,CHENG A G.Immune conditioning of arginine[J].Chinese Journal of Coal Industry Medicine,2007,10(2):121-122.(in Chinese)
[29]	MAYER B,HEMMENS B.Biosynthesis and action of nitric oxide in mammalian cells[J].Trends Biochem Sci,1997,22(12):477-481.
[30]	ALDERTON W K,COOPER C E,KNOWLES R G.Nitric oxide synthases:structure,function and inhibition[J].Biochem J,2001,357(3):593-615.
[31]	郭祎玮.精氨酸对肉仔鸡生长性能和免疫功能的影响及其机理研究[D].呼和浩特:内蒙古农业大学,2014.GUO Y W.Effects of arginine on growth performance and immune function in broilers and the underlying mechanism[D].Hohhot:Inner Mongolia Agricultural University,2014.(in Chinese)
[32]	韩杰.精氨酸对脂多糖和环磷酰胺刺激仔猪免疫功能的影响[D].武汉:武汉工业学院,2009.HAN J.Effect of arginine on immune function of piglets after lipopolysaccharide and cyclophosphamide challenge[D].Wuhan:Wuhan Polytechnic University,2009.(in Chinese)
[33]	KIRK S J,REGAN M C,BARBUL A.Cloned murine T lymphocytes synthesize a molecule with the biological characteristics of nitric oxide[J].Biochem Biophys Res Commun,1990,173(2):660-665.
[34]	WIEST R,GROSZMANN R J.The paradox of nitric oxide in cirrhosis and portal hypertension:too much,not enough[J].Hepatology,2002,35(2):478-491.
[35]	FENG D Y,CHENG R X,ZHENG H,et al.Effect of hepatitis C virus NS3 protein on expression of iNOS mRNA in hepatocarcinogenesis[J].Bulletin of Hunan Medical University,2001,26(3):192-194.
[36]	TACHE D E,STǍNCIULESCU C E,BANIŢǍ I M,et al.Inducible nitric oxide synthase expression (iNOS) in chronic viral hepatitis and its correlation with liver fibrosis[J].Rom J Morphol Embryol,2014,55(S2):539-543.
[37]	FU P,ZHANG Q,YI D Y,et al.Dehydrocavidine attenuates D-galactose induced learning and memory impairment in rats[J].Neurosci Lett,2018,665:200-205.
[38]	LOWENSTEIN C J,HILL S L,LAFOND-WALKER A,et al.Nitric oxide inhibits viral replication in murine myocarditis[J].J Clin Invest,1996,97(8):1837-1843.
[39]	MANNICK J B.The antiviral role of nitric oxide[J].Res Immunol,1995,146(9):693-697.
[40]	MANNICK J B,ASANO K,IZUMI K,et al.Nitric oxide produced by human B lymphocytes inhibits apoptosis and Epstein-Barr virus reactivation[J].Cell,1994,79(7):1137-1146.
[41]	SHARARA A I,PERKINS D J,MISUKONIS M A,et al.Interferon (IFN)-α activation of human blood mononuclear cells in vitro and in vivo for nitric oxide synthase (NOS) type 2 mRNA and protein expression:possible relationship of induced NOS2 to the anti-hepatitis C effects of IFN-α in vivo[J].J Exp Med,1997,186(9):1495-1502.
[42]	MUNIR K,MUNEER M A,MASAOUD E,et al.Dietary arginine stimulates humoral and cell-mediated immunity in chickens vaccinated and challenged against hydropericardium syndrome virus[J].Poult Sci,2009,88(8):1629-1638.
[43]	LEE J E,AUSTIC R E,NAQI S A,et al.Dietary arginine intake alters avian leukocyte population distribution during infectious bronchitis challenge[J].Poult Sci,2002,81(6):793-798.
[44]	谭建庄.日粮精氨酸对不同疾病模型肉鸡免疫功能的调节作用与机理研究[D].北京:中国农业大学,2014.TAN J Z.Effects of dietary L-Arginine supplementation on immune functions in broiler chickens challenged with different pathogens[D].Beijing:China Agricultural University,2014.(in Chinese)