猴头菇多糖对MDRV感染雏番鸭主要免疫器官细胞凋亡及血清免疫相关指标的影响
|
摘要
为探讨猴头菇多糖(HEP)对番鸭呼肠孤病毒(MDRV)感染番鸭主要免疫器官的细胞凋亡及免疫功能的影响,本研究将160只1日龄MDRV抗原、抗体阴性雏番鸭随机平均分为四组:空白对照组(NCG)、同居感染MDRV对照组(CICG)、HEP饮水对照组(HCG)和HEP饮水给药预防MDRV感染组(HPG)。四组试验雏番鸭分别于同居感染后第2、6、9、12天每组随机抽取5只鸭无菌采取肝脏、脾脏、胸腺和法氏囊,将采集的样品制成病理组织切片,并用原位末端标记法(TUNEL法)观察并计算凋亡指数,以及用免疫组织化学染色法检测Fas-L蛋白;同居感染后第12天,每组抽取10只番鸭心脏无菌采血分离血清,测定总抗氧化能力(T-AOC)、丙二醛(MDA)、IgG、IgM、IgA和补体C3、C4。结果显示:HPG组在MDRV感染前期(感染后第2～6天)鸭肝脏、脾脏、胸腺和法氏囊器官的凋亡率极显著(P<0.01)高于CICG组,而病毒感染后期(感染后第9～12天)则相反,同居感染后第12天血清各项检测指标值介于非病毒感染组与CICG组之间,其中T-AOC极显著(P<0.01)高于CICG组,而MDA含量极显著(P<0.01)低于CICG组,血清TP、ALB、GLO、IgA、IgM、IgG、C3、C4含量均显著(P<0.05)或极显著(P<0.01)高于CICG组。结果表明,HEP饮水预防用药可促进MDRV感染早期雏番鸭肝脏和主要免疫器官细胞凋亡,抑制病毒感染中后期的细胞凋亡,减少组织器官损伤,维持较正常的抗氧化能力,提高血清蛋白、抗体及补体含量,有效缓解MDRV感染导致的免疫抑制。
To investigate the effects of Hericium erinaceus polysaccharide(HEP) on apoptosis and immune function of Muscovy duck reovirus(MDRV)-infected Muscovy duck's main immune organs,one hundred and sixty one-day-old healthy Muscovy ducklings were divided randomly into four groups including normal control group(NCG),cohabitation infection control group(CICG),HEP control group(HCG) and HEP prevention group(HPG).Five Muscovy ducklings each group were respectively sacrificed on day 2,6,9 and12 post-infection(dpi) for samples collection and following detection,the liver,spleen,thymus and bursa each group were prepared for pathological tissue sections.The apoptotic index was observed and calculated by TUNEL method,and Fas-L protein was detected by immunohistochemical staining.Ten ducks were randomly selected from each group for the detection of total antioxidant capacity(T-AOC),malondialdehyde(MDA),IgG,IgM,IgA and complements C3 and C4 on day 12 post-infection.In result,the apoptosis rate of the liver,spleen,thymus and bursa of the HPG group was significantly increased(P<0.01) than the CICG group in the early stage of MDRV infection(2—6 dpi),but late in viral infection(9-12 dpi) were opposite.The serum values of the HPG group were between the NCG group and the CICG group on day 12 post-infection,and the T-AOC was extremely significant(P <0.01) increased than the CICG group,while the MDA content was extremely significant(P<0.01) decreased than the CICG group.TP,ALB,GLO,IgA,IgM,IgG,C3 and C4 levels of serum in the HPG group were significantly(P<0.05) or extremely significant(P<0.01) higher than the CICG group.The results showed that HEP can promote apoptosis of immune organs of Muscovy ducklings in early MDRV infection stage,while inhibiting cell apoptosis,inhibiting viral replication,reducing tissue and organ damage,maintaining normal antioxidant capacity,increasing serum pro tein,anti body and com plement levels,and relieving immunosuppression caused by MDRV infection in the middle and late stages of viral infection.
To investigate the effects of Hericium erinaceus polysaccharide(HEP) on apoptosis and immune function of Muscovy duck reovirus(MDRV)-infected Muscovy duck's main immune organs,one hundred and sixty one-day-old healthy Muscovy ducklings were divided randomly into four groups including normal control group(NCG),cohabitation infection control group(CICG),HEP control group(HCG) and HEP prevention group(HPG).Five Muscovy ducklings each group were respectively sacrificed on day 2,6,9 and12 post-infection(dpi) for samples collection and following detection,the liver,spleen,thymus and bursa each group were prepared for pathological tissue sections.The apoptotic index was observed and calculated by TUNEL method,and Fas-L protein was detected by immunohistochemical staining.Ten ducks were randomly selected from each group for the detection of total antioxidant capacity(T-AOC),malondialdehyde(MDA),IgG,IgM,IgA and complements C3 and C4 on day 12 post-infection.In result,the apoptosis rate of the liver,spleen,thymus and bursa of the HPG group was significantly increased(P<0.01) than the CICG group in the early stage of MDRV infection(2—6 dpi),but late in viral infection(9-12 dpi) were opposite.The serum values of the HPG group were between the NCG group and the CICG group on day 12 post-infection,and the T-AOC was extremely significant(P <0.01) increased than the CICG group,while the MDA content was extremely significant(P<0.01) decreased than the CICG group.TP,ALB,GLO,IgA,IgM,IgG,C3 and C4 levels of serum in the HPG group were significantly(P<0.05) or extremely significant(P<0.01) higher than the CICG group.The results showed that HEP can promote apoptosis of immune organs of Muscovy ducklings in early MDRV infection stage,while inhibiting cell apoptosis,inhibiting viral replication,reducing tissue and organ damage,maintaining normal antioxidant capacity,increasing serum pro tein,anti body and com plement levels,and relieving immunosuppression caused by MDRV infection in the middle and late stages of viral infection.
引文
[1]陈强,张英扬,蔡栋灵,等.番鸭呼肠孤病毒YB株?B蛋白的原核表达与生物信息学分析[J].中国兽医科学,2018.48(05).605-615.CHEN Qiang,ZHANG Ying-yang,CAI Dong-ling,et al.Prokaryotic expression and bioinformatic analysis of?B gene of Muscovy duck reovirus YB strain[J].Chinese Veterinary Science,2018.48(05).605-615.(in Chinese)
[2]吴宝成,姚金水,陈家祥,等.呼肠孤病毒B3分离株感染番鸭的病理组织学研究[J].福建农林大学学报(自然版),2001,30(4):514-517.WU Bao-cheng,YAO Jin-shui,CHEN Jia-xiang,et al.Pathology of infection with reovirus isolate B3 in Muscovy ducks[J].Journal of Fujian Agricultural University(NaturaI Science Edition),2001,30(4):514-517.(in Chinese)
[3]姚金水,吴宝成,陈文列,等.禽(番鸭)呼肠孤病毒诱导脾脏单核——巨噬细胞和淋巴细胞凋亡的超微观察[J].福建农林大学学报(自然版),2004,33(4):501-504.YAO Jin-shui,WU Bao-cheng,CHEN Wen-lie,et al.Ultrastructural observation on spleen cell,mononuclear macrophage and lymphocytes incurring apoptosis induced by avian(Muscovy duck)reovirus[J].JournaI of Fujian Agriculture and Forestry University(Natural Science Edition),2004,33(4):501-504.(in Chinese)
[4]王全溪,吴宝成,李国平.番鸭呼肠孤病毒诱导免疫抑制机制的初步研究[J].中国兽医科学,2009,39(09):769-773.WANG Quan-xi,WU Bao-cheng,LI Guo ping.Mechanism of immunosuppression induced by Muscovy duck reovirus[J].Chinese Veterinary Science,2009,39(09):769-773.(in Chinese)
[5]陈少莺,胡奇林,程晓霞,等.番鸭呼肠孤病毒病雏番鸭实质器官的超微结构[J].中国兽医学报,2006,26(6):662-664.CHEN Shao-ying,HU Qi-lin,CHENG Xiao-xia,et al.Ultrastructure observation of Muscovy ducks infected with Muscovy duck reovirus[J].Chinese Journal of Veterinary Science,2006,26(6):662-664.(in Chinese)
[6]徐杭民,谢志慧,张蔚羽,等.猴头菇多糖的免疫调节作用[J].中国中西医结合杂志,1994(7):427-428.XU Hang-min,XIE Zhi-hui,ZHANG Wei-yu,et al.Immunomodulatory function of Hericium erinaceus polysaccharide[J].Chinese Journal of Integrated Traditional and Western Medicine,1994(7):427-428.(in Chinese)
[7] HAN Z H,YE J M,WANG G F.Evaluation of in vivo antioxidant activity of Hericium erinaceus polysaccharides[J].International Journal of Biological Macromolecules,2013,(52):66-71.
[8]罗珍,黄萍,郭重仪,等.猴头菇多糖增强免疫功能的实验研究[J].中国实验方剂学杂志,2011,17(4):182-183.LUO Zhen,HUANG Ping,GUO Chong-yi,et al.Experimental study on immune function enhancemet of Hericium erinaceus polysaccharide[J].Chinese Journal of Experimental Traditional Medical Formulae,2011,17(4):182-183.(in Chinese)
[9]孙雪宁,陈景杰,黄诗杭,等.猴头菇多糖对MDRV感染雏番鸭肠道修复作用与MAdCAM-1分泌量关系的研究[J].中国家禽,2016,38(9):24-28.SUN Xue-ning,CHEN Jing-jie,HUANG Shi-hang,et al.Relationship between intestinal tract restoration by Hericium erinaceus polysaccharide and MAdCAM-1 secretion in MDRV-infected muscovy ducklings[J].China Poultry,2016,38(9):24-28.(in Chinese)
[10]陈新瑶,张建龙,董星,等.猴头菇多糖对氧化应激的IPEC-J2细胞抗氧化能力及紧密连接蛋白ZO-1的影响[J].畜牧兽医学报,2017,48(9):1769-1776.CHEN Xin-yao,ZHANG Jian-long,DONG Xing,et al.Effect of Hericium erinaceus polysaccharide on antioxidant ability and ZO-1 expression in IPEC-J2cells under oxidative stress[J].Acta Veterinaria et Zootechnica Sinica,2017,48(9):1769-1776.(in Chinese)
[11]吴异健.两种中药多糖对呼肠孤病毒感染番鸭的免疫调节作用[D].福州:福建农林大学,2010.WU Yi-jian.Immunoregulating effects of two chinese herbal polysaccharides on Muscovy duck infecting of MDRV[D].Fuzhou:Fujian Agriculture and Forestry University,2010.(in Chinese)
[12] WANG Q,LIU M,XU L,et al.Transcriptome analysis reveals the molecular mechanism of hepatic fat metabolism disorder caused by Muscovy duck reovirus infection[J].Avian Pathology,2018,47(2):127-139.
[13]陈少莺,陈仕龙,林锋强,等.新型鸭呼肠孤病毒的分离与鉴定[J].病毒学报,2012,28(3):224-230.CHEN Shao-ying,CHEN Shi-long,LIN Feng-qiang,et al.The isolation and identification of novel duck reovirus[J].Chinese Journal of Virology,2012,28(3):224-230.(in Chinese)
[14]陈宗艳,朱英奇,王世传,等.一株新型鸭源呼肠孤病毒(TH11株)的分离与鉴定[J].中国动物传染病学报,2012,20(1):10-15.CHEN Zong-yan,ZHU Ying-qi,WANG Shi-chuan,et al.Isolation and characterization of a new type of duck reovirus[J].Chinese Journal of Animal Infectious Diseases,2012,20(1):10-15.(in Chinese)
[15] MA G,WANG D,SHI J,et al.Complete genomic sequence of a reovirus isolate from Pekin ducklings in China[J].Journal of Virology,2012,86(23):13137.
[16] YUN T,YU B,NI Z,et al.Genomic characteristics of a novel reovirus from Muscovy duckling in China[J].Veterinary Microbiology,2014,168(2-4):261-271.
[17]包汉勋,祁保民,尚迎辉.免疫组化法检测番鸭呼肠孤病毒在番鸭体内的分布[J].中国畜牧兽医,2009,36(4):43-46.BAO Han-xun,QI Bao-min,SHANG Ying-hui.Distribution of Muscovy duck reovirus in muscovy ducks detected by immunohistochemical method[J].China Animal Husbandry and Veterinary Medicine,2009,36(4):43-46.(in Chinese)
[18]于可响,韩宏宇,刘存霞,等.绿头野鸭新型呼肠孤病毒的分离鉴定[J].中国家禽,2014,36(13):50-52.YU Ke-xiang,HAN Hong-yu,LIU Cun-xia,et al.Isolation and identification of a new reovirus from mallard[J].China Poultry,2014,36(13):50-52.(in Chinese)
[19]陈艺娟,吴异健,黄一帆,等.猴头菇多糖对呼肠孤病毒感染番鸭组织病变及细胞凋亡的影响[J].中国预防兽医学报,2010,32(1):14-18.CHEN Yi-juan,WU Yi-jian,HUANG Yi-fang,et al.Effects of Hericium erinaceus polysaccharide on reovirus-induced tissue necrosis and apoptosis in Muscovy ducks[J].Chinese Journal of Preventive Veterinary Medicine,2010,32(1):14-18.(in Chinese)
[20]黄丽娜.猴头菇多糖对MDRV致番鸭肠黏膜免疫抑制的调节作用及其机制[D].福州:福建农林大学,2015.HUANG Li-na.The regulation effect and mechanism of Hericium erinaceus polysaccharide on intestinal mucosal immune suppression of Muscovy ducks induced by reovirus[D].Fuzhou:Fujian Agriculture and Forestry University,2015.(in Chinese)
[21]姜慧慧.HPS对MDRV感染番鸭肠黏膜免疫屏障功能损伤的修复及免疫抑制的调节作用[D].福州:福建农林大学,2015.JIANG Hui-hui.The repairability and regulation of HEP on intestinal mucosal barrier dysfunction and immune suppression in MDRV-infected Muscovy ducks[D].Fuzhou:Fujian Agriculture and Forestry University,2015.(in Chinese)
[22]伊恩·蒂萨德(美).兽医免疫学[M].8版.北京:中国农业出版社,2012:158-168.Tizard I R(USA).Veterinary Immunology[M].8th ed.Beijing:China Agricultural Press,2012:158-168.(in Chinese)
[23]金伯泉.医学免疫学[M].5版.北京:人民卫生出版社,2008:49-59.JIN Bo-quan.Medical Immunology[M].5th ed.Beijing:People's Medical Publishing House,2008:49-59.(in Chinese)
[2]吴宝成,姚金水,陈家祥,等.呼肠孤病毒B3分离株感染番鸭的病理组织学研究[J].福建农林大学学报(自然版),2001,30(4):514-517.WU Bao-cheng,YAO Jin-shui,CHEN Jia-xiang,et al.Pathology of infection with reovirus isolate B3 in Muscovy ducks[J].Journal of Fujian Agricultural University(NaturaI Science Edition),2001,30(4):514-517.(in Chinese)
[3]姚金水,吴宝成,陈文列,等.禽(番鸭)呼肠孤病毒诱导脾脏单核——巨噬细胞和淋巴细胞凋亡的超微观察[J].福建农林大学学报(自然版),2004,33(4):501-504.YAO Jin-shui,WU Bao-cheng,CHEN Wen-lie,et al.Ultrastructural observation on spleen cell,mononuclear macrophage and lymphocytes incurring apoptosis induced by avian(Muscovy duck)reovirus[J].JournaI of Fujian Agriculture and Forestry University(Natural Science Edition),2004,33(4):501-504.(in Chinese)
[4]王全溪,吴宝成,李国平.番鸭呼肠孤病毒诱导免疫抑制机制的初步研究[J].中国兽医科学,2009,39(09):769-773.WANG Quan-xi,WU Bao-cheng,LI Guo ping.Mechanism of immunosuppression induced by Muscovy duck reovirus[J].Chinese Veterinary Science,2009,39(09):769-773.(in Chinese)
[5]陈少莺,胡奇林,程晓霞,等.番鸭呼肠孤病毒病雏番鸭实质器官的超微结构[J].中国兽医学报,2006,26(6):662-664.CHEN Shao-ying,HU Qi-lin,CHENG Xiao-xia,et al.Ultrastructure observation of Muscovy ducks infected with Muscovy duck reovirus[J].Chinese Journal of Veterinary Science,2006,26(6):662-664.(in Chinese)
[6]徐杭民,谢志慧,张蔚羽,等.猴头菇多糖的免疫调节作用[J].中国中西医结合杂志,1994(7):427-428.XU Hang-min,XIE Zhi-hui,ZHANG Wei-yu,et al.Immunomodulatory function of Hericium erinaceus polysaccharide[J].Chinese Journal of Integrated Traditional and Western Medicine,1994(7):427-428.(in Chinese)
[7] HAN Z H,YE J M,WANG G F.Evaluation of in vivo antioxidant activity of Hericium erinaceus polysaccharides[J].International Journal of Biological Macromolecules,2013,(52):66-71.
[8]罗珍,黄萍,郭重仪,等.猴头菇多糖增强免疫功能的实验研究[J].中国实验方剂学杂志,2011,17(4):182-183.LUO Zhen,HUANG Ping,GUO Chong-yi,et al.Experimental study on immune function enhancemet of Hericium erinaceus polysaccharide[J].Chinese Journal of Experimental Traditional Medical Formulae,2011,17(4):182-183.(in Chinese)
[9]孙雪宁,陈景杰,黄诗杭,等.猴头菇多糖对MDRV感染雏番鸭肠道修复作用与MAdCAM-1分泌量关系的研究[J].中国家禽,2016,38(9):24-28.SUN Xue-ning,CHEN Jing-jie,HUANG Shi-hang,et al.Relationship between intestinal tract restoration by Hericium erinaceus polysaccharide and MAdCAM-1 secretion in MDRV-infected muscovy ducklings[J].China Poultry,2016,38(9):24-28.(in Chinese)
[10]陈新瑶,张建龙,董星,等.猴头菇多糖对氧化应激的IPEC-J2细胞抗氧化能力及紧密连接蛋白ZO-1的影响[J].畜牧兽医学报,2017,48(9):1769-1776.CHEN Xin-yao,ZHANG Jian-long,DONG Xing,et al.Effect of Hericium erinaceus polysaccharide on antioxidant ability and ZO-1 expression in IPEC-J2cells under oxidative stress[J].Acta Veterinaria et Zootechnica Sinica,2017,48(9):1769-1776.(in Chinese)
[11]吴异健.两种中药多糖对呼肠孤病毒感染番鸭的免疫调节作用[D].福州:福建农林大学,2010.WU Yi-jian.Immunoregulating effects of two chinese herbal polysaccharides on Muscovy duck infecting of MDRV[D].Fuzhou:Fujian Agriculture and Forestry University,2010.(in Chinese)
[12] WANG Q,LIU M,XU L,et al.Transcriptome analysis reveals the molecular mechanism of hepatic fat metabolism disorder caused by Muscovy duck reovirus infection[J].Avian Pathology,2018,47(2):127-139.
[13]陈少莺,陈仕龙,林锋强,等.新型鸭呼肠孤病毒的分离与鉴定[J].病毒学报,2012,28(3):224-230.CHEN Shao-ying,CHEN Shi-long,LIN Feng-qiang,et al.The isolation and identification of novel duck reovirus[J].Chinese Journal of Virology,2012,28(3):224-230.(in Chinese)
[14]陈宗艳,朱英奇,王世传,等.一株新型鸭源呼肠孤病毒(TH11株)的分离与鉴定[J].中国动物传染病学报,2012,20(1):10-15.CHEN Zong-yan,ZHU Ying-qi,WANG Shi-chuan,et al.Isolation and characterization of a new type of duck reovirus[J].Chinese Journal of Animal Infectious Diseases,2012,20(1):10-15.(in Chinese)
[15] MA G,WANG D,SHI J,et al.Complete genomic sequence of a reovirus isolate from Pekin ducklings in China[J].Journal of Virology,2012,86(23):13137.
[16] YUN T,YU B,NI Z,et al.Genomic characteristics of a novel reovirus from Muscovy duckling in China[J].Veterinary Microbiology,2014,168(2-4):261-271.
[17]包汉勋,祁保民,尚迎辉.免疫组化法检测番鸭呼肠孤病毒在番鸭体内的分布[J].中国畜牧兽医,2009,36(4):43-46.BAO Han-xun,QI Bao-min,SHANG Ying-hui.Distribution of Muscovy duck reovirus in muscovy ducks detected by immunohistochemical method[J].China Animal Husbandry and Veterinary Medicine,2009,36(4):43-46.(in Chinese)
[18]于可响,韩宏宇,刘存霞,等.绿头野鸭新型呼肠孤病毒的分离鉴定[J].中国家禽,2014,36(13):50-52.YU Ke-xiang,HAN Hong-yu,LIU Cun-xia,et al.Isolation and identification of a new reovirus from mallard[J].China Poultry,2014,36(13):50-52.(in Chinese)
[19]陈艺娟,吴异健,黄一帆,等.猴头菇多糖对呼肠孤病毒感染番鸭组织病变及细胞凋亡的影响[J].中国预防兽医学报,2010,32(1):14-18.CHEN Yi-juan,WU Yi-jian,HUANG Yi-fang,et al.Effects of Hericium erinaceus polysaccharide on reovirus-induced tissue necrosis and apoptosis in Muscovy ducks[J].Chinese Journal of Preventive Veterinary Medicine,2010,32(1):14-18.(in Chinese)
[20]黄丽娜.猴头菇多糖对MDRV致番鸭肠黏膜免疫抑制的调节作用及其机制[D].福州:福建农林大学,2015.HUANG Li-na.The regulation effect and mechanism of Hericium erinaceus polysaccharide on intestinal mucosal immune suppression of Muscovy ducks induced by reovirus[D].Fuzhou:Fujian Agriculture and Forestry University,2015.(in Chinese)
[21]姜慧慧.HPS对MDRV感染番鸭肠黏膜免疫屏障功能损伤的修复及免疫抑制的调节作用[D].福州:福建农林大学,2015.JIANG Hui-hui.The repairability and regulation of HEP on intestinal mucosal barrier dysfunction and immune suppression in MDRV-infected Muscovy ducks[D].Fuzhou:Fujian Agriculture and Forestry University,2015.(in Chinese)
[22]伊恩·蒂萨德(美).兽医免疫学[M].8版.北京:中国农业出版社,2012:158-168.Tizard I R(USA).Veterinary Immunology[M].8th ed.Beijing:China Agricultural Press,2012:158-168.(in Chinese)
[23]金伯泉.医学免疫学[M].5版.北京:人民卫生出版社,2008:49-59.JIN Bo-quan.Medical Immunology[M].5th ed.Beijing:People's Medical Publishing House,2008:49-59.(in Chinese)