多糖-硒化多糖复方增强鸡免疫功能的比较研究
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摘要
[目的]验证多糖-硒化多糖复方的增强免疫活性,筛选出活性最强的复方,为研制新型免疫增强剂提供试验依据。[方法]在前期研究筛选出增强免疫活性较强的当归多糖(CAP)、党参多糖(CPP)、大蒜多糖(GP)、硒化当归多糖(sCAP)、硒化党参多糖(sCPP)和硒化大蒜多糖(s GP)的基础上,以其为组分药,按照多糖与硒化多糖质量比9∶1的比例组成CAP-sCPP、CAP-s GP、CPP-sCAP、CPP-s GP、GP-sCAP和GP-sCPP共6个复方,比较它们的体内/外增强免疫活性能力。体外试验,用MTT法测定6个复方对鸡外周血淋巴细胞增殖的影响;体内试验,进一步比较了筛选出的3个复方对新城疫苗免疫雏鸡淋巴细胞增殖,血清抗体效价,IL-2、IL-6和IFN-γ含量的影响。[结果]体外试验结果显示:单独或与植物血凝素(PHA)共同刺激时,6个复方均有不同程度的增强免疫活性,CAP-sCPP、GP-sCPP和CPP-sCAP的活性较强;体内试验结果显示:在免疫后7、14、21和28 d,GP-sCPP高剂量促进T淋巴细胞增殖的作用最强,血清抗体效价和IL-2、IL-6及IFN-γ含量均为最高,且均显著高于免疫对照组。[结论]GP-sCPP的增强免疫活性最强,可以考虑作为新型免疫增强剂的候选方。
[Objectives]In order to validate the immune-enhancing activity of polysaccharide-selenizing polysaccharide compounds,a compound with the strongest activity was selected and offer theoretical evidence for developing new-type immunopotentiators was offered. [Methods]On the basis of screening out the Chinese angelica polysaccharide( CAP),Codonopsis pilosula polysaccharide( CPP),garlic polysaccharide( GP),selenizing CAP( sCAP),selenizing CPP( sCPP) and selenizing GP( s GP) with stronger immuneenhancing activity in previous research,six polysaccharide-selenizing polysaccharide compounds,CAP-sCPP,CAP-s GP,CPP-sCAP,CPP-sGP,GP-sCAP and GP-sCPP,were composed by taking them as constituents according to 9 ∶ 1 ratio of polysaccharide to selenizing polysaccharide,and their immune-enhancing activities in vitro and in vivo were compared. In vitro test,the effects of6 compounds on chicken peripheral lymphocyte proliferation were determined by MTT method. In vivo test,the effects of3 compounds screened on lymphocyte proliferation,serum antibody titer,IL-2,IL-6 and IFN-γ contents were further compared in chickens vaccinated with Newcastle disease virus vaccine. [Results]The results in vitro test showed that in single or synergistic stimulation with phytohemagglutinin( PHA),all of six compounds possessed the immune-enhancing activities at different strength,and the activities of CAP-sCPP,GP-sCPP and CPP-sCAP were stronger. The results in vivo test showed that in GP-sCPP at high dose group,the actions of promoting T lymphocyte proliferation were strongest,and the serum antibody titer,IL-2,IL-6 and IFN-γcontents were the highest and significantly higher than those in vaccination control group at four time points after vaccination.[Conclusions]GP-sCPP possesses the strongest immune-enhancing activity and can be considered as a candidate of new-type immunopotentiators.
[Objectives]In order to validate the immune-enhancing activity of polysaccharide-selenizing polysaccharide compounds,a compound with the strongest activity was selected and offer theoretical evidence for developing new-type immunopotentiators was offered. [Methods]On the basis of screening out the Chinese angelica polysaccharide( CAP),Codonopsis pilosula polysaccharide( CPP),garlic polysaccharide( GP),selenizing CAP( sCAP),selenizing CPP( sCPP) and selenizing GP( s GP) with stronger immuneenhancing activity in previous research,six polysaccharide-selenizing polysaccharide compounds,CAP-sCPP,CAP-s GP,CPP-sCAP,CPP-sGP,GP-sCAP and GP-sCPP,were composed by taking them as constituents according to 9 ∶ 1 ratio of polysaccharide to selenizing polysaccharide,and their immune-enhancing activities in vitro and in vivo were compared. In vitro test,the effects of6 compounds on chicken peripheral lymphocyte proliferation were determined by MTT method. In vivo test,the effects of3 compounds screened on lymphocyte proliferation,serum antibody titer,IL-2,IL-6 and IFN-γ contents were further compared in chickens vaccinated with Newcastle disease virus vaccine. [Results]The results in vitro test showed that in single or synergistic stimulation with phytohemagglutinin( PHA),all of six compounds possessed the immune-enhancing activities at different strength,and the activities of CAP-sCPP,GP-sCPP and CPP-sCAP were stronger. The results in vivo test showed that in GP-sCPP at high dose group,the actions of promoting T lymphocyte proliferation were strongest,and the serum antibody titer,IL-2,IL-6 and IFN-γcontents were the highest and significantly higher than those in vaccination control group at four time points after vaccination.[Conclusions]GP-sCPP possesses the strongest immune-enhancing activity and can be considered as a candidate of new-type immunopotentiators.
引文
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[6]贾江滨,刘静,谭亚非.大蒜化学成分研究进展[J].广东药学,1999,9(1):1-5.Jia J B,Liu J,Tan Y F.Research progress of chemical constituents of garlic[J].Guangdong Pharmacy,1999,9(1):1-5(in Chinese).
[7]Gao Z Z,Chen J,Qiu S L,et al.Optimization of selenylation modification for garlic polysaccharide based on immune-enhancing activity[J].Carbohydrate Polymers,2016,136(7):560-569.
[8]Qin T,Chen J,Wang D Y,et al.Optimization of selenylation conditions for Chinese angelica polysaccharide based on immune-enhancing activity[J].Carbohydrate Polymers,2013,92(1):645-650.
[9]Qiu S L,Chen J,Qin T,et al.Effects of selenylation modification on immune-enhancing activity of garlic polysaccharide[J].PLo S ONE,2014,9(1):e86377.
[10]秦韬.硒化当归多糖和硒化党参多糖的增强免疫和抗氧化的活性研究[D].南京:南京农业大学,2013.Qin T.Immune-enhancing and antioxidant activities of selenizing Chinese angelica polysaccharide and selenizing Codonopsis pilosula polysaccharide[D].Nanjing:Nanjing Agricultural University,2013(in Chinese with English abstract).
[11]高建忠,秦顺义,黄克和.氢化物发生-原子荧光光谱法测定富硒酵母中的有机硒和无机硒[J].分析科学学报,2006,22(2):157-160.Gao J Z,Qin S Y,Huang K H.Assay of organic selenium and inorganic selenium of enriched yeast by hydride generation atomic fluorescence spectrometry method[J].Journal of Analytical Science,2006,22(2):157-160(in Chinese with English abstract).
[12]刘丽,季辉,彭麟,等.鸡肝原代细胞药物代谢模型的建立与优化[J].南京农业大学学报,2015,38(1):127-133.DOI:10.7685/j.issn.1000-2030.2015.01.019.Liu L,Ji H,Peng L,et al.The establishment and optimization of the drug metabolic model of adult chicken primary hepatocytes[J].Journal of Nanjing Agricultural University,2015,38(1):127-133(in Chinese with English abstract).
[13]林锋强,胡奇林,陈少莺,等.MTT法检测番鸭外周血淋巴细胞转化功能的研究[J].动物医学进展,2005,26(1):75-77.Lin F Q,Hu Q L,Chen S Y,et al.Studies on MTT colorimetric assay for transformation of muscovy duck peripheral blood lymphocytes[J].Progress in Veterinary Medicine,2005,26(1):75-77(in Chinese with English abstract).
[14]Guo L W,Liu J G,Hu Y L,et al.Astragalus polysaccharide and sulfated epimedium polysaccharide synergistically resist the immunosuppression[J].Carbohydrate Polymers,2012,90(2):1055-1060.
[15]Kapczynski D R,Afonso C L,Miller P J.Immune responses of poultry to Newcastle disease virus[J].Developmental and Comparative Immunology,2013,41(3):447-453.
[16]Irie T,Watarai S,Kodama H.Humoral immune response of carp(Cyprinus carpio)induced by oral immunization with liposome-entrapped antigen[J].Developmental and Comparative Immunology,2003,27(5):413-421.
[17]邱妍,胡元亮,崔保安,等.怀牛膝多糖对雏鸡疫苗免疫效果的影响[J].畜牧兽医学报,2007,38(7):723-727.Qiu Y,Hu Y L,Cui B A,et al.Effects of achyranthes bidentate polysaccharide on immune efficacy of vaccine in chickens[J].Acta Veterinari et Zoontechnica Sinica,2007,38(7):723-727(in Chinese with English abstract).
[18]邹云,谢红兵,禹琪芳,等.植物多糖对断奶仔猪淋巴细胞增殖和细胞因子分泌的影响[J].动物营养学报,2014,26(1):210-218.Zou Y,Xie H B,Yu Q F,et al.Plant polysaccharides:effects on lymphocytes proliferation and cytokines secretion of weaner piglets[J].Chinese Journal of Animal Nutrition,2014,26(1):210-218(in Chinese with English abstract).
[19]李蕴玉,李佩国,张艳英,等.白细胞介素-2(IL-2)在兽医中的应用进展(综述)[J].河北科技师范学院学报,2006,20(4):63-66.Li W Y,Li P G,Zhang Y Y,et al.Applying progress on interleukin-2(IL-2)in veterinary medicine(summary)[J].Journal of Hebei Normal University of Science and Tachnology,2006,20(4):63-66(in Chinese with English abstract).
[20]Schroder K,Hertzog P J,Ravasi T.Interferon-gamma:an overview of signals,mechanisms and functions[J].Journal of Leukocyte Biology,2004,75(2):163-189.
[21]李悦,张昊,杨敏馨,等.淀粉液化芽孢杆菌对脂多糖刺激肉鸡生长性能、肠道形态及免疫功能的影响[J].南京农业大学学报,2016,39(4):624-631.DOI:10.7685/jnau.201504024.Li Y,Zhang H,Yang M X,et al,Effects of Bacillus amyloliquefaciens on growth performance,intestinal morphology and immune function of lipopolysaccharide-challenge broilers[J].Journal of Nanjing Agricultural University,2016,39(4):624-631(in Chinese with English abstract).
[22]Matthes T,Manfroi B,Huard B.Revisiting IL-6 antagonism in multiple myeloma[J].Critical Reviews in Oncology/Hematology,2016,105:1-4.
[23]中国兽药典委员会.中华人民共和国兽药典(2015年版二部)[M].北京:中国农业出版社,2016:34,416.Commission of Chinese Veterinary Pharmacopeia.Veterinary Pharmacopoeia of the People's Republic of China,2015 PartⅡ[M].Beijing:China Agriculture Press,2016:34,416(in Chinese).
[24]陈文霞,张培,高霞,等.硒化纹党参多糖和其抗A549细胞的活性[J].中成药,2015,37(11):2408-2413.Chen W X,Zhang P,Gao X,et al.Selenylation of polysaccharides from Codonopsis pilosula and its inhibition against A549 cells[J].Chinese Traditional Patent Medicine,2015,37(11):2408-2413(in Chinese with English abstract).
[25]Gao Z Z,Zhang C,Tian W J,et al.The antioxidative and hepatoprotective effects comparison of Chinese angelica polysaccharide(CAP)and selenizing CAP(s CAP)in CCl4induced hepatic injury mice[J].International Journal of Biological Macromolecules,2017,97:46-54.
[26]Qiu S L,Chen J,Chen X,et al.Optimization of selenylation conditions for lycium barbarum polysaccharide based on antioxidant activity[J].Carbohydrate Polymers,2014,103(1):148-153.
[2]李曦,张丽宏,王晓晓,等.当归化学成分及药理作用研究进展[J].中药材,2013,36(6):1023-1028.Li X,Zhang L H,Wang X X,et al.Research progress on chemical constituents and pharmacological effect of Angelica sinensis[J].Journal of Chinese Medicinal Materials,2013,36(6):1023-1028(in Chinese).
[3]温悦,傅正毅,赖艳,等.当归多糖的药理作用研究进展[J].中国医药导报,2012,9(30):27-29.Wen Y,Fu Z Y,Lai Y,et al.Research progress of pharmacological action of angelica polysaccharides[J].China Medical Herrald,2012,9(30):27-29(in Chinese with English abstract).
[4]孙政华,邵晶,郭玫.党参化学成分及药理作用研究进展[J].安徽农业科学,2015,43(33):174-176.Sun Z H,Shao J,Guo M.Research progress of Codonopsis pilosula chemical component and pharmacological effects[J].Journal of Anhui Agricultural Science,2015,43(33):174-176(in Chinese with English abstract).
[5]陈克克,王喆之.党参多糖的研究进展[J].现代生物医学进展,2007,7(4):635-637.Chen K K,Wang Z Z.Progerss in polysaeeharides from radix codonopsis[J].Progress in Modern Biomeidicine,2007,7(4):635-637(in Chinese with English abstract).
[6]贾江滨,刘静,谭亚非.大蒜化学成分研究进展[J].广东药学,1999,9(1):1-5.Jia J B,Liu J,Tan Y F.Research progress of chemical constituents of garlic[J].Guangdong Pharmacy,1999,9(1):1-5(in Chinese).
[7]Gao Z Z,Chen J,Qiu S L,et al.Optimization of selenylation modification for garlic polysaccharide based on immune-enhancing activity[J].Carbohydrate Polymers,2016,136(7):560-569.
[8]Qin T,Chen J,Wang D Y,et al.Optimization of selenylation conditions for Chinese angelica polysaccharide based on immune-enhancing activity[J].Carbohydrate Polymers,2013,92(1):645-650.
[9]Qiu S L,Chen J,Qin T,et al.Effects of selenylation modification on immune-enhancing activity of garlic polysaccharide[J].PLo S ONE,2014,9(1):e86377.
[10]秦韬.硒化当归多糖和硒化党参多糖的增强免疫和抗氧化的活性研究[D].南京:南京农业大学,2013.Qin T.Immune-enhancing and antioxidant activities of selenizing Chinese angelica polysaccharide and selenizing Codonopsis pilosula polysaccharide[D].Nanjing:Nanjing Agricultural University,2013(in Chinese with English abstract).
[11]高建忠,秦顺义,黄克和.氢化物发生-原子荧光光谱法测定富硒酵母中的有机硒和无机硒[J].分析科学学报,2006,22(2):157-160.Gao J Z,Qin S Y,Huang K H.Assay of organic selenium and inorganic selenium of enriched yeast by hydride generation atomic fluorescence spectrometry method[J].Journal of Analytical Science,2006,22(2):157-160(in Chinese with English abstract).
[12]刘丽,季辉,彭麟,等.鸡肝原代细胞药物代谢模型的建立与优化[J].南京农业大学学报,2015,38(1):127-133.DOI:10.7685/j.issn.1000-2030.2015.01.019.Liu L,Ji H,Peng L,et al.The establishment and optimization of the drug metabolic model of adult chicken primary hepatocytes[J].Journal of Nanjing Agricultural University,2015,38(1):127-133(in Chinese with English abstract).
[13]林锋强,胡奇林,陈少莺,等.MTT法检测番鸭外周血淋巴细胞转化功能的研究[J].动物医学进展,2005,26(1):75-77.Lin F Q,Hu Q L,Chen S Y,et al.Studies on MTT colorimetric assay for transformation of muscovy duck peripheral blood lymphocytes[J].Progress in Veterinary Medicine,2005,26(1):75-77(in Chinese with English abstract).
[14]Guo L W,Liu J G,Hu Y L,et al.Astragalus polysaccharide and sulfated epimedium polysaccharide synergistically resist the immunosuppression[J].Carbohydrate Polymers,2012,90(2):1055-1060.
[15]Kapczynski D R,Afonso C L,Miller P J.Immune responses of poultry to Newcastle disease virus[J].Developmental and Comparative Immunology,2013,41(3):447-453.
[16]Irie T,Watarai S,Kodama H.Humoral immune response of carp(Cyprinus carpio)induced by oral immunization with liposome-entrapped antigen[J].Developmental and Comparative Immunology,2003,27(5):413-421.
[17]邱妍,胡元亮,崔保安,等.怀牛膝多糖对雏鸡疫苗免疫效果的影响[J].畜牧兽医学报,2007,38(7):723-727.Qiu Y,Hu Y L,Cui B A,et al.Effects of achyranthes bidentate polysaccharide on immune efficacy of vaccine in chickens[J].Acta Veterinari et Zoontechnica Sinica,2007,38(7):723-727(in Chinese with English abstract).
[18]邹云,谢红兵,禹琪芳,等.植物多糖对断奶仔猪淋巴细胞增殖和细胞因子分泌的影响[J].动物营养学报,2014,26(1):210-218.Zou Y,Xie H B,Yu Q F,et al.Plant polysaccharides:effects on lymphocytes proliferation and cytokines secretion of weaner piglets[J].Chinese Journal of Animal Nutrition,2014,26(1):210-218(in Chinese with English abstract).
[19]李蕴玉,李佩国,张艳英,等.白细胞介素-2(IL-2)在兽医中的应用进展(综述)[J].河北科技师范学院学报,2006,20(4):63-66.Li W Y,Li P G,Zhang Y Y,et al.Applying progress on interleukin-2(IL-2)in veterinary medicine(summary)[J].Journal of Hebei Normal University of Science and Tachnology,2006,20(4):63-66(in Chinese with English abstract).
[20]Schroder K,Hertzog P J,Ravasi T.Interferon-gamma:an overview of signals,mechanisms and functions[J].Journal of Leukocyte Biology,2004,75(2):163-189.
[21]李悦,张昊,杨敏馨,等.淀粉液化芽孢杆菌对脂多糖刺激肉鸡生长性能、肠道形态及免疫功能的影响[J].南京农业大学学报,2016,39(4):624-631.DOI:10.7685/jnau.201504024.Li Y,Zhang H,Yang M X,et al,Effects of Bacillus amyloliquefaciens on growth performance,intestinal morphology and immune function of lipopolysaccharide-challenge broilers[J].Journal of Nanjing Agricultural University,2016,39(4):624-631(in Chinese with English abstract).
[22]Matthes T,Manfroi B,Huard B.Revisiting IL-6 antagonism in multiple myeloma[J].Critical Reviews in Oncology/Hematology,2016,105:1-4.
[23]中国兽药典委员会.中华人民共和国兽药典(2015年版二部)[M].北京:中国农业出版社,2016:34,416.Commission of Chinese Veterinary Pharmacopeia.Veterinary Pharmacopoeia of the People's Republic of China,2015 PartⅡ[M].Beijing:China Agriculture Press,2016:34,416(in Chinese).
[24]陈文霞,张培,高霞,等.硒化纹党参多糖和其抗A549细胞的活性[J].中成药,2015,37(11):2408-2413.Chen W X,Zhang P,Gao X,et al.Selenylation of polysaccharides from Codonopsis pilosula and its inhibition against A549 cells[J].Chinese Traditional Patent Medicine,2015,37(11):2408-2413(in Chinese with English abstract).
[25]Gao Z Z,Zhang C,Tian W J,et al.The antioxidative and hepatoprotective effects comparison of Chinese angelica polysaccharide(CAP)and selenizing CAP(s CAP)in CCl4induced hepatic injury mice[J].International Journal of Biological Macromolecules,2017,97:46-54.
[26]Qiu S L,Chen J,Chen X,et al.Optimization of selenylation conditions for lycium barbarum polysaccharide based on antioxidant activity[J].Carbohydrate Polymers,2014,103(1):148-153.