禽网状内皮组织增生病病毒感染SPF雏鸡血液淋巴细胞增殖功能及其细胞周期相关因子变化
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摘要
为探索禽网状内皮组织增生病病毒(REV)感染对SPF雏鸡外周血液的淋巴细胞增殖功能及其细胞周期素D1、p27表达的影响。将80只1日龄SPF雏鸡随机分为REV感染组和对照组,应用CCK-8、RT-qPCR、Western Blot等方法对雏鸡外周血液上述被检指标的动态变化进行检测。结果表明,REV感染SPF雏鸡后,其外周血液中T、B淋巴细胞增殖能力明显低于对照组雏鸡,且p27mRNA水平显著(P<0.05)高于对照组雏鸡;病毒感染后期Cyclin D1mRNA表达及其蛋白质含量均显著高于(P<0.05)对照组雏鸡,然而p27转录显著低于对照组雏鸡(P<0.05)。REV感染SPF雏鸡导致的血液T、B细胞增殖能力下降、Cyclin D1mRNA转录及其蛋白质含量的增加而p27mRNA转录下降与REV感染所致的雏鸡免疫机能下降密切相关。
This experiment was conducted to explore the proliferation function of T,B lymphocytes and expression of Cyclin D1 and p27 in peripheral blood of SPF chicks infected with reticuloendotheliosis virus(REV).Eighty 1-day-old SPF chicks were randomly divided into REV infection group and control group.The T,B lymphocytes proliferation function and expression of Cyclin D1 at mRNA and protein level,transcription of p27 in peripheral blood were detected by CCK-8,RT-qPCR and Western Blot.The results showed that the proliferation ability of T and B lymphocytes in peripheral blood of SPF chicks infected with REV early was significantly lower than that of the control chicks,and the transcription of p27 mRNA was significantly higher than that of the control chicks.The expression of Cyclin D1 mRNA and protein content in the late stage of infection were significantly higher than that of the control chicks(P<0.05),but the transcription of p27 mRNA were significantly lower than those of control chicks(P<0.05).The above results showed that the decrease of T,B lymphocytes proliferative function and p27 mRNAtranscription,and increase of Cyclin D1 at mRNA and protein level in blood of SPF chicks were closely related to the decrease in immune function of chicks infected with REV.
This experiment was conducted to explore the proliferation function of T,B lymphocytes and expression of Cyclin D1 and p27 in peripheral blood of SPF chicks infected with reticuloendotheliosis virus(REV).Eighty 1-day-old SPF chicks were randomly divided into REV infection group and control group.The T,B lymphocytes proliferation function and expression of Cyclin D1 at mRNA and protein level,transcription of p27 in peripheral blood were detected by CCK-8,RT-qPCR and Western Blot.The results showed that the proliferation ability of T and B lymphocytes in peripheral blood of SPF chicks infected with REV early was significantly lower than that of the control chicks,and the transcription of p27 mRNA was significantly higher than that of the control chicks.The expression of Cyclin D1 mRNA and protein content in the late stage of infection were significantly higher than that of the control chicks(P<0.05),but the transcription of p27 mRNA were significantly lower than those of control chicks(P<0.05).The above results showed that the decrease of T,B lymphocytes proliferative function and p27 mRNAtranscription,and increase of Cyclin D1 at mRNA and protein level in blood of SPF chicks were closely related to the decrease in immune function of chicks infected with REV.
引文
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[11]李宏梅,成子强,刘建柱,等.REV与AJV-L共感染对肉鸡T淋巴细胞免疫功能与组织病理学的影响[J].中国农业科学,2009,42(9):3296-3304.LI H M,CHENG Z Q,LIU J Z,et al.Effect of coinfection with REV and ALV-J on T lymphocytes bioactivities and histopathology in broiler chickens[J].Scientia Agricultura Sinica,2009,42(9):3296-3304.(in Chinese)
[12]侯宁.REV感染对SPF雏鸡免疫器官细胞凋亡及Bcl-2、C-myc基因表达的影响[D].哈尔滨:东北农业大学,2016.HOU N.Influence of reticuloendotheliosis virus to cells apoptosis and gene expression of bcl-2and Cmyc in immune organs of SPF chickens[D].Harbin:Northeast Agricultural University,2016.(in Chinese)
[13]GOLIAS C H,CHARALABOPOULOS A,CHARALABOPOULOS K.Cell proliferation and cell cycle control:a mini review[J].Int J Clin Pract,2004,58(12):1134-1141.
[14]KIM T H,OH S,KIM S S.Recombinant human prothrombin kringle-2induces bovine capillary endothelial cell cycle arrest at G0-G1phase through inhibition of cyclin D1/CDK4complex:modulation of reactive oxygen species generation and up-regulation of cyclin-dependent kinase inhibitors[J].Angiogenesis,2006,8(4):307-314.
[15]FENG W G,ZHOU D F,MENG W,et al.Growth retardation induced by avian leukosis virus subgroup J associated with down-regulated Wnt/β-catenin pathway[J].Microb Pathog,2017,104:48-55,doi:10.1016/j.micpath.2017.01.013.
[16]徐彦波,金华,甘永华.鸡贫血病病毒感染雏鸡细胞周期的研究[J].中国预防兽医学报,1999,21(2):122-123.XU Y B,JIN H,GAN Y H.Cell cycle studies on chickens experimentally infected with chicken anemia virus[J].Chinese Journal of Preventive Veterinary Medicine,1999,21(2):122-123.(in Chinese)
[17]SANDA T,ASAMITSU K,OGURA H,et al.Induction of cell death in adult T-cell leukemia cells by a novel IκB kinase inhibitor[J].Leukemia,2006,20(4):590-598.
[18]李凌云.人疱疹病毒6型感染对HSB2细胞周期和细胞凋亡的影响及机制研究[D].南京:南京医科大学,2010.LI L Y.Effect of human herpes virus type 6infection on cell cycle and apoptosis of HSB2 cells and its mechanism[D].Nanjing:Nanjing Medical University,2010.(in Chinese)
[19]XU M,CHEUNG C C M,CHOW C,et al.Overexpression of PIN1enhances cancer growth and aggressiveness with Cyclin D1induction in EBV-associated nasopharyngeal carcinoma[J].PLoS One,2016,11(6):e0156833.
[20]MINARIKOVA P,BENESOVA L,HALKOVA T,et al.Prognostic importance of cell cycle regulators cyclin D1(CCND1)and cyclin-dependent kinase inhibitor 1B(CDKN1B/p27)in sporadic gastric cancers[J].Gastroenterol Res Pract,2016,2016:9408190,doi:10.1155/2016/9408190.
[21]刘润.p27与胃粘膜上皮细胞周期调控的研究进展[J].国外医学·消化系疾病分册,2003,23(1):52-55.LIU R.The research progress of p27and gastric mucosal cell cycle regulation[J].Foreign Medical Sciences.Section of Digestive Diseases,2003,23(1):52-55.(in Chinese)
[22]YU Z Q,GAO X L,LIU C N,et al.Analysis of microRNA expression profile in specific pathogen-free chickens in response to reticuloendotheliosis virus infection[J].Appl Microbiol Biotechnol,2017,101(7):2767-2777.
[23]陶金程,汉丽梅,刘佳,等.细胞周期蛋白激酶抑制因子p27 Kip1研究进展[J].动物医学进展,2011,32(5):112-115.TAO J C,HAN L M,LIU J,et al.Progress on cyclin-dependent kinase inhibitor p27Kip1[J].Progress in Veterinary Medicine,2011,32(5):112-115.(in Chinese)
[24]OMATSU M,KUNIMURA T,MIKOGAMI T,et al.Cyclin-dependent kinase inhibitors,p16and p27,demonstrate different expression patterns in thymoma and thymic carcinoma[J].Gen Thorac Cardiovasc Surg,2014,62(11):678-684.
[25]KRAVTSOV V G,SHAKHMUROV M G,SUKMANOV O V,et al.Expression of cycline-dependent kinase p27in the low differentiated gastric adenocarcinoma[J].Arkh Patol,2006,68(5):14-16.
[2]张文娟,裴宗飞,牛钟相.禽网状内皮组织增生症病毒env蛋白的表达及其在ELISA方法中的应用[J].畜牧兽医学报,2009,40(10):1521-1525.ZHANG W J,PEI Z F,NIU Z X.Expression of reticuloendotheliosis virus env protein and development of ienv-ELISA for detection of antibody to REV[J].Acta Veterinaria et Zootechnica Sinica,2009,40(10):1521-1525.(in Chinese)
[3]姜莉莉,祁小乐,高玉龙,等.野鸭源禽网状内皮组织增生症病毒的分离与鉴定[J].畜牧兽医学报,2013,44(8):1277-1282.JIANG L L,QI X L,GAO Y L,et al.Isolation and identification of reticuloendotheliosis virus in wild birds[J].Acta Veterinaria et Zootechnica Sinica,2013,44(8):1277-1282.(in Chinese)
[4]卢梦玲,闫超,赖多,等.Cyclin D1与细胞周期调控[J].生物技术通报,2011(10):55-59.LU M L,YAN C,LAI D,et al.Cyclin D1and cell cycle regulation[J].Biotechnology Bulletin,2011(10):55-59.(in Chinese)
[5]邹向阳,李连宏.细胞周期调控与肿瘤[J].国际遗传学杂志,2006,29(1):70-73.ZOU X H,LI L H.Cell cycle control and tumor[J].International Journal of Genetics,2006,29(1):70-73.(in Chinese)
[6]罗昆仑.p27与细胞周期调控[J].国外医学·肿瘤学分册,1998,25(5):262-265.LUO K L.P27and cell cycle regulation[J].Foreign Medical Sciences.Cancer Section,1998,25(5):262-265.(in Chinese)
[7]周雪红.病毒感染对细胞周期调控影响的研究进展[J].微生物学免疫学进展,2012,40(6):54-57.ZHOU X H.Research progress of cell cycle regulation during virus infection[J].Progress in Microbiology and Immunology,2012,40(6):54-57.(in Chinese)
[8]XUE M,SHI X M,ZHAO Y,et al.Effects of reticuloendotheliosis virus infection on cytokine production in SPF chickens[J].PLoS One,2013,8(12):e83918,doi:10.1371/journal.pone.0083918.
[9]胡峰.宿主波形蛋白在禽网状内皮组织增生病病毒感染中的作用机制研究[D].哈尔滨:中国农业科学院研究生院(哈尔滨兽医研究所),2015.HU F.The molecular mechanism of host vimentin protein contributing to reticuloendotheliosis virus infection[D].Harbin:Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,2015.(in Chinese)
[10]孔斌,陈雪锋,唐德宏,等.禽网状内皮组织增生症病毒感染肉鸡中鸡白细胞介素18的定量检测[J].畜牧兽医学报,2011,42(7):963-966.KONG B,CHEN X F,TANG D H,et al.Quantitative detection of chicken IL-18in broiler chickens infected with reticuloendotheliosis virus[J].Acta Veterinaria et Zootechnica Sinica,2011,42(7):963-966.(in Chinese)
[11]李宏梅,成子强,刘建柱,等.REV与AJV-L共感染对肉鸡T淋巴细胞免疫功能与组织病理学的影响[J].中国农业科学,2009,42(9):3296-3304.LI H M,CHENG Z Q,LIU J Z,et al.Effect of coinfection with REV and ALV-J on T lymphocytes bioactivities and histopathology in broiler chickens[J].Scientia Agricultura Sinica,2009,42(9):3296-3304.(in Chinese)
[12]侯宁.REV感染对SPF雏鸡免疫器官细胞凋亡及Bcl-2、C-myc基因表达的影响[D].哈尔滨:东北农业大学,2016.HOU N.Influence of reticuloendotheliosis virus to cells apoptosis and gene expression of bcl-2and Cmyc in immune organs of SPF chickens[D].Harbin:Northeast Agricultural University,2016.(in Chinese)
[13]GOLIAS C H,CHARALABOPOULOS A,CHARALABOPOULOS K.Cell proliferation and cell cycle control:a mini review[J].Int J Clin Pract,2004,58(12):1134-1141.
[14]KIM T H,OH S,KIM S S.Recombinant human prothrombin kringle-2induces bovine capillary endothelial cell cycle arrest at G0-G1phase through inhibition of cyclin D1/CDK4complex:modulation of reactive oxygen species generation and up-regulation of cyclin-dependent kinase inhibitors[J].Angiogenesis,2006,8(4):307-314.
[15]FENG W G,ZHOU D F,MENG W,et al.Growth retardation induced by avian leukosis virus subgroup J associated with down-regulated Wnt/β-catenin pathway[J].Microb Pathog,2017,104:48-55,doi:10.1016/j.micpath.2017.01.013.
[16]徐彦波,金华,甘永华.鸡贫血病病毒感染雏鸡细胞周期的研究[J].中国预防兽医学报,1999,21(2):122-123.XU Y B,JIN H,GAN Y H.Cell cycle studies on chickens experimentally infected with chicken anemia virus[J].Chinese Journal of Preventive Veterinary Medicine,1999,21(2):122-123.(in Chinese)
[17]SANDA T,ASAMITSU K,OGURA H,et al.Induction of cell death in adult T-cell leukemia cells by a novel IκB kinase inhibitor[J].Leukemia,2006,20(4):590-598.
[18]李凌云.人疱疹病毒6型感染对HSB2细胞周期和细胞凋亡的影响及机制研究[D].南京:南京医科大学,2010.LI L Y.Effect of human herpes virus type 6infection on cell cycle and apoptosis of HSB2 cells and its mechanism[D].Nanjing:Nanjing Medical University,2010.(in Chinese)
[19]XU M,CHEUNG C C M,CHOW C,et al.Overexpression of PIN1enhances cancer growth and aggressiveness with Cyclin D1induction in EBV-associated nasopharyngeal carcinoma[J].PLoS One,2016,11(6):e0156833.
[20]MINARIKOVA P,BENESOVA L,HALKOVA T,et al.Prognostic importance of cell cycle regulators cyclin D1(CCND1)and cyclin-dependent kinase inhibitor 1B(CDKN1B/p27)in sporadic gastric cancers[J].Gastroenterol Res Pract,2016,2016:9408190,doi:10.1155/2016/9408190.
[21]刘润.p27与胃粘膜上皮细胞周期调控的研究进展[J].国外医学·消化系疾病分册,2003,23(1):52-55.LIU R.The research progress of p27and gastric mucosal cell cycle regulation[J].Foreign Medical Sciences.Section of Digestive Diseases,2003,23(1):52-55.(in Chinese)
[22]YU Z Q,GAO X L,LIU C N,et al.Analysis of microRNA expression profile in specific pathogen-free chickens in response to reticuloendotheliosis virus infection[J].Appl Microbiol Biotechnol,2017,101(7):2767-2777.
[23]陶金程,汉丽梅,刘佳,等.细胞周期蛋白激酶抑制因子p27 Kip1研究进展[J].动物医学进展,2011,32(5):112-115.TAO J C,HAN L M,LIU J,et al.Progress on cyclin-dependent kinase inhibitor p27Kip1[J].Progress in Veterinary Medicine,2011,32(5):112-115.(in Chinese)
[24]OMATSU M,KUNIMURA T,MIKOGAMI T,et al.Cyclin-dependent kinase inhibitors,p16and p27,demonstrate different expression patterns in thymoma and thymic carcinoma[J].Gen Thorac Cardiovasc Surg,2014,62(11):678-684.
[25]KRAVTSOV V G,SHAKHMUROV M G,SUKMANOV O V,et al.Expression of cycline-dependent kinase p27in the low differentiated gastric adenocarcinoma[J].Arkh Patol,2006,68(5):14-16.