运输应激对肉牛肝脏和脾脏中应激相关蛋白的影响
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摘要
选取15头体况相似的健康肉牛作为研究对象,随机分为运输0h组、运输3h组和运输6h组,每组5头肉牛,采用公路运输方式运输(平均速度为80km/h)。采集各组牛肝脏和脾脏组织样本,用酶联免疫吸附试验(ELISA)和荧光定量PCR(qRT-PCR)2种试验方法检测各组肝脏和脾脏组织中应激蛋白-热休克蛋白70(HSP70)和急性期蛋白-C反应蛋白(CRP)含量的变化。qRT-PCR检测结果显示:与运输0h组相比,运输3h和6h肝脏组织中HSP70mRNA的表达量极显著升高(P<0.01);而脾脏组织中HSP70mRNA只在运输应激6h后表达量显著增加(P<0.05)。与0h组相比,运输应激后肝脏组织中CRP mRNA的表达量极显著升高(P<0.01),脾脏组织中CRP mRNA的表达量显著升高(P<0.05)。ELISA检测结果显示:与0h组相比,运输应激3h和6h肝脏中的HSP70质量浓度极显著升高(P<0.01),脾脏中HSP70的质量浓度在运输6h后显著升高(P<0.05);肝脏和脾脏CRP的表达在运输应激6h后极显著升高(P<0.01)。结果表明:不同运输时间的运输应激可导致肝脏和脾脏中HSP70和CRP的浓度和mRNA转录水平升高,且肝脏中HSP70和CRP的表达量高于脾脏。HSP70和CRP可作为肉牛运输应激候选诊断标志物。
In order to study the effect of transport stress on stress protein-HSP70 and acute phase protein-CRP in liver and spleen of beef cattle at different times.15 healthy beef cattle with similar body condition were selected as study subjects and randomly divided into transport 0 hgroup,transport 3 hgroup and transport 6 hgroup,each group of 5 beef cattle.The tested animals were transported by road transport(average speed of 80 km/h).After transportation the liver and spleen samples of each group were collected,and the levels of HSP70 and CRP in liver and spleen of each group were detected by enzyme-linked immunosorbent assay(ELISA)and quantitative PCR(qRT-PCR).The results of qRT-PCR showed that the mRNA expression of HSP70 in liver tissue was significantly increased at 3 hand 6 hafter transport compared with that at 0 h(P<0.01),while the expression of HSP70 in spleen was only increased at 6 hafter transport stress(P<0.05).Compared with 0 hgroup,the expression of CRP mRNA in liver tissue increased significantly(P<0.01)and the expression of CRP mRNA in spleen increased significantly(P<0.05).The results of ELISA showed that the HSP70 concentrations in liver were significantly increased at 3 h and 6 h after transport stress(P <0.01)compared with 0 h group.The HSP70 concentration in spleen increased significantly at 6 hafter transport(P<0.05).The expression of CRP in liver and spleen significantly increased at 6 hafter transport stress(P<0.01).Transport stress at different transport time could lead to the increase of HSP70 and CRP concentration and mRNA transcription in liver and spleen,and the HSP70 and CRP in liver are higher than those in spleen.HSP70 and CRP can be used as candidate diagnostic markers for stress in beef cattle.
In order to study the effect of transport stress on stress protein-HSP70 and acute phase protein-CRP in liver and spleen of beef cattle at different times.15 healthy beef cattle with similar body condition were selected as study subjects and randomly divided into transport 0 hgroup,transport 3 hgroup and transport 6 hgroup,each group of 5 beef cattle.The tested animals were transported by road transport(average speed of 80 km/h).After transportation the liver and spleen samples of each group were collected,and the levels of HSP70 and CRP in liver and spleen of each group were detected by enzyme-linked immunosorbent assay(ELISA)and quantitative PCR(qRT-PCR).The results of qRT-PCR showed that the mRNA expression of HSP70 in liver tissue was significantly increased at 3 hand 6 hafter transport compared with that at 0 h(P<0.01),while the expression of HSP70 in spleen was only increased at 6 hafter transport stress(P<0.05).Compared with 0 hgroup,the expression of CRP mRNA in liver tissue increased significantly(P<0.01)and the expression of CRP mRNA in spleen increased significantly(P<0.05).The results of ELISA showed that the HSP70 concentrations in liver were significantly increased at 3 h and 6 h after transport stress(P <0.01)compared with 0 h group.The HSP70 concentration in spleen increased significantly at 6 hafter transport(P<0.05).The expression of CRP in liver and spleen significantly increased at 6 hafter transport stress(P<0.01).Transport stress at different transport time could lead to the increase of HSP70 and CRP concentration and mRNA transcription in liver and spleen,and the HSP70 and CRP in liver are higher than those in spleen.HSP70 and CRP can be used as candidate diagnostic markers for stress in beef cattle.
引文
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[12]李玉保,裴兰英,鲍恩东,等.运输应激猪HSPs mRNA转录、分布及免疫器官病理性损伤相关性研究[J].中国农业科学,2008(6):1832-1837.
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[14]许利凡.运输对夏南牛血液主要APPs、激素及细胞因子水平的影响[D].河南郑州:河南农业大学,2014.
[15] YAKOB M,MEURMAN J H,JOGESTRAND T,et al.C-reactive protein in relation to early atherosclerosis and periodontitis[J].Clin Oral Invest,2012,16(1):259-265.
[2]裴志花,王开,马红霞,等.运输应激及其对动物免疫功能影响的研究进展[J].中国兽医杂志,2012,48(1):81-83.
[3]YU A,LI P,TANG T,et al.Roles of HSP70sin stress responses of microorganisms,plants,and animals[J].Biomed Res Int,2015(16):510319.doi:10.1155/2015/510319.
[4]COLLIER R J,RENQUIST B J,XIAO Y.A 100-year review:stress physiology including heat stress[J].J Dairy Sci,2017,100(12):10367-10380.
[5]MARQUES A T,NORDIO L,LECCHI C,et al.Widespread extrahepatic expression of acute-phase proteins in healthy chicken(Gallus gallus)tissues[J].Vet Immunol Immunopathol,2017,190:10-17.
[6]CRAY C,ZAIAS J,ALTMAN N H.Acute phase response in animals:a review[J].Comp Med,2009,59(6):517-526.
[7]DABROWSKI R,WAWRON W,KOSTRO K.Changes in CRP,SAA,and haptoglobin produced in response to ovariohysterectomy in healthy bitches and those with pyometra[J].Theriogenology,2007,67:321-327.
[8]MARTI S,WILDE R E,MOYA D,et al.Effect of rest stop duration during long-distance transport on welfare indicators in recently weaned beef calves[J].J Anim Sci,2017,95(2):636.
[9]丁亚军.肉牛运输应激综合征的发生与诊疗体会[C]//中国畜牧业协会.第八届中国牛业发展大会论文集.[S.l.]:中国畜牧业协会,2013.
[10] LEONARD B E.The HPA and immune axes in stress:the involvement of the serotonergic system[J].Eur Psychiat,2005(Suppl 3):302-306。
[11]连帅,司鸿飞,王迪,等.复合添加剂缓解出栏肉牛运输应激效果评价[J].中国兽医学报,2016,36(9):1603-1607.
[12]李玉保,裴兰英,鲍恩东,等.运输应激猪HSPs mRNA转录、分布及免疫器官病理性损伤相关性研究[J].中国农业科学,2008(6):1832-1837.
[13] KURASH J K,SHEN C N,TOSH D.Induction and regulation of acute phase prote in sintransdifferentiated hepatocytes[J].Exp Cell Res,2004,292:342-358。
[14]许利凡.运输对夏南牛血液主要APPs、激素及细胞因子水平的影响[D].河南郑州:河南农业大学,2014.
[15] YAKOB M,MEURMAN J H,JOGESTRAND T,et al.C-reactive protein in relation to early atherosclerosis and periodontitis[J].Clin Oral Invest,2012,16(1):259-265.
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