运输应激对猪脾脏IL-2、IL-6和IL-10 mRNAs表达的影响及其调控机制

【目的】探讨不同时间运输应激对猪脾脏中IL-2mRNA、IL-6mRNA、IL-10mRNA及其IL-2RmRNA、IL-6RmRNA表达的影响,并探讨P38MAPK-NF-κB信号途径对其表达的调控作用。【方法】将19头体重50kg左右的猪随机分为4组,分别进行0、1、2和4h公路运输,运输结束后立即扑杀取脾脏,液氮保存,用荧光定量PCR的方法对3种细胞因子及其IL-2RmRNA、IL-6RmRNA进行定量;另用ELISA方法检测试验猪脾脏组织匀浆液中P38MAPK含量的动态变化;激光扫描共聚焦显微镜观察NF-κB运输前后在脾脏细胞中的分布。【结果】经1、2和4h的运输应激后,脾脏中IL-10、IL-6及IL-6RmRNA的表达发生了显著的变化,P38MAPK变化不明显,NF-κB在脾脏中的表达是先上升,随后稍微下降,到4h时又升高并高于其它3个组;且NF-κB的入核率也呈现同样的变化趋势。【结论】运输应激可引起猪脾脏IL-6mRNA、IL-10mRNA和IL-6RmRNA表达的显著变化,且存在时效性;相关性分析表明,在所检测的3种细胞因子中,脾脏主要对运输过程中IL-6水平起重要的调节作用,结合细胞因子表达调控的信号机制表明,NF-κB可能对运输过程中猪脾脏相关细胞因子变化起重要的调控作用。

Effects of Transportation Stress on Expressions of IL-2,IL-6 and IL-10 mRNAs in Spleen and Regulation of Cytokines in Pigs

[Objective] The objective of this study was to explore expressions of IL-2, IL-6, IL-10 mRNAs, IL-2RmRNA and IL-6RmRNA in spleen and regulation of cytokines by P38MAPK-NF-κB in pigs during truck transportation. [Method] Nineteen pigs were randomly divided into 4 groups for 0 h,1 h, 2 h and 4 h transportaion when mean live weight was approximately (50±2) kg (mean±standard deviation; SD). At the end of the transportation trial, the spleen was removed and frozen in liquid nitrogen. SYBR Green I-based one-step real-time quantitative RT-PCR amplification was performed in quantitation of the three cytokines, IL-2RmRNA and IL-6RmRNA. P38MAPK in the homogenization sample of spleen was measured with homologous specific enzyme-linked immunosorbent assay (ELISA). The distribution of NF-κB in the spleen was examined using confocal laser scanning microscope (CLSM). [Result] After 1 h, 2 h and 4 h of transportation, the expression level of IL-6, IL-10 and IL-6RmRNAs in spleen were significantly changed compared with the control group during transportation stress. NF-κB in the spleen has a sharp rise at 1 h, followed by a slight decline, rise again at 4 h and higher than the other three groups. The rates of NF-κB entry nuclear showed a similar trend, but there was no significant change with respect to P38MAPK. [Conclusion] The IL-6mRNA, IL-10mRNA and the expression of IL-6RmRNA could be significantly changed by transportation stress and were also related to the time of post-stress in pigs. Correlation analysis showed that the spleen is an important regulator of IL-6 levels among the detected three kinds of cytokines during transportation Stress.Combination of expression and regulation of cytokine signaling mechanism shows that the activation of NF-κB in the spleen may be play an important role in the peripheral immune-related cytokines change in transport-stressed pigs.