刺激隐核虫感染诱导斜带石斑鱼MHCIα和β_2m基因的表达谱分析

Expression pattern analysis on MHC I α and β_2m genes in Epinephelus coioides infected by Cryptocaryon irritans

【目的】检验石斑鱼(Epinephelus coioides)MHCIα和β_2m基因是否参与诱导抗寄生虫免疫应答,为揭示鱼类免疫系统启动抗刺激隐核虫(Cryptocaryon irritans)感染的细胞免疫应答分子机制打下基础。【方法】以实时荧光定量PCR对刺激隐核虫感染斜带石斑鱼前后MHC Iα和β_2m基因在感染局部位点和免疫器官中的表达变化进行检测。【结果】MHCIα和β_2m基因在健康斜带石斑鱼的组织中均呈组成性表达,且以在外周血、腮、头肾和脾脏中的表达量较高。经刺激隐核虫感染后,MHCIα和β_2m基因在斜带石斑鱼皮肤中的表达明显上调,均在感染后第5 d达峰值,其中MHCIα基因的最大表达量为2.6倍、β_2m基因的最大表达量为6.1倍;在腮中二者呈波动性变化,而在头肾中以显著下调为主。在脾脏中,MHCIα基因在刺激隐核虫感染后第5 d显著上调表达,为对照组的2.0倍,至感染后第7 d其表达水平略微下调,为对照组的1.5倍;β_2m基因表达于感染后12 h即升高至对照组的4.0倍,此后呈下调表达趋势,至感染后第5 d其表达水平再次上调为对照组的3.4倍。【结论】在刺激隐核虫感染过程中,斜带石斑鱼MHCIα和β_2m基因的相对表达量在皮肤、鳃、头肾和脾脏中均发生了明显变化,即鱼类MHCⅠ类分子可能参与了机体的细胞免疫,在抗寄生虫感染的特异性免疫应答过程中发挥重要作用。

【Objective】The purpose of this study was to verify whether MHC I α and β2m genes in Epinephelus coioides were invovled in the induction of anti—parasite immune response and lay a foundation to reveal molecular mechanism of cellular imune response started of infected fish by Cryptocaryon irritans.【Method】Real-time quantitative PCR was applied to investigate expression levels of MHC I α and β2m genes before and after E.coioides infected by C.irritans in local infection sites and immune organs.【Result】MHC I α and β2m genes were both constilutively expressed in healthy tissues of E.coioides,with higher expressions in peripheral blood,gill,spleen and head kidney.After infected by C.irritans,significant up-regulations of MHC I α and β2m genes were detected in skin and reached the peak on day 5 post infection.The maximum expression of MHC I α gene was 2.6 times and β2m gene 6.1 times.They were both fluctuated in gill and their significant declines were observed in head kidney at most time points.In spleen,the expression of MHC I α gene was up-regulated on day 5 significantly after infected by C.irritans,which was as 2 limes as that in CK;and slightly declined on day 7,which was as 1.5 times as that in CK.In terms of β2m gene,the expression was un-regulated after 12 hours of infection and as high as 4.0 times of that in CK;later the expression was down-regulated until day 5,when the expression was up-regulated again to 3.4 times of that in CK.【Conclusion】During C.irritans infection process,the relative expressions of MHC I α and β2m genes in E.coioides are both found to be significantly regulated in skin,gill,spleen and head kidney.These results indicate that fish MHC I molecule may be involved in cellular immune and play an important role in activation of specific cellular immune response to parasites.