Zhao Beibei, Katagiri Takayuki, Kondo Hidehiro, Hirono Ikuo
Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato-ku, Tokyo 108-8477, Japan.
Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato-ku, Tokyo 108-8477, Japan.
Dev Comp Immunol. 2015 Sep;52(1):37-47. doi: 10.1016/j.dci.2015.04.011. Epub 2015 Apr 23.
A new type of CXCL8, named CXCL8_L1b, was identified in this research. Comparison of amino acid sequences of Japanese flounder CXCL8_L1b and CXCL8_L1a (BAB86884.1) showed only 41.2% identity. Transcripts of CXCL8_L1a were highly detected in spleen, kidney, gill and liver, while transcripts of CXCL8_L1b only were detected highly in spleen and kidney of apparently healthy fish. In fish challenged with E. tarda, transcripts of CXCL8_L1a were significantly increased at day 6, while no significant increase was detected in the mRNA level of CXCL8_L1b. On the other hand, fish infected by S. iniae significantly increased both transcripts of CXCL8_L1a and CXCL8_L1b at days 1 and 3. In VHSV-infected fish, only the transcripts of CXCL8_L1b were significantly induced at day 6. LPS and poly I:C stimulation of PBLs induced a high level of CXCL8_L1a transcripts, while CXCL8_L1b transcripts were significantly increased only post poly I:C treatment. To evaluate the chemotactic activity of CXCL8_L1a and CXCL8_L1b, Japanese flounder were intramuscularly injected with recombinant plasmids pCI-CXCL8_L1a and pCI-CXCL8_L1b. H & E staining showed that injections of both pCI-CXCL8_L1a and pCI-CXCL8_L1b caused strong immune responses in the form of intermuscular cell infiltration and capillary congestion. Injection of pCI-CXCL8_L1a and pCI-CXCL8_L1b significantly induced the expressions of genes related to inflammatory response such as IL-6 and CD8α on day 1 post-injection. The transcripts of IgM only significantly increased on day 7 post-injection of pCI-CXCL8_L1b.
本研究鉴定出一种新型的CXCL8，命名为CXCL8_L1b。对牙鲆CXCL8_L1b和CXCL8_L1a（BAB86884.1）的氨基酸序列进行比较，结果显示其一致性仅为41.2%。CXCL8_L1a的转录本在脾脏、肾脏、鳃和肝脏中高表达，而CXCL8_L1b的转录本仅在外观健康的鱼的脾脏和肾脏中高表达。在用迟缓爱德华氏菌攻击的鱼中，CXCL8_L1a的转录本在第6天显著增加，而CXCL8_L1b的mRNA水平未检测到显著增加。另一方面，感染海豚链球菌的鱼在第1天和第3天CXCL8_L1a和CXCL8_L1b的转录本均显著增加。在感染水疱性口炎病毒的鱼中，仅在第6天CXCL8_L1b的转录本被显著诱导。脂多糖和聚肌胞苷酸刺激外周血淋巴细胞诱导CXCL8_L1a转录本高水平表达，而CXCL8_L1b转录本仅在聚肌胞苷酸处理后显著增加。为了评估CXCL8_L1a和CXCL8_L1b的趋化活性，将重组质粒pCI-CXCL8_L1a和pCI-CXCL8_L1b肌肉注射到牙鲆体内。苏木精-伊红染色显示，注射pCI-CXCL8_L1a和pCI-CXCL8_L1b均引起强烈的免疫反应，表现为肌间细胞浸润和毛细血管充血。注射pCI-CXCL8_L1a和pCI-CXCL8_L1b在注射后第1天显著诱导了与炎症反应相关的基因如IL-6和CD8α的表达。仅在注射pCI-CXCL8_L1b后第7天IgM的转录本显著增加。
