Department of Marine Biology & Aquaculture, Institute of Marine Industry, College of Marine Science, Gyeongsang National University, 455, Tongyeong, Tongyong, Gyeongnam 650-160, Republic of Korea.
Fish Shellfish Immunol. 2013 May;34(5):1103-11. doi: 10.1016/j.fsi.2013.01.007. Epub 2013 Jan 30.
Chemokines are chemoattractant cytokines defined by the presence of four conserved cysteine residues. In mammals, these cytokines can be divided into four subfamilies depending on the arrangement of the first two conserved cysteines in the sequence, and include the CXC(α), CC(β), C(γ), and CX3C(δ) classes. We identified CXC chemokine cDNA, designated RbCXC, isolated using expressed sequence tag analysis of a lipopolysaccharide (LPS)-stimulated rock bream liver cDNA library. The full-length RbCXC cDNA (742 bp) contained an open reading frame of 342 bp encoding 114 amino acids. Results from phylogenetic analysis showed that RbCXC was strictly separated into a distinct clade compared to other known CXC chemokine subgroups. RbCXC was significantly expressed in the trunk kidney, liver, spleen, gill, peripheral blood leukocytes (PBLs), and head kidney. Rock bream PBLs were stimulated with several mitogens, including LPS and polyinosinic-polycytidylic acid (poly I:C), which significantly induced the expression of RbCXC mRNA. RbCXC mRNA expression was examined in several tissues under conditions of bacterial and viral challenge. Experimental challenges revealed that all examined tissues from fish infected with Edwardsiella tarda and red sea bream iridovirus showed significant increases in RbCXC expression compared to the control. In the case of Streptococcus iniae infection, RbCXC mRNA expression was markedly upregulated in the kidney, spleen, and liver. In addition, a maltose binding protein fusion recombinant RbCXC (~53 kDa) was produced in an Escherichia coli expression system and purified. Subsequently, the addition of purified recombinant RbCXC (rRbCXC) to kidney leukocytes was examined to investigate the impact of proliferative and chemotactic activity. The rRbCXC induced significant kidney leukocyte proliferation and attraction at concentrations ranging from 10 to 300 μg/mL, suggesting that it can be utilised as an immune stimulant and/or molecular adjuvant to enhance the immunological effects of vaccines.
趋化因子是一类具有化学引诱作用的细胞因子，其特征为存在四个保守的半胱氨酸残基。在哺乳动物中，根据序列中前两个保守半胱氨酸的排列方式，这些细胞因子可分为四个亚家族，包括 CXC(α)、CC(β)、C(γ)和 CX3C(δ)类。我们利用脂多糖（LPS）刺激的牙鲆肝 cDNA 文库的表达序列标签分析，鉴定出了趋化因子 cDNA，命名为 RbCXC。RbCXC 的全长 cDNA（742bp）包含一个 342bp 的开放阅读框，编码 114 个氨基酸。系统进化分析结果表明，与其他已知的 CXC 趋化因子亚群相比，RbCXC 严格地分为一个独特的分支。RbCXC 在躯干肾脏、肝脏、脾脏、鳃、外周血白细胞（PBLs）和头肾中均有显著表达。牙鲆 PBLs 被几种有丝分裂原刺激，包括 LPS 和聚肌胞苷酸（poly I:C），这些刺激显著诱导了 RbCXC mRNA 的表达。在细菌和病毒挑战条件下，我们在几种组织中检测了 RbCXC mRNA 的表达。实验性挑战表明，与对照组相比，感染迟钝爱德华氏菌和红鳍东方鲀虹彩病毒的鱼的所有检测组织中，RbCXC 表达均显著增加。在感染无乳链球菌的情况下，肾脏、脾脏和肝脏中 RbCXC mRNA 的表达明显上调。此外，在大肠杆菌表达系统中产生了麦芽糖结合蛋白融合重组 RbCXC（~53kDa）并进行了纯化。随后，研究了向肾脏白细胞中添加纯化的重组 RbCXC（rRbCXC）以研究其增殖和趋化活性的影响。rRbCXC 在 10 至 300μg/mL 的浓度范围内诱导显著的肾脏白细胞增殖和趋化作用，表明它可用作免疫刺激剂和/或分子佐剂，以增强疫苗的免疫效果。
