Abstract：
CD49d, encoded by the gene Integrin α4, is a significant member of cell adhesion receptors, which is widely expressed in various immune cells to trigger immune responses against invading pathogens. In the present study, the expression of CgCD49d and its regulatory role in TNF expression were investigated in the Pacific oyster Crassostrea gigas. There were five Int-alpha domains, an Integrin_alpha2 region and a unique FG-GAP repeat region inserted identified in CgCD49d. CgCD49d transcript was specifically expressed in haemocytes, and its mRNA expression level in haemocytes increased after LPS and Vibrio splendidus stimulation. After CgCD49d was blocked by using its antibody, the phosphorylation level of CgJNK in the MAPK signaling pathway and CgTNF transcripts decreased significantly post V. splendidus stimulation. After phosphorylation level of CgJNK was inhibited by using its inhibitor, the nuclear translocation of CgRel was restrained and CgTNF transcripts also decreased significantly post V. splendidus stimulation. Furthermore, CgCD49d was found to be mainly expressed in the agranulocyte subpopulation, and Alexa Fluor 488-conjugated CgCD49d antibody labeled agranulocytes with a circle of green fluorescence signals on CgCD49d+ agranulocyte surface under Confocal microscopy, which accounted for 24.9 ± 4.53% of total haemocytes. Collectively, these results suggested that CgCD49d promoted TNF expression in oyster haemocytes against bacterial invasion by mediating MAPK pathway, and it could be used as a surface marker to type and sort a subset of agranulocyte subpopulation among haemocytes.
摘要：
CD49d,由基因整合素α4编码，是细胞粘附受体的重要成员，它在各种免疫细胞中广泛表达，以触发针对入侵病原体的免疫反应。在本研究中,在太平洋牡蛎Crassostreagigas中研究了CgCD49d的表达及其对TNF表达的调节作用。有五个Int-alpha域,在CgCD49d中鉴定的整合素_α2区和插入的独特FG-GAP重复区。CgCD49d转录本在血细胞中特异性表达，LPS和脾弧菌刺激后，其在血细胞中的mRNA表达水平增加。CgCD49d用其抗体阻断后，脾静脉刺激后,MAPK信号通路中CgJNK的磷酸化水平和CgTNF转录本显著降低。在使用其抑制剂抑制CgJNK的磷酸化水平后，CgRel的核易位受到抑制，CgTNF转录本在脾V.刺激后也显着降低。此外，发现CgCD49d主要在粒细胞亚群中表达，和AlexaFluor488缀合的CgCD49d抗体标记的粒细胞，在共聚焦显微镜下在CgCD49d+粒细胞表面上有一圈绿色荧光信号，占血细胞总数的24.9±4.53％。总的来说，这些结果表明，CgCD49d通过介导MAPK通路促进牡蛎血细胞中TNF的表达抵抗细菌侵袭，它可以用作表面标记，以对血细胞中的粒细胞亚群进行分型和分类。
