Two hemocyte sub-populations of kuruma shrimp Marsupenaeus japonicus
Two hemocyte sub-populations of kuruma shrimp Marsupenaeus japonicus
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DOI：
10.1016/j.molimm.2017.01.024
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发表时间：
2017-05-01
影响因子：
3.6
通讯作者：
Hirono, Ikuo
中科院分区：
文献类型：
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作者：
Koiwai, Keiichiro;Alenton, Rod Russel R.;Hirono, Ikuo
血淋巴中的血细胞在对虾的免疫应答中起着重要的作用。以往的研究是基于血细胞的整个人口的基因表达分析，因此可能错过了不同的血细胞亚群的免疫反应。本研究采用Percoll梯度离心法将血细胞分为两个亚群，分别采用May Giemsa染色、流式细胞仪和FACSCalibur分析了各亚群的形态学特征。结果显示，血细胞分为上层嗜碱性血细胞和下层嗜酸性血细胞。嗜碱性血细胞比嗜酸性血细胞更大，嗜酸性血细胞比嗜碱性血细胞更颗粒化。然后通过Miseq的RNA-seq分析进行转录组分析，其揭示了两个亚群之间的16个差异表达的转录物。上层血细胞高表达的转录本与免疫相关基因同源，提示该层血细胞可能是免疫系统的调节者，控制其他细胞的作用，从而消灭病原体。另一方面，在较低层中高度表达的转录物与抗菌肽（AMP）crustin同源，这支持该层上的血细胞具有颗粒，因为crustin通常从血细胞颗粒分泌。crustin在下层的高表达也提供了对抗微生物功能机制的深入了解，其中血细胞在其颗粒中产生和储存AMP。这些差异表达的基因是潜在的血细胞分子标记，其中我们鉴定了来自上层的血细胞中的高表达基因之一（c11736-g1），其被预测为表面分子，这是分子标记的共同特征，是有希望的候选分子标记。(C)2017爱思唯尔有限公司版权所有。
Hemocytes in the circulating hemolymph play important roles for immune responses in shrimp. Previous studies on immune responses by hemocytes in penaeid shrimp were based on gene expression analyses of the entire population of hemocytes and thus may have missed different immune responses of different hemocyte sub-populations. In this study, we separated hemocytes into two sub-populations by Percoll gradient centrifugation, morphological characteristics of each population were then analyzed by May Giemsa staining, flow cytometry, and FACSCalibur. Results showed hemocytes were divided into an upper layer basophilic, and lower layer of eosinophilic hemocytes. Basophilic hemocytes were larger in size compared to eosinophilic hemocytes, which were more granulated than the basophilic hemocytes. Transcriptome analysis was then conducted through RNA-seq analysis by Miseq, which revealed 16 differentially-expressed transcripts between the two sub-populations. In the upper-layer, the highly expressed transcripts that were homologous to immune-related genes that suggest hemocytes from this layer may play as the regulator of immune system and control the action of other cells to eliminate pathogen. On the other hand, transcripts that were highly expressed in the lower-layer were homologous to the antimicrobial peptide (AMP) crustin, which supports that hemocytes on this layer have granules as crustins are normally secreted from hemocyte granules. The high expression of crustin in the lower-layer also provides insight on the mechanism of the anti-microbial function, where hemocytes produce and store AMPs in its granules. These differentially expressed genes are potential hemocyte molecular markers, and among them we identified one of the highly expressed genes in the hemocytes from the upper-layer (c11736-g1) to be a promising candidate molecular marker predicted to be a surface molecule, which is a common characteristic for molecular markers. (C) 2017 Elsevier Ltd. All rights reserved.