Laboratory of Ecotoxicology, University of Le Havre, EA 3222, IFRMP 23, BP 540, 76058 Le Havre cedex, France.
Fish Shellfish Immunol. 2010 Feb;28(2):372-86. doi: 10.1016/j.fsi.2009.11.011. Epub 2009 Nov 26.
In bivalve molluscs, defence against pathogens mainly relies on fast tissue infiltration by immunocompetent hemocytes that migrate from circulating hemolymph to sites of infection, in order to deliver, in situ, an effective immune response. In the present work, we have investigated dynamics of hemocyte subpopulations motility by combining flow cytometry coupled to Coulter-type cell volume determination, Hoffman modulation contrast microscopy, time-lapse imaging and off-line analysis of cell shape changes. Our results revealed fast modifications of hemocyte aspect in vitro, with bidirectional transitions from spread outlines to condensed cell body morphologies, in the minute range. Amoeboid or non-amoeboid types of locomotion were observed, depending on the cell shapes and on the cell subtypes, with velocities reaching up to 30 mum min(-1). Correlations between motion profiles, Hemacolor staining and flow cytometry analysis on living cells help to propose a functional mussel hemocyte classification including the motile properties of these cells. In particular, basophils were shown to be involved in dynamic hemocyte-hemocyte interactions and in the constitution of aggregation cores. Physiological implications, in terms of immune response in organisms devoid of endothelium-closed vascular system, and potential applications of hemocyte motility studies for the development and the interpretation of experiments involving hemocytes in the field of marine ecotoxicology are discussed.
在双壳贝类软体动物中，防御病原体主要依赖于免疫活性血细胞的快速组织浸润，这些血细胞从循环的血淋巴迁移到感染部位，以便就地提供有效的免疫反应。在本工作中，我们通过结合流式细胞术与库尔特型细胞体积测定、霍夫曼调制对比显微镜、延时成像和细胞形状变化的离线分析，研究了血细胞亚群运动的动力学。我们的结果显示，体外血细胞的形态在数分钟内迅速发生变化，从伸展的轮廓到浓缩的细胞体形态发生双向转变。根据细胞的形状和细胞亚型，可以观察到阿米巴样或非阿米巴样的运动方式，速度可达 30 微米/分钟。运动谱、Hemacolor 染色和活细胞流式细胞术分析之间的相关性有助于提出一种功能性贻贝血细胞分类，包括这些细胞的运动特性。特别是，嗜碱性粒细胞被证明参与了动态血细胞-血细胞相互作用和聚集核心的形成。讨论了在没有内皮封闭血管系统的生物体中，免疫反应的生理意义，以及在海洋生态毒理学领域中涉及血细胞的实验中，血细胞运动研究的潜在应用。
