School of Marine and Atmospheric Sciences, Stony Brook University, Stony Brook, NY, 11794, United States.
School of Marine and Atmospheric Sciences, Stony Brook University, Stony Brook, NY, 11794, United States.
Fish Shellfish Immunol. 2018 Jul;78:18-25. doi: 10.1016/j.fsi.2018.04.019. Epub 2018 Apr 7.
Hemocytes associated with the mucus lining of pallial (mantle, gill) surfaces of the oyster Crassostrea virginica have been recently suggested to facilitate infection by the Alveolate parasite Perkinsus marinus by mediating the uptake and dispersion of parasite cells. These "pallial hemocytes", which are directly exposed to microbes present in surrounding seawater, are able to migrate bi-directionally between mucosal surfaces and the circulatory system, potentially playing a sentinel role. Interestingly, P. marinus was shown to increase trans-epithelial migration of hemocytes suggesting it may regulate cell motility to favor infection establishment. The purpose of this study was to investigate the effect of P. marinus on hemocyte motility and identify specific molecular mechanisms potentially used by the parasite to regulate hemocyte migration. In a first series of experiments, various components of P. marinus (live P. marinus cells, extracellular products, fragments of P. marinus cell membrane, membrane-modified live P. marinus cells, heat-killed P. marinus) along with components of the opportunistic bacterial pathogen Vibrio alginolyticus (bacterial cells and extracellular products) were investigated for their effects on hemocyte motility. In a second series of experiments, inhibitors of specific molecular pathways involved in motility regulation (Y-27632: inhibitor of Rho-associated protein kinase, RGDS: integrin inhibitor, CK-666: Arp2/3 inhibitor) were used in conjunction with qPCR gene expression experiments to identify pathways regulated by P. marinus exposure. Results showed a specific increase in hemocyte motility following exposure to live P. marinus cells. The increase in motility induced by P. marinus was suppressed by RGDS and CK-666 implicating the involvement of integrins and Arp2/3 in cell activation. Gene expression data suggest that Arp2/3 is possibly regulated directly by an effector produced by P. marinus. The implications of increased hemocyte motility prompted by P. marinus during the early stage of the infection process are discussed.
牡蛎（贻贝、鳃）表面黏液衬里相关的血细胞最近被认为通过介导寄生虫细胞的摄取和扩散来促进有孔虫寄生虫海栖派琴虫的感染。这些“鳃血细胞”直接暴露于周围海水中存在的微生物，能够在黏膜表面和循环系统之间双向迁移，可能发挥哨兵作用。有趣的是，海栖派琴虫被证明增加了血细胞的跨上皮迁移，表明它可能调节细胞迁移性以利于感染建立。本研究旨在研究海栖派琴虫对血细胞迁移性的影响，并确定寄生虫可能用于调节血细胞迁移的特定分子机制。在一系列实验中，研究了海栖派琴虫的各种成分（活海栖派琴虫细胞、细胞外产物、海栖派琴虫细胞膜片段、膜修饰的活海栖派琴虫细胞、热灭活的海栖派琴虫）以及机会性病原体溶藻弧菌的成分（细菌细胞和细胞外产物）对血细胞迁移性的影响。在一系列实验中，使用了参与运动调节的特定分子途径抑制剂（Rho 相关蛋白激酶抑制剂 Y-27632、整合素抑制剂 RGDS、Arp2/3 抑制剂 CK-666），并结合 qPCR 基因表达实验，鉴定了海栖派琴虫暴露调节的途径。结果表明，暴露于活海栖派琴虫细胞后，血细胞迁移性特异性增加。RGDS 和 CK-666 抑制了海栖派琴虫诱导的迁移性增加，表明整合素和 Arp2/3 参与了细胞激活。基因表达数据表明，Arp2/3 可能直接受到海栖派琴虫产生的效应物的调节。讨论了海栖派琴虫在感染早期过程中引起的血细胞迁移性增加的意义。
