ABSTRACT
In vertebrates, the generation of superoxide reactive oxygen species (ROS) via activation of the Nox/Duox family of NADPH oxidases is a prototypical feature of the pathogen-induced defensive responses of activated professional phagocytes. To understand the role of the rainbow trout (Oncorhynchus mykiss) Phox oxidase from a phylogenetic and functional perspective we describe the cloning, sequencing and expression analysis of multiple NADPH components in cultured macrophages. Phylogenetic analyses support the notion of the emergence of Phox-related components before the diversification of basal euteleosts and add to the limited collection of teleost NADPH oxidases. Expression studies using lipopolysaccharide, polyinosine-polycytidylic acid and zymosan to mimic the onset of inflammatory responses in trout macrophages suggest differences in regulation of the NADPH complex throughout the maturation/differentiation period of culture and between different treatments.
Subject(s)
Adjuvants, Immunologic/pharmacology , Gene Expression Regulation , Macrophages/enzymology , NADPH Oxidases/immunology , Oncorhynchus mykiss/immunology , Amino Acid Sequence , Animals , Cells, Cultured , Gene Expression Regulation/drug effects , Lipopolysaccharides/pharmacology , Macrophages/immunology , Molecular Sequence Data , NADPH Oxidases/chemistry , NADPH Oxidases/genetics , Oncorhynchus mykiss/classification , Phylogeny , Poly I-C/pharmacology , Sequence Alignment , Time Factors , Zymosan/pharmacologyABSTRACT
BACKGROUND: The response of the trout, O. mykiss, head kidney to bacterial lipopolysaccharide (LPS) or active and attenuated infectious hematopoietic necrosis virus (IHNV and attINHV respectively) intraperitoneal challenge, 24 and 72 hours post-injection, was investigated using a salmonid-specific cDNA microarray. RESULTS: The head kidney response to i.p. LPS-induced inflammation in the first instance displays an initial stress reaction involving suppression of major cellular processes, including immune function, followed by a proliferative hematopoietic-type/biogenesis response 3 days after administration. The viral response at the early stage of infection highlights a suppression of hematopoietic and protein biosynthetic function and a stimulation of immune response. In fish infected with IHNV a loss of cellular function including signal transduction, cell cycle and transcriptional activity 72 hours after infection reflects the tissue-specific pathology of IHNV infection. attIHNV treatment on the other hand shows a similar pattern to native IHNV infection at 24 hours however at 72 hours a divergence from the viral response is seen and replace with a recovery response more similar to that observed for LPS is observed. CONCLUSION: In conclusion we have been able to identify and characterise by transcriptomic analysis two different types of responses to two distinct immune agents, a virus, IHNV and a bacterial cell wall component, LPS and a 'mixed' response to an attenuated IHNV. This type of analysis will lead to a greater understanding of the physiological response and the development of effective immune responses in salmonid fish to different pathogenic and pro-inflammatory agents.
