Acute phase response and plasma lectins of rainbow trout (Oncorhynchus mykiss)

The innate immune system, including the acute phase response (APR), is an essential component of defense in all organisms and plays a critical role recognizing and eliminating infectious agents. The objectives of this research were to (1) identify acute phase proteins in trout that could be used as sensitive indicators of adverse health and (2) to describe the primary structure, expression and immunohistochemical localization of two plasma pattern recognition receptors (PRRs); rainbow trout ladderlectin (RTLL) and intelectin (RTInt). Following the induction of inflammation or injection with LPS/FIA, a plasma protein panel of eight increased or induced proteins (p9.5, p10.5, p24a, p24b, p24c, p25a, p36 and p37), one decreased (16d) and two (p28a, p28b) unaltered proteins was constructed. Proteins previously identified included, pre-cerebellin (p24a, p24b), transferrin (p37), and apolipoprotein (p10.5, p28a, p28b). The first plasma PRR characterized was RTLL. Two cDNA sequences (RTLL-1(504) and RTLL-2(546bp)) were obtained and identified as group VII mannose binding C-type lectins. The genomic sequence of RTLL encompassed 6 exons and 5 introns, with exon 2 encoding 14 amino acids which were exclusive to RTLL-2. Binding assays and Western blots demonstrated multiple, 16-18kDa, isoforms of RTLL which bound chitin and intact bacteria. A second PRR (RTInt) was a 37kDa protein which exhibited calcium-dependent binding to N-acetylglucosamine and mannose conjugated matrices and bound similar relevant intact bacteria and chitin. A 975bp cDNA sequence obtained from the liver and spleen encoded a 325 amino acid secretory protein with homology to human, murine and carp intelectins. Both RTLL and RTInt were localized to the cytoplasm of branchial and intestinal epithelial cells, polymorphonuclear leukocytes, monocytes/macrophages, and lymphocytes found in blood vessels, hepatic sinusoids, renal interstitium, mucosal epithelium and sub-mucosa of normal intestine. Clinically diseased rainbow trout tissue had increased immunohistochemical staining compared to healthy fish. Both lectins were detected in extra-cellular spaces surrounding bacteria, fungi and protozoa and within inflammatory leukocytes of infected tissues. This research has added to the known range of acute phase reactants in rainbow trout; has characterized the biochemical and binding properties of two PRRs and has expanded the understanding of the innate defense of teleost fish.