Induction and persistence of Mx protein in tissues, blood and plasma of Atlantic salmon parr, Salmo salar, injected with poly I:C.
ABSTRACT The expression of Mx transcripts and Mx protein was monitored at weekly intervals for 7 weeks, by qRT-PCR and immunohistochemistry, in the kidney, liver, gill and blood of Atlantic salmon parr following injection of poly I:C. Elevated levels of Mx transcripts compared to PBS injected control fish were found in the tissues at week 1. Background levels were then found up to week 7, with the exception of week 4 when high levels were again found in poly I:C injected fish as well as control fish. Immunostaining for Mx protein in the kidney, liver and gill of poly I:C injected fish was higher than in control fish from weeks 1-4, but little staining was found in the tissues of both poly I:C treated and control fish thereafter. Blood monocytes stained consistently in all fish, suggesting that this leucocyte type constitutively expressed Mx protein. From weeks 2-4, lymphocytes of both groups consistently stained for Mx protein but the consistency decreased at weeks 5-7. Staining of neutrophils was also inconsistent. Western blots of plasma showed an immunoreactive band of 76 kDa typical of salmon Mx protein. Semi-quantitative measurements of dot blots showed poly I:C injected fish to have higher levels of plasma Mx protein than controls on weeks 1-4 with very low levels on weeks 5-7. The results indicate that following induction of an interferon response with poly I:C, Atlantic salmon parr maintain elevated levels of Mx protein in tissues, leucocytes and blood plasma for about 4 weeks. Production of Mx protein by blood monocytes appears to be constitutive, though production by lymphocytes and neutrophils was less consistent.
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ABSTRACT: Microcystins (MCs) are secondary metabolites produced by cyanobacteria. Oxidative stress is considered the major cytotoxic mechanism of microcystin-LR (MCLR). Quercetin (QE) is a flavonoid that can eliminate reactive oxygen species (ROS) and elicit anti-inflammatory and anti-apoptotic effects. This study determined the regulatory effect of QE on the cytotoxicity and oxidative stress of Carassius auratus lymphocytes induced by 1 μg/L MCLR in vitro after 24 h. MCLR-mediated cytotoxicity and ROS formation in fish lymphocytes were suppressed by QE in a concentration-dependent manner. In addition, QE enhanced the endogenous antioxidant defense system and the Bax/Bcl-2 ratio to protect fish lymphocytes against oxidative stress and apoptosis induced by MCLR. Glutathione levels and catalase activities increased by approximately 3.9- and 2-fold, respectively, in the QE treatment group (1000 μg/L) compared with the MCLR treatment group. The percentage of apoptosis in the only MCLR treatment group was 59% whereas that in the control group was 23%. The percentage of apoptosis in the high-dose QE treatment group (1000 μg/L) was 29%, lower by nearly half compared with the only MCLR treatment group. QE (1000 μg/L) effectively inhibited the expression of caspase-3 protein by nearly 43% compared with the only MCLR treatment group. The results obtained clearly indicate that QE can effectively prevent MCLR-induced immunotoxicity by eliminating oxidative stress and blocking the mitochondrial apoptotic pathway in fish lymphocytes.Fish & Shellfish Immunology 04/2014; · 3.03 Impact Factor
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ABSTRACT: Nodularin (NOD) is a hazardous material widely detected in water blooms. Fish immune cells are extremely vulnerable to NOD-induced oxidative stress. Oligomeric proanthocyanidin complex (OPC), extracted from grapeseed, was used as an antioxidant to eliminate reactive oxygen species and prevent apoptotic effects. Carassius auratus lymphocytes were treated with different concentrations (0, 10, 100, and 1000μg/L) of OPC and a constant dose (100μg/L) of NOD for 12h in vitro. OPC inhibited mitosis by decreasing intracellular levels of oxidative stress, regulating antioxidant enzymes (CAT, SOD, GPx, GR, and GST), mediating bcl-2 family proteins, and deactivating caspase-3. Glutathione (GSH) levels in group V (NOD 100μg/L; OPC 1000μg/L) showed a twofold increase compared with corresponding levels in group II (NOD 100μg/L). Structure parameters of NOD and NOD-GSH were calculated using SYBYL 7.1 software. ClogP and HINK logP values of NOD-GSH decreased by 10.4- and 2.3-fold, respectively, compared with corresponding values of NOD. OPC-stimulated GSH can lower the lipophilicity and polarity of NOD. OPC, as a protective agent, can alleviate NOD-induced toxicity in C. auratus lymphocytes by regulating oxidative stress and inducing NOD-GSH detoxification.Journal of hazardous materials 04/2014; 274C:247-257. · 4.33 Impact Factor
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ABSTRACT: Infectious pancreatic necrosis virus (IPNV) causes high incidence of disease in salmonids during the first period after SW transfer. During this period as well as during periods of stress, cortisol levels increase and indications of a relationship between IPNV susceptibility and cortisol have been suggested. The intestine is an entry route and a target tissue for IPNV displaying severe enteritis and sloughing of the mucosa in infected fish. The mechanisms behind effects of the virus on the intestinal tissue and the impact of cortisol on the effect remain unclear. In the present study, Atlantic salmon post smolts treated with or without slow release cortisol implants were subjected to a cohabitant IPNV challenge. Analysis of genes and proteins related to the innate and acquired immune responses against virus was performed 6 days post-challenge using qPCR and immunohistochemistry. An increased mRNA expression of anti-viral cytokine interferon type I was observed in the proximal intestine and head kidney as a response to the viral challenge and this effect was suppressed by cortisol. No effect was seen in the distal intestine. T-cell marker CD3 as well as MHC-I in both intestinal regions and in the head kidney was down regulated at the mRNA level. Number of CD8α lymphocytes decreased in the proximal intestine in response to cortisol. On the other hand, mRNA expression of Mx and IL-1β increased in the proximal intestine and head kidney in IPNV challenged fish in the presence of cortisol suggesting that the immune activation shifts in timing and response pathway during simulated stress. The present study clearly demonstrates that IPNV infection results in a differentiated epithelial immune response in the different intestinal regions of the Atlantic salmon. It also reveals that the epithelial immune response differs from the systemic, but that both are modulated by the stress hormone cortisol.PLoS ONE 05/2014; 9(5):e94288. · 3.53 Impact Factor