In vitro immune-depression and anti-inflammatory activities of cantharidin on gilthead seabream (Sparus aurata) leucocytes activated by λ-carrageenan.

Cantharidin is a natural compound with known therapeutic applications in humans. The aim of this study was to investigate the in vitro effects of cantharidin on gilthead seabream (Sparus aurata) head kidney leucocytes (HKL) stimulated with λ-carrageenan. HKLs were incubated for 24 h with cantharidin (0, 2.5 and 5 µg mL-1) and λ-carrageenan (0 and 1000 µg mL-1). The results showed that HKL viability only decreased by 15.2% after incubated with 5 µg mL-1 of cantharidin and λ-carrageenan. Cantharidin increased the peroxidase activity of HKLs only when incubated in combination with λ-carrageenan. Besides this, cantharidin inhibited the respiratory burst and phagocytic activities. Furthermore, cantharidin induced morphological changes in HKLs (apoptotic and vacuolization signs) that were enhanced when incubated with λ-carrageenan. Considering the analysis of the selected gene expression studied in HKLs [NF-κB subunits (rela, relb, crel, nfkb1, nfkb2), proinflammatory cytokines (il1b, tnfa), anti-inflammatory cytokines (il10, tgfb) and caspases (casp1, casp3, casp8, casp9)], although λ-carrageenan up-regulated the expression of the proinflammatory gene il1b, λ-carrageenan and cantharidin down-regulated its expression in HKLs. In addition, cantharidin up-regulated casp3 and casp9 expression. The casp3 and casp9 gene expression was down-regulated while casp1 gene expression was up-regulated in HKLs incubated with both cantharidin and λ-carrageenan. All the effects of cantharidin are related to its inhibitory effect on protein phosphatases, which induce apoptosis at long exposure times, and minimize the effects of λ-carrageenan. The present results provide detailed insight into the immune-depressive and anti-inflammatory properties of cantharidin on immune cells, which could be of interest to the aquaculture sector.

Serum proteinogram of gilthead seabream (Sparus aurata) and European sea bass (Dicentrarchus labrax) as a new useful approach for detecting loss of haemostasis.

Proteinograms, a semiquantitative analytical method that separates proteins into multiple bands, have not been explored in teleosts for diagnostic or prognostic purposes. This study aimed to establish reference values for proteinograms in the serum of gilthead seabream (Sparus aurata) and European sea bass (Dicentrarchus labrax), two important farmed fish species in the Mediterranean region. Serum proteins were studied using SDS-PAGE, electropherogram, and HPLC-mass spectrometry. SDS-PAGE analysis revealed four major bands of proteins around 11, 25, 70, and 100 kDa in the serum of gilthead seabream and European sea bass. Electropherogram results showed that a protein with a molecular weight of 76.8 kDa was the most abundant protein in the serum of gilthead seabream, while a peak of 75.5 kDa was the most abundant in European sea bass. HPLC-mass spectrometry detected 87 proteins and 119 proteins in the serum of gilthead seabream and European sea bass, respectively, including α1-globulins, α2-globulins, ß-globulins, and γ-globulins. Notably, the albumin sequence was not detected in either of the two species. These results help to characterize the serum protein profile and to establish reference proteinograms for these two fish species. They also provide a basis for the development of novel approaches for the rapid detection of loss of haemostasis due to stress, health disorders or disease in farmed fish.

Short-term induced swimming activity enhanced innate immune parameters and antioxidant status of European eel (Anguilla anguilla).

The swimming activity, although an essential trait in the life cycle of fish, is still poorly understood in farmed fish. The current study aimed to investigate the impact of short-term induced swimming on the immune and antioxidant defence systems in European eel (Anguilla anguilla). Sixteen male yellow European eels (total length: 39.9 ± 0.7 cm; body weight: 108.8 ± 6.1 g) were individually placed in swimming flumes and divided into two groups: i) no swimming (n = 8); and ii) induced-swimming (n = 8) at 0.3 body lengths (BL)·s-1 for 7 h. Swimming resulted in a 2-fold lower cortisol concentration in plasma, whereas plasma glucose, lactate, and several immune-related parameters did not present variations between groups. Interestingly, swimming led to higher lysozyme, peroxidase, and protease activities in skin mucus, whereas bactericidal activity did not show differences among groups. Additionally, the gene expression of interleukin 1 beta showed an up-regulation in the skin of fish with induced swimming, while no differences were observed in the head-kidney or gills. Furthermore, modulation of the antioxidant status was observed in the liver and posterior skeletal muscle after induced swimming. Fish subjected to swimming showed lower lipid peroxidation and higher reduced glutathione levels, increasing the reduced/oxidized glutathione ratio. However, no variations in the antioxidant status were observed between groups in the anterior skeletal muscle. This study showed modulation of immune and oxidative stress markers in European eels upon short-term induced swimming compared to non-swimming fish.

Short-term swimming up-regulates pro-inflammatory mediators and cytokines in gilthead seabream (Sparus aurata).

Aerobic swimming exercise in fish has been shown to improve robustness of some species. However, the optimal conditions to be applied and the mechanisms underlying remain unknown. We investigated the effects of 6 h of induced swimming on the immune response of gilthead seabream (Sparus aurata), by analysing markers related to immune status in plasma, skin mucus, gills, heart and head-kidney. Forty fish were individually exercised in swim tunnels by applying different water currents: steady low (SL, 0.8 body lengths (BL) s-1), steady high (SH, 2.3 BL s-1), oscillating low (OL, 0.2/0.8 BL s-1) and oscillating high (OH, 0.8/2.3 BL s-1) velocities, including a non-exercised group with minimal water flow (MF, <0.1 BL s-1). Swimming conditions did not trigger a stress response or anaerobic metabolism, suggested by similar levels of cortisol, lactate, and glucose in plasma among groups. Blood haemoglobin and innate immune parameters in plasma and skin mucus also remained unaltered. However, decreased blood haematocrit was observed in fish swimming on the OL condition. Interestingly, gene expression analysis revealed that the OL condition led to the up-regulation of pro-inflammatory mediators (nfκb1 and mapk3) and cytokines (tnfα, il1ß and il6) in gills. A similar response occurred in heart, with an up-regulation of nfκb1, tnfα, il6 and cox2 in the OL condition. Gene expression of these cytokines was unaltered in the head-kidney. The inflammatory response in gills and heart of gilthead seabream triggered by the OL condition highlights the importance of establishing suitable rearing conditions to improve welfare of cultured fish.

In vitro effects of λ-carrageenin in the head-kidney leucocytes of gilthead seabream (Sparus aurata).

The λ-carrageenin is a sulphated mucopolysaccharide that has been used for decades to induce experimental inflammation in mammals. However, it has been little considered in fish. We studied the in vitro effects of λ-carrageenin on gilthead seabream (Sparus aurata L.) head-kidney leucocytes (HKLs). For this purpose, HKLs were incubated with serial concentrations (from 0 to 1,000 µg mL-1) of λ-carrageenin for 3, 6, 12 and 24 h to assess its influence on cell viability and morphology, cell activity and modulation of several selected inflammation-related genes. The viability results demonstrated that λ-carrageenin has no negative effects on HKLs. The respiratory burst activity and phagocytic ability of HKLs after being incubated with λ-carrageenin (100 and 1,000 µg mL-1) for 24 h were increased, whereas the phagocytic capacity was inhibited by the higher dose at the same experimental time compared with control samples. However, the peroxidase activity of HKLs was not changed by incubation with λ-carrageenin. According to transmission electron microscopy results, incubation of HKLs with the higher dose of λ-carrageenin appeared to activate the cells being evident different morphological changes without sign of cell death. Furthermore, up-regulation of three proinflammatory cytokines (il1b, tnfa, and il6) and down-regulation of anti-inflammatory genes (tgfb) were denoted in HKLs incubated with carrageenin. The present results provide a detailed approach to the effects of λ-carrageenin on fish leucocytes, which could have some impact on how we understand the response of these cells when inducing an inflammatory process in fish.

In vitro effects of cantharidin on gilthead seabream (Sparus aurata) head-kidney leucocytes.

Cantharidin is a toxic vesicant terpene used in folk and traditional medicine due to its various therapeutic effects. Since there are no previous data on the effect of cantharidin in fish, this study aimed to investigate the in vitro related-inflammatory effects of cantharidin in gilthead seabream (Sparus aurata L.) head-kidney leucocytes (HKLs). In the first experiment, the HKLs were incubated with 0, 5 and 10 µg mL-1 of cantharidin for 24 h to delimit its possible toxic effects. In a second experiment, leucocytes were incubated with ranging concentrations from 0 to 10 µg mL-1 for 3, 6, or 12 h. Cell viability was higher in acidophilic granulocytes than in monocytes/macrophages and lymphocytes. Cantharidin caused apoptosis as was evidenced by transmission electron microscopy. In addition, cantharidin produced a time- and dose-dependent decrease of respiratory burst and phagocytic activities in HKLs, while their peroxidase activity was increased at 24 h of incubation with 5 and 10 µg mL-1 of cantharidin. Different changes in the gene expression were observed after incubation with cantharidin. While the gene expression of tnfa, il1b and crel was up-regulated in HKLs, the nfkb1 and igmh genes were down-regulated in comparison to the expression found in control HKLs. Present results offer a first view of the possible effects and action mechanisms of cantharidin in HKLs, as well as its implication in the inflammatory process, which could be of interest not only for basic research but also in the aquaculture sector.

Ultrasonography and X-ray micro-computed tomography characterization of the effects caused by carrageenin in the muscle of gilthead seabream (Sparus aurata).

The current work aimed to carry out an in vivo study of the λ-carrageenin-induced inflammation in the skin of gilthead seabream (Sparus aurata). The fish were injected intramuscularly with phosphate-buffered saline (PBS, as control) or λ-carrageenin (1% in PBS), and the injection zone was evaluated by real-time ultrasonography (Vevo Lab, VisualSonics) at 1.5, 3, 6, 12, and 24 h post-injection (p.i.). Results demonstrated that the skin thickness was increased in fish injected with λ-carrageenin and sampled at 1.5, 3, and 6 h p.i. However, the skin thickness of the injected area decreased to the normal values in those fish sampled at 12 and 24 h p.i. In addition, fish injected with λ-carrageenin and analysed at 1.5, 3, and 6 h p.i. showed, in the underlying muscle at the injection place, several hyperechoic small foci surrounded by an anechoic area which were not observed in control fish. Furthermore, the fish were analysed by X-ray micro-computed tomography (micro-CT). The analysis of the micro-CT acquisitions revealed also a dark area in the place of the injection with λ-carrageenin at 1.5, 3, and 6 h. These areas were smaller in fish analysed at longer times (12 h p.i.) and were almost disappeared in fish sampled at 24 h p.i. These areas had an average density of -850 to -115 HU, which did not correspond with any tissue density of the rest of the body. Furthermore, similar dark areas at the injection zones were never observed in control fish. Present results support the use of both non-invasive techniques to study the inflammatory process in fish of commercial interest such as gilthead seabream.

Implication of adipocytes from subcutaneous adipose tissue and fatty acids in skin inflammation caused by λ-carrageenin in gilthead seabream (Sparusaurata).

The role of subcutaneous adipose tissue adipocytes and the effects of fatty acids on carrageenan-induced skin inflammation in gilthead seabream (Sparus aurata) were studied. Fish were injected intramuscularly with phosphate-buffered saline (control) or λ-carrageenin (1%), and skin samples collected at the injection site at 3 and 6 h post-injection (p.i.) were processed for histological study. In addition, the presence and levels of lipid classes, fatty acid methyl esters (FAME) and eicosanoids were evaluated in the skin samples obtained from the injected areas. Histological results indicated an increase in adipocyte area in fish sampled at 3 h p.i. with λ-carrageenin compared to fish in the control group. Furthermore, the frequency of adipocytes between 4500 and 5000 µm2 was increased at 6 h in the λ-carrageenin group compared to the control group. Analysis of lipid classes found that fish injected with λ-carrageenan showed increased free fatty acid (FFA) and sphingomyelin content at 3 and 6 h, respectively, compared to the control group. An increase in saturated fatty acids (SFA), n-6 polyunsaturated fatty acids (PUFA), and a decrease in the values of monounsaturated fatty acids (MUFA), n-3 PUFA and minor fatty acids were observed in fish skin at 6 h after λ-carrageenin injection, with respect to the values obtained in the control group. Regarding the analysis of eicosanoids, an increase in hydroxyeicosatetraenoic acid (5-HETE) was detected in the skin of fish at 6 h post-carrageenin injection compared to the control group. The presented results indicate the contribution of adipocytes and fatty acids in the development and regulation of the inflammatory response triggered by λ-carrageenin in gilthead seabream skin.

Implication of mucus-secreting cells, acidophilic granulocytes and monocytes/macrophages in the resolution of skin inflammation caused by subcutaneous injection of λ/κ-carrageenin to gilthead seabream (Sparus aurata) specimens.

To date, the mechanisms of inflammation have been poorly studied in fish of commercial interest, due to the lack of development of appropriate experimental models. The current study evaluated a local inflammation triggered by a polymeric carrageenin mixture (a mucopolysaccharide derived from the red seaweed Chondrus crispus) in the skin of gilthead seabream (Sparus aurata). Fish were injected subcutaneously with phosphate-buffered saline (as control) or λ/κ-carrageenin (1%), and skin samples from the injection sites were collected 1.5, 3 and 6 hr post-injection, processed for inclusion in paraplast and stained with haematoxylin-eosin, Alcian blue or periodic acid-Schiff. Furthermore, immunohistochemistry and expression analyses of several cells' markers and proinflammatory genes were also analysed in samples of the injected sites. Microscopic results indicated an increased number of skin mucus-secreting cells and acidophilic granulocytes in the skin of fish studied at 1.5 hr and 3 hr post-injection with carrageenin, respectively, with respect to the data obtained in control fish. Otherwise, both the gene expression of the non-specific cytotoxic cell marker (granzyme B, grb) and the proinflammatory cytokine (interleukin-1ß, il-1ß) were up-regulated at 1.5 hr in the skin of fish injected with carrageenin compared with the control fish, whilst the gene expression of acidophilic granulocyte markers (NADPH oxidase subunit Phox22 and Phox40, phox22 and phox40) was up-regulated at 3 and 6 hr in the carrageenin group, compared with the control group. In addition, the gene expression of myeloperoxidase (mpo) was also up-regulated at 6 hr in the skin of fish injected with carrageenin in comparison with control samples. The present results indicate the chronological participation of two important immune cells involved in the resolution of the inflammation in the skin of gilthead seabream.

Optimization of an Analytical Protocol for the Extraction of Microplastics from Seafood Samples with Different Levels of Fat.

Marine organisms are affected by the ubiquitous occurrence of microplastics (MPs) in the environment. Several protocols have been described to extract and quantify MPs in seafood, although their complex matrices, with high level of fat, can compromise the efficiency of MPs extraction. To solve this issue, the present study aimed to develop a detailed methodology suitable to process seafood samples with different levels of fat, namely fish and molluscs, from fresh and canned sources, including the immersive liquids from the cans. Sample digestion was tested using different solutions (10% KOH, 30% H2O2), temperatures (40 °C, 65 °C) and incubation times (24, 48, 72 h). For fat removal, three detergents (two laboratory surfactants and a commercial dish detergent) and 96% ethanol were tested, as well as the manual separation of fat. The methodology optimized in this study combined a digestion with 30% H2O2 at 65 °C, during 24 to 48 h, with a manual separation of the fat remaining after the digestion. All steps from the present methodology were tested in six types of polymers (PE-LD, PET, PE, AC, PS, and lycra), to investigate if these procedures altered the integrity of MPs. Results showed that the optimized methodology will allow for the efficient processing of complex seafood samples with different fat levels, without compromising MPs integrity (recoveries rate higher than 89% for all the polymers tested).

Acute inflammatory response in the skin of gilthead seabream (Sparus aurata) caused by carrageenin.

Although inflammation is a well-characterized process in mammals, few studies have dealt with the mechanisms involved in this process in fish. The present study evaluated the expression of inflammation-related genes in the skin of fish injected with carrageenin, which has previously been used in inflammatory models in mammals. In our case, fish were injected subcutaneously with PBS (as control) or carrageenin (1%), and skin samples from the injection site were collected 1.5, 3 and 6 h post-injection. The gene expression of inflammatory markers (csfr1, mhc-ii and phox40), several pro-inflammatory cytokines (il1b, tnfa, il6, il8 and il18) and other molecules related (such as myd88 and c-rel) were up-regulated at 1.5 and 3 h in fish injected with carrageenin compared with control levels. By contrast, the gene expression of anti-inflammatory molecules (nlrx1, nlrc5 isoform 1, ctsd and ctss) was down-regulated in fish injected with carrageenin and sampled 3 h post injection, again compared to the gene expression in control fish. According to our results, carrageenin can be considered not only a good stimulator to study skin inflammation in gilthead seabream but also this method might be use to study the modulation of fish inflammatory process caused by internal or external factors.

Role of mucosal immune response and histopathological study in European eel (Anguilla anguilla L.) intraperitoneal challenged by Vibrio anguillarum or Tenacibaculum soleae.

The external mucus layer that covers fish skin contains numerous immune substances scarcely studied that act as the first line of defence against a broad spectrum of pathogens. This study aimed to characterize and describe for the first time several humoral immune defence parameters in the skin mucus of the European eel (Anguilla anguilla) after intraperitoneal injection with Vibrio anguillarum or Tenacibaculum soleae. This study evaluated several immune-related enzymes and bactericidal activity against fish pathogenic bacteria in the skin mucus of European eels at 24, 48, and 72 h post-challenge. The results demonstrated that European eel skin mucus showed significant increments in peroxidase and lysozyme activity at 48 and 72 h after V. anguillarum challenge, compared to other experimental groups. In the case of antiprotease activity, an increase was observed at 24 h in the skin mucus of fish challenged with V. anguillarum compared to unchallenged fish, while this activity was undetected at 48 and 72 h. In contrast, protease activity had decreased at 48 and 72 h in the skin mucus of fish challenged with V. anguillarum compared to the unchallenged group. Regarding bactericidal activity, a high growth capacity of T. soleae was observed in the skin mucus of all experimental groups. Interestingly, the skin mucus from fish challenged with V. anguillarum exhibited increased bactericidal activity against this bacterium at 48 h, compared to unchallenged fish. Finally, severe histopathological alterations were observed in the gills and liver at the end of the trial (72 h), whereas the skin showed only an overspread presence of goblet cells in the challenged fish compared to unchallenged fish. The present results may give new insights into the mucosal immune system of this primitive species with potential applications in aquaculture.

Yarrowia lipolytica, health benefits for animals.

The yeast Yarrowia lipolytica has been industrially adopted for docosahexaenoic acid and eicosapentaenoic acid production under good manufacturing practices over 2 decades. In recent years, it has claimed attention for novel biotechnological applications, such as a functional feed additive for animals. Studies have demonstrated that this yeast is safe and has probiotic and nutritional properties for mammals, birds, fish, crustaceans, and molluscs. Animals fed Y. lipolytica enhanced productive and immune parameters, as well as modulated microbiome, fatty acid composition, and biochemical profiles. Additionally, some Y. lipolytica-derived compounds have improved productive performance, immune status, and disease resistance in animals. Therefore, the aim of this review is to identify and discuss research advances on the potential use of this yeast for animals of economic interest. Challenges, opportunities, and trends were identified and envisioned in the near future for this industrially produced yeast. KEY POINTS: • Yarrowia lipolytica has probiotic and nutritional effects in animals. • Lipase2, EPA, and ß-glucan from Y. lipolytica have health benefits for animals. • Y. lipolytica is envisioned in terrestrial and aquatic animal production systems.

Effects of subcutaneous injection of λ/κ-carrageenin on the immune and liver antioxidant status of gilthead seabream (Sparus aurata).

This study investigated the acute inflammatory response induced by subcutaneous injection of carrageenin (1%) or phosphate-buffered saline (control) in gilthead seabream (Sparus aurata). Skin mucus, serum, head kidney (HK) and liver were sampled at 1.5, 3 and 6 hr post-injection (p.i.) to determine the immune and antioxidant status of this fish species. The skin mucus of the carrageenin group showed increased superoxide dismutase and peroxidase activities, lysozyme abundance, bactericidal activity against Vibrio anguillarum and Photobacterium damselae, and total immunoglobulins compared with those of the control group. However, the carrageenin-injected fish sampled at 6 hr p.i. showed decreased protease activity in the skin mucus and peroxidase activity in the HK leucocytes compared with the control. Moreover, the carrageenin injection had no effects on the systemic immune system, but it reduced the liver catalase activities at both 3 and 6 hr in the carrageenin group relative to those in the control group. The expression levels of several proinflammatory and cell marker genes in the HK and liver were also determined. In the HK, the expression levels of interleukin-1ß and prostaglandin D synthase 1 were upregulated at 1.5 and 3 hr, respectively, in the carrageenin group compared with those in the control group. Contrarily, the expression of the NADPH oxidase subunit phox40 (an acidophilic granulocyte marker) in the carrageenin group at 6 hr was downregulated compared with that in the control group. These results suggested that subcutaneous injection of κ/λ-carrageenin in gilthead seabream triggered an acute skin inflammation characterized by the rapid recruitment of acidophilic granulocytes and the release of humoral mediators into the skin mucus.

In silico and gene expression analysis of the acute inflammatory response of gilthead seabream (Sparus aurata) after subcutaneous administration of carrageenin.

Inflammation is one of the main causes of loss of homeostasis at both the systemic and molecular levels. The aim of this study was to investigate in silico the conservation of inflammation-related proteins in the gilthead seabream (Sparus aurata L.). Open reading frames of the selected genes were used as input in the STRING database for protein-protein interaction network analysis, comparing them with other teleost protein sequences. Proteins of the large yellow croaker (Larimichthys crocea L.) presented the highest percentages of identity with the gilthead seabream protein sequence. The gene expression profile of these proteins was then studied in gilthead seabream specimens subcutaneously injected with carrageenin (1%) or phosphate-buffered saline (control) by analyzing skin samples from the injected zone 12 and 24 h after injection. Gene expression analysis indicated that the mechanisms necessary to terminate the inflammatory response to carrageenin and recover skin homeostasis were activated between 12 and 24 h after injection (at the tested dose). The gene analysis performed in this study could contribute to the identification of the main mechanisms of acute inflammatory response and validate the use of carrageenin as an inflammation model to elucidate these mechanisms in fish.

Unitary matrix approach for a precise voltage dependent characterization of reflective liquid crystal devices by average Stokes polarimetry.

Precise characterization of parallel-aligned liquid crystal on silicon microdisplays has an important impact in many advanced photonics applications. We show liquid crystal on silicon (LCoS) modeled as a non-absorbent reciprocal device. Combined with time-average Stokes polarimetry, LCoS enables us to demonstrate robust measurements across the whole applied voltage range for the retardance and its flicker, and also as a novelty for the director orientation. We obtain that the director orientation changes across the voltage range, especially at larger applied voltages. This is a small effect, but it may provide a deeper insight into the internal dynamics in the liquid crystal layer, and in sensitive phase-only applications will produce a coupling between amplitude and phase.

Influence of skin wounds on the intestinal inflammatory response and barrier function: Protective role of dietary Shewanella putrefaciens SpPdp11 administration to gilthead seabream (Sparus aurata L.).

The effects of skin wounds on the intestinal barrier function and the beneficial effects of the dietary administration of Shewanella putrefaciens (known as SpPdp11) in gilthead seabream (Sparus aurata L.) were studied. Two replicates of fish were fed a commercial diet (control, CON) or CON diet enriched with 109 cfu g-1 SpPdp11 (SP diet) for 30 days. After this time, half of the fish were sampled, while the others were injured below the lateral line (wounded fish, W) and fed the same diets for an extra week before sampling (CON + W and SP + W groups). The intestinal histology and gene expression of different genes relevant for the intestinal barrier function were studied. The results showed that injured fish had a disordered enterocyte nucleus disposition, a more intense infiltration of mixed leucocytes and a thicker lamina propria in the intestine compared to the control fish. However, the fish in the SP + W group did not present these pathological symptoms in the intestine. No significant variations in the number of goblet cells were detected among the different experimental groups. Pro-inflammatory cytokines (colony-stimulating factor receptor 1, CSF1R, myeloperoxidase, MPO and interleukin-1ß, IL-1ß), mucins (intestinal mucin, IMUC and mucin 2, MUC2), and immunoglobulin T heavy chain (IGHT) were up-regulated, while tight junction protein occludin was down-regulated in the intestine from fish of the CON + W group. Similarly, the dietary administration of SpPdp11 markedly depressed the gene expression of pro-inflammatory cytokines, MUC2 and IGHT, but increased the gene expression of anti-inflammatory cytokine transforming growth factor-ß1 (TGF-ß1) and the tight junction proteins tricellulin and occluding after wounding. In brief, the skin wounds provoked an intestinal inflammatory response that included changes in the mucus layer and tight junction disruptions. Besides this, preventive administration of SpPdp11 alleviated the intestinal dysfunctions caused by skin wounds in gilthead seabream.

The alleviation of skin wound-induced intestinal barrier dysfunction via modulation of TLR signalling using arginine in gilthead seabream (Sparus aurata L).

The present study sought to investigate the effect of arginine on the involvement of toll-like receptors (TLRs) in skin wound-induced intestinal barrier dysfunction in gilthead seabream (Sparus aurata L.). Two replicates of fish (n = 8) were fed a commercial diet (CON, total 2.75% arginine), CON diet enriched with 1% arginine (ARG1, total 3.65% arginine) and 2% arginine (ARG2, total 4.53% arginine) for 30 days. Half of the fish were sampled, whereas the others were injured and sampled 7 days post-wounding. The intestinal histology results showed that a more intense infiltration of mixed leucocytes was evident in the wounded fish, which was remarkably reduced in fish that were fed the ARG1 diet. Serum IgM levels were significantly higher in the ARG1 group than levels in the CON group at 7 days post-wounding. Compared with the fish in the CON group after wounding, dietary administration of 1% arginine markedly downregulated the gene expression of TLRs (TLR2 and TLR5), MyD88, and proinflammatory cytokines (CSF1R, IL-1ß, and TNFα), but significantly enhanced the gene expression of IκBα, the anti-inflammatory cytokine TGF-ß1, and tight junction proteins (tricellulin and occludin) in wounded fish. Furthermore, the ARG2 diet demonstrated no additional benefits on intestinal cells, compared to both the ARG1 and the CON diets, and it even appeared to induce negative effects. In summary, dietary administration of 1% arginine significantly inhibited intestinal inflammatory response and tight junction disruption in skin-wounded gilthead seabream by modulating TLR signalling in the intestine.

Skin wound healing in gilthead seabream (Sparus aurata L.) fed diets supplemented with arginine.

Dietary administration of arginine on the wound healing process of gilthead seabream was studied. Two replicates of fish (n = 8) were fed with either a commercial diet [control diet (CON), no arginine added] and the CON diet supplemented with 1% arginine (ARG1) or with 2% arginine (ARG2) for 30 days. Afterward, half of the fish were sampled while the other half were injured and continued to be fed the same diet for an extra week. Results by image analysis showed that the wound closure rate was significantly improved in fish that were fed the ARG1 diet, compared with those in the CON group. After seven days of wound healing, the aminotransferase and creatine kinase levels in the serum and the protease and peroxidase activities in the skin mucus were down-regulated, while the immunoglobulin M level in the skin mucus was up-regulated in the ARG1 group after wounding and in the CON group before wounding. Compared with the CON diet, the ARG1 diet remarkedly depressed the gene expression of mpo, il-8, and tnf-α, and enhanced the gene expression of tgf-ß1, igf-1, pcna, krt2, mmp9, fn1α, and colIα and the antioxidant enzyme cat in the skin tissues after wounding. Furthermore, compared with both the ARG1 and the CON groups, negative effects of the ARG2 diet on wound healing were demonstrated. In conclusion, a 1% arginine supplementation facilitates skin wound healing and prevents a systemic inflammation reaction by alleviating the inflammatory response and enhancing the re-epithelialization and ECM biosynthesis in skin wound sites.

Combining average molecular tilt and flicker for management of depolarized light in parallel-aligned liquid crystal devices for broadband and wide-angle illumination.

We demonstrate a complete semiphysical and analytical model describing the angular and wavelength dependencies not only of retardance, but also its flicker, in parallel aligned liquid crystal (PA-LC) devices. It relies on the fitting of the molecules' equivalent tilt angle as a function of applied voltage. The wide range of calculations it offers without requiring extensive characterization makes the model unique. We focus on PA-LCoS application as a polarization state generator across the visible spectrum and for a wide range of incidence angles. This approach offers novel capabilities for managing arbitrary states of both full and partial polarization. To highlight the richness of situations with PA-LCoS devices, we provide results for two different digital addressing sequences producing different levels of flicker.