Effects of Dietary Fish Meal Replacement with Alternative Protein Ingredients and Their Combinations on Growth Performance, Feed Utilization, Fillet Composition, and Biochemical Parameters of Red Seabream (Pagrus major).

The experiment was conducted to evaluate alternative protein ingredients in a low-fish meal (FM) diet for red seabream (Pagrus major). Twelve experimental diets were formulated. Control diet (CON) was designed to contain 60% FM. Other experimental diets were formulated by replacing 50% of FM from the CON with soy protein concentrate (SPC), corn gluten (CG), meat meal (MM), and/or chicken byproduct meal (CBM). Four diets were designed including one of SPC, CG, MM, or CBM as FM replacer and designated as SPC, CG, MM, and CBM. Six other diets were formulated by adding two ingredients as SPC and CG, SPC and MM, SPC and CBM, CG and MM, CG and CBM, or MM and CBM, and designated as SCG, SMM, SCM, CMM, CCM, and MCM, respectively. The 12th diet (MIX) was formulated by including SPC, CGM, MM, and CBM. Triplicate fish groups (50.2 ± 0.1 g) were hand-fed for 12 weeks. Weight gain (WG) of fish was significantly improved by MM and MCM diets compared to CG, SCG, CMM, and CCM diets. WG of CON, SPC, CM, SMM, SCM, and MIX groups were comparable with MM and MCM groups. The lowest WG was observed in CG and CMM groups. Feed efficiency (FE) was significantly higher in MM group compared to SPC, CG, SGC, and CMC groups. FE of MCM group was significantly higher than CG and SCG groups. Fillet linolenic acid (C18:2n-6) level in CG group was significantly higher than CON, MM, CM, SCM, CCM, and MCM groups. Serum lysozyme activity was significantly higher in MCM and MIX groups. Therefore, a high level of dietary CG reduces the growth performance and feed utilization of red seabream. A mixture of MM and CBM seems to be more efficient in replacing FM from red seabream diet.

Immuno-stimulatory effects of sulfated polysaccharides isolated from Codium fragile in olive flounder, Paralichthys olivaceus.

Sulfated polysaccharides isolated from Codium fragile have been previously demonstrated to possess immune-stimulating effects on murine cell lines and the fraction F2 (F2) isolated by ion exchange chromatography was the most effective. In this study, the effects of the fraction F2 were evaluated on the expressions of immune genes including IL-1ß, TNF-α, IL-8, IFN-γ and lysozyme in vitro and in vivo as well as lysozyme and complement activities in serum of olive flounder, Paralichthys olivaceus. In vitro, these gene expressions were up-regulated by F2 in head kidney cells. In vivo, IL-1ß and IL-8 gene expressions were up-regulated in peritoneal cells, head kidney, liver, gill and spleen, while TNF-α, IFN-γ and lysozyme gene expressions were mostly up-regulated but varied depending on tissue types or time points. Indeed, lysozyme and complement activities in serum were increased. Overall, these results indicate that the sulfated polysaccharides from C. fragile have immuno-stimulatory effects on olive flounder and may be used to enhance immunity during aquaculture.

Immune-Enhancement and Anti-Inflammatory Activities of Fatty Acids Extracted from Halocynthia aurantium Tunic in RAW264.7 Cells.

Halocynthia aurantium, an edible ascidian species, has not been studied scientifically, even though tunicates and ascidians are well-known to contain several unique and biologically active materials. The current study investigated the fatty acid profiles of the H. aurantium tunic and its immune-regulatory effects on RAW264.7 macrophage cells. Results of the fatty acid profile analysis showed a difference in ratios, depending on the fatty acids being analysed, including those of saturated fatty acids (SFA), monounsaturated fatty acids (MUFA), and polyunsaturated fatty acids (PUFA). In particular, omega-3 fatty acids, such as eicosatrienoic acid n-3 (ETA n-3), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), were much higher than omega-6 fatty acids. Moreover, the H. aurantium tunic fatty acids, significantly and dose-dependently, increased the NO and prostaglandin E2 (PGE2) production in RAW264.7 cells, for immune-enhancement without cytotoxicity. In addition, these fatty acids regulated the transcription of immune-associated genes, including iNOS, IL-1ß, IL-6, COX-2, and TNF-α. These actions were activated and deactivated via Mitogen-activated protein kinase (MAPK)and NF-κB signaling, to regulate the immune responses. Conversely, the H. aurantium tunic fatty acids effectively suppressed the inflammatory cytokine expressions, including iNOS, IL-1ß, IL-6, COX-2, and TNF-α, in LPS-stimulated RAW264.7 cells. Productions of COX-2 and PGE2, which are key biomarkers for inflammation, were also significantly reduced. These results elucidated the immune-enhancement and anti-inflammatory mechanisms of the H. aurantium tunic fatty acids in macrophage cells. Moreover, the H. aurantium tunic might be a potential fatty acid source for immune-modulation.

Comparison of neuroprotective effects of extract and fractions from Agarum clathratum against experimentally induced transient cerebral ischemic damage.

UNLABELLED: CONTEXTS: Agarum clathratum (Laminariaceae), a typical brown algae, has been identified by National Plant Quarantine Service in Korea. The extract of A. clathratum has antioxidant activities. OBJECTIVE: We investigated the neuroprotective effects of crude-extract, ethyl acetate (EA)-, n-butanol (BU)-, dichloromethane (DCM)- and n-hexane (Hx)-fractions from A. clathratum on ischemic damage in the gerbil hippocampal CA1 region (CA1) after 5 min of transient cerebral ischemia. MATERIALS AND METHODS: Agarum clathratum was collected in Kangwon province (South Korea) and treated with 95% ethanol. The ethanol extract was suspended in distilled water and subjected to a series of partitions with EA, BU, DCM and Hx. Each of extract and fraction was orally administered with 50 mg/kg once a day for one week before ischemia--reperfusion (I-R). RESULT: In the crude-extract-, EA- and BU-fraction-treated ischemia groups, we found strong neuroprotection in the CA1--about 80-89% of CA1 pyramidal neurons survived. However, in the DCM- and Hx-fraction-treated ischemia groups, we did not find any significant neuroprotection. In addition, we observed changes in astrocytes and microglia in the ischemic CA1. In the crude-extract, EA- and BU-fraction-treated ischemia groups, the distribution pattern and activity of the glial cells were similar to that found in the sham group. DISCUSSION: Repeated supplements of crude-extract, EA- and BU-fractions of A. clathratum could protect neurons from I-R injury in the hippocampal CA1 induced by transient cerebral ischemia via decrease of glial activation.

Investigation of the formation mechanism of the pepper starch-piperine complex.

In this study, a new carrier for loading piperine was prepared using pepper starch, and its interaction mechanism was investigated. The porous pepper starch-piperine complex (PPS-PIP) showed higher loading efficiency (76.15 %) compared to the porous corn starch-piperine complex (PCS-PIP (52.34 %)). This may be ascribed to the hemispherical shell structure of porous pepper starch (PPS) compared to the porous structure of porous corn starch (PCS) based on the SEM result. PPS-PIP had smaller particle size (10.53 µm), higher relative crystallinity (38.95 %), and better thermal stability (87.45 °C) than PCS-PIP (17.37 µm, 32.17 %, 74.35 °C). Fourier transform infrared spectroscopy (FTIR) results implied that piperine not only forms a complex with amylose but may also be physically present in porous starch. This was demonstrated by the short-range order and X-ray type. Molecular dynamics simulations confirmed that hydrogen bonding is the primary interaction between amylose and piperine. Besides the formation of the amylose-piperine complex, some of the piperine is also present in physical form.

Quantification of Neuronal Cell-Released Hydrogen Peroxide Using 3D Mesoporous Copper-Enriched Prussian Blue Microcubes Nanozymes: A Colorimetric Approach in Real Time and Anticancer Effect.

Despite the effectiveness and selectivity of natural enzymes, their instability has paved the way for developing nanozymes with high peroxidase activity using a straightforward technique, thereby expanding their potential for multifunctional applications. Herein, meso-copper-Prussian blue microcubes (Meso-Cu-PBMCs) nanozymes were successfully prepared via a cost-effective hydrothermal route. It was found that the Cu-PBMCs nanozymes, with three-dimensional (3D) mesoporous cubic morphologies, exhibited an excellent peroxidase-like property. Based on the high affinity of Meso-Cu-PBMCs toward H2O2 (Km = 0.226 µM) and TMB (Km = 0.407 mM), a colorimetric sensor for in situ H2O2 detection was constructed. On account of the high catalytic activity, affinity, and cascade strategy, the Meso-Cu-PBMCs nanozyme generated rapid multicolor displays at varying H2O2 concentrations. Under optimized conditions, the proposed sensor exhibits a preferable sensitivity of 18.14 µA µM-1, a linear range of 10 nM-25 mM, and a detection limit of 6.36 nM (S/N = 10). The reliability of the sensor was verified by detecting H2O2 in spiked human blood serum and milk samples, as well as by detecting in situ H2O2 generated from the neuron cell SH-SY5Y. Besides, the Meso-Cu-PBMCs nanozyme facilitated the catalysis of H2O2 in cancer cells, generating •OH radicals that induce the death of cancer cells (HCT-116 colon cancer cells), which holds substantial potential for application in chemodynamic therapy (CDT). This proposed strategy holds promise for simple, rapid, inexpensive, and effective intracellular biosensing and offers a novel approach to improve CDT efficacy.

Molecular properties of water-unextractable proteoglycans from Hypsizygus marmoreus and their in vitro immunomodulatory activities.

Four proteoglycans were sequentially extracted from Hypsizygus marmoreus using 0.1 M NaOH (alkali-soluble proteoglycans [F1] and alkali-insoluble proteoglycans [F3]) and 0.1 M HCl (acid-soluble proteoglycans [F2] and acid-insoluble proteoglycans [F4]), and their structures and immunomodulatory activities were investigated. The proteoglycans were found to contain carbohydrates (19.8-82.4%) with various amounts of proteins (7.7-67.3%), and glucose was the major monosaccharide unit present, along with trace amounts of galactose. The molecular weights (Mw) and the radius of gyration (Rg) of these proteoglycans showed ranges of 16 × 10(4)-19,545 × 10(4) g/mol and 35-148 nm, respectively, showing significant variations in their molecular conformations. The backbones of F1 and F2 were mainly connected through a-(1→3), (1→4) and b-(1→6)-glycosidic linkages with some branches. The F1 and F2 proteoglycans significantly stimulated Raw264.7 cells to release nitric oxide (NO), prostaglandin E2 (PGE(2)) and various cytokines, such as IL-1ß, TNF-α and IL-6 by inducing their mRNA expressions.

Phosphine residues and physicochemical stability of Hwangtae after fumigation.

This study detected phosphine residues and the qualitative effect of phosphine fumigation on Hwangtae (yellowish-dried Alaska pollock). Four types of Hwangtae products commercially purchased were investigated to assess phosphine residue. Hwangtae was fumigated at both laboratory scale, at an aluminum phosphide rate of 33.6 g/m3, and large scale (1.68 g/m3) to evaluate phosphine residue and dissipation. Further, nutritional composition analyses between pre- and post-fumigated Hwangtae were conducted. The concentration of phosphine residues was lower than the detection limit (0.005 mg/kg) in all Hwangtae products. After fumigation in laboratory scale, phosphine residue was 2.47 mg/kg, and after fumigation in large scale, the residue was 3.25 mg/kg. After 3-d aeration in the open air, there was no residue detected from fumigated Hwangtae. Nutritional composition, including proximate, mineral, and amino acid compositions, did not differ (P > 0.05) between pre- and post-fumigated Hwangtae. Overall, Hwangtae did not demonstrate a phosphine residue problem after the proper aeration process, and phosphine did not alter the nutritional composition, suggesting the use of phosphine as a fumigant to protect Hwangtae from insect pests.

MT2 melatonin receptor immunoreactivity in neurons is very high in the aged hippocampal formation in gerbils.

Melatonin exerts many physiological functions via its G protein-coupled receptors. In the present study, we investigated age-related changes in MT2 melatonin receptor immunoreactivity and its levels in the gerbil hippocampus during normal aging. In the postnatal month 1 (PM 1) group, MT2 immunoreaction was well observed in neurons in all subregions of the gerbil hippocampus. In the PM 3 and 6 groups, MT2 immunoreactivity in neurons was decreased compared to that in the PM 1 group. Thereafter, MT2 immunoreactivity in neurons was increased. In the PM 18 and 24 groups, MT2 immunoreactivity in neurons was strong in all subregions of the gerbil hippocampus. In addition, the number of MT2 immunoreactive cells was lowest at PM 3 and highest at PM 24. From western blot analysis, age-dependent change pattern in MT2 level in the gerbil hippocampus was similar to the immunohistochemical result. These results indicate that MT2 immunoreactivity and levels are altered in the gerbil hippocampus during normal aging; lowest at young adult stage and highest at aged stage.

Relationship between oversulfation and conformation of low and high molecular weight fucoidans and evaluation of their in vitro anticancer activity.

Low and high molecular weight fucoidans (F(5-30K) and F(>30K)) were chemically modified through the addition of sulfate groups, and the effect of oversulfation on the in vitro anticancer activity was investigated. After the addition of sulfate groups, a considerable increase of 35.5 to 56.8% was observed in the sulfate content of the F(5-30K) fraction, while the sulfate content of the F(>30K) fraction increased to a lesser extent (from 31.7 to 41.2%). Significant differences in anticancer activity were observed between the oversulfated F(5-30K) and F(>30K) fractions, with activities of 37.3-68.0% and 20.6-35.8%, respectively. This variation in the anticancer activity of oversulfated fucoidan derivatives was likely due to differences in their sulfate content. The results suggest that the molecular conformation of these molecules is closely related to the extent of sulfation in the fucan backbones and that the sulfates are preferably substituted when the fucoidan polymers are in a loose molecular conformation.

Co-immunomodulatory Activities of Anionic Macromolecules Extracted from Codium fragile with Red Ginseng Extract on Peritoneal Macrophage of Immune-Suppressed Mice.

In this study we investigated the immune effects of oral administration of anionic macromolecules extracted from Codium fragile (CFAM) and red ginseng extract mixture on the peritoneal macrophage cells in immune-suppressed mice. Cyclophosphamide (CY) induces the immune-suppressed condition. CY-treated mice were orally fed with different concentrations of CFAM supplemented with red ginseng extract and the peritoneal macrophages collected. CY treatment significantly decreased the immune activities of peritoneal macrophages, compared to the normal mice. The administration of CFAM mixed with red ginseng extract significantly boosted the viability of macrophage cells and nitric oxide production of peritoneal macrophages. Further, the oral administration of CFAM mixed with red ginseng extract up-regulated the expression of iNOS, COX-2, and TLR-4 as well as cytokines such as IL-1ß, IL-6, TNF-α, and IFN-γ more than the red ginseng-treated group. This study showed that CFAM enhanced the immune activity of red ginseng extract in the peritoneal macrophage cells of immune-suppressed mice. Furthermore, CFAM might be used as a co-stimulant of red ginseng extract through the regulation of macrophage cells for the enhancement of human health and immunity.

Immune Enhancement Effects of Codium fragile Anionic Macromolecules Combined with Red Ginseng Extract in Immune-Suppressed Mice.

Codium fragile is an edible seaweed in Asian countries that has been used as a thrombolytic, anticoagulant, antioxidant, anti-inflammatory, and immune-stimulatory agent. Ginseng has also been known to maintain immune homeostasis and to regulate the immune system via enhancing resistance to diseases and microorganisms. In this study, anionic macromolecules extracted from C. fragile (CFAM) were orally administered with red ginseng extract (100 mg/kg body weight) to cyclophosphamide-induced immunosuppressed male BALB/c mice to investigate the immune-enhancing cooperative effect of Codium fragile and red ginseng. Our results showed that supplementing CFAM with red ginseng extract significantly increased spleen index, T- and B-cell proliferation, NK cell activity, and splenic lymphocyte immuneassociated gene expression compared to those with red ginseng alone, even though a high concentration of CFAM with red ginseng decreased immune biomarkers. These results suggest that CFAM can be used as a co-stimulant to enhance health and immunity in immunosuppressed conditions.

Molecular characteristics and immunomodulatory activities of water-soluble sulfated polysaccharides from Ulva pertusa.

Sulfated polysaccharides isolated from Ulva pertusa and fractionated using anion-exchange chromatography were investigated to determine their molecular characteristics and bioactivities. The crude and fractionated polysaccharides (F(1), F(2), and F(3)) were mainly composed of carbohydrates (59.9-65.9%), sulfates (11.6-15.3%), and uronic acid (7.30-16.4%) with small amounts of proteins (1.40-4.80%). Rhamnose (62.5-80.7%) was the major monosaccharide unit of these polysaccharides, with different levels of glucose (13.5-27.4%) and xylose (2.74-11.5%). The polysaccharides contained one or two major subfractions with weight-average molecular mass ranging from 51.1×10(3) to 1,690×10(3) g/mol. The relatively low in vitro anticancer activity of the polysaccharides (22.3-42.4%) suggested that they had little cytotoxicity against the cancer cell line used (AGS). On the other hand, the polysaccharides significantly stimulated Raw 264.7 cells, inducing considerable amounts of nitric oxide and various cytokines production, which suggested that they could be strong immunostimulators.

Taurine is a liver X receptor-α ligand and activates transcription of key genes in the reverse cholesterol transport without inducing hepatic lipogenesis.

SCOPE: Taurine, which is abundant in seafood, has antiatherogenic activities in both animals and humans; however, its molecular target has been elusive. We examined whether taurine could activate liver X receptor-α (LXR-α), a critical transcription factor in the regulation of reverse cholesterol transport in macrophages. METHODS AND RESULTS: Taurine bound directly to LXR-α in a reporter gene assay, time-resolved fluorescence resonance energy transfer analysis, and limited protease digestion experiment. Macrophage cells incubated with taurine showed reduced cellular cholesterol and induced medium cholesterol in a dose-dependent manner with the induction of ATP-binding cassette transporter A1 and G gene and protein expression. In hepatocytes, taurine significantly induced Insig-2a levels and delayed nuclear translocation of the sterol regulatory element-binding protein 1 (SREBP-1) protein, resulting in a dose-dependent reduction in the cellular lipid levels without inducing the expression of fatty acid synthesis genes. CONCLUSION: Taurine is a direct LXR-α ligand, represses cholesterol accumulation, and modulates the expression of genes involved in reverse cholesterol transport in macrophages, without inducing hepatic lipogenesis. The induction of Insig-2a suppressed the nuclear translocation of SREBP-1c.

In vitro and in vivo immunomodulatory activity of sulfated polysaccharides from Enteromorpha prolifera.

Water-soluble sulfated polysaccharides extracted from Enteromorpha prolifera and fractionated using ion-exchange chromatography (crude, F(1), F(2) and F(3) fractions) were investigated to determine their in vitro and in vivo immunomodulatory activities. The sulfated polysaccharides, especially the F(1) and F(2) fractions, stimulated a macrophage cell line, Raw 264.7, inducing considerable nitric oxide (NO) and various cytokine production via up-regulated mRNA expression. The in vivo experiment results show that the sulfated polysaccharides (the crude and F(2) fractions) significantly increased Con A-induced splenocyte proliferation, revealing their potential comitogenic activity. In addition, IFN-γ and IL-2 secretions were considerably increased by the F(2) fraction without altering the release of IL-4 and IL-5. This implies that the F(2) fraction can activate T cells by up-regulating Th-1 response and that Th-1 cells might be the main target cells of the F(2) fraction. These in vitro and in vivo results suggest that the sulfated polysaccharides are strong immunostimulators.

Butanol extract of Ecklonia cava prevents production and aggregation of beta-amyloid, and reduces beta-amyloid mediated neuronal death.

Beta-amyloid (Aß) is a major pathogenic peptide for Alzheimer's disease (AD) and is generated by the processing of amyloid precursor protein (APP). The Aß monomers aggregate into oligomeric and fibrillar forms which have been implicated as the toxic species inducing the neuronal dysfunction. Brown algae Ecklonia cava is known for its anti-oxidant and anti-inflammatory functions. Therefore, we tested the effect of E. cava extract on the production and aggregation of Aß peptides. The butanol extract of E. cava reduced Aß secretion from HEK293 cells expressing APP with Swedish mutation and increased soluble APPα and C-terminal fragment-α (CTFα), of which activity was similar to BACE (ß-site of APP cleaving enzyme) inhibitors. Furthermore, the extract inhibited Aß oligomerization, particularly mid-size oligomer formation, confirmed by the ultrastructural morphology. Congo red, thioflavin T assays, and electron microscopy showed that the extract inhibited Aß fibril formation effectively. Finally, the extract protected primary cortical neurons from various Aß-induced cell deaths, especially oligomer-induced death. Although further study is needed to test the effectiveness of the extract in vivo, our results demonstrate, for the first time, that the butanol extract of E. cava could be used as an anti-Aß agent for AD therapeutics.

Leptin's neuroprotective action in experimental transient ischemic damage of the gerbil hippocampus is linked to altered leptin receptor immunoreactivity.

Circulating leptin crosses blood-brain barrier to provide control of feeding behavior and energy balance. We investigated changes in leptin and leptin receptor (ObR) in the gerbil hippocampal CA1 region (CA1) after transient cerebral ischemia, and examined effects of leptin on ischemic damage. In vehicle-treated ischemia (vehicle-ischemia) group, the number of survived neurons in the CA1 was 16.4% compared to vehicle-treated sham (vehicle-sham) group; however, in 1 mg/kg leptin-treated ischemia (leptin-ischemia) group, 77.5% of neurons of the CA1 has survived. In the vehicle-sham group, weak leptin immunoreactivity was detected in CA1 neurons. From 4 days post-ischemia, moderate leptin immunoreactivity was expressed in CA1 neurons. In the leptin-ischemia group, leptin immunoreactivity at 5 days post-ischemia was higher than the sham group. ObR immunoreaction in the sham group was hardly detected in any cells. From 2 days post-ischemia, ObR immunoreaction was expressed in microglia, showing the highest immunoreactivity at 5 days post-ischemia. Microglial activation in the leptin-ischemia group was hardly detected at 5 days post-ischemia; however, astrocytes in the group were slightly increased compared to the vehicle-ischemia group. These suggest that treatment of leptin has neuroprotective effects against ischemic damage, showing that ObR immunoreactivity is distinctly changed in the ischemic CA1.

Difference of fibroblast growth factor receptor 1 expression among CA1-3 regions of the gerbil hippocampus after transient cerebral ischemia.

Fibroblast growth factors are important regulators of neuronal development. In this study, we observed fibroblast growth factor receptor 1 (FGFR1) immunoreactivity and its protein levels in the hippocampus proper (CA1-3 regions) of the gerbil at various time points after ischemia/reperfusion. In the sham-operated group, FGFR1 immunoreaction was not detected in the hippocampus proper. FGFR1 immunoreaction was first detected in non-pyramidal neurons in the CA1-3 region at 12h and 1day after ischemia/reperfusion. From 2days after ischemia/reperfusion, FGFR1 immunoreaction was found in astrocytes, not in microglial cells, in the CA1 region: FGFR1 immunoreactivity and the number of astrocytes were significantly increased at 5days post-ischemia. Western blot analysis revealed that FGFR1 protein levels were also increased from 1day after ischemia/reperfusion. These results indicate that increase of FGFR1 in astrocytes of the ischemic CA1 region may be associated with gliosis followed by delayed neuronal death.