New insight into the biological activity of Salmo salar NK-lysin antimicrobial peptides.

NK-lysin is a potent antimicrobial peptide (AMP) with antimicrobial activity against bacteria, fungi, viruses, and parasites. NK-lysin is a type of granulysin, a member of the saposin-like proteins family first isolated from a pig's small intestine. In previous work, for the first time, we identified four variants of nk-lysin from Atlantic salmon (Salmo salar) using EST sequences. In the present study, we reported and characterized two additional transcripts of NK-lysin from S. salar. Besides, we evaluated the tissue distribution of three NK-lysins from S. salar and assessed the antimicrobial, hemolytic, and immunomodulatory activities and signaling pathways of three NK-lysin-derived peptides. The synthetic peptides displayed antimicrobial activity against Piscirickettsia salmonis (LF-89) and Flavobacterium psychrophilum. These peptides induced the expression of immune genes related to innate and adaptive immune responses in vitro and in vivo. The immunomodulatory activity of the peptides involves the mitogen-activated protein kinases-mediated signaling pathway, including p38, extracellular signal-regulated kinase 1/2, and/or c-Jun N-terminal kinases. Besides, the peptides modulated the immune response induced by pathogen-associated molecular patterns (PAMPs). Our findings show that NK-lysin could be a highly effective immunostimulant or vaccine adjuvant for use in fish aquaculture.

Adverse effects of fenpropathrin on the intestine of common carp (Cyprinus carpio L.) and the mechanism involved.

Fenpropathrin (FEN), a pyrethroid pesticide, is frequently detected in natural water bodies, unavoidable pose adverse effects to aquatic organisms. However, the harmful effects and potential mechanisms of FEN on aquatic species are poorly understood. In this study, common carp were treatment with FEN at 0.45 and 1.35 µg/L for 14 d, and the toxic effects and underlying mechanisms of FEN on the intestine of carp were revealed. RNA-seq results showed that FEN exposure cause a wide range of transcriptional alterations in the intestine and the differentially expressed genes were mainly enrichment in the pathways related to immune and metabolism. Specifically, FEN exposure induced pathological damage and altered submicroscopic structure of the intestine, elevated the levels of Bacteroides fragilis enterotoxin, altered the contents of claudin-1, occludin, and zonula occluden-1 (ZO-1), and causing injury to the intestinal barrier. In addition, inflammation-related index TNF-α in the serum and IL-6 in the intestinal tissues were generally increased after FEN exposure. Moreover, FEN exposure promoted an increase in reactive oxygen species (ROS), altered the levels of superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH), upregulated the contents of malondialdehyde (MDA) in the intestines. The apoptosis-related parameter cytochrome c, caspase-9, and caspase-3 were significantly altered, indicating that inflammation reaction, oxidative stress, and apoptosis may be involved in the toxic mechanism of FEN on carp. Moreover, FEN treatment also altered the intestinal flora community significantly, which may affect the intestinal normal physiological function and thus affect the growth of fish. Overall, the present study help to clarify the intestinal reaction mechanisms after FEN treatment, and provide a basis for the risk assessment of FEN.

Fibrous matrices facilitate pleurocidin killing of wound associated bacterial pathogens.

Conventional wound infection treatments neither actively promote wound healing nor address the growing problem of antibacterial resistance. Antimicrobial peptides (AMPs) are natural defense molecules, released from host cells, which may be rapidly bactericidal, modulate host-immune responses, and/or act as endogenous mediators for wound healing. However, their routine clinical use has hitherto been hindered due to their instability in the wound environment. Here we describe an electrospun carrier system for topical application of pleurocidin, demonstrating sufficient AMP release from matrices to kill wound-associated pathogens including Acinetobacter baumannii and Pseudomonas aeruginosa. Pleurocidin can be incorporated into polyvinyl alcohol (PVA) fiber matrices, using coaxial electrospinning, without major drug loss with a peptide content of 0.7% w/w predicted sufficient to kill most wound associated species. Pleurocidin retains its activity on release from the electrospun fiber matrix and completely inhibits growth of two strains of A. baumannii (AYE; ATCC 17978) and other ESKAPE pathogens. Inhibition of P. aeruginosa strains (PAO1; NCTC 13437) is, however, matrix weight per volume dependent, with only larger/thicker matrices maintaining complete inhibition. The resulting estimation of pleurocidin release from the matrix reveals high efficiency, facilitating a greater AMP potency. Wound matrices are often applied in parallel or sequentially with the use of standard wound care with biocides, therefore the presence and effect of biocides on pleurocidin potency was tested. It was revealed that combinations displayed additive or modestly synergistic effects depending on the biocide and pathogens which should be considered during the therapy. Taken together, we show that electrospun, pleurocidin-loaded wound matrices have potential to be investigated for wound infection treatment.

Polysaccharides derived from Spirulina platensis inhibited Singapore grouper iridovirus by impeding the entry of viral particles.

Attributable to the rapid dissemination and high lethality of Singapore grouper iridovirus (SGIV), it has caused significant economic losses for marine fish aquaculture in China and Southeast Asian nations. Hence, there is an urgent need to find antiviral drugs that are both safe and effective. In this study, a novel heteropolysaccharide named Spirulina platensis polysaccharides (SPP) was purified and characterized from S. platensis. The molecular weight of SPP is 276 kDa and it mainly consists of Glc and Rha, followed by minor components such as Gal, Xyl, and Fuc. The backbone of SPP was determined to be →2) -ß-Rhap-(1 â†’ 4) -α-Fucp-(1 â†’ [2) -α-Rhap-(1] 2[→6)-α-Glcp-(1] 4[→ 4) -α-Glcp-(1] 8[→ 4) -ß-Glcp-(1]2→, with branches of ß-Galp, α-Xylp and α-Glcp. SPP significantly inhibited SGIV-induced cytopathic effects (CPEs), viral gene replication and viral protein expression. The antiviral mechanism of SPP was associated with the disruption of SGIV entry to host cells. Furthermore, it was not observed that SPP made statistically significant impact on the expression of interferon-related cytokines. Our results offered novel insights into the potential utilization of spirulina polysaccharides for combating aquatic animal viruses.

Difenoconazole disrupts carp intestinal physical barrier and causes inflammatory response via triggering oxidative stress and apoptosis.

As a common fungicide, difenoconazole (DFZ) is widespread in the natural environment and poses many potential threats. Carp makes up a significant proportion of China's freshwater aquaculture population and are vulnerable to the DFZ. Therefore, this study investigated the effects of DFZ (0.488 mg/L and 1.953 mg/L) exposure for 4 d on the intestinal tissues of carp and explored the mechanisms. Specifically, DFZ exposure caused pathological damage to the intestinal tissues of carp, reducing the expression levels of intestinal tight junction proteins, and leading to damage to the intestinal barrier. In addition, DFZ exposure activated the NF-κB signaling pathway, increasing the levels of pro-inflammatory factors (TNF-α, IL-1ß, IL-6) and decreasing the levels of anti-inflammatory factors (IL-10, TGF-ß1). As disruption of the intestinal barrier is closely linked to oxidative stress and apoptosis, we have conducted research in both areas for this reason. The results showed that DFZ exposure elevated reactive oxygen species in carp intestines, decreased antioxidant enzyme activity, and suppressed the expression of oxidative stress-related genes. TUNEL results showed that DFZ induced the onset of apoptosis. In addition, the expression levels of apoptosis-related genes and proteins were examined. Western blotting results showed that DFZ could upregulate the protein expression levels of Bax, Cytochrome C and downregulate the protein levels of Bcl-2. qPCR results showed that DFZ could upregulate the transcript levels of Bax, Caspase-3, Caspase-8 and Caspase-9 and downregulate the transcript levels of Bcl-2 transcript levels. This suggests that DFZ can induce apoptosis of mitochondrial pathway in carp intestine. In conclusion, DFZ can induce oxidative stress and apoptosis in carp intestine, leading to the destruction of intestinal physical barrier and the occurrence of inflammation. Our data support the idea that oxidative stress and apoptosis are important triggers of pesticide-induced inflammatory bowel illness.

Scorpionfish BPI is highly active against multiple drug-resistant Pseudomonas aeruginosa isolates from people with cystic fibrosis.

Chronic pulmonary infection is a hallmark of cystic fibrosis (CF) and requires continuous antibiotic treatment. In this context, Pseudomonas aeruginosa (Pa) is of special concern since colonizing strains frequently acquire multiple drug resistance (MDR). Bactericidal/permeability-increasing protein (BPI) is a neutrophil-derived, endogenous protein with high bactericidal potency against Gram-negative bacteria. However, a significant range of people with CF (PwCF) produce anti-neutrophil cytoplasmic antibodies against BPI (BPI-ANCA), thereby neutralizing its bactericidal function. In accordance with literature, we describe that 51.0% of a total of 39 PwCF expressed BPI-ANCA. Importantly, an orthologous protein to human BPI (huBPI) derived from the scorpionfish Sebastes schlegelii (scoBPI) completely escaped recognition by these autoantibodies. Moreover, scoBPI exhibited high anti-inflammatory potency towards Pa LPS and was bactericidal against MDR Pa derived from PwCF at nanomolar concentrations. In conclusion, our results highlight the potential of highly active orthologous proteins of huBPI in treatment of MDR Pa infections, especially in the presence of BPI-ANCA.

Cystic fibrosis is a genetic disorder that makes people produce unusually thick and sticky mucus that clogs their lungs and airways. This inevitably leads to recurring bacterial infections, particularly those caused by the Gram-negative bacterium Pseudomonas aeruginosa. Antibiotics are needed to treat these infections. However, over time most bacteria build modes of resistance to these drugs and, once multiple drug-resistant bacteria colonize the lung, very limited treatment options are left. Therefore, new therapeutic approaches are desperately needed. Notably, humans themselves express a highly potent antimicrobial protein called BPI (short for Bactericidal/permeability­increasing protein) that attacks Gram-negative bacteria, including multiple drug-resistant strains of P. aeruginosa. Unfortunately, many people with cystic fibrosis also generate antibodies that bind to BPI and interfere with its antimicrobial function. Faced with this conundrum, Holzinger et al. set out to find BPIs made by other animals which might not be recognized by human antibodies and also display a high potential to attack Gram-negative bacteria. Based on specific selection criteria, Holzinger et al. focused their attention on BPI made by scorpionfish, a type of venomous fish that live near coral reefs. Compared to other BPI proteins they investigated, the one produced by scorpionfish appeared to be the most capable of binding to P. aeruginosa via a prominent surface molecule exclusively found on Gram-negative bacteria. Furthermore, when Holzinger et al. tested whether the antibodies present in people with cystic fibrosis could recognize scorpionfish BPI, they found that the BPI completely evaded detection. The scorpionfish BPI was also able to pre-eminently attack P. aeruginosa. In fact, it was even able to potently kill drug-resistant strains of the bacteria that had been isolated from people with cystic fibrosis. This study suggests that scorpionfish BPI could serve as an alternative to antibiotics in people with cystic fibrosis that have otherwise untreatable bacterial infections. Drug-resistant bacteria which cause life threatening conditions are on the rise across the globe, and scorpionfish BPI could be a potential candidate to treat affected patients. In the future, animal experiments will be needed to explore how highly potent non-human BPIs function in whole living organisms.

Antibacterial Activity and Mechanisms of TroHepc2-22, a Derived Peptide of Hepcidin2 from Golden Pompano (Trachinotus ovatus).

Hepcidin, a cysteine-rich antimicrobial peptide, has a highly conserved gene structure in teleosts, and it plays an essential role in host immune response against various pathogenic bacteria. Nonetheless, few studies on the antibacterial mechanism of hepcidin in golden pompano (Trachinotus ovatus) have been reported. In this study, we synthesized a derived peptide, TroHepc2-22, from the mature peptide of T. ovatus hepcidin2. Our results showed that TroHepc2-22 has superior antibacterial abilities against both Gram-negative (Vibrio harveyi and Edwardsiella piscicida) and Gram-positive (Staphylococcus aureus and Streptococcus agalactiae) bacteria. Based on the results of a bacterial membrane depolarization assay and propidium iodide (PI) staining assay in vitro, TroHepc2-22 displayed antimicrobial activity by inducing the bacterial membrane depolarization and changing the bacterial membrane permeability. Scanning electron microscopy (SEM) visualization illustrated that TroHepc2-22 brought about membrane rupturing and the leakage of the cytoplasm for the bacteria. In addition, TroHepc2-22 was verified to have hydrolytic activity on bacterial genomic DNA in view of the results of the gel retardation assay. In terms of the in vivo assay, the bacterial loads of V. harveyi in the tested immune tissues (liver, spleen, and head kidney) were significantly reduced in T. ovatus, revealing that TroHepc2-22 significantly enhanced the resistance against V. harveyi infection. Furthermore, the expressions of immune-related genes, including tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin 1-ß (IL-1ß), IL-6, Toll-like receptor 1 (TLR1), and myeloid differentiation factor 88 (MyD88) were significantly increased, indicating that TroHepc2-22 might regulate inflammatory cytokines and activate immune-related signaling pathways. To summarize, TroHepc2-22 possesses appreciable antimicrobial activity and plays a vital role in resisting bacterial infection. The observation of our present study unveils the excellent application prospect of hepcidin as a substitute for antibiotics to resist pathogenic microorganisms in teleosts.

Enhancement of the Antioxidant Effect of Natural Products on the Proliferation of Caco-2 Cells Produced by Fish Protein Hydrolysates and Collagen.

A large amount of fish side streams are produced each year, promoting huge economic and environmental problems. In order to address this issue, a potential alternative is to isolate the high-added-value compounds with beneficial properties on human health. The objectives of this study were to determine the effect of hydrolyzed fish protein and collagen samples on cell proliferation, as well as to determine the specific influence of minerals and metals on this effect and whether dietary antioxidants can enhance cell proliferation. The results of hydrolyzed fish protein and collagen samples showed negative effects on Caco-2 cell proliferation at the highest concentrations tested. Moreover, the pre-treatment of these hydrolyzates with vitamin C and E, quercetin and resveratrol increased the proliferation of bioaccessible fractions of hydrolyzated fish protein and collagen samples compared to the bioaccessible fractions without pre-treatment. The highest mineral concentrations were found for P, Ca and Mg. The metals found in the pure hydrolyzates were As, Cd, Hg and Pb; however, they appeared at almost undetectable levels in bioavailable fractions. It can be concluded that the consumption of hydrolyzates of fish by-products is an interesting strategy for complying with EFSA recommendations regarding fish consumption while at the same time reducing fish waste.

NKHs27, a sevenband grouper NK-Lysin peptide that possesses immunoregulatory and antimicrobial activity.

As an effective and broad-spectrum antimicrobial peptide, NK-Lysin is attracted more and more attention at present. However, the functions and action mechanism of NK-Lysin peptides are still not comprehensive enough at present. In this study, a sevenband grouper (Hyporthodus septemfasciatus) NK-Lysin peptide, NKHs27, was identified and synthesized, and its biological functions were studied. The results indicated that NKHs27 shares 44.44%∼88.89% overall sequence identities with other teleost NK-Lysin peptides. The following antibacterial activity assay exhibited that NKHs27 was active against both Gram-negative and Gram-positive bacteria, including Staphylococcus aureus, Listonella anguillarum, Vibrio parahaemolyticus and Vibrio vulnificus. Additionally, NKHs27 showed a synergistic effect when it was combined with rifampicin or erythromycin. In the process of interaction with the L. anguillarum cells, NKHs27 changed the cell membrane permeability and retained its morphological integrity, then penetrated into the cytoplasm to act on genomic DNA or total RNA. Then, in vitro studies showed that NKHs27 could enhance the respiratory burst ability of macrophages and upregulate immune-related genes expression in it. Moreover, NKHs27 incubation improved the proliferation of peripheral blood leukocytes significantly. Finally, in vivo studies showed that administration of NKHs27 prior to bacterial infection significantly reduced pathogen dissemination and replication in tissues. In summary, these results provide new insights into the function of NK-Lysin peptides in teleost and support that NKHs27, as a novel broad-spectrum antibacterial peptide, has potential applications in aquaculture against pathogenic infections.

Structural characteristic and molecular docking simulation of fish protein-derived peptides: Recent updates on antioxidant, anti-hypertensive and anti-diabetic peptides.

Over the decade, fish protein-derived peptides (FPDP) have been evaluated for various biological activities including their mechanism of action through structure-activity relationship (SAR) and molecular simulation. SAR studies are known to provide the basic structural information of the active site which can be used for designing synthetic bioactive peptides for application in therapeutics and medicinal purposes. In light of the above discussion, this review discusses the mechanism of action and SAR of the FPDP with a focus on three widely studied bioactive properties including antioxidant, antihypertensive and anti-diabetic activities. The emphasis is given to the recently purified and identified FPDP from various seafood resources. A brief discussion has been made on their structural characteristics and mechanism of action towards antioxidant, angiotensin-I converting enzyme (ACE) inhibition, and dipeptidyl peptidase-IV (DPP-IV) inhibitory activities. Additionally, the importance and future perspective of SAR of food-derived bioactive peptides have been addressed.

A Non-Canonical Teleost NK-Lysin: Antimicrobial Activity via Multiple Mechanisms.

NK-lysin (NKL) is a family of antimicrobial proteins with an important role in innate and adaptive immunity. In this study, a non-canonical NK-lysin (NKLnc) was identified in the Japanese flounder (Paralichthys olivaceus), which shares low sequence identities (15.8-20.6%) with previously reported fish NKLs and was phylogenetically separated from the canonical NKLs in teleost. NKLnc expression was upregulated in flounder tissues during bacterial infection, and interference with NKLnc expression impaired the ability of flounder cells to eliminate invading bacteria. When expressed in Escherichia coli, NKLnc was detrimental to the host cells. P35, a peptide derived from the saposin B domain (SapB) of NKLnc, bound major bacterial surface molecules and killed both Gram-negative and Gram-positive bacteria by inflicting damage to bacterial cell structure and genomic DNA. The bactericidal activity, but not the bacteria-binding capacity, of P35 required the structural integrity of the alpha 2/3 helices in SapB. Furthermore, P35 induced the migration of flounder peripheral blood leukocytes, inhibited bacterial dissemination in fish tissues, and facilitated fish survival after bacterial challenge. Together our study reveals that NKLnc plays an important part in flounder immune defense, and that NKLnc peptide exerts an antimicrobial effect via multiple mechanisms by targeting both bacteria and fish cells.

Fish proteins as potential precursors of taste-active compounds: an in silico study.

BACKGROUND: Fish protein is a good source of amino acids and peptides with sensory properties. Theoretically, the type of protein affects the taste quality of the protein hydrolysates. To better use fish protein in the food ingredients industry, an in silico approach was adopted to evaluate the potential of fish protein to release taste-active compounds. RESULTS: Six types of protein from seven commercial fishes were screened from the Uniprot knowledge base. The results showed that a remarkable number of umami fragments presented in myosin and parvalbumin (PB), such as glutamic acid (Glu), aspartic acid (Asp), and Asp- and Glu- containing peptides, whereas sweet amino acids and bitter peptides (e.g., Pro- and Gly- containing peptides) were mainly found in collagen (CGI) in all fish samples. After the in silico proteolysis by papain, a difference in the profile of taste-active fragments was observed among the six types of proteins. Amino acids were the main hydrolysis products of these proteins, especially umami, sweet, and bitter amino acids, significantly contributing to the taste formation of protein hydrolysates. Besides, the myosin and CGI hydrolysates were abundant in taste active peptides both in types and quantities. CONCLUSION: Myosin is a promising protein source for producing umami fragments, and CGI seems to be a good precursor of sweet and bitter fragments. Different types of protein have an essential effect on the taste of protein hydrolysates. © 2022 Society of Chemical Industry.

LcCCL28-25, Derived from Piscine Chemokine, Exhibits Antimicrobial Activity against Gram-Negative and Gram-Positive Bacteria In Vitro and In Vivo.

Antimicrobial peptides (AMPs) are currently recognized as potentially promising antibiotic substitutes. Fish are an important seawater/freshwater medicinal biological resource, and the antimicrobial peptides and proteins that are key components of their innate immune systems are potential candidates for the development of novel antibacterial agents. The rainbow trout Oncorhynchus mykiss chemokine CK11 (omCK11), classified in the C-C motif chemokine ligand 27/28 (CCL27/28) family, is the only CC-type chemokine reported to play a direct antibacterial role in the immune response; however, its antibacterial domain remains unknown. In this study, we analyzed the structure-activity relationship of omCK11 and identified the antibacterial C-terminal domain. Additionally, we performed structure-function analyses of CCL27/28 proteins from different, representative freshwater and seawater fishes, revealing their shared C-terminal antibacterial domains. Surprisingly, a synthesized cationic peptide (named lcCCL28-25), derived from the large yellow croaker Larimichthys crocea CCL28, exhibited broad-spectrum and the most acceptable bactericidal activity, as well as antibiofilm activity and negligible hemolytic and cytotoxic activity in vitro. Additionally, lcCCL28-25 conferred a protective effect in the thighs of neutropenic mice infected with Staphylococcus aureus. SYTOX green fluorescence and electron microscopy experiments revealed that lcCCL28-25 was capable of rapidly destroying the integrity and permeability of the bacterial cell membrane. Overall, this study aided in the advancement of antibacterial CC-type chemokine research and also suggested a new strategy for exploring novel AMPs. Additionally, the efficacy of lcCCL28-25 in in vivo antibacterial activity in a mammalian model revealed that this compound could be a promising agent for the development of peptide-based antibacterial therapeutics. IMPORTANCE The primary function of chemokines has been described as recruiting and activating leukocytes to participate in the immune response. Some chemokines are also broad-spectrum antibacterial proteins in mammals. The Oncorhynchus mykiss chemokine CK11 (omCK11) is the first reported and currently the only CC-type antibacterial chemokine. The present study identified the antibacterial domain of omCK11. Structure-function analysis of various fish CCL27/28 proteins identified a novel antibacterial peptide (lcCCL28-25) from Larimichthys crocea CCL28 that exhibited broad-spectrum and the most acceptable bactericidal activity in vitro, as well as a protective effect in a Staphylococcus aureus infection mouse model. The antibacterial mechanisms included membrane disruption and permeation. This study advanced the field of antibacterial chemokine research in fish and also suggested a new strategy for exploring novel AMPs. The novel peptide lcCCL28-25 may prove to be an effective antibacterial agent.

Investigations on the Wound Healing Potential of Tilapia Piscidin (TP)2-5 and TP2-6.

Wound healing is a highly orchestrated process involving many cell types, such as keratinocytes, fibroblasts and endothelial cells. This study aimed to evaluate the potential application of synthetic peptides derived from tilapia piscidin (TP)2, TP2-5 and TP2-6 in skin wound healing. The treatment of HaCaT keratinocytes with TP2-5 and TP2-6 did not cause cytotoxicity, but did enhance cell proliferation and migration, which could be attributed to the activation of epidermal growth factor receptor signaling. In CCD-966SK fibroblasts, although TP2-5 (31.25 µg/mL) and TP2-6 (125 µg/mL) showed cytotoxic effects, we observed the significant promotion of cell proliferation and migration at low concentrations. In addition, collagen I, collagen III, and keratinocyte growth factor were upregulated by the peptides. We further found that TP2-5 and TP2-6 showed pro-angiogenic properties, including the enhancement of human umbilical vein endothelial cell (HUVEC) migration and the promotion of neovascularization. In a murine model, wounds treated topically with TP2-5 and TP2-6 were reduced by day 2 post-injury and healed significantly faster than untreated wounds. Taken together, these findings demonstrate that both TP2-5 and TP2-6 have multifaceted effects when used as topical agents for accelerating wound healing.

Oxidative stress suppression in C. elegans by peptides from dogfish skin via regulation of transcription factors DAF-16 and HSF-1.

Functional peptides were obtained via enzymatic hydrolysis of smooth dogfish (Mustelus canis) skin. The enzyme-assisted process was optimized to achieve high yield of smooth dogfish skin peptides (SDSP). Fractions of SDSP (MW < 2 kDa, 2-5 kDa, 5-10 kDa and >10 kDa) showed in vitro antioxidant activities. The peptides <2 kDa (SDSP<2 kDa) significantly improved motility, reduced ROS and H2O2 levels of Caenorhabditis elegans, and increased its resistance to oxidative stress compared to the other peptide fractions. In vivo function of SDSP<2 kDa could be explained by their capacity to increase the expression of stress-response genes. The enhanced resistance to oxidative stress mediated by SDSP<2 kDa was dependent on DAF-16 and HSF-1. The amino acid residues and sequences of SDSP<2 kDa were characterized and revealed a higher content of hydrophobic versus polar amino acid contents. This study (especially the in vivo investigation) explored new potent antioxidant peptides derived from dogfish skin.

Dual-agonist incretin peptides from fish with potential for obesity-related Type 2 diabetes therapy - A review.

The long-acting glucagon-like peptide-1 receptor (GLP1R) agonist, semaglutide and the unimolecular glucose-dependent insulinotropic polypeptide receptor (GIPR)/GLP1R dual-agonist, tirzepatide have been successfully introduced as therapeutic options for patients with Type-2 diabetes (T2DM) and obesity. Proglucagon-derived peptides from phylogenetically ancient fish act as naturally occurring dual agonists at the GLP1R and the glucagon receptor (GCGR) with lamprey GLP-1 and paddlefish glucagon being the most potent and effective in stimulating insulin release from BRIN-BD11 clonal ß-cells. These peptides were also the most effective in lowering blood glucose and elevating plasma insulin concentrations when administered intraperitoneally to overnight-fasted mice together with a glucose load. Zebrafish GIP acts as a dual agonist at the GIPR and GLP1R receptors. Studies with the high fat-fed mouse, an animal model with obesity, impaired glucose-tolerance and insulin-resistance, have shown that twice-daily administration of the long-acting analogs [D-Ala2]palmitoyl-lamprey GLP-1 and [D-Ser2]palmitoyl-paddlefish glucagon over 21 days improves glucose tolerance and insulin sensitivity. This was associated with ß-cell proliferation, protection of ß-cells against apoptosis, decreased pancreatic glucagon content, improved lipid profile, reduced food intake and selective alteration in the expression of genes involved in ß-cell stimulus-secretion coupling. In insulin-deficient GluCreERT2;ROSA26-eYFP transgenic mice, the peptides promoted an increase in ß-cell mass with positive effects on transdifferentiation of glucagon-producing to insulin-producing cells. Naturally occurring fish dual agonist peptides, particularly lamprey GLP-1 and paddlefish glucagon, provide templates for development into therapeutic agents for obesity-related T2DM.

Molecular characterization and antibacterial ability of galectin-3 and galectin-9 in Onychostoma macrolepis.

Galectins belong to the ß-galactoside binding protein family, which have conserved carbohydrate-recognition domains (CRDs) and participate in innate and acquired immunity in animals. In this study, two galectin genes were cloned from Onychostoma macrolepis, OmGal-3 (galectin-3) and OmGal-9 (galectin-9). The open reading frames (ORFs) of OmGal-3 and OmGal-9 contain 732 and 978 base pairs, encoding 243 and 325 amino acids, respectively. OmGal-3 contains a C-terminal CRD, but OmGal-9 contains an N-terminal CRD and a C-terminal CRD. Two galectins were expressed at varying levels in all tissues examined, with the liver showing the highest expression. The relative gene expression levels of OmGal-3 and OmGal-9 following Aeromonas hydrophila infection were significantly up-regulated in the liver and spleen, and OmGal-9 had a greater increase than OmGal-3. The recombinant OmGal-3 and OmGal-9 proteins (rOmGal-3 and rOmGal-9) were authenticated and verified by SDS-PAGE and western blotting. ROmGal-3 and rOmGal-9 agglutinated all tested bacteria, including 3 g-positive bacteria (Aeromonas hydrophila, Escherichia coli, and Vibrio parahaemolyticus) and 3 g-negative bacteria (Streptococcus agalactiae, Staphylococcus aureus, and Bacillus cereus) in vivo without Ca2+. ROmGal-3 showed strong binding both to gram-positive and gram-negative bacteria and OmGal-9 had a stronger binding activity against gram-positive bacteria. Furthermore, rOmGal-3 and rOmGal-9 exhibited dose-dependent binding capability to two classic pathogens associated molecular pattern (LPS and PGN) and two sugars (d-lactose and d-galactose), and rOmGal-3 has better binding activity at lower concentrations in LPS and PGN than rOmGal-3. The integrated analyses indicate that the two galectins probably play an important role in innate immune defense by binding to bacterial cells via the CRD domain against pathogen infection.

Hypouricemic, hepatoprotective and nephroprotective roles of oligopeptides derived from Auxis thazard protein in hyperuricemic mice.

The oligopeptides derived from Auxis thazard protein (ATO) are a class of small peptides with molecular weight <1 kDa and good bioactivity. This paper aimed to explore the hypouricemic, hepatoprotective, and nephroprotective effects of ATO and its potential mechanisms in hyperuricemia in mice induced by potassium oxonate. The results showed that ATO significantly reduced serum UA, serum creatinine levels, inhibited XOD and ADA activities in the liver (p < 0.05), and accelerated UA excretion by downregulating the gene expression of renal mURAT1 and mGLUT9 and upregulating the gene expression of mABCG2 and mOAT1. ATO could also reduce the levels of liver MDA, increase the activities of SOD and CAT, and reduce the levels of IL-1ß, MCP-1 and TNF-α. Histological analysis also showed that ATO possessed hepatoprotective and nephroprotective activities in hyperuricemic mice. Thus, ATO could reduce the serum UA level in hyperuricemic mice by decreasing UA production and promoting UA excretion from the kidney, suggesting that ATO could be developed as a dietary supplement for hyperuricemia treatment.

Effect of Scomberomorus niphonius peptide on the characteristics of resveratrol.

Resveratrol has a variety of physiological activities, but its bioavailability in the body is low. In this study, the interaction between the peptide SH, prepared from Scomberomorus niphonius, and resveratrol was judged by fluorescence spectroscopy. Then, SHa1 was obtained by the purification of SH, and its effect on the characteristics of resveratrol was studied. SHa1 interacted with resveratrol at 37 °C for 30 min to obtain the complex SHa1-R, which then showed an obviously stronger inhibition on B16 cells than resveratrol using the MTT assay after in vitro gastrointestinal digestion. The solubility and digestive stability of SHa1-R were higher than that of free resveratrol. The intestinal absorption rate of SHa1-R was also increased compared with resveratrol according to the non-inverted rat intestinal sac model. The structure of SHa1 was analyzed by UPLC, auto amino acid analysis, and UPLC-MS/MS. The molecular weight of SHa1 was mainly concentrated under 1000 Da, and it was rich in glutamic acid, aspartic acid, lysine, and leucine. Eighteen possible peptides were identified from SHa1. The results suggested that the peptide SHa-1 may help to increase the bioavailability of resveratrol by increasing the solubility, digestive stability and intestinal absorption of resveratrol, thereby promoting its inhibitory effect on B16 cells.

Exploiting of Secondary Raw Materials from Fish Processing Industry as a Source of Bioactive Peptide-Rich Protein Hydrolysates.

Developing peptide-based drugs are very promising to address many of the lifestyle mediated diseases which are prevalent in a major portion of the global population. As an alternative to synthetic peptide-based drugs, derived peptides from natural sources have gained a greater attention in the last two decades. Aquatic organisms including plants, fish and shellfish are known as a rich reservoir of parent protein molecules which can offer novel sequences of amino acids in peptides, having unique bio-functional properties upon hydrolyzing with proteases from different sources. However, rather than exploiting fish and shellfish stocks which are already under pressure due to overexploitation, the processing discards, regarded as secondary raw material, could be a potential choice for peptide based therapeutic development strategies. In this connection, we have attempted to review the scientific reports in this area of research that deal with some of the well-established bioactive properties, such as antihypertensive, anti-oxidative, anti-coagulative, antibacterial and anticarcinogenic properties, with reference to the type of enzymes, substrate used, degree of particular bio-functionality, mechanism, and wherever possible, the active amino acid sequences in peptides. Many of the studies have been conducted on hydrolysate (crude mixture of peptides) enriched with low molecular bioactive peptides. In vitro and in vivo experiments on the potency of bioactive peptides to modulate the human physiological functions beneficially have demonstrated that these peptides can be used in the prevention and treatment of non-communicable lifestyle mediated diseases. The information synthesized under this review could serve as a point of reference to drive further research on and development of functionally active therapeutic natural peptides. Availability of such scientific information is expected to open up new zones of investigation for adding value to underutilized secondary raw materials, which in turn paves the way for sustainability in fish processing. However, there are significant challenges ahead in exploring the fish waste as a source of bioactive peptides, as it demands more studies on mechanisms and structure-function relationship understanding as well as clearance from regulatory and statutory bodies before reaching the end user in the form of supplement or therapeutics.