In-vitro immunomodulatory responses and antiviral activities of antimicrobial peptide octominin against fish pathogenic viruses.

Antimicrobial peptides (AMPs) are considered a novel approach to stimulate fish antiviral mechanisms for defense against a broad range of viral infections by enhancing immunomodulatory activities. Octominin is an AMP derived from the defense proteins of Octopus minor. In this study, preliminary screening of octominin against viral hemorrhagic septicemia virus (VHSV), infectious hematopoietic necrosis virus (IHNV), and infectious pancreatic necrosis virus (IPNV) was carried out. Moreover, immune responses upon octominin treatment and IHNV challenge were investigated using fathead minnow (FHM) cells. The CC50s of octominin for FHM and Chinook salmon embryo-214 (CHSE-214) cells were 2146.2 and 1865.2 µg/mL, respectively. With octominin treatment, EC50 resulted in 732.8, 435.1, and 925.9 µg/mL for VHSV, IHNV, and IPNV, respectively. The selectivity indices were 2.9, 4.9, and 2.0, respectively. The transcriptional analysis results demonstrated the induced transcription factors (Irf3; 143-fold, Irf7; 105-fold, and NF-κB; 8-fold), stress response gene (HspB8; 2-fold), and apoptosis functional gene (p53; 3-fold) in octominin treated (500 µg/mL) FHM cells for 48 h. Moreover, IHNV viral copy number was slightly decreased with the octominin treatment (500 µg/mL) in FHM cells. Overall results suggest that octominin could be a potential antiviral agent, although further studies are necessary to understand its mode of action and the mechanism of its antiviral activity.

Characterization and biocontrol efficacy of lytic phage (KPP-1) that infects multidrug resistant Klebsiella variicola.

Klebsiella variicola strain was identified from a natural water stream. Novel phage (KPP-1) infecting K. variicola was isolated and characterized. The biocontrol efficacy of KPP-1 against K. variicola-infected adult zebrafish was also investigated. The host K. variicola strain was resistant to six of the antibiotics tested and comprised the virulence genes kfuBC, fim, ureA, and Wza-Wzb-Wzccps. Morphological analysis by transmission electron microscopy revealed that KPP-1 has icosahedron head and tail structures. The latent period and burst size of KPP-1 were 20 min and 88 PFU per infected cell, respectively, at a multiplicity of infection of 0.1. KPP-1 was stable over a broad pH range (3-11), temperature (4-50 °C), and salinity (0.1-3%). KPP-1 inhibits the growth of K. variicola in vitro and in vivo. In the zebrafish infection model, treatment with KPP-1-infected K. variicola demonstrated 56% of cumulative survival. This suggests the possibility of developing KPP-1 as a potential biocontrol agent against multidrug-resistant K. variicola that belongs to the K. pneumoniae complex.

Hot Water Extract of Sasa borealis (Hack.) Makino & Shibata Abate Hydrogen Peroxide-Induced Oxidative Stress and Apoptosis in Kidney Epithelial Cells.

Sasa borealis (Hack.) Makino & Shibata or broad-leaf bamboo is famous for its richness of bioactive natural products and its uses in traditional medicine for its anti-inflammatory, diuretic, and antipyretic properties and preventive effects against hypertension, arteriosclerosis, cardiovascular disease, and cancer. The present study investigated the antioxidant activity of S. borealis hot water extract (SBH) and its effects in ameliorating hydrogen peroxide-induced oxidative stress, using an African green monkey kidney epithelial cell line (Vero). Known polyphenols in SBH were quantified by HPLC analysis. SBH indicated a dose-dependent increase for reducing power, ABTS+ (IC50 = 96.44 ± 0.61 µg/mL) and DPPH (IC50 = 125.78 ± 4.41 µg/mL) radical scavenging activities. SBH markedly reduced intracellular reactive oxygen species (ROS) generation in the Vero cells and increased the protective effects against H2O2-induced oxidative stress by reducing apoptosis. Other than the direct involvement in neutralizing ROS, metabolites in SBH were also found to induce NRF2-mediated production of antioxidant enzymes, HO-1, and NQO1. These findings imply that S. borealis hot water extract can be utilized to create nutraceutical and functional foods that can help to relieve the effects of oxidative stress in both acute and chronic kidney injury.

Moringa oleifera Hot Water Extract Protects Vero Cells from Hydrogen Peroxide-Induced Oxidative Stress by Regulating Mitochondria-Mediated Apoptotic Pathway and Nrf2/HO-1 Signaling.

The present study discloses the identification of phenolic compounds in Moringa oleifera hot water extract (MOH) and the evaluation of its antioxidant activity on H2O2-induced oxidative stress in Vero cells. Upon analysis, MOH was found to contain phenolic compounds and indicated 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS+) radical scavenging with IC50 values of 102.52 and 122.55 µg/mL, respectively. The ferric reducing antioxidant power (FRAP) of MOH indicated a dose-dependent increase with a maximum absorbance at 125 µg/mL and the oxygen radical absorbance capacity (ORAC) of MOH was 1004.95 µmol TE/mg. Results showed that MOH dose-dependently reduced intracellular ROS generation in H2O2-stimulated Vero cells while increasing the cell viability. Fluorescence microscopy and flowcytometric analyses have supported the above findings. MOH markedly suppressed the H2O2-induced mitochondrial depolarization and apoptosis through suppression of the mitochondrial-mediated apoptosis pathway and activated the Nrf2/HO-1 signaling pathway by possibly involving H2O2 generation in cell media. Findings of western blot were supported by immunocytochemistry of Nrf2 nuclear translocation. Thus, MOH bioactivity would potentiate its applications in manufacturing functional food.

(-)-Loliolide Isolated from Sargassum horneri Abate UVB-Induced Oxidative Damage in Human Dermal Fibroblasts and Subside ECM Degradation.

Ultraviolet (UV) B exposure is a prominent cause of skin aging and a contemporary subject of interest. The effects are progressing through the generation of reactive oxygen species (ROS) that alter cell signaling pathways related to inflammatory responses. The present study evaluates the protective effects of (7aR)-6-hydroxy-4,4,7a-trimethyl-6,7-dihydro-5H-1-benzofuran-2-one (HTT) isolated from the edible brown algae Sargassum horneri against UVB protective effects in human dermal fibroblasts (HDFs). HTT treatment dose-dependently suppressed intracellular ROS generation in HDFs with an IC50 of 62.43 ± 3.22 µM. HTT abated UVB-induced mitochondrial hyperpolarization and apoptotic body formation. Furthermore, UVB-induced activation of key nuclear factor (NF)-κB and mitogen-activated protein kinase signaling proteins were suppressed in HTT treated cells while downregulating pro-inflammatory cytokines (interleukin-1ß, 6, 8, 33 and tumor necrosis factor-α). Moreover, HTT treatment downregulated matrix metalloproteinase1, 2, 3, 8, 9 and 13 that was further confirmed by the inhibition of collagenase and elastase activity. The evidence implies that HTT delivers protective effects against premature skin aging caused by UVB exposure via suppressing inflammatory responses and degradation of extracellular matrix (ECM) components. Extensive research in this regard will raise perspectives for using HTT as an ingredient in UV protective ointments.

UVB protective effects of Sargassum horneri through the regulation of Nrf2 mediated antioxidant mechanism.

The present study aimed to evaluate the protective effect of a methanol extract of Sargassum horneri (SHM), which contains 6-hydroxy-4,4,7a-trimethyl-5,6,7,7a-tetrahydrobenzofuran-2(4H)-one (HTT) and apo-9'-fucoxanthinone, against ultraviolet B (UVB)-induced cellular damage in human keratinocytes and its underlying mechanism. SHM significantly improved cell viability of UVB-exposed human keratinocytes by reducing the generation of intracellular reactive oxygen species (ROS). Moreover, SHM inhibited UVB exposure-induced apoptosis by reducing the formation of apoptotic bodies and the populations of the sub-G1 hypodiploid cells and the early apoptotic cells by modulating the expression of the anti- and pro-apoptotic molecules, Bcl-2 and Bax, respectively. Furthermore, SHM inhibited NF-κB p65 activation by inducing the activation of Nrf2/HO-1 signaling. The cytoprotective and antiapoptotic activities of SHM are abolished by the inhibition of HO-1 signaling. In further study, SHM restored the skin dryness and skin barrier disruption in UVB-exposed human keratinocytes. Based to these results, our study suggests that SHM protects the cells against UVB-induced cellular damages through the Nrf2/HO-1/NF-κB p65 signaling pathway and may be potentially useful for the prevention of UVB-induced skin damage.

Effects of (-)-Loliolide against Fine Dust Preconditioned Keratinocyte Media-Induced Dermal Fibroblast Inflammation.

At present air pollution in parts of East Asia is at an alarming level due to elevated levels of fine dust (FD). Other than pulmonary complications, FD was found to affect the pathogenesis of ROS-dependent inflammatory responses via penetrating barrier-disrupted skin, leading to degradation of extracellular matrix components through the keratinocyte-fibroblast axis. The present study discloses the evaluation of human dermal fibroblast (HDF) responses to FD preconditioned human keratinocyte media (HPM) primed without and with (-)-loliolide (HTT). HPM-FD treatment increased the ROS level in HDFs and activated mitogen-activated protein kinase-derived nuclear factor (NF)-κB inflammatory signaling pathways with a minor reduction of viability. The above events led to cell differentiation and production of matrix metalloproteinases (MMP), increasing collagenase and elastase activity despite the increase of tissue inhibitors of metalloproteinases (TIMP). Media from HTT primed keratinocytes stimulated with FD indicated ameliorated levels of MMPs, inflammatory cytokines, and chemokines in HDFs with suppressed collagenase and elastase activity. Present observations help to understand the factors that affect HDFs in the microenvironment of FD exposed keratinocytes and the therapeutic role of HTT as a suppressor of skin aging. Further studies using organotypic skin culture models could broaden the understanding of the effects of FD and the therapeutic role of HTT.

Sargassum horneri (Turner) C. Agardh ethanol extract attenuates fine dust-induced inflammatory responses and impaired skin barrier functions in HaCaT keratinocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Sargassum horneri (Turner) C. Agardh is well known in East Asia as an edible brown alga rich in bioactive compounds. It has an ethnopharmacological significance in traditional Chinese medicine to treat inflammatory disorders varying from edema, furuncles, dysuria to cardiovascular diseases. AIM OF THE STUDY: Surge of fine dust (FD), in densely populated areas, have been reported to cause adverse health conditions ranging from respiratory diseases to inflammatory skin disorders. The current study investigates the protective effects of an ethanol extract from S. horneri (SHE) on FD-induced inflammatory responses and impaired skin hydration in HaCaT keratinocytes. MATERIALS AND METHODS: Intracellular reactive oxygen species (ROS) generation was evaluated with the 2',7'-Dichlorofluorescin diacetate (DCFH-DA) stain. Anti-inflammatory properties of SHE in FD-stimulated HaCaT keratinocytes were investigated for the suppression of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) pathways and downregulation of pro-inflammatory cytokines. As a means of studying FD-induced skin barrier disruption and the effects of SHE on stratum corneum hydration-controlling factors, tight junction regulatory mediators, and hyaluronic acid (HA) production were evaluated using keratinocytes. RESULTS: SHE suppressed the intracellular ROS production, simultaneously improving cell viability in FD-stimulated keratinocytes. Also, SHE upregulated anti-inflammatory cytokine interleukin (IL)-4 while downregulating inflammatory cytokines IL-1ß, IL-6, IL-8, tumor necrosis factor (TNF)-α; epidermal and epithelial cytokines IL-25, IL-33, and thymic stromal lymphopoietin (TSLP); thymus and activation-regulated chemokine (TARC), macrophage-derived chemokine (MDC) and regulated upon activation, normally T-expressed, and presumably secreted expression and suppressed (RANTES) chemokine, MAPK and NF-κB mediators in a dose-dependent manner. Furthermore, SHE ameliorated filaggrin, involucrin, lymphoepithelial Kazal-type-related inhibitor (LEKTI), signifying its beneficial effects on deteriorated skin hydration caused by FD-induced inflammation. SHE further exhibited its skin protective effects regulating the tight junction proteins; Occludin, zonula occludens (ZO)-1, claudin-1, claudin-4, claudin-7, and claudin-23 while increasing the production of HA minimizing skin damage. CONCLUSIONS: Anti-inflammatory effects of, SHE against FD-induced keratinocyte inflammation is attributable to the suppression of upstream MAPK and NF-κB mediators. SHE indicated potential anti-inflammatory properties attenuating deteriorated skin barrier function in HaCaT keratinocytes. The effects are attributable to the polyphenols and other antioxidant compounds in SHE. Further studies could envisage the use of SHE for developing rejuvenating cosmetics.

Fucoidan Fractionated from Sargassum coreanum via Step-Gradient Ethanol Precipitation Indicate Promising UVB-Protective Effects in Human Keratinocytes.

Fucoidans exhibit a wide range of bioactivities and receive significant attention in functional food and cosmetic research. Industrial applications of fucoidan are limited partially due to high extraction and purification costs. The present study implements an enzyme-assisted extraction and step-gradient ethanol precipitation for fractionating fucoidan from Sargassum coreanum based on its charge and molecular weight and evaluation of ultraviolet B (UVB) protective effects in human keratinocytes (HaCaT). The fucoidan fraction SCOC4 indicated higher fucose and sulfate contents with Fourier-transform infrared and 1H NMR spectral patterns resembling fucoidans. SCOC4 dose-dependently abated UVB-induced keratinocyte damage via suppressing intracellular reactive oxygen species, apoptotic body formation, DNA damage via suppressing mitochondria-mediated apoptosis. UVB-protective effects of SCOC4 were further attributable to the augmentation of nuclear factor erythroid 2-related factor 2 mediated cellular antioxidant defense enzymes. Step-gradient ethanol precipitation was a convenient approach of fractionating fucoidans based on molecular weight and charge (depend on the degree of sulfation). Further evaluation of seasonal variations, biocompatibility parameters, efficacy, and shelf life may widen the use of S. coreanum fucoidans in developing UVB-protective cosmetics and functional foods.

Low molecular weight fucoidan fraction ameliorates inflammation and deterioration of skin barrier in fine-dust stimulated keratinocytes.

Recently evidence linking the effects of fine-dust (FD) on skin inflammation is exaggerating. Fucoidan derived from brown algae has great potential for ameliorating oxidative stress and inflammation. Herein, a fucoidan fraction (SHC4-6) was purified from an enzymatic (Celluclast) extract of an invasive seaweed, Sargassum horneri following gradient ethanol precipitation and anion exchange chromatography. Effectiveness of SHC4-6 in ameliorating FD (from Beijing, China)-induced inflammatory responses in HaCaT keratinocytes and recovery of skin barrier dysfunction was evaluated. SHC4-6 was comprising of sulfated mannofucans with their molecular weights distributed around 45 kDa. SHC4-6 dose-dependently lowered ROS levels in FD-induced HaCaT keratinocytes, ameliorating viability at 50 µg mL-1. SHC4-6 downregulated inflammatory cytokines, tumor necrosis factor-α, interleukin (IL)-1ß, -5, -6, -8, -13, interferon-γ, and chemokines, macrophage-derived chemokine, eotaxin, and thymus and activation regulated chemokine by inhibiting mitogen-activated protein kinase and nuclear factor-κB pathways. SHC4-6 treatment ameliorated key tight junction proteins and skin hydration factors, depicting the effects of fucoidan in reducing FD-induced inflammation and skin barrier deterioration. With further studies in place, SHC4-6 could be used as an ingredient for developing cosmetics to relieve FD-induced skin inflammation.

Oral Administration of Sargassum horneri Improves the HDM/DNCB-Induced Atopic Dermatitis in NC/Nga Mice.

The present study investigated the protective effects of Sargassum horneri (S. horneri) ethanol extract (SHE) against atopic dermatitis (AD), known as an abnormal immune response in house dust mite (HDM)/2,4-dinitrochlorobenzene (DNCB)-stimulated NC/Nga mice. The oral administration of SHE attenuated the AD symptoms, including the skin dermatitis severity, transepidermal water loss (TEWL), and ear edema in HDM/DNCB-stimulated mice. Moreover, the histological analysis revealed that SHE improved epidermal hyperplasia and hyperkeratosis, and reduced the dermal infiltrations of mast cells and eosinophils. Moreover, SHE downregulated the expression levels of cytokines (interleukin (IL)-6, IL-10, and interferon (IFN)-γ) and chemokines (Regulated on Activation, Normal T Cell Expressed and Secreted (RANTES), Eotaxin, and Thymus and activation-regulated chemokine (TARC)) by decreasing the expression levels of atopic initiators (IL-25 and IL-33) in HDM/DNCB-stimulated skin. The oral administration of SHE decreased the spleen size, reducing expression levels of AD-related cytokines (IL-4, IL-5, IL-6, IL-10, IL-13, IFN-γ, and TARC) by regulating the expressions of Tbx21 (T-bet), GATA Binding Protein 3 (GATA-3), and Signal transducer and activator of transcription 3 (STAT3). Moreover, SHE significantly attenuated the serum immunoglobulin (Ig)G1 and IgG2a levels in HDM/DNCB-stimulated mice. Collectively, these results suggest that S. horneri could be an ingredient of functional food against abnormal immune response.

Fucoidan refined by Sargassum confusum indicate protective effects suppressing photo-oxidative stress and skin barrier perturbation in UVB-induced human keratinocytes.

Damage from ultraviolet (UV)B exposure is cumulative and proceeds via augmenting cellular oxidative stress. The present study investigates UVB-protective effects of Sargassum confusum derived fucoidans in human keratinocytes. Algae was extracted using Celluclast, and fucoidan fractions were recovered by step gradient ethanol precipitation. Refined fucoidan fractions were treated to human HaCaT keratinocytes and exposed to UVB (50 mJ cm-2). Among fucoidan fractions, SCFC4, the lowest molecular weight (MW) fraction indicated better UVB-protective effects with dose-dependent reduction of intracellular ROS levels, while recovering the cell viability. SCFC4 suppressed UVB-induced apoptotic body formation, sub-G1 cell content, and DNA damage by hindering the mitochondria-mediated apoptotic pathway. Also, SCFC4 repressed upstream mediators of UVB-induced inflammatory responses, which would impair stratum corneum hydration. The above therapeutic effects of SCFC4 could be attributed to suppression of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) pathways while activating nuclear factor erythroid 2-related factor 2 (Nrf2) mediated production of antioxidant enzymes. SCFC4 was identified as a fucoidan (MW ≈ 20 kDa) with a 64.64% fucose and 23.62% sulfate contents. Further analysis of shelf life, safety, and efficacy could promote SCFC4's use as a cosmetic ingredient.

Step gradient alcohol precipitation for the purification of low molecular weight fucoidan from Sargassum siliquastrum and its UVB protective effects.

Ultraviolet B (UVB) can induce oxidative damage to outermost layers of skin causing suntans, sunburns, and, in severe cases, blisters leading to photoaging. Low molecular weight (MW) fucoidan is renowned for possessing enhanced antioxidant activities. The present study discloses the use of step gradient ethanol precipitation in refining fucoidan fractions (SSQC1-SSQC4) from Sargassum siliquastrum and evaluation of their UVB-protective effects in human HaCaT keratinocytes. Among the fractions, SSQC4 indicated the best bioactive effects. 1H NMR, FTIR, monosaccharide composition by HPAEC-PAD analysis, MW estimation by agarose gel electrophoresis were used to characterize the fractions. SSQC4 was comprising of fucoidan, with an estimated MW distribution of 8-25 kDa. Exposure of UVB increased intracellular ROS, DNA damage, loss of mitochondrial membrane potential, apoptotic body formation causing cell death through the mitochondria-mediated apoptosis pathway. SSQC4 treatment could dose-dependently attenuate the ROS levels and suppress mitochondria-mediated apoptosis in UVB exposed keratinocytes. SSQC4 treatment enhanced cellular antioxidant defense by increasing Nrf2 mediated HO-1 generation, which was identified as the cause of observed bioactivities. The safety and stability of SSQC4 could be further evaluated to promote its use as a bioactive natural ingredient in UV-protective cosmetics.

(-)-Loliolide Isolated from Sargassum horneri Protects against Fine Dust-Induced Oxidative Stress in Human Keratinocytes.

The emergence of fine dust (FD) among air pollutants has taken a toll during the past few decades, and it has provided both controversy and a platform for open conversation amongst world powers for finding sustainable solutions and effective treatments for health issues. The present study emphasizes the protective effects of (-)-loliolide (HTT) isolated from Sargassum horneri against FD-induced oxidative stress in human HaCaT keratinocytes. The purification of (-)-loliolide was carried out by centrifugal partition chromatography. HTT did not show any cytotoxicity, and it further illustrated the potential to increase cell viability by reducing the reactive oxygen species (ROS) production in FD-stimulated keratinocytes. Furthermore, HTT suppressed FD-stimulated DNA damage and the formation of apoptotic bodies, and it reduced the population of cells in the sub-G1 apoptosis phase. FD-induced apoptosis was advancing through the mitochondria-mediated apoptosis pathway. The cytoprotective effects of the HTT against FD-stimulated oxidative damage is mediated through squaring the nuclear factor E2-related factor 2 (Nrf2)-mediated heme oxygenase-1 (HO-1) pathway, dose-dependently increasing HO-1 and NAD(P)H dehydrogenase (quinone) 1 (NQO1) levels in the cytosol while concomitantly improving the nuclear translocation of Nrf2. Future studies could implement the protective functionality of HTT in producing pharmaceuticals that utilize natural products and benefit the diseased.

Human Keratinocyte UVB-Protective Effects of a Low Molecular Weight Fucoidan from Sargassum horneri Purified by Step Gradient Ethanol Precipitation.

Ultraviolet B (UVB) radiation-induced oxidative skin cell damage is a major cause of photoaging. In the present study, a low molecular weight fucoidan fraction (SHC4) was obtained from Sargassum horneri by Celluclast-assisted extraction, followed by step gradient ethanol precipitation. The protective effect of SHC4 was investigated in human keratinocytes against UVB-induced oxidative stress. The purified fucoidan was characterized by Fourier-transform infrared spectroscopy (FTIR), 1H nuclear magnetic resonance (NMR), agarose gel-based molecular weight analysis and monosaccharide composition analysis. SHC4 had a mean molecular weight of 60 kDa, with 37.43% fucose and 28.01 ± 0.50% sulfate content. The structure was mainly composed of α-L-Fucp-(1→4) linked fucose units. SHC4 treatment dose-dependently reduced intracellular reactive oxygen species (ROS) levels and increased the cell viability of UVB exposed HaCaT keratinocytes. Moreover, SHC4 dose-dependently inhibited UVB-induced apoptotic body formation, sub-G1 accumulation of cells and DNA damage. Inhibition of apoptosis was mediated via the mitochondria-mediated pathway, re-establishing the loss of mitochondrial membrane potential. The UVB protective effect of SHC4 was facilitated by enhancing intracellular antioxidant defense via nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling. Further studies may promote the use of SHC4 as an active ingredient in cosmetics and nutricosmetics.