NIR-responsive carboxymethyl-cellulose hydrogels containing thioketal-linkages for on-demand drug delivery system.

Near-infrared (NIR) light-responsive hydrogels have emerged as a highly promising strategy for effective anticancer therapy owing to the remotely controlled release of chemotherapeutic molecules with minimal invasive manner. In this study, novel NIR-responsive hydrogels were developed from reactive oxygen species (ROS)-cleavable thioketal cross-linkers which possessed terminal tetrazine groups to undergo a bio-orthogonal inverse electron demand Diels Alder click reaction with norbornene modified carboxymethyl cellulose. The hydrogels were rapidly formed under physiological conditions and generated N2 gas as a by-product, which led to the formation of porous structures within the hydrogel networks. A NIR dye, indocyanine green (ICG) and chemotherapeutic doxorubicin (DOX) were co-encapsulated in the porous network of the hydrogels. Upon NIR-irradiation, the hydrogels showed spatiotemporal release of encapsulated DOX (>96 %) owing to the cleavage of thioketal bonds by interacting with ROS generated from ICG, whereas minimal release of encapsulated DOX (<25 %) was observed in the absence of NIR-light. The in vitro cytotoxicity results revealed that the hydrogels were highly cytocompatible and did not induce any toxic effect on the HEK-293 cells. In contrast, the DOX + ICG-encapsulated hydrogels enhanced the chemotherapeutic effect and effectively inhibited the proliferation of Hela cancer cells when irradiated with NIR-light.

Spatial distribution of meiofaunal and macrofaunal assemblages in the tidal flats of the southern Korean coast in relation to natural and anthropogenic impacts.

We investigated the spatial variability of macrofaunal and meiofaunal assemblages in intertidal flats on the southern coast of Korea. Abiotic and biotic samples were collected at five stations. The species richness, density, and composition of the assemblages differed significantly among stations. Nematoda and Annelida were the most dominant meiofaunal and macrofaunal taxa, respectively, although taxon dominance differed among stations. Distance-based linear models showed that sediment-related variables and heavy metals were the main environmental factors determining the spatial variability of the assemblages. Macrofauna had only sediment-related variables and heavy metals as major environmental factors, but meiofauna were also influenced by other environmental factors such as sea surface temperature, dissolved oxygen-related variables, and salinity. This study can provide basic ecological data for understanding the spatial distribution of macro-meiofaunal assemblages and aid in the development of marine environmental management strategies on the western south coast of Korea.

Dual cross-linked chitosan/alginate hydrogels prepared by Nb-Tz 'click' reaction for pH responsive drug delivery.

A novel physically and chemically double-crosslinked hydrogel derived from chitosan oligosaccharide/alginate (COS/Alg) was developed by using norbornene (Nb)-tetrazine (Tz) click reaction for ketoprofen delivery. The properties of the hydrogel were evaluated by rheological, FTIR, TGA, XRD, SEM, swelling and drug release studies. The Nb-Tz chemical cross-linking facilitated outstanding hydrophobic drug loading (44% wt/wt of ketoprofen) and sustained release through a hydrophobic interaction mechanism between the drug and the used polysaccharides. The COS/Alg electrostatics network (10/10 of NH2/COOH molar ratio) generated the pH responsiveness, suppressing the release in simulated gastric fluid (below 10% for 2 h) and enhancing the release in simulated intestinal fluids (up to 84% for 24 h). The prepared hydrogel was non-toxic to human HEK-293 cells (95% cell viability). This work opens up a potential approach for preparing hydrophilic hydrogels from natural polysaccharides that can be used in the delivery of hydrophobic drugs.

Multi-stimuli responsive hydrogels derived from hyaluronic acid for cancer therapy application.

One of the most promising strategies for the controlled release of therapeutic molecules is stimuli-responsive and biodegradable hydrogels developed from natural polymers. However, current strategies to development stimuli-responsive hydrogels lack precise control over drug release profile and use cytotoxic materials during preparation. To address these issues, multi-stimuli responsive hydrogels derived from hyaluronic acid and diselenide based cross-linker were developed for the controlled release of doxorubicin (DOX). Hydrogels were rapidly formed via an inverse electron demand Diels-Alder click chemistry and encapsulated DOX/indocyanine green (ICG) in their porous networks. The hydrogels showed a rapid release of DOX in acidic (pH 5), reducing (10 mmol DTT), and oxidizing medium (0.5% H2O2), and after NIR irradiation. The in vitro experiments demonstrated that hydrogels were highly cytocompatible and the DOX-loaded hydrogels induced similar anti-tumor effect as compared to that of the free-DOX. Furthermore, DOX + ICG loaded hydrogels increased the antitumor efficacy of DOX after NIR irradiation.

Dual pH-/thermo-responsive chitosan-based hydrogels prepared using "click" chemistry for colon-targeted drug delivery applications.

A dual pH-/thermo-responsive hydrogel was designed based on a polyelectrolyte complex of polyacrylic acid (PAA) and norbornene-functionalized chitosan (CsNb), which was synergized with chemical crosslinking using bistetrazine-poly(N-isopropyl acrylamide) (bisTz-PNIPAM). The thermo-responsive polymeric crosslinker, bisTz-PNIPAM, was synthesized via reversible addition-fragmentation transfer polymerization of NIPAM. FTIR, XRD, rheological and morphological analyses demonstrated the successful formation of the polyelectrolyte network. The highly porous structure generated through the in-situ "click" reaction between Tz and Nb resulted in a higher drug loading (29.35 %). The hydrogel (COOH/NH2 mole ratio of 3:1) exhibited limited drug release (8.5 %) of 5-ASA at a pH of 2.2, but it provided an almost complete release (92 %) at pH 7.4 and 37 °C within 48 h due to the pH responsiveness of PAA, hydrogel porosity, and shrinkage behavior of PNIPAM. The hydrogels were biodegradable and non-toxic against human fibroblast cells, suggesting their considerable potential for a colon-targeted drug delivery system.

Risk screening of the potential invasiveness of non-native marine fishes for South Korean coastal waters.

Risk screening tools are being increasingly used to identify the potential invasiveness and associated risks of non-native species. In this study, the Aquatic Species Invasiveness Screening Kit was used to evaluate the invasiveness risks of extant and horizon non-native marine fish species for the coastal waters of South Korea. In total, 57 marine fish species were screened and the threshold scores for the Basic Risk Assessment (BRA) and the BRA + Climate Change Assessment (BRA+CCA) (5.5 and 1.5, respectively) reliably distinguished those species carrying a high risk of invasiveness from those carrying a low to medium risk. For both the BRA and BRA+CCA, common lionfish Pterois miles was the highest-scoring species, followed by white perch Morone americana, red drum Sciaenops ocellatus, marbled spinefoot Siganus rivulatus and redcoat Sargocentron rubrum. The outcomes of this study will contribute to the management of non-native marine fish species for the conservation of the native ecosystems in the coastal waters of South Korea.

Changes in Hematological Parameters and Heat Shock Proteins in Juvenile Sablefish Depending on Water Temperature Stress.

Juvenile Sablefish Anoplopoma fimbria were used to assess the effects of water temperature (8, 10, 12, 14, 16, 18, and 20°C) on hematological parameters and heat shock proteins 70 and 90 for 4 months. Hematological parameters, including red blood cell count, hematocrit, and hemoglobin, were significantly decreased at 18°C. The inorganic plasma components calcium and magnesium were not altered by water temperature. The organic plasma components glucose and cholesterol were notably elevated at 18°C, whereas total protein was reduced. The enzymatic components, including aspartate aminotransferase, alanine aminotransaminase, and alkaline phosphatase, were notably elevated at 16°C or 18°C. The results of this study indicate that a temperature higher than the proper temperature affects the hematological parameters and heat shock proteins of juvenile Sablefish.

Meroterpenoid-Rich Fraction of the Ethanolic Extract from Sargassum serratifolium Suppressed Oxidative Stress Induced by Tert-Butyl Hydroperoxide in HepG2 Cells.

Sargassum species have been reported to be a source of phytochemicals, with a wide range of biological activities. In this study, we evaluated the hepatoprotective effect of a meroterpenoid-rich fraction of the ethanolic extract from Sargassum serratifolium (MES) against tert-butyl hydroperoxide (t-BHP)-treated HepG2 cells. Treatment with MES recovered the cell viability from the t-BHP-induced oxidative damage in a dose-dependent manner. It suppressed the reactive oxygen species production, lipid peroxidation, and glutathione depletion in the t-BHP-treated HepG2 cells. The activity of the antioxidants induced by t-BHP, including superoxide dismutase (SOD) and catalase, was reduced by the MES treatment. Moreover, it increased the nuclear translocation of nuclear factor erythroid 2-related factor 2, leading to the enhanced activity of glutathione S transferase, and the increased production of heme oxygenase-1 and NAD(P)H:quinine oxidoreductase 1 in t-BHP-treated HepG2 cells. These results demonstrate that the antioxidant activity of MES substituted the activity of the SOD and catalase, and induced the production of detoxifying enzymes, indicating that MES might be used as a hepatoprotectant against t-BHP-induced oxidative stress.

Toxic effects of juvenile sablefish, Anoplopoma fimbria by ammonia exposure at different water temperature.

Juvenile sablefish, Anoplopoma fimbria (mean length 17.1±2.4cm, and mean weight 75.6±5.7g) were used to evaluate toxic effects on antioxidant systems, immune responses, and stress indicators by ammonia exposure (0, 0.25, 0.75, and 1.25mg/L) at different water temperature (12 and 17°C) in 1 and 2 months. In antioxidant responses, superoxide dismutase (SOD) and catalase (CAT) were significantly increased by ammonia exposure, whereas glutathione (GSH) was decreased. In immune responses, lysozyme and phagocytosis activity were significantly increased by ammonia exposure. In stress indicators, plasma glucose, heat shock protein 70 (HSP 70), and cortisol were significantly increased. At high water temperature (17°C), alterations by ammonia exposure were more distinctly. The results of this study indicated that ammonia exposure can induce toxic effects in the sablefish, and high water temperature can affect the ammonia exposure toxicity.

Cyperus amuricus induces G1 arrest and apoptosis through endoplasmic reticulum stress and mitochondrial signaling in human hepatocellular carcinoma Hep3B cells.

ETHNOPHARMACOLOGY RELEVANCE: Cyperus amuricus (C. amuricus), belongs to the family Cyperaceae, was used to exert wound healing, diuretic, astringent and other intestinal problems in oriental medicine. However, only a few studies have reported its anticancer activities. AIM OF THE STUDY: In this study, we determined the activity of C. amuricus on ER stress-induced cell death and G1 cell cycle arrest in human hepatocellular carcinoma (HCC) Hep3B cells. MATERIALS AND METHODS: The in vitro cell proliferation assay of C. amuricus was tested on Hep3B and human embryonic kidney HEK293 cells. Subsequently, the cell cycle distribution in the indicated stages using flow cytometric analysis, the expression of cell cycle-related proteins by western blot analysis and immunofluorescence detection of p21CIP1/WAF1 were determined for the comprehensive identification of cell cycle arrest in Hep3B cells. The effect of C. amuricus on the expression of apoptosis-related proteins in Hep3B cells was also performed by western blot analysis. Furthermore, induction of the ER stress mediators in C. amuricus-treated Hep3B cells were observed by western blot analysis, intracellular Ca2+ mobilization assay and immunofluorescence detection of caspase-12. RESULTS: C. amuricus strongly exhibited cytotoxic activity on Hep3B cells, but not on HEK293 cells. C. amuricus affected the phosphorylation levels of endoplasmic reticulum sensors and increased the expression of GRP78/BiP and CHOP, resulting in the accumulation of unfolded proteins in the ER and triggering the unfolded protein response. These changes occurred by the elevation of intracellular Ca2+ concentrations, which contributed to ER stress-induced apoptosis in C. amuricus-treated Hep3B cells. C. amuricus also coordinated the stimulation of ER chaperones, which initiated G1 cell cycle arrest through the induction of CDKIs and the inhibition of cyclins and CDKs. Furthermore, C. amuricus triggered apoptosis through the activation of mitochondrial-dependent pathway in Hep3B cells. CONCLUSIONS: Our results suggest that C. amuricus is an effective apoptosis inducing agent for Hep3B cells via the G1 arrest, ER stress and mitochondrial mediated apoptotic pathways.

Antioxidant Responses, Neurotoxicity, and Metallothionein Gene Expression in Juvenile Korean Rockfish Sebastes schlegelii under Dietary Lead Exposure.

This study was conducted to assess toxic effects of dietary lead (Pb) exposure on Korean Rockfish Sebastes schlegelii. Juvenile rockfish were used to evaluate the oxidative stress, neurotoxicity, and metallothionein (MT) gene expression after dietary exposure to lead (as Pb2+; 0, 30, 60, 120 and 240 mg/kg). Superoxide dismutase (SOD) activity, a measure of oxidative stress, was substantially elevated in the livers and gills of fish given dietary Pb greater than 60 mg/kg. Glutathione S-transferase (GST) activity in the liver and gill was significantly increased by dietary Pb > 60 mg/kg. A significant decrease in glutathione (GSH) level was observed in fish liver after exposure to dietary Pb > 30 mg/kg and in the gill after treatment with dietary Pb > 120 mg/kg. Acecyltholinesterase (AChE) was noticeably decreased in the brain by dietary Pb > 120 mg/kg and in the muscle by dietary Pb > 60 mg/kg. Metallothionein gene expression in the liver was stimulated significantly by the Pb exposure. Because dietary Pb exposure had a toxic effect on antioxidant responses, a neurotransmitter, and a specific immune expression in rockfish, the results of this study can be used to determine potential useful markers of Pb toxicity. Received June 11, 2016; accepted March 10, 2017.

Growth performance, oxidative stress, and non-specific immune responses in juvenile sablefish, Anoplopoma fimbria, by changes of water temperature and salinity.

Juvenile sablefish, Anoplopoma fimbria (mean length 15.5 ± 1.9 cm, mean weight 68.5 ± 4.8 g), were used to evaluate the effects on growth, oxidative stress, and non-specific immune responses by changes of water temperature (8, 10, 12, 14, 16, 18, and 20 °C) and salinity (100 (35.0), 90 (31.5), 80 (28.0), 70 (24.5), 60 (21.0), 50 (17.5), and 40% (14.0) (‰)) for 4 months. The growth performance was significantly increased at the temperature of 12 and 14 °C, and the feed efficiency was notably decreased at the temperature of 18 °C. The growth performance and feed efficiency were also significantly decreased at low salinity. The antioxidant responses such as superoxide dismutase and catalase were significantly increased by the high temperature and decreased by the low salinity. The immune responses such as lysozyme and phagocytosis were elevated by the temperature of 18 °C and decreased by the salinity of 50%. The results of this study indicate that the growth performance of juvenile sablefish, A. fimbria, is influenced by the temperature and salinity, and the excessive temperature and salinity levels can affect the antioxidant and immune responses.

Anti-Inflammatory Effect of Ethanolic Extract of Sargassum serratifolium in Lipopolysaccharide-Stimulated BV2 Microglial Cells.

Sargassum serratifolium was found to contain high concentrations of meroterpenoids, having strong antioxidant, anti-inflammatory, and neuroprotective activities. This study aims to investigate the anti-inflammatory mechanisms of an ethanolic extract of S. serratifolium (ESS) using lipopolysaccharide (LPS)-stimulated BV2 microglial cells and to identify the anti-inflammatory components in ESS. The level of proinflammatory cytokines was measured by enzyme-linked immunosorbent assay. The expression of inflammation-related proteins and mRNA was evaluated by Western blot and reverse transcription-polymerase chain reaction analysis, respectively. Anti-inflammatory activities of isolated components from ESS were analyzed in LPS-stimulated BV2 cells. ESS inhibited LPS-induced nitric oxide (NO) and prostaglandin E2 and the expression of inducible NO synthase and cyclooxygenase-2. ESS also decreased the release of proinflammatory cytokines in a dose-dependent manner. LPS-induced nuclear factor-kappa B (κB) transcriptional activity and translocation into the nucleus were remarkably suppressed by ESS through the prevention of inhibitor κB-α degradation. The main anti-inflammatory components in ESS were identified as sargahydroquinoic acid, sargachromenol, and sargaquinoic acid based on the inhibition of NO production using LPS-stimulated BV2 cells. Furthermore, treatment with ESS significantly reduced levels of tumor necrosis factor-α and interleukin-1ß stimulated with LPS in mouse hippocampus. Our results indicate that ESS can be used as a functional food or therapeutic agent for the treatment of neuroinflammatory diseases.

The herbal medicine Cyperus amuricus inhibits proliferation of human hepatocellular carcinoma Hep3B cells by inducing apoptosis and arrest at the G0/G1 cell cycle phase.

Cyperus amuricus (C. amuricus) is one of the most common herbs in Oriental folk medicine for exerting astringent, diuretic, wound healing and other intestinal problems. However, little is known about the molecular mechanism of C. amuricus on anticancer activity. In the present study, the underlying mechanism of the anticancer effect of C. amuricus were elucidated. The methyl alcohol extract from the whole plant of C. amuricus exhibited cytotoxicity against Hep3B cells, but not against A549 and HaCaT cells. Consistent with an acceleration of the sub-G1 phase, downregulation of cdc25A, cyclin D1 and cyclin E, CDK4 and 2 as well as E2F-1, phospho-Rb, with concomitant of upregulation of p21CIP1/WAF1, p27KIPI and p16INK4a proteins, as evidenced by the appearance of cell cycle arrest, were detected in C. amuricus-treated Hep3B cells. Additionally, the sequential activation of various caspases (cleaved caspase-8, -9, -3, -7 and -6, and cleaved PARP) and the changed expression of other proteins related to the apoptosis pathway were observed after C. amuricus exposure. An increment in the pro-apoptotic proteins (Bim, tBid, Bax and Bak) and a reduction of anti-apoptotic protein (Bcl-2) regulate Hep3B cell death by controlling the permeability of mitochondrial membranes and the release of cytochrome c from mitochondria into the cytosol with Apaf-1 after C. amuricus treatment. This is the first study indicating the potential of C. amuricus as a complementary agent for prevention and treatment of human liver cancer.

Sargaquinoic acid attenuates inflammatory responses by regulating NF-κB and Nrf2 pathways in lipopolysaccharide-stimulated RAW 264.7 cells.

Myagropsis myagroides, a brown alga, showed strong anti-inflammatory activities in the previous studies. In this study, we isolated a strong anti-inflammatory compound, sargaquinoic acid (SQA), from M. myagroides and investigated the anti-inflammatory action using lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. SQA suppressed the production of inducible nitric oxide synthase and cyclooxygenase-2 in LPS-stimulated cells as well as that of reactive oxygen species. As a result, SQA inhibited the production of NO, prostaglandin E2, and pro-inflammatory cytokines. LPS-induced transcriptional activation of nuclear factor-κB (NF-κB) was remarkably inhibited by SQA treatment through the prevention of inhibitor κB-α degradation. The regulation of NF-κB activation was also mediated by the phosphorylation of ERK and Akt in LPS-stimulated RAW 264.7 cells. Moreover, SQA induced the production of heme oxygenase 1 via activation of transcription factor Nrf2. These results indicate that SQA inhibits the LPS-induced expression of inflammatory mediators via suppression of ERK and Akt-mediated NF-κB pathway as well as up-regulation of Nrf2/HO-1 pathway, indicating that SQA has a potential therapeutic and preventive application in various inflammatory diseases.

Sargaquinoic Acid Inhibits TNF-α-Induced NF-κB Signaling, Thereby Contributing to Decreased Monocyte Adhesion to Human Umbilical Vein Endothelial Cells (HUVECs).

Sargaquinoic acid (SQA) has been known for its antioxidant and anti-inflammatory properties. This study investigated the effects of SQA isolated from Sargassum serratifolium on the inhibition of tumor necrosis factor (TNF)-α-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). SQA decreased the expression of cell adhesion molecules such as intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 as well as chemotactic cytokines such as interleukin-8 and monocyte chemoattractant protein-1 in TNF-α-treated HUVECs. As a result, SQA prevented monocyte adhesion to TNF-α-induced adhesion. SQA also inhibited TNF-α-induced nuclear factor kappa B (NF-κB) translocation into the nucleus by preventing proteolytic degradation of inhibitor κB-α. Overall, SQA protects against TNF-α-induced vascular inflammation through inhibition of the NF-κB pathway in HUVECs. These data suggest that SQA may be used as a therapeutic agent for vascular inflammatory diseases such as atherosclerosis.

Hexane extract from Sargassum serratifolium inhibits the cell proliferation and metastatic ability of human glioblastoma U87MG cells.

The present study is the first to demonstrate the anticancer effects of a hexane extract from the brown algae Sargassum serratifolium (HES) on human cancer cell lines, including glioblastoma U87MG, cervical cancer HeLa and gastric cancer MKN-28 cells, as well as liver cancer SK-HEP 1 cells. Among these cancer cell lines, U87MG cells were most sensitive to the cell death induced by HES. HES exhibited a cytotoxic effect on U87MG cells at concentrations of 14-16 µg/ml, yet an effect was not observed in human embryonic kidney HEK293 cells. The antiproliferative effects of HES were regulated by inhibition of the MAPK/ERK signaling pathway which plays a pivotal role in the proliferation of glioblastoma U87MG cells. In addition, treatment with HES led to cell morphological changes and cell cytoskeleton degradation through regulation of actin dynamic signaling. Furthermore, migration and invasion of the U87MG cells were inhibited by HES via suppression of matrix metalloproteinase (MMP)-2 and -9 expression. Thus, our results suggest that HES is a potential therapeutic agent which has anticancer effects on glioblastoma.

Antimetastatic effects of gambogic acid are mediated via the actin cytoskeleton and NF-κB pathways in SK-HEP1 cells.

Hepatocellular carcinoma (HCC) is one of the most malignant and frequent cancers with a high metastatic potential. The prevention of HCC metastasis is a critical target for effective therapies in HCC. Gambogic acid (GA), a natural compound obtained from Garcinia hanburyi has reported anticancer activity in cell lines. However, the antimetastatic mechanisms of GA are unclear, particularly with respect to HCC. In this study, the influence of GA on migration and invasion of SK-HEP1 cells was evaluated. At concentrations above 0.6 µM, GA reduced cell proliferation in SK-HEP1 cells without affecting proliferation of noncancerous HEK-293 cells. GA also suppressed migration and invasion of SK-HEP1 cells. GA downregulated the expression of the integrin ß1/rho family GTPase signaling pathway, suppressed the actin rearrangement related to cell cytoskeleton and migration and decreased matrix metalloproteinases MMP-2, MMP-9, and NF-κB expression involved in cancer invasion. These results suggest that GA may be a potential lead in developing an antimetastatic therapeutic for the treatment of HCC.

Dieckol enhances the expression of antioxidant and detoxifying enzymes by the activation of Nrf2-MAPK signalling pathway in HepG2 cells.

Dieckol was previously reported to exhibit antioxidant and anticancer activities in vitro studies. In this study, we characterised the mechanism underlying the dieckol-mediated expression of antioxidant and detoxifying enzymes. Dieckol suppressed the production of intracellular reactive oxygen species in the presence or absence of H2O2 and increased glutathione level in HepG2 cells. Dieckol enhanced the activities of antioxidant enzymes, and the expression of detoxifying enzymes including heme oxygenase-1 (HO-1), NAD(P)H:quinine oxidoreductase 1 (NQO1), and glutathione S-transferase (GST) in HepG2 cells. Enhanced expression of antioxidant and detoxifying enzymes by dieckol was presumed to be the activation of the nuclear factor erythroid-derived 2-like 2 (Nrf2) demonstrated by its nuclear translocation and transcriptional activity via activation of mitogen-activated protein kinases in HepG2 cells. Furthermore, we demonstrated dieckol induced the expression of HO-1 in mouse liver. These results demonstrate that the dieckol-mediated cytoprotection in HepG2 cells is mediated through a ROS-independent up-regulation of antioxidant and detoxifying enzymes via Nrf2 activation as well as its intrinsic antioxidant activity, suggesting that dieckol may be used as a natural cytoprotective agent.