Jong-Shu Kim

Fucoidan from seaweed Fucus vesiculosus inhibits migration and invasion of human lung cancer cell via PI3K-Akt-mTOR pathways.

Background Recently there has been an increased interest in the pharmacologically active natural products associated with remedies of various kinds of diseases, including cancer. Fucoidan is a polysaccharide derived from brown seaweeds and has long been used as an ingredient in some dietary supplement products. Although fucoidan has been known to have anti-cancer activity, the anti-metastatic effects and its detailed mechanism of actions have been poorly understood. Therefore, the aims of this study were to demonstrate the anti-metastatic functions of fucoidan and its mechanism of action using A549, a highly metastatic human lung cancer cell line. Methods and Principal Findings Fucoidan inhibits the growth of A549 cells at the concentration of 400 µg/ml. Fucoidan treatment of non-toxic dose (0–200 µg/ml) exhibits a concentration-dependent inhibitory effect on the invasion and migration of the cancer cell via decreasing its MMP-2 activity. To know the mechanism of these inhibitory effects, Western blotting was performed. Fucoidan treatment down-regulates extracellular signal-related kinase 1 and 2 (ERK1/2) and phosphoinositide 3-kinase (PI3K)–Akt–mammalian target of rapamycin (PI3K-Akt-mTOR) pathways. Furthermore, fucoidan decreases the cytosolic and nuclear levels of Nuclear Factor-kappa B (p65). Conclusions/Significance The present study suggests that fucoidan exhibits anti-metastatic effect on A549 lung cancer cells via the down-regulation of ERK1/2 and Akt-mTOR as well as NF-kB signaling pathways. Hence, fucoidan can be considered as a potential therapeutic reagent against the metastasis of invasive human lung cancer cells.

Cytotoxicity and hemolytic activity of jellyfish Nemopilema nomurai (Scyphozoa: Rhizostomeae) venom

The recent bloom of a giant jellyfish Nemopilema nomurai has caused a danger to sea bathers and fishery damages in the waters of China, Korea, and Japan. The present study investigated the cytotoxic and hemolytic activities of crude venom extract of N. nomurai using a number of in vitro assays. The jellyfish venom showed a much higher cytotoxic activity in H9C2 heart myoblast than in C2C12 skeletal myoblast (LC(50)=2 microg/mL vs. 12 microg/mL, respectively), suggesting its possible in vivo selective toxicity on cardiac tissue. This result is consistent with our previous finding that cardiovascular function is a target of the venom. In order to determine the stability of N. nomurai venom, its cytotoxicity was examined under the various temperature and pH conditions. The activity was relatively well retained at low environmental temperature (<or=20 degrees C) and dramatically lost at high temperature (>or=60 degrees C). In pH stability test, the venom has abruptly lost its activity at low pH environment (pH<or=4). Interestingly enough, however, its activity was not significantly affected even at the highest pH environment tested (pH<or=12) in the present study. Additionally, hemolytic activity of the venom was examined using the erythrocytes of cat, dog, human, rabbit and rat. Venom concentration-dependent hemolysis could be observed from 10 microg/mL of protein equivalents or higher with variable potencies in different species, among which dog erythrocyte was the most susceptible to the venom (EC(50)=151 microg/mL). SDS-PAGE analysis of N. nomurai venom showed the molecules of 20-40 kDa and 10-15 kDa appeared to be the major protein components of the venom.

Brown alga Ecklonia cava attenuates type 1 diabetes by activating AMPK and Akt signaling pathways.

The antidiabetic therapeutic effect of Ecklonia cava, a brown alga, was investigated using streptozotocin-induced type 1 diabetes mellitus rats and C2C12 myoblasts. The methanol extract of E. cava (ECM), having a strong radical scavenging activity, significantly reduced plasma glucose level and increased insulin concentration in type 1 diabetes mellitus rats. Moreover, the elevation of plasma ALT in diabetic rats was dramatically restored near to normal range by the treatment of ECM, whereas AST level was not meaningfully altered in any group throughout the experiment. The characteristic indications of diabetes, such as polyphagia and polydipsia, were greatly improved by ECM treatment as well. The mechanism of action of ECM appears to be, at least partially, mediated by the activation of both AMP-activated protein kinase/ACC and PI-3 kinase/Akt signal pathways. Taken together, the present results suggest that E. cava has both in vivo and in vitro antidiabetic effects.

Differential expression of two functional serine/threonine protein kinases from soyabean that have an unusual acidic domain at the carboxy terminus

Abstract Two soybean cDNA clones, SPK-3 and SPK-4, encoding putative protein kinases were isolated and characterized. Both cDNAs encoded approximately 40-kDa serine/threonine kinases with unusual stretches of acidic amino acids in their carboxy-terminal regions, which are highly homologous to PKABA1 from wheat and ASKs from Arabidopsis. These kinases are encoded by one- or two-copy genes in the soybean genome. Notably, SPK-3 and -4 showed different patterns of expression in various soybean tissues. SPK-3 is highly expressed in dividing and elongating tissues of young seedlings but relatively weakly in tissues of mature plants. In contrast, SPK-4 showed relatively high and constitutive expression in all the tissues examined except for leaf tissues of mature plants. Although various stressors, such as dehydration and high salinity, increased the expression of both genes, the induction kinetics were different. The two genes also differed in their response to abscisic acid (ABA). SPK-3 was induced but SPK-4 was not affected by exogenously supplied abscisic acid. In accordance with these expression data analysis of the activity of a chimeric SPK-3 promoter::β-glucuronidase (GUS) reporter gene by transient expression in tobacco leaves confirmed the inducibility of SPK-3 by salt and ABA. Polyclonal antibodies raised against a recombinant SPK-4 protein produced in Escherichia coli specifically recognized both recombinant SPK-3 and -4 proteins. Kinase assays using affinity-purified SPK-4/antibody complexes with crude soybean extracts as substrate identified specific phosphorylation of two 41 and 170 kDa soybean proteins that were phosphorylated on serine residues. Taken together, our results suggest that SPK-3, and/or SPK-4 are functional serine protein kinase(s). Furthermore, SPK-3 and -4 may play different roles in the transduction of various environmental stresses.

Saffron (Crocus sativus L.) increases glucose uptake and insulin sensitivity in muscle cells via multipathway mechanisms

Saffron (Crocus sativus Linn.) has been an important subject of research in the past two decades because of its various biological properties, including anti-cancer, anti-inflammatory, and anti-atherosclerotic activities. On the other hand, the molecular bases of its actions have been scarcely understood. Here, we elucidated the mechanism of the hypoglycemic actions of saffron through investigating its signaling pathways associated with glucose metabolism in C(2)C(12) skeletal muscle cells. Saffron strongly enhanced glucose uptake and the phosphorylation of AMPK (AMP-activated protein kinase)/ACC (acetyl-CoA carboxylase) and MAPKs (mitogen-activated protein kinases), but not PI 3-kinase (Phosphatidylinositol 3-kinase)/Akt. Interestingly, the co-treatment of saffron and insulin further improved the insulin sensitivity via both insulin-independent (AMPK/ACC and MAPKs) and insulin-dependent (PI 3-kinase/Akt and mTOR) pathways. It also suggested that there is a crosstalk between the two signaling pathways of glucose metabolism in skeletal muscle cells. These results could be confirmed from the findings of GLUT4 translocation. Taken together, AMPK plays a major role in the effects of saffron on glucose uptake and insulin sensitivity in skeletal muscle cells. Our study provides important insights for the possible mechanism of action of saffron and its potential as a therapeutic agent in diabetic patients.

Evaluation of Antimicrobial Activity of the Methanol Extracts from 8 Traditional Medicinal Plants

The methanol extract of 12 medicinal plants were evaluated for its antibacterial activity against Gram-positive (5 strains) and Gram-negative bacteria (10 strains) by assay for minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) . The antibacterial activity was determined by an agar dilution method (according to the guidelines of Clinical and Laboratory Standard Institute) . All the compounds (12 extracts) of the 8 medicinal plants (leaf or root) were active against both Gram-negative and Gram-positive bacteria. Gram-negative showed a more potent action than Gram positive bacteria. The MIC concentrations were various ranged from 0.6 μg/ml to 5000 μg/ml. The lowest MIC (0.6 μg/ml) and MBC (1.22 μg/ml) values were obtained with extract on 4 and 3 of the 15 microorganisms tested, respectively.

Antimetastatic activity of polyphenol-rich extract of Ecklonia cava through the inhibition of the Akt pathway in A549 human lung cancer cells.

An ethyl acetate extract (ECE) of a brown alga, Ecklonia cava, was examined for its anti-metastatic effect, using A549 human lung carcinoma cells. ECE treatment significantly suppressed the migration and invasion of A549 cells in a concentration-dependent manner. It also strongly down-regulated the matrix metalloproteinase (MMP)-2 activity of the cancer cells by gelatin zymography assay. For elucidating its mechanism of action in cancer cell metastasis, ECE was further investigated for various cell signalling pathways, including JNK, ERK, p38, and Akt. In this, ECE showed an anti-metastatic effect in a concentration- and time-dependent manner by the mechanism of suppression of Akt and p38, but not JNK and ERK. These results, for the first time, suggest that ECE (a polyphenol-enriched, highly anti-oxidative fraction of brown alga, E. cava) may have therapeutic potential in metastatic lung cancer, based on its strong inhibitory effects on the migration and invasiveness of A549 human lung adenocarcinoma cells.

protective effect of tetracycline against dermal toxicity induced by Jellyfish venom.

Background Previously, we have reported that most, if not all, of the Scyphozoan jellyfish venoms contain multiple components of metalloproteinases, which apparently linked to the venom toxicity. Further, it is also well known that there is a positive correlation between the inflammatory reaction of dermal tissues and their tissue metalloproteinase activity. Based on these, the use of metalloproteinase inhibitors appears to be a promising therapeutic alternative for the treatment of jellyfish envenomation. Methodology and Principal Findings Tetracycline (a metalloproteinase inhibitor) has been examined for its activity to reduce or prevent the dermal toxicity induced by Nemopilema nomurai (Scyphozoa: Rhizostomeae) jellyfish venom (NnV) using in vitro and in vivo models. HaCaT (human keratinocyte) and NIH3T3 (mouse fibroblast) incubated with NnV showed decreases in cell viability, which is associated with the inductions of metalloproteinase-2 and -9. This result suggests that the use of metalloproteinase inhibitors, such as tetracycline, may prevent the jellyfish venom-mediated local tissue damage. In vivo experiments showed that comparing with NnV-alone treatment, tetracycline pre-mixed NnV demonstrated a significantly reduced progression of dermal toxicity upon the inoculation onto rabbit skin. Conclusions/Significance It is believed that there has been no previous report on the therapeutic agent of synthetic chemical origin for the treatment of jellyfish venom-induced dermonecrosis based on understanding its mechanism of action except the use of antivenom treatment. Furthermore, the current study, for the first time, has proposed a novel mechanism-based therapeutic intervention for skin damages caused by jellyfish stings.

Thimerosal induces oxidative stress in HeLa S epithelial cells.

Thimerosal is one of the most widely used preservatives and is found in a variety of biological products, including vaccines, contact lens cleaning solutions, and cosmetics. It has been reported to have harmful effects on epithelial tissues, such as causing conjunctivitis or contact dermatitis. However, the molecular mechanism of its toxicity has not been characterized using epithelial tissues. In the present study, we report that reactive oxygen species play a key role in thimerosal-induced cytotoxicity in HeLa S epithelial cells. Thimerosal significantly reduced HeLa S cell viability and it was associated with a decrease in intracellular glutathione levels. Flow cytometric cell cycle analysis showed a marked increase in the hypodiploidic cell population, indicating apoptosis of thimerosal-treated cells. The apoptotic cell death of epithelial cells was confirmed by observing a significant increase of caspase-3 activity in the cytosolic fraction of the treated cells. Thimerosal also induced a concentration-dependent increase of genomic DNA fragmentation, a biochemical hallmark of apoptosis. Hoechst 33342 nuclear staining demonstrated apoptotic-fragmented multinuclei in thimerosal-treated cells. All the thimerosal-mediated toxic responses observed in the present study were almost completely suppressed by pretreating cells with N-acetyl-l-cysteine, a radical scavenger. Taken together, these results suggest for the first time that epithelial cytotoxicity of thimerosal is mediated by oxidative stress.

Mutual synergistic toxicity between environmental toxicants: A study of mercury chloride and 4-nonylphenol.

Mercury chloride (HgCl(2)) and 4-nonylphenol (NP) are widespread environmental and industrial pollutants that are known to have toxic effects as well as endocrine disrupting activities. Although the individual effects of HgCl(2) and NP in liver have been relatively well recognized, little is known about the interaction of NP and HgCl(2) during the induction of their toxicity. In the current study, we investigated the synergism between HgCl(2) and NP using HepG2 cells. Surprisingly, the concurrent treatment of HepG2 with HgCl(2) and NP induced a significant cytotoxicity at concentrations where neither of them have any cytotoxic effect when treated alone. The cytotoxicity of NP is enhanced in the presence of HgCl(2) (a shift from 74.9 to 47.4μM in LC(50)) and vice versa (a shift from 94.9 to 66.3μM in LC(50)). Estrogen receptor antagonists such as ICI 182,780 did not protect HepG2 cells from these cytotoxic insults. Whereas the intracellular level of reduced form glutathione (GSH) was considerably decreased upon the co-treatment with NP and HgCl(2). Furthermore, the synergistic cytotoxicity was significantly inhibited by 20-mM N-acetylcysteine (NAC). These results indicate that the mutual synergistic cytotoxicity of HgCl(2) and NP on HepG2 cell is not associated with estrogen receptor signaling but mediated by reactive oxygen species (ROS) generation. In our real life, we are continuously and often simultaneously exposed to many different kinds of environmental pollutants. The present study suggests a mechanism of potential synergistic adverse effects of these toxic pollutants.