Toxicities of Four Parabens and Their Mixtures to and

The objective of this study was to determine toxicities of four parabens (methyl paraben, MP; ethyl paraben, EP; n-propyl paraben, PP; and n-butyl paraben; BP) and their mixtures to two aquatic microorganisms, Daphnia magna and Aliivibrio fischeri. Parabens are one of the widely used preservatives for personal care products, such as cosmetics, pharmaceuticals and food also. First, each paraben was treated to D. magna to measure the toxicity levels as LC20 and LC50. The results showed their value of MP (25.2 mg/L, 73.4 mg/L), EP (18.4 mg/L, 43.7 mg/L), PP (10.4 mg/L, 21.1 mg/L) and BP (3.3 mg/L, 11.2 mg/L). Then, each of the parabens was treated to A. fischeri and calculated their EC20 and EC50 by bioluminescence inhibition test. The results showed the values of MP (2.93 mg/L, 16.8 mg/L), EP (1.18 mg/L, 6.74 mg/L), PP (0.51 mg/L, 5.85 mg/L) and BP (0.21 mg/L, 2.34 mg/L). These four parabens belong to the group classified as being ‘harmful to aquatic organisms’ (above 10 mg/L, below 100 mg/L). After measuring the toxicity, EC20 values of two or more parabens were tested in order to investigate their toxicity. A total of ten combinations of four parabens were tested. As a result, the bioluminescence inhibition test of A. fischeri showed that the toxicity of mixture parabens was stronger than that of a single compound and combinations of three parabens showed the highest bioluminescence inhibition. These results showed that independent toxicity of paraben was maintained. Therefore, it can be predictable that the toxicity of paraben is getting stronger by the addition of other parabens

Analysis of lysosomal membrane proteins exposed to melanin in HeLa cells

Objectives There have been developed to use targeting ability for antimicrobial, anticancerous, gene therapy and cosmetics through analysis of various membrane proteins isolated from cell organelles. Methods It was examined about the lysosomal membrane protein extracted from lysosome isolated from HeLa cell treated by 100 ppm melanin for 24 hours in order to find associated with targeting ability to melanin using by 2-dimensional electrophoresis. Results The result showed 14 up-regulated (1.5-fold) and 13 down-regulated (2.0-fold) spots in relation to melanin exposure. Conclusions It has been found that lysosomal membrane proteins are associated with melanin to decolorize and quantity through cellular activation of lysosome

Toxic detection in mine water based on proteomic analysis of lysosomal enzymes in

Objectives Lysosome is the cell-organelle which is commonly used as biomonitoring tool in environmental pollution. In this study, the lysosomal proteomic of the yeast Saccharomyces cerevisiae was analyzed for utilization in the detection of toxic substances in mine water samples. Methods This work informs the expression of lysosomal proteomic in yeast in response with toxic chemicals, such as sodium meta-arsenite and tetracycline, for screening specific biomarkers. After that, a recombinant yeast contained this biomarker were constructed for toxic detection in pure toxic chemicals and mine water samples. Results Each chemical had an optimal dose at which the fluorescent protein intensity reached the peak. In the case of water samples, the yeast showed the response with sample 1, 3, 4, and 5; whereas there is no response with sample 2, 6, and 7. Conclusions The recombinant yeast showed a high ability of toxic detection in response with several chemicals such as heavy metals and pharmaceuticals. In the case of mine water samples, the response varied depending on the sample content

Anti-Inflammatory Action of an Antimicrobial Model Peptide That Suppresses the TRIF-Dependent Signaling Pathway via Inhibition of Toll-Like Receptor 4 Endocytosis in Lipopolysaccharide-Stimulated Macrophages.

Antimicrobial peptides (AMPs), also called host defense peptides, particularly those with amphipathic helical structures, are emerging as target molecules for therapeutic development due to their immunomodulatory properties. Although the antimicrobial activity of AMPs is known to be exerted primarily by permeation of the bacterial membrane, the mechanism underlying its anti-inflammatory activity remains to be elucidated. We report potent anti-inflammatory activity of WALK11.3, an antimicrobial model peptide with an amphipathic helical conformation, in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. This peptide inhibited the expression of inflammatory mediators, including nitric oxide, COX-2, IL-1β, IL-6, INF-β, and TNF-α. Although WALK11.3 did not exert a major effect on all downstream signaling in the MyD88-dependent pathway, toll-like receptor 4 (TLR4)- mediated pro-inflammatory signals were markedly attenuated in the TRIF-dependent pathway due to inhibition of the phosphorylation of STAT1 by attenuation of IRF3 phosphorylation. WALK11.3 specifically inhibited the endocytosis of TLR4, which is essential for triggering TRIF-mediated signaling in macrophage cells. Hence, we suggest that specific interference with TLR4 endocytosis could be one of the major modes of the anti-inflammatory action of AMPs. Our designed WALK11 peptides, which possess both antimicrobial and anti-inflammatory activities, may be promising molecules for the development of therapies for infectious inflammation

Effects of WALK11.3 on LPS-induced TLR4 endocytosis and DNP-HSA-induced FcεRI endocytosis.

<p>The RAW264.7 cells were incubated in the absence (A) or presence of dynasore (80 μM) (B), WALK11.3 (2 μM) (C), or WALK11.11 (2 μM) (D) for 1 h. After treatment with the LPS (100 ng/mL) for 30 min, flow cytometry was used to examine TLR4 endocytosis by measuring its surface expression on the cells. (E) The results are summarized as the mean fluorescence intensity of TLR4 staining at each time point. The error bars represent standard deviations from the average values of three independent experiments. (F) The endocytosis of FcεRI was also assessed by flow cytometry to detect the FcεRI level on the surface of the anti-DNP IgE-sensitized RBL-2H3 cells, which were stimulated with DNP-HSA (100 ng/mL) for 10 min, with or without pre-treatment (10 min) with WALK11.3 or dynasore. (G) The results are summarized as the mean fluorescence intensity of FcεRI/IgE staining. (H) DNP-HSA-induced degranulation in the RBL-2H3 cells was also estimated by measuring the activity of β-hexosaminidase in the cell culture media. The bar graphs present the mean values ± SEM of three independent experiments. *<i>p</i> < 0.05, **<i>p</i> < 0.01, or ***<i>p</i> < 0.001 compared with the DNP-HSA treated cells (ns, non-significant). [Student’s <i>t</i>-test (G and H) or a one-way ANOVA with Bonferroni’s multiple comparison test (E)].</p

Effects of WALK11.3 on the MyD88-dependent signaling pathway in LPS-stimulated RAW264.7 cells.

<p>The cells were pre-treated with WALK11.3 (2 μM) for 1 h before the treatment with LPS (100 ng/mL). After the designated incubation time with LPS, the phosphorylation levels of three MAPKs were determined (JNK, ERK, and p38) (A), SEK1/MKK4 phosphorylation levels (B) and degradation of IκB-α (C) by immunoblot analysis. The bar graphs present the mean values ± SEM of three independent experiments. *<i>p</i> < 0.05 or ***<i>p</i> < 0.001 versus the peptide-treated cells (ns, non-significant). [One-way ANOVA with Bonferroni’s multiple comparison test (A-C)]</p

Effects of WALK11.3 on the CD14 expression level and LPS binding on RAW264.7 cells.

<p>The cells, pre-treated with or without WALK11.3 (2 μM) for 1 h at 37°C, were stained with anti-CD14 and subsequently with FITC-conjugated anti-Rat IgG antibodies. (A) The cell surface expression of CD14 was then evaluated by flow cytometry. (B) The LPS binding was also analyzed with a Spectramax M2e system to detect the FITC-conjugated LPS (20 min treatment) on the RAW264.7 cells, with or without pre-, co-, or 10 min post-treatment with WALK11.3 for 20 min. The effect of pretreatment with PMB was also compared in the same manner. *<i>p</i> < 0.05 versus the FITC-LPS treated group (ns, non-significant). [Student’s <i>t</i>-test (B)].</p