11 research outputs found
Analysis of Compounds That Interfere with Herpes Simplex Virus–Host Receptor Interactions Using Surface Plasmon Resonance
The
entry of herpes simplex virus into host cells involves a complex
series of events that require concerted inputs from multiple HSV glycoproteins.
Among these glycoproteins, the gD protein of HSV-1 and HSV-2 plays
an important role for host receptor binding and membrane fusion. In
the present study, we evaluated the ability of different sulfated
saccharides to interfere with gD–host receptor (HVEM) interactions
using our recently reported molecular assay (Gopinath, S. C. B.; Hayashi, K.; Kumar, P. K. R. J. Virol. 2012, 86, 6732−6744). Initially, we tested the ability of heparan sulfate to interfere
with the HVEM–HSV-1 gD interaction and found that heparan sulfate
is able to interfere efficiently, with an apparent EC<sub>50</sub> of 2.1 μM. In addition, we tested different synthetic sulfated
polysaccharides and natural sulfated polysaccharides from an edible
alga, Sargassum horneri, after fractionation
into different sizes and sulfate and uronic acid contents. Six polysaccharides
isolated from S. horneri were found
to efficiently interfere with the HVEM–gD interaction. Three
others caused moderate interference, and five caused weak interference.
These results were confirmed with plaque assays, and good agreement
was found with the results of the SPR assay for the identification
of compounds that interfere with HVEM–HSV-1 gD binding. These
studies suggest that our molecular assay based on surface plasmon
resonance is not only useful for the analysis of viral–host
protein interactions but is also applicable for the routine screening
of compounds to identify those that interfere with the first step
of viral entry, thus facilitating the rapid development of novel antiviral
compounds that target HSV
Impact of Sulfur Fumigation on Ginger: Chemical and Biological Evidence
We previously found that sulfur fumigation, a commonly
used controversial
method for the post-harvest handling of ginger, induces the generation
of a compound in ginger, which was speculated to be a sulfur-containing
derivative of 6-shogaol based on its mass data. However, the chemical
and biological properties of the compound remain unknown. As a follow-up
study, here we report the chemical structure, systemic exposure, and
anticancer activity of the compound. Chromatographic separation, nuclear
magnetic resonance analysis, and chemical synthesis structurally elucidated
the compound as 6-gingesulfonic acid. Pharmacokinetics in rats found
that 6-gingesulfonic acid was more slowly absorbed and eliminated,
with more prototypes existing in the blood than 6-shogaol. Metabolism
profiling indicated that the two compounds produced qualitatively
and quantitatively different metabolites. It was further found that
6-gingesulfonic acid exerted significantly weaker antiproliferative
activity on tumor cells than 6-shogaol. The data provide chemical
and biological evidence that sulfur fumigation may impair the healthcare
functions of ginger
Structure Elucidation and Immunomodulatory Activity of A Beta Glucan from the Fruiting Bodies of <i>Ganoderma sinense</i>
<div><p>A polysaccharide named GSP-2 with a molecular size of 32 kDa was isolated from the fruiting bodies of <i>Ganoderma sinense</i>. Its structure was well elucidated, by a combined utilization of chemical and spectroscopic techniques, to be a β-glucan with a backbone of (1→4)– and (1→6)–Glc<i>p</i>, bearing <i>terminal-</i> and (1→3)–Glc<i>p</i> side-chains at <i>O</i>-3 position of (1→6)–Glc<i>p</i>. Immunological assay exhibited that GSP-2 significantly induced the proliferation of BALB/c mice splenocytes with target on only B cells, and enhanced the production of several cytokines in human peripheral blood mononuclear cells and derived dendritic cells. Besides, the fluorescent labeled GSP-2 was phagocytosed by the RAW 264.7 cells and induced the nitric oxide secretion from the cells.</p></div
Nitric oxide production and phagocytosis of GSP-2-treated RAW 264.7 cells.
<p>A: The levels of NO production of RAW264.7 cells were assessed after incubation with GSP-2 or LPS for 24 h using Griess assay. All data are expressed as mean ± SEM of three individual experiments (n = 12). Significant difference: *, P<0.05; **, P<0.01; ***, P<0.001 for difference from culture without treatment. B: The viable RAW 264.7 cell populations were gated (left) and the FITC-positive population was shown in fluorescent-GSP-2-labeled cells (lower right histogram). C: RAW 264.7 cells were incubated with 10 µg GSP-2 (fluro-labeled) for 24 h. The cells were imaged at the 1<sup>st</sup> and 24<sup>th</sup> hour (lower histograms). After 24 h incubation, the cells were collected, washed and resuspended in PBS and the fluorescence of the samples was detected by flow cytometry. (Upper histograms: viable RAW 264.7 cells without fluorescent labeled GSP-2, lower histograms, 1<sup>st</sup> hour: viable RAW 264.7 cells with fluorescent labeled GSP-2, 24<sup>th</sup> hour).</p
GC–MS test result for the methylated sugar moieties of the polysaccharide GSP-2 <sup>a</sup><sup>,</sup><sup>b</sup>.
a<p>The result were tested on DB-5 GC-MS column.</p>b<p>All the sugar residues were primarily identified by their MS spectrum and further confirmed by their relative retention time to 2,3,4,6-Me4-Glc.</p
Stimulating effect of GSP-2 on the proliferation of the mouse splenocytic B and T cells.
<p>Freshly fractionated splenocyte, splenic B (solid bars) and T (open bars) cells were stimulated with, or without (<i>Medium</i>), dextran or GSP (30 µg/ml). LPS (2 µg/ml) and ConA (2 µg/ml) were included as controls. In a parallel experiment, mouse splenocytes were stimulated with, or without (<i>Medium</i>), GSP (30 µg/ml), dextran (30 µg/ml) or LPS (10 µg/ml) in triplicate wells in the presence, or absence, of PMB. <sup>3</sup>H-TdR was added to the cultures for the last 8 hrs of incubation and then <sup>3</sup>H-TdR incorporation (CPM) of each well counted. A: Parallel experiment to exclude the influence of the endotoxin contamination; B: Stimulating effect of GSP-2 to the mouse splenocytic B and T cells. All results are presented as mean ± SEM, *, P<0.05; **, P<0.01; ***, P<0.001 for difference from culture without treatment. (n = 9, repeated 3 times).</p
Cytokine productions of GSP-2-treated PBMCs.
<p>Culture supernatants were collected 24-2 and the cytokines concentrations were specifically determined by ELISA. All data are expressed as mean ± SEM of three individual experiments (n = 12). Differences between the treated and untreated control groups were compared using one-way ANOVA. * P<0.05, ** P<0.01, *** P<0.001 for difference from culture without treatment.</p
Amino acid composition of the polysaccharide (GSP-2) isolated from the fruiting bodies of <i>Ganoderma sinense</i><sup>a</sup>.
a<p>All the result were tested by a HITACHI automatic amino acid analyzer.</p
HPLC profile, methylation analysis result and 1D NMR spectrums of GSP-2.
<p>A: HPLC profile of GSP-2. Samples (2 mg/mL, 10 µL) were analyzed on an Agilent 1100 system equipped with an ELSD detector and TSK GMPW<sub>XL</sub> gel filtration columns (7.8×300 mm×2), with 20 mM CH<sub>3</sub>COONH<sub>4</sub> as mobile phase at 0.6 mL/min and column temperature at 40°C. Commercially available T-series dextrans (MW 2000, 670, 410, 270, 150, 80, 50, 12, 5 and 1 kD). B: Methylation analysis result of GSP-2. GC-MS tests for methylation analysis were measured with a DB-5 column (30 m×0.25 mm×0.25 µm), and at temperatures programmed from 170–225 °C at 2 °C/min and then hold on 10 min, in the figure, a: <i>T</i>-Glc<i>p</i>, b: <i>1,3</i>-linked Glc<i>p</i>, c: <i>1,4</i>-linked Glc<i>p</i>, d: <i>1,6</i>-linked Glc<i>p</i>, e: <i>1,3,6</i>-linked Glc<i>p</i>. C-E: 1D NMR (<sup>1</sup>H, <sup>13</sup>C, DEPT) spectra of GSP-2 in D<sub>2</sub>O with TMS as external standard, obtained on a Bruker AM 500 spectrometer with a dual probe in the FT mode at room temperature.</p
2D NMR spectrums and proposed structure of the polysaccharides from the fruiting bodies of <i>G. sinense</i>.
<p>(A: HSQC spectrum, B: HMBC spectrum, C: NOESY spectrum, D proposed strcuture).</p