23 research outputs found
Synthesis of the MN Ring of Caribbean Ciguatoxin C‑CTX‑1 via Desymmetrization by Acetal Formation
The MN ring of Caribbean ciguatoxin C-CTX-1 was synthesized
from
a meso-syn-2,7-dimethyloxepane derivative
corresponding to the M ring via desymmetrization by acetal formation
with a camphor derivative, followed by construction of the N ring
via the Horner–Wadsworth–Emmons reaction and acetal
formation. The meso-syn-2,7-dimethyloxepane
derivative was synthesized via photoinduced electrocyclization of
a conjugated exo-diene under flow conditions, giving
a cyclobutene derivative, followed by ring expansion via oxidative
cleavage and diastereoselective reduction of a β-hydroxy ketone
Eliminated BMC transplantation-induced tissue recovery by HMGB1-inhibition.
<p>Reduced extracellular collagen deposition (<b>A–C;</b> picrosirius red = red), increased capillary density (<b>D–F;</b> Isolectin B4 = red), and increased proliferation (<b>G–I;</b> Ki67 = red; nuclei = blue; cTnT = green) were observed in the border areas at day 28 after BMC transplantation (BMC group), compared to the PBS control (CON group). These effects were all abolished by anti-HMGB1 antibody neutralization (AB group), but not by control IgG administration (IgG group). Representative images of only BMC and AB groups are present (see <b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0076908#pone.0076908.s002" target="_blank">Figure S2</a></b> for additional images). Scale bars = 50 µm in <b>A, B, G, H</b> and 30 µm in <b>D, E</b>. *:<i>p</i><0.05 <i>versus</i> the CON group, <sup>†</sup>:<i>p</i><0.05 <i>versus</i> the BMC group, <sup>‡</sup>:<i>p</i><0.05 <i>versus</i> the IgG group, mean±SEM for n = 5∼7 in each group.</p
The effects of amitriptyline on the mRNA expression of neurotrophic/growth factors.
<p>Cortical astrocyte cultures and neuron-enriched cultures were treated with 25 µM of amitriptyline (Ami) for the indicated periods of time, and the mRNA expression levels of FGF-2 (A), BDNF (B), VEGF (C), GDNF (D), and NGF (E) were analyzed by real time PCR. The values are shown as the ratio of each neurotrophic/growth factor mRNA to GAPDH mRNA. The data are expressed as the means ± S.E.M. *<i>p</i><0.05, **<i>p</i><0.01, or ***<i>p</i><0.001 vs. basal (Dunnett's test; n = 4–11).</p
Different distribution of retained BMMNC between ventricular layers.
<p>At 5 minutes after IC injection of 8x10<sup>6</sup> PKH67-labeled BMMNC, the number of donor cells within the endocardium-side myocardium, central myocardium and epicardium-side myocardium were counted in cross-section of immunohistolabelling samples. The concentration of cells progressively increased from the epicardium to the endocardium (n = 4, <i>p</i><0.0001, One-way ANOVA followed by Bonferroni post-hoc test).</p
Poor donor cell survival and HMGB1 leakage after BMC transplantation.
<p>(<b>A</b>) Quantitative PCR for the male specific <i>sry</i> gene showed that the survival of male donor cells in female hearts was poor similarly in the BMC (BMC injection), IgG (BMC+control IgG injection), and AB (BMC+anti-HMGB1 antibody injection) groups at both days 3 and 28; n = 5∼7 in each point. (<b>B</b>) Clusters of DiI-labeled (red) donor BMCs were detected in the heart at day 3 after BMC transplantation. A higher magnification image of the yellow frame is shown. Green = cardiomyocytes (cTnT); blue = nuclei (DAPI). Scale bar = 300 µm. (<b>C</b>) ELISA showed that the circulating HMGB1 level was increased at 1 hour in the BMC group compared to the PBS injection control (CON group). *:<i>p</i><0.05 <i>versus</i> the CON group, mean±SEM for n = 5 each.</p
NA, but not amitriptyline, increases the FGF-2 mRNA expression via α1 and β receptors.
<p>A, Astrocyte cultures were treated with 10 µM of NA or 5-HT for the indicated periods of time, and the FGF-2 mRNA expression was analyzed by real time PCR. The data are expressed as the means ± S.E.M. **<i>p</i><0.01, or ***<i>p</i><0.001 vs. basal (Dunnett's test; n = 3). B, The results of reverse transcriptase PCR analysis of adrenergic receptor mRNA expression. Each lane represented the cDNA fragments of the α1A, α1B, α1D, α2A, α2B, α2C, β1, β2, or β3 adrenergic receptor amplified from the RNA of cortical astrocyte cultures (AS) and the adult rat cortex (Cx). Size markers were shown in Lane M. Cx samples were used as positive controls. C, Astrocyte cultures were pretreated with or without prazosin (pra, 10 µM), yohimbine (yoh, 10 µM) or propranolol (pro, 10 µM) for 30 min,and treated with NA (10 µM) for 3 h. The FGF-2 mRNA expression was analyzed by real time PCR. The data are expressed as the means ± S.E.M. (% of NA alone) ***<i>p</i><0.001 vs. basal, n.s.; not significantly different from the control (NA only) group, and †<i>p</i><0.05 or †††<i>p</i><0.001 vs. NA alone (Tukey’s HSD test; n = 3–4). D, Astrocyte cultures were pretreated with or without both pra (10 µM) and pro (10 µM) for 30 min and treated with amitriptyline (Ami, 25 µM) for 24 h. The FGF-2 mRNA expression was analyzed by real time PCR. The data are expressed as the means ± S.E.M. (% of Ami alone) ***<i>p</i><0.001 vs. basal and n.s.; not significantly different from Ami alone (Tukey’s HSD test; n = 3).</p
Modulation of innate immunity by BMC transplantation via released HMGB1.
<p>Accumulation of CD68<sup>+</sup> pan-macrophages (<b>A</b>), CD86<sup>+</sup> classically-activated pro-inflammatory M1 macrophages (<b>B</b>), and CD163<sup>+</sup> alternatively-activated anti-inflammatory M2 macrophages (<b>C</b>) in the border areas at day 3 after each treatment was assessed by immunolabeling. See <b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0076908#pone.0076908.s003" target="_blank">Figure S3</a></b> for representative images. Myocardial expression of <i>IL-10</i> (<b>D</b>), <i>IL-1β</i> (<b>E</b>)), and <i>TNF-α</i> (<b>F</b>) at day 3 after each treatment was measured by quantitative RT-PCR. *:<i>p</i><0.05 <i>versus</i> the CON group, <sup>†</sup>:<i>p</i><0.05 <i>versus</i> the BMC group, mean±SEM for n = 5∼7 in each group.</p
Relationship of cell-surface proteins and retention of BMMNC.
<p>The expression profiles of cell-surface proteins, including integrin and selectin ligand, on pre-injection BMMNC and BMMNC in the coronary effluent (collected over the five-minute duration post-IC injection of 8x10<sup>6</sup> BMMNC) were compared by flow cytometric analysis. No difference was found in any of the surface proteins investigated (n = 6 hearts were studied, unpaired T-test), suggesting that these cell-surface proteins are not critical for retention in normal hearts.</p
Initial retention of MSC after IC injection.
<p>(<b>A</b>) Following IC injection of 1×10<sup>6</sup> bone marrow-derived rat MSC into a rat heart using the same model, reduced numbers of donor MSC were found in the coronary effluent within the first minute, in comparison with IC injection of the same number of BMMNC (refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0158232#pone.0158232.g001" target="_blank">Fig 1</a>). n = 8 in each cell-type, <i>p</i><0.001, Two-way ANOVA followed by Bonferroni post-hoc test; *<i>p</i><0.05 versus Epicardium, <sup>†</sup>p<0.05 versus Myocardium. (<b>B</b>) The coronary effluent flow rate decreased immediately following IC injection of MSC but gradually recovered to the base line by 10 minutes (n = 8). (<b>C</b>) The distribution of MSC diameters prior to injection and in the coronary effluent were quantified and expressed as fractions (n = 8). There appeared to be a leftwards shift in the diameters of the coronary effluent cell population. (<b>D</b>) With knowledge of the cell numbers of each cell diameter, the retention efficiency of MSC having each diameter was calculated (n = 8). Larger MSC subsets were likely to be more frequently retained, but the retention rate was plateaued at ~80% with cell diameters ≥ 9 μm.</p
Amitriptyline, but not NA, induces FGF-2 expression through <i>de novo</i> synthesis in astrocyte cultures.
<p>A, Cells were pretreated with or without cycloheximide (Cyclo) for 10 min and treated with amitriptyline (Ami, 25 µM) for 24 h. The FGF-2 mRNA expression was analyzed by real time PCR. The data are expressed as the means ± S.E.M. (% of Ami alone) *<i>p</i><0.05 vs. basal, and †<i>p</i><0.05 or ††<i>p</i><0.01 vs. Ami alone (Tukey’s HSD test; n = 3). B, Cells were pretreated with or without Cyclo for 10 min or prazosin (pra, 10 µM) and propranolol (pro, 10 µM) for 30 min and treated with NA (10 µM) for 3 h. The FGF-2 mRNA expression was analyzed by real time PCR. The data are expressed as the means ± S.E.M. (% of NA alone) ***<i>p</i><0.001 vs. basal, †††<i>p</i><0.001 vs. NA alone, and ###<i>p</i><0.001 vs. NA and Cyclo (1 µM) (Tukey’s HSD test; n = 3). C, Cells were pretreated with or without Cyclo (1 µM) for 10 min and treated with 25 µM of Ami for 72 h. The FGF-2 protein levels were analyzed by an immunoblotting analysis. The data are expressed as the means ± S.E.M. (% of Ami alone) *<i>p</i><0.05 or **<i>p</i><0.01 vs. basal, and ††<i>p</i><0.01 or †††<i>p</i><0.001 vs. Ami alone (Tukey’s HSD test; n = 8). D, Cells were pretreated with or without Cyclo (1 µM) for 10 min and treated with 10 µM of NA for 24 h. The FGF-2 protein levels were analyzed by an immunoblotting analysis. The data are expressed as the means ± S.E.M. (% of NA alone) **<i>p</i><0.01 or ***<i>p</i><0.001 vs. basal, and †<i>p</i><0.05 or †††<i>p</i><0.001 vs. NA alone (Tukey’s HSD test; n = 4–5).</p