Effect of various combinations of the Cdt subunits for their ability to induce G₂ arrest and effect of CdtA and CdtC on CdtB-induced G₂ arrest.

<p>The cells were analyzed for cell cycle distribution by treated with Cdt proteins alone or in various combinations (as indicated) following staining with propidium iodide. (A), PAM cell; (B), PK-15 cell; (C) Jurkat cell or (D) PAM cells were exposed to 200 µl CdtB alone (50 µg/ml) or in the presence of CdtA (50 µg/ml) or CdtC (50 µg/ml). The cells were analyzed for cell cycle distribution by flow cytometry based upon propidium iodide fluorescence. The data represent the mean ± SEM of three experiments; at least 20,000 cells were analyzed per sample.</p

Phylogenetic analysis of two Cdts in the strain SH0165 and 29755 on the basis of the ClustalW method in Lasergene software (DNASTAR).

<p>(A), CdtA; (B), CdtB; (C), CdtC; (D), the relationship between CdtB in strain SH0165 and 29755 and that in the other bacterium species produced CdtB, on the basis of the ClustalW method.</p

Positive-specific iterated BLAST alignment of CdtB and assessment of CdtB mutants for their ability to induce G₂ arrest.

<p>(A), The alignment is taken directly from the final iteration. M, metal ion-binding residues; C, catalytic residues; asterisk, DNA residues. (B), PAM cells were exposed to medium alone (Negative control) or 200 µl CdtA and CdtC (50 µg/ml) in the presence of CdtB^WT (50 µg/ml) (Positive control) or CdtB^R118A, CdtB^H161Q, CdtB^D235A, CdtB^D267A or CdtB^H268Q (50 µg/ml). Cells were analyzed for cell cycle distribution 24 h after exposure to toxin subunits using flow cytometric analysis of propidium iodide fluorescence. The numbers in each panel represent the percentages of cells in G₂/M. Results are representative of three experiments.</p

Analisi dell'integrazione delle tecnologie blockchain e Internet of Things

La tecnologia blockchain è emersa come un'innovazione dalle potenzialità di segnare un importante svolta nel modo di condividere le informazioni. Il punto di forza di tale tecnologia è quello di offrire soluzioni di consenso in modo da garantire la fiducia in ambienti distribuiti. Un'altra importante caratteristica della blockchain è quella di risolvere i problemi di sicurezza i quali rappresentano uno degli aspetti cruciali nell’IoT. Il lavoro svolto in questa tesi mira ad offrire una panoramica dettagliata delle due tecnologie in modo da capire quali siano i punti dove l’integrazione di esse possa essere più fattibile rispetto ad altri. Tuttavia, nonostante vi siano ancora molte sfide da affrontare, si può concludere che la combinazione delle due tecnologie possa fungere da apriporta a nuove opportunità di business, difficilmente immaginabili con gli strumenti di cui si disponeva qualche anno fa

Structural alignment and assessment of CRAC site mutants in CdtC subunits for their ability to induce G₂ arrest.

<p>(A), structural alignment of CRAC site in <i>H. parasuis</i>, CdtC in <i>A. actinomycetemcomitans</i> and the classical CRAC sites. (B), assessment of CRAC site mutants for their ability to induce G₂ arrest. The cells were incubated with medium alone, 50 µg/ml CdtABC^WT, 50 µg/ml CdtABC^V77Y or 50 µg/ml CdtABC^V77A for 24 h, stained with propidium iodide, and analyzed for cell cycle distribution by flow cytometry as described above.</p

Detection of the expression and Cdt activity of CdtB from 15 <i>H. parasuis</i> reference strains and 109 <i>H. parasuis</i> clinical isolates.

<p>(A), Whole cell proteins of 15 <i>H. parasuis</i> reference strains were applied to western bolt and detected by anti-CdtB specific antibody.1–15 represents 15 <i>H. parasuis</i> reference strains, 0165 strain was used as positive control. (B), Cells were incubated with medium alone (negative control), 50 µg/ml CdtABC^WT or 200 µg/ml whole cell proteins of 15 <i>H. parasuis</i> reference strains for 24 h, stained with propidium iodide, and analyzed for cell cycle distribution by flow cytometry as described above. (C), the whole cell proteins of 109 clinical isolates were applied to western bolt and detected by anti-CdtB specific antibody. M, marker; C, 0165 strain was used as positive control.</p

Hydroxylated Bisabolol Oxides: Evidence for Secondary Oxidative Metabolism in <i>Matricaria chamomilla</i>

German chamomile (<i>Matricaria chamomilla</i>) is one of the most popular medicinal plants used in Western herbal medicine. Among the various phytochemicals present in the essential oil of the flowers of German chamomile, bisabolol and its oxidized metabolites are considered as marker compounds for distinguishing different chemotypes. These compounds are influential in mediating the aroma of the essential oil of <i>M. chamomilla</i> and contribute to the therapeutic properties (anti-inflammatory, antibacterial, insecticidal, and antiulcer) of this species. In order to find other possible bisabolol derivatives as marker compounds for authentication of German chamomile in botanical and commercial products, an in-depth investigation using a GC-assisted fractionation procedure was performed on nonpolar fractions. As a result of this approach, three new hydroxylated derivatives of bisabolol oxides A and B (<b>1</b>–<b>3</b>) have been isolated from <i>M. chamomilla</i>. Plausible biogenetic pathways are presented

New Triterpenoid Saponins from Ilex cornuta and Their Protective Effects against H₂O₂‑Induced Myocardial Cell Injury

Five new triterpenoid saponins, <b>1</b>–<b>5</b>, together with 10 known ones, <b>6</b>–<b>15</b> were isolated from the aerial parts of Ilex cornuta. The structures of compounds <b>1</b>–<b>5</b> were determined as 3β-<i>O</i>-α-l-arabinopyranosyl-19α,23-dihydroxy-20α-urs-12-en-28-oic acid 28-<i>O</i>-β-d-glucopyranosyl ester, <b>1</b>; 3β-<i>O</i>-β-d-glucopyranosyl-(1→2)-α-l-arabinopyranosyl-19-hydroxy-20α-urs-12-en-28-oic acid 28-<i>O</i>-β-d-glucopyranosyl ester, <b>2</b>; 19α,23-dihydroxyurs-12-en-28-oic acid 3β-<i>O</i>-β-d-glucuronopyranoside-6-<i>O</i>-methyl ester, <b>3</b>; 19α,23-dihydroxyurs-12-en-28-oic acid 3β-<i>O</i>-[β-d-glucuronopyranoside-6-<i>O</i>-methyl ester]-28-<i>O</i>-β-d-glucopyranosyl ester, <b>4</b>; and 3β-<i>O</i>-[α-l-arabinopyranosyl-(1→2)-β-d-glucuronic acid]-oleanolic acid 28-<i>O</i>-β-d-glucopyranosyl ester, <b>5</b>, on the basis of spectroscopic analyses (IR, ESI-MS, HR-ESI-MS, 1D and 2D NMR) and chemical reactions. Protective effects of compounds <b>1</b>–<b>15</b> were tested against H₂O₂-induced H9c2 cardiomyocyte injury, and the data showed that compounds <b>1</b>, <b>4</b>, <b>6</b>, and <b>13</b> had significant cell-protective effects. No significant DPPH radical scavenging activity was observed for compounds <b>1</b>–<b>15</b>

Quality Evaluation of Terpinen-4-ol-Type Australian Tea Tree Oils and Commercial Products: An Integrated Approach Using Conventional and Chiral GC/MS Combined with Chemometrics

GC/MS, chiral GC/MS, and chemometric techniques were used to evaluate a large set (<i>n</i> = 104) of tea tree oils (TTO) and commercial products purported to contain TTO. Twenty terpenoids were determined in each sample and compared with the standards specified by ISO-4730-2004. Several of the oil samples that were ISO compliant when distilled did not meet the ISO standards in this study primarily due to the presence of excessive <i>p</i>-cymene and/or depletion of terpinenes. Forty-nine percent of the commercial products did not meet the ISO specifications. Four terpenes, viz., α-pinene, limonene, terpinen-4-ol, and α-terpineol, present in TTOs with the (+)-isomer predominant were measured by chiral GC/MS. The results clearly indicated that 28 commercial products contained excessive (+)-isomer or contained the (+)-isomer in concentrations below the norm. Of the 28 outliers, 7 met the ISO standards. There was a substantial subset of commercial products that met ISO standards but displayed unusual enantiomeric +/– ratios. A class predictive model based on the oils that met ISO standards was constructed. The outliers identified by the class predictive model coincided with the samples that displayed an abnormal chiral ratio. Thus, chiral and chemometric analyses could be used to confirm the identification of abnormal commercial products including those that met all of the ISO standards

New Triterpenoid Saponins from Green Vegetable Soya Beans and Their Anti-Inflammatory Activities

Ten compounds were isolated and identified from green vegetable soya beans, of which five are new triterpenoid saponins (<b>1</b>–<b>5</b>) and five are known compounds (<b>6</b>–<b>10</b>). The chemical structures of the five triterpenoid saponins (<b>1</b>–<b>5</b>) were elucidated to be 3<i>β,</i>24-dihydroxy-22β,30-epoxy-30-oxoolean-12-en 3-<i>O</i>-α-l-rhamnopyranosyl-(1 → 2)-β-d-xylopyranosyl-(1 → 2)-β-d-glucuronopyranoside, <b>1</b>; 3<i>β,</i>24-dihydroxy-22β,30-epoxy-30-oxoolean-12-en 3-<i>O</i>-α-l-rhamnopyranosyl-(1 → 2)-β-d-(3″-<i>O</i>-formyl)-galactopyranosyl-(1 → 2)-β-d-glucuronopyranoside, <b>2</b>; 22-keto-3β,24-dihydroxy oleanane-12-ene 3-<i>O</i>-α-l-rhamnopyranosyl-(1 → 2)-β-d-(3″-<i>O</i>-formyl)-galactopyranosyl-(1 → 2)-β-d-glucuronopyranoside, <b>3</b>; 3β,22β,24-trihydroxy oxyolean-18(19)-ene-29-acid 3-<i>O</i>-α-l-rhamnopyranosyl-(1 → 2)-β-d-galactopyranosyl-(1 → 2)-β-d-glucuronopyranoside, <b>4</b>; and punicanolic acid 3-<i>O</i>-α-l-rhamnopyranosyl-(1 → 2)-β-d-galactopyranosyl-(1 → 2)-β-d-glucuronopyranoside, <b>5</b> from the spectroscopic data (IR, GTC/FID, HR-ESI-MS, and 1D and 2D NMR). The nitric oxide release inhibitions of compounds <b>1</b>–<b>10</b> in LPS-stimulated RAW264.7 cells were evaluated, and the data suggested that compounds <b>1</b>, <b>2</b>, and <b>5</b> might possess moderate anti-inflammatory activities, with IC₅₀ values of 18.8, 16.1, and 13.2 μM, respectively