EV71 infection results in morphological changes in mitochondria.

<p>SF268 cells were mock- (A & C) or infected with (B, D–G) EV71 at an m.o.i. of 1.25 for 48 hr, and processed for electron microscopic examination. The mock-infected cells had typical nucleus (N) and mitochondria (M). In EV71-infected cells, a number of mitochondria underwent changes in morphology, characterized by deranged cristae (D & E). The developing viral replication site (RS) was lined with ribosomes and was in proximity to mitochondria (D). Numerous single or double membrane-bound vesicles (MV) developed in EV71-infected cells, and some contained virus particles (VP). For A & B, bar represents 5 µm; for F, bar represents 1 µm; for C, D, E & G, bar represents 0.2 µm.</p

Mitochondrial ROS are essential to EV71 replication.

<p>(A) SF268 cells were mock- (−) or infected (+) with EV71 at an m.o.i. of 1.25, and treated without (<i>Con</i>) or with indicated concentrations of Mito-TEMPO. Forty-eight hours later, cells were subject to MitoSOX Red staining and flow cytometric analysis. The mean fluorescence intensity (MFI) of MitoSOX of mock- and infected cells is expressed as the percentage of that of uninfected cells. The results are presented as mean ± SD, n = 3. *p<0.05 vs. infected Con group. (B) SF268 cells were mock- or infected with EV71 at an m.o.i. of 1.25, and treated without or with 200 µM of Mito-TEMPO. Forty-eight hours later, cells were harvested for western blotting with antibodies to phosphorylated eIF2α and total eIF2α, viral protein 3D, and actin. A representative experiment out of three is shown here. (C) SF268 cells were mock- (−) or infected (+) with EV71 at an m.o.i. of 1.25, and treated without (<i>Con</i>) or with 200 µM of Mito-TEMPO. Forty-eight hours later, cells were analyzed for levels of EV71 genomic RNA. The results are presented as means ± SD n = 3. *p<0.05 vs. infected Con group.</p

EV71 infection induces ROS in neural cells in a time-dependent manner.

<p>SF268 cells were infected with EV71 at m.o.i. of 1.25, 1.5 and 2 for 0, 12, 24, 36, 48, 60 and 72 hr, and were subject to H₂DCFDA staining and flow cytometric analysis. (A) Representative histograms of cell counts (<i>counts</i>) vs. DCF fluorescence (<i>FL1-H</i>) for cells infected at an m.o.i. of 1.25 at indicated times are shown. (B) The mean fluorescence intensity (MFI) of DCF of infected cells is expressed as fold change relative to that of uninfected cells. The results are presented as mean±SD of three separate experiments.</p

Mitochondrial mass increases and expression of mitochondrial proteins changes in response to EV71 infection.

<p>(A) SF268 cells were mock- (−) or infected (+) with EV71 at an m.o.i. of 1.25 for 48 hr, and were subject to Mitotracker dye staining and flow cytometric analysis as described in <i>Materials and Methods</i>. The MFI of the stained cells is expressed relative to that of control cells. Results are mean ± SD, n = 3. *p<0.05 vs. uninfected cells. (B & C) Cells were un- or infected under the similar condition, and mitochondria were isolated for SDS-PAGE electrophoresis and silver staining (B). In the silver-stained gel, the leftmost lane corresponds to protein markers with respective molecular weights indicated alongside the bands. (C) Cells similarly infected were harvested for western blotting with indicated antibodies. A representative experiment out of three is shown here.</p

EV71 infection-induced oxygen consumption is associated with a reduction in respiratory efficiency.

<p>(A) SF268 cells were mock- or infected with EV71 at an m.o.i. of 1.25 for indicated times. Oxygen concentration was assayed with Clark oxygen electrode, and oxygen consumption rate (10⁻² µg O₂/5×10⁵ cells/min) was calculated accordingly. Results are mean ± SD, n = 6. *p<0.05 vs. uninfected cells. (B) SF268 cells were mock- or infected with EV71 at an m.o.i. of 1.25, and were treated without or with oligomycin. Oxygen concentration was assayed with Clark oxygen electrode, and oxygen consumption rate (10⁻² µg O₂/5×10⁵ cells/min) was calculated. Results are mean ± SD, n = 6. *p<0.05 vs. uninfected cells; ^#p<0.05, oligomycin-treated vs. untreated cells. (C) Oligomycin-sensitive oxygen consumption rate was calculated as the difference in the absence and presence of oligomycin. (D & E) The oxygen consumption was measured as described in (A) and (B). Data were normalized to the relative mitochondrial mass unit (MMU) of control and infected cells. Results are mean ± SD, n = 6. *p<0.05 vs. uninfected cells. (F) Oligomycin-sensitive oxygen consumption rate was calculated as described in (C), and normalized to the relative mitochondrial mass unit (MMU) of control and infected cells. (G–I) SF268 cells were mock- (−) or infected (+) with EV71 at an m.o.i. of 1.25 for 48 hr, and mitochondria were isolated and assayed for oxygen consumption rates (10⁻¹ µg O₂/mg mitochondrial protein/min) during state 3 (G) and 4_o (H) respiration as described in <i>Materials and Methods</i>. RCRs were calculated accordingly, and are shown (I). Results are mean ± SD, n = 6. *p<0.05 vs. uninfected cells.</p

Levels of ATP, ADP and AMP in EV71-infected cells.

<p>SF268 cells were mock- (<i>Con</i>) or infected (<i>Infected</i>) with EV71 at an m.o.i. of 1.25 for 48 hr, and were harvested for UPLC-based analyses of ATP, ADP and AMP. These adenine nucleotides are normalized to cellular protein content. Levels of ATP, ADP and AMP (A) and total adenine nucleotides (B) are shown. Results are mean ± SD, n = 3. *p<0.05 vs. uninfected cells.</p

Ligand concentration-dependent supramolecular complexes with uncoordinated carbonyl groups based on a new pyrazole carboxylic acid ligand

<div><p>One new pyrazole-based ligand, 1-carboxymethyl-3,5-dimethyl-1H-pyrazole-4-carboxylic acid (H₂cmdpca), has been synthesized and characterized. Structural analysis reveals that H₂cmdpca crystallizes in the monoclinic system and adopts a 3-D supramolecular network via the interaction of intermolecular hydrogen bonds. The reactions of Cd(II) ions with H₂cmdpca and 4,4′-bipyridine (4,4′-bpy) afforded three metal complexes, [Cd(4,4′-bpy)(Hcmdpca)₂(H₂O)₃]·H₂O (<b>1</b>), [Cd(4,4′-bpy)(Hcmdpca)₂(H₂O)]·3H₂O (<b>2</b>), and [Cd(4,4′-bpy)(Hcmdpca)₂(H₂O)] (<b>3</b>). Structural analyses reveal that these complexes are all monoclinic and <b>1</b>, <b>2</b>, and <b>3</b> exhibit mononuclear, 1-D chain, and 1-D with binuclear loop structures, respectively, which are further assembled into 3-D supramolecular frameworks through non-covalent interactions. <b>1</b> and <b>2</b> are true supramolecular isomers, while <b>2</b> and <b>3</b> are “pseudo-supramolecular” isomers. In addition, the thermal stability and luminescent properties of the complexes are also investigated.</p></div

Proteome-wide Dysregulation by Glucose-6-phosphate Dehydrogenase (G6PD) Reveals a Novel Protective Role for G6PD in Aflatoxin B₁‑Mediated Cytotoxicity

Glucose-6-phosphate dehydrogenase (G6PD) is pivotal to reduced nicotinamide adenine dinucleotide phosphate (NADPH) production and cellular redox balance. Cells with G6PD deficiency are susceptible to oxidant-induced death at high oxidative stress. However, it remains unclear what precise biological processes are affected by G6PD deficiency due to altered cellular redox homeostasis, particularly at low oxidative stress. To further explore the biological role of G6PD, we generated G6PD-knockdown cell clones using lung cancer line A549. We identified proteins differentially expressed in the knockdown clones without the addition of exogenous oxidant by means of isobaric tags for relative and absolute quantification (iTRAQ) labeling coupled with multidimensional liquid chromatography–mass spectrometry (LC–MS/MS). We validated a panel of proteins that showed altered expression in G6PD-knockdown clones and were involved in metabolism of xenobiotic and glutathione (GSH) as well as energy metabolism. To determine the physiological relevancy of our findings, we investigated the functional consequence of G6PD depletion in cells treated with a prevalent xenobiotic, aflatoxin B₁ (AFB₁). We found a protective role of G6PD in AFB₁-induced cytotoxicity, possibly via providing NADPH for NADPH oxidase to induce epoxide hydrolase 1 (EPHX1), a xenobiotic-metabolizing enzyme. Collectively, our findings reveal for the first time a proteome-wide dysregulation by G6PD depletion under the condition without exogenous oxidant challenge, and we suggest a novel association of G6PD activity with AFB₁-related xenobiotic metabolism

Manganese-Based Layered Coordination Polymer: Synthesis, Structural Characterization, Magnetic Property, and Electrochemical Performance in Lithium-Ion Batteries

Manganese-based layered coordination polymer ([Mn­(tfbdc)­(4,4′-bpy)­(H₂O)₂], Mn–LCP) with microporous structure was synthesized by reaction of 2,3,5,6-tetrafluoroterephthalatic acid­(H₂tfbdc) and 4,4′-bipyridine­(4,4′-bpy) with manganese­(II) acetate tetrahydrate in water solution. Mn–LCP was characterized by elemental analysis, IR spectra, thermogravimetric analysis, X-ray single-crystal structure analysis, and powder X-ray diffraction. Magnetic susceptibility data from 300 to 1.8K show that there is a weak antiferromagnetic exchange between Mn­(II) ions in Mn–LCP. As anode material, the Mn–LCP electrode exhibits an irreversible high capacity in the first discharge process and a reversible lithium storage capacity of up to about 390 mA h/g from the fourth cycle. It might provide a new method for finding new electrode materials in lithium-ion batterie

Synthesis, crystal structures, and luminescent properties of Pb(II) and Sr(II) coordination polymers constructed by 5-methyl-1H-pyrazole-3-carboxylic acid

<div><p>Two new coordination polymers, {[Pb(HMPCA)₂]}_<i>n</i> (<b>1</b>) and {[Sr(HMPCA)₂(H₂O)₂]·H₂O}_<i>n</i> (<b>2</b>) (H₂MPCA = 5-methyl-1H-pyrazole-3-carboxylic acid), have been synthesized and characterized by elemental analysis, IR spectroscopy, X-ray diffraction, and thermogravimetric analysis. Compound <b>1</b> forms a one-dimensional (1-D) chain, and a two-dimensional (2-D) supramolecular framework is constructed by hydrogen bonding. The HMPCA^– in <b>1</b> adopts two coordination modes, a N,O-chelate and a <i>μ</i>₃-<i>κ</i>N, O: <i>κ</i>O, O′: <i>κ</i>O′ mode. The structure of <b>2</b> is a 1-D Sr–O–Sr chain built up by two waters bridging a pair of distrontium [Sr(H₂O)(HMPCA)(<i>μ</i>₂-HMPCA)]₂ units, and the chains are further extended into a 2-D supramolecular layer via hydrogen bonds. In addition to a N,O-chelate, the HMPCA^– in <b>2</b> exhibit a third coordination mode, <i>μ</i>₂-<i>κ</i>N, O: <i>κ</i>O. The thermal decomposition and luminescent properties of <b>1</b> and <b>2</b> in the solid state have also been investigated.</p></div