ZnO/CuO Heterojunction Branched Nanowires for Photoelectrochemical Hydrogen Generation

We report a facile and large-scale fabrication of three-dimensional (3D) ZnO/CuO heterojunction branched nanowires (b-NWs) and their application as photocathodes for photoelectrochemical (PEC) solar hydrogen production in a neutral medium. Using simple, cost-effective thermal oxidation and hydrothermal growth methods, ZnO/CuO b-NWs are grown on copper film or mesh substrates with various ZnO and CuO NWs sizes and densities. The ZnO/CuO b-NWs are characterized in detail using high-resolution scanning and transmission electron microscopies exhibiting single-crystalline defect-free b-NWs with smooth and clean surfaces. The correlation between electrode currents and different NWs sizes and densities are studied in which b-NWs with longer and denser CuO NW cores show higher photocathodic current due to enhanced reaction surface area. The ZnO/CuO b-NW photoelectrodes exhibit broadband photoresponse from UV to near IR region, and higher photocathodic current than the ZnO-coated CuO (core/shell) NWs due to improved surface area and enhanced gas evolution. Significant improvement in the photocathodic current is observed when ZnO/CuO b-NWs are grown on copper mesh compared to copper film. The achieved results offer very useful guidelines in designing b-NWs mesh photoelectrodes for high-efficiency, low-cost, and flexible PEC cells using cheap, earth-abundant materials for clean solar hydrogen generation at large scales

Arsenic induced angiogenesis.

<p>(A) A549 cells were treated without or with 5 µM of arsenic (As) for 5 h, then trypsinized, resuspended in serum-free medium (3×10⁷cells/ml, 0.1 ml), and mixed in 1∶1 ratio with Matrigel (Collaborative Biomedical Products, Bedford, MA). Aliquots of the mixture were then implanted onto the CAM of 9-day-old embryos. After 96 h incubation, the area around the implanted Matrigel was photographed with a Nikon digital camera. Bar: 2 mm (upper panel). The number of blood vessels was obtained by counting the branching of blood vessels, and the relative angiogenesis was obtained by normalizing to that of the control without arsenic treatment. The data represent the mean ± SD of the relative angiogenesis from eight different embryos (bottom panel). *, indicates that the relative angiogenesis index significantly increased in arsenic treatment group when compared with control group, <i>P</i><0.05. (B) BEAS-2B cells were treated with or without arsenic to perform tumor angiogenesis assay as above. Bar: 2 mm. *, indicates that the relative angiogenesis index significantly increased in arsenic treatment group when compared with control group, <i>P</i><0.05.</p

Arsenic treatment induced phospho-AKT and phospho-ERK1/2 activation, and increased HIF-1α and VEGF expression.

<p>(A) A549 and BEAS-2B cells were treated with different doses of arsenic (As) for 6 h, total proteins are subjected to Western blotting for HIF-1α and HIF-1β expression. A549 and BEAS-2B cells were cultured in serum-free medium for 24 h, then treated with different doses of arsenic for 2 h. Total proteins were subjected to Western blotting analysis for the levels of phospho-AKT, total AKT, phospho-ERK1/2, and ERK2 expression (upper panel). Relative densities of p-AKT, p-ERK1/2 and HIF-1α were analyzed by the ratio of p-AKT/AKT, p-ERK1/2/ERK2 and HIF-1α/HIF-1β using ImageJ software and normalized to those of control cells. The data represents the mean± SD from duplicate experiments (bottom panel). *, indicates significant increase when compared with the control cells, <i>P</i><0.05. (B) BEAS-2B cells were seeded in 12-well plate. Cells were co-transfected with VEGF reporter and β-galactosidase (β-gal) plasmids and cultured for 15 h. Arsenic at 0, 2.5, and 5 µM was added for 24 h. Luciferase assay was performed by using luciferase assay system. The activity of β-gal was used as internal control of transfection efficiency. The relative luciferase activity was calculated as the ratio of luciferase/β-gal activity, and normalized to the control group. *, indicates that the relative luc activity significantly increased in arsenic treatment group when compared with the control group, <i>P</i><0.05. <i>C,</i> BEAS-2B cells were treated without or with 5 µM of arsenic for 24 h. Total RNAs were extracted by Trizol and subjected to RT-PCR analysis of VEGF and GAPDH expression.</p

Arsenic induced ROS production in BEAS-2B cells, which was required for angiogenesis.

<p>(A) BEAS-2B cells were seeded into 6-well plates. Cells were treated with different doses of arsenic as indicated in serum-free medium. DCFH-DA at 5 µM was added to the cells for 15 min. Then the cells were washed and fixed, and the fluorescent images were captured using a fluorescent microscope (upper panel). The corresponding phase micrographs were shown in the bottom panel. (B) BEAS-2B cells were seeded into the 6-well plate. The cells were then cultured in serum-free medium with arsenic at 5 µM for different time points as indicated. DCFH-DA staining was performed as above. (C) BEAS-2B cells were infected with adenovirus carrying GFP (Ad-GFP) and catalase (Ad-catalase), respectively at 20 MOI. After 24 h, cells were treated with 5 µM arsenic for 5 h to perform angiogenesis assay. *, indicates that the relative angiogenesis index was significantly decreased when compared with Ad-GFP control group, <i>P</i><0.05.</p

AKT and ERK1/2 pathways are required for arsenic-inducing HIF-1α expression and angiogenesis.

<p>(A) BEAS-2B cells were cultured in serum-free medium for 24 h, then cells were pre-treated with LY294002 or U0126 at 20 µM for 30 min. Arsenic at 5 µM was added to the cells for 2 h (for p-AKT, AKT, p-ERK1/2, and ERK2 expression) and 6 h (for HIF-1α and HIF-1β expression), respectively. Total proteins were analyzed by Western blotting to detect the expression of proteins as indicated (left panel). Relative densities of p-AKT, p-ERK1/2 and HIF-1α were analyzed as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0020858#pone-0020858-g002" target="_blank">Fig. 2A</a> (right panel). *, indicates significant increase when compared with the control cells, <i>P</i><0.05. . #, indicates significant decrease when compared with the sodium arsenite treatment, <i>P</i><0.05. (B) BEAS-2B cells were treated with 5 µM of arsenic. Cells were trypsinized, mixed with equal volume of Matrigel with or without 15 µM of LY294002 or U0126. Equal volume of DMSO was added as a negative control. Angiogenesis assay was performed as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0020858#pone-0020858-g001" target="_blank">Fig. 1</a>. *, indicates that the relative angiogenesis index was significantly decreased when compared with DMSO control group, <i>P</i><0.05. (C) BEAS-2B cells were infected with adenovirus carrying GFP (Ad-GFP) or AKT dominant negative (Ad-AKT-DN) at 20 MOI (upper panel), or transfected with scrambled control of siRNA (siScramble) or siMAPK at 50 nM (bottom panel). After 24 h, cells were treated with arsenic and angiogenesis assay was performed as above. *, indicates that the relative angiogenesis index was significantly decreased when compared with Ad-GFP or siScramble control group, <i>P</i><0.05.</p

VEGF is required for arsenic-inducing angiogenesis.

<p>(A) BEAS-2B cells were treated with DPI or catalase for 30 min, then with 5 µM arsenic for 24 h. Total RNAs were extracted by Trizol, and analyzed by RT-PCR for VEGF and GAPDH expression (upper panel). Relative density of VEGF was analyzed by the ratio of VEGF/GAPDH using ImageJ software and normalized to control cells. The data represents the mean± SD from duplicate experiments (bottom panel). *, indicates significant increase when compared with the control cells, <i>P</i><0.05. . #, indicates significant decrease when compared with the sodium arsenite treatment alone, <i>P</i><0.05. (B) BEAS-2B cells were transfected with VEGF siRNA and scrambled siRNA (siVEGF and siScramble, respectively). After the transfection for 24 h, cells were treated with 5 µM arsenic for 5 h to perform angiogenesis assay. *, indicates that the relative angiogenesis index was significantly decreased in siVEGF treatment group when compared with siScramble group, <i>P</i><0.05.</p

ROS are required for AKT and ERK1/2 activation, HIF-1α expression, and angiogenesis.

<p>(A) BEAS-2B cells were cultured in serum-free medium for 24 h, then cells were pre-treated with DPI or catalase for 30 min. Arsenic at 5 µM was added to the cells for 2 h or 6 h as above. The specific proteins are analyzed by Western blotting (left panel). The relative densities of p-AKT, p-ERK1/2 and HIF-1α were determined as above. *, indicates significant increase when compared with the control cells, <i>P</i><0.05. . #, indicates significant decrease when compared with the sodium arsenite treatment alone, <i>P</i><0.05. (B) BEAS-2B cells were infected with adenovirus carrying GFP and HIF-1α siRNA (Ad-GFP and Ad-siHIF-1α, respectively) at 20 MOI for 24 h, then the cells were treated with 5 µM arsenic for 5 h and angiogenesis assay was performed as above. *, indicates that the relative angiogenesis index was significantly decreased when compared with the control group, <i>P</i><0.05.</p

3D Branched Nanowire Photoelectrochemical Electrodes for Efficient Solar Water Splitting

We report the systematic study of 3D ZnO/Si branched nanowire (b-NW) photoelectrodes and their application in solar water splitting. We focus our study on the correlation between the electrode design and structures (including Si NW doping, dimension of the trunk Si and branch ZnO NWs, and b-NW pitch size) and their photoelectrochemical (PEC) performances (efficiency and stability) under neutral conditions. Specifically, we show that for b-NW electrodes with lightly doped p-Si NW core, larger ZnO NW branches and longer Si NW cores give a higher <i>photocathodic</i> current, while for b-NWs with heavily doped p-Si NW trunks smaller ZnO NWs and shorter Si NWs provide a higher <i>photoanodic</i> current. Interestingly, the photocurrent turn-on potential decreases with longer p-Si NW trunks and larger ZnO NW branches resulting in a significant photocathodic turn-on potential shift of ∼600 mV for the optimized ZnO/p-Si b-NWs compared to that of the bare p-Si NWs. A photocathode energy conversion efficiency of greater than 2% at −1 V <i>versus</i> Pt counter electrode and in neutral solution is achieved for the optimized ZnO/p-Si b-NW electrodes. The PEC performances or incident photon-to-current efficiency are further improved using Si NW cores with smaller pitch size. The photoelectrode stability is dramatically improved by coating a thin TiO₂ protection layer using atomic-layer deposition method. These results provide very useful guidelines in designing photoelectrodes for selective solar water oxidation/reduction and overall spontaneous solar fuel generation using low cost earth-abundant materials for practical clean solar fuel production

Overexpression of HER2 and HER3 reverses miR-199a- and miR-125b-inhibited tumor angiogenesis.

<p>(A) Immunoblotting to confirm establishment of an A2780 cell line stably overexpressing HRE2 and OVCAR-3 cell line stably overexpressing HER3 using pBABEpuro and pReceiver-Lv105 vector, respectively. (B) Tube formation assay using HUVEC cells was described as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056647#pone-0056647-g002" target="_blank">Figure 2</a>. (C) Total tube lengths for each treatment were analyzed and presented as mean ± SE (millimeter) from six replicates for each treatment. *Significantly different vs vector+miR-control (P<0.05). (D, E) A2780 and OVCAR cells were transfected pre-miR-control, pre-miR-199a or pre-miR-125b, respectively; and implanted onto the CAMs to perform angiogenesis assay as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056647#pone-0056647-g002" target="_blank">Figure 2</a>. The representative images from each group were shown here. The total number of blood vessels in each group was quantified. *,**Significantly different compared with that of the same cell line transfected with pre-miR-control with *P<0.05 and **P<0.01.</p

miR-21 induced the activation of AKT and ERK1/2, and the expression of HIF-1α and VEGF.

<p>(A) DU145 cells were seeded into 60 mm dishes. When cells were 60% confluence, cells were transfected with pre-miRNA negative control or pre-miR-21 as described above. Total proteins were extracted 36 h after the transfection, and analyzed for the expression of phospho-AKT (p-AKT), total AKT, phospho-ERK1/2 (p-ERK), and ERK2 by immunoblotting. (B) DU145 cells were transfected with pre-miR-21 precursor as described above. After the transfection for 36 h, the cells were treated with solvent alone, 10 µM of LY294002, or 10 µM of U0126 for 6 h. The expression levels of p-AKT, total AKT, p-ERK, ERK2, HIF-1α and HIF-1β were analyzed by immunoblotting. (C-D) After the transfection of cells with pre-miR-21, the cells were cultured for 36 h, then treated with solvent alone, 10 µM of LY294002, or 10 µM of U0126 for 12 h. VEGF mRNA expression was analyzed by RT-PCR (C), and by real-time RT-PCR (D). ##, indicates the significant decrease when compared to that of the control (<i>p</i><0.01).</p