Effects of mutating Lys65 to Ala and Arg on α_2A-AR-mediated activation of ERK1/2.

<p>(A) HEK293 cells were transfected with wild-type (WT) α_2A-AR or its mutants K65A and K65R and then stimulated with increasing concentrations of UK14,304 for 5 min. ERK1/2 activation was determined by Western blot analysis using phospho-specific ERK1/2 antibodies. <i>Upper panel</i>, a representative blot of ERK1/2 activation; <i>Lower panel</i>, total ERK1/2 expression. (B) Quantitative data expressed as percentage of ERK1/2 activation obtained in cells transfected with α_2A-AR and stimulated with UK14304 at 1 µM and presented as the mean ± S.E. of three separate experiments. *, <i>p</i><0.05 <i>versus</i> WT α_2A-AR at the same concentration of UK14,304.</p

Effects of mutating Leu64 and Lys65 residues on the subcellular distribution of α_2A-AR.

<p>GFP-tagged wild-type (WT) α_2A-AR and its mutants L64A, K65A and LK-AA were transiently expressed in HEK293 (<i>upper panel</i>) and HeLa cells (<i>lower panel</i>) and their subcellular distribution of the receptors was revealed by detecting GFP fluorescence by confocal microscopy. The data shown are representative images of at least three independent experiments. <i>Green</i>, GFP-tagged receptors; <i>blue</i>, DNA staining by DAPI (nuclei). Scale bar, 10 µm.</p

Effects of mutating Lys65 to Arg, Glu and Gln on the cell-surface expression and subcellular distribution of α_2A-AR.

<p>(A) Quantification of the cell surface and total expression of α_2A-AR and its Lys mutants. HEK293 cells were transfected with α_2A-AR and its mutants. The cell-surface expression of the receptors was measured by intact cell binding assays using [³H]-RX821002 and total receptor expression by flow cytometry measuring the GFP signal as described in the legends of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#pone-0050416-g001" target="_blank">figure 1</a>. (B) Quantification of the cell-surface expression of α_2A-AR and its mutants by flow cytometry following staining with anti-HA antibodies in nonpermeabilized cells as described in the legends of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#pone-0050416-g001" target="_blank">figure 1</a>. The data shown in (A) and (B) are percentages of the mean value obtained from cells transfected with wild-type (WT) α_2A-AR and are presented as the mean ± S.E. of four experiments. *, <i>p</i><0.05 <i>versus</i> WT α_2A-AR. (C) Effect of mutation of Lys65 on the subcellular distribution of α_2A-AR. α_2A-AR and its mutants K65R, K65E and K65Q were tagged with GFP at their C-termini and transiently expressed in HEK293 (<i>upper panel</i>) and HeLa cells (<i>lower panel</i>). Their subcellular distribution was revealed by detecting GFP fluorescence by confocal microscopy. The data shown are representative images of at least three independent experiments. <i>Green</i>, GFP-tagged receptors; <i>blue</i>, DNA staining by DAPI (nuclei). Scale bar, 10 µm.</p

Effect of mutation of Lys65 on the colocalization of α_2A-AR with the ER marker DsRed2-ER.

<p>(A) HEK293 cells were transiently transfected with the GFP-tagged α_2A-AR or its Lys65 mutants together with pDsRed2-ER. The subcellular distribution and co-localization of the receptors with the ER marker DsRed2-ER were revealed by confocal fluorescence microscopy as described under “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#s2" target="_blank">Materials and Methods</a>”. <i>Green</i>, α_2A-AR or its mutants tagged with GFP; <i>red</i>, DsRed2-ER; <i>yellow</i>, co-localization of α_2A-AR or its mutants with the ER marker DsRed2-ER; <i>blue</i>, DNA staining by DAPI (nuclei). The data shown are representative images of at least three independent experiments. (B) Quantification of Pearson’s coefficient between the receptors and the ER marker. The data are presented as the mean ± S.E. of 20 cells from three different experiments. *, <i>p</i><0.05 <i>versus</i> WT α_2A-AR. Scale bar, 10 µm.</p

Effects of the mutation of Leu residues and their neighboring positively charged residues in the ICL1 on the cell-surface and total expression of α_2A-AR and α_2B-AR.

<p>(A) The sequence of the ICL1 of α_2A-AR, α_2B-AR and α_2C-AR. (B) Ligand dose-dependent binding of α_2A-AR in intact HEK293 cells. HEK293 cells cultured on 6-well plates were transfected with α_2A-AR and then split onto 24-well plates. The cells were incubated with increasing concentrations of [³H]-RX821002 (0.3125, 0.625, 1.25, 2.5, 5, 10, and 20 nM) and the ligand bound to the receptor was measured by liquid scintillation spectrometry as described in the “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#s2" target="_blank">Materials and Methods</a>”. The nonspecific binding was determined in the presence of rauwolscine (10 µM). Similar results were obtained in at least three different experiments. (C) Quantification of the cell-surface and total expression of α_2A-AR and its mutants in which Leu64 and Lys65 were mutated to Ala individually or in combination. (D) Quantification of the cell-surface and total expression of α_2B-AR and its mutants in which Leu48 and Arg49 were mutated to Ala. In (C) and (D), HEK293 cells were transfected with α₂-AR and the cell-surface expression of the receptors was measured by intact cell binding assays using [³H]-RX821002 at 20 nM. The mean values of specific [³H]-RX821002 binding were 27255±415 and 16785±452 cpm from cells transfected with wild-type (WT) α_2A-AR and α_2B-AR, respectively. Wild-type α_2A-AR and α_2B-AR and their mutants were tagged with GFP and transiently expressed in HEK293 cells. Total receptor expression was determined by flow cytometry measuring the GFP signal as described in the “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#s2" target="_blank">Materials and Methods</a>”. (E) Quantification of the cell-surface expression of α_2A-AR and its mutants by flow cytometry. HEK293 cells were transfected with HA-tagged α_2A-AR or its individual mutant and the cell-surface expression of the receptors was measured by flow cytometry following staining with anti-HA antibodies in nonpermeabilized cells as described in the “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050416#s2" target="_blank">Materials and Methods</a>”. The data shown in (C), (D), and (E) are percentages of the mean value obtained from cells transfected with wild-type α₂-AR and are presented as the mean ± S.E. of at least three different experiments. *, <i>p</i><0.05 <i>versus</i> respective WT α₂-AR.</p

Efficient and Stable Red Emissive Carbon Nanoparticles with a Hollow Sphere Structure for White Light-Emitting Diodes

Red-emissive solid-state carbon nanoparticles (CNPs) with a hollow sphere structure for white light-emitting diodes (WLEDs) were designed and synthesized by molecular self-assembly and microwave pyrolysis. Highly ordered graphite-like structures for CNPs were characterized by transmission electron microscopy, X-ray photoelectron spectroscopy, and ultraviolet–visible (UV–vis) spectroscopy. The emission mechanism of the red-emissive solid-state CNPs was investigated in detail by steady-state and time-resolved photoluminescence (PL) spectroscopy. The as-prepared CNPs showed a red emission band centered at 620 nm with excitation wavelength independence, indicating uniform size of sp² carbon domains in the CNPs. The CNPs also had a PL quantum yield (QY) of 17% under 380 nm excitation. Significantly, the PL QY of the organosilane-functionalized CNPs was 47%, which is the highest value recorded for red-emissive solid-state carbon-based materials under UV-light excitation. More importantly, the red-emissive CNPs exhibited a PL QY of 25% after storage in air for 12 months, indicating their excellent stability. The red-emissive CNP powders were used as environmentally friendly and low-cost phosphors on a commercial 460 nm blue GaN-based chip, and a pure white light with CIE coordinates of (0.35, 0.36) was achieved. The experimental results indicated that the red-emissive CNP phosphors have potential applications in WLEDs

Interfacial Emission Adjustment in ZnO Quantum Dots/p-GaN Heterojunction Light-Emitting Diodes

Ultraviolet (UV) light-emitting diodes (LEDs) were made by using ZnO quantum dots (QDs) as the emission layer. ZnO QDs with the diameter of 5 nm were fabricated by using a simple sintering method. By using p-GaN as the hole injection layer, a ZnO QDs/p-GaN heterojunction LED was constructed. Trap-controlled SCLC behavior of QDs led the LED to emit light mainly from the QDs layer, and the direct physical contact between ZnO QDs and GaN could effectively reduce the interfacial emission. As the result, a UV LED with the electroluminescence wavelength of 382 nm has been achieved

Bifunctional MoO₃–WO₃/Ag/MoO₃–WO₃ Films for Efficient ITO–Free Electrochromic Devices

Dielectric–metal–dielectric (DMD) trilayer films, served as both electrochromic (EC) film and transparent conductor (TC), have exhibited great potential application in low–cost, ITO–free electrochromic devices (ECDs). However, recent reports on the DMD–based ECDs revealed that the response time and the optical modulation properties were not very satisfactory. Here, the mixed MoO₃–WO₃ materials were first introduced as the dielectric layer to construct an EC–TC bifunctional MoO₃–WO₃/Ag/MoO₃–WO₃ (MWAMW) film, which demonstrates strong and broad–band optical modulation in the visible light region, fast color-switching time (2.7 s for coloration and 4.1 s for bleaching), along with high coloration efficiency (70 cm² C^–1). The electrical structure and electrochemical reaction kinetics analysis revealed that the improved EC performances are associated with the increased electron intervalence transition together with the fast charge–transfer and ion–diffusion dynamics

Comparison of the SWI phase values and R2* values in cirrhotic patients and healthy subjects.

<p>The SWI phase values (in radians) in high-iron areas of the 37 healthy subjects were significantly higher than those of the 87 cirrhotic patients (−0.161±0.010 vs. −0.266±0.155, respectively; <i>P</i><0.001) (a). The R2* values (Hz) of the 37 healthy subjects were significantly lower than those of the 87 cirrhotic patients (67.02±12.32 vs. 86.30±34.48, respectively; <i>P = </i>0.004) (b). The error bar represents the 95% confidence interval. NC: normal control; HC: hepatic cirrhosis patients.</p

Correlation between the SWI phase values and R2* values in vitro.

<p>In cases of MnCl₂ concentrations lower than 2.3 mM, the SWI phase values correlated linearly with the R2* values (r = −0.996, <i>P</i><0.001).</p