Transwell migration assay showed that PDGF-BB-induced ARPE19 cell migration was inhibited by resveratrol.

<p>Transwell inserts were coated with fibronectin (0.3 mg). ARPE19 cells (5×10⁴ in 200 µl) were seeded in the upper chamber in the absence or presence of resveratrol. The inserts were assembled in the lower chamber, which was filled with 600 µl serum-free medium without PDGF-BB (A) and containing PDGF-BB (20 ng/ml) (B), and preincubated with various concentrations of resveratrol for 30 mininutes at 37°C. After incubating for 5 hours at 37°C, fixation was performed. ARPE19 cells that migrated to the underside of filter membrane were photographed (A, B) and counted by phase contrast light microscope under high power field (magnification, 100×), (C). All experiments were conducted in duplicates and similar results were repeated four times. The results are expressed as percentage of control and represent mean ± standard errors (SE) of the eight experiments. *p<0.05 significantly differs from PDGF-BB-stimulated cells (the fourth bar).</p

Hollow Interior Structure of Spin-Coated Polymer Thin Films Revealed by ToF-SIMS Three-Dimensional Imaging

Surface patterns were observed on spin-coated poly­(bisphenol A decane ether) (BA-C10) films prepared with chloroform and tetrahydrofuran as the solvents. The interior structure of these surface patterns were analyzed using a time-of-flight secondary ion mass spectrometry (ToF-SIMS) equipped with a bismuth cluster source for ion imaging and a C₆₀⁺ cluster source for depth profiling. For the first time, the surface patterns have been shown to be hollow rather than solid using ToF-SIMS three-dimensional (3D) analysis and optical techniques. Moreover, the microarea depth profiling analysis indicated that the hollow structure was sandwiched between two polymer layers rather than sitting on the substrate. The height of the hollow structure and the thicknesses of the polymer layers above and below the hollow structure were also estimated from the depth profiling results

Hollow Interior Structure of Spin-Coated Polymer Thin Films Revealed by ToF-SIMS Three-Dimensional Imaging

Surface patterns were observed on spin-coated poly­(bisphenol A decane ether) (BA-C10) films prepared with chloroform and tetrahydrofuran as the solvents. The interior structure of these surface patterns were analyzed using a time-of-flight secondary ion mass spectrometry (ToF-SIMS) equipped with a bismuth cluster source for ion imaging and a C₆₀⁺ cluster source for depth profiling. For the first time, the surface patterns have been shown to be hollow rather than solid using ToF-SIMS three-dimensional (3D) analysis and optical techniques. Moreover, the microarea depth profiling analysis indicated that the hollow structure was sandwiched between two polymer layers rather than sitting on the substrate. The height of the hollow structure and the thicknesses of the polymer layers above and below the hollow structure were also estimated from the depth profiling results

PDGF-BB-induced PI3K and Akt phosphorylations were inhibited by resveratrol in a time- and concentration-dependant manner.

<p>ARPE19 cells were treated with the indicated lengths of time of PDGF-BB (20 ng/ml) and preincubated with or without resveratrol (10 µM) at 37°C (A). After being further preincubated for the indicated concentrations of resveratrol and incubated with or without PDGF-BB (20 ng/ml) at 37°C for 30 minutes, the cells were collected and their lysates were analyzed by Western blot analysis (B). The changes in phosphorylated PI3K and Akt expression were evaluated. The quantitative data of western blot are shown below the panels which are expressed as percentage of control and represent mean ± standard errors (SE) of the four independent experiments. *p<0.05 significantly differs from same indicated time of cells stimulated PDGF-BB only (A) and *p<0.05 significantly differs from PDGF-BB-stimulated cells (the fifth bar) (B).</p

Viability and cell adhesion of ARPE19 cells was not influenced by resveratrol.

<p>The cells were treated with different concentrations of resveratrol for 24 hours after being starved for 24 hours. Cell viability was determined by MTT assay (A). BCECF-labeled cells were treated with DMSO or resveratrol for 30 minutes. They were then seeded and allowed to adhere on plates with precoated fibronectin (fn) (15 µg/ml) at 37°C for 1 hour. Fluorescence was measured using excitation and emission wavelength of 485 and 535 nm, respectively (B). The results are expressed as percentage of control and represent the mean ± standard errors (SE) of four independent experiments.</p

PDGF-BB-induced ARPE19 cell migration was inhibited by resveratrol in an ECIS migration assay.

<p>The cells cultured in 8W1E ECIS arrays were treated with different combinations of PDGF-BB (20 ng/ml) and resveratrol (CTL indicates that it contained only DMSO) which were preincubated together at 37°C for 30 minutes. Cell migration was then assessed by continuous resistance measurements for 30 hours. Resveratrol (Res) (10 µM) did not increased cell migration when PDGF-BB was not present. In the well containing PDGF-BB but not resveratrol, the impedance, which corresponds to the number of cells migrated to the surface of the detective electrode, increased sharply during the first 10 hours. By contrast, the impedance in the well containing PDGF-BB and resveratrol increased slowly during the same time period.</p

PDGF-BB-induced ERK and P38 phosphorylations were inhibited by resveratrol in a time- and concentration-dependant manner.

<p>ARPE19 cells were treated with the indicated lengths of time of PDGF-BB (20 ng/ml) and preincubated with or without resveratrol (10 µM) at 37°C (A). After being further preincubated for the indicated concentrations of resveratrol and incubated with or without PDGF-BB (20 ng/ml) at 37°C for 30 minutes, the cells were collected and their lysates were analyzed by Western blot analysis (B). The changes in phosphorylated ERK, JNK and p38 expression were evaluated. The quantitative data of western blot are shown below the panels which are expressed as percentage of control and represent mean ± standard errors (SE) of the four independent experiments. *p<0.05 significantly differs from same indicated time of cells stimulated PDGF-BB only (A) and *p<0.05 significantly differs from PDGF-BB-stimulated cells (the fifth bar) (B).</p

PDGF-BB-induced cell migrations were inhibited by resveratrol and by suppression of PDGFR, PI3K/Akt and MAPK signaling.

<p>The plates with confluent monolayer of ARPE cells were pretreated with mitomycin-C (5 µg/ml) for 1 hour, then wounded with a linear scratching by a sterile 20- µl pipette tip. The cells were immediately washed and were incubated with PDGF-BB (20 ng/ml) only, PDGF-BB in combination with resveratrol (10 µM), AG1295 (10 µM), LY294002 (10 µM), U0126 (10 µM), SP600125 (3 µM) and SB203580 (3 µM) respectively. The wound closure was monitored for 16 h followed by photography under phase-contrast microscope (x100) (A). The quantitative data of the number of migrated cell in the wound area are expressed as percentage of control and represent mean ± standard errors (SE) of the four independent experiments. *p<0.05 significantly differs from PDGF-BB-stimulated cells (the second bar) (B).</p

Resveratrol did not directly interact with PDGF-BB in dot binding assay.

<p>Human recombinant PDGF-BB, phosphate buffer saline (PBS) and the indicated concentrations of resveratrol were applied onto the nitrocellulose (NC) membrane. The membrane was incubated with PDGF-BB in PBS and then developed by probing with Ab directed against PDGF-BB. The results presented are representative of four independent experiments.</p

Evidence of Enhanced Mobility at the Free Surface of Supported Polymer Films by in Situ Variable-Temperature Time-of-Flight-Secondary Ion Mass Spectrometry

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) spectra of polystyrene (PS) films supported on silicon wafers were obtained at temperatures ranging from room temperature to 100 °C. Principal component analysis (PCA) of the TOF-SIMS data revealed a transition temperature (<i>T</i>_T) at which the surface structure of PS was rearranged. The <i>T</i>_T of a 120-nm thick PS (weight-average molecular weight of 3 000 g/mol) thin film was determined to be about 36 °C, which is approximately 30 °C lower than the bulk glass transition temperature (<i>T</i>_g) of that PS. Similar <i>T</i>_Ts were observed on PSs with different molecular weights. As the <i>T</i>_T is strongly related to the <i>T</i>_g and dependent on the molecular weight, it is believed that the <i>T</i>_T determined by TOF-SIMS is related to the surface glass transition temperature (<i>T</i>_g^S) measured by other techniques. This suggests that TOF-SIMS combined with PCA can be used to determine the <i>T</i>_g^S of polymer films. Furthermore, the detailed PCA analyses indicate that the phenyl groups of PS tended to move away from the surface at temperatures above <i>T</i>_T. This conclusion was further confirmed by contact angle and XPS measurements