Transport of Asian ozone pollution into surface air over the western United States in spring

Many prior studies clearly document episodic Asian pollution in the western U.S. free troposphere. Here, we examine the mechanisms involved in the transport of Asian pollution plumes into western U.S. surface air through an integrated analysis of in situ and satellite measurements in May–June 2010 with a new global high-resolution (∼50 × 50 km2) chemistry-climate model (GFDL AM3). We find that AM3 with full stratosphere-troposphere chemistry nudged to reanalysis winds successfully reproduces observed sharp ozone gradients above California, including the interleaving and mixing of Asian pollution and stratospheric air associated with complex interactions of midlatitude cyclone air streams. Asian pollution descends isentropically behind cold fronts; at ∼800 hPa a maximum enhancement to ozone occurs over the southwestern U.S., including the densely populated Los Angeles Basin. During strong episodes, Asian emissions can contribute 8–15 ppbv ozone in the model on days when observed daily maximum 8-h average ozone (MDA8 O3) exceeds 60 ppbv. We find that in the absence of Asian anthropogenic emissions, 20% of MDA8 O3 exceedances of 60 ppbv in the model would not have occurred in the southwestern USA. For a 75 ppbv threshold, that statistic increases to 53%. Our analysis indicates the potential for Asian emissions to contribute to high-O3 episodes over the high-elevation western USA, with implications for attaining more stringent ozone standards in this region. We further demonstrate a proof-of-concept approach using satellite CO column measurements as a qualitative early warning indicator to forecast Asian ozone pollution events in the western U.S. with lead times of 1–3 days

A yeast-based chemical screen identifies a PDE inhibitor that elevates steroidogenesis in mouse Leydig cells via PDE8 and PDE4 inhibition.

A cell-based high-throughput screen (HTS) was developed to detect phosphodiesterase 8 (PDE8) and PDE4/8 combination inhibitors. By replacing the Schizosaccharomyces pombe PDE gene with the murine PDE8A1 gene in strains lacking adenylyl cyclase, we generated strains whose protein kinase A (PKA)-stimulated growth in 5-fluoro orotic acid (5FOA) medium reflects PDE8 activity. From our previously-identified PDE4 and PDE7 inhibitors, we identified a PDE4/8 inhibitor that allowed us to optimize screening conditions. Of 222,711 compounds screened, ∼0.2% displayed composite Z scores of >20. Additional yeast-based assays using the most effective 367 compounds identified 30 candidates for further characterization. Among these, compound BC8-15 displayed the lowest IC₅₀ value for both PDE4 and PDE8 inhibition in in vitro enzyme assays. This compound also displays significant activity against PDE10A and PDE11A. BC8-15 elevates steroidogenesis in mouse Leydig cells as a single pharmacological agent. Assays using BC8-15 and two structural derivatives support a model in which PDE8 is a primary regulator of testosterone production by Leydig cells, with an additional role for PDE4 in this process. BC8-15, BC8-15A, and BC8-15C, which are commercially available compounds, display distinct patterns of activity against PDE4, PDE8, PDE10A, and PDE11A, representing a chemical toolkit that could be used to examine the biological roles of these enzymes in cell culture systems

Development of a PDE8A inhibitor HTS.

<p><b>A)</b> Structure of PDE4/8 inhibitor BC69 used to optimize PDE8 inhibitor assay. <b>B)</b> A yeast-based screen finds no potent PDE8 inhibitors in a Known Bioactives Collection. PDE8A-expressing strain CHP1204 was screened against 2,640 known bioactive compounds in duplicate wells (plate A and B). Scatter plot of absorbance values from plate A against plate B is shown. Negative control wells contained 0.2% DMSO (black circles), positive control wells contained 10 or 20 µM BC69 (dark gray circles). Experimental wells are shown in light gray. The data set has a correlation value of 0.996.</p

Strain list.

<p>Strain list.</p

BC8-15 derivative elevation of progesterone release by MA-10 Leydig tumor cells corresponds with PDE8 inhibitory activity.

<p>MA-10 cells were treated with 20 µM of rolipram, BC8-15, BC8-15A or BC8-15C. Progesterone released into the media after 2 h was measured using a progesterone ELISA kit (Neogen). The data are representative of three independent experiments with 2–4 sample wells for each condition. Values are the mean of four experimental wells ± SEM. Data were analyzed with one-way ANOVA followed by a Tukey’s multiple comparison test (*** p<0.001). The response to BC8-15 is significantly different from that for BC8-15A (p<0.001) and both are significantly different from all other groups (p<0.001).</p

BC8-15 elevates testosterone release by primary Leydig cells.

<p>Leydig cells isolated from wild-type (Panel A) and PDE8A^−/−/B^−/− knock-out (Panel B) mice were treated with rolipram (20 µM) or BC8-15 (10 µM and 40 µM) for three hours. Testosterone released into the media was assayed in duplicate for each condition. Values represent the mean ± SEM. Data from wild type and knock-out samples were separately analyzed with one-way ANOVA followed by Tukey’s multiple comparison test. Significant differences are indicated in comparison to the DMSO control (* p<0.05, ** p<0.01, *** p<0.001).</p

Summary of the HTS data.

<p>The ability of 220,071 small molecules to promote 5FOA^R growth of a PDE8A-expressing strain was assessed by their composite Z scores (see Materials and Methods). The number of compounds within each indicated composite Z score interval is presented.</p

Inhibition of PDE4A, PDE7A and PDE8A by BC8-15 as determined by <i>in vitro</i> enzyme assays.

<p>BC8-15 inhibitory activity was measured by <i>in vitro</i> PDE assays as described in Materials and Methods. Substrate concentrations are 10 nM cAMP for PDE8A and PDE7A and 50 nM cAMP for PDE4A. Each assay was performed at 10 different compound concentrations in duplicate reaction tubes. IC₅₀± SD values are determined by performing non-linear regression analysis on three independent experiments.</p

IC₅₀ values for BC8-15 and related compounds.

<p>The structures of BC8-15 and two structurally-related derivatives are shown. The IC₅₀ values of each compound were determined by <i>in vitro</i> PDE assays (see Material and Methods). The values represent mean IC₅₀± SD determined from three independent experiments.</p