Ex vivo analyses of K17-dependent neutrophil recruitment.

<p><b>(A)</b> Parental and <i>KRT17</i> null A431 keratinocytes stained for K17 and nuclei (DAPI). Scale bars: 20 μm. <b>(B)</b> Migration of human primary neutrophils towards A431 conditioned medium (CM) “titrated” (i.e., diluted) as indicated using control medium (<i>n</i> = 2). <b>(C)</b> Migration of human primary neutrophils towards CM from GFP-transfected parental A431, GFP-transfected <i>KRT17</i> null A431, and GFP-K17-transfected <i>KRT17</i> null A431 cells. Individual symbols depict data using neutrophils from different donors (<i>n</i> = 6). Data reported as mean ± SEM. One-way ANOVA. <b>(D)</b> Transcript levels of CXCR3 in human primary neutrophils before (t = 0) and after treatment with buffer (mHBSS, 2 h), culture medium control (2 h), CM from acetone-Tx parental A431 (2 h), or CM from TPA-Tx parental A431 (2 h). <i>n</i> = 3. Data reported as mean ± SEM. One-way ANOVA. <b>(E, F)</b> ELISA measurements for selected chemokine and cytokine levels in A431 CM (reported as pg/ml). <i>n</i> = 3 for CXCL2 (2 technical replicates each), and <i>n</i> = 4 for CXCL9 (4 technical replicates for 2 measurements, 2 technical replicates for the other 2 measurements). Data reported as mean ± SEM. One-way ANOVA. <b>(G)</b> ELISA measurements of TNFα levels in A431 CM (pg/ml). <i>n</i> = 2 measurements (4 technical replicates each). Data are shown as mean ± SEM. One-way ANOVA. Acet, Acetone. <b>(H)</b> Migration of human primary neutrophils towards DMSO (vehicle control) or fMLF, with or without the addition of a CXCR2 antagonist, a CXCR3 antagonist, and infliximab (anti-TNFα). <i>n</i> = 5. Data reported as mean ± SEM. Paired <i>t</i> test. The source data used to derive the numerical values reported here can be found in <b><a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.3002779#pbio.3002779.s016" target="_blank">S1 Data</a></b>.</p> <p>(PDF)</p>