Journal: bioRxiv

Article Title: A porcine ex vivo lung perfusion model to investigate bacterial pathogenesis

doi: 10.1101/708503

Figure Lengend Snippet: A. CD163 surface expression in PBS or Kp52145-infected pBMDMs by flow cytometry. Values are shown as standard error of mean of two independent experiments in duplicate. **, p<0.01 determined by unpaired Student’s -test. B. Immunoblotting analysis of phosphorylation of STAT6 (PSTAT6) and tubulin in lysates of pBMDMs infected with Kp52145 for the indicated times or left uninfected (ni). Data is representative of three independent experiments. C. cd163 levels in pBMDMs non-infected (ni) or infected with Kp52145 pre-treated with STAT6 inhibitor (AS1517499, 50nM 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments. D. Arginase-1 levels in pBMDMs non-infected (ni) or infected with Kp52145 pre-treated with STAT6 inhibitor (AS 1517499, 50nM 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments. E. Immunoblotting analysis of phosphorylation of STAT6 (PSTAT6) and tubulin in lysates of pBMDMs infected with Kp52145 and the isogenic cps mutant, strain 52145-Δ wca K2 for the indicated times or left uninfected (ni). Data is representative of three independent experiments. F. cd163 levels in pBMDMs non-infected (ni) or infected with the cps mutant, strain 52145-Δ wca K2 , pre-treated with STAT6 inhibitor (AS1517499, 50nM 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments in duplicate. G. arginase-1 levels in pBMDMs non-infected (ni) or infected with cps mutant, strain 52145-Δ wca K2 , pre-treated with STAT6 inhibitor (AS 1517499, 50nM 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments in duplicate. In panels C, and D, ***p<0.001, **p<0.01, for the indicated comparisons using one-way ANOVA with Bonferroni correction.

Article Snippet: Primary antibodies included: phospho-STAT6 (Tyr641) (1:2000, #9361), total STAT6 (1:1000, BioRad #170-6516), phospho-STAT3 (Y705) (1:2000, #9145), total STAT3 (1:2000, #12640), phospho-ERK (p44/42) (1:2000; #91015), phopsho-p38 (T180/Y182) (1:2000, #4511), all from Cell Signalling Technologies.

Techniques: Expressing, Infection, Flow Cytometry, Western Blot, Mutagenesis

Journal: bioRxiv

Article Title: A porcine ex vivo lung perfusion model to investigate bacterial pathogenesis

doi: 10.1101/708503

Figure Lengend Snippet: A. il-10 levels in pBMDMs non-infected (ni) or infected with Kp52145 pre-treated with STAT6 inhibitor (AS 1517499, 50nM/ 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments. B. Immunoblotting analysis of phosphorylation of STAT3 (PSTAT3) and STAT3 in lysates of pBMDMs infected with Kp52145 for the indicated times or left uninfected (ni). Data is representative of three independent experiments. C. Immunoblotting analysis of phosphorylations of ERK (pERK), p38 (Pp38), and tubulin in lysates of pBMDMs infected with Kp52145 for the indicated times or left uninfected (ni). Data is representative of three independent experiments. D. il-10 levels in pBMDMs non-infected (ni) or infected with Kp52145 pre-treated with p38 inhibitor (SB203580, Tocris, 10 μg/mL, 2 h prior to infection), ERK inhibitor (U0126, LC laboratories, 20 μg/mL, 2 h prior to infection) or DMSO vehicle control. Values are shown as standard error of mean of three independent experiments. In panels A and D, ****p < 0.0001, ***p < 0.001, for the indicated comparisons using one-way ANOVA with Bonferroni correction.

Article Snippet: Primary antibodies included: phospho-STAT6 (Tyr641) (1:2000, #9361), total STAT6 (1:1000, BioRad #170-6516), phospho-STAT3 (Y705) (1:2000, #9145), total STAT3 (1:2000, #12640), phospho-ERK (p44/42) (1:2000; #91015), phopsho-p38 (T180/Y182) (1:2000, #4511), all from Cell Signalling Technologies.

Techniques: Infection, Western Blot