Immunomodulatory functions of FABP5 in primary NHBE cells.

<p>FABP5 exerts host defense and anti-inflammatory functions against <i>P. aeruginosa</i> bacterial infection indirectly by stimulating PPAR-γ activity. PPAR-γ activity increases β defensin-2 expression thus preventing bacterial growth and inhibits inflammatory cytokine (e.g., IL-8) production.</p

CS exposure impairs <i>P. aeruginosa</i> clearance.

<p><b>A.</b> CS exposure increases <i>P. aeruginosa</i> colony forming units (CFU) on primary NHBE cells. <b>B.</b> CS exposure significantly enhances IL-8 secretion in the basolateral supernatant of primary NHBE cell cultures. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

FABP5 down regulation increases inflammation but decreases innate immunity in primary NHBE cells.

<p><b>A.</b> CS exposure increases <i>P. aeruginosa</i> colony forming units (CFU) on FABP5 knocked down primary NHBE cells. <b>B.</b> CS exposure significantly enhances IL-8 secretion in the basolateral supernatant of primary NHBE cell cultures knocked down for FABP5. <b>C.</b> CS exposure decreases β-defensin 2 mRNA expression on FABP5 knocked down primary NHBE cells. FABP5 shRNA − indicates cells that were knocked down for Firefly luciferase and FABP5 shRNA + indicates cells that were knocked down for FABP5. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

CS exposure prevents bacteria-induced FABP5 expression.

<p><b>A.</b> FABP5 mRNA expression is increased post <i>P. aeruginosa</i> infection, however CS exposure decreases FABP5 mRNA expression and prevents bacteria-induced FABP5 mRNA expression. <b>B.</b> Representative Western blot detection of FABP5 and β-actin proteins shows that FABP5 protein levels increased post Pa infection but CS exposure decreases Pa-induced FABP5 protein levels. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

FABP5 expression is decreased in COPD airway epithelial cells.

<p><b>A.</b> Immunostaining of FABP5 protein (brown color) in airway epithelial cells from normal smoker. Bar represent 50 µm. <b>B.</b> Immunostaining of FABP5 protein (brown color) in airway epithelial cells from COPD smoker. Bar represent 50 µm. <b>C.</b> FABP5 protein was quantified by densitometry using the Scion Image software (Scion Corporation, Frederick, MD). Data are representative of 5 different subjects per group.</p

FABP5 expression modulates PPAR-γ activity.

<p><b>A.</b> FABP5 down regulation significantly decreases PPAR-γ activity in primary NHBE cell cultures. <b>B.</b> FABP5 up regulation significantly increases PPAR-γ activity in primary NHBE cell cultures. FABP5 shRNA − indicates cells that were knocked down for Firefly luciferase. FABP5 shRNA + indicates cells that were knocked down for FABP5. FABP5 − indicates cells overexpressing GFP. FABP5 + indicates cells overexpressing FABP5. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

FABP5 expression modulates TLR2 and TLR4 mRNA expression.

<p><b>A.</b> TLR2 mRNA expression was induced in cells knocked down for FABP5. <b>B.</b> TLR4 mRNA expression was induced in cells knocked down for FABP5. <b>C.</b> TLR2 mRNA expression was decreased in cells overexpressing FABP5. <b>D.</b> TLR2 mRNA expression was decreased in cells overexpressing FABP5. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

FABP5 overexpression decreases inflammation and increases innate immunity in primary NHBE cells.

<p><b>A.</b> CS exposure decreases <i>P. aeruginosa</i> colony forming units (CFU) on primary NHBE cells overexpressing FABP5. <b>B.</b> FABP5 overexpression blocks CS-induced IL-8 secretion in the basolateral supernatant of primary NHBE cell cultures. <b>C.</b> FABP5 overexpression augments β-defensin 2 mRNA expression in primary NHBE cells. FABP5 − indicates cells overexpressing GFP and FABP5 + indicates cells overexpressing FABP5. Data are representative of 3 independent experiments and are expressed as mean ± SEM.</p

CS inhibits <i>FABP5</i> promoter activity in human lung epithelial BEAS-2B cells.

<p><b>A.</b> Schematic overview of the wild type and mutant c-Jun binding site on the FABP5 promoter at position -495. <b>B.</b> Dual luciferase assay following 2% CSE and/or 1 μg/ml LPS treatment of the wild type <i>FABP5</i> promoter or the promoter mutated at position -495 in the c-Jun binding site. Data are representative of 3 independent experiments.</p

CS inhibits c-Jun activity in human lung epithelial BEAS-2B cells.

<p>Cells were treated with 2% CSE for 24 hours and then stimulated with 1 μg/ml LPS for the indicated time points. <b>A.</b> Phosphorylation of c-Jun at Ser63 and total c-Jun were determined by Western blot. <b>B.</b> Phosphorylation of c-Jun at Ser73 and total c-Jun were determined by Western blot. <b>C.</b> Ratio of total c-Jun and GAPDH densitometry determined by Western blot. <b>D.</b> Activation of c-Jun transcription factor was determined using an ELISA-based TransAM c-Jun activation assay. Data are representative of 6 independent experiments. * p<0.05, ** p<0.01 compared to no CSE and no LPS.</p