File S1 - EGF-Induced Bronchial Epithelial Cells Drive Neutrophil Chemotactic and Anti-Apoptotic Activity in Asthma

Supporting Methods. Figure S1, Shows that the PI (3)Kδ-selective inhibitor, IC87114, (10 µM; solid bars) did not significantly (one way ANOVA, p = 0.16) affect either basal-CM and EGF-CM-directed neutrophil chemotaxis compared with the corresponding non-drug treated controls. Vehicle-treated control cells were treated with DMSO (0.1% v/v). Neutrophil chemotaxis was assessed using calcein-loaded cells in a fluorescence-based chemotaxis microplate. Data represent mean ± SEM from PBECs derived from n = 6 different Mod/Sev asthma patients and using peripheral-blood neutrophils from healthy subjects each performed in duplicate. Figure S2, Schematic representation of potential regulation of neutrophil chemotactic and anti-apoptotic responses by an EGF-conditioned asthmatic epithelium. Unresolved airway neutrophilia and disordered airway epithelial function are pathobiological features in more severe forms of asthma. It is proposed that an EGF-conditioned asthmatic epithelium modulates neutrophil migration via a potential signaling mechanism involving RhoA and class IB PI (3)Kγ signaling. EGF-conditioned epithelium also delays neutrophil constitutive apoptosis through the production of GM-CSF and via activation of all class I PI (3)Ks in neutrophils. Table S1, Comparability of the absolute values representing neutrophil chemotactic and anti-apoptotic activity generated by PBEC-CM derived from patients with mild asthma, Mod/Sev asthma and healthy controls. Supporting References. (DOCX)