Fig. 2.

Increased phosphorylation of IκB-α and enhanced expression of TNF-α in PDMCs with ORMDL3 KO. A: IκB-α phosphorylation (pIκB-α) levels were determined by immunoblotting of whole-cell lysates from Ag-activated cells using the corresponding antibody (α). Loading control levels of IκB-α and β-actin were also determined. B: Quantification and statistical evaluation of pIκB-α levels as in (A) normalized to nonactivated WT (n = 5) and ORMDL3 KO (n = 8) PDMCs and corresponding β-actin load. C: RT-qPCR quantfication of mRNAs encoding TNF-α in nonactivated WT (n = 5) and ORMDL3 KO PDMCs (n = 7) and Ag-activated WT (n = 7) and ORMDL3 KO PDMCs (n = 7). D: Flow cytometry analysis of TNF-α production in nonactivated and Ag-activated WT and ORMDL3 KO PDMCs. E: Statistical evaluation of TNF-α data. From the PE positive quadrants as in (D); WT (n = 13) and ORMDL3 KO (n = 11). Quantitative data presented are mean ± SEM, calculated from n, which show numbers of biological replicates of independently isolated PDMCs. P values were determined by unpaired two-sided Student's t test.