Figure 6.

(A) NanoString analysis of inflammation related genes including NF-κB pathway, JAK/STAT pathway, cytokines and inflammasome was performed for mRNA from unstimulated PBMCs of three mutation carriers (IV:2, III:2 and II:2) and age/sex-matched controls. The average fold change in gene expression of inflammation related genes is illustrated. (B) NF-κB p65 Ser529 phosphorylation in CD3 lymphocytes. The NF-κB p65 Ser529 phosphorylation was measured by flow cytometry. Two mutation carriers (III:1 and III:2) and sex-matched controls´ PBMCs were unstimulated or stimulated with PMA or LPS for 15 minutes in vitro and stained for NF-κB p65 and CD3 markers. (C) Inflammasome activity assay. Mutation carrier and sex/age-matched control PBMCs were either unstimulated, ATP-stimulated, LPS-stimulated or LPS/ATP stimulated in vitro for 6 hours and 45 minutes (ATP added at 6 hours). The IL-1β production was measured from medium with ELISA. PBMCs from an autoinflammatory patient caused by NFKB1 p.Arg157X mutation were used for comparison to demonstrate uncontrolled IL-1β production.