Macrophage-Specific NF-κB Activation Dynamics Can Segregate Inflammatory Bowel Disease Patients

• Main Authors: Stamatia Papoutsopoulou, Michael D. Burkitt, François Bergey, Hazel England, Rachael Hough, Lorraine Schmidt, David G. Spiller, Michael H. R. White, Pawel Paszek, Dean A. Jackson, Vitor A. P. Martins Dos Santos, Gernot Sellge, D. Mark Pritchard, Barry J. Campbell, Werner Müller, Chris S. Probert
• Format: Article
• Language: English
• Published: Frontiers Media S.A. 2019-09-01
• Series: Frontiers in Immunology
• Subjects: inflammatory bowel disease; NF-κB; macrophages; cytokines; Crohn's disease; ulcerative colitis
• Online Access: https://www.frontiersin.org/article/10.3389/fimmu.2019.02168/full

The heterogeneous nature of inflammatory bowel disease (IBD) presents challenges, particularly when choosing therapy. Activation of the NF-κB transcription factor is a highly regulated, dynamic event in IBD pathogenesis. Using a lentivirus approach, NF-κB-regulated luciferase was expressed in patient macrophages, isolated from frozen peripheral blood mononuclear cell samples. Following activation, samples could be segregated into three clusters based on the NF-κB-regulated luciferase response. The ulcerative colitis (UC) samples appeared only in the hypo-responsive Cluster 1, and in Cluster 2. Conversely, Crohn's disease (CD) patients appeared in all Clusters with their percentage being higher in the hyper-responsive Cluster 3. A positive correlation was seen between NF-κB-induced luciferase activity and the concentrations of cytokines released into medium from stimulated macrophages, but not with serum or biopsy cytokine levels. Confocal imaging of lentivirally-expressed p65 activation revealed that a higher proportion of macrophages from CD patients responded to endotoxin lipid A compared to controls. In contrast, cells from UC patients exhibited a shorter duration of NF-κB p65 subunit nuclear localization compared to healthy controls, and CD donors. Analysis of macrophage cytokine responses and patient metadata revealed a strong correlation between CD patients who smoked and hyper-activation of p65. These in vitro dynamic assays of NF-κB activation in blood-derived macrophages have the potential to segregate IBD patients into groups with different phenotypes and may therefore help determine response to therapy.