Differences in immune cell population subsets in inflammatory bowel disease patients under anti-TNF treatment

Abstract

Background: In autoimmune diseases targets attacked by the immune system differ one from another and the immune system deregulation seems to be the main cause of these pathologies. The aim of this study was to determine the existence of a differential pattern in the immune system cells (in terms of cell percent, ability of cytokines production and transcription factor activation) in inflammatory bowel disease (IBD) patients under infliximab (IFX) therapy.

Methods: A pilot case–control study was performed. Inclusion criteria were IBD patients in clinical remission under IFX treatment. After informed consent, blood samples were obtained in IBD patients just before IFX infusions and in a healthy control. Patients were classified in different groups: Crohn’s disease (CD) without rheumatologic manifestations (Group 1), ulcerative colitis (UC) without rheumatologic manifestations (Group 2) and IBD patients with associated rheumatoid arthritis (RA) (Group 3). Blood samples were used to determine the immune cell status of patients and negative control. To investigate the immune system cell distribution, peripheral mononuclear blood cells were isolated from fresh blood to characterise: monocyte, dendritic cells (DC), Th1, Th17, Treg, and B cell. Cells were then incubated with specific fluorescent antibodies’ cocktails, then identified with flow cytometry. T cells ability to produce TNF-α, IL-17 and INF-γ was tested by performing intra cellular staining, while T-bet, Fox-P3, and Ror-γ t expression was tested trough intra nuclear staining. Data were collected with flow cytometry.

Results: Fifteen IBD patients (60% female, mean age 42) were consecutively included, 7 CD,5 UC and 3 IBD with RA. The surface staining demonstrated differences between the group’s cell subtype. CD and IBD-RA patients showed a decrease of CD25Hi CD127- Treg subset in comparison with negative control. Decrease of transitional B cell subset CD38Hi CD24Hi CD19+ was observed in CD patients, while UC patients maintain normal values. The cytokine production in T cell, showed a significative increase of TNF-α, especially in IL-17 with a five-fold increase, while no significant difference in IFN-γ production, in CD and UC patients. Regarding the transcription factor expression T-bet and Ror-γ t increased significantly in CD and UC vs. negative control. T-bet was more specifically expressed in UC, whereas Ror-γ t more in CD.

Conclusions: The immune system cell subset is highly modified by the disease type (CD, UC, IBD+RA). IFX treatment does not seem to unmask the immune system celĹs capability to produce proinflammatory cytokines. Transcription factors expression showed that patients are affected by a Th1 disease, due to the increase of T-bet.