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P008. Enhanced induction of regulatory macrophages upon azathioprine/infliximab combination treatment in vitro

P008. Enhanced induction of regulatory macrophages upon azathioprine/infliximab combination treatment in vitro

A.C.W. Vos, M. Duijvestein, M.E. Wildenberg, A.P. Verhaar, G.R. van den Brink, D.W. Hommes

Leiden University Medical Center, Leiden, The Netherlands

Introduction and Aim: We have previously shown that anti-TNF antibodies induce regulatory macrophages in an Fc region dependent manner in vitro (Gastroenterology 2010). Regulatory macrophages play an important role in wound healing and gut homeostasis and have anti-inflammatory properties. In addition, a recent study has shown that a combination of infliximab and azathioprine therapy is more effective than infliximab or azathioprine monotherapy in inducing and maintaining remission in Crohn's disease (CD) patients (New Engl J Med 2010). The aim of this study was to examine the effects of infliximab/azathioprine combination therapy on the induction and function of regulatory macrophages.

Methods: Mixed lymphocyte reactions (MLR) were established using PBMCs from two healthy donors in a 1:1 ratio and treated with either infliximab, azathioprine or both. Macrophage phenotype was evaluated by flow cytometry. Inhibition of T cell proliferation was measured in a secondary MLR containing macrophages and third party lymphocytes. Cytokine secretion was measured using a cytomatric bead assay.

Results: Alternatively activated macrophages were induced upon treatment of MLR with anti-TNF antibodies (infliximab or adalimumab). These macrophages showed anti-inflammatory properties in terms of their ability to inhibit proliferation of activated T cells, expression of the regulatory macrophage marker CD206, as well as production of IL-10 in response to LPS. When cells in an MLR were treated with a combination of infliximab and azathioprine, an enhanced induction of regulatory macrophages was observed. Not only was the number of regulatory macrophages increased upon combination treatment, the macrophages also had a more potent phenotype. Compared to regulatory macrophages induced by infliximab monotreatment, regulatory macrophages induced by combination treatment had higher expression of the regulatory macrophage marker CD206 and showed enhanced capacity to inhibit proliferation of activated T cells.

Conclusion: We show an enhanced induction of regulatory macrophages upon infliximab/azathioprine combination treatment in vitro. In addition, macrophages induced by combination treatment showed a stronger immunosuppressive phenotype. This mechanism of action may play a role in mucosal healing in patients with IBD and might explain the enhanced efficacy of infliximab/azathioprine combination therapy.