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P040 The cytokine milieu in patients with inflammatory bowel disease impacts the phenotype of mesenchymal stromal cells

M. Barnhoorn*1, K. Schepers2, H. Verspaget1, W. Fibbe2, L. Hawinkels1, M. van Pel2, A. van der Meulen - de Jong1

1Leiden University Medical Center, Gastroenterology and Hepatology, Leiden, The Netherlands, 2Leiden University Medical Center, Immunohematology and Blood Transfusion, Leiden, The Netherlands

Background

Mesenchymal stromal cells (MSCs) have the capacity to promote healing of refractory perianal fistulas in Crohn’s disease (CD) and pre-treatment with cytokines may enhance therapeutic efficacy. Furthermore, locally applied MSCs are under clinical development for treatment of refractory proctitis in ulcerative colitis (UC). Despite these clinical advances, the mechanism of action of MSC therapy is largely unknown. We hypothesise that the proinflammatory environment in the patient promotes the immunomodulatory properties of MSCs. Therefore, we analysed cytokine levels in inflamed tissues obtained from CD and UC patients. Next, we assessed the expression of immunomodulatory molecules by MSCs upon exposure to these cytokines.

Methods

U-plex cytokine assay and ELISA were used to measure the levels of 11 cytokines, including interferon-γ, interleukin (IL)-17 and IL-1β, in perianal fistula scraping of patients with CD (n = 20), colonic tissue samples (inflamed and non-inflamed) from patients with UC (n = 18) and adjacent healthy tissue from patients with colorectal carcinoma (n = 18). To determine the response of bone-marrow-derived MSCs to different proinflammatory environments, MSCs were exposed to defined (sets of these) cytokines and the expression of immunomodulatory molecules was determined by flow cytometric and qPCR analyses.

Results

Scrapings of perianal fistulas obtained from CD patients contained high levels of cytokines, including IL-1β and IL-17 (IL-1β 0.102 pg/µg vs. 0.012 pg/µg in normal colon tissue, p = 0.003, and IL-17 0.206 pg/µg vs. 0.009 pg/µg, p < 0.001). In contrast, inflamed colon of UC patients only showed the presence of a selected set of cytokines of which some, like IL-1β, were already present in non-inflamed colons. Next, we evaluated the response of MSCs to exposure of the individual cytokines and 4 different cytokine mixtures which resemble the complex proinflammatory milieus in inflammatory bowel disease. Interestingly, each cytokine mixture induced a unique expression pattern of intra –and extracellularly expressed immunomodulatory molecules in MSCs, including cyclo-oxygenase 2 and indoleamine 2,3-dioxygenase. Assays are ongoing to investigate the consequence of cytokine priming on the immunomodulatory function of MSCs.

Conclusion

The patient’s proinflammatory milieu is strongly dependent on the underlying disease. We found in vitro evidence that infusion of MSCs into inflamed UC tissue or CD fistulas induces up-regulation of immunomodulatory molecules in MSCs that are unique for the patient’s cytokine milieu and that play a role in the immunomodulatory properties of the cells. Differences in cytokine expression between patients may explain the different clinical efficacies that are observed following MSC therapy.