A differential expression of SLAM (Signaling and Activation of Lymphocytes molecule) is induced by gliadin on antigen presenting cells in patients with active Celiac Disease

Abstract

Background: Monocytes (Mo) recruited into the intestinal mucosa are source of macrophages and dendritic cells (DC). Under inflammatory conditions, lymphocytes (L) are co recruited and direct (cell to cell contact) or indirectly (by soluble factors) influence Mo outcome. In vitro interactions between blood cells and soluble factors produced by them may influence Mo activation and differentiation into DC. SLAM (Signaling and Activation of Lymphocytes molecule) and OX40 ligand (OX40-L) co-stimulatory molecules expressed on antigen presenting cells (APC) are involved in the interaction with activated L. Aim: to examine the influence of gliadin on APC in celiac patients (Ce) and healthy controls (Co) by measuring SLAM and OX40-L expression. Materials: 15 adults (> 18y) diagnosed as celiacs and 15 age-matched controls were enroled. Mo activation: Peripheral blood mononuclear cells (PBMC) were isolated, cultured at 106 cells/ml for 24h with complete RPMI-1640, gliadin (25 ug/ml), urea (2M) and LPS ( 1ug/ul), stained with conjugated mAb anti -CD14, -OX40-L and -SLAM, and analyzed by flow cytometry. (FC). Interleukin (IL) -8 was determined by ELISA in supernatants of Mo previously isolated by Percoll gradients. Mo-derive DC: Adherent PBMC were incubated with GM-CSF (800U/ml) and IL-4 (10ng/ml) for 5 days, induced to mature in the presence of gliadin (75ug/ml) for 2 days and analyzed for the expression of surface CD11c, HLA-DR, CD86, SLAM and OX40-L by FC. Median stimulation indexes (Ĩ) were defined as [gliadin-stimulated Mo or DC / basal Mo or DC] to measure IL-8 production and % of double positive Mo (CD14+ plus SLAM/OX40L) and DC (CD11c+ plus SLAM/OX40-L/ CD86/ HLADR). The Ĩ were compared by Mann-Whitney test. Results: IL-8 was spontaneously released by Mo from Co and Ce (p=NS) and further stimulated by gliadin (Ĩ IL-8 : 2.70 ± 0.70 vs. 1.40 ± 0.05, p=0,028; Ce vs. Co). SLAM expression was higher in gliadin-stimulated Mo (Ĩ% CD14+SLAM+: 10.60 ± 1.40 vs. 6.30 ± 0.90, p=0.009; Ce vs. Co) and DC (Ĩ CD11c+SLAM+: 6.30 ±1.10 vs.3.20 ±1.20, p= 0.028; Ce vs. Co). After gliadin treatment, similar Ĩ% CD14+CD86 and Ĩ% CD14+HLA-DR were calculated in mature DC from Ce and Co (p=NS). After gliadin treatments, OX40-L was similarly expressed on Mo and DC from Ce and Co (p=NS). Conclusions: Although we used both a well-defined cytokine cocktail and gliadin as definite inductors for Mo activation, differentiation and DC maduration, SLAM expression was enhanced on APC from Ce patients. Our results suggest that besides the presence of gliadin, a particular peripheral microenvironment might contribute to the amplification and specialization of the adaptive immune response in celiac disease. The functional involvement of SLAM is currently under investigation.