Gliadin stimulates SLAM expression on human monocytes?.

L.GUILLÉN; ; N. PERIOLO; M BARBOZA; A. CHERÑAVSKY

Iguassu Falls . Brasil.

Congreso; World Congress of Pediatric Gastroenterology, Hepatology and Nutrition. ( WCPGHAN3).; 2008

Latin American Society for Pediatric, Gastroenterology , Hepatology and Nutrition ( LASPGHAN)

Gliadin stimulates SLAM expression on human monocytes .Objectives: Since gliadin activates human blood monocytes by chemokine induction, activatedmonocytes contribute to immune response in Celiac Disease, and the expression of SLAM(Signaling Limphocytic Activation Molecule) might be a sign of innate immune activation ofmonocytes by different stimuli, we aimed to search for SLAM expression on monocytes (Mo)after treatment with gliadin.Materials/methods: Buffy coats from six normal healthy volunteers were processed for theisolation of monocytes using standard gradients of Ficoll-Hypaque followed by centrifugationby Percoll. Monocyte suspensions (70%) were cultured (106 cells/ml) for 24h in complete RPMI-1640 with peptic fragments of gliadin (5-100 μg/ml) digested with a-chemotripsyn 200:1 (w/w)in 0,1M NH4HCO3 / urea 2M, lipopolysaccharide (LPS)(1ug/ul), UREA 2M and zein(100ug/ml). Doble immunofluorescence staining with anti CD14 PE, - SLAM FITC and flowcytometry analysis was performed. Percentages of CD14+SLAM+ double positive cells andmean fluorescence intensity (MFI) were compared between different incubation conditions. Thelevel of IL-8 was determined in cell culture supernatants collected 24 hs of cultivation byELISA according to the manufacturer´s instructions (BD Biosciences Set). Reagnts were testedby LAL test for LPS.Resultados: The monocytes spontaneously released IL-8 (12244 pg/ml).Meanwhile IL-8 wasstimulated by addition of zein to the culture of monocytes (19542 pg/ml), peptides gliadinproduced 2.9-fold more IL-8 (57961 pg/ml). UREA did not induce production of IL-8 bymonoocytes (13946 pg/ml ). The expression of SLAM was evaluated on CD14+ subpopulation.SLAM was neither expresed on fresh monocytes (3,6 %; 18,03 FMI), nor monocytes 24 hcultures in RPMI (3,8%; 2,26 FMI) or with UREA 2M (1,8%; 29.6 FMI). SLAM waspreferentially induced by LPS (36,9 %; 102,72 IFM) and gliadin peptides (25ug/ml) (32,28%;102,72 IFM). By contrast, Zein did not induce SLAM on monocytes (2,4 %; 24.37 FMI).Conclusions: While food components (gliadin, zein) induced chemokine production bymonocytes, SLAM was only up regulated after gliadin stimulation. As was previously observedfor dendritic cells, gliadin also induced monocytes activation. SLAM might be involved in theknown association between innate response to gliadin and activation of pathogenic T cells inCD.