Isolation and Modulation of Pathogen-specific T cells from Patients with Inflammatory Bowel Diseases

RENATA CURCIARELLO; CANZIANI KARINA; SERRADELL MARÍA DE LOS ANGELES; MARTÍN RUMBO; AYELÉN HUGO; ANDRÉS ROCCA; SANTIAGO BRAYER; ALICIA SAMBUELLI; MARTÍN YANTORNO; GUSTAVO J. CORREA; NÉSTOR CHOPITA; GUILLERMO HORACIO DOCENA; MUGLIA CECILIA

Mar del Plata

Congreso; LXIV Reunion Anual de la Sociedad Argentina de Inmunologia; 2016

Sociedad Arg de Inmunología, Sociedad Argentina de Investigación clínica (SAIC) y Sociedad Argentina de Farmacología Experimental

Inflammatory bowel diseases (IBD) are a complex group of pathologies amongst which ulcerative colitis (UC) and Crohn´s disease (CD) are the most conspicuous. Lamina propria T cells (LPTC) are key cells in IBD pathogenesis, contributing to mucosal inflammation by secreting pro-inflammatory cytokines. It has been shown that LPTC in IBD are resistant to apoptosis. Kefir is a fermented milk that has been used in Eastern Europe with health-promoting properties. The aim of this work was to isolate and establish primary cultures of LPTC from IBD patients and modulate their pro-inflammatory response using microorganisms from kefir. Colonic biopsies from IBD patients (N=8, 5 CD and 3 UC) were washed in HBSS medium with EDTA and DTT, and digested by collagenase and DNAse treatment. In order to select pathogen-specific T lymphocytes, cells were cultured with extracts of enteroadhesive (EA) Escherichia coli and IL-2. Microorganisms from kefir (Lactobacillus kefiri and Enterococcus durans) were evaluated for their modulation capacity on LPTC by proliferation assays (flow cytometry with CFSE) and cytokine secretion (TNF and IL-10 by ELISA) of T cell lines stimulated with anti-CD3 and anti-CD28 followed by culture with probiotic bacteria. LPTC lines specific for EA E. coli were developed for all patients. Cell proliferation of activated lymphocytes decreased with the presence of L. kefiri and E. durans (proliferation index: 3.0±0.54 vs 1.3±0.33 and 0.56±0.28 respectively; unstimulated control: 1.02±0.03). Besides, TNF secretion was dampened in activated LPTC incubated with L. kefiri compared to medium (P˂0.05). No significant differences were observed for IL-10 secretion. Our results show that these probiotics from kefir modulate pathogen specific activated T cells form IBD patients. These promising results could provide the basis for future therapies in IBD patients.