Novel role of the serine protease inhibitor elafin in gluten-related disorders.

OBJECTIVES: Elafin, an endogenous serine protease inhibitor, modulates colonic inflammation. We investigated the role of elafin in celiac disease (CD) using human small intestinal tissues and in vitro assays of gliadin deamidation. We also investigated the potential beneficial effects of elafin in a mouse model of gluten sensitivity. METHODS: Epithelial elafin expression in the small intestine of patients with active CD, treated CD, and controls without CD was determined by immunofluorescence. Interaction of elafin with human tissue transglutaminase-2 (TG-2) was investigated in vitro. The 33-mer peptide, a highly immunogenic gliadin peptide, was incubated with TG-2 and elafin at different concentrations. The degree of deamidation of the 33-mer peptide was analyzed by liquid chromatography-mass spectrometry. Elafin was delivered to the intestine of gluten-sensitive mice using a recombinant Lactococcus lactis vector. Small intestinal barrier function, inflammation, proteolytic activity, and zonula occludens-1 (ZO-1) expression were assessed. RESULTS: Elafin expression in the small intestinal epithelium was lower in patients with active CD compared with control patients. In vitro, elafin significantly slowed the kinetics of the deamidation of the 33-mer peptide to its more immunogenic form. Treatment of gluten-sensitive mice with elafin delivered by the L. lactis vector normalized inflammation, improved permeability, and maintained ZO-1 expression. CONCLUSIONS: The decreased elafin expression in the small intestine of patients with active CD, the reduction of 33-mer peptide deamidation by elafin, coupled to the barrier enhancing and anti-inflammatory effects observed in gluten-sensitive mice, suggest that this molecule may have pathophysiological and therapeutic importance in gluten-related disorders.

Protective effects of lactococci strains delivering either IL-10 protein or cDNA in a TNBS-induced chronic colitis model.

BACKGROUND: Oral treatment with Lactococcus lactis strains secreting the anti-inflammatory cytokine interleukin (IL)-10 has previously shown success as a therapy for inflammatory bowel diseases (IBD). GOALS: Our aim was to compare the protective effects of IL-10, delivered by recombinant lactoccoci using 2 novel expression systems, in a murine colitis model mimicking the relapsing nature of IBD. The first system is based on a Stress-Inducible Controlled Expression system for the production and delivery of heterologous proteins at mucosal surfaces and the second allows the delivery to the host cells of an il-10 cDNA cassette, harbored in a eukaryotic DNA expression vector (pValac). STUDY: Colitis was induced in female BALB/c mice by intrarectal injection of 2,4,6-trinitrobenzenesulphonic acid (TNBS). Mice that recovered received one of the bacteria treatments or saline solution orally during 14 days. Colitis was reactivated 25 days after the first TNBS injection with a second TNBS challenge. Three days after colitis reactivation, cytokine profiles and inflammation in colon samples were evaluated. RESULTS: Animals (N=9) receiving L. lactis strains secreting IL-10 using Stress-Inducible Controlled Expression system or delivering pValac:il-10 plasmid showed lower weight loss (P<0.005), lower damage scores (P<0.005), and immune activation in their large intestines compared with inflamed nontreated mice. CONCLUSIONS: Our results confirm the protective effect of IL-10 delivered either as a protein or as a cDNA in a colitis model mimicking the relapsing nature of IBD and provides a step further in the "proof-of-concept" of genetically engineered bacteria as a valid system to deliver therapeutic molecules at mucosal level.

Selection of a novel strain of Christensenella minuta as a future biotherapy for Crohn's disease.

Microbiome-based therapies for inflammatory bowel diseases offer a novel and promising therapeutic approach. The human commensal bacteria of the species Christensenella minuta (C. minuta) have been reported consistently missing in patients affected by Crohn's disease (CD) and have been documented to induce anti-inflammatory effects in human epithelial cells, supporting their potential as a novel biotherapy. This work aimed at selecting the most promising strain of C. minuta for future development as a clinical candidate for CD therapy. Here, we describe a complete screening process combining in vitro and in vivo assays to conduct a rational selection of a live strain of C. minuta with strong immunomodulatory properties. Starting from a collection of 32 strains, a panel of in vitro screening assays was used to narrow it down to five preclinical candidates that were further screened in vivo in an acute TNBS-induced rat colitis model. The most promising candidate was validated in vivo in two mouse models of colitis. The validated clinical candidate strain, C. minuta DSM 33715, was then fully characterized. Hence, applying a rationally designed screening algorithm, a novel strain of C. minuta was successfully identified as the most promising clinical candidate for CD.