Oriol Juanola

Gut Bacterial DNA Translocation is an Independent Risk Factor of Flare at Short Term in Patients With Crohn's Disease.

OBJECTIVES:We aimed at evaluating bacterial DNA (bactDNA) presence in blood of Crohn’s disease (CD) patients in remission as an independent risk factor of flare at 6 months.METHODS:This is a prospective, multicenter study on CD patients with Crohn’s disease activity index (CDAI)<150. The primary end point was time-to-relapse as evaluated by CDAI>150 in the following 6 months. BactDNA in blood, the nucleotide-binding oligomerization domain containing 2 (NOD2) genotype, and serum cytokine levels were determined at baseline.RESULTS:A total of 288 patients were included. BactDNA was detected in 98 patients (34.0%). A variant-NOD2 genotype was identified in 114 patients (39.6%). Forty patients (14%) relapsed during follow-up. Multivariate survival analysis identified bactDNA as an independent risk factor of flare (hazard ratio (HR) 8.75 (4.02–19.06) 95% confidence interval (CI)). Hospitalization, surgery, switch of treatment, initiation and escalation of anti-tumor necrosis factor (TNF) therapy, steroids initiation, and increased fecal calprotectin levels at 6 months were associated with bactDNA at baseline. A logistic regression analysis showed bactDNA as an independent and significant predictive factor of hospitalization (odds ratio (OR) 11.9 (3.4–42.3); P<0.001), steroids startup (OR 8.5 (2.7–27.1); P<0.001), and switch of treatment (OR 3.5 (1.6–7.7); P=0.002) at 6 months. No relationship was observed between bactDNA and mucosal lesions in patients with colonoscopy at admission. Serum pro-inflammatory cytokines were significantly increased in patients with bactDNA or a variant-NOD2 genotype. The combination of both factors induced decreased anti-TNF-α levels and a higher percentage of patients on intensified anti-TNF therapy.CONCLUSIONS:BactDNA is an independent risk factor of relapse at 6 months in CD patients. BactDNA is also independently associated with an increased risk of hospitalization, switch of treatment, and steroids initiation.

Bifidobacterium pseudocatenulatum CECT7765 induces an M2 anti-inflammatory transition in macrophages from patients with cirrhosis

BACKGROUND & AIMS Patients with cirrhosis show recurrent access of bacterial products into the bloodstream inducing a multi-altered immunological status leading to relevant complications. We aimed at evaluating Bifidobacterium pseudocatenulatum CECT7765 effect on the hosts macrophage function. PATIENTS & METHODS Patients with cirrhosis and ascites were included. Granulocyte-macrophage colony-stimulating factor (GM-CSF) monocyte-derived and ascitic fluid (AF) macrophages were cultured with M-CSF, lipopolysaccharide (LPS) and/or the bifidobacterial strain. Pellets and supernatants were evaluated for gene expression of M1 and M2-related genes and cytokine secretion. Cell surface expression molecules were evaluated by flow cytometry. Kupffer cells from bile duct ligated and CCl4 rats were also evaluated. RESULTS Experiments were run on GM-CSF blood-derived and AF macrophages from 10 patients with cirrhosis and 10 healthy donors. Different macrophage morphology was observed by optical microscopy in cells stimulated with bifidobacteria vs. LPS. M2-like expression of CD206, CD163 and CD16 was significantly increased in macrophages after stimulation with the bifidobacterial strain vs. LPS. B. pseudocatenulatum CECT7765 was able to significantly change the cytokine secretion pattern of blood-derived and AF macrophages and Kupffer cells from bile duct ligated and CCl4 cirrhotic rats compared to that induced by LPS. B. pseudocatenulatum CECT7765 was also effective in inducing a phenotype transition and a functional change from an M1- to an M2-related gene expression and cytokine secretion pattern in AF macrophages even after LPS-pretreatment. B. pseudocatenulatum CECT7765 did not reduce AF macrophage bacterial killing capacity. CONCLUSION B. pseudocatenulatum CECT7765 induces a morphologic, phenotypic and functional transition towards an anti-inflammatory profile in GM-CSF monocyte-derived and AF macrophages from patients with cirrhosis that may help in controlling sustained inflammation in decompensated cirrhosis.

Toll-like receptor polymorphisms compromise the inflammatory response against bacterial antigen translocation in cirrhosis

Bacterial translocation is associated with clinically relevant complications in cirrhosis. We evaluated the effect of toll-like receptor polymorphisms in the soluble response against these episodes. Consecutive patients with cirrhosis and ascitic fluid were distributed by TLR2 rs4696480, TLR4 rs4986790, and TLR9 rs187084 single-nucleotide polymorphisms. Lipoteichoic acid, lipopolyssaccharide, bacterial-DNA, pro-inflammatory cytokines and nitric oxide levels were quantified in serum samples. In vitro response against specific ligands in variant TLR genotypes was evaluated. One hundred and fourteen patients were included. Variant TLR-2, TLR-4 and TLR-9 SNP genotypes were associated with significantly increased serum levels of LTA, LPS and bacterial-DNA. TNF-α, IL-6 and nitric oxide serum levels were significantly decreased in all variant TLR genotyped patients. Cytokine levels were significantly less upregulated in response to specific TLR-ligands in patients with all variant vs wildtype TLR genotypes. Although in vitro gene expression levels of all wildtype and variant TLRs were similar, MyD88 and NFkB were significantly downregulated in cells from TLR-variant genotyped patients in response to their ligands. Variant TLR genotypes are associated with an increased circulating antigen burden and a decreased proinflammatory response in cirrhosis. This immunodeficiency may facilitate bacteria-related complications in cirrhosis and enhance TLR targeting for its management.

Selective intestinal decontamination with norfloxacin enhances a regulatory T cell‐mediated inflammatory control mechanism in cirrhosis

Norfloxacin exerts immunomodulatory effects in cirrhosis beyond its bactericidal activity. We aimed at identifying the role of regulatory T (Treg) cells in the norfloxacin mechanism that compensates the inflammatory environment in cirrhosis.

Regulatory T Cells Restrict Permeability to Bacterial Antigen Translocation and Preserve Short-Chain Fatty Acids in Experimental Cirrhosis

Intestinal permeability to translocation of bacterial products is increased in cirrhosis. Regulatory T cells (Tregs) remain central to the interplay between the host and microbial milieu. We propose that Tregs are involved in promoting gut barrier integrity and a balanced interaction with gut microbiota–derived short‐chain fatty acids (SCFAs). Carbon tetrachloride cirrhosis was induced in wild‐type and recombination activating gene 1 (Rag1)‐/‐ mice. Naive T cells and Treg cells were transferred into Rag1‐/‐ mice. Intestinal permeability was assessed in vivo after lipopolysaccharide (LPS) oral administration, and bacterial DNA presence was evaluated in mesenteric lymph nodes. Transcript and protein levels of tight‐junction (TJ) proteins were measured in colonic tissue. Intestinal T helper profile in response to Escherichia coli (E. coli) was determined by flow cytometry. SCFAs were measured by gas chromatography–mass spectrometry in colonic content before and after E. coli challenge. Rag1‐/‐ mice showed significantly increased permeability to LPS and bacterial DNA translocation rate compared with control mice. Naive T and Treg cotransfer significantly reduced gut permeability to bacterial antigen translocation and restored TJ protein expression in Rag1‐/‐ mice. Naive T and Treg replenishment in Rag1‐/‐ mice restrained proinflammatory differentiation of intestinal lymphocytes in response to E. coli. The main SCFA concentration resulted in significant reduction in Rag1‐/‐ mice after E. coli administration but remained unaltered after naive T and Tregs cotransfer. The reduced expression of SCFA receptors induced by E. coli was reestablished following naive T and Treg reconstitution in Rag1‐/‐ mice. Conclusion: The restriction of gut permeability, local inflammatory differentiation, and loss of bacteria‐derived SCFAs foster the value of Tregs in preventing bacterial translocation in cirrhosis.