Peter D. Howdle

Newly identified genetic risk variants for celiac disease related to the immune response

Our genome-wide association study of celiac disease previously identified risk variants in the IL2–IL21 region. To identify additional risk variants, we genotyped 1,020 of the most strongly associated non-HLA markers in an additional 1,643 cases and 3,406 controls. Through joint analysis including the genome-wide association study data (767 cases, 1,422 controls), we identified seven previously unknown risk regions (P < 5 × 10−7). Six regions harbor genes controlling immune responses, including CCR3, IL12A, IL18RAP, RGS1, SH2B3 (nsSNP rs3184504) and TAGAP. Whole-blood IL18RAP mRNA expression correlated with IL18RAP genotype. Type 1 diabetes and celiac disease share HLA-DQ, IL2–IL21, CCR3 and SH2B3 risk regions. Thus, this extensive genome-wide association follow-up study has identified additional celiac disease risk variants in relevant biological pathways.

Decreased antioxidant status and increased lipid peroxidation in patients with septic shock and secondary organ dysfunction

OBJECTIVE To determine antioxidant vitamin concentrations, lipid peroxidation, and an index of nitric oxide production in patients in the intensive care unit (ICU) with septic shock and relate the findings to the presence of secondary organ failure. DESIGN A prospective, observational study. SETTING A nine-bed ICU in a University teaching hospital. PATIENTS Sixteen consecutive patients with septic shock, defined as: a) clinical evidence of acute infection; b) hypo- or hyperthermia (< 35.6 degrees C or > 38.3 degrees C); c) tachypnea (> 20 breaths/min or being mechanically ventilated); d) tachycardia (> 90 beats/min); e) shock (systolic pressure < 90 mm Hg) or receiving inotropes. Fourteen patients also had secondary organ dysfunction. INTERVENTIONS None. MEASUREMENTS AND MAIN RESULTS Antioxidant vitamin concentrations were significantly lower in the patients than the reference range obtained from a comparable group of healthy controls. The mean plasma retinol (vitamin A) concentration was 26.5 +/- 19.3 micrograms/dL compared with 73.5 +/- 18.3 micrograms/dL in healthy subjects (p < .01). Additionally, 13 (81%) patients had retinol values below the lower limit of our reference range (< 37.0 micrograms/dL). Tocopherol (vitamin E) plasma concentrations were below the reference range in all patients (< 9.0 mg/L), with a mean value of 3.6 +/- 2.0 mg/L compared with 11.5 +/- 1.3 mg/L in healthy subjects (p < .001). Plasma beta carotene and lycopene concentrations were undetectable (< 15 micrograms/L) in eight (50%) patients, and below our reference range (< 101 micrograms/L and < 154 micrograms/L, respectively) in the remaining patients. In the five patients with three or more dysfunctional secondary organs, plasma thiobarbituric acid-reactive substances were significantly increased (p < .05), suggesting increased lipid peroxidation. Concentrations of thiobarbituric acid-reactive substances correlated negatively with both plasma retinol and plasma tocopherol (r2 = .42, p < .01 and r2 = .48, p < .005, respectively). In the five patients from whom we were able to collect urine, nitrite excretion was increased approximately 400-fold (p < .001). CONCLUSIONS These data indicate decreased antioxidant status in the face of enhanced free radical activity, and suggest potential therapeutic strategies involving antioxidant repletion.

Guidelines for the investigation of chronic diarrhoea, 2nd edition

1.0 PREFACE 1.1 Purpose of guidelines These guidelines were compiled at the request of the Chairman of the British Society of Gastroenterology’s clinical services committee. The guidelines are directed at consultant gastroenterologists, specialist registrars in training, and general practitioners, and refer specifically to adult not paediatric gastroenterology. Their purpose is to provide guidance on the best available methods of investigating symptoms of chronic diarrhoea. Given this broad symptom based focus, the guidelines cover a wide range of gastroenterological conditions and are not intended as a comprehensive review of all aspects of the clinical conditions mentioned herein, but rather an attempt to rationalise the approach to investigation in the context of this common clinical scenario. 1.2 Development of guidelines The guidelines were prepared following a comprehensive literature search by Dr PD Thomas. This involved a review of electronic databases (Medline and PubMed) using keywords such as “diarrhea”, “chronic”, “diagnostic evaluation”, “investigation”, “malabsorption”, and terms related to the specific conditions mentioned in the text (for example, coeliac disease and small bowel bacterial overgrowth). Papers relating to diarrhoea in the context of immunodeficiency syndromes were specifically excluded from this review as this subject was felt to require a different investigative approach. A total of 530 key papers and relevant abstracts in English in peer reviewed journals were identified and read, and relevant work has been cited and referenced. An initial draft document was produced and subsequently reviewed and modified by a multidisciplinary group comprising clinical gastroenterologists, radiologists, and biochemists.

Contribution of the MHC region to the familial risk of coeliac disease

Susceptibility to coeliac disease is genetically determined by possession of specific HLA-DQ alleles, acting in concert with one or more non-HLA linked genes. The pattern of risk seen in sibs and twins in coeliac disease is most parsimonious with a multiplicative model for the interaction between the two classes of genes. Based on a sib recurrence risk for coeliac disease of 10% and a population prevalence of 0.0033, the sib relative risk is 30. To evaluate the contribution of the MHC region to the familial risk of coeliac disease, we have examined haplotype sharing probabilities across this region in 55 coeliac disease families. Based on these probabilities the sib relative risk of coeliac disease associated with the MHC region is 3.7. Combining these results with published data on allele sharing at HLA, the estimated sib relative risk associated with the MHC region is 3.3. Therefore, the MHC genes contribute no more than 40% of the sib familial risk of coeliac disease and the non-HLA linked gene (or genes) are likely to be the stronger determinant of coeliac disease susceptibility.

Coeliac disease-associated risk variants in TNFAIP3 and REL implicate altered NF-κB signalling

Objective: Our previous coeliac disease genome-wide association study (GWAS) implicated risk variants in the human leucocyte antigen (HLA) region and eight novel risk regions. To identify more coeliac disease loci, we selected 458 single nucleotide polymorphisms (SNPs) that showed more modest association in the GWAS for genotyping and analysis in four independent cohorts. Design: 458 SNPs were assayed in 1682 cases and 3258 controls from three populations (UK, Irish and Dutch). We combined the results with the original GWAS cohort (767 UK cases and 1422 controls); six SNPs showed association with p<1×10−04 and were then genotyped in an independent Italian coeliac cohort (538 cases and 593 controls). Results: We identified two novel coeliac disease risk regions: 6q23.3 (OLIG3-TNFAIP3) and 2p16.1 (REL), both of which reached genome-wide significance in the combined analysis of all 2987 cases and 5273 controls (rs2327832 p = 1.3×10−08, and rs842647 p = 5.2×10−07). We investigated the expression of these genes in the RNA isolated from biopsies and from whole blood RNA. We did not observe any changes in gene expression, nor in the correlation of genotype with gene expression. Conclusions: Both TNFAIP3 (A20, at the protein level) and REL are key mediators in the nuclear factor kappa B (NF-κB) inflammatory signalling pathway. For the first time, a role for primary heritable variation in this important biological pathway predisposing to coeliac disease has been identified. Currently, the HLA risk factors and the 10 established non-HLA risk factors explain ∼40% of the heritability of coeliac disease.

THE EFFECTS OF INTRAVENOUS ANTIOXIDANTS IN PATIENTS WITH SEPTIC SHOCK

Oxidative stress is implicated in septic shock. We investigated the effect of intravenous antioxidant therapy on antioxidant status, lipid peroxidation, hemodynamics and nitrite in patients with septic shock. Thirty patients randomly received either antioxidants (n-acetylcysteine 150 mg/kg for 30 min then 20 mg/kg/h plus bolus doses of 1 g ascorbic acid and 400 mg alpha-tocopherol) or 5% dextrose. Basal vitamin C was low and redox-reactive iron was elevated in all patients. In the 16 patients receiving antioxidants, vitamin C increased (p = .0002) but total antioxidant capacity was unaffected. Lipid peroxides were elevated in all patients but did not increase further in the patients receiving antioxidants. Plasma total nitrite also increased (p = .007) in the antioxidant group. Heart rate increased in patients receiving antioxidants at 60 min (p = .018) and 120 min (p = .004). Cardiac index also increased at 60 min (p = .007) and 120 min (p = .05). Systemic vascular resistance index decreased at 120 min in the antioxidant treated patients (p = .003). The effect of antioxidants on hemodynamic variables has not previously been reported. Antioxidant administration may be a useful adjunct to conventional approaches in the management of septic shock.

Evidence of high sugar intake, and low fibre and mineral intake, in the gluten-free diet

Aliment Pharmacol Ther 2010; 32: 573–581

Associations with tight junction genes PARD3 and MAGI2 in Dutch patients point to a common barrier defect for coeliac disease and ulcerative colitis.

Background: Coeliac disease (gluten-sensitive enteropathy; GSE) and inflammatory bowel disease (IBD) are common gastrointestinal disorders. Both display enhanced intestinal permeability, initiated by gluten exposure (GSE) or bacterial interactions (IBD). Previous studies showed the association of both diseases with variants in MYO9B, presumably involved in epithelial permeability. Aim: It was hypothesised that genetic variants in tight junction genes might affect epithelial barrier function, thus contributing to a shared pathogenesis of GSE and IBD. Methods: This hypothesis was tested with a comprehensive genetic association analysis of 41 genes from the tight junction pathway, represented by 197 tag single nucleotide polymorphism (SNP) markers. Results: Two genes, PARD3 (two SNPs) and MAGI2 (two SNPs), showed weak association with GSE in a Dutch cohort. Replication in a British GSE cohort yielded significance for one SNP in PARD3 and suggestive associations for two additional SNPs, one each in PARD3 and MAGI2. Joint analysis of the British and Dutch data further substantiated the association for both PARD3 (rs10763976, p = 6.4×10−5; OR 1.23, 95% CI 1.11 to 1.37) and MAGI2 (rs6962966, p = 7.6×10−4; OR 1.19, 95% CI 1.08 to 1.32). Association was also observed in Dutch ulcerative colitis patients with MAGI2 (rs6962966, p = 0.0036; OR 1.26, 95% CI 1.08 to 1.47), and suggestive association with PARD3 (rs4379776, p = 0.068). Conclusions: These results suggest that coeliac disease and ulcerative colitis may share a common aetiology through tight junction-mediated barrier defects, although the observations need further replication.

Advances in celiac disease.

Purpose of review To critically summarize recent research in celiac disease. Recent findings There are new serological markers with potential use not only in the diagnosis of celiac disease but also as important follow-up tools. As our understanding of celiac disease increases with further isolation of nonhuman leukocyte antigen genes and clarification of the intracellular pathways that underlie its pathogenesis, there are new modalities which will not only allow improved risk stratification of individuals but also facilitate the development of novel therapeutic agents. Summary Small bowel biopsy remains the gold standard for both diagnosis and monitoring. A gluten-free diet currently remains the only treatment option, with potential other options being discovered such as glutenases for predigestion of gluten.

Mycobacterium paratuberculosis DNA not detected in Crohn's disease tissue by fluorescent polymerase chain reaction.

The role of mycobacteria in the aetiology of Crohns disease has been a contentious subject for many years. Mycobacterium paratuberculosis is known to cause a chronic granulomatous enteritis in animals (Johnes disease) and has been implicated as a possible infectious cause of Crohns disease. However this fastidious organism is only rarely detected by conventional microbiological techniques. This study used oligonucleotide primers to the species-specific M paratuberculosis IS900 DNA insertion element and the polymerase chain reaction to amplify any M paratuberculosis DNA from intestinal tissue DNA extracts. One oligonucleotide primer was fluorochrome-labelled and the presence of fluorescent amplified product was determined using an automated DNA sequencer with a computerised gel-scanning laser. This method was shown capable of detecting 1-2 mycobacterial genomes. Intestinal tissue samples were obtained from 68 patients with histologically confirmed Crohns disease, 49 patients with histologically confirmed ulcerative colitis, and 26 non-inflammatory bowel disease controls. In no case was M paratuberculosis detected in any of the inflammatory bowel disease tissue samples and only one non-inflammatory bowel disease case was positive. These results do not support the hypothesis that M paratuberculosis has an aetiological role in Crohns disease.