Antitissue Transglutaminase Normalization Postdiagnosis in Children With Celiac Disease.

OBJECTIVES: Limited pediatric data exist examining the trend and predictors of antitissue transglutaminase (atTG) normalization over time in children with celiac disease (CD). We aimed to evaluate time to normalization of atTG in children after CD diagnosis, and to assess for independent predictors affecting this duration. METHODS: A retrospective chart review was completed in pediatric patients with CD diagnosed from 2007 to 2014 at the Stollery Children's Hospital Celiac Clinic (Edmonton, Alberta, Canada). The clinical predictors assessed for impact on time to atTG normalization were initial atTG, Marsh score at diagnosis, gluten-free diet compliance (GFDC), age at diagnosis, sex, ethnicity, medical comorbidities, and family history of CD. Kaplan-Meier survival analysis was completed to assess time to atTG normalization, and Cox regression to assess for independent predictors of this time. RESULTS: A total of 487 patients met inclusion criteria. Approximately 80.5% of patients normalized atTG levels. Median normalization time was 407 days for all patients (95% confidence interval [CI: 361-453]), and 364 days for gluten-free diet compliant patients (95% CI [335-393]). Type 1 diabetes mellitus (T1DM) patients took significantly longer to normalize at 1204 days (95% CI [199-2209], P < 0.001). Cox regression demonstrated T1DM (hazard ratio = 0.36 [0.24-0.55], P < 0.001) and higher baseline atTG (hazard ratio = 0.52 [0.43-0.63], P < 0.001) were significant predictors of longer atTG normalization time. GFDC was a significant predictor of earlier normalization (OR = 13.91 [7.86-24.62], P < 0.001). CONCLUSIONS: GFDC and lower atTG at diagnosis are predictors of earlier normalization. Patients with T1DM are less likely to normalize atTG levels, with longer normalization time. Additional research and education for higher-risk populations are needed.

Computational Analysis of DNA Modifications in Confocal Images.

Computational analysis of digital images provides a robust and unbiased way to compare and investigate the amount (pixel intensity) and spatial distribution of DNA modifications. The DNA modifications in the cells are visualized by fluorescence labeling and the images are captured by confocal microscopy. The key advantage of the confocal over conventional microscope is that it images only a thin optical section around the focal plane of the microscope therefore it can precisely record signals only from the focal plane inside the nucleus. In this chapter, we will describe in detail several analysis methods to visualize and quantify the DNA modification signals including how to investigate codistribution of such signals when using dual labeling.

Mechanism of induction of muscle protein loss by hyperglycaemia.

Treatment of murine myotubes with high glucose concentrations (10 and 25 mM) stimulated protein degradation through the ubiquitin-proteasome pathway, and also caused activation (autophosphorylation) of PKR (double-stranded-RNA-dependent protein kinase) and eIF2alpha (eukaryotic initiation factor 2alpha). Phosphorylation of PKR and eIF2alpha was also seen in the gastrocnemius muscle of diabetic ob/ob mice. High glucose levels also inhibited protein synthesis. The effect of glucose on protein synthesis and degradation was not seen in myotubes transfected with a catalytically inactive variant (PKRDelta6). High glucose also induced an increased activity of both caspase-3 and -8, which led to activation of PKR, since this was completely attenuated by the specific caspase inhibitors. Activation of PKR also led to activation of p38MAPK (mitogen activated protein kinase), leading to ROS (reactive oxygen species) formation, since this was attenuated by the specific p38MAPK inhibitor SB203580. ROS formation was important in protein degradation, since it was completely attenuated by the antioxidant butylated hydroxytoluene. These results suggest that high glucose induces muscle atrophy through the caspase-3/-8 induced activation of PKR, leading to phosphorylation of eIF2alpha and depression of protein synthesis, together with PKR-mediated ROS production, through p38MAPK and increased protein degradation.

The changing frequency of celiac disease diagnosed at the Stollery Children's Hospital.

BACKGROUND: Celiac disease (CD) is recognized as one of the most common and important autoimmune gastrointestinal disorders affecting children. There is evidence that a diagnosis of CD during childhood improves health outcomes. The increasing prevalence of CD is due to increased awareness of the wide range of extraintestinal symptoms associated with CD. OBJECTIVE: To determine whether there has been a temporal increase in the diagnosis of CD associated with an increased diagnosis of children without typical gastrointestinal symptoms at the Stollery Children's Hospital (Edmonton, Alberta). METHODS: Patients with biopsy-proven CD diagnosed at the Stollery Children's Hospital from 1998 to 2007, were identified by retrospective chart review. Baseline and follow-up data, including demographics, symptoms, risk factors, anthropometrics and laboratory investigations, were collected. RESULTS: An increase in the frequency of diagnosis of CD was noted during the study period, particularly from January 2003 onward. Before January 2003, nine children were diagnosed with CD - all with typical symptoms. Between January 2003 and January 2007, inclusive, 149 children were diagnosed with CD, of whom 46% had absent or atypical symptoms. At follow-up, 96% of patients reported improved symptoms, including 53% of individuals who reported being asymptomatic before diagnosis. CONCLUSIONS: In the last four years of the period studied, the number of children diagnosed with CD at Stollery Children's Hospital increased 11-fold. Screening children at risk for CD, and those with atypical presentations, contributed to the increased number of diagnoses. Identification of CD and establishment of lifelong, dietary gluten avoidance during childhood has important health benefits and should be encouraged.

Immunostaining for DNA Modifications: Computational Analysis of Confocal Images.

For several decades, 5-methylcytosine (5mC) has been thought to be the only DNA modification with a functional significance in metazoans. The discovery of enzymatic oxidation of 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) as well as detection of N6-methyladenine (6mA) in the DNA of multicellular organisms provided additional degrees of complexity to the epigenetic research. According to a growing body of experimental evidence, these novel DNA modifications may play specific roles in different cellular and developmental processes. Importantly, as some of these marks (e. g. 5hmC, 5fC and 5caC) exhibit tissue- and developmental stage-specific occurrence in vertebrates, immunochemistry represents an important tool allowing assessment of spatial distribution of DNA modifications in different biological contexts. Here the methods for computational analysis of DNA modifications visualized by immunostaining followed by confocal microscopy are described. Specifically, the generation of 2.5 dimension (2.5D) signal intensity plots, signal intensity profiles, quantification of staining intensity in multiple cells and determination of signal colocalization coefficients are shown. Collectively, these techniques may be operational in evaluating the levels and localization of these DNA modifications in the nucleus, contributing to elucidating their biological roles in metazoans.

A Canadian Study toward Changing Local Practice in the Diagnosis of Pediatric Celiac Disease.

Background. The European Society for Pediatric Gastroenterology, Hepatology and Nutrition endorses serological diagnosis (SD) for pediatric celiac disease (CD). The objective of this study was to pilot SD and to prospectively evaluate gastrointestinal permeability and mucosal inflammation at diagnosis and after one year on the gluten-free diet (GFD). We hypothesized that SD would be associated with similar short term outcomes as ED. Method. Children, 3-17 years of age, referred for possible CD were eligible for SD given aTTG level ≥200 U/mL, confirmed by repeat aTTG and HLA haplotypes. Gastrointestinal permeability, assessed using sugar probes, and inflammation, assessed using fecal calprotectin (FC), at baseline and after one year on a GFD were compared to patients who had ED. Results. Enrolled SD (n = 40) and ED (n = 48) patients had similar demographics. ED and SD groups were not different in baseline lactulose: mannitol ratio (L : M) (0.049 versus 0.034; p = 0.07), fractional excretion of sucrose (%FES; 0.086 versus 0.092; p = 0.44), or fecal calprotectin (FC; 89.6 versus 51.4; p = 0.05). At follow-up, urine permeability improved and was similar between groups, L : M (0.022 versus 0.025; p = 0.55) and %FES (0.040 versus 0.047; p = 0.87) (p > 0.05). FC improved but remained higher in the SD group (37.1 versus 15.9; p = 0.04). Conclusion. Patients on the GFD showed improved intestinal permeability and mucosal inflammation regardless of diagnostic strategy. This prospective study supports that children diagnosed by SD have resolving mucosal disease early after commencing a GFD.

Exploring anthropometric and laboratory differences in children of varying ethnicities with celiac disease.

BACKGROUND: Celiac disease (CD) is a common autoimmune disorder with an increasing prevalence, including in ethnic minorities. OBJECTIVE: To report the frequency of CD diagnosis in ethnic minorities presenting to a Canadian pediatric celiac clinic and to determine whether ethnic differences exist at diagnosis or follow-up. METHODS: Patients with biopsy-proven CD diagnosed at a multidisciplinary celiac clinic between 2008 and 2011 were identified through the clinic database. Data at referral, and six-month and 12-month follow-ups were collected. These included demographics, self-reported ethnicity, symptoms, anthropometrics and laboratory investigations, including serum immunoglobulin antitissue transglutaminase (aTTG). RESULTS: A total of 272 patients were identified; 80% (n = 218) were Caucasian (group 1) and 20% (n = 54) were other ethnicities. South Asians (group 2) comprised 81% (n = 44) of the minority population. No differences in age or sex were found between the two groups. Group 1 patients presented more often with gastrointestinal symptoms (71% versus 43%; P < 0.001), while patients in group 2 presented more often with growth concerns (21% versus 68%; P < 0.001). At diagnosis, serum aTTG level was consistently lower in group 1 compared with group 2 (367 IU/mL versus 834 IU/mL; P = 0.030). Both groups reported symptom improvement at six months and one year. At the end of one year, aTTG level was more likely to be normal in group 1 compared with group 2 (64% versus 29%; P < 0.001). CONCLUSION: Although they represent a minority group, South Asian children comprised a significant proportion of CD patients presenting to a Canadian celiac clinic. South Asian children were more likely to present with growth concerns, which has important implications for timely diagnosis in this population. In addition, the apparent delay in normalization of aTTG levels suggests that careful follow-up and culturally focused education supports should be developed for South Asian children with CD.

Patient and parent satisfaction with a dietitian- and nurse- led celiac disease clinic for children at the Stollery Children's Hospital, Edmonton, Alberta.

OBJECTIVE: To assess patient and parent satisfaction with a primarily nurse- and dietitian-led celiac disease clinic in a tertiary pediatric centre. METHODS: An online survey was sent to families and patients attending the Stollery Children's Hospital's Multidisciplinary Pediatric Celiac Clinic (Edmonton, Alberta) since 2007. The survey focused on clinic attendance, satisfaction with clinic structure, processes, and education and preference for alternatives to the current process. Respondents were asked to rank satisfaction or preference on a five-point Likert scale, with 1 being lowest and 5 being highest. RESULTS: Most satisfaction related to follow-up with serology (4.6) and with a dietitian (4.3). The most preferred changes included either meeting the entire multidisciplinary team after the biopsy (4.7), or meeting with only the dietitian and nurse after the biopsy (4.4). The preferred education resources were the Internet (4.3) and the dietitian (4.2). The mean overall satisfaction score of the Multidisciplinary Pediatric Celiac Clinic was 4.0. CONCLUSIONS: Results of the present survey suggested that patients and families value a multidisciplinary follow-up clinic for children with celiac disease. In particular, feedback based on repeat blood work and regular contact with a dietitian were highly valued. The present survey, outlining the most valued aspects of the clinic, may be useful for service delivery in other regions. In addition, it provides information on how to better support pediatric patients with celiac disease.