Prevalence of celiac disease in a large cohort of young patients with type 1 diabetes.

BACKGROUND: Serological screening for celiac disease (CD) allows the identification of individuals genetically predisposed, as type 1 diabetes mellitus (T1DM). However, the diagnosis is confirmed by intestinal biopsy. The aim was to determine the prevalence of immunoglobulin-A anti-tissue transglutaminase antibodies (IgA-tTG) and CD in a large cohort of young T1DM patients. METHODS: Screening for CD was randomly conducted in 881 T1DM by IgA-tTG and total IgA. Individuals with positive antibodies were referred to endoscopy/duodenal biopsy. RESULTS: The age of the cohort at the screening was 14.3 ± 5.9 years and at T1DM onset was 7.9 ± 4.4 years. The prevalence of positive serology was 7.7%. Median IgA-tTG levels were 117.7 U/mL (interquartile range [IQR] 35.7-131.5 U/mL). Of the 62 duodenal biopsy, CD was diagnosed in 79.0%, yielding an overall prevalence of 5.6%. The mean age of CD patients was 15.6 ± 6.5 years and, at T1DM onset was 6.3 years (4.0-9.9 years). The modified Marsh-Oberhuber histological classification was 22.5% (3a), 36.7% (3b), and 40.8% (3c). In the biopsy-proven patients, T1DM onset occurred at slightly younger ages (6.3 vs 9.7 years, P = 0.1947), gastrointestinal (GI) manifestations, predominantly abdominal pain and distension, were more prevalent (71.4% vs 38.5%, P = 0.027) and higher IgA-tTG titers (128.0 vs 26.3 U/mL, P = 0.0003) were found than in those with negative-biopsies. CONCLUSION: Our results demonstrate the prevalence of 7.7% of IgA-tTG and 5.6% of CD in T1DM patients in South Brazil and, emphasize the importance of the screening in high-risk individuals. Furthermore, the presence of GI manifestations and higher IgA-tTG titers strongly suggest the diagnosis of CD.

Antifibrogenic effect of melatonin in rats with experimental liver cirrhosis induced by carbon tetrachloride.

BACKGROUND AND AIM: Liver diseases are a major public health problem, accounting for a significant number of hospital visits and admissions and an increasing mortality rate. Melatonin (MLT) is a powerful antioxidant molecule that has been shown to be beneficial under various conditions. The objective was to evaluate the effect of MLT on experimental liver cirrhosis induced by carbon tetrachloride (CCl4) in rats. METHODS: Twenty male Wistar rats (230-250 g) were divided into four groups. I: control group (CO); II: CO + MLT; III: CCl4; and IV: CCl4 + MLT. CCl4 was administered intraperitoneally (i.p.) as follows: 10 doses every 5 days, 10 doses every 4 days, and 7 doses every 3 days. MLT was administered i.p. at a dose of 20 mg/kg from the 10th week to the end of the experiment (16th week). RESULTS: In the CCl4 + MLT group, we found that MLT caused a decrease in the level of F2-isoprostanes and NQO1 expression. We also found that MLT reduced the inflammatory process as shown by decreased expressions of NF-KB/p65 and inducible nitric oxide synthase (iNOS) and a smaller amount of inflammatory infiltrate. MLT reduced the expression of transforming growth factor beta1 (TGF-ß1), alpha-smooth muscle actin (α-SMA), and vascular endothelial growth factor (VEGF). Picrosirius staining showed that MLT decreases fibrosis. CONCLUSION: MLT has a potent antifibrogenic effect, modulating the parameters of oxidative stress, angiogenesis, and inflammation.

Genetic variants underlying vitamin D metabolism and VDR-TGFß-1-SMAD3 interaction may impact on HCV progression: a study based on dbGaP data from the HALT-C study.

Vitamin D deficiency is prevalent in liver disease and vitamin D has been shown to decrease hepatic fibrosis through an anti-TGFß-1/SMAD3 effect mediated by the vitamin D receptor. Thus, we hypothesized that genetic variants involved in vitamin D metabolism and/or VDR/TGFß-1/SMAD3 interaction could impact on the progression of chronic HCV. We obtained or imputed genotypes for 40 single nucleotide polymorphisms (SNPs) located in genes implicated in vitamin D metabolism from the HALT-C cohort via dbGaP. The HALT-C study followed 692 chronic HCV patients over 4 years, evaluating clinical outcomes including worsening of fibrosis, hepatic decompensation (gastric/esophageal bleeding, CTP>7, ascites, spontaneous bacterial peritonitis and encephalopathy), development of hepatocellular carcinoma, and liver death. We tested the selected SNPs for association with these outcomes in 681 HALT-C subjects. Eleven SNPs presented tendency towards significance (P<0.05): four SNPs in DHCR7 related to with hepatic decompensation (rs4944957, rs12800438, rs3829251 and rs4945008); two in GC to worsening of fibrosis and liver death (rs7041 and rs222020); two in CYP2R1 to ascites and hepatocellular carcinoma (rs7116978 and rs1562902); two in VDR to gastric/esophageal bleeding and hepatocellular carcinoma (rs4516035 and rs2239186); and one in SMAD3 to worsening of fibrosis and encephalopathy (rs2118610). Only rs1800469 in TGFB1 was statistically associated with hepatic decompensation after Bonferroni's correction (P<0.00125). In conclusion, rs1800469 in TGFB1 was associated to hepatic decompensation in chronic hepatitis C, while the other 11 described polymorphisms must be evaluated in a larger cohort to determine the possible role of vitamin D in hepatitis C.

Brain-Derived Neurotrophic Factor in Children and Adolescents with Cirrhosis Due to Biliary Atresia.

BACKGROUND: The nutritional status in patients with cirrhosis is not so easy to assess properly. Considering the relationship between brain-derived neurotrophic factor (BDNF) and energy homeostasis, the main aim of this study was to evaluate the concentration of BDNF in children and adolescents with cirrhosis due to biliary atresia (BA) and correlate it with their nutritional status. METHODS: Fifty-three children and adolescents with cirrhosis due to BA and 33 healthy controls were enrolled in this study. Nutritional status was evaluated using anthropometric parameters, and serum BDNF was measured by ELISA. Spearman coefficient was used to evaluate the correlation between variables. RESULTS: In the cirrhosis group, 28.8% were undernourished and in the control group, 100% were well-nourished. BDNF median values for the control and cirrhosis group were 28.5 and 9.0 pg/ml respectively. BDNF and platelets were positively associated with both Standard Deviation Score (SDS) for height-for-age ratio and SDS for triceps skinfold thickness-for-age ratio. CONCLUSIONS: Considering these associations, BDNF may be an indirect biomarker of nutritional status in children and adolescents with chronic liver disease. Further studies must be conducted to clarify the role of BDNF in this population.

Angiopoietin 1 and angiopoietin 2 are associated with medial thickening of hepatic arterial branches in biliary atresia.

BACKGROUND: Biliary atresia (BA) is an infantile disorder characterized by progressive sclerosing cholangiopathy leading to biliary obstruction. First-line treatment of BA is hepatoportoenterostomy, the prognosis of which is related to age at surgery and to histological variables such as extent of fibrosis and ductular reaction. Hepatic arterial medial thickening (MT) suggests an arteriopathy in BA pathogenesis. We evaluated the expression of angiopoietin (ANGPT)/tyrosine kinase with immunoglobulin-like and epidermal growth factor-like domains 2 (TIE2) system in liver samples obtained from patients with BA, correlating it with MT, variables associated with disease severity, and postoperative prognosis. METHODS: ANGPT1, ANGPT2, and TIE2 expression levels were assessed by quantitative PCR in liver samples obtained from BA patients (n = 23) at portoenterostomy and age-matched infants with intrahepatic cholestasis (IHC; n = 7). Histological variables were morphometrically assessed. RESULTS: ANGPT1 and ANGPT2 were overexpressed in BA in comparison with IHC (P = 0.024 and P = 0.029, respectively). In BA, ANGPTs expression was positively correlated with MT (ANGPT1: rs = 0.59, P = 0.013; ANGPT2: rs = 0.52, P = 0.032), not with the variables associated with disease severity. TIE2 and ANGPTs expression levels were negatively correlated (ANGPT1: rs = -0.73, P < 0.001; ANGPT2: rs = -0.54, P = 0.007). CONCLUSION: In BA, there is overexpression of both ANGPT1 and ANGPT2, which is correlated with MT but not with age at portoenterostomy or with the histological variables associated with disease severity at the time of procedure.

Blood collection for biochemical analysis in adult zebrafish.

The zebrafish has been used as an animal model for studies of several human diseases. It can serve as a powerful preclinical platform for studies of molecular events and therapeutic strategies as well as for evaluating the physiological mechanisms of some pathologies. There are relatively few publications related to adult zebrafish physiology of organs and systems, which may lead researchers to infer that the basic techniques needed to allow the exploration of zebrafish systems are lacking. Hematologic biochemical values of zebrafish were first reported in 2003 by Murtha and colleagues who employed a blood collection technique first described by Jagadeeswaran and colleagues in 1999. Briefly, blood was collected via a micropipette tip through a lateral incision, approximately 0.3 cm in length, in the region of the dorsal aorta. Because of the minute dimensions involved, this is a high-precision technique requiring a highly skilled practitioner. The same technique was used by the same group in another publication in that same year. In 2010, Eames and colleagues assessed whole blood glucose levels in zebrafish. They gained access to the blood by performing decapitations with scissors and then inserting a heparinized microcapillary collection tube into the pectoral articulation. They mention difficulties with hemolysis that were solved with an appropriate storage temperature based on the work Kilpatrick et al. When attempting to use Jagadeeswaran's technique in our laboratory, we found that it was difficult to make the incision in precisely the right place as not to allow a significant amount of blood to be lost before collection could be started. Recently, Gupta et al. described how to dissect adult zebrafish organs, Kinkle et al. described how to perform intraperitoneal injections, and Pugach et al. described how to perform retro-orbital injections. However, more work is needed to more fully explore basic techniques for research in zebrafish. The small size of zebrafish presents challenges for researchers using it as an experimental model. Furthermore, given this smallness of scale, it is important that simple techniques are developed to enable researchers to explore the advantages of the zebrafish model.

Amplification of bacterial DNA does not distinguish patients with ascitic fluid infection from those colonized by bacteria.

OBJECTIVE: To evaluate 16S ribosomal RNA (rRNA) gene amplification to diagnose spontaneous bacterial peritonitis (SBP). PATIENTS AND METHODS: According to a retrospective protocol, 31 patients with portal hypertensive ascites (serum to ascites albumin gradient > or = 1.1 g/dL) were studied. Ascitic fluid was analyzed as follows: Gram stain, aerobic and anaerobic cultures, polymorphonuclear cell count, and biochemical tests. Bacterial DNA was detected by polymerase chain reaction. RESULTS: There were 8 episodes of SBP and 4 episodes of bacterascites (BA). Culture was positive in 4 of 8 cases of SBP and bacterial DNA was positive in 7 of 8 cases of SBP. Bacterial DNA was positive in 3 of 4 cases of BA and in 8 of 28 cases of culture-negative non-neutrocytic ascites (CNNNA). The PELD score, serum to albumin ascites gradient, and mortality showed no statistical difference between patients with CNNNA and the result of the bacterial DNA analysis. CONCLUSIONS: Although the 16S rRNA gene amplification was better than culture to diagnose SBP, bacterial DNA does not seem to allow a distinction between ascites infection and ascites colonization.

Infected and noninfected ascites in pediatric patients.

OBJECTIVES: To determine the prevalence of spontaneous bacterial peritonitis, ascites with bacterial infection and noninfected ascites in pediatric patients with portal hypertensive ascites and to compare the clinical and laboratory features of infected and noninfected ascites. METHODS: Forty-one episodes of portal hypertensive ascites (serum-ascites albumin gradient >1.1 g/dL) in 31 patients were studied. Median age was 2.9 years. Twenty-four (77.4%) patients were cirrhotic and 20 (83.3%) were classified as Child-Pugh C. Median pediatric end-stage liver disease score was 18.5. The following ascites features were assessed: polymorphonuclear neutrophil cell count, cytology, pH, concentration of glucose, lactic dehydrogenase, total protein and albumin, Gram stain and bacteriological culture. Blood was sampled for complete blood count, coagulation studies, liver and renal function tests. Groups were compared by Mann-Whitney and chi tests (P < 0.05). RESULTS: Noninfected ascites were observed in 29 of 41 samples, spontaneous bacterial peritonitis in eight of 41 and ascites with bacterial infection in four of 41. The most prevalent clinical features were fever, voluminous ascites and encephalopathy, but there were no significant differences in the clinical features of the groups. All patients with infected ascites were cirrhotic. There was no statistical difference in Child-Pugh or pediatric end-stage liver disease status between patients with infected and noninfected ascites. Culture of ascetic fluid was positive in four of eight cases of spontaneous bacterial peritonitis. Gram-negative rods were the most prevalent bacteria cultured. Except for serum albumin, no statistical differences in biochemical markers were observed between patients with infected and noninfected ascites. CONCLUSIONS: The prevalence of infected ascites was 29.2%. With the exception of serum albumin, there were no differences in the clinical and biochemical features of patients with infected ascites and noninfected ascites.

Chronic functional constipation in adolescents: clinical findings and motility studies.

OBJECTIVE: To evaluate the clinical findings and colonic transit time in adolescents with chronic functional constipation. METHODS: Forty-eight consecutive adolescents with chronic functional constipation referred to the Gastroenterology Service at the Hospital de Clínicas de Porto Alegre, Brazil were studied. Clinical parameters were assessed using a questionnaire. Total and segmental colonic transit time were measured with radiopaque markers. RESULTS: Mean age at first visit was 14 +/- 2 years, and age at onset of constipation was 6 +/- 4 years; 90% of patients depended on laxatives, and 86% on intermittent enemas; 76% had a family history of constipation. There was no statistical difference in the amount of daily fiber ingested by patients and controls. Measurements of colonic transit time revealed that 60% of patients had slow transit constipation, 13% had pelvic floor dysfunction, 10% had slow transit constipation associated with pelvic floor dysfunction, and 17% had a normal colonic transit time. Decreased frequency of evacuation and palpable abdominal fecal mass were significantly associated with slow transit constipation. CONCLUSIONS: Functional constipation in adolescence consists of a heterogeneous group of colonic functional disorders. The identification of these different functional disorders in adolescents will guide specific treatment, which may prevent the progression of this symptom into adult life.

[Blood levels of vitamin D in children and adolescents with chronic cholestasis]. / Níveis plasmáticos de vitamina D em crianças e adolescentes com colestase.

OBJECTIVE: To verify blood levels of vitamin D in patients with chronic cholestasis, and relate them to nutritional status, length of time since the onset of cholestasis and use of vitamin supplement. METHODS: Controlled cross-sectional study with chronic cholestasis as study factor and blood levels of vitamin D as outcome. The study included patients aged between 4 months and 18 years, who were cared for at the Pediatric Gastroenterology Unit of Hospital de Clínicas de Porto Alegre. Controls were eutrophic children in the same age range. Blood was collected for radioimmunoassay. Anthropometric analyses were performed, as was determination of the length of time since the onset of cholestasis and use of vitamin supplement. RESULTS: 22 patients and 17 controls were evaluated. Average vitamin D level in patients was 13.7 -/+ 8.39 ng/ml, compared to 25.58 -/+ 16.73 ng/ml in controls (p = 0.007). Prevalence of hypovitaminosis D in patients was 36%. Median of period of time since the onset of cholestasis was 1 year with variation of 6 months to the 25th percentile and 3.9 years to the 75th percentile. Anthropometric evaluation (NCHS) showed 36% of malnutrition by weight and 41% by height. Anthropometric evaluation according to Z score showed 33.3% and 23.8% prevalence of malnutrition for the criteria height/age and weight/age, respectively. The evaluation of weight regarding height did not show values below two standard deviations. No relationship was found between nutritional state, use of oral vitamin supplement and blood levels of vitamin D. CONCLUSIONS: Blood levels of vitamin D in patients with cholestasis were lower than those of controls, but were not related to nutritional status, period of time since the onset of cholestasis or use of vitamin supplement.

[Digital morphometric and stereologic analysis of small intestinal mucosa in well-nourished and malnourished children with persistent diarrhea]. / Estudo morfométrico e estereológico digital da mucosa do intestino delgado de crianças eutróficas e desnutridas com diarréia persistente.

OBJECTIVE: To test the hypothesis that the proximal small intestines of children with persistent diarrhea present morphometric and stereologic changes proportional to their nutritional status, using microscope images stored in a computer. METHODS: Cross-sectional study with 65 pediatric patients, whose ages ranged from 4 months to 5 years, with persistent diarrhea for over 14 days. The nutritional assessment was performed according to the z-scores for weight/age (W/A), weight/height (W/H) and height/age (H/A) ratios, divided into: well-nourished = z > or =2SD and malnourished = z<2SD; well-nourished = z > or =2SD, nutritional risk = z<1SD and malnourished = z<-2SD; and continuously, in descending order, using the NCHS charts. After obtaining the computer images using the software Scion Image, villous height, crypt depth, mucosal thickness, total mucosal thickness, and villous/crypt ratio were measured in the fragments of the small intestinal mucosa, enlarged 100 times. When images were enlarged 500 times, enterocyte height, nuclear height and brush-border height were measured. Stereologic analysis was performed using cycloid arcs. RESULTS: For W/A, W/H and H/A z-scores, divided into two nutritional status categories, no statistically significant difference was observed in regard to villous height, crypt depth, mucosal thickness, total mucosal thickness and villous/crypt ratio. Enterocyte height presented the most significant difference between well-nourished and malnourished groups, for W/A and W/H ratios, with a 500x enlargement, although this difference was not statistically significant. When z-scores were subdivided into three nutritional status categories, a digital morphometric analysis showed a statistically significant difference for villous/crypt ratio between the well-nourished and slightly malnourished group and the well-nourished and mild to severe malnourished group (p=0.048). The villous/crypt ratio was higher among well-nourished children. Using the Spearman coefficient, the variables enterocyte height, height of enterocyte nucleus and brush-border height presented a clear association with the W/A ratio (r=0.25; p=0.038), W/H ratio (r=0.029; p=0.019). The height of the enterocyte and the brush-border height were associated with W/H ratio. CONCLUSION: The observed associations between nutritional status and the analyzed small intestinal mucosa variables showed a positive correlation with patients' weight. Although these associations were of a slight to moderate magnitude, we observed a tendency of enterocyte size reduction, as well as a reduction in the size of its nucleus and brush-border, as the level of malnutrition increases.

[Prevalence of hepatitis A antibody in children and adolescents with chronic liver disease]. / Prevalência do anticorpo da hepatite A em crianças e adolescentes com hepatopatia crônica.

OBJECTIVE: To evaluate the prevalence of Hepatitis A virus (HAV) in children and adolescents with chronic liver disease in a unit of pediatric hepatology. METHODS: Between May 1999 and February 2001, we studied the prevalence of anti-HAV in 60 children and adolescents with chronic liver disease, aged between 1 and 16 years, from the Unit of Pediatric Hepatology of the Hospital de Clínicas de Porto Alegre. The total anti-HAV was determined by a commercially available competitive ELISA method (Abbott), and compared with age, sex, race, etiologic diagnosis and family income of each patient. RESULTS: A one-year old child was not included in the study because she presented twice with undetermined anti-HAV results. Among the other 59 patients, 14 (24%) presented a positive result of total anti-HAV. The ages of test-positive subjects varied between 1 and 16 years old (mean=7.7 years, median=8.5). The differences between positive and negative groups in relation to age, sex and race were not statistically significant. Family income was lower in anti-HAV positive patients, but this difference was not significant. The differences between the etiologies of liver diseases were probably more related to the age than to the etiologies of the diseases. CONCLUSIONS: In the studied population, the majority (76%) of children and adolescents with chronic liver disease are susceptible to hepatitis A virus infection and, consequently, they could present a more severe disease or even fulminant hepatitis A. We strongly suggest that these subjects receive Hepatitis A inactivated vaccine.