Visceral and subcutaneous adipose tissue as markers of local and systemic inflammation: a comparison between celiac and obese patients using MRI.

BACKGROUND: Celiac disease (CD) is a systemic inflammatory disease, which primarily affects the gastrointestinal tract. It has been recently demonstrated that adipose-tissue infiltration by proinflammatory immune cells causes a chronic low-grade inflammation in obese patients. Magnetic resonance imaging (MRI) has already proved to be useful in evaluation of inflammatory states. The aim of the present study was to determine whether alterations of visceral and subcutaneous adipose tissue, identified with MRI, could serve as markers of local and systemic inflammation in patients with CD. METHODS: A pilot study was conducted comparing alterations in visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) in CD patients vs obese patients and healthy controls. Fifty patients were enrolled and assigned to one of the following groups: Group A: 11 active CD patients; Group B: 11 CD patients in remission; Group C: 16 obese patients; Group D: 12 healthy controls. A 3-T MRI unit was used and T2-weighted TSE images of VAT and SAT were obtained in specific regions of interest. Serum cytokine concentrations (TNF-α, IL-6, adiponectin, leptin, IL-2, IFN-γ) were determined. RESULTS: There was a significant difference in VAT T2 relaxation time between Group A and B (p < 0.001), A and D (p < 0.01), B and C (p < 0.001). There was a statistically significant difference in SAT T2 relaxation time between Group A and B (p < 0.001), A and C (p < 0.05), A and D (p < 0.001), B and C (p < 0.01). In addition, VAT/SAT T2 relaxation time ratio showed a statistically significant difference between Group A and C (p < 0.05) and between Group B and C (p < 0.01). Only TNF-α and IL-6 significantly correlated with both VAT and VAT/SAT ratio in active CD. CONCLUSIONS: MRI showed similar increased visceral inflammatory signals in patients with active CD and obese patients. However, subcutaneous inflammatory signals were higher in active CD than in all the other groups. These data show that there is a systemic inflammatory state in active CD, whereas chronic inflammation appears confined to VAT in obesity. These data were only partially confirmed by serological cytokine profiles, which showed less specificity than MRI.

Nuclear fluorescence serum reactivity on monkey oesophagus: a new antibody for the follow-up of coeliac disease?

We have identified previously a nuclear fluorescence reactivity (NFR) pattern on monkey oesophagus sections exposed to coeliac disease (CD) patients' sera positive for anti-endomysium antibodies (EMA). The aim of the present work was to characterize the NFR, study the time-course of NFR-positive results in relation to gluten withdrawal and evaluate the potential role of NFR in the follow-up of CD. Twenty untreated, 87 treated CD patients and 15 healthy controls were recruited and followed for 12 months. Their sera were incubated on monkey oesophagus sections to evaluate the presence of NFR by indirect immunofluorescence analysis. Duodenal mucosa samples from treated CD patients were challenged with gliadin peptides, and thus the occurrence of NFR in culture supernatants was assessed. The NFR immunoglobulins (Igs) reactivity with the nuclear extract of a human intestinal cell line was investigated. Serum NFR was present in all untreated CD patients, persisted up to 151 ± 37 days from gluten withdrawal and reappeared in treated CD patients under dietary transgressions. Serum NFR was also detected in two healthy controls. In culture supernatants of coeliac intestinal mucosa challenged with gliadin peptides, NFR appeared before EMA. The Igs responsible for NFR were identified as belonging to the IgA2 subclass. The NFR resulted differently from EMA and anti-nuclear antibodies, but reacted with two nuclear antigens of 65 and 49 kDa. A new autoantibody, named NFR related to CD, was described. Furthermore, NFR detection might become a valuable tool in monitoring adherence to a gluten-free diet and identifying slight dietary transgressions.

Antitissue transglutaminase antibodies in acute coronary syndrome: an alert signal of myocardial tissue lesion?

BACKGROUND AND AIM: Antitransglutaminase, previously considered identical to antiendomysial in coeliac sprue (CS), have been reported in end-stage heart failure. To clarify the above-mentioned data, we evaluated these antibodies in a cohort of cardiological patients with respect to troponin I, creatine kinase (CK), MB fraction creatine kinase (CK-MB mass) and myoglobin. METHODS: Forty-one patients with acute coronary syndrome (ACS), 39 with dilated cardiomyopathy (DCM), 45 with CS and 58 blood donors (BDs) were evaluated. Antitransglutaminase and antiendomysial antibodies were tested in serum of the patients being studied. RESULTS: High-positive antitransglutaminase values were found in CS, whilst low-positive values were also found in ACS and DCM. In patients at the second ACS, antibody levels were higher than in those at the first cardiac event. In patients with infarct Q, antitransglutaminase were higher than those in infarct non-Q, in which antibody levels were higher than those in unstable angina. A correlation between antitransglutaminase and troponin I, CK, CK-MB mass and myoglobin was found. Finally, antibody levels rose to reach a peak at 30 days from the cardiac event, whereas after further 150 days, approached the values of BDs. Antiendomysial were detectable only in CS. CONCLUSIONS: Data highlight that antitransglutaminase can occur in cardiological patients, and that these antibodies are related to the severity/extent of the myocardial tissue lesion. This feature suggests a loss of specificity for antitransglutaminase in CS. Furthermore, the possibility of employing these antibodies in the long-term follow-up of ACS, could become an object of interesting discussion.

Detection of anti-endomysial and anti-tissue transglutaminase autoantibodies in media following culture of oral biopsies from patients with untreated coeliac disease.

BACKGROUND AND AIMS: Coeliac disease is an autoimmune disorder characterised by high levels of anti-endomysial and anti-tissue transglutaminase autoantibodies in sera and media of cultured intestinal mucosa biopsies from affected patients. In this study, we wished to investigate whether anti-endomysial and anti-tissue transglutaminase antibodies can also be detected in culture media of oral mucosa specimens, and whether the mouth can be used as an area of immunological testing for coeliac disease. METHODS: Small intestine and cheek biopsy samples taken from 16 patients with active coeliac disease and from 11 controls were cultured in vitro for 48 h at 37 degrees C in presence of medium alone. Anti-endomysial and anti-tissue transglutaminase were detected in sera and in supernatants of these cultured biopsy samples by indirect immunofluorescence and enzyme immunoassay (EIA), respectively. RESULTS: Anti-endomysial and anti-tissue transglutaminase were positive in sera of 15/16 coeliac disease patients. Culture media of intestinal mucosa samples from 14/16 coeliac disease patients were anti-endomysial positive, while the same antibodies were positive in supernatants of cultured oral mucosa samples from 15/16 coeliac disease patients. Anti-tissue transglutaminase were positive in both intestinal and oral culture media of 15/16 coeliac disease patients. Neither anti-endomysial nor anti-tissue transglutaminase were found in sera or in culture supernatants of both intestinal and oral biopsy samples from 11 controls. CONCLUSIONS: Our study suggests a new immunological site to detect the pathognomonic autoantibodies of coeliac disease and confirms that the mouth is involved in this illness.

Impact of gluten-free diet on quality of life in celiac patients.

BACKGROUND AND AIM: Celiac disease (CD) is a common gluten-related disorder, whose only treatment is a gluten-free diet (GFD). Since a unique view on psychological consequences of a GFD still lacks, our aim was to assess the quality of life (QoL) and the depression state in symptomatic CD patients after GFD. Socio-demographic features were considered. PATIENTS AND METHODS: 210 adult CD patients were recruited and divided into 3 groups : 70 newly diagnosed patients (​Group A​),70 patients who have been on GFD for 6-12 months (​Group B​), and 70 patients who have been on GFD for more than 12 months (​Group C​). We recruited 210 healthy controls (​Group D). Psychological General Well-Being Index (PGWBI) and Beck Depression Inventory (BDI) questionnaires were administered. Each group was evaluated according to age, gender and school ranking. RESULTS: Groups A ​and B showed lower PGWBI scores compared with both Group C ​and D (p <0.001 for each comparison). Moreover, Groups A and B showed higher BDI scores compared with both ​Group C ​and D (p <0.001 for each comparison).Women, the elderly and the poorly educated seemed to suffer more psychological stress. CONCLUSION: GFD induces an improvement of well-being and a decrease of depression state after 12 months of strict GFD. Negative psychological implications were observed only in specific risk categories. (Acta gastroenterol. belg., 2016, 79, 447-453).

Abdominal CT findings may suggest coeliac disease.

BACKGROUND: Coeliac disease is the most common gastrointestinal immunological disorder in the western countries. Many adult patients present non-specific symptoms and signs of malabsorption such as chronic diarrhoea, anaemia, weight loss and abdominal distention. In non-specific and doubtful conditions, computed tomography is often the first medical examination performed. In a clinical practice, a critical review of computed tomography signs is therefore mandatory. AIMS: To evaluate the abdominal computed tomography findings, which are useful to suggest the presence of coeliac disease in adult patients. PATIENTS AND METHODS: The computed tomography studies of 28 coeliac patients were reviewed, looking for any intestinal and extraintestinal abnormality. The computed tomography findings evaluated were: abnormalities of intestinal fold pattern, bowel dilatation, fluid and air excess, duodenal abnormalities, intestinal intussusception, bowel wall thickening, lymphadenopathy, ascites, intestinal stenosis, mesenteric vascular changes. The abdominal computed tomography of a group of 30 normal subjects was also analysed. RESULTS: Intestinal fold pattern abnormalities were seen in 23/28 patients. Intestinal dilatation was seen in 21/28. Fluid excess in 18/28 and lymphadenopathy was seen in 12/28 patients; engorgement of mesenteric vessels in 7/28. Bowel wall thickening was observed in 6/28 patients and transient intussusception was observed in 6/28 patients. Increased air content within the bowel in 4/28 and ascites in 2/28 patients. Bowel dilatation together with fluid excess was observed in 18/28 patients. None of the above mentioned abnormalities abnormalities were seen in normal subjects. CONCLUSIONS: Data of the present study show that several abdominal computed tomography findings may be seen in coeliac adult patients; these findings should be taken into consideration with a high in level of suspicion by radiologists, to avoid diagnostic delay and unnecessary diagnostic and therapeutic procedures.

Immunologic evidence of no harmful effect of oats in celiac disease.

BACKGROUND: It was recently shown that antiendomysial antibodies (EMAs), which are highly sensitive and specific for celiac disease, are produced by intestinal mucosa. Furthermore, EMAs were detected previously in supernatant fluid from cultured duodenal mucosa specimens collected from untreated celiac disease patients and in culture media of biopsy specimens collected from treated celiac disease patients after an in vitro challenge with gliadin. Moreover, it was recently shown in vivo that oats are not toxic to celiac disease patients, suggesting the safety of oats in a gluten free-diet. OBJECTIVE: The objective was to better define the controversial role of oats in celiac disease to determine whether oats can be safely included in a gluten-free diet. DESIGN: We used an in vitro model to test whether oats induce EMA production in supernatant fluid from cultured duodenal mucosa specimens collected from 13 treated celiac disease patients. The biopsy specimens were cultured with and without peptic-tryptic digest (PT) of gliadin and avenin (from oats) and in medium alone. Samples from 5 of the 13 patients were cultured with the C fraction of PT-avenin. Indirect immunofluorescence was used to detect EMAs. RESULTS: EMAs were detected in specimens from all 13 patients after the challenge with gliadin but not after culture in medium alone. By contrast, no EMAs were detected in any of the specimens cultured with PT-avenin and its C fraction. CONCLUSIONS: Because the in vitro challenge with PT-avenin and its C fraction did not induce EMA production in treated celiac disease patients, it appears that oats have no harmful effect on celiac disease. Therefore, oats can be safely included in a gluten-free diet.

Evidence of a selective epitope loss of anti-transglutaminase immunoreactivity in gluten-free diet celiac sera: a new tool to distinguish disease-specific immunoreactivities.

The aim of the present study was to evaluate the epitope specific humoral human tissue transglutaminase (tTG) immunoreactivity against 3 different human recombinant tTG constructs [(full-length tTG (a.a. 1-687), tTG (a.a. 227-687); tTG (a.a. 473-687)] before and after the introduction of a gluten-free diet (GFD). To this end, sera from 64 celiac disease (CD) subjects on a gluten-containing diet (44 f, 20 m) and after 0.6 +/- 0.3 years and 2.1 +/- 1.3 years of GFD were studied using a quantitative radioimmunoprecipitation assay. All 64 CD patients at diagnosis were full-length anti-tTG (a.a. 1-687)Ab positive. These Abs significantly decreased in frequency and titer after 6 months and 2 years of GFD. However, at low titers, 64.1% (41/64) of CD patients were still fl-tTG (a.a. 1-687)Ab positive after 2 years of GFD. At disease diagnosis, 70.3% (45/64) of the CD patients had Abs directed against fragments (227-687) and/or (473-687) of the tTG protein. This percentage, after 2 years of GFD, significantly decreased to 18.7%, whereas almost 50% of GFD patients had no tTG (227-687) and tTG (473-687) fragment reactivity, but only persistent, low-titer full-length tTG (1-687)Abs. We suggest that the selective loss of immunoreactivity against tTG (227-687) and tTG (473-687) fragments in CD patients with a GFD, could be due to quantitative decrease of autoreactivity driven by tTG-gliadin interaction underlying celiac disease pathogenesis.

Anti-endomysial antibody of IgG1 isotype detection strongly increases the prevalence of coeliac disease in patients affected by type I diabetes mellitus.

A strong association between type 1 insulin-dependent diabetes mellitus (IDDM1) and coeliac disease (CD) is well documented, but it is known that prevalence values are underestimated. Serum anti-endomysial antibodies (EMA), considered diagnostic for CD because of their high sensitivity and specificity, belong to the IgA class, but the existence of EMA of IgG1 isotype in the presence or absence of IgA deficiency was reported. In order to re-evaluate the occurrence of CD in IDDM1 patients we performed a screening in IDDM1 patients using EMA of both isotypes. Ninety-four adults affected by IDDM1 (unaffected by CD before enrolling) were enrolled and 83 blood donors as controls. All subjects were on a gluten-containing diet. Histology and biopsy culture were performed. EMA IgA and IgG1 in sera and culture supernatants were detected. Serum EMA were positive in 13 of 94 IDDM1 patients (13.8%). Six of 13 presented IgA-EMA, seven of 13 presented IgG1-EMA. No EMA were found in the control population. Total intestinal atrophy was found in all six patients with serum IgA-EMA and in five of seven with serum IgG1-EMA. Diagnosis of CD was confirmed by histology and organ culture in all 13 patients with serum EMA. The prevalence of CD in the patients affected by IDDM1 was 6.4% for IgA-EMA-positive and 7.4% for IgG1-EMA-positive patients. We confirmed the prevalence of CD in the IDDM1 population obtained with IgA-EMA screening only (6.4%). This prevalence value increases dramatically to 13.8% when IgG1-EMA are also used in the screening. We conclude that IgG1-EMA should also be sought whenever an IDDM1 patient undergoes screening for CD.

Blockage of T-cell costimulation inhibits T-cell action in celiac disease.

BACKGROUND & AIMS: Celiac disease is an exemplary model of T cell-mediated pathology. Therefore, therapeutic approaches that target T cells may successfully control this disease. CTLA-4 immunoglobulin (CTLA-4Ig) can inhibit T-cell activation by blocking the engagement of CD28. We took advantage of this tool to define the pathogenic role of gliadin-specific T cells in the induction of celiac disease. METHODS: Duodenal biopsy specimens from 7 treated celiac patients were challenged in vitro with gliadin and CTLA-4Ig or CD40-Ig. After 24 hours, the biopsy specimens were analyzed for the presence of characteristic modifications induced by gliadin challenge. RESULTS: CTLA-4Ig down-regulated the expression of CD25, intercellular adhesion molecule 1, interleukin 2, and interferon gamma (stained lamina propria mononuclear cells/mm2; P < 0.05) induced by gliadin challenge, caused apoptosis of gliadin-specific T cells (apoptotic T cells/mm2; P < 0.05), and inhibited the production of antiendomysial antibody (P < 0.01). However, it did not control intraepithelial T-cell migration (P = NS) and Fas expression by enterocytes. Conversely, CD40-Ig only controlled production of antiendomysial antibody. CONCLUSIONS: In an organ culture model, CTLA-4Ig controls many but not all of the immunologic features of celiac disease.

Celiac disease diagnosis in misdiagnosed children.

Antiendomysial antibodies (EMA) are today considered the most sensitive and specific serological marker of celiac disease (CD). The aim of the present study was to assess the occurrence of EMA of IgG isotype in EMA IgA negative children with clinical suspicion of malabsorption and their relationship with CD. Serum EMA IgG1 determination was performed on 30 EMA IgA negative children with clinical suspicion of CD. Total serum IgA levels were further investigated. Sixty children with gastroenterological diseases other than CD were used as control disease patients and 63 healthy children were evaluated as the control group. Eighteen out of 30 children in the study showed EMA IgG1 positivity in sera and a villous height/crypt depth ratio <3:1 as index of intestinal atrophy. It is noticeable that a selective IgA deficiency was present in only 9 of 18 EMA IgG1 positive children. In addition, clinical symptoms, EMA IgG1, and mucosal atrophy disappeared after 8-10 mo on a gluten-free diet. Neither EMA IgA nor EMA IgG1 were detected in the children in the control groups. The other 12 children in study group showed no histologic abnormalities and were EMA IgG1 negative. In this study, we reveal a group of EMA IgG1 CD children without IgA deficiency. The diagnosis was based on the presence of gluten-dependent typical serological and histologic features of CD. Our data suggest that EMA IgG1 determination could be a new tool in the diagnostic workup of CD, useful in avoiding possible misdiagnosis.

Celiac disease-associated transglutaminase autoantibody target domains at diagnosis are age and sex dependent.

The contribution of age and/or sex to the transglutaminase (tTG) autoantibody response in celiac disease (CD) is not known. To gain insights into transglutaminase humoral autoimmunity at CD diagnosis, our aim was to characterize the autoimmune response against three tTG constructs [(full-length tTG(a.a.1-687), tTG(a.a.227-687), and tTG(a.a.473-687)] and to investigate into its relationship with CD patients' age and sex. One hundred seventy-five newly diagnosed CD patients (115 females and 60 males), subdivided into different groups according to age and sex, were studied using a serum 35S-radioimmunoassay. We found that among full-length tTG autoantibody-positive CD subjects (175/175), 50.9% (89/175) and 83.4% (146/175) had autoantibodies against tTG(227-687) and tTG(473-687) domains, respectively. Female patients of less than 4 years expressed tTG(227-687)Abs in significantly higher percentage and mean autoantibody titers vs. all other groups investigated, and tTG(473-687)Abs in significantly higher titers with respect to adult female patients. Our data identify a subset of CD patients showing a strong humoral tTG immunoreactivity at diagnosis, thus suggesting that age and sex influence the anti-tTG autoantibody response.

Quantitative analysis of stool losses in adult celiac disease: use of near-infrared analysis reconsidered.

BACKGROUND: An attempt has been made to establish whether near-infrared stool analysis is more suitable for quantifying malabsorption than the traditional stool fat analysis. A group of celiac disease (CD) patients was used as index population. METHODS: Stool fat, nitrogen, and water were measured with near-infrared analysis of 1- and 3-day stool collections in 96 celiac disease patients on a free diet (in 39 also on gluten-free diet) and in 96 matched controls and 14 patients with latent CD. RESULTS: The fecal output of fat, nitrogen, and water was significantly increased in free-diet CD, whereas their percentage content was only slightly modified compared with controls. None of the variables under consideration differed significantly between the 24-h and 72-h stool specimens. CONCLUSION: Our data show that the high value of fecal fat, nitrogen, and water, in celiac disease, are mainly due to the fecal weight, whereas the percentage composition of stool does not offer additional diagnostic information. Furthermore, 3-day stool collection is not necessary to confirm or rule out malabsorption in most patients. Near infrared analysis of 24-h specimens is time- and cost-effective and may increase the use of stool analysis and be usefully employed to monitor the clinical follow-up of patients with chronic diarrhea.

Identification of a new coeliac disease subgroup: antiendomysial and anti-transglutaminase antibodies of IgG class in the absence of selective IgA deficiency.

OBJECTIVE: The aim of the present study was to increase the sensitivity of the antiendomysial antibody (EMA) test by evaluating also EMAs of IgG1 isotype. DESIGN AND SUBJECTS: Over the last 2 years, serum EMAs IgA and IgG1 were determined in 1399 patients, referred to our gastrointestinal unit due to clinical suspicion of malabsorption. Serum anti-tissue transglutaminase (tTG) antibodies IgA and IgG, as well as total IgA levels, were also investigated. Furthermore, EMAs IgA and IgG1 were evaluated in biopsy culture supernatants. Biopsy specimens were also admitted to histological and immunohistochemical evaluation. Twenty-six patients with gastroenterological disease other than coeliac disease (CD) were used as a disease control group. Ninety-nine blood donors were used as a healthy control group. RESULTS: Diagnosis of CD was based on histological findings in the 110/1399 patients showing EMA IgA positivity, and in a further 56/1399 patients presenting both EMA IgA and IgG1 positivity in sera as well as in culture supernatants. Of the remaining 1233 EMA IgA-negative patients, 60 showed only EMA IgG1 positivity both in sera and in culture supernatants. It is noteworthy that anti-tissue transglutaminase antibodies IgG (anti-tTG) were positive in all 60 EMA IgG1-positive patients as well. By contrast, a selective IgA deficiency was found in only 11 out of the 60 EMA IgG1-positive patients. Villous height/crypt depth ratio was < 3:1 in 38 of the 60 EMA IgG1-positive patients (63.3%), whilst overexpression of ICAM-1 and CD25 was observed in all these patients. CONCLUSIONS: In this study, we observed a group of CD patients who were EMA IgG1-positive even in the absence of EMA IgA positivity and IgA deficiency. The diagnosis was based on the finding of the gluten-dependent clinical and histological features typical of CD. Data emerging from the present investigation thus suggest that the prevalence of CD should be reassessed and that the determination of EMA IgG1 could offer a new tool in the diagnostic armamentarium of CD.

31-43 amino acid sequence of the alpha-gliadin induces anti-endomysial antibody production during in vitro challenge.

BACKGROUND: Wheat gliadin is the culprit antigen of coeliac disease (CD). Two short sequences of NH2-terminal portion of gliadin seem to be responsible for CD. Antiendomysial antibodies (EMA), highly sensitive and specific for CD, are detectable in the culture media from treated CD patients, after in vitro challenge with peptic-tryptic (PT) digest of gliadin. In this study we detected EMA production after in vitro challenge with 31-43 peptide. We used 56-68 peptide, lacking toxic sequences, as a negative control. METHODS: Duodenal samples from 11 treated CD patients and 9 control patients were cultured with 31-43 and 56-68 peptides and PT gliadin. Indirect immunofluorescence analysis was used for EMA detection. RESULTS: EMA were detected in culture media of 10 of 11 specimens challenged with PT-gliadin and in the media of all specimens challenged with 31-43 peptide. No EMA were detectable in any treated patients cultured with 56-68 peptide or with medium alone. No EMA were observed in cultures of control specimens. DISCUSSION: The ability of the 31-43 sequence of the alpha-gliadin to induce EMA production suggests its involvement in the pathogenesis of CD. Furthermore, it may be a more useful antigenic substance than PT gliadin for both in vitro and in vivo studies of CD.

Antibodies to bovine beta-casein in diabetes and other autoimmune diseases.

Cow's milk is thought to be an environmental trigger for autoimmune response in Type 1 diabetes. In the present study, our aim was to investigate the antibody response to bovine beta-casein in different immune- and non-immune-mediated diseases and to establish whether such an antibody response is specific to Type 1 diabetes. We measured antibodies to bovine beta-casein using an enzyme-linked immunosorbent assay in a total of 519 sera from subjects as follows: 71 patients with Type 1 diabetes, 33 patients with coeliac disease, 100 patients with latent autoimmune diabetes in adults (LADA), 50 patients with autoimmune thyroid disease (ATD), 50 patients with Type 2 diabetes, 24 patients with multiple sclerosis (MS), and 3 different groups of controls (n = 191). Significantly increased levels of antibodies to beta-casein were found in patients with Type 1 diabetes, coeliac disease and in LADA compared to age-matched controls (p = 0.01, p = 0.02 and p = 0.01, respectively). No differences were observed in beta-casein antibody titres between patients with other disease conditions (MS, and ATD) and age-matched controls. The highest antibody response to beta-casein in Type 1 diabetic patients and in patients with coeliac disease could reflect the gut mucosal immune disorders common to Type 1 diabetes and coeliac disease. Furthermore, the elevated beta-casein antibody levels found in LADA patients suggest that the antibody response to this protein may be relevant in autoimmune diabetes.