Antibodies against human tissue transglutaminase and endomysium in diagnosing and monitoring coeliac disease.

BACKGROUND: Coeliac disease (CD) patients often present a variety of uncharacteristic symptoms and therefore sensitive and specific screening tests are needed as an aid in making an accurate diagnosis. A recently developed ELISA, using human recombinant tissue transglutaminase (tTG) as antigen, was evaluated for its significance in the diagnosis of CD. The patient's compliance to a gluten-free diet and the serological reaction during gluten challenge were also monitored. The results were compared with IgA-endomysium antibody (EMA) results. METHODS: Sera previously collected from 365 patients (0.4-76 years) with jejunal biopsy on a gluten-containing diet and from 41 patients on a gluten-free diet or challenge were tested for IgA anti-human tTG antibodies (IgA tTG ab) with Celikey (Pharmacia Diagnostics). The study population comprised 208 CD patients and 157 controls. The diagnostic performance and cut-off for the assay were estimated with ROC analysis. EMA was analysed by indirect immunofluorescence microscopy on cryostat sections of monkey oesophagus. RESULTS: 200/208 patients with CD had positive IgA tTG ab (median >100 U/ml), while only 1/157 of the control patients were positive (median 1.67 U/ml). The area under the ROC curve was 98.3% and the sensitivity and specificity of the test were 96% and 99% for the study population. Only 4/365 patients (1%) presented discordant IgA tTG ab and EMA results, 2 of them had only IgA tTG ab and 2 only EMA. The IgA tTG ab levels and the EMA titres were closely correlated to the duration of gluten-free diet and gluten challenge, respectively. CONCLUSION: IgA tTG ab can be used as an accurate observer-independent alternative to EMA in diagnosing or monitoring CD.

[Quality assurance in diagnostics: are there normal values for IgG-antibodies to cow's milk proteins?]. / Zur Sicherung der Prozessqualität in der Diagnostik: Gibt es Normwerte für die IgG-Antikörper gegen Kuhmilchproteine?

BACKGROUND: The value of IgG-cow's milk antibodies is heavily disputed. Therefore, aim of the study was to establish normal values for these IgG-antibodies and to determine their diagnostic significance especially for gastrointestinal allergic manifestations. PATIENTS AND METHODS: 702 newborns were included prospectively. They received different feedings: mothers milk, cow's milk based formula and a partially or an extensively hydrolyzed formula. All children were examined annually or on occasion of allergic manifestations. An indirect immunofluorescent test was used for the assessment of IgG-antibodies against casein, beta-lactoglobulin, alpha-lactalbumin and bovine serum albumin. Values were expressed as geometric mean titers. RESULTS: IgG-antibody titers rose slowly in the exclusively breast-fed children and peaked rapidly in the cow's milk formula-fed group, especially during the first year of life. There were no significant differences between children with and without allergic manifestations. Because of a large overlap between asymptomatic and symptomatic children it was impossible to establish normal values for the IgG-cow's milk antibodies. CONCLUSIONS: The occurrence of IgG-cow's milk antibodies is a physiologic phenomenon without diagnostic significance. The costs of these determinations can be saved to improve process quality.

The diagnostic significance of IgG cow's milk protein antibodies re-evaluated.

UNLABELLED: The effect of different feeding regimens, notably the use of hydrolysed cow's milk formulas, on the development of allergic reactions and the development of cow's milk protein-IgG antibodies is still disputed. We prospectively compared the development of allergic manifestations and cow's milk protein-IgG antibodies in a total of 702 infants who were divided into six groups: 1. exclusively breast milk for at least 4 weeks (n = 206). 2. Breast milk plus initial partially hydrolysed formula (n = 104). 3. Breast milk plus extensively hydrolysed formula (n = 50). 4. Breast milk plus initial conventional cow's milk formula (n = 73). 5. Conventional cow's milk with or without breast milk throughout (n = 187). 6. Extensively hydrolysed cow's milk formula for 2 months, followed by conventional cow's milk (n = 82). Cow's milk protein antibodies were determined by an indirect immunofluorescent test. Antibody titres rose slowly in groups 1, 3 and 6. Children in group 5 showed two high peaks. There were no significant differences in the frequency and type of allergic manifestations between the groups. Introduction of cow's milk formula during the first trimenon resulted in elevated antibody titres in all breast fed infants compared with introduction at a later date. CONCLUSION: In contrast to a previous study from the same laboratory, there is no diagnostic significance of cow's milk protein-IgG antibodies for allergic manifestations. The occurrence of these antibodies is a physiological phenomenon: the shorter the breast feeding period and the earlier cow's milk formula is introduced, the higher the antibody levels.

Occurrence of large granular lymphocytes and natural killer cells in the epithelium of the gut distinguishes two different coeliac diseases.

In a longterm study, we have divided coeliac disease into two distinct entities (abortive and permanent) based on the occurrence of large granular lymphocytes and natural killer cells within the epithelium of the gut. The natural killer and large granular lymphocytes cells were characterised by either immunohistochemical or phase contrast microscopical procedures on the initial biopsies from 15 children with coeliac disease. They were compared with seven individuals with partial villus atrophy and eight with normal villous morphology. Although the histological findings were similar in the initial biopsies of all patients with coeliac disease, the patients with permanent coeliac disease had a significantly lower number (0.41(0.61)cells/mm2) of large granular lymphocytes and natural killer cells compared with those patients with abortive coeliac disease (11.93 (6.23) cells/mm2). Those in the permanent group developed a significantly more pronounced flat mucosa after gluten challenge or provocation compared with the abortive group and had to remain on a strict gluten free diet in contrast with those in the abortive group. Thus, the occurrence of intraepithelial large granular lymphocytes and natural killer cells characterises two distinctly different coeliac diseases. Based on our results neither the histological evaluation of the biopsy nor the utilisation of the revised European Society for Paediatric Gastroenterology and Nutrition (ESPGAN) Criteria are adequate in diagnosing the two types of coeliac disease.

Ultrastructural binding sites of endomysium antibodies from sera of patients with dermatitis herpetiformis and coeliac disease.

The ultrastructural binding sites of endomysium antibodies, specific serological markers of gluten sensitive enteropathy, were investigated in the rabbit oesophagus using the immunogold technique. Endomysium antibodies from sera of patients with dermatitis herpetiformis and with coeliac disease bound in an identical manner in a non-fibrillar material closely associated with fine collagenous-reticulin fibrils and also with similar fibrils connecting smooth muscle cells and elastic tissue in the endomysial connective tissue. These observations suggest that IgA antibodies in sera from patients with dermatitis herpetiformis and coeliac disease recognise a common antigen in an amorphous component associated with the reticular connective tissue of oesophageal lamina muscularis mucosae and thus confirm the probable identity of IgA class endomysium and jejunal antibodies.

Antigliadin and antiendomysium antibody determination for coeliac disease.

The value of IgG and IgA gliadin antibodies (AGA) was compared with that of IgA endomysium antibodies (EMA) for the diagnosis of coeliac disease. Three hundred and six of 340 (90%) children with untreated coeliac disease (flat mucosa) had EMA and 338/340 (99.4%) had IgG AGA and/or IgA AGA. Only 1/340 (a 7 year old boy with selective IgA deficiency) had neither AGA nor EMA. Absence of EMA is more frequent in coeliac patients younger than 2 years than in older patients (32/277 compared with 1/62). EMA were present in 4/211 (2%) of comparison subjects (normal mucosa), IgA AGA in 12/211 (6%), and IgG AGA in 74/211 (35%). The specificity of AGA cannot be calculated from these figures as they are biased. The combined determination of AGA and EMA, taking advantage of the high sensitivity of AGA and the high specificity of EMA, gives an excellent prediction of the condition of the mucosa: 247/248 patients (99.6%) with positive EMA and positive IgG AGA and IgA AGA had a flat mucosa, whereas 136/137 patients (99.3%) with neither AGA nor EMA had a normal mucosa. During a gluten free diet EMA and AGA disappear. Their presence or absence is therefore an indicator of dietary compliance. After reintroduction of gluten into the diet 110/134 (82%) of the patients who had a flat mucosa at diagnosis relapsed, but 24/134 still had a normal mucosa after 2-15 years of challenge. All these patients without a morphological relapse were less than 2 years old at diagnosis so we conclude that patients who are young at diagnosis should be challenged. AGA often reappear earlier than EMA. After one month of challenge 93% of patients are AGA and 69% EMA positive. After more than three years of gluten intake the percentage of AGA positive patients decreased to about 50% whereas the percentage of EMA positive sera was then highest (93%). Therefore EMA are more sensitive for the detection of 'silent' relapse after prolonged periods of gluten intake.

Binding to human jejunum of serum IgA antibody from children with coeliac disease.

Jejunal histology and the presence of serum IgA antibodies (JAB) binding to human jejunum in vitro were studied in 139 children with severe malabsorptive symptoms. Among 33 children with confirmed coeliac disease (ESPGAN criteria), 13 (93%) of 14 sampled before starting on a gluten-free diet had JAB, none of 21 sampled had JAB while on a gluten-free diet of long duration, and 90% of 30 sampled during gluten challenge had JAB. 53 children had severe jejunal villous atrophy (probable coeliac disease): 71% of those younger than 2 years and 94% of those aged 2-18 years had JAB during gluten intake. JAB could not be detected in 53 disease control patients (normal jejunal histology) and in 3 coeliac disease patients with selective IgA deficiency. Simultaneous determination of antigliadin (AGA) and antiendomysium (EMA) levels, and gliadin and tissue absorption studies, showed that JAB and AGA are different, whereas JAB and EMA are probably identical. IgA JAB could be the target-organ-related autoantibodies in coeliac disease.

[Incidence and symptoms of cow's milk protein intolerance following acute gastroenteritis in young infants. Effect of a hypoallergenic diet]. / Inzidenz und Symptome der Kuhmilchprotein-Intoleranz nach akuter Gastroenteritis bei jungen Säuglingen. Einfluss hypoallergener Diät.

In a prospective randomized study we investigated in 28 mainly bottle-fed infants younger than 60 days whether in acute gastroenteritis a hypoallergenic formula could prevent the development of cow's milk protein intolerance. Group 1 (14 infants) was fed with a formula adapted to human milk, Group 2 (14 infants) got a semi-elementary formula (Alfaré). After 3 months group II was exposed to cow's milk protein with a standardized challenge and the incidence of CMPI in both groups was calculated. All cases with the acute form of CMPI occurred in group II (5/12) whereas in group I only one infant suffered from the protracted mild form of the disease. Inspite of the relatively small number of probands we conclude from our results that in infants who are not totally breast-fed in the post-enteritic period feeding with a formula adapted to human milk is preferable to hypoallergenic semi-elementary preparations. An allergen free period of 3 months seems to induce symptoms of cow's milk intolerance, probably as a booster-effect to early sensibilisation.

IgG, IgA and IgE gliadin antibody determinations as screening test for untreated coeliac disease in children, a multicentre study.

The diagnostic value of gliadin IgG, IgA and IgE antibody (AB) determinations using the fluorescent immunosorbent test was examined in 586 children with malabsorptive disorders and/or failure to thrive. All patients underwent jejunal biopsy and were on a gluten-containing diet. IgG AB were found in all patients (331/331) with untreated coeliac disease (CD) in our study, but IgA AB in only 295/331 (89%). Therefore a screening test based only on IgA AB determinations is not recommended. By contrast, 203 (80%) of 255 children with other malabsorptive disorders had no gliadin AB, 43 (16.5%) had only IgG AB and only 9 (3.5%) had IgG and IgA AB. IgE AB proved to be of no additional value as a diagnostic tool because they were found in a quarter of the children without CD. Statistical evaluation of combined IgG and IgA AB determination showed at least 96% sensitivity and a specificity of 97%. The subjective ("Bayesian") probability that an actual patient with a given AB test result has CD, is considered: a patient very probably has CD in the case of positive IgG and IgA AB, and no CD in the case of a negative AB result. In the case of negative IgA AB but positive IgG AB the physician's judgement ("prior probability") influences the ("posterior") probability of CD for an actual patient. In contrast to IgG AB, IgA AB decline rapidly after the introduction of a gluten-free diet and may be used for diet control after diagnosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Prevalence of coeliac disease in diabetic children and adolescents. A multicentre study.

Screening for coeliac disease (CD) with serum antigliadin antibodies (AGA) was performed in 1032 diabetic children and adolescents. In 8 children CD had been diagnosed before study entry. Of the remaining 1024 children, 33 had an elevated AGA titre in the first serum sample. On follow-up an elevated AGA titre was confirmed in only 17 of 31 patients. Nine of the repeatedly positive patients underwent jejunal biopsy, and CD was diagnosed in two asymptomatic patients; both were positive for IgG- and IgA-AGA. Among 10 AGA-positive patients in whom biopsies could not be performed, only 1 showed IgA-AGA and thus carried a high risk for CD. From our results we estimate a prevalence of CD in Swiss and German diabetic children between 1.1% and 1.3%. False-positive AGA titres occurred significantly more often in patients with diabetes duration of less than 1 year. AGA testing reached a specificity of 99% if performed at least 1 year after the onset of diabetes. Children suffering from both diabetes and CD showed a diabetes manifestation at a significantly younger age than non-coeliac patients, whereas CD tended to be diagnosed at a remarkably late age.

Coexistent coeliac disease, Graves' disease and diabetes mellitus type 1 in a patient with Down syndrome.

A 17-year-old girl with Down syndrome is presented who developed coeliac disease, Graves' disease and diabetes type 1. Her HLA type was A3, A9, B8, B15, DR3, DR5.

The fluorescent immunosorbent test for IgG gliadin antibodies and the leucocyte migration inhibition test in coeliac disease; comparison of diagnostic value.

The diagnostic value of the fluorescent immunosorbent test for IgG gliadin antibodies (FIST) has been investigated in comparison with the LIF test--the competence of the gluten subfractions B2 and B3 in releasing lymphokines from peripheral lymphocytes in vitro--in 96 patients with coeliac disease (CD) under various dietary conditions. In untreated children with CD during their first 2 years of life, the FIST showed 100% sensitivity with 95% specificity, whilst the LIF test showed only 70% sensitivity and 73% specificity. Therefore it can be concluded that the FIST as a screening test is superior to the LIF. In older children with a proved recurrence of the mucosal abnormality after reintroduction of a normal diet, only 44% showed increased IgG gliadin antibody titres whereas 70% proved to be positive in the LIF test. Under a controlled gluten challenge all six patients reacted with a distinct increase in gliadin antibody titres whereas the LIF test changed from positive to negative and vice versa without following any clear principle. These results emphasize the inadequacy of the LIF test as a diagnostic method, both in untreated CD and under controlled gluten challenge.

A reliable screening test for childhood celiac disease: fluorescent immunosorbent test for gliadin antibodies. A prospective multicenter study.

The diagnostic value of gliadin antibody determination using the fluorescent immunosorbent test was examined in a prospective multicenter study comprising 251 children with malabsorptive disorders. Antibodies to gliadin were found in all 72 patients (100%) with active celiac disease (29 children with celiac disease proved by challenge, 43 with probable celiac disease). All children up to the age of 7 years had antibodies in high titers. By contrast, 96 (84%) of 114 children with other malabsorptive disorders and a normal mucosa or with partial villous atrophy had no gliadin antibodies, 14 (12%) had a low titer, and only four (3.5%) showed moderate to high titers. Four children with gastrointestinal tract symptoms of cow milk intolerance and a flat mucosa also showed no antibodies. In 24 of 29 children (83%) with cystic fibrosis and six of seven children with Crohn disease (biopsies not performed in either group), no antibodies could be detected. The others had low or elevated titers. In 25 children with acute gastroenteritis (not biopsied) antibodies were not found at hospital admission nor six weeks later after reintroduction of gluten. The determination of antibodies to gliadin with the fluorescent immunosorbent test is a reliable screening test for childhood celiac disease. In our series there were no false negative results in children with untreated celiac disease. A positive gliadin antibody titer is not proof of celiac disease. In each child the diagnosis must be confirmed by small intestinal biopsy even if the gliadin antibody titer is high. The detection of high titers of cow milk antibodies in 27% of patients with celiac disease is of no value.