Small-intestinal TG2-specific plasma cells at different stages of coeliac disease.

BACKGROUND: In coeliac disease, ingestion of gluten induces the production of transglutaminase 2 (TG2)-targeted autoantibodies by TG2-specific plasma cells present at high frequency in the small intestinal mucosa in untreated disease. During treatment with a gluten-free diet (GFD), the number of these cells decreases considerably. It has not been previously investigated whether the cells are also present prior to development of villous atrophy, or in non-responsive patients and those with dietary lapses. We aimed to define the frequency of small bowel mucosal TG2-specific plasma cells in coeliac disease patients with varying disease activity, and to investigate whether the frequency correlates with serum and small intestinal TG2-targeting antibodies as well as mucosal morphology and the number of intraepithelial lymphocytes. RESULTS: Mucosal TG2-specific plasma cells were found in 79% of patients prior to development of mucosal damage, in all patients with villous atrophy, and in 63% of the patients after 1 year on GFD. In these disease stages, TG2-specific plasma cells accounted for median of 2.3, 4.3, and 0.7% of all mucosal plasma cells, respectively. After long-term treatment, the cells were present in 20% of the patients in clinical remission (median 0%) and in 60% of the patients with poor dietary adherence (median 5.8%). In patients with non-responsive coeliac disease despite strict GFD, the cells were found in only one (9%) subject; the cells accounted for 2.4% of all plasma cells. A positive correlation between the percentage of TG2-specific plasma cells and serum TG2 antibody levels (rS = 0.69, P < 0.001) and the intensity of mucosal TG2-targeting IgA deposits (rS = 0.43, P < 0.001) was observed. CONCLUSIONS: Our results show that TG2-specific plasma cells are already detectable prior to villous atrophy, and that generally their frequency increases during overt disease. By contrast, on GFD, the percentage of these cells decreases. Overall, the presence of TG2-specific plasma cells in the small bowel mucosa mirrors the presence of gluten in the diet, but the frequency is not always parallel to the level of serum or intestinal TG2 antibodies. These findings increase the knowledge about the development of the TG2 plasma cell responses especially in the early phases of coeliac disease.

Injection of celiac disease patient sera or immunoglobulins to mice reproduces a condition mimicking early developing celiac disease.

UNLABELLED: Typical features of celiac disease are small-bowel villus atrophy, crypt hyperplasia, and inflammation which develop gradually concomitant with ingestion of gluten. In addition, patients have anti-transglutaminase 2 (TG2) autoantibodies in their serum and tissues. The aim of this study was to establish whether celiac disease can be passively transferred to mice by serum or immunoglobulins. Serum aliquots or purified immunoglobulins (Ig) were intraperitoneally injected into Hsd:Athymic Nude-Foxn1nu mice for 8 or 27 days. As mice do not have proper IgA transport from peritoneum to blood, sera with a high content of IgG class anti-TG2 antibodies from untreated IgA-deficient celiac patients were used. Mouse sera were tested for celiac disease-specific autoantibodies, and several tissues were analyzed for autoantibody deposits targeted to TG2. Morphological assessment was made of the murine small intestinal mucosa. Injection of celiac disease patient sera or total IgG led to a significant delay in weight gain and mild diarrhea in a subset of mice. The mice injected with celiac patient sera or IgG had significantly decreased villus height crypt depth (Vh/CrD) ratios and celiac disease-specific autoantibody deposits targeted to TG2 in several tissues, including the small intestine. None of these features were observed in control mice. We conclude that administration of IgA-deficient celiac disease patient serum or total IgG induces both deterioration of the intestinal mucosa and clinical features of celiac disease in mice. The experimentally induced condition in the mice injected with patient serum or IgG resembles early developing celiac disease in humans. KEY MESSAGE: Celiac disease patient sera or total IgG was injected into athymic mice. A significant delay in weight gain and mild diarrhea was observed. Mice evinced significantly decreased villus height crypt depth ratios. Celiac disease-specific autoantibody deposits were present in several tissues. The condition in mice resembles early stage celiac disease in humans.

Impaired epithelial integrity in the duodenal mucosa in early stages of celiac disease.

The small-bowel mucosal damage characteristic of celiac disease (CD) develops from normal villus morphology to inflammation and finally to villus atrophy with crypt hyperplasia. Patients with early stage CD may already suffer from abdominal symptoms before the development of villus atrophy. Although epithelial junctional integrity is compromised in overt disease, the appearance of such changes in early phases of the disorder is not known. We investigated whether alterations in epithelial junction protein expression occur already in early stage CD with normal mucosal morphology, and whether this correlates with inflammation indicators and clinical symptoms. The study involved 10 patients with early stage and 10 patients with overt villus atrophy that were followed yearly according to the study protocol. As controls, 20 nonceliac subjects were included. The expression of junction proteins (occludin, claudin 3, zonula occludens 1, and E-cadherin) was studied in small-intestinal biopsies using immunohistochemistry and Western blot. The correlation between junctional proteins and mucosal morphology, autoantibodies, the number of intraepithelial lymphocytes (IELs), and gastrointestinal symptoms was assessed. The expression of all junction proteins was already decreased in early stage CD when compared with nonceliac controls (P < 0.05). Junction protein expression correlated positively with mucosal villus morphology and negatively with the number of IELs, the intensity of small-intestinal autoantibody deposits, and serum autoantibodies. The expression of claudin 3 showed a negative correlation with diarrheal score (R = -0.314, P = 0.04). These findings show that the mucosal epithelial integrity is disrupted already in early stage CD before the disorder progresses to full-blown enteropathy.

Small bowel transglutaminase 2-specific IgA deposits in dermatitis herpetiformis.

Dermatitis herpetiformis (DH) is an extraintestinal manifestation of coeliac disease. Untreated coeliac disease patients are known to have transglutaminase 2 (TG2)-targeted IgA deposits in the small bowel mucosa. To evaluate whether similar intestinal IgA deposits are also present in DH and whether the deposits disappear with gluten-free diet, 47 untreated and 27 treated DH patients were studied. Seventy-nine percent of untreated and 41% of the treated DH patients had TG2-specific IgA deposits in the small bowel, and the presence of the deposits showed a significant association with the degree of small bowel villous atrophy (p < 0.001). Other coeliac-disease related inflammatory markers were also investigated, and the density of small bowel mucosal intraepithelial γδ(+) T cells was increased in 91% of untreated and 73% of treated DH patients. The results show that the majority of untreated DH patients have similar gluten-dependent TG2-specific IgA deposits the small bowel mucosa as coeliac disease patients.

Validation of morphometric analyses of small-intestinal biopsy readouts in celiac disease.

BACKGROUND: Assessment of the gluten-induced small-intestinal mucosal injury remains the cornerstone of celiac disease diagnosis. Usually the injury is evaluated using grouped classifications (e.g. Marsh groups), but this is often too imprecise and ignores minor but significant changes in the mucosa. Consequently, there is a need for validated continuous variables in everyday practice and in academic and pharmacological research. METHODS: We studied the performance of our standard operating procedure (SOP) on 93 selected biopsy specimens from adult celiac disease patients and non-celiac disease controls. The specimens, which comprised different grades of gluten-induced mucosal injury, were evaluated by morphometric measurements. Specimens with tangential cutting resulting from poorly oriented biopsies were included. Two accredited evaluators performed the measurements in blinded fashion. The intraobserver and interobserver variations for villus height and crypt depth ratio (VH:CrD) and densities of intraepithelial lymphocytes (IELs) were analyzed by the Bland-Altman method and intraclass correlation. RESULTS: Unevaluable biopsies according to our SOP were correctly identified. The intraobserver analysis of VH:CrD showed a mean difference of 0.087 with limits of agreement from -0.398 to 0.224; the standard deviation (SD) was 0.159. The mean difference in interobserver analysis was 0.070, limits of agreement -0.516 to 0.375, and SD 0.227. The intraclass correlation coefficient in intraobserver variation was 0.983 and that in interobserver variation 0.978. CD3(+) IEL density countings in the paraffin-embedded and frozen biopsies showed SDs of 17.1% and 16.5%; the intraclass correlation coefficients were 0.961 and 0.956, respectively. CONCLUSIONS: Using our SOP, quantitative, reliable and reproducible morphometric results can be obtained on duodenal biopsy specimens with different grades of gluten-induced injury. Clinically significant changes were defined according to the error margins (2SD) of the analyses in VH:CrD as 0.4 and in CD3(+)-stained IELs as 30%.

An update on the diagnostics of celiac disease.

In celiac disease, highly sensitive and specific serum endomysial and transglutaminase 2 antibody tests are widely used in identifying patients for diagnostic endoscopy and small-bowel biopsy. In addition, the recently developed deamidated gliadin peptide antibody tests show promise in celiac disease diagnostics. In view of these apparent problems attending the diagnostic gold standard, gluten-induced small-bowel mucosal villous atrophy with crypt hyperplasia, other diagnostic approaches beyond conventional histology have been introduced. Furthermore, the diagnostic criteria for celiac disease are currently under revision with an eye also to noninvasive diagnostic strategies.

IgA-class autoantibodies against neuronal transglutaminase, TG6 in celiac disease: no evidence for gluten dependency.

BACKGROUND: It has been suggested that antibodies against transglutaminase (TG) 6 could serve as a biomarker to identify a subgroup of gluten-sensitive patients who may be at risk of developing neurological disease. We here investigated whether TG6-targeted autoantibodies are a characteristic feature of celiac patients, especially those with neurological symptoms, and further, whether such antibodies are gluten-dependent. METHODS: Serum IgA-class TG6 autoantibodies were measured in untreated and treated celiac patients with and without neurological manifestions and in non-celiac controls. The results were compared to TG2 autoantibody levels. RESULTS: During a gluten-containing diet the number of TG6 autoantibody-positive celiac patients with neurological problems (25%) did not significantly differ from that of TG6-seropositive patients without neurological impairment (16%) or from non-celiac controls (15%). This was in contrast to our finding in TG2 autoantibody-positive individuals, whose numbers differed significantly between all three study groups. On a gluten-free diet the levels of TG6 autoantibodies remained unchanged. CONCLUSIONS: We conclude that the serum IgA-class TG6 autoantibody assay is not able to distinguish gluten-sensitive patients with neurological manifestations from celiac patients without neurological problems or from control subjects, and further, that TG6 autoantibodies are not gluten-dependent.

Serodiagnostic assays for celiac disease based on the open or closed conformation of the autoantigen, transglutaminase 2.

The autoantigen of celiac disease, transglutaminase 2 (TG2), adopts an open conformation during enzymatic activation. We studied diagnostic accuracy of serodiagnostic assays using TG2 in its open and closed conformation as antigens in patients with diagnostic difficulties. The open TG2 antibody (TG2ab) test identified 93% of untreated celiac patients in contrast to 44%, 27%, and 68% detected by closed and conventional TG2ab and endomysial antibody (EmA) tests, respectively. The assay was able to detect 60% of non-responding celiac patients seronegative for conventional TG2ab and EmA. The titers of the open TG2abs were higher than those of the closed TG2abs. The serological test utilizing TG2 in an open conformation was more accurate than the other assays in finding active celiac disease even in patients having negative or borderline conventional celiac autoantibodies and in revealing poor dietary response non-invasively. It thus offers a promising tool in the diagnostics and follow-up of celiac disease.

Coeliac disease--a diagnostic and therapeutic challenge.

During the past 20 years the diagnosis of coeliac disease has improved significantly. However, at the same time the true prevalence of the condition has doubled, involving more than 2% of the population in some countries. Due to mild or atypical symptoms, the diagnosis remains a challenge for the health care system. Highly sensitive and specific serum endomysial and transglutaminase-2 antibody tests are helpful in identifying patients for diagnostic endoscopy and small-bowel biopsy. The diagnosis of the disease is still based on the demonstration of gluten-induced small-bowel mucosal villous atrophy with crypt hyperplasia. However, coeliac disease may manifest itself before the development of the overt small-intestinal lesion. Positive endomysial and transglutaminase antibodies in patients with normal small-bowel mucosal villous architecture may indicate early stage coeliac disease. Currently, the only effective treatment for the condition is a life-long strict gluten-free diet. Long-term regular follow-up of patients is recommended in order to maintain good adherence to the diet.

Celiac disease without villous atrophy in children: a prospective study.

OBJECTIVE: To establish whether children who are endomysial antibody (EmA) positive and have normal small-bowel mucosal villous morphology are truly gluten-sensitive and may benefit from early treatment with a gluten-free diet. STUDY DESIGN: Children who were EmA positive with normal small-bowel mucosal villi were compared with children who were seropositive with villous atrophy by using several markers of untreated celiac disease. Thereafter, children with normal villous structure either continued on a normal diet or were placed on a gluten-free diet and re-investigated after 1 year. Seventeen children who were seronegative served as control subjects for baseline investigations. RESULTS: Normal villous morphology was noted in 17 children who were EmA positive, and villous atrophy was noted in 42 children who were EmA positive. These children were comparable in all measured variables regardless of the degree of enteropathy, but differed significantly from the seronegative control subjects. During the dietary intervention, in children who were EmA positive with normal villi, the disease was exacerbated in children who continued gluten consumption, whereas in all children who started the gluten-free diet, both the gastrointestinal symptoms and abnormal antibodies disappeared. CONCLUSIONS: The study provided evidence that children who are EmA positive have a celiac-type disorder and benefit from early treatment despite normal mucosal structure, indicating that the diagnostic criteria for celiac disease should be re-evaluated.

Intestinal transglutaminase 2 specific antibody deposits in non-responsive coeliac disease.

BACKGROUND AND AIMS: The diagnosis of coeliac disease is problematic in individuals not responding to a gluten-free diet. Small-bowel villous atrophy occurs in other enteropathies and non-responsive patients are often seronegative. We investigated whether small-bowel mucosal transglutaminase-2 specific autoantibody deposits distinguish non-responsive coeliac disease from other enteropathies. METHODS: Small-bowel mucosal autoantibody deposits were determined in 27 non-responsive, 28 responsive coeliac patients and 10 controls with other enteropathies. Of the non-responsive coeliac patients six were adhering poorly and 21 strictly to the diet; six of the 21 had enteropathy-associated lymphoma, five refractory coeliac disease and 10 otherwise persistent villous atrophy. The presence of mucosal autoantibody deposits was compared to serology, villous morphology, densities of intraepithelial lymphocytes (IELs) and markers of refractory coeliac disease. RESULTS: Twenty out of 21 well-adhering, all six poorly adhering non-responsive and all 28 untreated responsive coeliac patients had small-bowel mucosal autoantibody deposits present, while controls with other enteropathies were negative. Small-bowel mucosal autoantibody deposits were more accurate in detecting coeliac disease than serology or IEL densities. Refractory coeliac markers revealed only cases with the most severe condition. CONCLUSIONS: Small-bowel mucosal autoantibody deposits differentiate coeliac disease from other enteropathies, enabling the design of appropriate therapeutic strategies.

Usefulness of small-bowel mucosal transglutaminase-2 specific autoantibody deposits in the diagnosis and follow-up of celiac disease.

BACKGROUND: Diagnosis of celiac disease may be problematic in that small-bowel villous atrophy sometimes occurs in conjunction with other enteropathies, develops gradually and may be patchy. Furthermore, as the often compromised quality of biopsy specimens renders diagnosis difficult, new diagnostic tools are warranted. GOALS: As the celiac disease-specific autoantibodies are found deposited at their production site, in the small-bowel mucosa, they may be useful in diagnostics, especially in problematic cases. We therefore systematically assessed the occurrence of celiac-specific autoantibody deposits in a large cohort of celiac patients, and established how IgA deposits decline after initiation of a gluten-free diet. METHODS: Transglutaminase-2 specific mucosal IgA autoantibody deposits were determined from small-bowel mucosal biopsies in 261 untreated, 71 short-term (1 y), and 105 long-term (2 to 41 y) treated celiac disease patients and in 78 nonceliac controls. The presence of the deposits was compared with celiac serology, mucosal villous morphology and density of intraepithelial lymphocytes. RESULTS: All untreated celiac disease patients had mucosal autoantibody deposits and their intensity was moderate or strong in 90% of cases. In contrast, 18% of the controls had weak depositions. During a gluten-free diet the intensity of the deposits diminished, but was still faintly positive in 56% of long-term treated celiac patients. The efficiency of the test in determining mucosal autoantibody deposits was superior to serology and inflammatory markers. CONCLUSIONS: Mucosal transglutaminase-2 specific autoantibody deposits proved to be accurate gluten-dependent markers of celiac disease and would thus be of value in the diagnostics and dietary monitoring of this disorder.

Oats do not induce systemic or mucosal autoantibody response in children with coeliac disease.

OBJECTIVES: A gluten-free diet omitting wheat, rye, and barley is the only effective treatment for coeliac disease. The necessity of excluding oats from the diet has remained controversial. We studied the toxicity of oats in children with coeliac disease during a 2-year follow-up by investigating jejunal transglutaminase 2 (TG2)-targeted IgA-class autoantibody deposits, a potentially more sensitive disease marker than serum antibodies or conventional histology. PATIENTS AND METHODS: Twenty-three coeliac children in remission were randomized to undergo oat or gluten challenge with wheat, rye, barley, and oats. When jejunal histological relapse was evident after gluten challenge, patients excluded wheat, rye, and barley but continued with oats. Mucosal morphology and TG2-targeted autoantibody deposits were studied in jejunal biopsies taken at baseline and after 6 and 24 months. Furthermore, serum IgA-class TG2 antibodies were measured. RESULTS: At baseline, serum TG2 antibodies were negative in all 23 patients, but 7 of them had minor mucosal deposits. In the oats group, there was no significant change in the intensity of the deposits within 2 years. In contrast, during the gluten challenge, the intensity of the deposits clearly increased and decreased again when wheat, rye, and barley were excluded but consumption of oats was continued; this was in line with serum autoantibodies. The intensity of the mucosal deposits correlated well with both villous morphology and serum autoantibody levels. CONCLUSIONS: Consumption of oats does not induce TG2 autoantibody production at mucosal level in children with coeliac disease. Measurement of small-intestinal mucosal autoantibody deposits is suitable for monitoring treatment in coeliac patients.

Gluten-dependent small bowel mucosal transglutaminase 2-specific IgA deposits in overt and mild enteropathy coeliac disease.

OBJECTIVES: In coeliac disease, immunoglobulin (Ig)A-class autoantibodies against transglutaminase-2 are produced in the small intestinal mucosa, where they are deposited extracellularly. It remains unclear whether positive intestinal transglutaminase-2-targeted IgA deposits in subjects having normal small bowel mucosal morphology are signs of early-stage coeliac disease. We evaluated the gluten dependency of these deposits in overt and mild enteropathy coeliac disease. PATIENTS AND METHODS: All together 48 subjects suspected of coeliac disease but having normal small bowel mucosal villi were enrolled; 28 of them had latent coeliac disease. The remaining 20 having positive intestinal IgA deposits adopted a gluten-free diet before villous atrophy had developed. For comparison, 13 patients with overt coeliac disease and 42 noncoeliac controls were studied. Small bowel mucosal transglutaminase-2-specific autoantibodies were compared with villous morphology, intraepithelial lymphocyte densities, and serum coeliac autoantibodies. RESULTS: Intestinal IgA deposits were seen in all but 1 of the patients with latent coeliac disease, when the morphology was still intact; the intensity of these deposits increased as villous atrophy developed and decreased again on a gluten-free diet. In 20 patients with intestinal IgA deposits in normal villi, the intensity of the deposits decreased with the diet similarly to that seen in patients with overt coeliac disease. Mucosal IgA deposits were seen initially only in 5% of noncoeliac controls and in 8% after extended gluten consumption. CONCLUSIONS: The response of small bowel mucosal transglutaminase-2-specific IgA deposits for dietary intervention was similar in overt and mild enteropathy coeliac disease. Detection of such IgA deposits thus offers a good diagnostic tool to uncover early-stage coeliac disease.

Family type and criminal behaviour of male offspring: the Northern Finland 1966 Birth Cohort Study.

BACKGROUND: Unstable family environment during childhood is known to predispose to juvenile delinquency. AIMS: This study explored whether childhood family structure is associated with violent behaviour of adult offspring. METHODS: We used a large, unselected general population birth cohort (n = 5589 males) linked with the national crime registers (up to the age of 32 years). The Ministry of Justice provided information on registered offences for all subjects. A logistic regression analysis was performed to examine the association between family type and criminality (violent and non-violent crimes). RESULTS: We found that single-parent family "at birth" (adj. OR 3.6, 95% CI 1.8-7.0) and "all time" (up to the age of 14 years) (adj. OR 5.2, 95% CI 2.5-10.6) were risk factors for violent offences of an adult offspring. Also parental death (adj. OR 2.2, 95% CI 1.3-3.6) and divorce (adj. OR 2.5, 95% CI 1.6-3.7) doubled the risk for violence. Non-violent offences were associated only with parental death and, divorce. CONCLUSIONS: A single-parent family of origin is strongly associated with later violent criminality of male offspring. Further studies are needed to explore the psychosocial aspects of single-parent family environment which may promote the vulnerability to violent offending in adulthood.

Is occupation relevant in suicide?

BACKGROUND: The seasonality of suicide rates and methods of suicide may be related to changes in weather and conditions of employment. Particularly the amount of occupational outdoor exposure could show differences in the distribution of suicides by season and the selection of suicide method, in addition to age at the time of death. METHODS: The data consisted of all death certificates (n=1359) of completed suicides in the province of Oulu, Finland, during the years 1988-1999. For male subjects included in this study, four occupational groups were identified according to decreasing occupational outdoor exposure. The mean ages, the distribution of suicide methods and the seasonal variation in suicides for each occupational group were analyzed. RESULTS: Farmers were significantly older at the time of suicide than construction or indoor workers, and farmers employed significantly more violent methods than the other occupational groups. In the spring, farmers had a significant peak in the rate of suicides. In the winter, forest workers had a significant trough in the rate of suicides. In the summer, indoor workers had a significant peak in the rate of suicides. LIMITATIONS: The analyses were restricted to males due to the low number of females in the study population. CONCLUSIONS: The novel finding in this study was that the seasonality of violent suicides was most strongly seen as a spring peak and a winter trough among outdoor workers. The recognition of typical risk factors of different occupations, such as outdoor exposure, and occupational-related susceptibility towards certain suicide methods could benefit in the prevention of suicides.