Katri Lindfors

Live probiotic Bifidobacterium lactis bacteria inhibit the toxic effects induced by wheat gliadin in epithelial cell culture

Wheat gliadin induces severe intestinal symptoms and small‐bowel mucosal damage in coeliac disease patients. At present, the only effective treatment for the disease is a strict life‐long gluten‐free diet. In this study we investigated whether probiotics Lactobacillus fermentum or Bifidobacterium lactis can inhibit the toxic effects of gliadin in intestinal cell culture conditions. The ability of live probiotics to inhibit peptic‐tryptic digested gliadin‐induced damage to human colon cells Caco‐2 was evaluated by measuring epithelial permeability by transepithelial resistance, actin cytoskeleton arrangements by the extent of membrane ruffling and expression of tight junctional protein ZO‐1. B. lactis inhibited the gliadin‐induced increase dose‐dependently in epithelial permeability, higher concentrations completely abolishing the gliadin‐induced decrease in transepithelial resistance. The same bacterial strain also inhibited the formation of membrane ruffles in Caco‐2 cells induced by gliadin administration. Furthermore, it also protected the tight junctions of Caco‐2 cells against the effects of gliadin, as evinced by the pattern of ZO‐1 expression. We conclude thus that live B. lactis bacteria can counteract directly the harmful effects exerted by coeliac‐toxic gliadin and would clearly warrant further studies of its potential as a novel dietary supplement in the treatment of coeliac disease.

Persistent small bowel mucosal villous atrophy without symptoms in coeliac disease

Background  Refractory sprue with malabsorption carries a risk of lymphoma.

The duodenal microbiota composition of adult celiac disease patients is associated with the clinical manifestation of the disease.

Background:Celiac disease is classically manifested in the gastrointestinal (GI) tract but extraintestinal symptoms, such as dermatitis herpetiformis (DH), are also common. Besides several well-known shared genetic risk factors and an environmental trigger, gliadin, factors determining the clinical outcome of the disease are not known. In this study, the role of duodenal microbiota in the celiac disease outcome was studied by analyzing mucosa-associated microbiota in celiac disease patients with a variety of intestinal and extraintestinal symptoms. Methods:Microbiota in duodenal biopsy samples obtained from 33 patients with celiac disease with GI, DH, anemia, or mixed symptoms, as well as screen-detected asymptomatic celiac disease and 18 control subjects were analyzed using PCR denaturing gradient gel electrophoresis and a subset of samples additionally by the 16S ribosomal RNA gene sequencing. Results:The composition and diversity of mucosal microbiota was associated with the manifestation of celiac disease when analyzed using PCR denaturing gradient gel electrophoresis and the 16S ribosomal RNA gene sequencing. The patients with celiac disease with GI symptoms or anemia had lower microbial diversity than those with DH. Moreover, the patients with GI symptoms had different intestinal microbiota composition and structure, dominated by Proteobacteria, in comparison to those with DH or control subjects (patients with dyspepsia). The relatively similar intestinal microbiota composition in the control subjects and those with DH was characterized by the high abundance of Firmicutes. Conclusions:The two common outcomes of celiac disease, classical GI and extraintestinal manifestations, had marked differences on the diversity and composition of intestinal microbiota. This association suggested that intestinal microbiota may have a role in the manifestation of the disease.

Altered Duodenal Microbiota Composition in Celiac Disease Patients Suffering From Persistent Symptoms on a Long-Term Gluten-Free Diet

Objectives:A significant fraction of celiac disease patients suffer from persistent symptoms despite a long-term gluten-free diet (GFD) and normalized small bowel mucosa. The commonly suggested reasons, such as inadvertent gluten-intake or presence of other gastrointestinal disease, do not explain the symptoms in all these patients. Recently, alterations in intestinal microbiota have been associated with autoimmune disorders, including celiac disease. This led us to test a hypothesis that abnormal intestinal microbiota may be associated with persisting gastrointestinal symptoms in treated celiac disease patients.Methods:Duodenal microbiota was analyzed in 18 GFD-treated patients suffering from persistent symptoms and 18 treated patients without symptoms by 16S rRNA gene pyrosequencing. The celiac disease patients had been following a strict GFD for several years and had restored small bowel mucosa and negative celiac autoantibodies. Their symptoms on GFD were assessed with Gastrointestinal Symptom Rating Scale.Results:The results of several clustering methods showed that the treated celiac disease patients with persistent symptoms were colonized by different duodenal microbiota in comparison with patients without symptoms. The treated patients with persistent symptoms had a higher relative abundance of Proteobacteria (P=0.04) and a lower abundance of Bacteroidetes (P=0.01) and Firmicutes (P=0.05). Moreover, their microbial richness was reduced. The results indicated intestinal dysbiosis in patients with persistent symptoms even while adhering to a strict GFD.Conclusions:Our findings indicate that dysbiosis of microbiota is associated with persistent gastrointestinal symptoms in treated celiac disease patients and open new possibilities to treat this subgroup of patients.

Coeliac disease-specific autoantibodies targeted against transglutaminase 2 disturb angiogenesis

Coeliac disease is characterized by immunoglobulin‐A (IgA)‐class autoantibodies targeted against transglutaminase 2 (TG2), a multi‐functional protein also with a role in angiogenesis. These antibodies are present in patient serum but are also found bound to TG2 below the epithelial basement membrane and around capillaries in the small intestinal mucosa. Based on these facts and the information that the mucosal vasculature of coeliac patients on a gluten‐containing diet is disorganized, we studied whether the coeliac disease‐specific autoantibodies targeted against TG2 would disturb angiogenesis. The effects of coeliac disease‐specific autoantibodies on in vitro angiogenesis were studied in angiogenic cell cultures. The binding of the antibodies to cells, endothelial sprouting, migration of both endothelial and vascular mesenchymal cells, the integrity of the actin cytoskeleton in both cell types and the differentiation of vascular mesenchymal cells were recorded. In vitro, IgA derived from coeliac disease patients on a gluten‐containing diet binds to surface TG2 on endothelial and vascular mesenchymal cells and this binding can be inhibited by the removal of TG2. In addition, coeliac disease‐specific autoantibodies targeting TG2 disturb several steps of angiogenesis: endothelial sprouting and the migration of both endothelial and vascular mesenchymal cells. Furthermore, the autoantibodies cause disorganization of the actin cytoskeleton in both capillary cell types that account most probably for the defective cellular migration. We conclude that coeliac disease‐specific autoantibodies recognizing TG2 inhibit angiogenesis in vitro. This disturbance of the angiogenic process could lead in vivo to the disruption of the mucosal vasculature seen in coeliac disease patients on a gluten‐containing diet.

Transcription factors GATA-4 and GATA-6 in normal and neoplastic human gastrointestinal mucosa

BackgroundHuman gastrointestinal mucosa regenerates vigorously throughout life, but the factors controlling cell fate in mature mucosa are poorly understood. GATA transcription factors direct cell proliferation and differentiation in many organs, and are implicated in tumorigenesis. GATA-4 and GATA-6 are considered crucial for the formation of murine gastrointestinal mucosa, but their role in human gastrointestinal tract remains unexplored. We studied in detail the expression patterns of these two GATA factors and a GATA-6 down-stream target, Indian hedgehog (Ihh), in normal human gastrointestinal mucosa. Since these factors are considered important for proliferation and differentiation, we also explored the possible alterations in their expression in gastrointestinal neoplasias. The expression of the carcinogenesis-related protein Indian hedgehog was also investigated in comparison to GATA factors.MethodsSamples of normal and neoplastic gastrointestinal tract from children and adults were subjected to RNA in situ hybridization with 33P labelled probes and immunohistochemistry, using an avidin-biotin immunoperoxidase system. The pathological tissues examined included samples of chronic and atrophic gastritis as well as adenomas and adenocarcinomas of the colon and rectum.ResultsGATA-4 was abundant in the differentiated epithelial cells of the proximal parts of the gastrointestinal tract but was absent from the distal parts. In contrast, GATA-6 was expressed throughout the gastrointestinal epithelium, and in the distal gut its expression was most intense at the bottom of the crypts, i.e. cells with proliferative capacity. Both factors were also present in Barretts esophagus and metaplasia of the stomach. GATA-6 expression was reduced in colon carcinoma. Ihh expression overlapped with that of GATA-6 especially in benign gastrointestinal neoplasias.ConclusionThe results suggest differential but overlapping functions for GATA-4 and GATA-6 in the normal gastrointestinal mucosa. Furthermore, GATA-4, GATA-6 and Ihh expression is altered in premalignant dysplastic lesions and reduced in overt cancer.

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.

Association study of FUT2 (rs601338) with celiac disease and inflammatory bowel disease in the Finnish population

Homozygosity for a nonsense mutation in the fucosyltransferase 2 (FUT2) gene (rs601338G>A) leads to the absence of ABH blood groups (FUT2 non-secretor status) in body fluids. As the secretor status has been shown to be a major determinant for the gut microbial spectrum, assumed to be important in the gut immune homeostasis, we studied the association of rs601338-FUT2 with celiac disease (CelD) and inflammatory bowel disease (IBD) in the Finnish population. Rs601338 was genotyped in CelD (n = 909), dermatitis herpetiformis (DH) (n = 116), ulcerative colitis (UC) (n = 496) and Crohns disease (CD) (n = 280) patients and healthy controls (n = 2738). CelD showed significant genotypic [P = 0.0074, odds ratio (OR): 1.28] and recessive (P = 0.015, OR: 1.28) association with the rs601338-AA genotype. This was also found in the combined CelD+DH dataset (genotype association: P = 0.0060, OR: 1.28; recessive association: P < 0.011, OR: 1.28). The A allele of rs601338 showed nominal association with dominant protection from UC (P = 0.044, OR: 0.82) and UC+CD (P = 0.035, OR: 0.84). The frequency of non-secretors (rs601338-GG) in controls, CelD, DH, UC and CD datasets was 14.7%, 18%, 18.1%, 14.3% and 16.1%, respectively. No association was evident in the DH or CD datasets alone. In conclusion, FUT2 non-secretor status is associated with CelD susceptibility and FUT2 secretor status may also play a role in IBD in the Finnish population.

A single conformational transglutaminase 2 epitope contributed by three domains is critical for celiac antibody binding and effects

The multifunctional, protein cross-linking transglutaminase 2 (TG2) is the main autoantigen in celiac disease, an autoimmune disorder with defined etiology. Glutamine-rich gliadin peptides from ingested cereals, after their deamidation by TG2, induce T-lymphocyte activation accompanied by autoantibody production against TG2 in 1–2% of the population. The pathogenic role and exact binding properties of these antibodies to TG2 are still unclear. Here we show that antibodies from different celiac patients target the same conformational TG2 epitope formed by spatially close amino acids of adjacent domains. Glu153 and 154 on the first alpha-helix of the core domain and Arg19 on first alpha-helix of the N-terminal domain determine the celiac epitope that is accessible both in the closed and open conformation of TG2 and dependent on the relative position of these helices. Met659 on the C-terminal domain also can cooperate in antibody binding. This composite epitope is disease-specific, recognized by antibodies derived from celiac tissues and associated with biological effects when passively transferred from celiac mothers into their newborns. These findings suggest that celiac antibodies are produced in a surface-specific way for which certain homology of the central glutamic acid residues of the TG2 epitope with deamidated gliadin peptides could be a structural basis. Monoclonal mouse antibodies with partially overlapping epitope specificity released celiac antibodies from patient tissues and antagonized their harmful effects in cell culture experiments. Such antibodies or similar specific competitors will be useful in further functional studies and in exploring whether interference with celiac antibody actions leads to therapeutic benefits.

Antibodies in celiac disease: implications beyond diagnostics

Celiac disease is a multisystemic dietary, gluten-induced autoimmune disorder characterized by the presence of transglutaminase (TG) 2 serum autoantibodies. Distinct autoantibodies targeting members of the TG family (TG2, TG3 and TG6) are found deposited in small-bowel mucosa and in extraintestinal tissues affected by the disease. Serum autoantibodies against other self-antigens also emerge in untreated celiac disease patients. Although villous atrophy and crypt hyperplasia in small-bowel biopsy samples are still the gold standards in diagnostics, celiac disease-specific antibodies are widely used as diagnostic aids. Gluten-induced small-bowel mucosal T-cell response is the cornerstone in the pathogenesis of the disorder, but humoral immunity may also play a central role. This review article is focused on the autoantibodies that occur in the context of celiac disease. The article summarizes the diagnostic utility of different celiac-related antibodies and discusses their roles in the pathogenesis of the disease.