BACKGROUND

Celiac disease (CD) has emerged as a common, but largely undiagnosed health problem. Numerous studies examined the influence of infant nutrition on the development of diagnosed CD. However, results are still inconsistent. In addition, the effect of infant feeding practices on the development of potential forms of CD might be different.

OBJECTIVE

The objective was to examine whether the timing of gluten introduction and breastfeeding duration are associated with CD autoimmunity (CDA) in children at the age of 6 y.

METHODS

This study was embedded in the Generation R Study, a population-based prospective cohort study. Participants included 1679 Dutch children who were positive for human leukocyte antigen (HLA) DQ2/DQ8. Data on the timing of gluten introduction (<6 mo compared with ≥6 mo) and duration of breastfeeding (<6 mo compared with ≥6 mo) were obtained by questionnaire. Serum samples were analyzed for anti-tissue transglutaminase (anti-tTG) concentrations at age 6 y. Anti-tTG concentrations were categorized into negative (<7 U/mL) and positive (≥7 U/mL) values. Positive anti-tTG concentrations were further categorized based on ≥10 times the upper limit of normal (ULN) values of the test kit (≥7-70 and ≥70 U/mL). Multivariable logistic regression analyses were performed.

RESULTS

Positive anti-tTG concentrations were found in 43 children, 26 of whom had concentrations ≥10 times the ULN (≥70 IU/mL). The introduction of gluten from the age of 6 mo onward and breastfeeding for ≥6 mo were not significantly associated with positive anti-tTG concentrations. In addition, the timing of gluten introduction and duration of breastfeeding were not significantly associated with positive anti-tTG concentrations below or above 10 times the ULN.

CONCLUSIONS

Delayed introduction of gluten beyond the age of 6 mo does not increase the risk of CDA. In addition, breastfeeding for ≥6 mo does not decrease the risk of CDA in children at 6 y of age.

Alzheimer's disease is characterized by progressive dementia, cortical atrophy with synaptic loss, and the accumulation of neurofibrillary tangles and senile plaques containing beta-amyloid. The beta-amyloid protein precursor (beta-APP), may normally be involved in cell adhesion related to synaptic maintenance. Loss of synapses correlates with dementia, suggesting that synaptic deficits may underlie the disease. Synapse stability may depend on the action of tissue transglutaminase (tTG), an enzyme capable of crosslinking large, multi-domain extracellular glycoproteins, that is active and present at synapses. We now show that beta-APP is a substrate for tTG in vitro that results in dimers and multimers by silver staining and immunoblotting. This novel post-translational modification suggests further roles for beta-APP in synaptic function as well as in Alzheimer's disease.

BACKGROUND

Duodenal intraepithelial lymphocytosis (D-IEL) is an early marker for celiac disease (CD). However, the majority of cases are due to non-CD-related conditions.

OBJECTIVE

To identify the predictors of CD when presented with D-IEL.

METHODS

A total of 215 adult patients with D-IEL had undergone prospective and systematic evaluation for CD and other recognized associations.The gold-standard diagnosis of CD was based upon the presence of HLA-DQ2 and/or DQ8, persistence or progression of D-IEL following a gluten challenge, followed by symptomatic improvement on a gluten-free diet.Binary logistic regression models, adjusting for age and sex, were subsequently performed to compare presenting variables between CD and non-CD cases, and to determine their sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV).

RESULTS

CD was diagnosed in 48 cases (22%) and non-CD in 167 cases (78%). There was no statistical difference in baseline demographics, clinical symptoms (ie, diarrhea, weight loss, abdominal pain), anemia, or hematinics between the CD and non-CD group.Patients with CD, in comparison with non-CD, were significantly more likely to have a positive family history of CD (21% vs. 3.6%, OR 6.73; PPV 62.5%, NPV 81%, specificity 96.4%), positive HLA-DQ status (100% vs. 49.1%; PPV 36.4%, NPV 100%, specificity 50.9%), and presence of endomysial antibody (EMA) (48% vs. 0%; PPV 100%, NPV 87%, specificity 100%); all P≤0.001.A normal tissue transglutaminase antibody (TTG) level was seen in 29.2% CD and 83.2% non-CD cases (OR 0.084, P<0.001; PPV 9.2%). There was no difference in the prevalence of TTG levels 1 to 2×upper limit of normal (ULN) between the groups (29.2% CD vs. 14.4% non-CD; PPV 33% to 38%). However, TTG levels between 3 and 20×ULN were significantly more prevalent in the CD group (33.3% vs. 2.4%, PPV 66.6% to 89%), whereas a TTG)20×ULN was exclusive to CD (8.3%, P<0.001, PPV 100%).

CONCLUSIONS

In the setting of D-IEL, only the presence of a positive EMA or TTG)20×ULN at the outset can be used to make an immediate diagnosis of CD. Gastrointestinal symptoms, family history, anemia, or other celiac serology results do not reliably distinguish CD from non-CD without further investigations.

Celiac disease (CD) is an autoimmune disorder that is triggered by an immune response to gluten in genetically predisposed individuals. Although considered a primary gastrointestinal disease, CD is now known to have widespread systemic manifestations. We attempted to define the nature and role of systemic cytokine levels in the pathophysiology of CD. Multiplex cytokine assays were performed on four different groups of adult patients; patients with active CD (ACD), patients on a gluten-free diet (GFD) with positive TTG IgA antibodies, patients on a GFD with negative antibodies, and those with refractory CD (RCD). The results were compared with values in healthy adult controls. Patients with active CD and those on GFD with positive antibodies had significantly higher levels of proinflammatory cytokines, such as interferon-gamma, interleukin (IL)-1beta, tumor necrosis factor-alpha, IL-6 and IL-8, and also T(h)-2 cytokines such as IL-4 and IL-10, compared with normal controls and patients on GFD without antibodies. Interestingly patients on GFD for less than 1 year had significantly higher levels of both proinflammatory cytokines and T(h)2 cytokines compared with the patients on GFD for more than 1 year. In addition, a statistically significant correlation between levels of TTG IgA titers and serum levels of T(h)-2 cytokines IL-4 (p < 0.001), IL-10 (p < 0.001) and inflammatory cytokines such as IL-1alpha (p < 0.001), IL-1beta (p < 0.005), and IL-8 (p < 0.05) was observed.

OBJECTIVE

We evaluated the diagnostic performance of an ELISA test for anti-gliadin IgA and IgG antibodies, which uses synthetic deamidated gliadin peptides (anti-gliadin antibodies, AGAs) as coating; the results were compared with a test that uses extracted gliadin (AGAe).

METHODS

The study was conducted on the sera of 144 patients suffering from celiac disease (CD), including 20 patients with IgA deficiency and 9 who were following a gluten-free diet (GFD), and 129 controls.

RESULTS

In the 115 CD patients (without IgA deficiency), the sensitivity of AGAe IgA and IgG was 32.2 and 60.9%, whereas that of AGAs IgA and IgG was 59.1 and 72.2%. The specificity for AGAe IgA and IgG, and AGAs IgA and IgG was 93.8 and 89.9%, and 96.9% and 99.2%, respectively. Of the 20 patients with CD and IgA deficiency, 7 tested positive for AGAe IgG and 14 for AGAs IgG. The test using deamidated gliadin peptides performed better in terms of sensitivity and specificity than the AGA tests with extracted antigen.

CONCLUSIONS

The very high specificity of the AGAs IgG test (99.2%) also suggests that patients who test positive with this assay require a thorough followup, even if the anti-tissue transglutaminase antibodies (anti-tTG) and anti-endomysial autoantibodies (EMA) assays are negative.

We are using trichome formation on the plant Arabidopsis thaliana as a model for the study of plant cell determination and differentiation. Several of the genes that are required for trichome formation are defined by mutations. Two mutations, ttg and gl1, prevent the initiation of trichome differentiation. Thus, these mutations define products that are involved in the signalling of trichome determination. Other mutations, gl2, gl3, dis1, and dis2, define genes that are involved in trichome maturation. Our immediate goal has been to isolate the genes defined by these mutations and determine the role that they play in trichome formation. Our general goals are (1) to identify counterparts to these genes that are involved in other cell type determination and differentiation processes; (2) to manipulate cell development by altering the normal expression of these genes; and (3) to determine if this information can be used to improve crop plants. Presently, most of our progress has centered on the GL1 gene, which has been isolated and characterized. We have found that GL1 is a myb-related gene that is uniquely required for trichome initiation. As in other plants, Arabidopsis has a family of myb-related genes. We are currently investigating the possibility that some of these other myb-related genes are also uniquely required for other types of cell determination events.

The geodynamic environment in which Earth's first continents formed and were stabilized remains controversial. Most exposed continental crust that can be dated back to the Archaean eon (4 billion to 2.5 billion years ago) comprises tonalite-trondhjemite-granodiorite rocks (TTGs) that were formed through partial melting of hydrated low-magnesium basaltic rocks; notably, these TTGs have 'arc-like' signatures of trace elements and thus resemble the continental crust produced in modern subduction settings. In the East Pilbara Terrane, Western Australia, low-magnesium basalts of the Coucal Formation at the base of the Pilbara Supergroup have trace-element compositions that are consistent with these being source rocks for TTGs. These basalts may be the remnants of a thick (more than 35 kilometres thick), ancient (more than 3.5 billion years old) basaltic crust that is predicted to have existed if Archaean mantle temperatures were much hotter than today's. Here, using phase equilibria modelling of the Coucal basalts, we confirm their suitability as TTG 'parents', and suggest that TTGs were produced by around 20 per cent to 30 per cent melting of the Coucal basalts along high geothermal gradients (of more than 700 degrees Celsius per gigapascal). We also analyse the trace-element composition of the Coucal basalts, and propose that these rocks were themselves derived from an earlier generation of high-magnesium basaltic rocks, suggesting that the arc-like signature in Archaean TTGs was inherited from an ancestral source lineage. This protracted, multistage process for the production and stabilization of the first continents-coupled with the high geothermal gradients-is incompatible with modern-style plate tectonics, and favours instead the formation of TTGs near the base of thick, plateau-like basaltic crust. Thus subduction was not required to produce TTGs in the early Archaean eon.

GGTenul, a recently described genetic murine model of gamma-glutamyl transferase (GGT) deficiency, was induced by the point mutagen N-ethyl-N-nitrosourea and is inherited as an autosomal recessive trait. The phenotype of systemic GGT deficiency suggested a mutation site within the cDNA coding region which is common in all GGT transcripts. To identify this site, total lung and kidney RNA was isolated from normal and mutant mice, amplified by RT-PCR using GGT-specific primers, cloned as two overlapping approximately 1 kb GGT cDNA fragments, sequenced and compared with that in the literature. A single base pair substitution was identified in the coding region at position 237, where thymidine became adenine, and this mutation replaced a leucine codon, TTG, with a termination codon, TAG. This mutation site was confirmed in mutant genomic DNA by PCR using primers that flanked the predicted site and spanned the intron between the common GGT non-coding exon and the first GGT coding exon. This PCR product was sequenced directly with the secondary 3' PCR primer, the mutation site identified and the protocol then utilized to genotype animals. In addition to this mutation, the steady-state level of GGT mRNA in mutant kidney is reduced 3-fold compared with the control. Heterodimeric GGT protein is not detectable by western blot in either whole kidney homogenate or a microsomal membrane fraction. The steady-state mRNA level of gamma-glutatmyl cysteinyl synthetase was unchanged in mutant mice compared with normal, but that of heme oxygenase-1 and Cu,Zn-SOD was induced 4- and 3-fold, respectively. Hence, the GGTenul mouse model of GGT deficiency results from a single point mutation in the first coding exon of GGT mRNA and the resulting impairment in glutathione turnover induces oxidative stress in the kidney.

BACKGROUND

Tuberculosis (TB) is a major public health problem with high mortality and morbidity rates, especially in low-income countries. Disturbingly, the emergence of multidrug resistant (MDR) and extensively drug resistant (XDR) TB cases has worsened the situation, raising concerns of a future epidemic of virtually untreatable TB. Indeed, the rapid diagnosis of MDR TB is a critical issue for TB management. This study is an attempt to establish a rapid diagnosis of MDR TB by sequencing the target fragments of the rpoB gene which linked to resistance against rifampicin and the katG gene and inhA promoter region, which are associated with resistance to isoniazid.

METHODS

For this purpose, 133 sputum samples of TB patients from Morocco were enrolled in this study. One hundred samples were collected from new cases, and the remaining 33 were from previously treated patients (drug relapse or failure, chronic cases) and did not respond to anti-TB drugs after a sufficient duration of treatment. All samples were subjected to rpoB, katG and pinhA mutation analysis by polymerase chain reaction and DNA sequencing.

RESULTS

Molecular analysis showed that seven strains were isoniazid-monoresistant and 17 were rifampicin-monoresistant. MDR TB strains were identified in nine cases (6.8%). Among them, eight were traditionally diagnosed as critical cases, comprising four chronic and four drug-relapse cases. The last strain was isolated from a new case. The most recorded mutation in the rpoB gene was the substitution TCG>> TTG at codon 531 (Ser531 Leu), accounting for 46.15%. Significantly, the only mutation found in the katG gene was at codon 315 (AGC to ACC) with a Ser315Thr amino acid change. Only one sample harbored mutation in the inhA promoter region and was a point mutation at the -15p position (C>> T).

CONCLUSIONS

The polymerase chain reaction sequencing approach is an accurate and rapid method for detection of drug-resistant TB in clinical specimens, and could be of great interest in the management of TB in critical cases to adjust the treatment regimen and limit the emergence of MDR and XDR strains.

OBJECTIVE

Celiac disease, an autoimmune enteropathy that affects the proximal small intestine, is characteristically seen in people who have a genetic susceptibility to gluten sensitivity. Celiac patients' first-degree relatives are more at risk of acquiring the disease. The objective of the present study was consequently to determine the prevalence of celiac disease in a group of first-degree relatives of our patients with celiac disease.

METHODS

First-degree relatives of 195 patients with celiac disease attending a gastroenterology unit underwent serologic screening. Antitissue transglutaminase (anti-tTG) immunoglobulin A (IgA) and total serum IgA tests were used for first-level screening. Duodenal biopsy was recommended to subjects showing positive results to anti-tTG IgA testing. Biopsy samples were obtained by endoscopy, and biopsy specimens were evaluated and classified according to Marsh classification.

RESULTS

Positive anti-tTG IgA was found in 46 first-degree relatives (9.5%), whereas serum IgA levels were normal. Of 46 serology-positive relatives, 34 agreed to the endoscopy procedure. Histological changes characteristic of celiac disease were found in 23 subjects. The prevalence of celiac disease among the first-degree relatives was found to be at least 4.8%. Of 34 subjects that underwent biopsy, 11 were evaluated as Marsh 0, 5 as Marsh 1, 4 as Marsh 2, 12 as Marsh 3, and 2 as Marsh 4. Of the biopsy-positive subjects, 3 were mothers, 1 was a father, and 19 were siblings.

CONCLUSIONS

The present study identified 23 undiagnosed cases of celiac disease among 484 first-degree relatives of 195 patients with celiac disease, confirming the high prevalence (4.8%) of the disease in this specific group. It is suggested that an extensive screening policy be mandatory for these subjects.

The present study compared the genetic variation among 19 different isolates of Fasciola hepatica from cattle and sheep in different areas of Iran using sequence data for mitochondrial DNA gene, the subunit 1 of cytochrome C oxidase gene (CO1). Four different CO1 genotypes were detected among F. hepatica isolates that showed five variable nucleotide positions (accession nos.; GQ398051, GQ398052, GQ398053, GQ398054). Nucleotide sequence variation among 19 isolates for CO1 analyzed in this study ranged from 0% to 0.98% in Iran. Among the five polymorphism sites identified in this study, only one (T to G at position 51 in 5'end of GQ175362) resulted in putative amino acid alteration of phenylalanine (TTT) to leucine (TTG) in CO1. A phylogenetic analysis of the sequence data revealed that host associations and geographic location are likely not useful markers for Fasciola genotype classification. In addition, morphological analysis showed that the ratios of body length and body width of some (n = 5) of the 19 examined F. hepatica isolates were intermediate between F. hepatica and Fasciola gigantica, representing the substantial polymorphism of the F. hepatica species and the difficulty in the accurate recognition based on morphological features. In conclusion, Iranian F. hepatica exhibited the presence of considerable genetic diversity at CO1.

Systemic lupus erythematosus (SLE) and coeliac disease (CD) are diseases of an autoimmune origin that share the human leukocyte HLA-B8 and HLA-DR3 histocompatibility antigens, yet the co-association of CD with SLE is mainly based on case reports. Thus, the real prevalence of CD in SLE is unclear. The aim of this study was to determine the prevalence of antitissue transglutaminase (anti-tTG) in SLE and the relation between SLE and CD. In this case-control study, 100 patients with SLE, and 120 healthy subjects were studied. Sera from all participants were analysed for the presence of IgA and IgG anti-tTG antibodies using a human recombinant tissue transglutaminase (tTG) immuno-enzymatic assay. Anti-tTG positive patients and controls were further tested for antiendomysial (EMA) antibodies by an indirect immunofluorescence and HLA typing (DQalpha1*0501-DQbeta1*0201 allele determination). Subjects who had EMA or the mentioned allele, underwent duodenal biopsy to confirm a possible diagnosis of CD. Anti-tTG antibodies (IgA or IgG isotypes) were found in three of the 100 SLE patients (overall prevalence of 3%): one had the IgA and two the IgG isotypes. Only 1 of 120 healthy subjects (0.8%) had a low positive reaction for IgA anti-tTG. Only the IgA anti-tTG positive SLE patient was diagnosed as having CD based on a positive IgA-EMA and small bowel biopsy findings. The two IgG anti-tTG positive SLE patients and the IgA anti-tTG positive healthy subject were classified as false positives (EMA negative and HLA DQalpha1*0501-DQbeta1*0201 allele negative). In conclusion, anti-tTG antibodies were found at a low rate in SLE patients and mostly did not indicate the presence of CD. Thus, serological screening for CD is not recommended in SLE, unless a clinical suspicion of CD is present.

Schistosomiasis, one of the most devastating parasitic diseases, is caused by Schistosoma japonicum (Sj) infection resulting in serious liver fibrosis. Interleukin- (IL-) 13, which is produced by TH2 cells, is a critical profibrotic cytokine found in various organs, including the liver. Tissue transglutaminase (tTG), a group of multifunctional enzymes, serves a central function in the pathogenesis of chronic liver diseases. However, the relationship between IL-13, tTG, and liver fibrosis during Schistosoma infection has not been established. This study investigated the involvement of IL-13 and tTG in liver fibrogenesis during Sj infection in mice. Five weeks after Sj infection, granuloma and fibrosis development in the liver coincided with an increase in IL-13 and tTG in the liver and the upregulation of serum IL-13 in infected mice. Administration of cystamine, an inhibitor of tTG, abrogated the increase in both tTG and IL-13 in infected mice and ameliorated liver fibrogenesis and granuloma development. This result establishes a novel link among IL-13, tTG, and liver granuloma and fibrosis under Sj infection. Based on their important functions in liver fibrosis induced by Sj infection, IL-13 and tTG could be promising potential drug targets against schistosomiasis.

Gluten is known to be the main triggering factor for celiac disease (CeD), an immune-mediated disorder. CeD is therefore managed using a strict and lifelong gluten-free diet (GFD), the only effective treatment available currently. However, the GFD is restrictive. Hence, efforts are being made to explore alternative therapies. Based on their mechanisms of action on various molecular targets involved in the pathogenesis of CeD, these therapies may be classified into one of the following five broad approaches. The first approach focuses on decreasing the immunogenic content of gluten, using strategies like genetically modified wheat, intra-intestinal gluten digestion using glutenases, microwave thermal treatment of hydrated wheat kernels, and gluten pretreatment with either bacterial/ fungal derived endopeptidases or microbial transglutaminase. The second approach involves sequestering gluten in the gut lumen before it is digested into immunogenic peptides and absorbed, using binder drugs like polymer p(HEMA-co-SS), single chain fragment variable (scFv), and anti- gluten antibody AGY. The third approach aims to prevent uptake of digested gluten through intestinal epithelial tight junctions, using a zonulin antagonist. The fourth approach involves tissue transglutaminase (tTG) inhibitors to prevent the enhancement of immunogenicity of digested gluten by the intestinal tTG enzyme. The fifth approach seeks to prevent downstream immune activation after uptake of gluten immunogenic peptides through the intestinal mucosal epithelial layer. Examples include HLA-DQ2 blockers that prevent presentation of gluten derived- antigens by dendritic cells to T cells, immune- tolerizing therapies like the vaccine Nexvax2 and TIMP-Glia, cathepsin inhibitors, immunosuppressants like corticosteroids, azathioprine etc., and anti-cytokine agents targeting TNF-α and interleukin-15. Apart from these approaches, research is being done to evaluate the effectiveness of probiotics/prebiotics, helminth therapy using Necator americanus, low FODMAP diet, and pancreatic enzyme supplementation in CeD symptom control; however, the mechanisms by which they play a beneficial role in CeD are yet to be clearly established. Overall, although many therapies being explored are still in the pre-clinical phase, some like the zonulin antagonist, immune tolerizing therapies and glutenases have reached phase II/III clinical trials. While these potential options appear exciting, currently they may at best be used to supplement rather than supplant the GFD.

We have previously described that tissue transglutaminase (tTG) is a high level phenotypic biomarker in prostate cancer, which is down regulated in prostate cancer and surrounding premalignant field compared to benign prostate glands. To understand the function of tTG in prostate cancer, we sought to identify proteins that interact with the transglutaminase moiety of tTG using a human prostate cancer complementary deoxyribonucleic acid library in a Yeast 2-Hybrid system. The Yeast 2-Hybrid experiments identified a strong and novel interaction between the transglutaminase moiety and protein kinase A anchor protein 13 (AKAP13), which was quantified by beta-galactosidase assay, confirmed in vitro by immunoprecipitation experiments using PC3 prostate cancer cell lysates, and in vivo colocalization was confirmed by immunofluorescence studies in PC3 cells. Because AKAP plays a major role in protein kinase A and Rho protein mediated signaling, functional studies are underway to elucidate the significance of tTG-AKAP13 interaction in prostate cancer.

BACKGROUND

Diabetic nephropathy (DN) is the leading cause of chronic kidney failure, however the mechanisms underlying the characteristic expansion of the extracellular matrix (ECM) in diabetic kidneys remain controversial and unclear. In non-diabetic kidney scarring the protein crosslinking enzyme tissue transglutaminase (tTg) has been implicated in this process by the formation of increased epsilon-(gamma-glutamyl)lysine bonds between ECM components in both experimental and human disease. Studies in db+/db+ diabetic mice and in streptozotocin-treated rats have suggested a similar mechanism, although the relevance of this to human disease has not been addressed.

METHODS

We have undertaken a retrospective analysis of renal biopsies from 16 DN patients with type 2 diabetes mellitus using an immunohistochemical and immunofluorescence approach, with tTg and epsilon-(gamma-glutamyl)lysine crosslink quantified by confocal microscopy.

RESULTS

Immunofluorescent analysis of human biopsies (confocal microscopy) showed increases in levels of tTg (+1,266%, p < 0.001) and epsilon-(gamma-glutamyl)lysine (+486%, p < 0.001) in kidneys with DN compared to normal. Changes were predominantly in the extracellular periglomerular and peritubular areas. tTg staining correlated with epsilon-(gamma-glutamyl)lysine (r = 0.615, p < 0.01) and renal scarring (Masson's trichrome, r = 0.728, p < 0.001). Significant changes in epsilon-(gamma-glutamyl)lysine were also noted intracellularly in some (< or =5%) tubular epithelial cells. This is consistent with cells undergoing a novel transglutaminase-mediated cell death process in response to Ca2+ influx and subsequent activation of intracellular tTg.

CONCLUSIONS

Changes in tTg and epsilon-(gamma-glutamyl)lysine occur in human DN. Cellular export of tTg may therefore be a factor in the perpetuation of DN by crosslinking and stabilisation of the ECM, while intracellular activation may lead to cell death contributing towards tubular atrophy.

BACKGROUND

Duodenal biopsy may be unnecessary to confirm celiac disease in patients with high tissue-transglutaminase antibody level.

OBJECTIVE

To define a cut-off value of tissue-transglutaminase antibody with high positive likelihood ratio for duodenal atrophy in patients with suspected celiac disease.

METHODS

We retrospectively identified 945 patients with suspected celiac disease and classified according to the method used for tissue-transglutaminase antibody assay: Group A (n=393, Eu-tTG® Eurospital), Group B (n=263; Eu-tTG® Eurospital) and Group C (n=289; Celikey® Phadia). Duodenal histology was graded according to Marsh. Sensitivity, specificity, and positive likelihood ratio were used to evaluate cut-off points of tissue-transglutaminase antibody as predictor of villous atrophy.

RESULTS

100% specificity and ∞ positive likelihood ratio for duodenal atrophy was observed at a cut-off value of tissue-transglutaminase antibody 5 times higher than the upper limit of normal. CD diagnosis was confirmed by concordance with antiendomysial antibodies, and by reduction of t-TG titre in all patients and improvement of duodenal histology in 80% during gluten-free diet.

CONCLUSIONS

Tissue-transglutaminase antibody level 5-folds the upper limit of normal is 100% specific for duodenal atrophy and using this cut-off biopsy could by avoided in 1/3 of patients. Diagnostic criteria of celiac disease in adults need revision.

OBJECTIVE

Analysis of antibodies against tissue transglutaminase (tTG) has been shown valuable in the diagnosis of coeliac disease (CD) but how quickly serum titres decrease after introduction of a gluten-free diet (GFD) is not known in adults. CD is a well-recognized disorder amongst the general population and many persons try a GFD for fairly vague symptoms before they seek medical advice. Therefore, it is important to determine the time that the serologic tests remain predictive of the disease after the introduction of a GFD.

METHODS

Sera were taken from 22 consecutively biopsy-proven adult patients with CD in connection with the diagnostic biopsy. The patients were followed for 1 year and sera were taken after 1, 3, 6 and 12 months after start of a GFD. Sera were stored at -20 degrees C and analysed for IgA antibodies against gliadin, endomysium and two different commercial tTG assays based on recombinant human tTG (tTGrh) and guinea-pig liver (tTGgp).

RESULTS

Twenty patients could be followed during GFD and all antibody titres fell sharply within 1 month after introduction of a GFD and continued to decline during the survey interval. Thirty days after beginning the diet only 58, 84, 74 and 53% of all patients had positive antibody levels of tTGrh, tTGgp, EmA and AGA respectively.

CONCLUSIONS

As the antibodies used to confirm the diagnosis of CD fall rapidly and continue to decline following the introduction of a GFD, it is important that health care providers carefully inquire about the possibility of self-prescribed diets before patients sought medical attention.

In stressful conditions, bacteria enter into the viable but non-culturable (VBNC) state; in this state, they are alive but fail to grow on conventional media on which they normally grow and develop into colonies. The molecular basis underlying this state is unknown. We investigated the role of the alternative sigma factor RpoS (σ(38)) in the VBNC induction using Salmonella Dublin, Salmonella Oranienburg and Salmonella Typhimurium LT2. VBNC was induced by osmotic stress in LT2 and Oranienburg. Dublin also entered the VBNC state, but more slowly than LT2 and Oranienburg did. The LT2 rpoS gene was initiated from an alternative initiation codon, TTG; therefore, LT2 had smaller amounts of RpoS than Dublin and Oranienburg. Oranienburg had a single amino acid substitution (D118N) in RpoS (RpoS(SO)). Disruption of rpoS caused rapid VBNC induction. VBNC induction was significantly delayed by Dublin-type RpoS (RpoS(SD)), but only slightly by RpoS(SO). These results indicate that RpoS delays VBNC induction and that the rapid induction of VBNC in LT2 and Oranienburg may be due to lower levels of RpoS and to the D118N amino acid substitution, respectively. Reduced RpoS intracellular level was observed during VBNC induction. During the VBNC induction, Salmonella might regulate RpoS which is important for maintenance of culturablity under stresses.

Cerebral amyloid angiopathy (CAA) is a key histopathological hallmark of Alzheimer's disease (AD) and hereditary cerebral hemorrhage with amyloidosis of the Dutch type (HCHWA-D). CAA is characterized by amyloid-beta (Aβ) depositions and remodeling of the extracellular matrix (ECM) in brain vessels and plays an important role in the development and progression of both AD and HCHWA-D. Tissue transglutaminase (tTG) modulates the ECM by molecular cross-linking of ECM proteins. Here, we investigated the distribution pattern, cellular source, and activity of tTG in CAA in control, AD, and HCHWA-D cases. We observed increased tTG immunoreactivity and colocalization with Aβ in the vessel wall in early stage CAA, whereas in later CAA stages, tTG and its cross-links were present in halos enclosing the Aβ deposition. In CAA, tTG and its cross-links at the abluminal side of the vessel were demonstrated to be either of astrocytic origin in parenchymal vessels, of fibroblastic origin in leptomeningeal vessels, and of endothelial origin at the luminal side of the deposited Aβ. Furthermore, the ECM proteins fibronectin and laminin colocalized with the tTG-positive halos surrounding the deposited Aβ in CAA. However, we observed that in situ tTG activity was present throughout the vessel wall in late stage CAA. Together, our data suggest that tTG and its activity might play a differential role in the development and progression of CAA, possibly evolving from direct modulation of Aβ aggregation to cross-linking of ECM proteins resulting in ECM restructuring.

BACKGROUND

This study investigated whether a dot immunoassay (DIA) can provide simultaneous detection of anti-tissue transglutaminase (tTG), anti-deamidated gliadin (DG) and total IgA antibodies, as required in the work-up of celiac disease (CD) patients.

METHODS

Celiac disease patients (n=111) consecutively diagnosed from 2001 to 2011 at the Children's Hospital and Institute of Immunology (Technical University Dresden) were tested for anti-tTG, anti-DG and total IgA by enzyme-linked immunosorbent assay (ELISA) and DIA retrospectively. Blood donors (n=45) and non-CD individuals with low IgA serum levels (n=8) were included as controls. Antibodies to endomysial antigens (EmA) were assessed by indirect immunofluorescence (IIF).

RESULTS

Four (3.6%) of 111 CD patients demonstrated an IgA deficiency with total IgA below 50 mg/L by ELISA. Total IgA of the 107 IgA-non-deficient CD patients varied from 70 to 6000 mg/L. All four IgA-deficient CD patients were detected by a reduced reaction control of DIA and demonstrated positive anti-tTG or anti-DG IgG by DIA or ELISA. Detection of anti-tTG and anti-DG by DIA and ELISA showed a very good agreement (IgA: κ=0.972, 0.856, respectively; IgG: 0.921, 0.895, respectively).

CONCLUSIONS

Immunodot assay is a reliable and easy-to-use technique for the detection of IgA-deficient CD patients. Simultaneous assessment of anti-tTG and anti-DG IgA antibodies, and IgA deficiency by DIA can improve the efficacy of CD serology.

BACKGROUND

the extracellular matrix (ECM) is an important determinant of plaque instability. Since tissue transglutaminase (tTG) and elafin act as stabilizing factors, they might play a crucial role in the pathogenesis of acute coronary syndrome. We examined their expression in human coronary arteries and the regulation of tTG expression in cultured vascular smooth muscle cells (SMCs).

RESULTS

immunohistochemical studies on autopsy samples of human coronary arteries revealed the expression of tTG and elafin in the endothelium, medial SMCs, and the ECM in non-atherosclerotic coronary arteries. Their expression in SMCs, endothelium, and ECM was enhanced in atherosclerotic coronary arteries. In contrast, they were hardly detectable in accumulating macrophages or at the lipid core. Double staining demonstrated that elafin was co-localized with tTG. Moreover, some tTG-expressing cells were positive for TNF-alpha, suggesting that this cytokine might play an important role in the regulation of tTG. Treatment of cultured rat aortic SMCs with TNF-alpha increased their tTG mRNA, protein expression and enzyme activity.

CONCLUSIONS

the expression of tTG and elafin increased in atherosclerotic coronary arteries. The investigation with cultured SMCs suggested that TNF-alpha might mediate the upregulation of tTG. Our findings may provide new insights into the mechanism of plaque instability and the pathogenesis of acute coronary syndrome.

The mitochondrial genome of a flat bug, Brachyrhynchus hsiaoi (Blöte), is a typical circular DNA molecule of 15,250 bp with 37 genes and 70.4% A + T content. The gene order is different from that of the putative ancestral arrangement of insects with a tRNA gene rearrangement, trnQ-trnI. This rearrangement has been found in other sequenced flat bugs and is likely synapomorphic for the Aradidae or some subgroup within this family. Ten protein-coding genes start with ATN codon and others use TTG. All the 22 tRNAs, ranging from 61 to 70 bp, have the clover-leaf structure except for the dihydrouridine (DHU) arm of trnS1 forms a simple loop. The sizes of the large and small ribosomal RNA genes are 1245 and 808 bp, respectively. The control region is located between rrnS and trnQ with 703 bp in length and 69.8% A + T content.