Lymph node fibroblastic reticular cells preserve a tolerogenic niche in allograft transplantation through laminin α4.

Lymph node (LN) fibroblastic reticular cells (FRCs) define LN niches and regulate lymphocyte homeostasis through producing diverse extracellular matrix (ECM) components. We examined the role of ECM laminin α4 (Lama4) using FRC-Lama4 conditional KO Pdgfrb-Cre-/- × Lama4fl/fl mice. Single-cell RNA-sequencing (scRNA-Seq) data showed the promoter gene Pdgfrb was exclusively expressed in FRCs. Depleting FRC-Lama4 reduced Tregs and dendritic cells, decreased high endothelial venules, impaired the conduit system, and downregulated T cell survival factors in LNs. FRC-Lama4 depletion impaired the homing of lymphocytes to LNs in homeostasis and after allografting. Alloantigen-specific T cells proliferated, were activated to greater degrees in LNs lacking FRC-Lama4, and were more prone to differentiate into effector phenotypes relative to the Treg phenotype. In murine cardiac transplantation, tolerogenic immunosuppression was not effective in FRC-Lama4 recipients, which produced more alloantibodies than WT. After lung transplantation, FRC-Lama4-KO mice had more severe graft rejection with fewer Tregs in their LNs. Overall, FRC-Lama4 critically contributes to a tolerogenic LN niche by supporting T cell migration, constraining T cell activation and proliferation, and promoting Treg differentiation. Hence, it serves as a therapeutic target for immunoengineering.

Celiac Disease and Celiac Antibodies in DM1 Patients: When Are Screening and Biopsy Recommended?

INTRODUCTION: The prevalence of celiac disease (CD) is increased in diabetes mellitus type 1 (DM1) patients. In most cases, CD is diagnosed in asymptomatic patients and hence periodic screening tests are recommended, but the timing, frequency of tests and indication for duodenal biopsy is unclear. The purpose of this study was to investigate the dynamics of CD serology in DM1 and identify risk factors for CD. METHODS: Celiac serology and duodenal biopsy results from 1990 until 2015 were collected from patients with DM1. The outcome of positive celiac serology, the incidence and risk factors for CD in DM1 patients were investigated. RESULTS: A total of 314 DM1 patients who had celiac serology were identified, with follow-up period up to 23 years. Of 31 patients (9.9%) with positive celiac serology, 11(35.4%) had spontaneous normalization after various time periods. Eighteen patients were diagnosed with CD (58.1% of positive celiac serology, 5.73% of the study cohort). Age under 4.5 years was a risk factor for CD, but not family background of autoimmune diseases or gender. All patients with CD diagnosis were diagnosed during the first 6 years following DM1 diagnosis. CONCLUSION: Screening asymptomatic DM1 patients for CD beyond 6 years after diagnosis is not recommended. Spontaneous normalization of CD serology occurs, and hence, serologic follow-up may be performed. In children with DM1 diagnosis under the age of 4.5 years or with positive CD serology at DM1 diagnosis, there is an increased risk for CD and therefore positive serology should lead to biopsy.

Testing for antireticulin antibodies in patients with celiac disease is obsolete: a review of recommendations for serologic screening and the literature.

Celiac disease (CD) is an autoimmune disorder that occurs in genetically susceptible individuals of all ages and is triggered by immune response to gluten and related proteins. The disease is characterized by the presence of HLA-DQ2 and/or -DQ8 haplotypes, diverse clinical manifestations, gluten-sensitive enteropathy, and production of several autoantibodies of which endomysial, tissue transglutaminase, and deamidated gliadin peptide antibodies are considered specific. Although antireticulin antibodies (ARA) have historically been used in the evaluation of CD, these assays lack optimal sensitivities and specificities for routine diagnostic use. This minireview highlights the advances in CD-specific serologic testing and the rationale for eliminating ARA from CD evaluation consistent with recommendations for diagnosis.

[Serological diagnosis of celiac disease]. / Diagnostic sérologique de la maladie cœliaque.

Screening studies using high-sensitivity and specificity markers indicate a prevalence of celiac disease of up to 1% in European and North-American populations. Celiac disease is a frequent condition that has become an important public health issue. Yet the majority of cases remain undiagnosed due to the polymorphism of its clinical manifestations. The new insight in the pathogenesis of celiac disease has lead to the development of new diagnostic tools. Early screening of symptomatic patients and pre-identified at-risk groups significantly improves the quality of life while reducing morbidity and mortality. However, prophylactic benefits of early diagnosis by assessing the general population have not been shown in any study. French and Northern American scientific societies have introduced serological testing in their newly revised strategies to diagnose celiac disease. Older markers judged insufficiently accurate like anti-gliadin and anti-reticulin antibodies have recently been withdrawn from the list of reimbursed medical expenses in France. Anti-endomysium and tissue transglutaminase IgA antibodies have proven to be at this day the most sensitive and specific markers for the diagnosis and follow-up of patients on gluten-free diet, at the exception of IgA-deficient patients. Assays testing for IgG antibodies are recommended upon IgA-deficiency. Although very accurate, a better standardisation of current assays may enable serological testing to replace in a near future histological confirmation brought by small bowel biopsies which remains today the gold standard test to diagnose celiac disease. Indeed, serological testing represents and attractive alternative as it is less invasive, less expansive, laboursaving and more objective in interpretation.

The histological characteristics of the aggregated lymphoid nodules area in abomasum of Bactrian camels (Camelus bactrianus) of different ages.

The aggregated lymphoid nodules area (ALNA) in abomasum of Bactrian camels is a special immune structure discovered only in Bactrian camels in recent years (2003). The anatomy research found that there was a close relationship between degree of development, anatomical characteristics and age. To further establish the relationship between histological characteristics of this special structure and animal age, 24 Alashan Bactrian camels of the following four age groups were studied: young (1-2 years), pubertal (3-5 years), middle-aged (6-16 years) and old (17-20 years). Mucosal-associated lymphoid tissue (MALT) of ALNA in abomasum was particularly observed and analyzed by histology, histochemistry and statistical methods. The results showed that the average number of lymphoid nodules in reticular mucosal folds region of ALNA in abomasum from young group to old group was in order of 26.8, 32.7, 17.6 and 7.8, and in longitudinal mucosal folds region was 20.1, 26.0, 10.3 and 5.1. The number of lymphoid nodules in the four experimental groups first increased and then decreased with increasing age (P<0.01). In young and pubertal camels lymphoid nodules were distributed evenly on both sides of the axis of mucosal folds and mostly displayed round, oval or wedge shape. The number of lymphoid nodules, follicle-associated epithelium (FAE), reticular fibers and plasmocytes in mucosal folds gradually increased from 1 to 2 years and peaked at puberty. There were up to 37 visible lymphoid nodules in a mucosal fold. However, ALNA of middle-aged and old camels gradually degenerated as aging. Lymphoid nodules were unevenly distributed on both sides of the axis of mucosal folds, which mostly displayed oval or irregular shape. Lymphoid tissue in old camels mostly existed as diffuse form. Although germinal centers of the lymphoid nodules were still obvious, the number of reticular fiber and plasmocyte and lymphoid nodules gradually decreased. The results indicated that in accord with the anatomical results, there was a close relationship between histology characteristics of lymphoid tissue of ALNA in abomasum and animal age. In summary, the lymphoid tissue of ALNA in abomasums gradually increased from young to pubertal groups with increasing age, peaked in 3-5 year-old camels, and subsequently declined with age and when 17-20 years old this immunity structure had severely atrophied.

HLA-DQ types of celiac disease in Libyan children with type 1 diabetes mellitus.

OBJECTIVE: To determine the genetic profile of celiac disease (CD) in Libyan children with type 1 diabetes as there are no data on the frequency of human leukocyte antigen (HLA)-related CD-predisposing genes in diabetic patients in Libya. METHODS: We randomly studied 218 Libyan type 1 diabetic children. The mean age was 12.2±4.6 years; 56% were female patients. The mean duration of diabetes was 4.7±4.0 years. All patients were screened for CD with IgA tissue-transglutaminase (tTG) and endomysium antibodies. Patients with positive immunological screen were programmed for a small-bowel biopsy. HLA-DRB1* and HLA-DQB1* were genotyped in all tTG-positive patients. RESULTS: Twenty-seven (12.4%) out of 218 patients with type1 diabetes had positive tTG, and 20 (9.2%) of these patients were positive for endomysium antibodies. Five patients (5/27) were already known cases of biopsy-proven CD. Biopsy was not performed in two patients. One biopsy result was normal, whereas 19 biopsies demonstrated morphological changes consistent with CD. Forty-eight percent of the anti-tTG-positive group were homozygous for HLA-DQ2, whereas 75% of biopsy-proven CD patients had HLA-DQ2, 21% had HLA-DQ2/DQ8, and 4% had HLA-DQ8. In addition, the majority (70%) carried HLA-DQ2 linkage with HLA-DRB1*03. CONCLUSION: Overall, biopsy-confirmed prevalence of CD was 11% (24 of 218). The present study confirms that CD in the Libyan type 1 diabetic population is high when compared with European and US studies, and for the first time we document that this population shares similar HLA-DQ2 genotype. This supports the theory regarding the role of the environment as an important factor in CD development in this part of the world.

T-cell movement on the reticular network.

The idea that the apparently random motion of T cells in lymph nodes is a result of movement on a reticular network (RN) has received support from dynamic imaging experiments and theoretical studies. We present a mathematical representation of the RN consisting of edges connecting vertices that are randomly distributed in three-dimensional space, and models of lymphocyte movement on such networks including constant speed motion along edges and Brownian motion, not in three-dimensions, but only along edges. The simplest model, in which a cell moves with a constant speed along edges, is consistent with mean-squared displacement proportional to time over intervals long enough to include several changes of direction. A non-random distribution of turning angles is one consequence of motion on a preformed network. Confining cell movement to a network does not, in itself, increase the frequency of cell-cell encounters.

The lymph node revisited: development, morphology, functioning, and role in triggering primary immune responses.

Scenarios have been proposed to explain how lymphoid components of a lymph node favor the encounter of a drained antigen with a circulating competent naïve lymphocyte to trigger a primary immune response. However, these scenarios rest on incorrect concepts about the organ. This situation resulted from a loss of interest for studies on in vivo lymphoid organs due to a widespread switch, decades ago, to work on suspended lymphoid cells. However, an in vivo holistic study of the organ continued in our laboratory. The present review synthesizes resulting knowledge on lymph node morphology and global functioning. We show that the opening of an afferent lymphatic vessel into the subcapsular sinus is the focal point from which the related portion of a lymph node-a node compartment-is developed. As to the formation of a compartment's lymphoid components, it is neonatally orchestrated by the dichotomic nature and distribution of antigens in this subcapsular sinus, which determines a dichotomic recruitment of circulating cells and the compartment's architectural complexity. The transport process of an antigen from a given tissue territory into restricted sites of the draining compartment further defines its local morphological features and activities, while providing the possibility to reduce the wandering of a short-lived naïve cell through innumerable target-devoid sites. We also explain that the nodal lymphoid components are not implicated in the triggering of primary responses, but are rather products of such responses. Scenarios for the triggering of primary responses, consistent with real node morphology and functioning, are proposed.

Diagnostic methods beyond conventional histology in coeliac disease diagnosis.

BACKGROUND: Coeliac disease diagnostic criteria currently require the detection of small bowel mucosal villous atrophy and crypt hyperplasia. AIMS: To compare conventional histological examination to the determination of small bowel mucosal intraepithelial lymphocytes (IELs) and to serum and intestinal coeliac autoantibodies in untreated coeliac disease with villous atrophy and in mild enteropathy coeliac disease. PATIENTS AND METHODS: Study comprised consecutive adult patients with coeliac disease suspicion; villous height-crypt depth ratio (Vh/CrD), the densities of CD3+, gammadelta+ and villous tip IELs and serum and intestinal transglutaminase 2 (TG2)-targeted autoantibodies were studied. Coeliac disease was diagnosed in 223 and excluded in 608 patients. Further, 66 patients were considered to suffer from mild enteropathy coeliac disease. Control group consisted of 138 patients. RESULTS: Vh/CrD determination detected 77% of untreated coeliac disease patients. Serum coeliac autoantibodies had 84% sensitivity for untreated coeliac disease with villous atrophy and 70% sensitivity for mild enteropathy coeliac disease; the specificity was 100%. Intestinal TG2-targeted autoantibodies had sensitivities of 100% and 93%, and 100% specificity, respectively. gammadelta+ and villous tip IELs proved more reliable than CD3+ IELs. CONCLUSIONS: Conventional histological examination as the golden standard in coeliac disease diagnosis is questionable. Serum and especially intestinal TG2-targeted autoantibodies seem promising in future coeliac disease diagnostics.

[Specific immune response in diagnosis of celiac disease with chronic diarrhea].

High concentration of Ab Gl, Ab tTG, Ab Em and Ab Rt and increased amount of immunocompetent cells in the SIM in coeliac disease is an index of the developing specific immune response to gliadin and tissue antigens. High specificity and sensitivity of Ab tTG and Ab Em were determined in celiac disease.

The conduit system of the lymph node.

The lymphoid compartment of lymph nodes is impermeable to many molecules that are delivered via afferent lymphatic vessels. In the lymphoid compartment, fibroblast reticular cells form an interconnected network-the conduit system. This network has a structural function supporting tightly packed lymphocytes and antigen-presenting cells; however, it also has an important function as a molecular sieve, since it contains tubules that are the only entry point for fluid and allow only small molecules and particles (including antigens) to flow along the network. This size exclusion may prevent pathogens entering the blood from lymph. Dendritic cells can sample antigens from the conduit system and present them to nearby lymphocytes; this may be particularly important in initiating immune responses. The importance of larger antigen transport via macrophages or other cells is unclear. Lymphocytes and antigen-presenting dendritic cells actively move and interact along the conduit system, perhaps in response to chemokines or cytokines transported by the conduit system; these molecules may also be transported to high endothelial venules and regulate the attraction of blood leukocytes to the lymph nodes. The conduit system is also important for fluid distribution between afferent lymphatics and blood, but the mechanisms are not yet established.

[Positive tests for serum anti-endomysium antibodies (AEA) and anti-jejunal antibodies (JAB) and histopathological diagnosis of celiac disease in children]. / Pozitivita sérových protilátok proti endomýziu, jejunu a histopatologická diagnostika celiakie u detí.

Non-invasive examination methods are increasingly important in diagnosing celiac disease. New options for diagnosing celiac disease have been discovered in addition to the established biochemical, hematological and other methods as a result expansive progress ofclinical genetics and immunology. At the same time, detection of circulating auto-antibodies is becoming ever more frequent in clinical practice. As a result, many new, clinically highly heterogeneous cases of the disease have been diagnosed and consequently the prevalence of the disease in both child and adult population has grown. Detection of anti-endomysial antibodies (AEA), characteristic for their high sensitivity and specificity, plays an important role in diagnosing and monitoring celiac disease in pediatric practice. Nevertheless, histopathological diagnosis remains the critical tool for definitive diagnosis of the disease. The article refers to relations between the degree of positivity of AEA and JAB antibodies in the IgA class and the respective grade in the Marsh grading system. The objective of the study was to examine AEA and JAB antibodies and the histological picture of the duodenal mucosa in 20 children and adolescents with celiac disease aged from 2 to 18 years. The authors developed a semiquantitative scale of positivity of both the antibodies, which they compared trying to find a correlation between these and the histopathological picture of the duodenal mucosa. The authors point out the need of timely determination of AEA and t-TG (tissue transglutaminase) in patients whose anamnesis, clinical picture or laboratory results may be indicative of celiac disease.

Performance of a new rapid whole blood coeliac test in adult patients with low prevalence of endomysial antibodies.

BACKGROUND: In coeliac disease endomysial and transglutaminase autoantibodies are directed against the human autoantigen, transglutaminase. The conventional coeliac antibody tests are performed from serum samples in centralized laboratories. AIMS: To evaluate a rapid and easy immunoglobulin A-class whole blood point-of-care test and its commercial application, the Biocard test, in coeliac autoantibody detection. METHODS: In the whole blood point-of-care test transglutaminase is liberated from the red blood cells by haemolysis. Transglutaminase antibodies, if present, complex with the liberated antigen, and are visualized. Altogether 51 biopsy-proven untreated coeliac adult patients, 48 of the same patients after treatment, and 36 controls were tested. The point-of-care test results were compared with serum endomysial and transglutaminase antibody and Biocard test results and histology. RESULTS: The whole blood point-of-care test was as sensitive (82%) as the serum endomysium test (80%) in detecting untreated coeliac disease while the serum transglutaminase antibody test was superior (88%). The tests had 100% specificity. A positive point-of-care test result seroconverted or the test reaction weakened in 90% of the treated coeliac patients. Biocard test-positive were 22 of the 24 tested untreated coeliac patients. Biocard test-negative were 15 of 19 controls. CONCLUSIONS: The whole blood rapid tests are as reliable as the conventional serological tests in detecting untreated coeliac disease and in coeliac disease diet monitoring.

Fate of five celiac disease-associated antibodies during normal diet in genetically at-risk children observed from birth in a natural history study.

OBJECTIVES: To explore the natural history of antibodies against tissue transglutaminase (TGA), endomysium (EMA), reticulin (ARA), and gliadin (AGA-IgG and AGA-IgA) in children carrying HLA-conferred risk for celiac disease (CD) and observed frequently from birth. METHODS: TGA was measured in serum samples obtained between years 2000 and 2003 from 1,320 children carrying genetic CD risk. If a sample was TGA positive, all five antibodies were analyzed in all banked and forthcoming samples from that child, and a duodenal biopsy was recommended. At the end of this observation, in August 2004, the age of the children was from 1 to 9.5 yr (mean 4.1 yr). RESULTS: Forty-nine children (3.7%) were TGA positive. In these children, AGA-IgG had emerged at the mean age (+/- SD, range) of 2.0 +/- 1.5, 0.5-6.6 yr, while TGA, EMA, and ARA all emerged concurrently somewhat later (TGA at 3.2 +/- 1.5, 1.0-7.0 yr, P < 0.001 when compared to AGA-IgG). Despite continuing gluten exposure, positive TGA, EMA, ARA, AGA-IgA, and AGA-IgG values were spontaneously lost in 49%, 45%, 43%, 41%, and 32% of the children, respectively. CD was diagnosed by biopsy in 20 of the 26 TGA-positive children who consented to a biopsy. CONCLUSIONS: Potential CD trigger(s) other than only gluten probably function before AGA-IgG emerges, i.e., > or =3 months earlier than the transglutaminase-associated antibodies appear. In a remarkable proportion of the children, antibodies disappear spontaneously suggesting that regulatory immune phenomena under favorable circumstances are able to extinguish incipient CD in genetically at-risk children even without exclusion of gluten from the diet.

Rheumatoid Arthritis and anti-endomysial antibodies.

BACKGROUND: Several autoimmune diseases may occur in the same patient. Celiac disease (CD) is found in patients with diabetes mellitus type-1 and thyroiditis. Few studies have addressed the association between CD and rheumatic disorders such as rheumatoid arthritis (RA). OBJECTIVE: To study the prevalence of anti-endomysial antibodies in RA patients. METHODS: The presence of IgA anti-endomysial antibodies (EmA-IgA) was evaluated in 85 RA patients and 97 healthy controls through indirect immunoflourescence using human umbilical cord as substrate. RESULTS: None of the RA patients or healthy controls was positive for EmA-IgA. CONCLUSION: We could not find an association between RA and anti-endomysial antibodies in the studied population.

Prevalence of IgA-antiendomysial antibody in a patient cohort with idiopathic low bone mineral density.

AIM: To investigate the frequency of serum IgA-antiendomysial antibody positivity in patients with low bone mineral density and to assess the risk group for screening of celiac disease. METHODS: One hundred and thirty-five patients (14 male, 121 female) with idiopathic low bone mineral density were evaluated. The median age was 57.2 years (24-81). Antiendomysial antibody was determined by the immunofluorescence method using a commercial kit (INOVA Diagnostics Inc., CA, USA), which employs a 5 microm thin cryostat section of monkey esophagus as a substrate. RESULTS: Of the 135 patients evaluated, 13 were found to have positive IgA antiendomysial antibody test (9.6%) response. None of the patients had IgA deficiency. Endoscopic appearance and histological examination were normal in all of these patients. Seropositive patients had significantly lower age (48.9 +/- 4.3 vs 59.2 +/- 6.2, P < 0.05), higher ratio of male gender (61.5% vs 4.9%, P < 0.01) and pre-menopausal status (8.7% vs 1.3%, P < 0.01). Lumbar spine and femoral neck z-scores, but not t-scores were significantly lower in seropositive patients. Seropositive patients had lower serum 25 (OH) vitamin D, calcium and higher serum parathormone levels than seronegative patients. CONCLUSION: The screening of celiac disease in idiopathic osteoporosis should be restricted to patients without classical risk factors (younger, pre-menopausal, male gender) for osteoporosis. Bone mineral density measurements using z-scores should be considered for identifying risk groups for celiac disease.

Oral mucosa of coeliac disease patients produces antiendomysial and antitransglutaminase antibodies: the diagnostic usefulness of an in vitro culture system.

BACKGROUND: Antiendomysial (EmA) and antitransglutaminase (anti-tTG) antibodies are the most specific indirect marker of coeliac disease (CD). It is not known whether the oral mucosa of patients with CD is able to produce these antibodies or not. AIMS: To evaluate the ability of the oral mucosa of patients with CD to produce antibodies in an in vitro culture system. PATIENTS AND METHODS: Twenty-eight patients with new diagnosis of CD (15 adults and 13 children) and 14 adult subjects with other diseases (controls) were studied. All underwent oral mucosa biopsy and subsequent EmA and anti-tTG assays on the mucosa culture medium. RESULTS: Sensitivity and specificity of EmA and anti-tTG assayed in the oral mucosa culture medium for CD diagnosis were 54% and 100% and 57% and 100%, respectively. The CD clinical presentation, such as the presence of oral mucosa lesions, did not influence the results of the EmA and anti-tTG assays in the oral mucosa culture medium. There was an association between positivity of antibodies and greater severity of the oral mucosa lymphocyte infiltration. CONCLUSION: This study demonstrates that the oral mucosa contributes to EmA and anti-tTG production in untreated patients with CD.