Peptide IgG Research Articles

Caspase-1 inhibitor ameliorates experimental autoimmune myasthenia gravis by innate dendric cell IL-1-IL-17 pathway.

BackgroundIL-1β has been shown to play a pivotal role in autoimmunity. Cysteinyl aspartate-specific proteinase-1 (caspase-1) inhibitor may be an important drug target for autoimmune diseases. However, the effects of caspase-1 inhibitor on myasthenia gravis (MG) remain undefined.MethodsTo investigate the effects of caspase-1 inhibitor on experimental autoimmune myasthenia gravis (EAMG), an animal model of MG, caspase-1 inhibitor was administered to Lewis rats immunized with region 97–116 of the rat AChR α subunit (R97-116 peptide) in complete Freund’s adjuvant. The immunophenotypical characterization by flow cytometry and the levels of autoantibody by ELISA were carried out to evaluate the neuroprotective effect of caspase-1 inhibitor.ResultsWe found that caspase-1 inhibitor improved EAMG clinical symptom, which was associated with decreased IL-17 production by CD4+ T cells and γδ T cells, lower affinity of anti-R97-116 peptide IgG. Caspase-1 inhibitor decreased expression of CD80, CD86, and MHC class II on DCs, as well as intracellular IL-1β production from DCs. In addition, caspase-1 inhibitor treatment inhibited R97-116 peptide-specific cell proliferation and decreased follicular helper T cells relating to EAMG development.ConclusionsOur results suggest that caspase-1 inhibitor ameliorates experimental autoimmune myasthenia gravis by innate DC IL-1-IL-17 pathway and provides new evidence that caspase-1 is an important drug target in the treatment of MG and other autoimmune diseases.

Humoral Responses to Diverse Autoimmune Disease-Associated Antigens in Multiple Sclerosis.

To compare frequencies of autoreactive antibody responses to endogenous disease-associated antigens in healthy controls (HC), relapsing and progressive MS and to assess their associations with clinical and MRI measures of MS disease progression.MethodsThe study analyzed 969 serum samples from 315 HC, 411 relapsing remitting MS (RR-MS), 128 secondary progressive MS (SP-MS), 33 primary progressive MS (PP-MS) and 82 patients with other neurological diseases for autoantibodies against two putative MS antigens CSF114(Glc) and KIR4.1a and KIR4.1b and against 24 key endogenous antigens linked to diseases such as vasculitis, systemic sclerosis, rheumatoid arthritis, Sjogren’s syndrome, systemic lupus erythematosus, polymyositis, scleroderma, polymyositis, dermatomyositis, mixed connective tissue disease and primary biliary cirrhosis. Associations with disability and MRI measures of lesional injury and neurodegeneration were assessed.ResultsThe frequencies of anti-KIR4.1a and anti-KIR4.1b peptide IgG positivity were 9.8% and 11.4% in HC compared to 4.9% and 7.5% in RR-MS, 8.6% for both peptides in SP-MS and 6.1% for both peptides in PP-MS (p = 0.13 for KIR4.1a and p = 0.34 for KIR4.1b), respectively. Antibodies against CSF114(Glc), KIR4.1a and KIR4.1b peptides were not associated with MS compared to HC, or with MS disease progression. HLA DRB1*15:01 positivity and anti-Epstein Barr virus antibodies, which are MS risk factors, were not associated with these putative MS antibodies.ConclusionsAntibody responses to KIR4.1a and KIR4.1b peptides are not increased in MS compared to HC nor associated with MS disease progression. The frequencies of the diverse autoreactive antibodies investigated are similar in MS and HC.

Evaluating diagnostic accuracy of anti-tissue Transglutaminase IgA antibodies as first screening for Celiac Disease in very young children

BackgroundSmall bowel biopsy is the gold standard for Celiac Disease (CD) diagnosis, nevertheless serum assays are the first step in ascertaining a diagnosis of CD. New ESPGHAN Criteria 2012 (European Society of Pediatric Gastroenterology Hepatology and Nutrition) suggest using exclusively anti-tissue Transglutaminase IgA antibodies (anti-tTGA) as initial approach to symptomatic subjects. The aim of our study was to evaluate the diagnostic accuracy of anti-tTGA as initial screening assay for CD in a large cohort of pediatric patients. MethodsWe selected 730 subjects aged between 6months and 4years (“Group A”) and 348 subjects younger than 2years (which are part of the 730 subjects) (“Group B”). We performed anti-Deamidated Gliadin Peptides IgA and IgG antibodies (a-DGP IgA/IgG) and anti-tTGA assays by ELISA test. We evaluated the agreement between anti-tTGA and a-DGP IgA/IgG assays and compared the diagnostic accuracy of a-DGP IgA/IgG with that of anti-tTGA in both groups of patients. ResultsThere was a substantial agreement between anti-tTGA and a-DGP IgA in “Group A” and an almost perfect agreement in “Group B”; the strength of agreement between anti-tTGA and a-DGP IgG was moderate in “Group A” and substantial in “Group B”.anti-tTGA were more sensitive and specific than a-DGP IgA/IgG in both groups. Conclusionsanti-tTGA could be used as initial screening assay for CD in all subjects from 6months of age according to ESPGHAN Criteria 2012.

Rare Case of a Patient Presenting with Parathyroid Hormone-Related Peptide Mediated Hypercalcemia and IgG Heavy Chain Disease: Case Report and Review of Literature

IgG Heavy Chain Disease (γHCD) is a rare plasma cell disorder. Hypercalcemia related to plasma cell dyscrasias is related to non-PTHrP related mechanisms. Here we describe the first case of a patient with γHCD and PTHrP related hypercalcemia. Methods: Patient case derived from chart review from 2011 to 2015. Literature review performed searching PubMed 1968-current. Results: The patient was diagnosed with hypercalcemia with elevated PTHrP and exclusion of other etiologies of hypercalcemia. She was diagnosed with (γHCD) by M-spike 0.64 g/dL, IFE showing a broad band of IgG heavy chain, without associated light chains and severe depression of the non-mono-clonal IgG. Serum immunoglobulins demonstrated elevated IgG (2110 mg/dL), normal IgA (46 mg/dL) and decreased IgM (<21 mg/dL). Bone marrow biopsy showed 5% PCs, non-clonal by kappa/lambda, but exclusive for IgG by IHC, without any staining for IgA or IgM. The patient was started on therapy with improved hypercalcemia and PTHrP levels. Conclusions: This is the first reported case of γHCD presenting with PTHrP related hypercalcemia. Given that skeletal involvement is uncommon in γHCD, hypercalcemia secondary to γHCD may at times be a PTHrP driven phenomenon and we recommend that this test be ordered in such cases.

Celiac Disease Screening Rates Vary by Race in Pediatric Patients With Type 1 Diabetes Mellitus at a Tertiary Care Center

Introduction: Screening for celiac disease (CD) is advocated in high-risk groups such as patients with type 1 diabetes mellitus (T1D) who carry similar genetic risk factors. However, little is known about screening and serological positivity in African Americans (AAs) with T1D. The aim of this study was to compare the rates of CD screening and antibody positivity in pediatric African American (AA) and Caucasian (Ca) patients with T1D at a single tertiary care center. Methods: A retrospective chart review was performed for patients seen between February 1999 and June 2013. Medical records were abstracted according to ICD-9 codes for T1D. Patients aged 18 years and younger were included, and self-identified race was recorded. Results for tissue transglutaminase (TTG) IgA, endomysial (EMA) IgA, and deaminated gliadin peptide (DGP) IgA and IgG serologies were obtained. Demographic data, screening frequency, and serological positivity were compared between AA and Ca patients by Student’s t test, Fischer’s exact test, and Chi-squared analysis with a significance level of p<0.05. Results: A total of 935 patients with T1D aged 18 years and younger were identified. Of the 798 patients with data on race, there were 380 AA and 405 Ca patients with T1D. There were no differences in mean age between AA and Ca, though AA patients were more commonly female (58.7% vs. 49.9%, p=0.015). The overall screening rate with any antibody test was 23.1%. Testing rates for TTG IgA (45/380 in AA vs. 180/405 in Ca, p<0.0001), EMA IgA (7/380 vs. 36/405, p<0.0001), DPG IgA (4/380 vs. 36/405, p<0.0001), and DGP IgG (4/380 vs. 34/405, p<0.0001) were all significantly lower in AA versus Ca. Positivity rates for TTG IgA (3/45 vs. 30/180, p=0.10), EMA IgA (0/7 vs. 13/36, p=0.08), DGP IgA (0/4 vs. 7/36, p=1.0), and DGP IgG (1/4 vs. 13/34, p=1.0) were not significantly different between the 2 groups. Conclusion: Despite recommendations to screen patients with T1D for CD, we found low screening rates in a pediatric T1D population overall. AA patients with T1D were significantly less likely to be screened than Ca patients. Interestingly, there were no differences in rates of serological positivity between AA and Ca, though this may be limited by a small sample size. Reasons for lower screening in AA T1D patients are not known but might be due to differences in perception of disease risk, symptomatology, lack of knowledge about screening recommendations, or referral bias. This study suggests the need for improvement in CD screening for pediatric patients with T1D and better estimates of CD prevalence in AA patients with T1D.

Can celiac serology alone be used as a marker of duodenal mucosal recovery in children with celiac disease on a gluten-free diet?

Assessment of treatment response in children with celiac disease (CD) after commencing a strict gluten-free diet (GFD) is generally based on the resolution of clinical features and normalization of serology. Recent adult studies have shown that serologic markers do not correlate with mucosal recovery. We aimed (i) to determine whether anti-tissue transglutaminase immunoglobulin (Ig)A (tTG) and anti-deamidated gliadin peptide IgG (DGP) antibodies are sensitive and specific markers of mucosal recovery in children with CD on a GFD for at least 12 months, and (ii) to determine whether a validated dietary questionnaire of compliance can identify patients with mucosal recovery. A total of 150 children with biopsy-proven CD were prospectively evaluated with duodenal biopsies at ≥12 months on GFD, paired with repeat tTG and DGP serology. The biopsies were reviewed in a blinded manner by two histopathologists and graded by Marsh criteria. A validated questionnaire of dietary compliance was also administered. Of 150 children recruited, 27 (18%) had positive serology, 97 (65%) had negative serology, and 26 (17%) had equivocal serology. Of the 97 children with negative serology, none had Marsh type 3 enteropathy. Of the 27 patients with positive serology, only 6 had Marsh type 3 changes. The sensitivity and specificity of serology as a marker of significant mucosal pathology was 75 and 85%, respectively, with a positive predictive value of 22% but a negative predictive value of 98%. Of the 129 (86%) questionnaires completed, 88% reported good or excellent compliance with a GFD (negative predictive value 97%). This study suggests that follow-up using two serological tests in children with CD on a GFD may obviate the need for repeat mucosal biopsy in the majority of patients. A standardized dietary questionnaire may be useful in identifying patients who require further evaluation.

Type 1 diabetes and celiac disease: clinical overlap and new insights into disease pathogenesis.

Type 1 diabetes (T1D) and celiac disease (CD) are autoimmune diseases with clinical and pathogenic overlap. The mean prevalence of CD in patients with T1D is about 8 %. Classic intestinal symptoms of CD may not be present in T1D leading to the recommendation for active case finding in this higher risk group. Screening is done with sensitive and specific serologies including tissue transglutaminase (tTG) IgA and deaminated gliadin peptide (DGP) IgA and IgG. Positive serologies are confirmed by the presence of villous atrophy and increased intraepithelial lymphocytes on duodenal biopsy. A strict gluten free diet is recommended, although this can pose challenges for T1D patients who already have dietary restrictions. In aggregate, it appears as if the gluten free diet may help T1D management. T1D and CD have overlapping genetic and environmental risk factors. Among these, non-HLA genetic factors and the gut microbiome are among recent developments that will be discussed in this review.

Ultrasound Detection of Myocardial Ischemic Memory Using an E-Selectin Targeting Peptide Amenable to Human Application

Vascular endothelial leukocyte adhesion molecules, such as E-selectin, are acutely upregulated in myocardial ischemia/reperfusion and are thus "ischemic memory" biomarkers for recent cardiac ischemia. We sought to develop an ultrasound molecular imaging agent composed of microbubbles (MBs) targeted to E-selectin to enable the differential diagnosis of myocardial ischemia in patients presenting with chest pain of unclear etiology. Biodegradable polymer MBs were prepared bearing a peptide with specific human E-selectin affinity (MBESEL). Control MBs had scrambled peptide (MBCTL) or nonspecific IgG (MBIgG). MBESEL adhesion to activated rat endothelial cells (ECs) was confirmed in vitro in a flow system and in vivo with intravital microscopy of rat cremaster microcirculation. Ultrasound molecular imaging of recent myocardial ischemia was performed in rats 4 hours after transient (15 minutes) coronary occlusion. MBESEL adhesion was higher to inflamed versus normal ECs in vitro; there was no difference in MBCTL or MBIgG adhesion to inflamed versus normal ECs. There was greater adhesion of MBESEL to inflamed versus noninflamed microcirculation and minimal adhesion of MBCTL or MBIgG under any condition. Ultrasound imaging after injection of MBSEL demonstrated persistent contrast enhancement of the previously ischemic region. Videointensity in postischemic myocardium after MBESEL was higher than that in the nonischemic bed (11.6 ± 2.7 dB vs 3.6 ± 0.8 dB, p < .02) and higher than that after MBCTL (4.0 ± 1.0 dB, p < .03) or MBIgG (1.7 ± 0.1 dB, p < .03). MBs targeted to E-selectin via a short synthetic peptide with human E-selectin binding affinity enables echocardiographic detection of recent ischemia, setting the stage for clinical myocardial ischemic memory imaging to identify acute coronary syndromes.

Dynamic and equilibrium performance of sensors based on short peptide ligands for affinity adsorption of human IgG using surface plasmon resonance

This paper characterizes the potential of novel hexameric peptide ligands for on-line IgG detection in bioprocesses. Surface Plasmon Resonance (SPR) was used to study the binding of human IgG to the hexameric peptide ligand HWRGWV, which was covalently grafted to alkanethiol self-assembled monolayers (SAM) on gold surfaces. Peptide coupling on SAMs was verified, followed by covalent grafting of peptides with a removable Fmoc or acetylated N-termini via their C-termini to produce active peptide SPR sensors that were tested for IgG binding. The dynamics and extent of peptide–IgG binding were compared with results from a conventional system using protein A attached on a gold surface via disulfide monolayers. IgG binding to protein A on disulfide monolayers yielded equilibrium dissociation constants of 1.4×10–7M. The corresponding dissociation constant value for the acetylated version of the peptide (Ac-HWRGWV) supported on alkanethiol SAM was 5.8×10–7M and that for HWRGWV on the alkanethiol SAM (after de-protection of Fmoc-HWRGWVA) was 1.2×10–6M. Maximum IgG binding capacities, Qm of 6.7, 3.8, and 4.1mgm−2 were determined for the protein A and the two forms of HWRGWV-based biosensors, respectively. Real-time data for the kinetics of adsorption were used to determine the apparent rate constants for adsorption and desorption. The results were analyzed to understand the mechanism of IgG binding to the protein and peptide ligands. It was found that the peptide–IgG binding was reaction controlled, however the protein A–IgG binding mechanism was partially mass transfer (diffusion) controlled. The adsorption rate constants, ka, for the protein A ligand increased with decreasing concentration of analyte and the peptide ligand ka values was constant at different IgG concentrations and flow rates.

A 10-year Retrospective Study of Celiac Disease (CD) and other Gluten Sensitive Patients at the University of California at San Diego (UCSD)

Purpose: To assess diagnostic testing of gluten sensitive patients seen at UCSD. Methods: We retrospectively evaluated patients seen at UCSD in the past 10 years diagnosed with CD or NCGS according to Olso criteria(1). Demographic factors, lab studies and pathology reports were reviewed. All serological testing analyzed was obtained while patients were consuming gluten. Results: Three major categories were identified; celiac disease (CD) with modified Marsh 3 histology [n=116, 73% female (F)]; non-celiac gluten sensitive (NCGS) with CD-like symptoms without histological CD while consuming gluten [n=68, 75%F]; and undiagnosed patients with gluten sensitive symptoms, without biopsies obtained prior to avoiding dietary gluten [n=32, 81%F]. Mean ages were similar in the 3 groups. Tissue transglutaminase (TTG) IgA levels were compared to modified Marsh 3 histology. Sensitivity/specificity of TTG IgA IgA>19 AU were 81%/93%, respectively. When TTG IgA was three times greater than the upper limit of normal (19 AU), sensitivity/specificity were 74%/100%. PPV and NPV were 100% and 75%, respectively. We determined how often deamidated gliadin peptide (DGP) IgA and IgG correctly predicted CD when TTG IgA levels were not elevated. When TTG IgA was <20AU in CD patients (n=12), two had DGP IgA>19, three had DGP IgG>19. Combined sensitivity of TTG IgA and DGP IgA or DGP IgG was 84% or 85%, respectively; specificity was 93% or 91%. At UCSD, a TTG IgA order set includes a reflexive measurement of endomysial antibody (EMA). We determined how often this changed management using Cohen's kappa to measure the level of agreement between the two tests. This value was 0.908 (P <0.001) indicating high agreement. We examined how often TTG IgA levels were predictive of IgA deficiency. A Spearman's rank correlation was 0.29 (P=0.009), indicating a statistically significant relationship. The average value of TTG IgA in CD patients with IgA deficiency was 1.33 AU, significantly lower than the average value of TTG IgA in those with normal IgA levels, 54.9 AU (P< 0.001). Conclusion: Gender and mean age were similar in the three different categories of gluten sensitive patients at UCSD. Specificity of TTG IgA was comparable to previous reports, while sensitivity was lower. As expected, using TTG IgA levels three times greater than the upper limit of normal decreased sensitivity but specificity was 100%. Adding DGP IgA/G testing to TTG IgA enhanced the detection of CD only slightly. We showed that reflexive EMA testing did not improve detection of CD beyond TTG IgA alone. Very low TTG IgA levels were predictive of IgA deficiency.

Prevalence of Small Bowel Bacterial Overgrowth (SIBO) in Decompensated Cirrhosis with Portal Hypertension: A Clinical Pilot Study

Purpose: Decompensated cirrhosis is associated with portal hypertension and its complications, like spontaneous bacterial peritonitis (SBP), hepato renal syndrome, and minimal and overt encephalopathy (HE), which accelerate mortality and affect quality of life (QOL). Portal hypertension causes transmural edema and dysbiosis, due to altered splanchnic hemodynamics promoting translocation of microbes across the bowel wall, causing SBP and precipitating HE (with substantially reduced survival rates). This study evaluates the prevalence of SIBO in decompensated cirrhotics (without overt HE) and the recurrence of SBP, both leading to a challenging clinical course. Methods: Total of 120 patients were recruited. Group A: (n=30) healthy controls, group B: (n=45) chronic liver disease {(ALD 12/45 (27%), NASH 5/45 (11%), HCV 18/45 (40%)}, group C: (n=45) decompensated cirrhosis with mean MELD of 16. Laboratories studies included: Stool H. pylori antigen, ova and parasites, initial lactoferrin, calprotectin, serum ammonia, anti-sacromyces cervecesiae antibody IgA and IgG, tissue transglutaminase antibody IgA and IgG, endomysial antibody IgA and IgG, de aminated gliadine peptide IgG, and breath resting for SIBO lactose at base-line and after three months. Group C underwent abdominal sonogram-guided baseline paracentesis, which was repeated in 3 months to evaluate the recurrence of SBP in SIBO. Exclusion: Diabetes mellitus, renal failure, IBS, celiac, IBD, parkinsonism, idiopathic dysmotility disorders, hepatic encephalopathy, TIPS, drugs-gut specific antibiotics, anti-cholinergics, tricyclics, methadone, narcotics, and PPis. Results: Group A 2/30 (6%) were positive for SIBO on breath testing, and 1/30 (3%) ASCA IgA-positive. Group B 5/45 (11%) were positive on breath testing, and 4/45 (8%) were ASCA IgA-positive. Group C: 33/45 (73%) were positive on breath testing, and 38/45 (84%) were ASCA IgA-positive. Recurrence of SBP was 28/33 (85%). Conclusion: This observational clinical trial reveals SIBO to be associated with decompensated cirrhosis. SBP with SIBO has independent morbidity with recurrence. Prophylactic antibiotics should be considered to avoid recurrent SBP and encephalopathy in cirrhotics with SIBO. A larger study needs to validate the role of SIBO in decompensated cirrhotics with portal hypertension.

PK modulation of haptenylated peptides via non-covalent antibody complexation

We applied noncovalent complexes of digoxigenin (Dig) binding antibodies with digoxigeninylated peptide derivatives to modulate their pharmacokinetic properties. A peptide derivative which activates the Y2R receptor was selectively mono-digoxigeninylated by reacting a NHS-Dig derivative with an ε-amino group of lysine 2. This position tolerates modifications without destroying receptor binding and functionality of the peptide. Dig-peptide derivatives can be loaded onto Dig-binding IgGs in a simple and robust reaction, thereby generating peptide–IgG complexes in a defined two to one molar ratio. This indicates that each antibody arm becomes occupied by one haptenylated peptide. In vitro receptor binding and signaling assays showed that Dig-peptides as well as the peptide–antibody complexes retain better potency than the corresponding pegylated peptides. In vivo analyses revealed prolonged serum half-life of antibody-complexed peptides compared to unmodified peptides. Thus, complexes are of sufficient stability for PK modulation. We observed more prolonged weight reduction in a murine diet-induced obesity (DIO) model with antibody-complexed peptides compared to unmodified peptides. We conclude that antibody–hapten complexation can be applied to modulate the PK of haptenylated peptides and in consequence improve the therapeutic efficacy of therapeutic peptides.

Atorvastatin-modified dendritic cells in vitro ameliorate experimental autoimmune myasthenia gravis by up-regulated Treg cells and shifted Th1/Th17 to Th2 cytokines

Conventional therapies for autoimmune diseases produce nonspecific immune suppression, which are usually continued lifelong to maintain disease control, and associated with a variety of adverse effects. In this study, we found that spleen-derived dendritic cells (DCs) from the ongoing experimental autoimmune myasthenia gravis (EAMG) rats can be induced into tolerogenic DCs by atorvastatin in vitro. Administration of these tolerogenic DCs to EAMG rats on days 5 and 13 post immunization (p.i.) resulted in improved clinical symptoms, which were associated with increased numbers of CD4(+)CD25(+) T regulatory (Treg) cells and Foxp3 expression, decreased lymphocyte proliferation among lymph node mononuclear cells (MNC), shifted cytokine profile from Th1/Th17 to Th2 type cytokines, decreased level of anti-R97-116 peptide (region 97-116 of the rat acetylcholine receptor α subunit) IgG antibody in serum. These tolerogenic DCs can migrate to spleen, thymus, popliteal and inguinal lymph nodes after they were injected into the EAMG rats intraperitoneally. Furthermore, these tolerogenic DCs played their immunomodulatory effects in vivo mainly by decreased expression of CD86 and MHC class II on endogenous DCs. All these data provided us a new strategy to treat EAMG and even human myasthenia gravis (MG).

Comparison between enzyme-linked immuno-sorbent assay and multiplex assay for serological markers of Celiac Disease detection

Event Abstract Back to Event Comparison between enzyme-linked immuno-sorbent assay and multiplex assay for serological markers of Celiac Disease detection Lydia Martín-Martín1*, Laura Herráiz-Nicuesa1, Yaciris Oleo-Pérez1, Silvia Sánchez-Ramón2 and Margarita Rodriguez-Mahou1 1 Hospital General Universitario Gregorio Marañón, Spain 2 Hospital Clínico San Carlos, Spain Objective: To compare ELISA with multiplex assay for anti-deamidated gliadin peptide (anti-DGP) IgG and anti-tissue transglutaminase (anti-tTG) IgA antibodies detection in order to identify the best according to sensivity and specificity. Methods: 101 patients with suspicion or diagnostic of celiac disease (CD) were selected. Each serum was tested by ELISA (Orgentec Diagnostika) and multiplex assay (Bioplex 2200, Bio-Rad) for the anti-DGP IgG and anti-tTG IgA autoantibodies quantification. The association with clinical features was examinated by systematical review of the clinical histories. Also previous CD markers determinations, biopsy and HLA features were studied. Results: Compared to the clinical, biopsy and HLA features, the ELISA showed a sensitivity of 96.15% and a specificity of 38.09 (Positive predictive value (PPV): 79.36%; Negative predictive value (NPV): 80%), whereas multiplex assay had a sensitivity of 79.59% and a specificity of 100% (PPV: 100%; NPV: 68.75%) for anti-DGP IgG antibodies detection. Furthermore, the ELISA showed a sensitivity of 68.42% and a specificity of 100% (PPV: 100%; NPV: 93.18%), and multiplex assay had a sensitivity of 100% and a specificity of 96.34% (PPV: 86.36%; NPV: 100%) for anti-tTG IgA antibodies testing. Conclusions: Taking into consideration the previous CD markers determination and clinical, biopsy and HLA features, ELISA showed a higher sensitivity for anti-DGP IgG antibodies, but specificity was better for multiplex assay. In addition, ELISA showed a higher specificity and multiplex had a better sensitivity for anti-tTG IgA antibodies detection. Keywords: Celiac Disease, ELISA, Multiplex assay, Anti-deamidated gliadin peptide IgG antibodies, Anti-tissue transglutaminase IgA antibodies Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Immune-mediated disease pathogenesis Citation: Martín-Martín L, Herráiz-Nicuesa L, Oleo-Pérez Y, Sánchez-Ramón S and Rodriguez-Mahou M (2013). Comparison between enzyme-linked immuno-sorbent assay and multiplex assay for serological markers of Celiac Disease detection. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.01057 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 10 Jul 2013; Published Online: 22 Aug 2013. * Correspondence: Miss. Lydia Martín-Martín, Hospital General Universitario Gregorio Marañón, Madrid, Spain, lydia.martin@salud.madrid.org Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Lydia Martín-Martín Laura Herráiz-Nicuesa Yaciris Oleo-Pérez Silvia Sánchez-Ramón Margarita Rodriguez-Mahou Google Lydia Martín-Martín Laura Herráiz-Nicuesa Yaciris Oleo-Pérez Silvia Sánchez-Ramón Margarita Rodriguez-Mahou Google Scholar Lydia Martín-Martín Laura Herráiz-Nicuesa Yaciris Oleo-Pérez Silvia Sánchez-Ramón Margarita Rodriguez-Mahou PubMed Lydia Martín-Martín Laura Herráiz-Nicuesa Yaciris Oleo-Pérez Silvia Sánchez-Ramón Margarita Rodriguez-Mahou Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Profiling anti-cyclic citrullinated peptide antibodies in patients with juvenile idiopathic arthritis

BackgroundAnti-citrullinated protein/peptide antibodies (ACPA), have high specificity for rheumatoid arthritis (RA). Some children with juvenile idiopathic arthritis (JIA), phenotypically resemble RA and test positive for rheumatoid factor (RF) a characteristic biomarker of RA. We investigated the prevalence of ACPA and its relationship to other serologic markers associated with RA in a well-characterized JIA cohort.MethodsCases were 334 children with JIA, 30 of whom had RF + polyarticular JIA. Sera from all cases and 50 healthy pediatric controls were investigated by ELISA at a single time point for anti-cyclic citrullinated peptide (anti-CCP) IgG, RF IgM, IgA and IgG, anti-RA33 IgG, and antinuclear antibodies (ANA). Comparisons between cases and controls were made using Chi-square or Fisher exact tests and T-tests.ResultsThe prevalence of RF was 8% among controls, and 12% among cases (ns). The prevalence of ACPA was 2% in controls and 14.3% in cases (OR 8.2, p <0.01). Children who were ACPA-positive and RF-negative (n = 23) had a significantly earlier onset-age (4.6 years vs. 12.1 years, p <0.00001) and had fewer HLA-DRB1 shared epitope alleles than those positive for both RF and ACPA (n = 25). Prevalence of anti-RA33 was not different between cases and controls.ConclusionsACPAs are detectable in 14% of children with JIA. Children with positive ACPA but negative RF are frequent, and may define a distinct subset of children with JIA. ACPA testing should be included in the classification of JIA.

Diagnostic Value of Anti‐deamidated Gliadin Peptide IgG Antibodies for Celiac Disease in Children and IgA‐deficient Patients

The aim of the study was to analyze the diagnostic performance of anti-deamidated gliadin peptide (dGp) immunoglobulin (Ig) G and IgA regarding the age at celiac disease (CD) diagnosis and the anti-dGp IgG usefulness for diagnosing CD IgA-deficient patients. Anti-dGp IgG and IgA and anti-native gliadin (nGlia) IgA were determined by enzyme fluoroimmunoassay in 100 newly diagnosed anti-tissue transglutaminase (tTG) IgA-positive pediatric and adult patients with CD and in 100 age-matched patients with other digestive pathologies. Anti-dGp IgG was evaluated in 6 CD IgA-deficient patients. When analyzing all of the patients, the anti-dGp IgG assay showed higher diagnostic accuracy (area under receiver operating characteristic curve), specificity, and positive predictive value than anti-dGp IgA and anti-nGlia IgA. All of the diagnostic parameters corresponding to anti-dGp IgG reached the same values as anti-tTG IgA in children 7 years or younger. In older patients, both anti-dGp isotypes showed an inverse behavior, IgG having a higher specificity and positive predictive value but a lower sensitivity and negative predictive value than IgA. Anti-dGp levels were associated with the severity of intestinal lesions, and an inverse association was found regarding age at diagnosis. Both anti-dGp IgG and IgA were found to be positive in the 9 patients with minimal intestinal changes included in the study. All of the patients with CD with IgA deficiency were positive for anti-dGp IgG. The diagnostic performance of anti-dGp depends on the antibody isotype and on the age at CD diagnosis, anti-dGp IgG being a serological marker at least as useful as anti-tTG IgA for detecting CD in children ages 7 years or younger. Our data also indicate that anti-dGp IgG can improve the diagnosis of IgA-deficient patients.

Sa1345 Increased Duodenal IL12A mRNA Expression in Celiac Disease Patients Might Lead to Increased IL-35 Production

BACKGROUND. Celiac disease (CD) is an autoimmune enteropathy that affects 1% of the general population. The association between CD and other autoimmune disorders is well established and increased prevalence of CD has been found in patients with type 1 diabetes (7%) and autoimmune thyroiditis (2%). Some reports on co-morbidity of CD and alopecia areata (AA), an autoimmune skin disorder that causes hair loss, have shown a variable hair re-growth upon institution of a gluten-free diet. No studies have addressed the association between CD and the different clinical subtypes of AA, which vary in duration and degree of hair loss. The aim of this study was to investigate the prevalence of CD across the phenotypic spectrum of AA using serological screening methods that have high sensitivity and specificity for CD diagnosis. METHODS. Sera obtained through the National Alopecia Areata Registry of the United States from 16 patients with transient AA (AAT), 24 with patchy persistent AA (AAP), 27 with total scalp hair loss (AT) and 32 with complete body hair loss (AU), were screened for CD using a commercial ELISA kit against combined antitissue transglutaminase and anti-deamidated gliadin peptide IgA and IgG antibodies (antitTG/DGP) from INOVA Diagnostics Inc. Titers >20 IU were considered positive and titers >30 IU were considered strongly positive and suggestive of CD. RESULTS. Of 99 samples (females=64, males=35), a total of 9 (9%) patients (all female) were positive for anti-tTG/ DGP antibodies (range 20.4 61.7 IU) and 4% (2 AT, 1 AU and 1 AAP) were strongly positive. By type of presentation, anti-tTG/DGP titers were elevated in 15% AT, 9% AU and 8% AAP patients but in none of AAT patients. CONCLUSIONS. These results demonstrate that AA patients have a higher prevalence of CD-associated antibodies (9%) than the general population (~3%). CD seropositivity is present in the more severe forms of AA, where hair loss is experienced for more than one year and/or results in total loss of hair from the scalp and/or body. Patients with the least severe form of the disease, transient hair loss lasting less than one year, do not appear to have an elevated risk for CD. Screening for CD is recommended in AA for early detection and institution of a gluten-free diet.

Atorvastatin inhibits the inflammatory response caused by anti-M3 peptide IgG in patients with primary Sjögren’s syndrome

Experimental and clinical investigations have revealed that statins can down-regulate acute and chronic inflammatory processes. Whether statins express anti-inflammatory activities in the salivary glands in patients with primary Sjögren's syndrome (pSS) is not known. The in vitro and in vivo effect of atorvastatin on rat submandibular gland treated with anti-M(3) peptide IgG purified from SS patients was studied. The anti-inflammatory effects of atorvastatin were assessed by measuring the levels of IL-1β, PGE(2) and MMP-3 by ELISA. Atorvastatin inhibited the increase in the production of IL-1β, PGE(2) and MMP-3 in submandibular glands treated with anti-M(3) peptide IgG. A positive correlation between IL-1β production with accumulation of PGE(2) and MMP-3 was observed. Rats pre-treated orally with atorvastatin (30 mg kg(-1)) or vehicle (phosphate-buffered solution) once a day for three consecutive days impaired the increment in the production of IL-1β, PGE(2) and MMP-3 in the submandibular gland in the presence of anti-M(3) peptide IgG. In conclusion, the anti-inflammatory effects of atorvastatin are dependent upon inhibition of production of a pro-inflammatory cytokine (IL-1β) and pro-inflammatory mediators such as PGE(2) and MMP-3. These data suggest that atorvastatin may constitute an anti-inflammatory effect in SS.

Anti-M3 peptide IgG from Sjögren’s syndrome triggers apoptosis in A253 cells

Primary Sjögren’s syndrome (pSS) is an autoimmune disease that targets salivary and lachrymal glands, characterized by anti-cholinergic autoantibodies directed against the M3 muscarinic acetylcholine receptor (mAChR). The aim of this work was to evaluate if cholinergic autoantibodies contained in IgG purified from Sjögren sera could trigger apoptosis of A253 cell line. We also determined if caspase-3 and matrix metalloproteinase-3 (MMP-3) are involved in the induction of A253 cell death. Our results demonstrated that anti-cholinergic autoantibodies stimulate apoptosis and inositol phosphate (InsP) accumulation accompanied by caspase-3 activation and MMP-3 production. All of these effects were blunted by atropine and J104794, indicating that M3 mAChRs are impacted by the anti-cholinergic autoantibodies. The intracellular pathway leading to autoantibody-induced biological effects involves phospholipase C (PLC), calcium/calmodulin (CaM) and extracellular calcium as demonstrated by treatment with U-73122, W-7, verapamil, BAPTA and BAPTA-AM, all of which blocked the effects of the anti-cholinergic autoantibodies. In conclusion, anti-cholinergic autoantibodies in IgG purified from pSS patient’s sera mediates apoptosis of the A253 cell line in an InsP, caspase-3 and MMP-3 dependent manner.

Gene Therapy Research at the Frontiers of Viral Immunology

Gene Therapy Research at the Frontiers of Viral Immunology