Detection of antibodies directed to the N-terminal region of GAD is dependent on assay format and contributes to differences in the specificity of GAD autoantibody assays for type 1 diabetes

Autoantibodies to glutamate decarboxylase (GADA) are sensitive markers of islet autoimmunity and type 1 diabetes. They form the basis of robust prediction models and are widely used for recruitment of subjects at high risk of type 1 diabetes to prevention trials. However GADA are also found in many individuals at low risk of diabetes progression. To identify the sources of diabetes irrelevant GADA reactivity therefore, we analyzed data from the 2009 and 2010 Diabetes Autoantibody Standardization Program GADA workshop and found that binding of healthy control sera varied according to assay type. Characterization of control sera found positive by radiobinding assay, but negative by ELISA showed that many of these sera reacted to epitopes in the N-terminal region of the molecule. This finding prompted development of an N-terminally truncated GAD65 radiolabel, (35)S-GAD65(96-585), which improved the performance of most GADA radiobinding assays (RBAs) participating in an Islet Autoantibody Standardization Program GADA substudy. These detailed workshop comparisons have identified a source of disease-irrelevant signals in GADA RBAs and suggest that N-terminally truncated GAD labels will enable more specific measurement of GADA in type 1 diabetes

Autoantibodies toward ATP4A and ATP4B subunits of gastric proton pump H+,K+-ATPase are reliable serological pre-endoscopic markers of corpus atrophic gastritis

INTRODUCTION: Noninvasive assessment of corpus atrophic gastritis (CAG), a condition at increased risk of gastric cancer, is based on the measurement of pepsinogens, gastrin, and Helicobacter pylori antibodies. Parietal cell autoantibodies (PCAs) against the gastric proton pump (ATP4) are potential serological biomarkers of CAG. The purpose of this study was to compare the diagnostic performance of PCA and pepsinogen I tests in patients with clinical suspicion of CAG with the histopathological evaluation of gastric biopsies as reference standard. METHODS: A prospective case-finding study was performed on 218 naive adult patients (131 women, median age 65 years) who underwent gastric biopsies to confirm/exclude CAG. Patients with histopathological CAG were defined as cases, conversely as controls. Autoantibodies against the individual alpha (ATP4A) and beta (ATP4B) subunits of ATP4 were measured by luciferase immunoprecipitation, and global PCA and pepsinogen I by enzyme-linked immunosorbent assay. RESULTS: Histopathology classified 107 subjects (49%) as cases (CAG+, autoimmune 81.2%, and multifocal extensive 18.8%) and 111 subjects (51%) as controls (CAG−). In cases, ATP4A, ATP4B, and PCA titers were increased compared with controls, whereas pepsinogen I was reduced (P < 0.0001 for all). ATP4B, ATP4A, and pepsinogen I tests showed sensitivities of 77%, 75%, and 73% and specificities of 88%, 88%, and 80%, respectively. The receiver operating characteristic (ROC) area under the ROC curve (AUC) of these serological biomarkers confirmed their ability to discriminate cases from controls (ATP4B = 0.838, ATP4A = 0.826, pepsinogen I = 0.775, and PCA = 0.805), whereas the partial ROC-pAUC90 analysis showed that the ATP4B test had the best diagnostic performance (P = 0.008 vs ATP4; P = 0.0002 vs pepsinogen I). The presence of autoimmune or extensive gastritis was not significantly different between ATP4B positive or negative cases (P = 0.217). DISCUSSION: PCAs are promising serological biomarkers for the identification of CAG in high-risk individuals, particularly in an autoimmune pattern but also in an extensive-multifocal atrophy pattern

The first 142 amino acids of glutamate decarboxylase do not contribute to epitopes recognized by autoantibodies associated with Type 1 diabetes

Human adenovirus (HAdV) E1B-55K is a multifunctional regulator of productive viral replication and oncogenic transformation in nonpermissive mammalian cells. These functions depend on E1B-55K's posttranslational modification with the SUMO protein and its binding to HAdV E4orf6. Both early viral proteins recruit specific host factors to form an E3 ubiquitin ligase complex that targets antiviral host substrates for proteasomal degradation. Recently, we reported that the PML-NB associated factor Daxx represses efficient HAdV productive infection and is proteasomally degraded via a SUMO-E1B-55K-dependent, E4orf6-independent pathway, the details of which remained to be established. RNF4, a cellular SUMO-targeted ubiquitin ligase (STUbL), induces ubiquitinylation of specific SUMOy lated proteins and plays an essential role during DNA repair. Here, we show that E1B-55K recruits RNF4 to the insoluble nuclear matrix fraction of the infected cell to support RNF4/Daxx association, promoting Daxx PTM and thus inhibiting this antiviral factor. Removing RNF4 from infected cells using RNA interference resulted in blocking the proper establishment of viral replication centers and significantly diminished viral gene expression. These results provide a model for how HAdV antagonize the antiviral host responses by exploiting the functional capacity of cellular STUbLs. Thus, RNF4 and its STUbL function represent a positive factor during lytic infection and a novel candidate for future therapeutic antiviral intervention strategies.IMPORTANCE Daxx is a PML-NB-associated transcription factor that was recently shown to repress efficient HAdV productive infection. To counteract this antiviral measurement during infection, Daxx is degraded via a novel pathway including viral E1B-55K and host proteasomes. This virus-mediated degradation is independent of the classical HAdV E3 ubiquitin ligase complex, which is essential during viral infection to target other host antiviral substrates. To maintain a productive viral life cycle, HAdV E1B-55K early viral protein inhibits the chromatin-remodeling factor Daxx in a SUMO-dependent manner. In addition, viral E1B-55K protein recruits the STUbL RNF4 and sequesters it into the insoluble fraction of the infected cell. E1B-55K promotes complex formation between RNF4-and E1B-55K-targeted Daxx protein, supporting Daxx posttranslational modification prior to functional inhibition. Hence, RNF4 represents a novel host factor that is beneficial for HAdV gene expression by supporting Daxx counteraction. In this regard, RNF4 and other STUbL proteins might represent novel targets for therapeutic intervention

A novel LIPS assay for insulin autoantibodies

Insulin autoantibodies (IAA) are often the first marker of autoimmunity detected in children in the preclinical phase of type 1 diabetes (T1D). Currently, the vast majority of laboratories adopt the radiobinding micro-assay (RBA) for measuring IAA. Our aim was to replace RBA with a novel non-radioactive IAA Luciferase Immuno Precipitation System (LIPS) assay with improved performance. We developed (pro)insulin antigens with alternative placements of a NanoLuc (TM) luciferase reporter (NLuc). Performance in LIPS was evaluated by testing sera from new onset T1D (n = 80), blood donors (n = 123), schoolchildren (n = 186), first-degree relatives (FDRs) from the Bart's Oxford family study (n = 53) and from the Belgian Diabetes Registry (n = 136), coded sera from the Islet Autoantibody Standardization Program (IASP) (T1D n = 50, blood donors n = 90). IAA LIPS based on B chain-NLuc proinsulin or B chain-NLuc insulin, in which NLuc was fused at the C-terminus of the insulin B chain, required only 2 mu L of serum and a short incubation time, showed high concordance with RBA (Spearman r = 0.866 and 0.833, respectively), high assay performance (B chain-NLuc proinsulin ROC-AUC = 0.894 and B chain-NLuc insulin ROC-AUC = 0.916), and an adjusted sensitivity at 95% specificity ranking on par with the best assays submitted to the two most recent IASP workshops. In FDRs, the IAA LIPS showed improved discrimination of progressors to T1D compared to RBA. We established a novel high-performance non-radioactive IAA LIPS that might replace the current gold standard RBA and find wide application in the study of the IAA response in T1D

Implementation and performances of the IPbus protocol for the JUNO Large-PMT readout electronics

The Jiangmen Underground Neutrino Observatory (JUNO) is a large neutrino detector currently under construction in China. Thanks to the tight requirements on its optical and radio-purity properties, it will be able to perform leading measurements detecting terrestrial and astrophysical neutrinos in a wide energy range from tens of keV to hundreds of MeV. A key requirement for the success of the experiment is an unprecedented 3% energy resolution, guaranteed by its large active mass (20 kton) and the use of more than 20,000 20-inch photo-multiplier tubes (PMTs) acquired by high-speed, high-resolution sampling electronics located very close to the PMTs. As the Front-End and Read-Out electronics is expected to continuously run underwater for 30 years, a reliable readout acquisition system capable of handling the timestamped data stream coming from the Large-PMTs and permitting to simultaneously monitor and operate remotely the inaccessible electronics had to be developed. In this contribution, the firmware and hardware implementation of the IPbus based readout protocol will be presented, together with the performances measured on final modules during the mass production of the electronics

Mass testing of the JUNO experiment 20-inch PMTs readout electronics

The Jiangmen Underground Neutrino Observatory (JUNO) is a multi-purpose, large size, liquid scintillator experiment under construction in China. JUNO will perform leading measurements detecting neutrinos from different sources (reactor, terrestrial and astrophysical neutrinos) covering a wide energy range (from 200 keV to several GeV). This paper focuses on the design and development of a test protocol for the 20-inch PMT underwater readout electronics, performed in parallel to the mass production line. In a time period of about ten months, a total number of 6950 electronic boards were tested with an acceptance yield of 99.1%

Validation and integration tests of the JUNO 20-inch PMTs readout electronics

The Jiangmen Underground Neutrino Observatory (JUNO) is a large neutrino detector currently under construction in China. JUNO will be able to study the neutrino mass ordering and to perform leading measurements detecting terrestrial and astrophysical neutrinos in a wide energy range, spanning from 200 keV to several GeV. Given the ambitious physics goals of JUNO, the electronic system has to meet specific tight requirements, and a thorough characterization is required. The present paper describes the tests performed on the readout modules to measure their performances.Comment: 20 pages, 13 figure

Potential of Core-Collapse Supernova Neutrino Detection at JUNO

JUNO is an underground neutrino observatory under construction in Jiangmen, China. It uses 20kton liquid scintillator as target, which enables it to detect supernova burst neutrinos of a large statistics for the next galactic core-collapse supernova (CCSN) and also pre-supernova neutrinos from the nearby CCSN progenitors. All flavors of supernova burst neutrinos can be detected by JUNO via several interaction channels, including inverse beta decay, elastic scattering on electron and proton, interactions on C12 nuclei, etc. This retains the possibility for JUNO to reconstruct the energy spectra of supernova burst neutrinos of all flavors. The real time monitoring systems based on FPGA and DAQ are under development in JUNO, which allow prompt alert and trigger-less data acquisition of CCSN events. The alert performances of both monitoring systems have been thoroughly studied using simulations. Moreover, once a CCSN is tagged, the system can give fast characterizations, such as directionality and light curve

Detection of the Diffuse Supernova Neutrino Background with JUNO

As an underground multi-purpose neutrino detector with 20 kton liquid scintillator, Jiangmen Underground Neutrino Observatory (JUNO) is competitive with and complementary to the water-Cherenkov detectors on the search for the diffuse supernova neutrino background (DSNB). Typical supernova models predict 2-4 events per year within the optimal observation window in the JUNO detector. The dominant background is from the neutral-current (NC) interaction of atmospheric neutrinos with 12C nuclei, which surpasses the DSNB by more than one order of magnitude. We evaluated the systematic uncertainty of NC background from the spread of a variety of data-driven models and further developed a method to determine NC background within 15\% with {\it{in}} {\it{situ}} measurements after ten years of running. Besides, the NC-like backgrounds can be effectively suppressed by the intrinsic pulse-shape discrimination (PSD) capabilities of liquid scintillators. In this talk, I will present in detail the improvements on NC background uncertainty evaluation, PSD discriminator development, and finally, the potential of DSNB sensitivity in JUNO

Real-time Monitoring for the Next Core-Collapse Supernova in JUNO

Core-collapse supernova (CCSN) is one of the most energetic astrophysical events in the Universe. The early and prompt detection of neutrinos before (pre-SN) and during the SN burst is a unique opportunity to realize the multi-messenger observation of the CCSN events. In this work, we describe the monitoring concept and present the sensitivity of the system to the pre-SN and SN neutrinos at the Jiangmen Underground Neutrino Observatory (JUNO), which is a 20 kton liquid scintillator detector under construction in South China. The real-time monitoring system is designed with both the prompt monitors on the electronic board and online monitors at the data acquisition stage, in order to ensure both the alert speed and alert coverage of progenitor stars. By assuming a false alert rate of 1 per year, this monitoring system can be sensitive to the pre-SN neutrinos up to the distance of about 1.6 (0.9) kpc and SN neutrinos up to about 370 (360) kpc for a progenitor mass of 30$M_{\odot}$ for the case of normal (inverted) mass ordering. The pointing ability of the CCSN is evaluated by using the accumulated event anisotropy of the inverse beta decay interactions from pre-SN or SN neutrinos, which, along with the early alert, can play important roles for the followup multi-messenger observations of the next Galactic or nearby extragalactic CCSN.Comment: 24 pages, 9 figure