| 1. |
- Al-Rabadi, Laith Farah, et al.
(författare)
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Serine Protease HTRA1 as a Novel Target Antigen in Primary Membranous Nephropathy
- 2021
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Ingår i: Journal of the American Society of Nephrology. - : American Society of Nephrology (ASN). - 1046-6673 .- 1533-3450. ; 32:7, s. 1666-1681
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Tidskriftsartikel (refereegranskat)abstract
- Background Identification of target antigens PLA2R, THSD7A, NELL1, or Semaphorin-3B can explain the majority of cases of primary membranous nephropathy (MN). However, target antigens remain unidentified in 15%-20% of patients. Methods A multipronged approach, using traditional and modern technologies, converged on a novel target antigen, and capitalized on the temporal variation in autoantibody titer for biomarker discovery. Immunoblotting of human glomerular proteins followed by differential immunoprecipitation and mass spectrometric analysis was complemented by laser-capture microdissection followed by mass spectrometry, elution of immune complexes from renal biopsy specimen tissue, and autoimmune profiling on a protein fragment microarray. Results These approaches identified serine protease HTRA1 as a novel podocyte antigen in a subset of patients with primary MN. Sera from two patients reacted by immunoblotting with a 51-kD protein within glomerular extract and with recombinant human HTRA1, under reducing and nonreducing conditions. Longitudinal serum samples from these patients seemed to correlate with clinical disease activity. As in PLA2R- and THSD7A- associated MN, anti-HTRA1 antibodies were predominantly IgG4, suggesting a primary etiology. Analysis of sera collected during active disease versus remission on protein fragment microarrays detected significantly higher titers of anti-HTRA1 antibody in active disease. HTRA1 was specifically detected within immune deposits of HTRA1-associated MN in 14 patients identified among three cohorts. Screening of 118 "quadruple-negative" (PLA2R-, THSD7A-, NELL1-, EXT2-negative) patients in a large repository of MN biopsy specimens revealed a prevalence of 4.2%. Conclusions Conventional and more modern techniques converged to identify serine protease HTRA1 as a target antigen in MN.
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| 2. |
- Dou, Diana R., et al.
(författare)
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Xist ribonucleoproteins promote female sex-biased autoimmunity
- 2024
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Ingår i: Cell. - : Elsevier BV. - 0092-8674 .- 1097-4172. ; 187:3, s. 16-733
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Tidskriftsartikel (refereegranskat)abstract
- Autoimmune diseases disproportionately affect females more than males. The XX sex chromosome complement is strongly associated with susceptibility to autoimmunity. Xist long non-coding RNA (lncRNA) is expressed only in females to randomly inactivate one of the two X chromosomes to achieve gene dosage compensation. Here, we show that the Xist ribonucleoprotein (RNP) complex comprising numerous autoantigenic components is an important driver of sex-biased autoimmunity. Inducible transgenic expression of a non-silencing form of Xist in male mice introduced Xist RNP complexes and sufficed to produce autoantibodies. Male SJL/J mice expressing transgenic Xist developed more severe multi-organ pathology in a pristane-induced lupus model than wild-type males. Xist expression in males reprogrammed T and B cell populations and chromatin states to more resemble wild-type females. Human patients with autoimmune diseases displayed significant autoantibodies to multiple components of XIST RNP. Thus, a sex-specific lncRNA scaffolds ubiquitous RNP components to drive sex-biased immunity.
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| 3. |
- Fuentes, Manuel, et al.
(författare)
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Exploring High-Throughput Immunoassays for Biomarker Validation in Rheumatic Diseases in the Context of the Human Proteome Project
- 2023
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Ingår i: Journal of Proteome Research. - : American Chemical Society. - 1535-3893 .- 1535-3907. ; 22:4, s. 1105-1115
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Tidskriftsartikel (refereegranskat)abstract
- Rheumatic diseases are high prevalence pathologies with different etiology and evolution and low sensitivity in clinical diagnosis. Therefore, it is necessary to develop an early diagnosis method which allows personalized treatment, depending on the specific pathology. The biology/disease initiative, at Human Proteome Project, is an integrative approach to identify relevant proteins in the human proteome associated with pathologies. A previously reported literature data mining analysis, which identified proteins related to osteoarthritis (OA), rheumatoid arthritis (RA), and psoriatic arthritis (PSA) was used to establish a systematic prioritization of potential biomarkers candidates for further evaluation by functional proteomics studies. The aim was to study the protein profile of serum samples from patients with rheumatic diseases such as OA, RA, and PSA. To achieve this goal, customized antibody microarrays (containing 151 antibodies targeting 121 specific proteins) were used to identify biomarkers related to early and specific diagnosis in a screening of 960 serum samples (nondepleted) (OA, n = 480; RA, n = 192; PSA, n = 288). This functional proteomics screening has allowed the determination of a panel (30 serum proteins) as potential biomarkers for these rheumatic diseases, displaying receiver operating characteristics curves with area under the curve values of 80-90%.
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| 4. |
- Hober, Sophia, Professor, 1965-, et al.
(författare)
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Systematic evaluation of SARS-CoV-2 antigens enables a highly specific and sensitive multiplex serological COVID-19 assay
- 2021
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Ingår i: Clinical & Translational Immunology. - : Wiley. - 2050-0068. ; 10:7
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Tidskriftsartikel (refereegranskat)abstract
- Objective. The COVID-19 pandemic poses an immense need for accurate, sensitive and high-throughput clinical tests, and serological assays are needed for both overarching epidemiological studies and evaluating vaccines. Here, we present the development and validation of a high-throughput multiplex bead-based serological assay. Methods. More than 100 representations of SARS-CoV-2 proteins were included for initial evaluation, including antigens produced in bacterial and mammalian hosts as well as synthetic peptides. The five best-performing antigens, three representing the spike glycoprotein and two representing the nucleocapsid protein, were further evaluated for detection of IgG antibodies in samples from 331 COVID-19 patients and convalescents, and in 2090 negative controls sampled before 2020. Results. Three antigens were finally selected, represented by a soluble trimeric form and the S1-domain of the spike glycoprotein as well as by the C-terminal domain of the nucleocapsid. The sensitivity for these three antigens individually was found to be 99.7%, 99.1% and 99.7%, and the specificity was found to be 98.1%, 98.7% and 95.7%. The best assay performance was although achieved when utilising two antigens in combination, enabling a sensitivity of up to 99.7% combined with a specificity of 100%. Requiring any two of the three antigens resulted in a sensitivity of 99.7% and a specificity of 99.4%. Conclusion. These observations demonstrate that a serological test based on a combination of several SARS-CoV-2 antigens enables a highly specific and sensitive multiplex serological COVID-19 assay.
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| 5. |
- Hong, Mun-Gwan, et al.
(författare)
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Profiles of histidine-rich glycoprotein associate with age and risk of all-cause mortality
- 2020
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Ingår i: Life Science Alliance. - : Life Science Alliance, LLC. - 2575-1077. ; 3:10, s. e202000817-
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Tidskriftsartikel (refereegranskat)abstract
- Despite recognizing aging as a common risk factor of many human diseases, little is known about its molecular traits. To identify age-associated proteins circulating in human blood, we screened 156 individuals aged 50–92 using exploratory and multiplexed affinity proteomics assays. Profiling eight additional study sets (N = 3,987), performing antibody validation, and conducting a meta-analysis revealed a consistent age association (P = 6.61 × 10−6) for circulating histidine-rich glycoprotein (HRG). Sequence variants of HRG influenced how the protein was recognized in the immunoassays. Indeed, only the HRG profiles affected by rs9898 were associated with age and predicted the risk of mortality (HR = 1.25 per SD; 95% CI = 1.12–1.39; P = 6.45 × 10−5) during a follow-up period of 8.5 yr after blood sampling (IQR = 7.7–9.3 yr). Our affinity proteomics analysis found associations between the particular molecular traits of circulating HRG with age and all-cause mortality. The distinct profiles of this multipurpose protein could serve as an accessible and informative indicator of the physiological processes related to biological aging.
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| 6. |
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| 7. |
- Lourido, L., et al.
(författare)
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PRESENCE OF FOUR SERUM AUTOANTIBODIES ASSOCIATES WITH THE ACPA STATUS IN EARLY RHEUMATOID ARTHRITIS
- 2021
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Ingår i: Annals of the Rheumatic Diseases. - : BMJ Publishing Group Ltd. - 0003-4967 .- 1468-2060. ; 80, s. 425-426
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The presence of anti-citrullinated protein antibodies (ACPAs) is a hallmark of rheumatoid arthritis (RA) that precede the development of the disease by years and is used for its clinical diagnosis. However, there are RA subjects that test negative for ACPA and thus the early diagnosis on these patients may be delayed. Furthermore, the presence or absence of ACPA in RA supports the hypothesis that on these two subsets of patients underlie different pathogenesis and clinical outcomes.Objectives:In this work, we searched for serum autoantibodies useful to assist the early diagnosis of ACPA-seronegative RA and its management.Methods:We profiled the serum autoantibody repertoire of 80 ACPA-seronegative and 80 ACPA-seropositive RA subjects from the Swedish population-based Epidemiological Investigation of RA (EIRA) cohort. A suspension bead array platform built on protein fragments within Human Protein Atlas and selected from an initial untargeted screening using arrays containing 2660 total antigens was employed to identify IgG and IgA serum autoantibodies. A validation phase on antigen suspension bead arrays was carried out on another set of samples from EIRA containing 386 ACPA-seropositive, 358 ACPA-seronegative and 372 randomly selected control subjects of the same age and sex. A sample-specific threshold based on 20 times the median absolute deviation plus the median of all signals was selected to determine the reactivity of samples. The Wilcoxon rank sum test and Fisher’s test were applied for the comparison of autoantibody levels and reactivity frequencies between the groups.Results:Our data revealed four antigens associated with the ACPA status (Table 1). Testis-specific Y-encoded-like protein 4 (TSPYL4) showed significantly higher IgG reactivity frequency in ACPA-seronegative subjects compared to ACPA-seropositive (8% vs. 3%; P<0.05). Significant differences at IgG autoantibody levels (P<0.05) were also observed between ACPA-seronegative subjects and controls for this specific antigen. Significantly higher IgG autoantibody levels (P<0.05) towards another antigen, dual specificity mitogen-activated protein kinase kinase 6 (MAP2K6), were also observed in ACPA-seronegative subjects compared to ACPA-seropositive and controls. In contrast, we found significantly higher IgG autoantibody levels (P<0.05) in ACPA-seropositive individuals compared to ACPA-seronegative and controls towards two antigens, anosmin-1 (ANOS-1) and muscle related coiled-coil protein (MURC). ANOS-1 shows also significantly higher IgG reactivity frequency in ACPA-seropositive individuals compared to ACPA-seronegative and controls (22%, 9% and 6% respectively; P<0.05). Interestingly, three out of the four antigens discovered to be associated with the ACPA status in early RA are highly expressed in lungs and heart, two of the main extraarticular sites affected in RA. No significant differences were observed at IgA levels for any of the antigens analyzed.Table 1.Scheme of the different phases of the study, the features within each phase and the results. The reactivity to four antigens allows to distinguish ACPA-seronegative (ACPA-), ACPA seropositive (ACPA+) and controls.PhasesUntargeteddiscoveryTargeteddiscoveryTargetedvalidationNumber of samples80 ACPA-80 ACPA-358 ACPA-372 Controls80 ACPA+80 ACPA+386 ACPA+Antigen arrayplatformPlanararraysSuspensionbead array 1Suspensionbead array 2Number of antigens26606227Number of candidatebiomarkers6227 4 (TSPYL4,MAP2K6,ANOS1,MURC)Conclusion:Upon further validation in other early RA sample cohorts, our data suggest the measurement of these four autoantibodies may be useful for the early diagnosis of ACPA-seronegative RA and give insight into the pathogenesis of the different RA subsets.Characters from table content including title and footnotes:Disclosure of Interests:None declared
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| 8. |
- Mikus, Maria, et al.
(författare)
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Allergome-wide peptide microarrays enable epitope deconvolution in allergen-specific immunotherapy
- 2020
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Ingår i: Journal of Allergy and Clinical Immunology. - : Mosby Inc.. - 0091-6749 .- 1097-6825.
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Tidskriftsartikel (refereegranskat)abstract
- Background: The interaction of allergens and allergen-specific IgE initiates the allergic cascade after crosslinking of receptors on effector cells. Antibodies of other isotypes may modulate such a reaction. Receptor crosslinking requires binding of antibodies to multiple epitopes on the allergen. Limited information is available on the complexity of the epitope structure of most allergens. Objectives: We sought to allow description of the complexity of IgE, IgG4, and IgG epitope recognition at a global, allergome-wide level during allergen-specific immunotherapy (AIT). Methods: We generated an allergome-wide microarray comprising 731 allergens in the form of more than 172,000 overlapping 16-mer peptides. Allergen recognition by IgE, IgG4, and IgG was examined in serum samples collected from subjects undergoing AIT against pollen allergy. Results: Extensive induction of linear peptide-specific Phl p 1– and Bet v 1–specific humoral immunity was demonstrated in subjects undergoing a 3-year-long AIT against grass and birch pollen allergy, respectively. Epitope profiles differed between subjects but were largely established already after 1 year of AIT, suggesting that dominant allergen-specific antibody clones remained as important contributors to humoral immunity following their initial establishment during the early phase of AIT. Complex, subject-specific patterns of allergen isoform and group cross-reactivities in the repertoires were observed, patterns that may indicate different levels of protection against different allergen sources. Conclusions: The study highlights the complexity and subject-specific nature of allergen epitopes recognized following AIT. We envisage that epitope deconvolution will be an important aspect of future efforts to describe and analyze the outcomes of AIT in a personalized manner.
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| 9. |
- Olofsson, Jennie, et al.
(författare)
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Array-Based Multiplex and High-Throughput Serology Assays
- 2023
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Ingår i: Methods in Molecular Biology. - New York, NY : Springer Nature. - 1064-3745 .- 1940-6029. ; 2628, s. 535-553
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Tidskriftsartikel (refereegranskat)abstract
- The detection of antibody responses using serological tests provides means to diagnose infections, follow disease transmission, and monitor vaccination responses. The coronavirus disease 2019 (COVID-19) pandemic, caused by the SARS-CoV-2 virus, highlighted the need for rapid development of robust and reliable serological tests to follow disease spreading. Moreover, the rise of SARS-CoV-2 variants emphasized the need to monitor their transmission and prevalence in the population. For this reason, multiplex and flexible serological assays are needed to allow for rapid inclusion of antigens representing new variants as soon as they appear. In this chapter, we describe the generation and application of a multiplex serological test, based on bead array technology, to detect anti-SARS-CoV-2 antibodies in a high-throughput manner, using only a few microliters of sample. This method is currently expanding to include a multi-disease antigen panel that will allow parallel detection of antibodies towards several infectious agents.
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| 10. |
- Rudberg, Ann-Sofie, et al.
(författare)
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SARS-CoV-2 exposure, symptoms and seroprevalence in healthcare workers in Sweden.
- 2020
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- SARS-CoV-2 may pose an occupational health risk to healthcare workers. Here, we report the seroprevalence of SARS-CoV-2 antibodies, self-reported symptoms and occupational exposure to SARS-CoV-2 among healthcare workers at a large acute care hospital in Sweden. The seroprevalence of IgG antibodies against SARS-CoV-2 was 19.1% among the 2149 healthcare workers recruited between April 14th and May 8th 2020, which was higher than the reported regional seroprevalence during the same time period. Symptoms associated with seroprevalence were anosmia (odds ratio (OR) 28.4, 95% CI 20.6-39.5) and ageusia (OR 19.2, 95% CI 14.3-26.1). Seroprevalence was also associated with patient contact (OR 2.9, 95% CI 1.9-4.5) and covid-19 patient contact (OR 3.3, 95% CI 2.2-5.3). These findings imply an occupational risk for SARS-CoV-2 infection among healthcare workers. Continued measures are warranted to assure healthcare workers safety and reduce transmission from healthcare workers to patients and to the community.
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| 11. |
- Thörnqvist, Linnea, et al.
(författare)
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Linear Epitope Binding Patterns of Grass Pollen-Specific Antibodies in Allergy and in Response to Allergen-Specific Immunotherapy
- 2022
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Ingår i: Frontiers in Allergy. - : Frontiers Media SA. - 2673-6101. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Allergic diseases affect many individuals world-wide and are dependent on the interaction between allergens and antibodies of the IgE isotype. Allergen-specific immunotherapy (AIT) can alter the development of the disease, e.g., through induction of allergen-specific IgG that block allergen-IgE interactions. The knowledge of epitopes recognized by allergy-causing and protective antibodies are limited. Therefore, we developed an allergome-wide peptide microarray, aiming to track linear epitope binding patterns in allergic diseases and during AIT. Here, we focused on immune responses to grass pollen allergens and found that such epitopes were commonly recognized before initiation of AIT and that AIT commonly resulted in increased antibody production against additional epitopes already after 1 year of treatment. The linear epitope binding patterns were highly individual, both for subjects subjected to and for individuals not subjected to AIT. Still, antibodies against some linear epitopes were commonly developed during AIT. For example, the two rigid domains found in grass pollen group 5 allergens have previously been associated to a diversity of discontinuous epitopes. Here, we present evidence that also the flexible linker, connecting these domains, contains regions of linear epitopes against which antibodies are developed during AIT. We also describe some commonly recognized linear epitopes on Phl p 2 and suggest how antibodies against these epitopes may contribute to or prevent allergy in relation to a well-defined stereotyped/public IgE response against the same allergen. Finally, we identify epitopes that induce cross-reactive antibodies, but also antibodies that exclusively bind one of two highly similar variants of a linear epitope. Our findings highlight the complexity of antibody recognition of linear epitopes, with respect to both the studied individuals and the examined allergens. We expect that many of the findings in this study can be generalized also to discontinuous epitopes and that allergen peptide microarrays provide an important tool for enhancing the understanding of allergen-specific antibodies in allergic disease and during AIT.
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