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Frontiers in Immunology 2024Interstitial lung disease is a common complication of anti-synthetase syndrome (ASS), and lymphocytic infiltration is often observed in the lesion. We have recently...
Interstitial lung disease is a common complication of anti-synthetase syndrome (ASS), and lymphocytic infiltration is often observed in the lesion. We have recently reported that disease-specific autoantibodies are produced by infiltrating lymphocytes in some autoimmune diseases. Here, we investigate the antigen specificity of B cells in the lung lesions of ASS patients. A total of 177 antibodies were produced from antibody-secreting cells in bronchoalveolar fluid (BALF) of three each of serum anti-Jo-1 and serum anti-EJ antibody-positive patients. Twelve to 30% and 50 to 62% of these antibodies were disease-specific autoantibodies, respectively. These autoantibodies recognized conformational epitopes of the whole self-antigen and had affinity maturations, indicating that self-antigens themselves are the target of humoral immunity. In addition, 100 antibodies were produced from two salivary gland tissues, obtained by chance, of ASS patients. Salivary glands are not generally recognized as lesions of ASS, but unexpectedly, ASS-related autoantibody production was also observed similar to that of BALF. Immunostaining confirmed the presence of ASS-related autoantibody-producing cells in salivary glands. Our results suggest that disease-specific autoantibody production at lesion sites is a common pathogenesis of autoimmune diseases, and that tissue-specific production of autoantibodies can provide insights regarding the distribution of organ manifestations in autoimmune diseases.
Topics: Humans; Salivary Glands; Autoantibodies; Myositis; Female; Male; Lung; Middle Aged; Bronchoalveolar Lavage Fluid; Adult; B-Lymphocytes; Lung Diseases, Interstitial; Autoantigens; Antibodies, Antinuclear; Aged
PubMed: 38938569
DOI: 10.3389/fimmu.2024.1265792 -
Acta Biomaterialia Jun 2024The emergence of antimicrobial-resistant bacterial infections poses a significant threat to public health, necessitating the development of innovative and effective...
The emergence of antimicrobial-resistant bacterial infections poses a significant threat to public health, necessitating the development of innovative and effective alternatives to antibiotics. Photodynamic therapy (PDT) and immunotherapy show promise in combating bacteria. However, PDT's effectiveness is hindered by its low specificity to bacteria, while immunotherapy struggles to eliminate bacteria in immunosuppressive environments. In this work, we introduce an innovative near-infrared antimicrobial nanoplatform (ZFC) driven by bacterial metabolism. ZFC, comprising D-cysteine-functionalized pentafluorophenyl bacteriochlorin (FBC-Cy) coordinated with Zn, is designed for antimicrobial photodynamic-immune therapy (aPIT) against systemic bacterial infections. By specifically targeting bacteria via D-amino acid incorporation into bacterial surface peptidoglycans during metabolism, ZFC achieves precise bacterial clearance in wound and pulmonary infections, exhibiting an antimicrobial efficacy of up to 90% with minimal damage to normal cells under 750 nm light. Additionally, ZFC enhances the activation of antigen-presenting cells by 3.2-fold compared to control groups. Furthermore, aPIT induced by ZFC triggers systemic immune responses and establishes immune memory, resulting in a 1.84-fold increase in antibody expression against bacterial infections throughout the body of mice. In conclusion, aPIT prompted by ZFC presents a approach to treating bacterial infections, offering a broad-spectrum solution for systemic bacterial infections. STATEMENT OF SIGNIFICANCE: The new concept demonstrated focuses on an innovative near-infrared antimicrobial nanoplatform (ZFC) for antimicrobial photodynamic-immune therapy (aPIT), highlighting its reliance on bacterial metabolism and its non-damaging effect on normal tissues. ZFC efficiently targets deep-tissue bacterial infections by harnessing bacterial metabolism, thereby enhancing therapeutic efficacy while sparing normal tissues from harm. This approach not only clears bacterial infections effectively but also induces potent adaptive immune responses, leading to the eradication of distant bacterial infections. By emphasizing ZFC's unique mechanism driven by bacterial metabolism and its tissue-sparing properties, this work underscores the potential for groundbreaking advancements in antimicrobial therapy. Such advancements hold promise for minimizing collateral damage to healthy tissues, thereby improving treatment outcomes and mitigating the threat of antimicrobial resistance. This integrated approach represents a significant progress forward in the development of next-generation antimicrobial therapies with enhanced precision and efficacy.
PubMed: 38936751
DOI: 10.1016/j.actbio.2024.06.024 -
Veterinary Journal (London, England :... Jun 2024African swine fever (ASF) is an acute, febrile, and highly lethal infectious disease in pigs caused by the African swine fever virus (ASFV). Effective detection methods...
African swine fever (ASF) is an acute, febrile, and highly lethal infectious disease in pigs caused by the African swine fever virus (ASFV). Effective detection methods and strict biosecurity measures are crucial for preventing and controlling ASF, especially since there are currently no commercially available vaccines or antiviral drugs to combat ASFV infection effectively. However, the emergence of low-virulence strains of ASFV in recent years has led to false-positive results, highlighting the importance of early-produced antibody detection methods. Therefore, detecting antibodies against ASFV produced early in the infection can facilitate the prompt identification of infected pigs. This study focused on the p30 protein, an early expressed protein during ASFV infection, to develop an indirect ELISA. This method was established using the HEK293F suspension cell expression system, which has the ability to produce large quantities of correctly folded proteins with normal functionality. In this study, we developed an indirect ELISA test utilizing the p30 recombinant protein produced by the HEK293F suspension cell expression system as the antigen coating. The concentration of the p30 protein obtained from the HEK293F suspension cell expression system was measured at 4.668mg/mL, serving as the foundation for establishing the indirect ELISA. Our findings indicate that the indirect ELISA method exhibits a sensitivity of 1:12800. Furthermore, it demonstrates high specificity and excellent reproducibility. Comparing our results to those obtained from the commercial kit, we found a coincidence rate of 98.148% for the indirect ELISA. In summary, we have developed a sensitive method for detecting ASFV, providing a valuable tool for monitoring ASFV infection in pig herds.
PubMed: 38936461
DOI: 10.1016/j.tvjl.2024.106186 -
The American Journal of Gastroenterology Jun 2024Life-long adherence to gluten-free diet (GFD) and its assessment is essential for celiac disease (CeD) patients. We have developed and validated a tool for assessing...
OBJECTIVE
Life-long adherence to gluten-free diet (GFD) and its assessment is essential for celiac disease (CeD) patients. We have developed and validated a tool for assessing adherence to GFD which can be used by both physicians and dietitians.
DESIGN
Phase-I: Development, content validation and assessment of reliability of tool. Phase-II: Validation of tool against standard dietary evaluation (SDE) (gold standard), IgA anti-tissue transglutaminase antibody (IgA-anti-tTG Ab) and gluten immunogenic peptides in urine (Urine-GIP). Overall, 380 biopsy-confirmed CeD patients [Derivation-cohort: n=100(Phase:I), n=210(Phase:II) and Independent validation cohort, n=70)were recruited.
RESULTS
Of initial 90-point questionnaire, 84-items (CD-CAT.v1) were retained after content validation and pilot testing. In Phase:I, upon administering CD-CAT.v1on 100 patients, a comprehensive 35-items tool (CD-CAT.v2;α=0.86) was obtained after removing items with low test-retest reliability and item-rest correlation values. In Phase:II, upon administering CD-CAT.v2 on 210patients,22 items were removed having low correlation values (R<0.4) with SDE. Finally, a 13-item tool (CD-CAT.v3;α=0.84) was obtained with high criterion validity with SDE (r=0.806, p<0.001), moderate convergent validity with CDAT (r=0.602, p=0.007) and moderate to weak correlation with urine GIP (r=0.46, p=0.001) and IgA anti-tTG Ab (r=0.39, p=0.008), respectively. The final 13-item tool also strongly correlated with SDE (r=0.78, p<0.001) in an independent validation cohort of 70 CeD patients. Principal component analysis identified three relevant sub-scales with a cumulative-variance of 62%. The sensitivity and specificity of CD-CAT.v3 were 80% and 91%, respectively, with an area under curve of 0.905 with SDE. The obtained cut-off score of <19 from ROC-curve was further categorized as: 13=excellent,14-18=very good, 19-28=average and >28=poor adherence to GFD.
CONCLUSION
CD-CAT is a new and rapid tool for monitoring dietary adherence to GFD with high sensitivity and specificity which can be administered by both physicians and dietitians.
PubMed: 38934507
DOI: 10.14309/ajg.0000000000002911 -
Pathogens & Immunity 2024Fcγ-receptor (FcγR)-independent enhancement of SARS-CoV-2 infection mediated by N-terminal domain (NTD)-binding monoclonal antibodies (mAbs) has been observed , but...
BACKGROUND
Fcγ-receptor (FcγR)-independent enhancement of SARS-CoV-2 infection mediated by N-terminal domain (NTD)-binding monoclonal antibodies (mAbs) has been observed , but the functional significance of these antibodies is less clear.
METHODS
We characterized 1,213 SARS-CoV-2 spike (S)-binding mAbs derived from COVID-19 convalescent patients for binding specificity to the SARS-CoV-2 S protein, VH germ-line usage, and affinity maturation. Infection enhancement in a vesicular stomatitis virus (VSV)-SARS-CoV-2 S pseudovirus (PV) assay was characterized in respiratory and intestinal epithelial cell lines, and against SARS-CoV-2 variants of concern (VOC). Proteomic deconvolution of the serum antibody repertoire was used to determine functional attributes of secreted NTD-binding mAbs.
RESULTS
We identified 72/1213 (5.9%) mAbs that enhanced SARS-CoV-2 infection in a PV assay. The majority (68%) of these mAbs recognized the NTD, were identified in patients with mild and severe disease, and persisted for at least 5 months post-infection. Infection enhancement by NTD-binding mAbs was not observed in intestinal and respiratory epithelial cell lines and was diminished or lost against SARS-CoV-2 VOC. Proteomic deconvolution of the serum antibody repertoire from 2 of the convalescent patients identified, for the first time, NTD-binding, infection-enhancing mAbs among the circulating immunoglobulins directly isolated from serum. Functional analysis of these mAbs demonstrated robust activation of FcγRIIIa associated with antibody binding to recombinant S proteins.
CONCLUSIONS
Functionally active NTD-specific mAbs arise frequently during natural infection and can last as major serum clonotypes during convalescence. These antibodies display functional attributes that include FcγR activation, and may be selected against by mutations in NTD associated with SARS-CoV-2 VOC.
PubMed: 38933606
DOI: 10.20411/pai.v9i2.679 -
Frontiers in Immunology 2024Poisoning by widow-spider (genus ) bites occurs worldwide. The illness, termed latrodectism, can cause severe and persistent pain and can lead to muscle rigidity,...
Poisoning by widow-spider (genus ) bites occurs worldwide. The illness, termed latrodectism, can cause severe and persistent pain and can lead to muscle rigidity, respiratory complications, and cardiac problems. It is a global health challenge especially in developing countries. Equine serum-derived polyclonal anti-sera are commercially available as a medication for patients with latrodectism, but the use of sera imposes potential inherent risks related to its animal origin. The treatment may cause allergic reactions in humans (serum sickness), including anaphylactic shock. Furthermore, equine-derived antivenom is observed to have batch-to-batch variability and poor specificity, as it is always an undefined mix of antibodies. Because latrodectism can be extremely painful but is rarely fatal, the use of antivenom is controversial and only a small fraction of patients is treated. In this work, recombinant human antibodies were selected against alpha-latrotoxin of the European black widow () by phage display from a naïve antibody gene library. Alpha-Latrotoxin (α-LTX) binding scFv were recloned and produced as fully human IgG. A novel alamarBlue assay for venom neutralization was developed and used to select neutralizing IgGs. The human antibodies showed neutralization efficacy both as single antibodies and antibody combinations. This was also confirmed by electrophysiological measurements of neuronal activity in cell culture. The best neutralizing antibodies showed nanomolar affinities. Antibody MRU44-4-A1 showed outstanding neutralization efficacy and affinity to α-LTX. Interestingly, only two of the neutralizing antibodies showed cross-neutralization of the venom of the Southern black widow (). This was unexpected, because in the current literature the alpha-latrotoxins are described as highly conserved. The here-engineered antibodies are candidates for future development as potential therapeutics and diagnostic tools, as they for the first time would provide unlimited supply of a chemically completely defined drug of constant quality and efficacy, which is also made without the use of animals.
Topics: Humans; Animals; Black Widow Spider; Antibodies, Neutralizing; Spider Venoms; Antivenins; Single-Chain Antibodies; Spider Bites; Immunoglobulin G
PubMed: 38933276
DOI: 10.3389/fimmu.2024.1407398 -
Journal of Personalized Medicine Jun 2024The early reliable detection and quantification of autoantibodies play an important role in autoimmune disease diagnosis and in disease-course monitoring. New...
BACKGROUND
The early reliable detection and quantification of autoantibodies play an important role in autoimmune disease diagnosis and in disease-course monitoring. New technologies, such as the multiplexed determination of autoantibodies, have recently been introduced and are being adopted more frequently. The aim of this study was to evaluate the ability of a new microdot array-based multiparametric assay (ZENIT AMiDot CTD panel, A. Menarini Diagnostics, Firenze, Italy) to correctly classify patients with autoimmune rheumatic diseases (ARDs) and compare it to a fluorescence enzyme immunoassay (FEIA) for the detection of anti-ENAs.
METHODS
The study included 69 consecutive samples from patients with ARDs that were analyzed using two different methods (FEIA and AMiDot) to detect anti-CENP B and six anti-ENA antibodies: anti-Scl-70, anti-SSB/La, anti-Jo-1, anti-U1-RNP, anti-Ro52, and anti-Ro60. The control group sera came from sixty-eight blood donors. Tests were run on the automated slide processor ZENIT FLOW, and then the slides were imaged and analyzed using ZENIT fast.
RESULTS
Since the samples were selected for at least one antibody positivity with an ARD diagnosis, we did not calculate clinical sensitivity but only specificity, which was 98.53%, ranging from 90% for anti-SSB/La antibodies to 100% for anti-CENP B ones. Mean agreement among the methods assessed by Cohen's kappa was 0.816 ± 0.240.
CONCLUSIONS
The assay demonstrated good clinical performance and may be considered a valuable aid in detecting ARD patients, offering an alternative to methods such as FEIA which are largely in use today.
PubMed: 38929828
DOI: 10.3390/jpm14060607 -
Foods (Basel, Switzerland) Jun 2024Soybean agglutinin (SBA) is a primary antinutritional factor in soybeans that can inhibit the growth of humans and mammals, disrupt the intestinal environment, and cause...
Soybean agglutinin (SBA) is a primary antinutritional factor in soybeans that can inhibit the growth of humans and mammals, disrupt the intestinal environment, and cause pathological changes. Therefore, detecting and monitoring SBA in foods is essential for safeguarding human health. In this paper, M13 phage-displayed nanobodies against SBA were isolated from a naive nanobody library. An M13 phage-displayed nanobody-based competitive enzyme-linked immunosorbent assay (P-cELISA) was then established for SBA analysis using biotinylated anti-M13 phage antibody (biotin-anti-M13) and streptavidin poly-HRP conjugate (SA-poly-HRP). The biotin-anti-M13@SA-poly-HRP probe can easily amplify the detection signal without the chemical modifications of phage-displayed nanobodies. The established P-cELISA presented a linear detection range of 0.56-250.23 ng/mL and a limit of detection (LOD) of 0.20 ng/mL, which was 12.6-fold more sensitive than the traditional phage-ELISA. Moreover, the developed method showed good specificity for SBA and acceptable recoveries (78.21-121.11%) in spiked wheat flour, albumen powder, and whole milk powder. This study proposes that P-cELISA based on biotin-anti-M13@SA-poly-HRP may provide a convenient and effective strategy for the sensitive detection of SBA.
PubMed: 38928834
DOI: 10.3390/foods13121893 -
International Journal of Molecular... Jun 2024The objective of the study was to evaluate the profile and diagnostic significance of serum autoantibodies in infertile patients with premature ovarian insufficiency...
The objective of the study was to evaluate the profile and diagnostic significance of serum autoantibodies in infertile patients with premature ovarian insufficiency (POI). The pilot study included 26 patients of reproductive age with POI and diminished ovarian reserve who received complex treatment using new surgical technologies (Group 1) and 18 patients without POI (Group 2). The profile of serum autoantibodies, including anti-ovarian antibodies, antibodies against thyroid peroxidase (TPO), steroidogenic enzymes, and steroid and gonadotropic hormones, was studied using modified ELISAs and human recombinant steroidogenic enzymes (CYP11A1, CYP19A1, CYP21A2). Patients in Group 1 had higher levels of IgG autoantibodies against steroidogenic enzymes, estradiol, progesterone, and TPO than those in Group 2. Tests for IgG antibodies against CYP11A1, CYP19A1, and CYP21A2 exhibited high sensitivity (65.4-76.9%), specificity (83.3-89.9%), and AUC values (0.842-0.910) for POI, the highest in the first test. Three-antibodies panel screening showed higher diagnostic accuracy (84.1% versus 75-79.6%). The levels of these antibodies correlated with menstrual irregularities and a decrease in the antral follicle count. Thus, antibodies against CYP11A1, CYP19A1, and CYP21A2 have a high diagnostic value for POI. Three-antibody panel screening may improve the accuracy of POI diagnosis and be useful for identifying high-risk groups, early stages of the disease, and predicting POI progression.
Topics: Humans; Female; Autoantibodies; Primary Ovarian Insufficiency; Adult; Infertility, Female; Cholesterol Side-Chain Cleavage Enzyme; Aromatase; Steroid 21-Hydroxylase; Iodide Peroxidase; Pilot Projects; Immunoglobulin G; Biomarkers; Progesterone; Estradiol
PubMed: 38928251
DOI: 10.3390/ijms25126545 -
Journal of Thrombosis and Haemostasis :... Jun 2024Scientific and clinical interest in extracellular vesicles (EVs) is growing. EVs that expose tissue factor (TF) bind factor VII/VIIa and can trigger coagulation. Highly...
BACKGROUND
Scientific and clinical interest in extracellular vesicles (EVs) is growing. EVs that expose tissue factor (TF) bind factor VII/VIIa and can trigger coagulation. Highly procoagulant TF-exposing EVs are detectable in the circulation in various diseases, such as sepsis, COVID-19 or cancer. Many in-house and commercially available assays have been developed to measure EV-TF activity and antigen but only a few studies have compared some of these assays. The ISTH SSC Subcommittee on Vascular Biology initiated a multicenter study to compare the sensitivity, specificity and reproducibility of these assays.
MATERIALS AND METHODS
Platelet-depleted plasma samples were prepared from blood of healthy donors. The plasma samples were spiked either with EVs from human milk, or EVs from TF-positive and TF-negative cell lines. Plasma was also prepared from whole human blood with or without LPS stimulation. Twenty-one laboratories measured EV-TF activity and antigen in the prepared samples using their own assays representing 18 functional and 9 antigenic assays.
RESULTS
There was a large variability in the absolute values for the different EV-TF activity and antigen assays. Activity assays had higher specificity and sensitivity compared to antigen assays. In addition, there was a large intra-assay and inter-assay variability. Functional assays that used a blocking anti-TF antibody or immunocapture were the most specific and sensitive. Activity assays that used immunocapture had a lower coefficient of variation compared to assays that isolated EVs by high-speed centrifugation.
CONCLUSION
Based on this multicenter study, we recommend measuring EV-TF using a functional assay in the presence of an anti-TF antibody.
PubMed: 38925490
DOI: 10.1016/j.jtha.2024.05.037