Clinical and experimental immunology最新文献

Our group previously demonstrated that NETs were involved in interstitial lung diseases (ILD) among patients with idiopathic inflammatory myopathies (IIM) and the experimental autoimmune myositis (EAM) mouse model, and that NETs activated lung fibroblasts through the TLR9-miR7-Smad2 axis. This study aimed to establish a novel mouse model of myositis-associated interstitial lung disease (MAILD) by using a TLR9 agonist (ODN2395). ODN2395 and muscle homogenate were used to induce MAILD in BALB/c mice. MAILD was evaluated using histopathology, immunohistochemistry, serum NETs determination, and myositis-specific antibody profile. Furthermore, TLR9 and IRF3 were examined in a lung biopsy tissue from a dermatomyositis patient with ILD. MAILD mice developed inflammatory myopathy with positive expression of myositis specific antibodies. ILD occurred in all mice of MAILD group. ODN2395 at doses of 5μg, 10μg or 20μg induced ILD, with increasing severity as the dose increased, but 20μg ODN2395 was not recommended due to non-specific damage to lungs. ILD could occur as early as one week after immunization and was most pronounced by the fourth/fifth week. MAILD process was accompanied by NETs infiltration and TLR9 activation. TLR9 activation was demonstrated in the patient with DM-ILD. Serum levels of Cit-H3 were elevated in the MAILD group. Skeletal muscle homogenate and ODN2395 induced neutrophils to form NETs in vitro. Combined with muscle homogenate, ODN2395 induced a novel MAILD mouse model with NETs infiltration and TLR9 activation, which are similar to pathogenesis of IIM-ILD, suggesting that MAILD model could replace EAM model in IIM-ILD research.

Vaccines are crucial for protecting health globally; however, their widespread use relies on rigorous clinical development programmes. This includes Phase 3 randomised controlled trials (RCTs) to confirm their safety, immunogenicity, and efficacy. Traditionally, such trials used fixed designs with predetermined assumptions, lacking the flexibility to change during the trial or stop early due to overwhelming evidence of either efficacy or futility. Modern vaccine trials benefit from innovative approaches like adaptive designs, allowing for planned trial adaptations based on accumulating data. Here, we provide an overview of the evolution of Phase 3 vaccine trial design and statistical analysis methods from traditional to more innovative contemporary methods. This includes adaptive trial designs, which offer ethical advantages and enable early termination if indicated; Bayesian methods, which combine prior knowledge and observed trial data to increase efficiency and enhance result interpretation; modern statistical analysis methods, which enable more accurate and precise inferences; the estimand framework, which ensures the primary question of interest is addressed in a trial; novel approaches using machine learning methods to assess heterogeneity of treatment effects; and statistical advances in safety analysis to evaluate reactogenicity and clinical adverse events. We conclude with insights into the future direction of vaccine trials, aiming to inform clinicians and researchers about conventional and novel RCT design and analysis approaches to facilitate the conduct of efficient, timely trials.

HYCOs are hybrid molecules consisting of activators of the transcription factor Nrf2 conjugated to carbon monoxide (CO)-releasing moieties. These "dual action" compounds have been designed to mimic the activity of heme oxygenase-1 (HO-1), a stress inducible cytoprotective enzyme that degrades heme to CO which expression is regulated by Nrf2. HYCOs have recently shown efficacy in ameliorating experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. However, the mechanism(s) of action of HYCOs still remain to be fully investigated. Here, we assessed the effects of HYCO-3, a prototype of these hybrids, on myeloid-derived cells, microglial cells and T lymphocytes obtained from EAE-immunized mice. HYCO-3 exerted immunomodulatory effects on all the examined cell populations by inhibiting the generation of pro-inflammatory cytokines and nitric oxide, and downregulating antigen-presenting capacity of these cells. The observed effects support the view that HYCOs are promising candidates to be developed for the treatment of autoimmune and chronic inflammatory disorders.

Adult-onset immunodeficiency with antibodies to interferon-γ (AOID with AIGA) is a rare, acquired immunodeficiency causing susceptibility to disseminated non-tuberculous mycobacteria and other intracellular opportunistic infections. The diagnosis depends on demonstrating the presence of endogenous anti-interferon-γ antibodies (AIGA) that suppress Th1 cell-mediated immunity. Bioluminescent immunoassays are a newly emerging immunoassay format which utilize the action of bioluminescent enzymes on a substrate for specific analyte detection. In short, detecting antibodies are conjugated with a bioluminescent enzyme. The detecting antibodies bind the analyte of interest and produce light (luminescence) after addition of a substrate. The purpose of this study was to evaluate two newly developed bioluminescent immunoassays using Lumit® (Promega) technology as a diagnostic test for AOID with AIGA. Specific aims included the clinical validation of a new inhibition bioluminescent immunoassay technique to detect AIGA which block detection of interferon-γ (IFN-γ) in vitro and correlation of inhibition bioluminescent immunoassay results with AOID with AIGA disease status. Two bioluminescent inhibition immunoassays were developed. One which adapted an existing kit from Promega (Lumit® Human IFN-γ Immunoassay) and the other which was developed in house. Eighty-seven healthy controls and 48 patients with previously diagnosed AOID with AIGA were recruited and tested using these two methods. Results showed both bioluminescent inhibition immunoassays were able to clearly discriminate between AOID with AIGA patients and healthy controls. The mean inhibition percentage between patient groups correlated with disease activity. Both assays appeared to be more sensitive when compared to the existing inhibition ELISA.

Acquired aplastic anemia (AA) often results from immune destruction of hematopoietic stem and progenitor cells. However, only 60%-70% of patients with AA respond to immunosuppressive therapy (IST). There is a lack of strong predictive markers for response to IST which can help therapy. Our study sought to pinpoint unique immune markers in AA patients and validate established predictors for response to IST. We enrolled 51 severe AA patients and analyzed 57 immunological parameters via flow cytometry. Additionally, we measured paroxysmal nocturnal hemoglobinuria (PNH) clone, telomere length, and thrombopoietin (TPO) levels prior to IST. After a 6-month follow-up, a response was observed. Patients with AA had a distinct immunological signature characterized by absolute lymphopenia, skewed CD4/CD8 ratio with expansion of CD8 T cells with activated and senescent phenotype. Treg counts were reduced, while the proportion of Treg A and B was comparable to controls. Treatment response was correlated with elevated absolute neutrophil count (ANC), absolute reticulocyte count (ARC), and reduced CD57+ CD8+ naive cells and B cell % before therapy. However, predictors like TPO, telomere length, and PNH did not emerge as indicators of treatment response. Identifying predictors for treatment response in AA is challenging due to abnormal hematopoiesis, genetic mutations, and treatment variables.

Cellular phenotype and function are altered in different microenvironments. For targeted therapies it is important to understand site-specific cellular adaptations. Juvenile idiopathic arthritis (JIA) is characterized by autoimmune joint inflammation, with frequent inadequate treatment responses. To comprehensively assess the inflammatory immune landscape, we designed a 37-parameter spectral flow cytometry panel delineating mononuclear cells from JIA synovial fluid (SF) of autoimmune inflamed joints, compared to JIA and healthy control blood. Synovial monocytes and NK cells (CD56bright) lack Fc-receptor CD16, suggesting antibody-mediated targeting may be ineffective. B cells and DCs, both in small frequencies in SF, undergo maturation with high 4-1BB, CD71, CD39 expression, supporting T-cell activation. SF effector and regulatory T cells were highly active with newly described co-receptor combinations that may alter function, and suggestion of metabolic reprogramming via CD71, TNFR2, and PD-1. Most SF effector phenotypes, as well as an identified CD4-Foxp3+ T-cell population, were restricted to the inflamed joint, yet specific SF-predominant CD4+ Foxp3+ Treg subpopulations were increased in blood of active but not inactive JIA, suggesting possible recirculation and loss of immunoregulation at distal sites. This first comprehensive dataset of the site-specific inflammatory landscape at protein level will inform functional studies and the development of targeted therapeutics to restore immunoregulatory balance and achieve remission in JIA.

Psoriasis is a chronic, inflammatory skin disease characterized by a dysregulated immune response and systemic inflammation. Up to one-third of patients with psoriasis have psoriatic arthritis (PsA). Targeted treatment with antibodies neutralizing tumor necrosis factor can ameliorate both diseases. We here explored the impact of long-term infliximab treatment on the composition and activity status of circulating immune cells involved in chronic skin and joint inflammation. Immune cells were analyzed by multicolor flow cytometry. We measured markers of immune activation in peripheral blood mononuclear cell populations in 24 infliximab-treated patients with psoriasis/PsA compared to 32 healthy controls. We observed a significant decrease in the frequency of both peripheral natural killer (NK) cells and their subset CD56dimCD16+ NK cells in PsA compared to healthy controls and patients with psoriasis. The latter had a strong-positive correlation with psoriasis area severity index (PASI) in these patients, while CD56brightCD16- NK cells were negatively correlated with PASI. In addition, we observed an upregulation of CD69+ intermediate CD14+CD16+ and CD69+ classical CD14+CD16- monocytes in PsA and increased activity of CD38+ intermediate CD14+CD16+ monocytes in patients with psoriasis. Compared to healthy controls, psoriasis patients demonstrated shifts of the three B-cell subsets with a decrease in transitional CD27-CD38high B cells. Our exploratory study indicates a preserved pathophysiological process including continuous systemic inflammation despite clinical stability of the patients treated with infliximab.

The recent pandemic was caused by the emergence of a new human pathogen, SARS-CoV-2. While the rapid development of many vaccines provided an end to the immediate crisis, there remains an urgent need to understand more about this new virus and what constitutes a beneficial immune response in terms of successful resolution of infection. Indeed, this is key for development of vaccines that provide long lasting protective immunity. The interferon lambda (IFNL) family of cytokines are produced early in response to infection and are generally considered anti-viral and beneficial. However, data regarding production of IFNL cytokines in coronavirus disease 2019 (COVID-19) patients is highly variable, and generally from underpowered studies. In this study, we measured all three IFNL1, IFNL2, and IFNL3 cytokines in plasma from a well characterized, large COVID-19 cohort (n = 399) that included good representation from patients with a more indolent disease progression, and hence a beneficial immune response. While all three cytokines were produced, they differed in both the frequency of expression in patients, and the levels produced. IFNL3 was produced in almost all patients but neither protein level nor IFNL3/IFNL4 single nucleotide polymorphisms were associated with clinical outcome. In contrast, both IFNL1 and IFNL2 levels were significantly lower, or absent, in plasma of patients that had a more severe disease outcome. These data are consistent with the concept that early IFNL1 and IFNL2 cytokine production is protective against SARS-CoV-2 infection.

Clinical manifestations, as distinct from thrombotic and obstetric morbidity, were recently included in the update of classification criteria of the antiphospholipid syndrome (APS). However, the existence of several patients with clinical manifestations suggestive of APS, but negative for criteria antiphospholipid antibodies (aPLs) [anti-cardiolipin antibodies (aCL), anti-β2-glycoprotein I antibodies (aβ2-GPI), and lupus anticoagulant] may suggest an update of diagnostic criteria. In this study, we analysed the prevalence of six non-criteria aPLs in a large monocentric cohort of patients with seronegative APS (SN-APS), to investigate their possible diagnostic role. aCL IgA, aβ2-GPI IgA, and aβ2-GPI Domain 1 antibodies were detected by chemiluminescence, anti-phosphatidylserine/prothrombin (aPS/PT) IgG, anti-vimentin/cardiolipin (aVim/CL) IgG, and anti-carbamylated-β2-glycoprotein I (aCarb-β2-GPI) IgG by ELISA in sera from 144 SN-APS patients. In SN-APS patients, aCL IgA was detected in 4/144 (2.77%), aβ2-GPI IgA in 2/144 (1.39%), aβ2-GPI-Domain 1 in 1/144 (0.69%), aPS/PT in 16/144 (11.11%), aVim/CL in 37/144 (25.69%), and aCarb-β2-GPI in 43/144 patients (29.86%). Patients negative for all non-criteria aPL assays were 77/144 (53.47%). Notably, the Venn diagram showed that aCarb-β2-GPI together with aVim/CL represented the prevalent combination of positive antibodies. In SN-APS patients, aCL IgA were associated with recurrent thrombosis (OR 11.48; P = 0.03); in obstetric SN-APS patients, aPS/PT were significantly associated with foetal deaths (OR 4.84; P = 0.01), aVim/CL with spontaneous abortions (OR 2.71; P = 0.016). This study indicates that aPS/PT, aVim/CL and aCarb-β2-GPI antibodies may represent useful tools to identify 'seronegative' APS patients, who are negative for criteria aPLs, supporting the need to make testing for non-criteria aPLs more accessible in patients with SN-APS.

Neutrophil extracellular traps (NETs) released by neutrophils are web-like DNA structures adhered to granulin proteins with bactericidal activity and can be an important mechanism for preventing pathogen dissemination or eliminating microorganisms. However, they also play important roles in diseases of other systems, such as the central nervous system. We tracked the latest advances and performed a review based on published original and review articles related to NETs and neurological diseases. Generally, neutrophils barely penetrate the blood-brain barrier into the brain parenchyma, but when pathological changes such as infection, trauma, or neurodegeneration occur, neutrophils rapidly infiltrate the central nervous system to exert their defensive effects. However, neutrophils may adversely affect the host when they uncontrollably release NETs upon persistent neuroinflammation. This review focused on recent advances in understanding the mechanisms and effects of NETs release in neurological diseases, and we also discuss the role of molecules that regulate NETs release in anticipation of clinical applications in neurological diseases.