Scandinavian Journal of Immunology最新文献

Current characterisation of cat dander/hair extracts used for allergy diagnosis or allergen-specific immunotherapy is mainly standardised towards the major allergen Fel d 1, while other allergens such as the lipocalin Fel d 7 are insufficiently characterised in such allergen products. This study aimed to produce recombinant Fel d 7 (rFel d 7) and murine IgG monoclonal antibodies (mAbs) specific to it for quantification in allergen extracts and assess the potential of the mAbs in inhibiting patients' IgE in functional assays. rFel d 7 was expressed in E. coli, purified by Ni-affinity and ion exchange chromatography, and physicochemically characterised by circular dichroism, Fourier-transform infrared spectroscopy, dynamic light scattering and mass spectrometry. Twenty hybridoma cell lines producing Fel d 7-specific mAbs were generated and sandwich ELISA was established for the quantitation of Fel d 7. Six different cat allergen extracts from different manufacturers and prepared from different sources were analysed and the concentration ranged from 0.02 μg/mg to 22.59 μg/mg. A mAb pool recognising non-overlapping epitopes inhibited the binding of human IgE-antigen complex formation (63.7% highest inhibition) and IgE-Fel d 7 cross-linking and consequent degranulation of basophilic cells (57.2% highest inhibition). We demonstrate the vast difference of Fel d 7 content in different cat allergen extracts, highlighting the necessity of improved standardisation of cat allergen extracts. The inhibition results showed that the analysed mAbs effectively inhibit rFel d 7 binding to human IgE, an assay we recommend for assessing the potency of allergen extracts as part of extract standardisation.

Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease characterised by heterogeneous clinical manifestations and varying degrees of organ involvement. The CD40-CD40 Ligand (CD40L) pathway is implicated in autoimmune responses, with elevated levels of soluble CD40L (sCD40L) observed in SLE patients. This study investigates sCD40L as a biomarker for disease activity and its utility in stratifying patients for CD40L-targeted therapies. Plasma levels of sCD40L were quantified in SLE patients (n = 169) and healthy controls (n = 100). Correlations between sCD40L levels and disease activity measures were analysed using Spearman's correlation and logistic regression. K-means clustering grouped patients based on sCD40L concentrations, and principal component analysis (PCA) was performed to assess relationships among disease activity variables. SLE patients exhibited significantly higher sCD40L levels (median 2.2 ng/mL) than healthy controls (median 0.81 ng/mL; p = 0.0079). Elevated sCD40L levels correlated weakly with higher SLE Disease Activity Index (SLEDAI) scores (p = 0.0418), positive Anti-Nuclear Antibody status (p = 0.0133), increased IgG levels (p = 0.0148) and decreased lymphocyte counts (p = 0.0327). Clustering analysis and PCA revealed that patients with higher sCD40L levels tended to have increased disease activity, elevated anti-dsDNA antibody concentrations, and higher C-reactive protein (CRP) levels. Elevated sCD40L levels in SLE patients correlate with disease activity markers, suggesting its potential as a biomarker for monitoring disease progression and severity. Additionally, sCD40L may aid in stratifying patients who could benefit from CD40L-targeted therapies within a treat-to-target framework. Further longitudinal studies with larger cohorts are warranted to validate and explore the role of sCD40L in personalised SLE treatment strategies.

Acute cellular rejection (ACR) remains a common complication post-liver transplantation despite standardised immunosuppressive regimens. Identifying patients at high risk of ACR is challenging, and new tools for prediction are needed. This study aimed to investigate if proportions of circulating T cells as well as different stages of maturation, exhaustion, and activation of T cells pre-transplantation are associated with ACR in liver transplant recipients within the first year post-transplantation. In the prospective ImmuneMO:SOT cohort study, we recruited participants listed for liver transplantation at Copenhagen University Hospital-Rigshospitalet who subsequently underwent liver transplantation. Before transplantation, we collected blood for extensive immune cell profiling using the standardised DURAClone flow cytometry panel and investigated if proportions of CD4⁺ and CD8⁺ T cells, including Th17, Treg, and memory T cell subsets, as well as their activation and exhaustion states were associated with ACR within 1 year post-transplantation. We included 40 liver transplant recipients, of whom 60% were male and had a median age of 50 years. ACR occurred in 37.5% of liver transplant recipients. We found no associations between investigated proportions of T cell subsets, including CD4⁺ (p = 0.25), CD8⁺ (p = 0.43), Th17 (p = 0.61), or Treg cells (p = 0.54) and ACR. This was consistent across activated and exhausted T cells as well as memory T cell subsets. Our findings suggest that proportions of pre-transplant circulating T cells have limited predictive value for post-transplantation ACR. Investigating if other circulating immune cell populations pre-transplantation are associated with ACR after liver transplantation is warranted.

Traditional organ transplantation relies on the Self-Non-self (SNS) model of immunity, focusing on donor-recipient compatibility and aggressive immunosuppression to prevent acute rejection. Although effective early, this strategy does not prevent chronic rejection and cannot account for operational tolerance, failure of perfectly HLA-matched grafts, or the occasional spontaneous acceptance of a fully mismatched organ. The adaptation model of immunity offers a different lens. In the thymus, "central adaptation" programs T cells to recognise self-peptide-MHC (pMHC) so they can later recognise different tissues to facilitate tissue repair and homeostasis. Whether a graft thrives or fails depends on how quickly this self-oriented circuitry can operate. Autografts and isografts arrive with their own extracellular-matrix (ECM) "memory", and recipient T cells immediately recognise their pMHC, triggering tissue-remodelling responses. Allografts must adapt to new ECM-a transition that is associated higher levels of graft injury allowing indirect antigen presentation. Until adaptation is complete, recipient T cells mount cytotoxic rather than reparative responses because of antigen cross-presentation, during which the graft relies on donor-derived tissue-resident memory T cells (T_RM) to maintain integrity. Therapeutically, interventions that preserve or expand graft-borne T_RM, or that pharmacologically enhance adaptation-receptor signalling, could hasten this donor-to-host reprogramming. By replacing blanket immunosuppression with targeted promotion of tissue-remodelling immunity, the adaptation model charts a path toward long-term graft survival without the lifelong risks of today's regimens.

B cells are essential in the immune system, driving antibody production, cytokine secretion and antigen presentation. Studies in mouse models have illuminated key mechanisms underlying B-cell activation, differentiation, class-switch recombination and somatic hypermutation. However, the extent to which these findings translate to human biology remains unclear. To address this, we developed a human primary B-cell culture system using feeder cells engineered to express CD40L, supplemented with the cytokines BAFF, IL-4 and IL-21. Using a Design of Experiments (DOE) approach, we optimised critical parameters and dissected the individual contributions of each specific factor. Our results reveal that BAFF plays a negligible role, and IL-21 has more subtle effects, whereas CD40L and IL-4 are critical determinants of cell viability, proliferation and IgE class-switching. Furthermore, we find that engineered feeder cells can serve equally well as a source of cytokines, but providing these in purified form increases the flexibility of the system. This platform enables detailed investigation of human B-cell biology, offering insights into intrinsic and extrinsic regulators of antibody responses and providing a foundation for in vitro production of human primary antibodies.

Mast cells have long been recognised for their pivotal role in the development of allergic inflammation. However, their contributions extend far beyond this well-established function. Accumulating evidence supports that mast cells play essential roles in regulating both innate and adaptive immune responses by releasing a diverse array of potent immune mediators and cytokines. Importantly, mast cells are nowadays appreciated to possess the capacity to mount effective host defence responses against a variety of pathogens. One of the most notable features of mast cells is their abundance in mucosal tissues, which serve as primary entry points for many infectious agents, as well as common allergens. This strategic positioning enables mast cells to respond quickly to both harmful pathogens and allergens, making them key players in the body's first line of defence. In this review, we aim to summarise the current understanding of how mast cells contribute to the immune defence against infections. We will explore the mechanisms by which these cells respond to various pathogens and how their activation affects the overall immune response. Furthermore, we will discuss innovative strategies for harnessing mast cell activation to enhance vaccine efficacy. By exploiting the adjuvant properties of mast cell activators, we can potentially improve the quality of vaccination against infectious diseases. This exploration highlights the dual role of mast cells not only in allergic responses but also as vital components in the fight against infections, underscoring their importance in both immunology and therapeutic strategies.

A key mechanism of tumour immune escape from CD8⁺ cytotoxic T lymphocytes occurs via downregulation of NLRC5, an IFNγ-induced transcriptional activator of MHC class-I. As NLRC5 deficiency does not abrogate CD8⁺ T cell development, we investigated whether NLRC5-dependent antitumour immune mechanisms are required for immune surveillance. We studied the development of 3-methylcholanthrene (MCA)-induced endogenous fibrosarcoma in Nlrc5^-/- mice with Nlrc5^+/+ and Rag1^-/- mice serving as controls. Nlrc5^-/- and Rag1^-/- mice showed increased propensity to develop MCA-induced tumours with a higher growth rate compared to Nlrc5^+/+ mice and displayed significantly reduced survival. Tumours from Nlrc5^+/+ and Nlrc5^-/- mice, but not from Rag1^-/- mice, contained necrotic areas and displayed T cell infiltration. Tumour cell lines established from MCA-induced tumours were evaluated for their sensitivity to immune-mediated growth control following implantation into immunocompetent C57BL/6 and immunodeficient Rag1^-/- hosts. Tumours formed by Nlrc5^+/+ tumour cell lines progressed unhindered in C57BL/6 hosts that reflected their immunoedited status, whereas cell lines from Nlrc5^-/- and Rag1^-/- tumours were efficiently controlled, indicating their non-immunoedited status. Proteomic analysis by mass spectrometry followed by pathway analysis revealed enrichment of granzyme-mediated cytolytic pathway in Nlrc5^+/+ tumours that were absent in Nlrc5^-/- tumours, which showed enrichment of humoral and innate immune pathways. Overall, our findings show that NLRC5 is required for robust tumour immune surveillance and tumour immunoediting and that compensatory humoral and innate immune mechanisms activated by the loss of NLRC5 are insufficient for cancer immune surveillance and cancer immunoediting.

Controlling excessive inflammation remains an unmet clinical need, for example, during sepsis or after severe injuries. Platelet-activating factor (PAF) is central in thromboinflammatory processes. However, its role in the interaction of platelets and neutrophils requires further insights. Therefore, we elucidated PAF-related neutrophil activation, including platelet-neutrophil complex (PNC) formation and investigated potential strategies to modulate PAF-related inflammation. For the translation of the PAF-mediated inflammation, we applied an animal-free human ex vivo whole blood model. The neutrophil phenotype, its function, and PNC formation were studied by flow cytometry and platelet-related activity was assessed by light microscopy and aggregometry. PAF induced a rapid and dose-dependent change in neutrophil phenotype, as evidenced by CD10, CD11b, and CD66b upregulation and CD62L downregulation. Moreover, PAF increased the generation of reactive oxygen species (ROS), phagocytic activity and PNC formation. Interestingly, PNCs displayed significantly enhanced ROS formation and phagocytosis compared to neutrophils without attached platelets, whereas these differences were not observed regarding phenotype changes. Furthermore, the findings were confirmed in a clinically relevant ex vivo whole blood model of lipopolysaccharide- or PAF-driven inflammation. In summary, the present study elucidates PAF-driven effects on neutrophils and their interaction with platelets. The findings might help in developing therapeutic approaches to modulate PAF-related thromboinflammation, for example, during sepsis.

Burkitt lymphoma (BL) is an aggressive non-Hodgkin B-cell lymphoma characterised by chromosomal translocations involving the MYC gene, leading to its overexpression and driving uncontrolled proliferation. BL is categorised into endemic, sporadic, and immunodeficiency-associated subtypes, each with distinct clinical and epidemiological features. TSPAN32, a member of the tetraspanin family, plays a key role in B cell development and immune regulation. In this study, we investigated the regulation of TSPAN32 expression in BL subtypes. Our results show that TSPAN32 expression is significantly downregulated in endemic, sporadic, and HIV-associated BL. Notably, this downregulation is independent of Epstein-Barr virus (EBV) infection, as no significant differences in TSPAN32 expression were observed between EBV-positive and EBV-negative BL clones. Functional studies revealed that overexpression of a wild-type ID3 gene, a known repressor of TCF3, and knockdown of TCF3, both led to a significant upregulation of TSPAN32, particularly in BL41 and Daudi cells, which harbour ID3 mutations. Supporting this, ChIP-seq analysis identified TCF3 binding peaks on the TSPAN32 gene, providing mechanistic evidence of its regulation by TCF3. These findings shed light on the complex transcriptional network regulating TSPAN32 and its dysregulation in BL. Overall, our study suggests that TSPAN32 may serve as both a biomarker and a potential therapeutic target for this disease.

The lack of diagnostic and monitoring tools for postsurgical immunological complications such as systemic inflammation can contribute to a poor outcome despite modern intensive care. The need for reliable immune monitoring has been emphasised. Polymorphonuclear leukocytes (PMNs) play an important role in postsurgical inflammation. A subgroup of PMNs is particularly interesting, because they are released shortly after (iatrogenic) trauma: immature PMNs, characterised by, for example, their low CD10 expression. Therefore, we investigated the role of CD10^low PMNs in a non-interventional exploratory study by including patients undergoing scheduled, highly standardised cardiac surgery with extracorporeal circulation. We were able to demonstrate that the number of CD10^low PMNs released shortly after the beginning of surgery correlated with different fluid phase markers of inflammation and organ damage postsurgically. Among these parameters were CRP, IL-6, NGAL, CK-MB, and troponin-T. Noteworthy, the amount of CD10^low PMNs increased as early as 24 h before these well-established markers, suggesting superiority of CD10^low PMNs as an early diagnostic marker. Comparing CD10^low immature PMNs with CD10^high mature PMNs revealed potential involved mechanisms, including lower CD11b expression and a significant decrease in the formation of platelet-neutrophil complexes (PNCs) by CD10^low PMNs. In conclusion, we propose CD10^low PMNs as a potential early cellular biomarker to assess the postsurgical inflammatory response. In comparison to clinically established markers like CRP or IL-6 and scoring systems such as the SOFA-Score, CD10^low PMNs reflect a potential candidate for future immune monitoring to determine the risk of excessive inflammation and organ impairment more rapidly.