Early IL-1 signaling promotes iBALT induction after influenza virus infection

Inducible bronchus-associated lymphoid tissue (iBALT) is a long lasting tertiary lymphoid tissue that can be induced following influenza A virus (IAV) infection. Previous studies have shown that iBALT structures containing germinal center (GC) B cells protect against repeated infection by contributing locally to the cellular and humoral immune response. If we are to exploit this in vaccination strategies, we need a better understanding on how iBALT structures are induced. One hypothesis is that the strength of the initial innate response dictates induction of iBALT. In the present study, we investigated the role of interleukin (IL)-1 and IL-1R signaling on iBALT formation. Mice lacking the IL-1R had a delayed viral clearance and, thus, a prolonged exposure to viral replication, leading to increased disease severity, compared to wild-type mice. Contradictorily, iBALT formation following clearance of the virus was heavily compromised in Il1r1-/- mice. Quantification of gene induction after IAV infection demonstrated induction of IL-1α and to a much lesser extent of IL-1β. Administration of recombinant IL-1α to the lungs of wild-type mice, early but not late, after IAV infection led to more pronounced iBALT formation and an increased amount of GC B cells in the lungs. Bone marrow chimeric mice identified the stromal compartment as the crucial IL-1 responsive cell for iBALT induction. Mechanistically, Q-PCR analysis of lung homogenates revealed a strongly diminished production of CXCL13, a B cell-attracting chemokine, in Il1r-/- mice during the early innate phase of IAV infection. These experiments demonstrate that appropriate innate IL-1α-IL-1R signaling is necessary for IAV clearance and at the same time instructs the formation of organized tertiary lymphoid tissues through induction of CXCL13 early after infection. These findings are discussed in the light of recent insights on the pathogenesis of tertiary lymphoid organ formation in the lung in various diseases where the IL-1 axis is hyperactive, such as rheumatoid arthritis and COPD

Anti-human PD-L1 Nanobody for immuno-PET imaging : validation of a conjugation strategy for clinical translation

Immune checkpoints, such as programmed death-ligand 1 (PD-L1), limit T-cell function and tumor cells use this ligand to escape the anti-tumor immune response. Treatments with monoclonal antibodies blocking these checkpoints have shown long-lasting responses, but only in a subset of patients. This study aims to develop a Nanobody (Nb)-based probe in order to assess human PD-L1 (hPD-L1) expression using positron emission tomography imaging, and to compare the influence of two different radiolabeling strategies, since the Nb has a lysine in its complementarity determining region (CDR), which may impact its affinity upon functionalization. The Nb has been conjugated with the NOTA chelator site-specifically via the Sortase-A enzyme or randomly on its lysines. [68Ga]Ga-NOTA-(hPD-L1) Nbs were obtained in >95% radiochemical purity. In vivo tumor targeting studies at 1 h 20 post-injection revealed specific tumor uptake of 1.89 ± 0.40%IA/g for the site-specific conjugate, 1.77 ± 0.29%IA/g for the random conjugate, no nonspecific organ targeting, and excretion via the kidneys and bladder. Both strategies allowed for easily obtaining 68Ga-labeled hPD-L1 Nbs in high yields. The two conjugates were stable and showed excellent in vivo targeting. Moreover, we proved that the random lysine-conjugation is a valid strategy for clinical translation of the hPD-L1 Nb, despite the lysine present in the CDR

Double-negative T resident memory cells of the lung react to influenza virus infection via CD11chi dendritic cells

Immunity to Influenza A virus (IAV) is controlled by conventional TCR alpha beta(+) CD4(+) and CD8(+) T lymphocytes, which mediate protection or cause immunopathology. Here, we addressed the kinetics, differentiation, and antigen specificity of CD4(-) CD8(-) double-negative (DN) T cells. DNT cells expressed intermediate levels of TCR/CD3 and could be further divided in gamma delta T cells, CD1d-reactive type I NKT cells, NK1.1(+) NKT-like cells, and NK1.1(-) DNT cells. NK1.1(-) DNT cells had a separate antigen-specific repertoire in the steady-state lung, and expanded rapidly in response to IAV infection, irrespectively of the severity of infection. Up to 10% of DNT cells reacted to viral nucleoprotein. Reinfection experiments with heterosubtypic IAV revealed that viral replication was a major trigger for recruitment. Unlike conventional T cells, the NK1.1(-) DNT cells were in a preactivated state, expressing memory markers CD44, CD11a, CD103, and the cytotoxic effector molecule FasL. DNT cells resided in the lung parenchyma, protected from intravascular labeling with CD45 antibody. The recruitment and maintenance of CCR2(+) CCR5(+) CXCR3(+) NK1.1(-) DNT cells depended on CD11c(hi) dendritic cells (DCs). Functionally, DNT cells controlled the lung DC subset balance, suggesting they might act as immunoregulatory cells. In conclusion, we identify activation of resident memory NK1.1(-) DNT cells as an integral component of the mucosal immune response to IAV infection

Early IL-1α signaling is required and sufficient for iBALT induction after influenza virus infection

Inducible Bronchus Associated Lymphoid Tissue (iBALT) is a long lasting tertiary lymphoid tissue that can be induced following influenza A virus (IAV) infection. Previous studies have shown that iBALT structures containing germinal center (GC) B cells protect against repeated infection by contributing locally to the cellular and humoral immune response. If we are to exploit this in vaccination strategies, we need a better understanding on how iBALT structures are induced. One hypothesis is that the strength of the initial innate response dictates induction of iBALT. In the present study, we investigated the role of IL-1 and IL-1R signalling on iBALT formation.Mice lacking the IL-1R, had a delayed viral clearance and thus a prolonged exposure to viral replication, leading to increased disease severity compared to wild type mice. Contradictorily, iBALT formation following clearance of the virus was heavily compromised in Il1r1-/- mice. Quantification of gene induction after IAV infection demonstrated induction of IL-1α and to a much lesser extent of IL-1β. Administration of recombinant IL-1α to the lungs of wild type mice early and late after IAV infection, led to more pronounced iBALT formation and an increased amount of GC B cells in the lungs. Bone marrow chimeric mice identified the stromal compartment as the crucial IL-1 responsive cell for iBALT induction. Mechanistically, Q-PCR analysis of lung homogenates revealed a strongly diminished production of CXCL13, a B cell attracting chemokine, in Il1r-/- mice during the early innate phase of IAV infection. These experiments demonstrate that appropriate innate IL 1α - IL 1R signalling is necessary for IAV clearance and at the same time instructs the formation of organized tertiary lymphoid tissues through induction of CXCL13 early after infection. These findings are discussed in the light of recent insights on the pathogenesis of TLO formation in the lung in various diseases where the IL-1 axis is hyperactive such as rheumatoid arthritis and COPD

Tertiary lymphoid organs in infection and autoimmunity

The lymph nodes (LNs) and spleen have an optimal structure that allows the interaction between T cells, B cells and antigen-presenting dendritic cells (DCs) on a matrix made up by stromal cells. Such a highly organized structure can also be formed in tertiary lymphoid organs (TLOs) at sites of infection or chronic immune stimulation. This review focuses on the molecular mechanisms of TLO formation and maintenance, the controversies surrounding the nature of the inducing events, and the functions of these structures in infection, transplantation and autoimmunity

Conventional and monocyte-derived CD11b⁺ dendritic cells initiate and maintain T helper 2 cell-mediated immunity to house dust mite allergen

Dendritic cells (DCs) are crucial for mounting allergic airway inflammation, but it is unclear which subset of DCs performs this task. By using CD64 and MAR-1 staining, we reliably separated CD11b(+) monocyte-derived DCs (moDCs) from conventional DCs (cDCs) and studied antigen uptake, migration, and presentation assays of lung and lymph node (LN) DCs in response to inhaled house dust mite (HDM). Mainly CD11b(+) cDCs but not CD103(+) cDCs induced T helper 2 (Th2) cell immunity in HDM-specific T cells in vitro and asthma in vivo. Studies in Flt3l(-/-) mice, lacking all cDCs, revealed that moDCs were also sufficient to induce Th2 cell-mediated immunity but only when high-dose HDM was given. The main function of moDCs was the production of proinflammatory chemokines and allergen presentation in the lung during challenge. Thus, we have identified migratory CD11b(+) cDCs as the principal subset inducing Th2 cell-mediated immunity in the LN, whereas moDCs orchestrate allergic inflammation in the lung

Inflammatory type 2 cDCs acquire features of cDC1s and macrophages to orchestrate immunity to respiratory virus infection

The phenotypic and functional dichotomy between IRF8+ type 1 and IRF4+ type 2 conventional dendritic cells (cDC1s and cDC2s, respectively) is well accepted; it is unknown how robust this dichotomy is under inflammatory conditions, when additionally monocyte-derived cells (MCs) become competent antigen-presenting cells (APCs). Using single-cell technologies in models of respiratory viral infection, we found that lung cDC2s acquired expression of the Fc receptor CD64 shared with MCs and of IRF8 shared with cDC1s. These inflammatory cDC2s (inf-cDC2s) were superior in inducing CD4+ T helper (Th) cell polarization while simultaneously presenting antigen to CD8+ T cells. When carefully separated from inf-cDC2s, MCs lacked APC function. Inf-cDC2s matured in response to cell-intrinsic Toll-like receptor and type 1 interferon receptor signaling, upregulated an IRF8-dependent maturation module, and acquired antigens via convalescent serum and Fc receptors. Because hybrid inf-cDC2s are easily confused with monocyte-derived cells, their existence could explain why APC functions have been attributed to MCs