T cell receptor reversed polarity recognition of a self-antigen major histocompatibility complex.

Central to adaptive immunity is the interaction between the αß T cell receptor (TCR) and peptide presented by the major histocompatibility complex (MHC) molecule. Presumably reflecting TCR-MHC bias and T cell signaling constraints, the TCR universally adopts a canonical polarity atop the MHC. We report the structures of two TCRs, derived from human induced T regulatory (iT(reg)) cells, complexed to an MHC class II molecule presenting a proinsulin-derived peptide. The ternary complexes revealed a 180° polarity reversal compared to all other TCR-peptide-MHC complex structures. Namely, the iT(reg) TCR α-chain and ß-chain are overlaid with the α-chain and ß-chain of MHC class II, respectively. Nevertheless, this TCR interaction elicited a peptide-reactive, MHC-restricted T cell signal. Thus TCRs are not 'hardwired' to interact with MHC molecules in a stereotypic manner to elicit a T cell signal, a finding that fundamentally challenges our understanding of TCR recognition.

Tolerogenic dendritic cells impede priming of naïve CD8⁺ T cells and deplete memory CD8⁺ T cells.

Type 1 diabetes is a T-cell-mediated autoimmune disease in which autoreactive CD8(+) T cells destroy the insulin-producing pancreatic beta cells. Vitamin D3 and dexamethasone-modulated dendritic cells (Combi-DCs) loaded with islet antigens inducing islet-specific regulatory CD4(+) T cells may offer a tissue-specific intervention therapy. The effect of Combi-DCs on CD8(+) T cells, however, remains unknown. To investigate the interaction of CD8(+) T cells with Combi-DCs presenting epitopes on HLA class I, naive, and memory CD8(+) T cells were co-cultured with DCs and proliferation and function of peptide-specific T cells were analyzed. Antigen-loaded Combi-DCs were unable to prime naïve CD8(+) T cells to proliferate, although a proportion of T cells converted to a memory phenotype. Moreover, expansion of CD8(+) T cells that had been primed by mature monocyte-derived DCs (moDCs) was curtailed by Combi-DCs in co-cultures. Combi-DCs expanded memory T cells once, but CD8(+) T-cell numbers collapsed by subsequent re-stimulation with Combi-DCs. Our data point that (re)activation of CD8(+) T cells by antigen-pulsed Combi-DCs does not promote, but rather deteriorates, CD8(+) T-cell immunity. Yet, Combi-DCs pulsed with CD8(+) T-cell epitopes also act as targets of cytotoxicity, which is undesirable for survival of Combi-DCs infused into patients in therapeutic immune intervention strategies.

Infectious tolerance as candidate therapy for type 1 diabetes: transfer of immunoregulatory properties from human regulatory T cells to other T cells and proinflammatory dendritic cells.

Type 1 diabetes is a T-cell-mediated autoimmune disease in which the insulin-producing cells in the islets of Langerhans are selectively destroyed. Although symptomatic insulin therapy is able to control glucose levels in the blood, many patients do not obtain the desired glycemic control, which increases the risk of diabetic complications. While many immunotherapeutic efforts to intervene in the disease process focus on systemic immune-suppressive therapies, antigen-specific immune modulation represents an attractive alternative. Dendritic cells modulated with 1.25(OH)2 vitamin D3, and dexamethasone (tolerogenic DCs) loaded with islet antigens induce Ag-specific regulatory CD4 T cells (iaTregs), offering a tissue-specific intervention therapy. iaTregs exert their function via linked suppression to diminish effector cells by modulating pro-inflammatory DCs to upregulate inhibitory receptors. In turn, these re-educated mature DCs induce IL-10-producing cells from the naïve T-cell pool. Thus, tolerogenic DCs transfer regulatory properties to pro-inflammatory DCs via iaTregs (i.e., infectious tolerance). In this review, we describe the current knowledge regarding regulatory mechanisms of these tolerogenic DCs and the Tregs that they induce, and we propose that cell therapy with human tolerogenic DCs provides new opportunities for immune intervention in patients with autoimmune diseases.

Tailored anticoagulant treatment after a first venous thromboembolism: protocol of the Leiden Thrombosis Recurrence Risk Prevention (L-TRRiP) study - cohort-based randomised controlled trial.

INTRODUCTION: Patients with a first venous thromboembolism (VTE) are at risk of recurrence. Recurrent VTE (rVTE) can be prevented by extended anticoagulant therapy, but this comes at the cost of an increased risk of bleeding. It is still uncertain whether patients with an intermediate recurrence risk or with a high recurrence and high bleeding risk will benefit from extended anticoagulant treatment, and whether a strategy where anticoagulant duration is tailored on the predicted risks of rVTE and bleeding can improve outcomes. The aim of the Leiden Thrombosis Recurrence Risk Prevention (L-TRRiP) study is to evaluate the outcomes of tailored duration of long-term anticoagulant treatment based on individualised assessment of rVTE and major bleeding risks. METHODS AND ANALYSIS: The L-TRRiP study is a multicentre, open-label, cohort-based, randomised controlled trial, including patients with a first VTE. We classify the risk of rVTE and major bleeding using the L-TRRiP and VTE-BLEED scores, respectively. After 3 months of anticoagulant therapy, patients with a low rVTE risk will discontinue anticoagulant treatment, patients with a high rVTE and low bleeding risk will continue anticoagulant treatment, whereas all other patients will be randomised to continue or discontinue anticoagulant treatment. All patients will be followed up for at least 2 years. Inclusion will continue until the randomised group consists of 608 patients; we estimate to include 1600 patients in total. The primary outcome is the combined incidence of rVTE and major bleeding in the randomised group after 2 years of follow-up. Secondary outcomes include the incidence of rVTE and major bleeding, functional outcomes, quality of life and cost-effectiveness in all patients. ETHICS AND DISSEMINATION: The protocol was approved by the Medical Research Ethics Committee Leiden-Den Haag-Delft. Results are expected in 2028 and will be disseminated through peer-reviewed journals and during (inter)national conferences. TRIAL REGISTRATION NUMBER: NCT06087952.

Transfer of regulatory properties from tolerogenic to proinflammatory dendritic cells via induced autoreactive regulatory T cells.

Infectious tolerance is a term generally assigned to the process through which regulatory T cells (Tregs) transfer immunoregulatory properties to other T cells. In this study, we demonstrated that a similar process applies to human dendritic cells (DCs), albeit through a different mechanism. We induced and cloned proinsulin-specific Tregs using tolerogenic DCs and investigated mechanisms by which induced Ag-specific regulatory T cells (iaTregs) endorse the suppressive effects. iaTregs expressed FOXP3, programmed death-1, and membrane-bound TGF-ß and upregulated IL-10 and CTLA-4 after stimulation with the cognate Ag. The iaTregs suppressed effector T cells only when both encountered the cognate Ags on the same APCs (linked suppression). This occurred independently of IL-10, TGF-ß, programmed death-1, or CTLA-4. Instead, iaTregs used a granzyme B-mediated mechanism to kill B cells and monocytes, whereas proinflammatory DCs that resisted being killed were induced to upregulate the inhibitory receptors B7 (family) homolog 3 and ICOS ligand. These re-educated mature monocyte-derived dendritic cells (mDCs) suppressed effector T cells and induced IL-10-producing cells from the naive T cell pool. Our data indicated that human tolerogenic DCs confer infectious tolerance by inducing Ag-specific Tregs, which, in turn, re-educate proinflammatory mature DCs into DCs with regulatory properties.

Differential protein pathways in 1,25-dihydroxyvitamin d(3) and dexamethasone modulated tolerogenic human dendritic cells.

Tolerogenic dendritic cells (DC) that are maturation-resistant and locked in a semimature state are promising tools in clinical applications for tolerance induction. Different immunomodulatory agents have been shown to induce a tolerogenic DC phenotype, such as the biologically active form of vitamin D (1,25(OH)(2)D(3)), glucocorticoids, and a synergistic combination of both. In this study, we aimed to characterize the protein profile, function and phenotype of DCs obtained in vitro in the presence of 1,25(OH)(2)D(3), dexamethasone (DEX), and a combination of both compounds (combi). Human CD14(+) monocytes were differentiated toward mature DCs, in the presence or absence of 1,25(OH)(2)D(3) and/or DEX. Cells were prefractionated into cytoplasmic and microsomal fractions and protein samples were separated in two different pH ranges (pH 3-7NL and 6-9), analyzed by 2D-DIGE and differentially expressed spots (p < 0.05) were identified after MALDI-TOF/TOF analysis. In parallel, morphological and phenotypical analyses were performed, revealing that 1,25(OH)(2)D(3)- and combi-mDCs are closer related to each other than DEX-mDCs. This was translated in their protein profile, indicating that 1,25(OH)(2)D(3) is more potent than DEX in inducing a tolerogenic profile on human DCs. Moreover, we demonstrate that combining 1,25(OH)(2)D(3) with DEX induces a unique protein expression pattern with major imprinting of the 1,25(OH)(2)D(3) effect. Finally, protein interaction networks and pathway analysis suggest that 1,25(OH)(2)D(3), rather than DEX treatment, has a severe impact on metabolic pathways involving lipids, glucose, and oxidative phosphorylation, which may affect the production of or the response to ROS generation. These findings provide new insights on the molecular basis of DC tolerogenicity induced by 1,25(OH)(2)D(3) and/or DEX, which may lead to the discovery of new pathways involved in DC immunomodulation.

Critical role for TNF in the induction of human antigen-specific regulatory T cells by tolerogenic dendritic cells.

TNF is a pleiotropic cytokine with differential effects on immune cells and diseases. Anti-TNF therapy was shown to be effective in rheumatoid arthritis but proved inefficient or even detrimental in other autoimmune diseases. We studied the role of TNF in the induction of Ag-specific regulatory T cells (Tregs) by tolerogenic vitamin D3-modulated human dendritic cells (VD3-DCs), which previously were shown to release high amounts of soluble TNF (sTNF) upon maturation with LPS. First, production of TNF by modulated VD3-DCs was analyzed upon maturation with LPS or CD40L with respect to both secreted (cleaved) TNF (sTNF) and expression of the membrane-bound (uncleaved) form of TNF (mTNF). Next, TNF antagonists were tested for their effect on induction of Ag-specific Tregs by modulated DCs and the subsequent functionality of these Tregs. VD3-DCs expressed greater amounts of mTNF than did control DCs (nontreated DCs), independent of the maturation protocol. Inhibition of TNF with anti-TNF Ab (blocking both sTNF and mTNF) during the priming of Tregs with VD3-DCs prevented generation of Tregs and their suppression of proliferation of CD4(+) T cells. In contrast, sTNF receptor II (sTNFRII), mainly blocking sTNF, did not change the suppressive capacity of Tregs. Blocking of TNFRII by anti-CD120b Ab during Treg induction similarly abrogated their subsequent suppressive function. These data point to a specific role for mTNF on VD3-DCs in the induction of Ag-specific Tregs. Interaction between mTNF and TNFRII instructs the induction of suppressive Tregs by VD3-DCs. Anti-TNF therapy may therefore act adversely in different patients or disease pathways.

Induction of Treg by monocyte-derived DC modulated by vitamin D3 or dexamethasone: differential role for PD-L1.

Specific therapy with modulated DC may restore immunological tolerance, thereby obviating the need for chronic immunosuppression in transplantation or autoimmunity. In this study we compared the tolerizing capacity of dexamethasone (Dex)- and 1 alpha,25-dihydroxyvitamin D3 (VD3)-modulated DC. Treatment of monocytes with either VD3 or Dex resulted in DC with stable, semi-mature phenotypes compared with standard DC, with intermediate levels of co-stimulatory and MHC class II molecules, which remained unaltered after subsequent pro-inflammatory stimulation. IL-12p70 secretion was lost by VD3- and Dex-DC, whereas IL-10 secretion was unaffected. VD3-DC distinctly produced large amounts of TNF-alpha. Both VD3- and Dex-DC possessed the capacity to convert CD4 T cells into IL-10-secreting Treg potently suppressing the proliferation of responder T cells. However, only Treg induced by VD3-DC exhibited antigen specificity. VD3-, but not Dex-, DC expressed significant high levels of PD-L1 (programmed death-1 ligand), upon activation. Blockade of PD-L1 during priming redirected T cells to produce IFN-gamma instead of IL-10 and abolished acquisition of regulatory capacity. Our findings demonstrate that both VD3- and Dex-DC possess durable but differential tolerogenic features, acting via different mechanisms. Both are potentially useful to specifically down-regulate unwanted immune responses and induce immune tolerance. These modulated DC appear suitable as adjuvant in antigen-specific clinical vaccination intervention strategies.