Accelerated waning of immunity to SARS-CoV-2 mRNA vaccines in patients with immune-mediated inflammatory diseases.

BACKGROUNDLimited information is available on the impact of immunosuppressants on COVID-19 vaccination in patients with immune-mediated inflammatory diseases (IMID).METHODSThis observational cohort study examined the immunogenicity of SARS-CoV-2 mRNA vaccines in adult patients with inflammatory bowel disease, rheumatoid arthritis, ankylosing spondylitis, or psoriatic disease, with or without maintenance immunosuppressive therapies. Ab and T cell responses to SARS-CoV-2, including neutralization against SARS-CoV-2 variants, were determined before and after 1 and 2 vaccine doses.RESULTSWe prospectively followed 150 subjects, 26 healthy controls, 9 patients with IMID on no treatment, 44 on anti-TNF, 16 on anti-TNF with methotrexate/azathioprine (MTX/AZA), 10 on anti-IL-23, 28 on anti-IL-12/23, 9 on anti-IL-17, and 8 on MTX/AZA. Ab and T cell responses to SARS-CoV-2 were detected in all participants, increasing from dose 1 to dose 2 and declining 3 months later, with greater attrition in patients with IMID compared with healthy controls. Ab levels and neutralization efficacy against variants of concern were substantially lower in anti-TNF-treated patients than in healthy controls and were undetectable against Omicron by 3 months after dose 2.CONCLUSIONSOur findings support the need for a third dose of the mRNA vaccine and for continued monitoring of immunity in these patient groups.FUNDINGFunded by a donation from Juan and Stefania Speck and by Canadian Institutes of Health (CIHR)/COVID-Immunity Task Force (CITF) grants VR-1 172711 and VS1-175545 (to THW and ACG), CIHR FDN-143250 (to THW), GA2-177716 (to VC, ACG, and THW), and GA1-177703 (to ACG) and the CIHR rapid response network to SARS-CoV-2 variants, CoVaRR-Net (to ACG).

T Cell-Intrinsic CX3CR1 Marks the Most Differentiated Effector CD4+ T Cells, but Is Largely Dispensable for CD4+ T Cell Responses during Chronic Viral Infection.

CD4+ T cells play critical roles during chronic viral infections, but the factors that regulate these responses remain incompletely defined. During chronic infection of mice with lymphocytic choriomeningitis virus clone 13 (LCMV13), the TNFR family member GITR plays a critical CD4+ T cell-intrinsic role in allowing T cell accumulation and viral control. Previously, RNA sequencing of GITR+/+ and GITR-/- T cells sorted from the spleen of mice at day 3 of LCMV13 infection identified the chemokine receptor CX3CR1 as increased by GITR signaling in CD4+ T cells. In this study, we evaluated the role of CX3CR1 on CD4+ T cells during LCMV13 infection. CX3CR1 expression is induced on Ag-specific CD4+ T cells upon Ag stimulation, and GITR signaling further increases the level of CX3CR1 expression. CX3CR1 marks the most differentiated T-bethi, Th1 effector population. Adoptively transferred CX3CR1-/- SMARTA cells had slightly reduced expression of T-bet and IFN-γ per cell compared with their CX3CR1+/+ counterparts but showed no deficit in accumulation in the spleen, lung, or liver. In mixed-radiation chimeras reconstituted with CX3CR1+/+ and CX3CR1-/- bone marrow, CX3CR1+/+ CD4+ T cells showed a marginal deficit in tissue-resident memory T cell numbers compared with the CX3CR1-/- T cells. CX3CR1 may limit acquisition of the tissue-resident memory T cell phenotype because of its effects on increasing T-bet expression, albeit these small effects are unlikely to be of major biological significance. Taken together, these studies show that CX3CR1 marks the most highly differentiated CD4+ Th1 effector population but is largely dispensable for CD4+ T cell responses during chronic viral infection.

GITR differentially affects lung effector T cell subpopulations during influenza virus infection.

Tissue resident memory T cells (Trm) are critical for local protection against reinfection. The accumulation of T cells in the tissues requires a post-priming signal from TNFR superfamily members, referred to as signal 4. Glucocorticoid-induced TNFR-related protein (GITR; TNFRSF18) signaling is important for this post-priming signal and for Trm formation during respiratory infection with influenza virus. As GITR signaling impacts both effector T cell accumulation and Trm formation, we asked if GITR differentially affects subsets of effector cells with different memory potential. Effector CD4+ T cells can be subdivided into 2 populations based on expression of lymphocyte antigen 6C (Ly6C), whereas effector CD8+ cells can be divided into 3 populations based on Ly6C and CX3CR1. The Ly6Chi and CX3CR1hi T cell populations represent the most differentiated effector T cells. Upon transfer, the Ly6Clo CD4+ effector T cells preferentially enter the lung parenchyma, compared to the Ly6Chi CD4+ T cells. We show that GITR had a similar effect on the accumulation of both the Ly6Chi and Ly6Clo CD4+ T cell subsets. In contrast, whereas GITR increased the accumulation of all three CD8+ T cell subsets defined by CX3CR1 and Ly6C expression, it had a more substantial effect on the least differentiated Ly6Clo CX3CR1lo subset. Moreover, GITR selectively up-regulated CXCR6 on the less differentiated CX3CR1lo CD8+ T cell subsets and induced a small but significant increase in CD127 selectively on the Ly6Clo CD4+ T cell subset. Thus, GITR contributes to accumulation of both differentiated effector cells as well as memory precursors, but with some differences between subsets.

Monocyte-Derived Cells in Tissue-Resident Memory T Cell Formation.

There is currently much interest in how different dendritic cell and macrophage populations contribute to T cell-mediated immunity. Although conventional dendritic cell subsets have received much attention for their role in T cell priming, there is emerging evidence for a role for monocyte-derived APC (MoAPC) in tissue-resident memory T cell (Trm) formation. Cells of the monocyte/macrophage lineage play a key role in providing chemokines and cytokines for the localization, differentiation, and survival of Trm and Trm precursors. In addition, inflammatory MoAPC are the key providers of TNF superfamily costimulatory signals, a signal we refer to as signal 4 for T cell activation. Recent evidence suggests that signal 4 from MoAPC occurs postpriming and substantially increases Trm formation. Key questions remain, such as the Ag dependence of signal 4 and the specific mechanisms by which MoAPC-Trm interactions affect the long-term maintenance of Trm.

Phosphoinositide-3 kinase gamma regulates caspase-1 activation and leukocyte recruitment in acute murine gout.

This study investigates the participation of PI3Kγ in the development of joint inflammation and dysfunction in an experimental model of acute gout in mice. Acute gout was induced by injection of monosodium urate (MSU) crystals into the tibiofemoral joint of mice. The involvement of PI3Kγ was evaluated using a selective inhibitor and mice deficient for PI3Kγ (PI3Kγ-/- ) or with loss of kinase activity. Neutrophils recovered from the inflamed joint were quantified and stained for phosphorylated Akt (pAkt) and production of reactive oxygen species (ROS). The adherence of leukocytes to the joint microvasculature was assessed by intravital microscopy and cleaved caspase-1 by Western blot. Injection of MSU crystals induced massive accumulation of neutrophils expressing phosphorylated Akt. In the absence of PI3Kγ, there was reduction of pAkt expression, chemokine production, and neutrophil recruitment. Genetic or pharmacological inhibition of PI3Kγ reduced the adherence of leukocytes to the joint microvasculature, even in joints with established inflammation. Neutrophils from PI3Kγ-/- mice produced less ROS than wild-type neutrophils. There was decreased joint damage and dysfunction in the absence of PI3Kγ. In addition, in the absence of PI3Kγ activity, there was reduction of cleaved caspase-1 and IL-1ß production in synovial tissue after injection of MSU crystals and leukotriene B4 . Our studies suggest that PI3Kγ is crucial for MSU crystal-induced acute joint inflammation. It is necessary for regulating caspase-1 activation and for mediating neutrophil migration and activation. Drugs that impair PI3Kγ function may be useful to control acute gout inflammation.

4-1BB Regulates Effector CD8 T Cell Accumulation in the Lung Tissue through a TRAF1-, mTOR-, and Antigen-Dependent Mechanism to Enhance Tissue-Resident Memory T Cell Formation during Respiratory Influenza Infection.

The TNFR superfamily member 4-1BB is important in the establishment of tissue-resident memory T cells (Trm) in the lung tissue following influenza infection. Moreover, supraphysiological boosting of 4-1BB in the airways during the boost phase of a prime-boost immunization regimen increases the long-lived Trm population, correlating with increased protection against heterotypic challenge. However, little is known about how 4-1BB contributes to the establishment of the lung Trm population. In this study, we show that effects of 4-1BB on lung Trm accumulation are already apparent at the effector stage, suggesting that the major role of 4-1BB in Trm formation is to allow persistence of CD8 T effector cells in the lung as they transition to Trm. Using supraphysiological stimulation of 4-1BB in the boost phase of a prime-boost immunization, we show that the effect of 4-1BB on Trm generation requires local delivery of both Ag and costimulation, is inhibited by rapamycin treatment during secondary CD8 effector T cell expansion, and is dependent on the signaling adaptor TRAF1. The decrease in lung Trm following early rapamycin treatment is accompanied by increased circulating memory T cells, as well as fewer effectors, suggesting a role for mammalian target of rapamycin (mTOR) in the formation of Trm through effects on the accumulation of effector precursors. Taken together, these data point to an important role for 4-1BB, TRAF1, and mTOR in the persistence of CD8 effector T cells in the lung parenchyma, thereby allowing the transition to Trm.

The Long Pentraxin 3 Contributes to Joint Inflammation in Gout by Facilitating the Phagocytosis of Monosodium Urate Crystals.

The purpose of this study was to investigate the role of pentraxin 3 (PTX3), a pivotal component of the innate immune system, in gout. Levels of PTX3 and IL-1ß in human samples were evaluated by ELISA. Development of murine gout was evaluated through the levels of cytokines (PTX3, CXCL1, and IL-1ß) and neutrophil recruitment into the joint cavity. Phagocytosis of monosodium urate (MSU) crystals and caspase-1 activation were determined by flow cytometer. Acute gout patients showed elevated concentration of PTX3 in plasma and synovial fluid as compared with healthy and osteoarthritic subjects. Moreover, there was a positive correlation between intra-articular PTX3 and IL-1ß levels. PTX3 was induced in the periarticular tissue of mice postinjection of MSU crystals. Importantly, Ptx3-deficient mice showed reduced inflammation in response to MSU crystal injection compared with wild-type mice, including reduction of neutrophil recruitment into the joint cavity and IL-1ß and CXCL1 production. Interestingly, addition of PTX3 in vitro enhanced MSU crystal phagocytosis by monocytes and resulted in higher production of IL-1ß by macrophages. This contribution of PTX3 to the phagocytosis of MSU crystals and consequent production of IL-1ß occurred through a mechanism mainly dependent on FcγRIII. Thus, our results suggest that PTX3 acts as a humoral pattern recognition molecule in gout facilitating MSU crystal phagocytosis and contributing to the pathogenesis of gouty arthritis.

GITRL on inflammatory antigen presenting cells in the lung parenchyma provides signal 4 for T-cell accumulation and tissue-resident memory T-cell formation.

T-cell responses in the lung are critical for protection against respiratory pathogens. TNFR superfamily members play important roles in providing survival signals to T cells during respiratory infections. However, whether these signals take place mainly during priming in the secondary lymphoid organs and/or in the peripheral tissues remains unknown. Here we show that under conditions of competition, GITR provides a T-cell intrinsic advantage to both CD4 and CD8 effector T cells in the lung tissue, as well as for the formation of CD4 and CD8 tissue-resident memory T cells during respiratory influenza infection in mice. In contrast, under non-competitive conditions, GITR has a preferential effect on CD8 over CD4 T cells. The nucleoprotein-specific CD8 T-cell response partially compensated for GITR deficiency by expansion of higher affinity T cells; whereas, the polymerase-specific response was less flexible and more GITR dependent. Following influenza infection, GITR is expressed on lung T cells and GITRL is preferentially expressed on lung monocyte-derived inflammatory antigen presenting cells. Accordingly, we show that GITR+/+ T cells in the lung parenchyma express more phosphorylated-ribosomal protein S6 than their GITR-/- counterparts. Thus, GITR signaling within the lung tissue critically regulates effector and tissue-resident memory T-cell accumulation.

Conserved and Differential Features of TNF Superfamily Ligand Expression on APC Subsets over the Course of a Chronic Viral Infection in Mice.

There is currently much interest in how different APC subsets shape the immune response. We recently described a division of labor between classical dendritic cells (cDC) and inflammatory monocyte-derived APC in provision of costimulatory ligands to T cells early during chronic lymphocytic choriomeningitis clone 13 (LCMV 13) infection in mice. At day 2 of LCMV 13 infection, cDC preferentially express CD80 and CD86, whereas TNF superfamily ligands GITRL, 4-1BBL, CD70, and OX40L are preferentially induced by type I IFN on inflammatory monocyte-derived APC, with minimal expression on cDC. In this study, we further investigate the expression of TNF and B7 family ligands on APC over the course of LCMV 13 infection. OX40L and 4-1BBL remain above baseline through the chronic stage of infection, with predominant expression on inflammatory APC compared with cDC in the spleen, partially blocked by anti-IFN-γR Ab pretreatment. Conversely, CD70, like GITRL, returns to baseline on the APC within a few days postinfection. In the lung, TNF family ligands were also preferentially expressed on inflammatory monocyte-derived APC. CD86 was generally higher on cDC than inflammatory APC in the spleen, but in the lung CD86 was highest on inflammatory APC. Moreover, in the spleen, CD80 levels on different APC subsets fluctuated over the course of the infection. We also show that LPS induction of TNF superfamily ligands is largely mediated through type I IFN. This study highlights the importance of IFNs and monocyte-derived APC in TNF superfamily ligand expression in both secondary lymphoid organs and tissues during chronic viral infection.

Tumor Necrosis Factor, but Not Neutrophils, Alters the Metabolic Profile in Acute Experimental Arthritis.

Metabolic alterations are associated with arthritis apart from obesity. However, it is still unclear which is the underlying process behind these metabolic changes. Here, we investigate the role of tumor necrosis factor (TNF) in this process in an acute model of antigen-induced arthritis (AIA). Immunized male BALB/c mice received an intra-articular injection of PBS (control) or methylated bovine serum albumin (mBSA) into their knees, and were also pre-treated with different drugs: Etanercept, an anti-TNF drug, DF2156A, a CXCR1/2 receptor antagonist, or a monoclonal antibody RB6-8C5 to deplete neutrophils. Local challenge with mBSA evoked an acute neutrophil influx into the knee joint, and enhanced the joint nociception, along with a transient systemic metabolic alteration (higher levels of glucose and lipids, and altered adipocytokines). Pre-treatment with the conventional biological Etanercept, an inhibitor of TNF action, ameliorated the nociception and the acute joint inflammation dominated by neutrophils, and markedly improved many of the altered systemic metabolites (glucose and lipids), adipocytokines and PTX3. However, the lessening of metabolic changes was not due to diminished accumulation of neutrophils in the joint by Etanercept. Reduction of neutrophil recruitment by pre-treating AIA mice with DF2156A, or even the depletion of these cells by using RB6-8C5 reduced all of the inflammatory parameters and hypernociception developed after AIA challenge, but could not prevent the metabolic changes. Therefore, the induction of joint inflammation provoked acute metabolic alterations which were involved with TNF. We suggest that the role of TNF in arthritis-associated metabolic changes is not due to local neutrophils, which are the major cells present in this model, but rather due to cytokines.

Antioxidative and immunomodulatory effects of tributyrin supplementation on experimental colitis.

Tributyrin (TBT) is a TAG composed of three butyric acids that has beneficial effects on ulcerative colitis due to its trophic, anti-inflammatory, pro-apoptotic and anti-carcinogenic properties. The goal of the present study was to evaluate the efficacy and mechanisms of action of TBT supplementation in the prevention of mucosal damage in experimental colitis. Mice received either a control diet or a TBT-supplemented diet for 15 d. Colitis was induced by dextran sodium sulphate administration during the last 7 d. Mucosal damage and the activation of immune cells and cytokines were determined by histological score, flow cytometry and ELISA. Leucocyte rolling and adhesion were assessed by intravital microscopy. Oxidative stress was determined by monitoring hydroperoxide concentration and evaluating superoxide dismutase (SOD) and catalase activities. Intestinal permeability was analysed using diethylenetriaminepentaacetate acid (99mTcDTPA). Compared with the colitis group, the animals in the colitis+TBT group had reduced mucosal damage and neutrophil and eosinophil mucosal infiltration, which were associated with a higher percentage of regulatory T cells (Treg) and higher levels of transforming growth factor ß and IL-10 in the lamina propria. The level of in vivo leucocyte adhesion in the colon microvasculature was reduced after TBT supplementation. A lower level of hydroperoxide and higher levels of SOD and catalase activities were associated with TBT supplementation. TBT-supplemented mice showed reduced intestinal permeability to the levels intermediate between the control and colitis groups. In conclusion, the present results show that TBT has positive effects on colonic restructuring in experimental colitis. Additionally, TBT supplementation changes the immune response by controlling inflammation and regulating the expression of anti-inflammatory cytokines and Treg.

The combination of high-fat diet-induced obesity and chronic ulcerative colitis reciprocally exacerbates adipose tissue and colon inflammation.

BACKGROUND: This study evaluated the relationship between ulcerative colitis and obesity, which are both chronic diseases characterized by inflammation and increases in immune cells and pro-inflammatory cytokines. METHODS: Mice with chronic ulcerative colitis induced by 2 cycles of dextran sodium sulfate (DSS) in the first and fourth week of the experiment were fed a high-fat diet (HFD) to induce obesity by 8 weeks. The animals were divided into 4 \ groups (control, colitis, HFD and colitis + HFD). RESULTS: Obesity alone did not raise histopathology scores, but the combination of obesity and colitis worsened the scores in the colon compared to colitis group. Despite the reduction in weight gain, there was increased inflammatory infiltrate in both the colon and visceral adipose tissue of colitis + HFD mice due to increased infiltration of macrophages, neutrophils and lymphocytes. Intravital microscopy of VAT microvasculature showed an increase in leukocyte adhesion and rolling and overexpression of adhesion molecules compared to other groups. Moreover, circulating lymphocytes, monocytes and neutrophils in the spleen and cecal lymph nodes were increased in the colitis + HFD group. CONCLUSION: Our results demonstrated the relationship between ulcerative colitis and obesity as aggravating factors for each disease, with increased inflammation in the colon and adipose tissue and systemic alterations observed in the spleen, lymph nodes and bloodstream.