Decreased Tim-3 and its correlation with Th1 cells in patients with immune thrombocytopenia

Here we evaluate the expression of Tim-3 in CD4 + T cells from patients with immune thrombocytopenia (ITP). We also counted platelets and evaluated plasma IFN-γ, IL-18 and IL-4 levels in patients with active ITP (n = 26), patients with ITP in remission (n = 23) and in healthy subjects (n = 34) by enzyme linked immunosorbent assay (ELISA). Using real-time quantitative polymerase chain reaction (RT-PCR), the mRNA expression of IL-18, IFN-γ, IL-4, T-box (T-bet) and Tim-3 was studied in the blood. The CD4 + Tim-3 + cells in blood were evaluated by flow cytometry and are expressed as a percentage of the total number of CD4 + cells. The Tim-3 positive cells within the circulating CD4 + population of newly diagnosed patients were significantly decreased compared to controls. However, T-bet, IL-18 and IFN-γ levels were significantly elevated in patients. Tim-3 mRNA expression was significantly lower in the peripheral mononuclear cells (PBMCs) of ITP patients compared to controls. The decreased levels of Tim-3 during active stages of disease suggest a possible role in the pathogenesis and course of ITP and restoration of Tim-3 may be a reasonable therapeutic strategy for ITP.     

Distinct expression of Tim-3 during different stages of rat experimental autoimmune neuritis

Experimental autoimmune neuritis (EAN) is a well-known animal model of human demyelinating polyneuropathies and is characterized by inflammation and demyelination in the peripheral nervous system. Tim-3 had been identified as a Th1-specific marker negatively regulating autoimmunity or inflammatory diseases. Here we have studied by immunohistochemistry the spatiotemporal accumulation of Tim-3(+) cells in sciatic nerves of EAN rats, particularly focusing on its association with alternatively activated macrophages. Our results showed that time course of Tim-3(+) cell accumulation correlated positively with disease progression of EAN; but distinct major cellular resources of Tim-3 were observed at different disease stages of EAN: during the early phase of EAN, the main cellular resource were T cells, but at the peak and during recovery phase of EAN, Tim-3 was mostly expressed on CD68(+) macrophages or CD163(+) cells. Further investigation suggested that accumulation of CD163(+) cells, particularly their relative abundances to activated macrophages at different time points, were in accordance with the recovery from EAN. Therefore, Tim3(+) cells might include a distinct macrophage population, which may be involved in anti-inflammatory effect and recovery from EAN.     

Inflammatory cells, microglia and MHC class II antigen-positive cells in the spinal cord of Lewis rats with acute and chronic relapsing experimental autoimmune encephalomyelitis

Chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) was induced in Lewis rats by inoculation with guinea pig spinal cord and adjuvants and treatment with low dose cyclosporin A (CsA). Acute EAE was induced by the same method without CsA treatment. Immunocytochemistry and flow cytometry were used to assess inflammatory cells and MHC class II (Ia) antigen expression in the central nervous system of these rats. The inflammatory infiltrate was composed mainly of CD4⁺ T cells and macrophages, and αß T cells constituted about 65% of the CD2⁺ T cells. After recovery from acute EAE and during the first remission of CR-EAE, the number of T cells was significantly less than in the preceding episodes. The number of T cells was higher in the second episode of CR-EAE than in the first remission. Throughout the course of CR-EAE, the majority of the CD2⁺ T cells were CD45RC⁻. The ratio of IL-2R⁺ cells to CD2⁺ cells ranged from 10.5 to 24.0%. The ratio of CD4⁺ T cells to B cells was lower in the later episodes of CR-EAE than in the first episode. Ia antigen was expressed on filtrating round cells at all stages of CR-EAE and on microglial cells (identified by dendritic morphology) with increasing intensity throughout the course of CR-EAE. With flow cytometry, the number of Ia⁺ cells obtained from the spinal cord rose throughout the course of CR-EAE. The number of FSC^lowOX1^low cells, which we consider represent microglia, also increased during the course of CR-EAE.     

Oligodendrocyte lysis by CD4+ T cells independent of tumor necrosis factor

The capacity of human CD4⁺ T cells to lyse heterologous human oligodendrocytes in an 18-hour chromium 51–release assay was compared to that of systemic blood-derived macrophages and central nervous system–derived microglia. CD4⁺ T cells, activated with either phytohemagglutinin, anti-CD3 antibody, or antigen (myelin basic protein), could induce lysis of the oligodendrocytes whereas macrophages and microglia, activated with interferon-γ and lipopolysaccharide, could not. The CD4⁺ T-cell effect was not inhibited with an anti–tumor necrosis factor-α–neutralizing antibody. Both the CD4⁺ T cells and the macrophages could induce lysis of tumor necrosis factor–sensitive rodent cell lines, Wehi 164, and L929; these effects were inhibited with anti–tumor necrosis factor antibody. Pretreatment of the CD4⁺ T cells with cyclosporine or mitomycin C did not inhibit oligodendrocyte lysis. These results indicate that at least in vitro, CD4⁺ T cells can induce a form of oligodendrocyte injury that is not reproduced by macrophages or microglia or by tumor necrosis factor. The non–major histocompatibility complex (MHC)–restricted injury of oligodendrocytes induced by both myelin antigen–reactive and mitogen-stimulated T cells may provide a basis whereby cytotoxic CD4⁺ T cells could interact with a target cell that does not express MHC class II molecules. Our results suggest that immune-mediated oligodendrocyte/myelin injury, as is postulated to occur in the disease multiple sclerosis, may involve multiple effector mechanisms.     

Difficulty diagnosing chronic cryptococcal meningitis in idiopathic CD4+ lymphocytopenia

A 64-year-old man with idiopathic CD4⁺ lymphocytopenia developed cognitive impairment and gait ataxia with isolated obstructive hydrocephalus, which was fatal. Cerebrospinal fluid showed mild pleocytosis, but the etiology was not revealed by extensive analysis. At autopsy, inflammatory cells, CD8⁺ lymphocytes and abundant macrophages but not CD4⁺ lymphocytes were infiltrating the meninges at the base of the brain. Electron microscopy demonstrated that inflammation was caused by Cryptococcus neoformans, which was localized exclusively within macrophages, where it grew with budding. Our study suggests that, in idiopathic CD4⁺ lymphocytopenia, macrophages can efficiently phagocytize but inefficiently digest C. neoformans, thus representing a vehicle of chronic intracellular infection.     

Peripheral leukocyte profile in people with temporal lobe epilepsy reflects the associated proinflammatory state

IntroductionMarkers of low-grade peripheral inflammation have been reported amongst people with epilepsy. The mechanisms underlying this phenomenon are unknown. We attempted to characterize peripheral immune cells and their activation status in people with temporal lobe epilepsy (TLE) and healthy controls.Methods and resultsTwenty people with TLE and 19 controls were recruited, and peripheral blood lymphocyte and monocyte subsets evaluated ex vivo by multi-color flow cytometry. People with TLE had higher expression of HLA-DR, CD69, CTLA-4, CD25, IL-23R, IFN-γ, TNF and IL-17 in CD4⁺ lymphocytes than controls. Granzyme A, CTLA-4, IL-23R and IL-17 expression was also elevated in CD8⁺ T cells from people with TLE. Frequency of HLA-DR in CD19⁺ B cells and regulatory T cells CD4⁺CD25⁺Foxp3⁺ producing IL-10 was higher in TLE when compared with controls. A negative correlation between CD4⁺ expressing co-stimulatory molecules (CD69, CD25 and CTLA-4) with age at onset of seizures was found. The frequency of CD4⁺CD25⁺Foxp3⁺ cells was also positively correlated with age at onset of seizures.ConclusionImmune cells of people with TLE show an activation profile, mainly in effector T cells, in line with the low-grade peripheral inflammation.     

Clinical significance of Tim3-positive T cell subsets in patients with multiple sclerosis

The present study evaluated associations between the percentages of T cell immunoglobulin and mucin domain 3 (Tim3)-positive T cells and related cytokines and multiple sclerosis (MS). We collected peripheral blood samples from 30 MS patients and 30 healthy controls. Flow cytometry was used to determine the proportions of CD3⁺Tim3⁺, CD4⁺Tim3⁺, and CD4⁺CD25⁺Tim3⁺ in peripheral blood mononuclear cells (PBMCs) and related cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ also were determined using enzyme-linked immunosorbent assays (ELISA). The percentages of Tim3-positive T cells in CD4⁺ and CD4⁺CD25⁺ T cell subsets were significantly lower among MS patients than among controls. This difference was particularly evident in the CD4⁺CD25(high) T cell subset. The proportions of CD4⁺Tim3⁺ and CD4⁺CD25⁺Tim3⁺ cells in PBMCs were significantly lower in the MS group than in the control group, whereas no significant differences were detected regarding the percentages of CD3⁺Tim3⁺ in PBMCs and T cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ all were increased in MS patients compared with healthy controls. Our results support that Tim3 and related cytokines may be involved in the onset of MS.     

Dopamine receptor D2 on CD4+ T cells is protective against neuroinflammation and neurodegeneration in a mouse model of Parkinson's disease

Parkinson's disease (PD) is a chronic neurodegenerative disease. Recently, neuroinflammation driven by CD4⁺ T cells has been involved in PD pathophysiology. Human and murine lymphocytes express all the five subtypes of dopamine receptors (DRs), DRD1 to DRD5. However, roles of DRs particularly DRD2 expressed on CD4⁺ T cells in PD remain elucidated. Global Drd1- or Drd2-knockout (Drd1^−/− or Drd2^−/−) mice or CD4⁺ T cell-specific Drd2-knockout (Drd2^fl/fl/CD4^Cre) mice were intraperitoneally injected with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to induce PD with the different mutants. On the 7th day following MPTP injection, mice were assessed for dopaminergic neurodegeneration, locomotor impairments, microglial activation, as well as CD4⁺ T-cell differentiation and function. Furthermore, in vitro CD4⁺ T cells were exposed to DRD2 agonist and antagonist and then differentiation and function of the cells were determined. MPTP induced dopaminergic neuronal loss in the nigrostriatal system, motor coordinative and behavioral impairments, microglial activation, and CD4⁺ T-cell polarization to pro-inflammatory T-helper (Th)1 and Th17 phenotypes. Importantly, either Drd2^−/− or Drd2^fl/fl/CD4^Cre mice manifested more severe dopaminergic neurodegeneration, motor deficits, microglial activation, and CD4⁺ T-cell bias towards Th1 and Th17 phenotypes in response to MPTP, but Drd1^−/− did not further alter MPTP intoxication. DRD2 agonist sumanirole inhibited shift of CD4⁺ T cells obtained from MPTP-intoxicated mice to Th1 and Th17 phenotypes and DRD2 antagonist L-741,626 reversed sumanirole effects. These findings suggest that DRD2 expressed on CD4⁺ T cells is protective against neuroinflammation and neurodegeneration in PD. Thus, developing a therapeutic strategy of stimulating DRD2 may be promising for mitigation of PD.     

Reduction of rat brain CD8+ T‐cells by levodopa/benserazide treatment after experimental stroke

The activation of inflammatory cascades in the ischemic hemisphere impairs mechanisms of tissue reorganization with consequences for recovery of lost neurological function. Recruitment of T‐cell populations to the post‐ischemic brain occurs and represents a significant part of the inflammatory response. This study was conducted to investigate if treatment with levodopa, potentially acting as an immunomodulator, affects the T‐cell accumulation in the post‐ischemic brain. Male Sprague–Dawley rats were subjected to transient occlusion of the middle cerebral artery (tMCAO) for 105 min followed by levodopa/benserazide treatment (20 mg/kg/15 mg/kg) for 5 days initiated on day 2 post‐stroke. One week after tMCAO, T‐cell populations were analysed from brains, and levels of interleukin (IL)‐1β, chemokine (C‐X‐C motif) ligand 1, IL‐4, IL‐5, interferon gamma and IL‐13 were analysed. After levodopa/benserazide treatment, we found a significant reduction of cytotoxic T‐cells (CD3⁺CD8⁺) in the ischemic hemisphere together with reduced levels of T‐cell‐associated cytokine IL‐5, while other T‐cell populations (CD3⁺, CD3⁺CD4⁺, CD3⁺CD4⁺CD25⁺) were unchanged compared with vehicle‐treated rats. Moreover, a reduced number of cells was associated with reduced levels of intercellular adhesion molecule 1, expressed in endothelial cells, in the infarct core of levodopa/benserazide‐treated animals. Together, we provide the first evidence that dopamine can act as a potential immunomodulator by attenuating inflammation in the post‐ischemic brain.     

Repeated bouts of aerobic exercise enhance regulatory T cell responses in a murine asthma model

We have reported previously that moderate intensity aerobic exercise training attenuates airway inflammation in a murine asthma model. Recent studies implicate regulatory T (Treg) cells in decreasing asthma-related airway inflammation; as such, the current study examined the effect of exercise on Treg cell function in a murine asthma model. Mice were sensitized with ovalbumin (OVA) prior to the start of exercise training at a moderate intensity 3×/week for 4 weeks; exercise was performed as treadmill running (13.5 m/min, 0% grade). Mice were OVA challenged repeatedly throughout the exercise protocol. At protocol completion, mice were analyzed for changes in the number and suppressive function of CD4⁺CD25⁺Foxp3⁺ cells isolated from lungs, mediastinal lymph nodes, and spleens. Results show that exercise increased significantly the number of Foxp3⁺ cells within the lungs and mediastinal lymph nodes, but not the spleens, of OVA-treated mice as compared with sedentary controls. Exercise also enhanced the suppression function of CD4⁺CD25⁺Foxp3⁺ Treg cells derived from OVA-treated mice as compared with sedentary controls. Specifically, Treg cells from exercised, OVA-treated mice more effectively suppressed CD4⁺CD25^? cell proliferation and Th2 cytokine production in vitro. Enhanced suppression was associated with increased protein levels of TGF-β and lesser amounts of IL-10 and IL-17; however, blocking TGF-β had no effect on suppressive functions. These data demonstrate that exercise-mediated increases in Treg cell function may play a role in the attenuation of airway inflammation. Further, these results indicate that moderate intensity aerobic exercise training may alter the Treg cell function within the asthmatic airway.     

Acute aerobic exercise in humans increases cytokine expression in CD27− but not CD27+ CD8+ T-cells

Exercise alters the percentage of CD8⁺ T-cells in the bloodstream expressing type I and type II cytokines. It is unknown if this reflects a change in cytokine expression within individual cells, or whether these observations result from the exercise-induced shift in the proportions of early/intermediate (CD27⁺) and late (CD27⁻) differentiated cells, which have vastly different cytokine profiles. 16 males cycled for 60 min at 95% maximal steady state. Mononuclear cells isolated from blood collected before, immediately after, and 1 h after exercise were cultured overnight with and without phytohaemagglutinin stimulation. CD8⁺ T-cells were assessed for differentiation markers and intracellular cytokine expression by flow cytometry. The numbers and percentage of CD27⁻CD8⁺ T-cells increased immediately after exercise and fell below pre-exercise values 1 h later. At 1 h after exercise, an increased number and percentage of CD8⁺ T-cells expressing IL-2, IFN-γ, TNF-α, IL-6, IL-4, and IL-10 was observed in both stimulated and unstimulated cells. The cytokine response to exercise was confined to CD27⁻CD8⁺ T-cells, although cytokine expression among CD8⁺ T-cells was highest when the proportion of CD27⁻CD8⁺ T-cells was lowest. Moreover, the cytokine response to exercise could be predicted by the number of late cells in resting blood: cytokine expression was highest among those with low resting proportions of late cells. We conclude that exercise-induced changes in the percentage of CD8⁺ T-cells expressing cytokines are not due to proportional shifts in early/intermediate and late differentiated T-cells. Exercise may prime late-differentiated blood CD8⁺ T-cells to initiate effector functions in preparation for their extravasation into the tissues.     

AMPA receptors shape Ca2+ responses in cortical oligodendrocyte progenitors and CG-4 cells

Intracellular calcium signals triggered by glutamate receptor activation were studied in primary cortical oligodendrocyte lineage cells and in the oligodendrocyte cell line CG-4. Glutamate, kainate, and AMPA (30-300 μM) increased [Ca ²⁺]_i in both types of cells at the stage of oligodendrocyte progenitors (O-2A; GD3⁺) or pro-oligodendroblasts (04⁺). The peak amplitude of Ca²⁺ responses to glutamate receptor agonists was significantly larger in cortical cells. In CG-4 and in cortical cells, the majority (more than 90%) of bipolar GD3⁺ or multipolar 04⁺ cells responded to kamate. In all the cells analyzed, kainate was more efficacious than AMPA and glutamate. The percentage of bipolar or multipolar cells responding to glutamate was significantly lower in the CG-4 cell line than in primary cultures. Cellular responses typical of metabotropic glutamate receptor activation were observed in 20% of the cortical O-2A progenitors, but in none of the CG-4 cells. The AMPA-selective antagonist GYKI 52466 blocked kainate-induced Ca²⁺ responses in cortical O-2A cells. The selective AMPA receptor modulator cyclothiazide (30 μM) greatly potentiated the effects of AMPA (30-100 μM) on [Ca ²⁺]_i in cortical and CG-4 cells. Our findings indicate that Ca²⁺ responses in cells of the oligodendrocyte lineage are primarily shaped by functional AMPA receptors. © 1995 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

Expression of FcR2/CD23 and p55 IL-2R/CD25 on peripheral blood macrophages/monocytes in multiple sclerosis

Macrophages have been found histologically to be activated in multiple sclerosis. We analyzed the expression of CD23 and CD25 on monocytes/macrophages in peripheral blood obtained from patients with multiple sclerosis (MS) to investigate their role in the demyelinating process. Peripheral blood mononuclear cells were obtained from 30 patients with MS including for Baló's diseases (24 with acute relapsing type disease, six with chronic progressive type disease) and 12 healthy controls. The percentage of CD14⁺ CD23⁺ monocytes/macrophages and CD14⁺ CD25⁺ monocytes/macrophages were determined by two-color flow cytometry. The percentage of CD14⁺ CD23⁺ monocytes was significantly higher in patients with MS in the active phase as compared with controls (P < 0.01). Six patients with acute relapsing MS, who had received no therapy, had higher CD14⁺ CD23⁺ cells than did controls (P < 0.0001). CD14⁺ CD25⁺ monocytes/macrophages were not detected in peripheral blood monocytes/macrophages of patients with MS except Baló's concentric sclerosis. The four patients with Baló's concentric sclerosis had markedly elevated levels of CD14⁺ CD25⁺ monocytes/macrophages. Our findings suggest that monocytes/macrophages are activated during an exacerbatiion of MS. and that they may play an important role in the process of demyelination.     

Role of resveratrol-induced CD11b(+) Gr-1(+) myeloid derived suppressor cells (MDSCs) in the reduction of CXCR3(+) T cells and amelioration of chronic colitis in IL-10(-/-) mice

Resveratrol, a naturally occurring polyphenol has received significant attention as a potent anti-inflammatory agent. Inflammatory bowel disease (IBD) is a chronic intestinal inflammation caused by hyperactivated effector immune cells that produce proinflammatory cytokines. Myeloid derived suppressor cells (MDSCs) are a heterogeneous population characterized by the co-expression of CD11b⁺ and Gr-1⁺ and have long been known for their immunosuppressive function. We report that resveratrol effectively attenuated overall clinical scores as well as various pathological markers of colitis in IL-10^−/− mice by down regulating Th1 responses. Resveratrol lessened the colitis-associated decrease in body weight and increased levels of serum amyloid A (SAA), CXCL10 and colon TNF-α, IL-6, RANTES, IL-12 and IL-1β concentrations. After resveratrol treatment, the percentage of CXCR3 expressing T cells was decreased in the spleen, mesenteric lymph nodes (MLN), and intestinal lamina propria (LP). However, the percentage and absolute numbers of CD11b⁺ and Gr-1⁺cells in the lamina propria (LP) and spleen were increased after resveratrol treatment as compared with the vehicle treatment. Co-culture of resveratrol-induced CD11b⁺ Gr-1⁺ cells with T cells, attenuated T cell proliferation, and most importantly reduced IFN-γ and GM-CSF production by LP derived T cells from vehicle treated IL-10^−/− mice with chronic colitis. The current study suggests that administration of resveratrol into IL-10^−/− mice induces immunosuppressive CD11b⁺ Gr-1⁺ MDSCs in the colon, which correlates with reversal of established chronic colitis, and down regulation of mucosal and systemic CXCR3⁺ expressing effector T cells as well as inflammatory cytokines in the colon. The induction of immunosuppressive CD11b⁺ Gr-1⁺ cells by resveratrol during colitis is unique, and suggests an as-yet-unidentified mode of anti-inflammatory action of this plant polyphenol.     

Tim-3在肿瘤免疫中的研究进展

免疫检查点分子是现在肿瘤免疫治疗讨论的热点。本文通过对免疫检查点T细胞免疫球蛋白粘蛋白3(Tim-3)的相关研究,从以下三个方面讨论它抑制肿瘤免疫的作用：第一,与不同的配体Gal-9、HMGB1、CEACAM-1结合来发挥作用;第二,可以负性调控CD4⁺T/CD8⁺T/NK等重要的免疫细胞;第三,针对Tim-3的抗肿瘤免疫治疗。表明Tim-3具有通过与不同配体的特异性结合,抑制固有免疫和适应性免疫细胞的能力,使肿瘤产生免疫逃逸。旨在阐述Tim-3免疫抑制的特异性,为免疫靶向治疗提供新思路和依据。     

Methylmercury chloride exposure aggravates proinflammatory mediators and Notch-1 signaling in CD14+ and CD40+ cells and is associated with imbalance of neuroimmune function in BTBR T+ Itpr3tf/J mice

Autism spectrum disorder (ASD) is a severe neurodevelopmental disorder characterized by deficits in social interaction, communication, and repetitive behaviors. A key role for immune dysfunction has been suggested in ASD. Recent studies have indicated that inflammatory mediators and Notch-1 signaling may contribute to the development of ASD. Methylmercury chloride (MeHgCl) is an environmental pollutant that primarily affects the central nervous system, causing neurological alterations. Its effects on immunological responses have not been fully investigated in ASD. In this study, we examined the influence of MeHgCl exposure on inflammatory mediators and Notch-1 signaling in BTBR T⁺ Itpr3tf/J (BTBR) mice, a model of ASD. We examined the effects of MeHgCl on the IL-6-, GM-CSF-, NF-κB p65-, Notch-1-, and IL-27-producing CD14⁺ and CD40⁺ cells in the spleen. We assessed the effect of MeHgCl on IL-6, GM-CSF, NF-κB p65, Notch-1, and IL-27 mRNA levels in brain tissue. We also measured IL-6, GM-CSF, and NF-κB p65 protein expression levels in brain tissue. MeHgCl exposure of BTBR mice significantly increased IL-6-, GM-CSF-, NF-κB p65-, and Notch-1-, and decreased IL-27-producing CD14⁺, and CD40⁺ cells in the spleen. MeHgCl exposure of BTBR mice upregulated IL-6, GM-CSF, NF-κB p65, and Notch-1, and decreased IL-27 mRNA expression levels in brain tissue. Moreover, MeHgCl resulted in elevated expression of the IL-6, GM-CSF, and NF-κB p65 proteins in brain tissue. Taken together, these results indicate that MeHgCl exposure aggravates proinflammatory mediators and Notch-1 signaling which are associated with imbalance of neuroimmune function in BTBR mice.