scholarly journals Oxidative stress response in regulatory and conventional T cells: a comparison between patients with chronic coronary syndrome and healthy subjects

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Anna K. Lundberg ◽  
Rosanna W. S. Chung ◽  
Louise Zeijlon ◽  
Gustav Fernström ◽  
Lena Jonasson
Keyword(s):  
Oxidative Stress ◽  
T Cells ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Thioredoxin Reductase ◽  
Healthy Controls ◽  
Antioxidant Genes ◽  
Consistent Finding ◽  
Coronary Syndrome ◽  
Thioredoxin Reductase 1

Abstract Background Inflammation and oxidative stress form a vicious circle in atherosclerosis. Oxidative stress can have detrimental effects on T cells. A unique subset of CD4+ T cells, known as regulatory T (Treg) cells, has been associated with atheroprotective effects. Reduced numbers of Treg cells is a consistent finding in patients with chronic coronary syndrome (CCS). However, it is unclear to what extent these cells are sensitive to oxidative stress. In this pilot study, we tested the hypothesis that oxidative stress might be a potential contributor to the Treg cell deficit in CCS patients. Methods Thirty patients with CCS and 24 healthy controls were included. Treg (CD4+CD25+CD127−) and conventional T (CD4+CD25−, Tconv) cells were isolated and treated with increasing doses of H2O2. Intracellular ROS levels and cell death were measured after 2 and 18 h, respectively. The expression of antioxidant genes was measured in freshly isolated Treg and Tconv cells. Also, total antioxidant capacity (TAC) was measured in fresh peripheral blood mononuclear cells, and oxidized (ox) LDL/LDL ratios were determined in plasma. Results At all doses of H2O2, Treg cells accumulated more ROS and exhibited higher rates of death than their Tconv counterparts, p < 0.0001. Treg cells also expressed higher levels of antioxidant genes, including thioredoxin and thioredoxin reductase-1 (p < 0.0001), though without any differences between CCS patients and controls. Tconv cells from CCS patients were, on the other hand, more sensitive to oxidative stress ex vivo and expressed more thioredoxin reductase-1 than Tconv cells from controls, p < 0.05. Also, TAC levels were lower in patients, 0.97 vs 1.53 UAE/100 µg, p = 0.001, while oxLDL/LDL ratios were higher, 29 vs 22, p = 0.006. Conclusion Treg cells isolated from either CCS patients or healthy controls were all highly sensitive to oxidative stress ex vivo. There were signs of oxidant-antioxidant imbalance in CCS patients and we thus assume that oxidative stress may play a role in the reduction of Treg cells in vivo.

scholarly journals Oxidative Stress Response in Regulatory and Conventional T Cells: A Comparison Between Patients with Chronic Coronary Syndrome and Healthy Subjects

2020 ◽  
Author(s):  
Anna K Lundberg ◽  
Rosanna WS Chung ◽  
Louise Zeijlon ◽  
Gustav Fernström ◽  
Lena Jonasson
Keyword(s):  
Oxidative Stress ◽  
T Cells ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Thioredoxin Reductase ◽  
Healthy Controls ◽  
Antioxidant Genes ◽  
Consistent Finding ◽  
Coronary Syndrome ◽  
Thioredoxin Reductase 1

Abstract BackgroundInflammation and oxidative stress form a vicious circle in atherosclerosis. Oxidative stress can have detrimental effects on T cells. A unique subset of CD4+ T cells, known as regulatory T (Treg) cells, has been associated with atheroprotective effects. Reduced numbers of Treg cells is a consistent finding in patients with chronic coronary syndrome (CCS). However, it is unclear to what extent these cells are sensitive to oxidative stress. In the present study, we tested the hypothesis that oxidative stress might be a potential contributor to the Treg cell deficit in CCS patients. MethodsThirty patients with CCS and 24 healthy controls were included. Treg (CD4+CD25+CD127-) and conventional T (CD4+CD25-, Tconv) cells were isolated and treated with increasing doses of H2O2. Intracellular ROS levels and cell death were measured after 2 and 18 h, respectively. The expression of antioxidant genes was measured in freshly isolated Treg and Tconv cells. Alxo, total antioxidant capacity (TAC) was measured in fresh peripheral blood mononuclear cells. ResultsAt all doses of H2O2, Treg cells accumulated more ROS and exhibited higher rates of death than their Tconv counterparts, p < 0.0001. Treg cells also expressed higher levels of antioxidant genes, including thioredoxin and thioredoxin reductase-1 (p < 0.0001), though without any differences between CCS patients and controls. Tconv cells from CCS patients were, on the other hand, more sensitive to oxidative stress ex vivo and expressed more thioredoxin reductase-1 than Tconv cells from controls, p < 0.05. Also, TAC levels were lower in patients, 0.97 vs 1.53 UAE/100 µg, p = 0.001. ConclusionTreg cells isolated from either CCS patients or healthy controls were all highly sensitive to oxidative stress ex vivo. There were however signs of oxidant-antioxidant imbalance in CCS patients and we thus assume that oxidative stress plays a role in the reduction of Treg cells in vivo.

scholarly journals Targeting CD38 is lethal to Breg-like chronic lymphocytic leukemia cells and Tregs, but restores CD8+ T-cell responses

2020 ◽  
Vol 4 (10) ◽  
pp. 2143-2157 ◽  
Author(s):  
Alak Manna ◽  
Timothy Kellett ◽  
Sonikpreet Aulakh ◽  
Laura J. Lewis-Tuffin ◽  
Navnita Dutta ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Chronic Lymphocytic Leukemia ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Transforming Growth Factor ◽  
Ex Vivo ◽  
Xenograft Model ◽  
Lymphocytic Leukemia ◽  
Dependent Manner

Abstract Patients with chronic lymphocytic leukemia (CLL) are characterized by monoclonal expansion of CD5+CD23+CD27+CD19+κ/λ+ B lymphocytes and are clinically noted to have profound immune suppression. In these patients, it has been recently shown that a subset of B cells possesses regulatory functions and secretes high levels of interleukin 10 (IL-10). Our investigation identified that CLL cells with a CD19+CD24+CD38hi immunophenotype (B regulatory cell [Breg]–like CLL cells) produce high amounts of IL-10 and transforming growth factor β (TGF-β) and are capable of transforming naive T helper cells into CD4+CD25+FoxP3+ T regulatory cells (Tregs) in an IL-10/TGF-β-dependent manner. A strong correlation between the percentage of CD38+ CLL cells and Tregs was observed. CD38hi Tregs comprised more than 50% of Tregs in peripheral blood mononuclear cells (PBMCs) in patients with CLL. Anti-CD38 targeting agents resulted in lethality of both Breg-like CLL and Treg cells via apoptosis. Ex vivo, use of anti-CD38 monoclonal antibody (mAb) therapy was associated with a reduction in IL-10 and CLL patient-derived Tregs, but an increase in interferon-γ and proliferation of cytotoxic CD8+ T cells with an activated phenotype, which showed an improved ability to lyse patient-autologous CLL cells. Finally, effects of anti-CD38 mAb therapy were validated in a CLL–patient-derived xenograft model in vivo, which showed decreased percentage of Bregs, Tregs, and PD1+CD38hiCD8+ T cells, but increased Th17 and CD8+ T cells (vs vehicle). Altogether, our results demonstrate that targeting CD38 in CLL can modulate the tumor microenvironment; skewing T-cell populations from an immunosuppressive to immune-reactive milieu, thus promoting immune reconstitution for enhanced anti-CLL response.

mRNA expression analysis confirms CD44 splicing impairment in systemic lupus erythematosus patients

Lupus ◽  
2021 ◽  
pp. 096120332110047
Author(s):  
Andrea Latini ◽  
Lucia Novelli ◽  
Fulvia Ceccarelli ◽  
Cristiana Barbati ◽  
Carlo Perricone ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
T Cells ◽  
Mrna Expression ◽  
Lupus Erythematosus ◽  
Mononuclear Cells ◽  
Direct Sequencing ◽  
Healthy Controls ◽  
Systemic Lupus ◽  
Peripheral Tissues ◽  
Variant Isoforms

Background Systemic Lupus Erythematosus (SLE) is a complex chronic autoimmune disease characterized by several immunological alterations. T cells have a peculiar role in SLE pathogenesis, moving from the bloodstream to the peripheral tissues, causing organ damage. This process is possible for their increased adherence and migration capacity mediated by adhesion molecules, such as CD44. Ten different variant isoforms of this molecule have been described, and two of them, CD44v3 and CD44v6 have been found to be increased on SLE T cells compared to healthy controls, being proposed as biomarkers of disease and disease activity. The process of alternative splicing of CD44 transcripts is not fully understood. We investigated the mRNA expression of CD44v3 and CD44v6 and also analyzed possible CD44 splicing regulators (ESRP1 molecule and rs9666607 CD44 polymorphism) in a cohort of SLE patients compared to healthy controls. Methods This study involved 18 SLE patients and 18 healthy controls. Total RNA and DNA were extracted by peripheral blood mononuclear cells. The expression study was conducted by quantitative RT-polymerase chain reaction, using SYBR Green protocol. Genotyping of rs9666607 SNP was performed by direct sequencing. Results CD44v6 mRNA expression was higher in SLE patients compared to healthy controls (p = 0.028). CD44v3/v6 mRNA ratio in healthy controls was strongly unbalanced towards isoform v3 compared to SLE patients (p = 0.002) and decreased progressively from healthy controls to the SLE patients in remission and those with active disease (p = 0.015). The expression levels of CD44v3 and CD44v6 mRNA correlated with the disease duration (p = 0.038, Pearson r = 0.493 and p = 0.038, Pearson r = 0.495, respectively). Splicing regulator ESRP1 expression positively correlated with CD44v6 expression in healthy controls (p = 0.02, Pearson r = 0.532) but not in SLE patients. The variant A allele of rs9666607 of CD44 was associated with higher level of global CD44 mRNA (p = 0.04) but not with the variant isoforms. Conclusions In SLE patients, the increase in CD44v6 protein correlates with a higher transcript level of this isoform, confirming an impairment of CD44 splicing in the disease, whose regulatory mechanisms require further investigation.

scholarly journals The Anthrax Vaccine Adsorbed Vaccine Generates Protective Antigen (PA)-Specific CD4+ T Cells with a Phenotype Distinct from That of Naïve PA T Cells

2008 ◽  
Vol 76 (10) ◽  
pp. 4538-4545 ◽  
Author(s):  
William W. Kwok ◽  
Junbao Yang ◽  
Eddie James ◽  
John Bui ◽  
Laurie Huston ◽  
...  
Keyword(s):  
T Cells ◽  
Mononuclear Cells ◽  
Protective Antigen ◽  
Ex Vivo ◽  
Cytokine Profile ◽  
T Cell Epitopes ◽  
Peripheral Blood Mononuclear ◽  
Anthrax Vaccine ◽  
Cellular Immune

ABSTRACT Cellular immune responses against protective antigen (PA) of Bacillus anthracis in subjects that received the anthrax vaccine adsorbed (AVA) vaccine were examined. Multiple CD4+ T-cell epitopes within PA were identified by using tetramer-guided epitope mapping. PA-reactive CD4+ T cells with a CD45RA− phenotype were also detected by direct ex vivo staining of peripheral blood mononuclear cells (PBMC) with PA-specific tetramers. Surprisingly, PA-specific T cells were also detected in PBMC of nonvaccinees after a single cycle of in vitro PA stimulation. However, PA-reactive CD4+ T cells in nonvaccinees occurred at lower frequencies than those in vaccinees. The majority of PA-reactive T cells from nonvaccinees were CD45RA+ and exhibited a Th0/Th1 cytokine profile. In contrast, phenotyping and cytokine profile analyses of PA-reactive CD4+ T cells from vaccinees indicated that vaccination leads to commitment of PA-reactive T cells to a Th2 lineage, including generation of PA-specific, pre-Th2 central memory T cells. These results demonstrate that the current AVA vaccine is effective in skewing the development of PA CD4+ T cells to the Th2 lineage. The data also demonstrated the feasibility of using class II tetramers to analyze CD4+ cell responses and lineage development after vaccination.

scholarly journals Cytokine production in ex-vivo stimulated fresh and cryopreserved T-cells

2021 ◽  
Vol 67 (2) ◽  
pp. 95-101
Author(s):  
Monica Vuță ◽  
Ionela-Maria Cotoi ◽  
Ion Bogdan Mănescu ◽  
Doina Ramona Manu ◽  
Minodora Dobreanu
Keyword(s):  
T Cells ◽  
Peripheral Blood ◽  
Cd4 T Cells ◽  
Mononuclear Cells ◽  
Cytokine Production ◽  
Ex Vivo ◽  
Gradient Centrifugation ◽  
Intracellular Cytokine Staining ◽  
Interleukin 2 ◽  
Middle Aged

Abstract Objective: In vitro cytokine production by peripheral blood mononuclear cells (PBMCs) is an important and reliable measure of immunocompetence. PBMC can be stimulated directly after isolation or frozen for later use. However, cryopreservation may affect cell recovery, viability and functionality. This study aims to investigate cytokine synthesis in ex-vivo stimulated fresh and cryopreserved CD4+ and CD4- T cells. Methods: PBMCs were obtained by Ficoll gradient centrifugation from heparinized peripheral blood of 6 middle-aged clinically healthy subjects. Half of these cells (labeled “Fresh”) was further processed and the other half (labeled “Cryo”) was cryopreserved at -140°C for up to 3 months. Fresh-PBMCs were activated with Phorbol-Myristate-Acetate/Ionomycin/Monensin for 5 hours immediately after isolation while Cryo-PBMCs were identically activated after thawing and cell resting. Activated cells were fixed, permeabilized and intracellular cytokine staining was performed using Phycoerythrin (PE)-conjugated antibodies for Interleukin-2 (IL-2), Tumor Necrosis Factor-alpha (TNF-a), and Interferon-gamma (IFN-g). All samples were analyzed within 24 hours by flow cytometry. Results: Both Fresh and Cryo CD3+CD4+/CD3+CD4- sub-populations partially produced each of the three cytokines. A higher percentage of CD4+ T cells produced IL-2 and TNF-a and a greater percentage of CD4- T cells were found to produce IFN-g. A significantly higher percentage of Cryo-lymphocytes was shown to produce TNF-a in both CD3+CD4+ (31.4% vs 24.9%, p=0.031) and CD3+CD4- (22.7% vs 17.9%, p=0.031) subpopulations. No notable difference was found for IL-2 and IFN-g production between Fresh and Cryo T cells. Conclusion: Cryopreservation for up to 3 months significantly increases TNF-a production of T-cells in clinically healthy middle-aged subjects.

scholarly journals Acetylated Thioredoxin Reductase 1 Resists Oxidative Inactivation

2021 ◽  
Vol 9 ◽  
Author(s):  
David. E. Wright ◽  
Nikolaus Panaseiko ◽  
Patrick O’Donoghue
Keyword(s):  
Oxidative Stress ◽  
Thioredoxin Reductase ◽  
Oxygen Species ◽  
Site Specific ◽  
Oxidative Inactivation ◽  
Thioredoxin Reductase 1 ◽  
Lysine Residues ◽  
Canonical Amino Acid ◽  
Genetic Code Expansion ◽  
Low Activity

Thioredoxin Reductase 1 (TrxR1) is an enzyme that protects human cells against reactive oxygen species generated during oxidative stress or in response to chemotherapies. Acetylation of TrxR1 is associated with oxidative stress, but the function of TrxR1 acetylation in oxidizing conditions is unknown. Using genetic code expansion, we produced recombinant and site-specifically acetylated variants of TrxR1 that also contain the non-canonical amino acid, selenocysteine, which is essential for TrxR1 activity. We previously showed site-specific acetylation at three different lysine residues increases TrxR1 activity by reducing the levels of linked dimers and low activity TrxR1 tetramers. Here we use enzymological studies to show that acetylated TrxR1 is resistant to both oxidative inactivation and peroxide-induced multimer formation. To compare the effect of programmed acetylation at specific lysine residues to non-specific acetylation, we produced acetylated TrxR1 using aspirin as a model non-enzymatic acetyl donor. Mass spectrometry confirmed aspirin-induced acetylation at multiple lysine residues in TrxR1. In contrast to unmodified TrxR1, the non-specifically acetylated enzyme showed no loss of activity under increasing and strongly oxidating conditions. Our data suggest that both site-specific and general acetylation of TrxR1 regulate the enzyme’s ability to resist oxidative damage.

scholarly journals Interplay between cytosolic disulfide reductase systems and the Nrf2/Keap1 pathway

2015 ◽  
Vol 43 (4) ◽  
pp. 632-638 ◽  
Author(s):  
Edward E. Schmidt
Keyword(s):  
Oxidative Stress ◽  
Thioredoxin Reductase ◽  
Reducing Power ◽  
Nuclear Factor ◽  
Independent Source ◽  
Nrf2 Pathway ◽  
Thioredoxin Reductase 1 ◽  
Disulfide Reductase ◽  
The Status ◽  
Thioredoxin 1

NADPH transfers reducing power from bioenergetic pathways to thioredoxin reductase-1 (TrxR1) and glutathione reductase (GR) to support essential reductive systems. Surprisingly, it was recently shown that mouse livers lacking both TrxR1 and GR (‘TR/GR-null’) can sustain redox (reduction-oxidation) homoeostasis using a previously unrecognized NADPH-independent source of reducing power fuelled by dietary methionine. The NADPH-dependent systems are robustly redundant in liver, such that disruption of either TrxR1 or GR alone does not cause oxidative stress. However, disruption of TrxR1 induces transcription factor Nrf2 (nuclear factor erythroid-derived 2-like-2) whereas disruption of GR does not. This suggests the Nrf2 pathway responds directly to the status of the thioredoxin-1 (Trx1) system. The proximal regulator of Nrf2 is Keap1 (Kelch-like ECH-associated protein-1), a cysteine (Cys)-rich protein that normally interacts transiently with Nrf2, targeting it for degradation. During oxidative stress, this interaction is stabilized, preventing degradation of newly synthesized Nrf2, thereby allowing Nrf2 accumulation. Within the Trx1 system, TrxR1 and peroxiredoxins (Prxs) contain some of the most reactive nucleophilic residues in the cell, making them likely targets for oxidants or electrophiles. We propose that Keap1 activity and therefore Nrf2 is regulated by interactions of Trx1 system enzymes with oxidants. In TR/GR-null livers, Nrf2 activity is further induced, revealing that TrxR-independent systems also repress Nrf2 and these might be induced by more extreme challenges.

scholarly journals Different Profile of Interleukin-10 Production in Circulating T Cells from Atopic Asthmatics Compared with Healthy Subjects

2004 ◽  
Vol 11 (1) ◽  
pp. 33-38 ◽  
Author(s):  
K Matsumoto ◽  
S Narita ◽  
T Rerecich ◽  
DP Snider ◽  
PM O'Byrne
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Healthy Subjects ◽  
Mononuclear Cells ◽  
Interleukin 10 ◽  
Healthy Controls ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Circulating T Cells ◽  
Normal Controls

BACKGROUND:Interleukin (IL)-10 is a pleiotropic cytokine released from various cells, including T cells. Although IL-10 is suggested to inhibit allergic responses, its role in asthma remains uncertain. The purpose of the present study was to compare the profile of IL-10 in circulating T cells from stable atopic asthmatics, atopic nonasthmatics and healthy controls.METHODS:Peripheral blood mononuclear cells were isolated, stained with anti-CD3 and CD4/CD8 antibodies, and then processed for intracellular IL-10 detection by flow cytometry.RESULTS:A kinetic study in healthy controls showed that stimulation with phorbol 12-myristate 13-acetate and ionomycin significantly increased the frequencies of IL-10-producing CD3+, CD4+and CD8+cells. Without stimulation, the frequencies of IL-10-producing CD3+, CD4+and CD8+cells were significantly higher in asthmatics than in healthy controls, while a similar trend was observed in atopic nonasthmatics. Stimulation for 24 h significantly increased IL-10-producing CD3+, CD4+and CD8+cells in healthy controls and atopic nonasthmatics, but not in asthmatics.CONCLUSIONS:The frequency of IL-10-producing T cells is increased in the circulation of stable atopic asthmatics compared with normal controls. The lack of enhancement in their frequency by phorbol 12-myristate 13-acetate and ionomycin in asthmatics suggests that the circulating T cells of asthmatic subjects are maximally stimulated with regards to IL-10 production; alternatively, IL-10 production by T cells from asthmatics may be regulated differently than T cells from other subjects.

scholarly journals Ex Vivo Expanded Human Vγ9Vδ2 T-Cells Can Suppress Epithelial Ovarian Cancer Cell Growth

2019 ◽  
Vol 20 (5) ◽  
pp. 1139 ◽  
Author(s):  
Tsui Mao ◽  
Carol Miao ◽  
Yi Liao ◽  
Ying Chen ◽  
Chia Yeh ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Γδ T Cells ◽  
Ovarian Cancer Cells ◽  
Cytotoxic Activities ◽  
Antitumor Effects ◽  
Peripheral Blood Mononuclear

γδ-T-cells have attracted attention because of their potent cytotoxicity towards tumors. Most γδ-T-cells become activated via a major histocompatibility complex (MHC)-independent pathway by the interaction of their receptor, Natural Killer Group 2 Member D (NKG2D) with the tumor-specific NKG2D ligands, including MHC class I-related chain A/B (MICA/B) and UL16-binding proteins (ULBPs), to kill tumor cells. However, despite their potent antitumor effects, the treatment protocols specifically targeting ovarian tumors require further improvements. Ovarian cancer is one of the most lethal and challenging female malignancies worldwide because of delayed diagnoses and resistance to traditional chemotherapy. In this study, we successfully enriched and expanded γδ-T-cells up to ~78% from peripheral blood mononuclear cells (PBMCs) with mostly the Vγ9Vδ2-T-cell subtype in the circulation. We showed that expanded γδ-T-cells alone exerted significant cytotoxic activities towards specific epithelial-type OVCAR3 and HTB75 cells, whereas the combination of γδ-T cells and pamidronate (PAM), a kind of aminobisphosphonates (NBPs), showed significantly enhanced cytotoxic activities towards all types of ovarian cancer cells in vitro. Furthermore, in tumor xenografts of immunodeficient NSG mice, γδ-T-cells not only suppressed tumor growth but also completely eradicated preexisting tumors with an initial size of ~5 mm. Thus, we concluded that γδ-T-cells alone possess dramatic cytotoxic activities towards epithelial ovarian cancers both in vitro and in vivo. These results strongly support the potential of clinical immunotherapeutic application of γδ-T-cells to treat this serious female malignancy.

Sign in / Sign up

Export Citation Format

Share Document