Abstract 001: Axl Expression by CD4+ T Lymphocytes Promotes Salt-Dependent Hypertension

Hypertension ◽  
2014 ◽  
Vol 64 (suppl_1) ◽  
Author(s):  
Kristine M Wadosky ◽  
Sri N Batchu ◽  
Angie Hughson ◽  
Kathy Donlon ◽  
Craig N Morrell ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cd4 T Cells ◽  
White Blood Cells ◽  
Interleukin 4 ◽  
Th2 Cells ◽  
Salt Hypertension ◽  
Ifn Γ

Introduction: Our laboratory has shown that Axl, a receptor tyrosine kinase, is important in both vascular and immune functions during deoxycorticosterone acetate (DOCA)-salt hypertension. We hypothesized that Axl activity specifically in T lymphocytes could explain the dependence of hypertension on Axl. Methods and Results: We did adoptive transfers of either Axl+/+ or Axl-/- CD4+ T cells to RAG1-/- mice that lack mature T cells. Once CD4+ T cell repopulations were confirmed, we induced DOCA-salt hypertension for 6 weeks. Systolic blood pressure (BP, mmHg) increased by 20±5 in Axl+/+RAG-/- mice after DOCA-salt, but Axl-/- RAG-/- mice had increases in BP by only 6+3 after 6 weeks of DOCA-salt. We isolated naïve CD4+ T cells from both Axl+/+ and Axl-/- littermates and primed them under either Th1 or Th2 polarizing conditions in culture. Production of interferon gamma (IFN-γ ng/mL) was significantly decreased (-23%, p<0.05) in Axl-/- (396±23) compared to Axl+/+ (512±42) under Th1-priming. However, Axl had no effect on interleukin 4 (IL-4, ng/mL) production under Th2 polarizing conditions. Intracellular staining of the Th1/Th2 cells with IFN-γ and IL-4 antibodies by flow cytometry confirmed expression of cytokines in culture media. Complete blood counts showed that Axl-/- mice had significantly lower white blood cells due to decreased numbers of lymphocytes (4.5±0.7x10 9 ) compared to Axl+/+ mice (7.8±0.7x10 9 ). We found a higher population of AnnexinV (marker of early apoptosis)-positive peripheral leukocytes in Axl-/- mice (10±1%) compared to Axl+/+ (4±1%) by flow cytometry; while the percentages of dead cells (~10%) were similar between Axl+/+ and Axl-/- mice. Conclusions: Altogether we show that expression of Axl by T cells drives salt-induced hypertension. The mechanism of Axl-dependent effects on T cells occurs via T-cell-dependent expression of the pro-inflammatory cytokine IFN-γ. In addition, Axl plays a role in inhibiting lymphocyte apoptosis in the circulation. Future work will focus on how Axl expression in T cells affects T cell-dependent vascular remodeling during hypertension.

scholarly journals CD4 T Cell Cytokine Differentiation: The B Cell Activation Molecule, OX40 Ligand, Instructs CD4 T Cells to Express Interleukin 4 and Upregulates Expression of the Chemokine Receptor, Blr-1

1998 ◽  
Vol 188 (2) ◽  
pp. 297-304 ◽  
Author(s):  
Sarah Flynn ◽  
Kai-Michael Toellner ◽  
Chandra Raykundalia ◽  
Margaret Goodall ◽  
Peter Lane
Keyword(s):  
T Cells ◽  
T Cell ◽  
Chemokine Receptor ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Interleukin 4 ◽  
Th2 Cells ◽  
Ox40 Ligand ◽  
Ifn Γ

This report investigates the role of OX40 ligand (OX40L) and its receptor, OX40, expressed on activated B and T cells, respectively, in promoting the differentiation of T helper type 2 (Th2) CD4 T cells. These molecules are expressed in vivo by day 2 after priming with T cell– dependent antigens. Their expression coincides with the appearance of immunoglobulin (Ig)G switch transcripts and mRNA for interleukin (IL)-4 and interferon (IFN)-γ, suggesting that this molecular interaction plays a role in early cognate interactions between B and T cells. In vitro, we report that costimulation of naive, CD62Lhigh CD4 T cells through OX40 promotes IL-4 expression and upregulates mRNA for the chemokine receptor, blr-1, whose ligand is expressed in B follicles and attracts lymphocytes to this location. Furthermore, T cell stimulation through OX40 inhibits IFN-γ expression in both CD8 T cells and IL-12–stimulated CD4 T cells. Although this signal initiates IL-4 expression, IL-4 itself is strongly synergistic. Our data suggest that OX40L on antigen-activated B cells instructs naive T cells to differentiate into Th2 cells and migrate into B follicles, where T cell–dependent germinal centers develop.

scholarly journals Enteral Immunization with Attenuated Recombinant Listeria monocytogenes as a Live Vaccine Vector: Organ-Dependent Dynamics of CD4 T Lymphocytes Reactive to a Leishmania major Tracer Epitope

2003 ◽  
Vol 71 (3) ◽  
pp. 1083-1090 ◽  
Author(s):  
Hélène Saklani-Jusforgues ◽  
Elisabeth Fontan ◽  
Neirouz Soussi ◽  
Geneviève Milon ◽  
Pierre L. Goossens
Keyword(s):  
T Cells ◽  
Listeria Monocytogenes ◽  
T Cell ◽  
Cd4 T Cells ◽  
Leishmania Major ◽  
Cd4 T Cell ◽  
Interleukin 4 ◽  
Cell Immune Response ◽  
Mesenteric Lymph Nodes ◽  
Ifn Γ

ABSTRACT Listeria monocytogenes is considered as a potential live bacterial vector, particularly for the induction of CD8 T cells. The CD4 T-cell immune response triggered after enteral immunization of mice has not yet been thoroughly characterized. The dynamics of gamma interferon (IFN-γ)- and interleukin-4 (IL-4)-secreting CD4 T cells were analyzed after priming through intragastric delivery of an attenuated ΔactA recombinant L. monocytogenes strain expressing the Leishmania major LACK protein; a peptide of this protein, LACK158-173 peptide (pLACK), is a well-characterized CD4 T-cell target in BALB/c mice. Five compartments were monitored: Peyer's patches, mesenteric lymph nodes (MLN), spleen, liver, and blood. A single intragastric inoculation of ΔactA-LACK-LM in BALB/c mice led to colonization of the MLN and spleen at a significant level for at least 3 days. Efficient priming of IFN-γ-secreting pLACK-reactive CD4 T cells was observed in all tested compartments. Interestingly, IL-4-secreting pLACK-reactive CD4 T cells were detectable at day 6 or 7 only in blood and liver. The absence of translocation of viable bacteria through the intestinal epithelium after further ΔactA-LACK-LM inoculations was concomitant with the absence of an increase in the level of IFN-γ secreted by the MLN, blood, and splenic pLACK-reactive Th1 T cells, although the levels remained significantly above the basal level. No change in this population size was detected in the spleen. However, an increase in the number of intragastric inoculations had a clinical beneficial effect in L. major-infected BALB/c mice. L. monocytogenes thus presents the potential of an efficient vector for induction of CD4 T cells when administered by the enteral route.

scholarly journals Interleukin 12 and Interleukin 4 Control T Cell Adhesion to Endothelial Selectins through Opposite Effects on α1,3-fucosyltransferase VII Gene Expression

1998 ◽  
Vol 188 (12) ◽  
pp. 2225-2231 ◽  
Author(s):  
Amy J. Wagers ◽  
Christopher M. Waters ◽  
Lloyd M. Stoolman ◽  
Geoffrey S. Kansas
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Single Gene ◽  
Interleukin 4 ◽  
Th2 Cells ◽  
Interleukin 12 ◽  
Th 1

The α1,3-fucosyltransferase, FucT-VII, is crucial for the formation of ligands for all three selectins, and its expression regulates the synthesis of these ligands. Short-term polarized T helper (Th)1, but not Th2 or naive CD4+ T cells, can home to sites of inflammation, but the molecular basis for this difference has remained unclear. Here we show that naive CD4+ T cells do not express FucT-VII and fail to bind vascular selectins. We also show that when CD4+ T cells are activated in the presence of the Th1 polarizing cytokine interleukin (IL)-12, levels of FucT-VII mRNA and binding to E- and P-selectin are significantly augmented. In contrast, activation of CD4+ T cells in the presence of IL-4, a Th2 polarizing cytokine, inhibited FucT-VII expression and binding to vascular selectins. T cell activation upregulated expression of the Core2 transferase, C2GnT, equivalently regardless of the presence or absence of polarizing cytokines. These data indicate that the selective ability of Th1 cells, as opposed to Th2 cells or naive CD4+ T cells, to recognize vascular selectins and home to sites of inflammation is controlled principally by the expression of a single gene, FucT-VII.

scholarly journals Modulation of Murine Lyme Borreliosis by Interruption of the B7/CD28 T-Cell Costimulatory Pathway

1998 ◽  
Vol 66 (1) ◽  
pp. 266-271 ◽  
Author(s):  
Marie-Claude Shanafelt ◽  
Insoo Kang ◽  
Stephen W. Barthold ◽  
Linda K. Bockenstedt
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lyme Borreliosis ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interleukin 4 ◽  
Th2 Cells ◽  
Th Cell ◽  
Ifn Γ ◽  
Costimulatory Pathway

ABSTRACT Recent studies have implicated cytokines associated with Th2 cells in the genetic resistance to murine Lyme borreliosis. Because the B7/CD28 costimulatory pathway has been shown to influence the differentiation of Th-cell subsets, we investigated the contribution of the B7 molecules CD80 and CD86 to the Th2 cytokine profile and development of arthritis in BALB/c mice infected with Borrelia burgdorferi. Effective blockade of CD86/CD28 interaction was demonstrated by elimination of interleukin 4 (IL-4) and upregulation of gamma interferon (IFN-γ) responses by B. burgdorferi-specific T cells and by reduction of B. burgdorferi-specific immunoglobulin G. Despite the shift toward a Th1 cytokine pattern, which others have associated with disease susceptibility, the severity of arthritis was unchanged. Moreover, combined CD80/CD86 blockade by using anti-CD80 and anti-CD86 monoclonal antibodies or CTLA-4Ig enhanced IFN-γ production over that seen with CD86 blockade alone, yet augmentation of this Th1-associated cytokine did not enhance disease. These results demonstrate that IL-4 production by T cells in B. burgdorferi-infected BALB/c mice is dependent upon CD86/CD28 interaction and that this cytokine does not contribute significantly to host resistance to the development of arthritis. In addition, combined CD80/CD86 blockade resulted in preferential expansion of IFN-γ-producing T cells in B. burgdorferi infection, suggesting that costimulatory pathways other than B7/CD28 may contribute to T-cell activation during continuous antigen stimulation. These studies may provide insight into the role of the B7/CD28 pathway in other infectious and autoimmune diseases in which deviation of Th cell immune responses occurs and antigen is persistently present.

PD-1/PD-L Pathway Potentially Involved in ITP Immunopathogenesis

2019 ◽  
Vol 119 (05) ◽  
pp. 758-765 ◽  
Author(s):  
Mu Nie ◽  
Yang Liu ◽  
Xiu-xiu Li ◽  
Ya-nan Min ◽  
Dan-dan Yang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cell Apoptosis ◽  
Cd4 T Cells ◽  
Peripheral Tolerance ◽  
Healthy Controls ◽  
T Cell Apoptosis ◽  
Ifn Γ

AbstractThe binding of programmed death 1 (PD-1) to its ligands PD-L1 and PD-L2 on antigen-presenting cells turns off autoreactive T cells and induces peripheral tolerance. Aberrant PD-1/PD-L signalling could result in a breakdown of peripheral tolerance and lead to autoimmune diseases. In this study, we detected PD-1 and PD-L expression on T cells and dendritic cells (DCs) in immune thrombocytopenia (ITP) patients with active disease by flow cytometry. The effects of PD-L1-Fc fusion protein (PD-L1-Fc) on T cells and on secretion of interferon-γ (IFN-γ) and interleukin-2 (IL-2) were detected by flow cytometry and enzyme-linked immunosorbent assay, respectively. Compared with healthy controls, PD-1 expression was significantly increased in CD4+ T cells and CD8+ T cells from patients with active ITP. However, PD-L1 expression on monocyte-derived DCs was lower in patients with active ITP than in healthy controls. In vitro assays revealed that PD-L1-Fc increased T cell apoptosis, inhibited activation and proliferation of CD4+ T cells and CD8+ T cells and decreased IFN-γ and IL-2 secretion in patients with active ITP. These results suggest that the aberrant PD-1/PD-L negative co-stimulatory pathway may play a role in ITP. Enhancing PD-1/PD-L signalling might be a promising therapeutic approach for ITP patients by enhancing T cell apoptosis, inhibiting T cell activation and proliferation and reducing secretion of inflammatory factors.

Detecting 24 Kinds of Cytokines Via Flow Cytometry Aimplex Kit Is an Effective Way to Monitor CRS after CAR-T Cells Infusion

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 50-51
Author(s):  
Dongchu Wang ◽  
Xiang Li ◽  
Liyuan Qiu ◽  
Shizhe Fu ◽  
Lingyun Cui ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Early Prevention ◽  
Tnf Α ◽  
Car T ◽  
Variation Curve ◽  
Ifn Γ ◽  
The Relationship

Introduction: Cytokine release syndrome (CRS) is a serious side effect of chimeric antigen receptor-T cell (CAR-T) therapy. Effective cytokine monitoring can provide support for early prevention and treatment. As more and more promising markers have been identified to be associated with CRS responses, traditional ELISA method is inapplicable because it's sample-consuming, low flexibility, and difficult to detect multiple cytokines simultaneously. Aimplex kit, a new high-throughput technique based on flow cytometry(FCM) and micro beads has been testified a effective way to monitor multiple cytokines in peripheral blood(PB) samples. If a new suitable cytokines panel is established to monitor the variation curve of multiple cytokines in patients' PBs post CAR-T therapy, study the relationship between clinical symptoms and the lab results, even build a cytokines database of CAR-T therapy, it would offer a promising support to prevent and handle CRS. On the other hand, by studying the relationship between cytokines and activated cells, we might investigate mechanisms of CRS and explore more related markers. Methods: A two tubes panel was designed to detect 24 cytokines, including IFN-γ/IL-1β/IL-2/IL-4/IL-5/IL-6/IL-8/IL-10/IL-12p70/IL-17A/IL-17F/IL-22/TNF-α/TNF-β, and sCD25/GM-CSF/IL-15/MCP-1/GranzymeB/Reg3A/ST2/TNFRSF1A/Elafin/MIP-1 alpha. 50 PB samples from complete response(CR) patients without CAR-T therapy were detested as normal controls to establish the normal values of 24 cytokines. 81 patients who infused CAR-T cell in Hebei Yanda Ludaopei Hospital from January to June 2020 were selected. Serum were collected on 0d, 4d, 7d, 11d, 15d, 20d and 30d after infusion, and 24 cytokines were detected by FCM Aimplex. Of which 31 patients were selected for methodological comparison between FCM and ELISA by detecting four routine cytokines, IL-6, IFN-γ, TNF-α and sCD25. 66 Patients were divided into 2 groups according to clinical manifestations, 60 patients without or mild CRS were classified as low grade CRS group, 6 patients with grade 2 or 3 CRS were classified as high grade CRS group. Results: The comparative test showed that a good relationship between the results of ELISA and Aimplex kit, showing similar time-cytokines variation curve of PBs after CAR-T treatment. The concentrations of IL-2, IL-10, IL-12p70, IL-17A, IL-17F, MCP-1, ST-2s, IL-5, IL-6 and IFN-γ were significantly higher in high CRS grade group. By comparing the proportion of CAR-T+, CD8+ and CD4+ T cells in CD3+ cells and the concentration of cytokines in PB specimens, there were obvious positive correlations between percentages of CAR-T+, CD8+ T cells and most cytokines, and negative correlation between proportion of CD4+ T cells and most cytokines. Conclusions: Detecting 24 kinds of cytokines by Aimplex kit is a promising method, which is time and specimens saving, and can cost-effectively reflect the cytokine situation in PBs from patients post-CAR-T treatment. Increasing the sample size and establishing a cytokines database will be helpful for monitoring, early prevention and control of side effects after CAR-T treatment. Disclosures No relevant conflicts of interest to declare.

T Helper Type 1-Bias in a Regulatory T Cell-Deficient Mouse Model for Immune Thrombocytopenia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 524-524
Author(s):  
Tetsuya Nishimoto ◽  
Fumiaki Kumagai ◽  
Masayoshi Monno ◽  
Tsutomu Takeuchi ◽  
Masataka Kuwana
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Autoantibody Response ◽  
Ifn Γ ◽  
Culture Supernatants ◽  
Deficient Mice ◽  
Platelet Autoantibodies

Abstract Abstract 524 Background: Immune thrombocytopenia (ITP) is a T cell-mediated autoimmune disorder, in which IgG autoantibodies to platelet surface glycoproteins promote platelet clearance in the reticuloendthelial system. CD4+CD25+Foxp3+ regulatory T cells (Tregs) are known to play a crucial role in the maintenance of immune homeostasis to self-antigens. Several lines of recent evidence have shown that Tregs are decreased in number and are functionally impaired in patients with ITP. Recently, we have found that approximately one third of Treg-deficient mice spontaneously develop thrombocytopenia with increased platelet-associated IgG and proportion of reticulated platelets. Platelets eluates and culture supernatants of splenocytes prepared from thrombocytopenic mice contain IgG antibodies capable of binding to intact platelets, which are not detected in non-thrombocytopenic mice. The main target of anti-platelet autoantibodies is GPIb, and some mice also produce anti-GPIIIa antibodies. However, detailed mechanisms that elicit ITP during immune reconstitution through homeostatic proliferation in the absence of Tregs remain uncertain. Purpose: To evaluate T-helper (Th) cell balance that promotes anti-platelet autoantibody response in a Treg-deficient mouse model for ITP. Methods: Treg-deficient mice were prepared by inoculation of Treg-depleted CD4+ T cells obtained from BALB/c mice into syngeneic T cell-deficient nude mice. Platelet count was determined using flow cytometry 4 weeks after inoculation, and Treg-deficient mice with platelet count < 0.33 × 106/ul were regarded as ITP mice. Treg-deficient mice without thrombocytopenia were also used as a control. To evaluate cytokine profiles of Th cells, proportions of Th subsets in the freshly prepared splenic CD4+ T cells were evaluated by intracellular staining for IFN-γ, IL-4, and IL-17 followed by flow cytometry. Th1, Th2, and Th0 cells were defined as IFN-γ+IL-4−, IFN-γ−IL-4+, IFN-γ+IL-4+ cells, respectively, and Th17 and Th1/17 cells were defined as IFN-γ−IL-17+ and IFN-γ+IL-17+, respectively. In addition, CD4+ T cells were isolated from splenocytes using magnetic activated cell sorting, and were stimulated with phorbol 1,2-myristate 1,3-acetate and ionomycin for 4 days. The culture supernatants were subjected to a cytokine bead array to measure levels of interleukin (IL)-2, IL-4, IL-6, IL-10, IL-17, interferon (IFN)-γ, and tumor necrosis factor (TNF). Finally, to determine IgG subclasses of anti-platelet autoantibodies, splenocyte culture supernatants were incubated with platelets derived from BALB/c mice, followed by incubation with fluorescence-conjugated antibodies to IgG1, IgG2a, IgG2b, or IgG3. Then, the antibodies bound to platelets was detected by flow cytometry. Results: Fourteen ITP mice and 8 control mice were used at 6–8 weeks after inoculation. The proportions of Th1, Th2, and Th0 cells did not differ significantly between ITP and control mice, while the Th1/Th2 ratio was significantly increased in ITP mice than in control mice (8.3 versus 3.2, p < 0.01). The proportions of Th17 and Th1/17 cells were comparable between ITP and control mice. There was no difference in the in vitro production levels of cytokines except IL-4, which was lower in ITP mice compared to control mice (140 versus 600 pg/ml, p = 0.02). Increase in the IFN-γ/IL-4 ratio was noted in the culture supernatants from ITP mice, compared to those from control mice (15.6 versus 9.2, p = 0.04). The Th1/Th2 ratio detected by flow cytometric measurement and the IFN-γ/IL-4 ratio in in vitro cultures were correlated with each other (r = 0.85, p < 0.01). IgG subclasses of anti-platelet autoantibodies were heterogeneous among individual ITP mice, but IgG2a was the predominant subclass in the majority of ITP mice. Interestingly, a high Th1/Th2 ratio was associated with production of IgG2b anti-platelet antibodies, while the mice with a low Th1/Th2 ratio produced IgG1 anti-platelet antibodies. Conclusions: These findings suggest that induction of IgG anti-platelet autoantibody response in Treg-deficient mice is associated with Th1 bias, which is analogous to the Th balance in patients with primary ITP. The Th1/Th2 balance may modulate the autoimmune responses during expansion of CD4+ T cells in the absence of Tregs. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Populations of Human T Lymphocytes That Traverse the Vascular Endothelium Stimulated by Borrelia burgdorferi Are Enriched with Cells That Secrete Gamma Interferon

2004 ◽  
Vol 72 (3) ◽  
pp. 1530-1536 ◽  
Author(s):  
Edna I. Gergel ◽  
Martha B. Furie
Keyword(s):  
T Cells ◽  
T Lymphocytes ◽  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Vascular Endothelium ◽  
Human Peripheral Blood ◽  
Gamma Interferon ◽  
Interleukin 4 ◽  
Ifn Γ

ABSTRACT Some diseases are characterized by prevalence in the affected tissues of type 1 T lymphocytes, which secrete gamma interferon (IFN-γ) and other proinflammatory cytokines. For example, type 1 T cells predominate in the lesions of patients with Lyme disease, which is caused by the bacterium Borrelia burgdorferi. We used an in vitro model of the blood vessel wall to test the premise that the vascular endothelium actively recruits circulating type 1 T cells to such lesions. When T lymphocytes isolated from human peripheral blood were examined, the populations that traversed monolayers of resting human umbilical vein endothelial cells (HUVEC) or HUVEC stimulated by interleukin-1β or B. burgdorferi were markedly enriched for T cells that produced IFN-γ compared to the initially added population of T cells. No enrichment was seen for cells that produced interleukin-4, a marker for type 2 T lymphocytes. Very late antigen-4 and CD11/CD18 integrins mediated passage of the T cells across both resting and stimulated HUVEC, and the endothelium-derived chemokine CCL2 (monocyte chemoattractant protein 1) was responsible for the enhanced migration of T cells across stimulated HUVEC. These results suggest that the vascular endothelium may contribute to the selective accumulation of type 1 T cells in certain pathological lesions, including those of Lyme disease.

scholarly journals CRTAM determines the CD4+ cytotoxic T lymphocyte lineage

2015 ◽  
Vol 213 (1) ◽  
pp. 123-138 ◽  
Author(s):  
Arata Takeuchi ◽  
Mohamed El Sherif Gadelhaq Badr ◽  
Kosuke Miyauchi ◽  
Chitose Ishihara ◽  
Reiko Onishi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cytotoxic Activity ◽  
Cd4 T Cells ◽  
Intracellular Signaling ◽  
Ectopic Expression ◽  
T Cell Subsets ◽  
Naive T Cells ◽  
Naïve T Cells ◽  
Ifn Γ

Naive T cells differentiate into various effector T cells, including CD4+ helper T cell subsets and CD8+ cytotoxic T cells (CTL). Although cytotoxic CD4+ T cells (CD4+CTL) also develop from naive T cells, the mechanism of development is elusive. We found that a small fraction of CD4+ T cells that express class I–restricted T cell–associated molecule (CRTAM) upon activation possesses the characteristics of both CD4+ and CD8+ T cells. CRTAM+ CD4+ T cells secrete IFN-γ, express CTL-related genes, such as eomesodermin (Eomes), Granzyme B, and perforin, after cultivation, and exhibit cytotoxic function, suggesting that CRTAM+ T cells are the precursor of CD4+CTL. Indeed, ectopic expression of CRTAM in T cells induced the production of IFN-γ, expression of CTL-related genes, and cytotoxic activity. The induction of CD4+CTL and IFN-γ production requires CRTAM-mediated intracellular signaling. CRTAM+ T cells traffic to mucosal tissues and inflammatory sites and developed into CD4+CTL, which are involved in mediating protection against infection as well as inducing inflammatory response, depending on the circumstances, through IFN-γ secretion and cytotoxic activity. These results reveal that CRTAM is critical to instruct the differentiation of CD4+CTL through the induction of Eomes and CTL-related gene.

scholarly journals Sunitinib Exerts In Vitro Immunomodulatory Activity on Sarcomas via Dendritic Cells and Synergizes With PD-1 Blockade

2021 ◽  
Vol 12 ◽  
Author(s):  
Darina Ocadlikova ◽  
Mariangela Lecciso ◽  
Javier Martin Broto ◽  
Katia Scotlandi ◽  
Michele Cavo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Cell Lines ◽  
Effector T Cells ◽  
Sarcoma Cell ◽  
Ifn Γ ◽  
Sarcoma Cells

BackgroundHigh-grade sarcomas are a heterogeneous group of aggressive tumors arising in bone and soft tissues. After relapse, treatment options are limited. The multi-targeted receptor tyrosine kinase inhibitors (TKIs) sunitinib and inhibitor of PD-1 (anti-PD-1) nivolumab have shown antitumor activity in selected subtypes. In this study, we examine the role of TKIs and PD-1 based therapy in in vitro cocultures of sarcoma.MethodsThe human osteosarcoma (SaOS-2) and synovial sarcoma (SYO-1) cell lines were treated with sunitinib. After cell death and proliferation assessment, expression of PD-L1 was analyzed by flow cytometry. Sunitinib-treated sarcoma cells were cocultured with dendritic cells (DCs), and the phenotype of mature DCs was determined by flow cytometry. Mature DCs were cultured with autologous T cells. PD-1 expression on T cells, their proliferation, T regulatory cell (Tregs) induction and IFN-γ production, before and after nivolumab exposure, were analyzed.ResultsAlong with its anti-proliferative and direct pro-apoptotic effect on sarcoma cell lines, sunitinib prompted PD-L1 upregulation on sarcoma cells. Interestingly, sunitinib-treated sarcoma cells drive DCs to full maturation and increase their capacity to induce sarcoma-reactive T cells to produce IFN-γ. Conversely, no effect on T cell proliferation and T cell subpopulation composition was observed. Moreover, both bone and synovial sarcoma cell lines induced Tregs through DCs but sunitinib treatment completely abrogated Treg induction. Finally, sarcoma cell lines induced PD-1 upregulation on both effector T cells and Tregs when loaded into DCs, providing a rationale for using PD-1 blockade. Indeed, PD-1 blockade by nivolumab synergized with sunitinib in inducing IFN-γ-producing effector T cells.ConclusionsTaken together, our in vitro data indicate that the treatment of sarcoma cells with sunitinib can exert significant changes on immune cell subsets toward immune activation, leading to DC-based cross-priming of IFN-γ-producing effector T cells and reduced Treg induction. PD-1 blockade with nivolumab has a synergistic effect with sunitinib, supporting the use of TKI and anti-PD-1 approach in sarcomas, and perhaps in other cancers. DC-targeted drugs, including toll-like receptor 3 inhibitors and CD47 inhibitors, are under development and our preclinical model might help to better design their clinical application.

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