scholarly journals Critical roles for Rac GTPases in T-cell migration to and within lymph nodes

Blood ◽  
2010 ◽  
Vol 116 (25) ◽  
pp. 5536-5547 ◽  
Author(s):  
Mustapha Faroudi ◽  
Miroslav Hons ◽  
Agnieszka Zachacz ◽  
Celine Dumont ◽  
Ruth Lyck ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Lymph Nodes ◽  
Chemokine Receptors ◽  
White Pulp ◽  
Multiple Defects ◽  
Rac Gtpases ◽  
Sphingosine 1 Phosphate

Abstract Naive T cells continuously recirculate between secondary lymphoid tissue via the blood and lymphatic systems, a process that maximizes the chances of an encounter between a T cell and its cognate antigen. This recirculation depends on signals from chemokine receptors, integrins, and the sphingosine-1-phosphate receptor. The authors of previous studies in other cell types have shown that Rac GTPases transduce signals leading to cell migration and adhesion; however, their roles in T cells are unknown. By using both 3-dimensional intravital and in vitro approaches, we show that Rac1- and Rac2-deficient T cells have multiple defects in this recirculation process. Rac-deficient T cells home very inefficiently to lymph nodes and the white pulp of the spleen, show reduced interstitial migration within lymph node parenchyma, and are defective in egress from lymph nodes. These mutant T cells show defective chemokine-induced chemotaxis, chemokinesis, and adhesion to integrin ligands. They have reduced lateral motility on endothelial cells and transmigrate in-efficiently. These multiple defects stem from critical roles for Rac1 and Rac2 in transducing chemokine and sphingosine-1-phosphate receptor 1 signals leading to motility and adhesion.

scholarly journals The Reduced Expression of 6ckine in the plt Mouse Results from the Deletion of One of Two 6ckine Genes

1999 ◽  
Vol 190 (8) ◽  
pp. 1183-1188 ◽  
Author(s):  
Galya Vassileva ◽  
Hortensia Soto ◽  
Albert Zlotnik ◽  
Hideki Nakano ◽  
Terutaka Kakiuchi ◽  
...  
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
T Cell ◽  
T Lymphocytes ◽  
Lymph Nodes ◽  
Lymphoid Organs ◽  
Peyer's Patches ◽  
White Pulp ◽  
Naive T Lymphocytes

6Ckine is an unusual chemokine capable of attracting naive T lymphocytes in vitro. It has been recently reported that lack of 6Ckine expression in lymphoid organs is a prominent characteristic of mice homozygous for the paucity of lymph node T cell (plt) mutation. These mice show reduced numbers of T cells in lymph nodes, Peyer's patches, and the white pulp of the spleen. The genetic reason for the lack of 6Ckine expression in the plt mouse, however, has remained unknown. Here we demonstrate that mouse 6Ckine is encoded by two genes, one of which is expressed in lymphoid organs and is deleted in plt mice. A second 6Ckine gene is intact and expressed in the plt mouse.

MiR-155 Impacts T Cell Migration in Acute Graft-Versus-Host Disease (aGVHD)

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3080-3080 ◽  
Author(s):  
Nina C. Zitzer ◽  
Patricia A. Taylor ◽  
Apollinaire Ngankeu ◽  
Yvonne A. Efebera ◽  
Steven M. Devine ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Chemokine Receptors ◽  
Ccr5 Expression ◽  
Post Transplant ◽  
T Cell Migration ◽  
T Cell Infiltration ◽  
Donor T Cells

Abstract Introduction: We reported that microRNA-155 (miR-155) expression is upregulated in donor T cells during aGVHD and mice receiving miR-155 knock-out (KO) donor splenocytes do not exhibit lethal GVHD and have improved survival as compared to mice receiving wild type (WT) splenocytes.1 While we showed that miR-155 does not affect the allo-reactive proliferative potential of T cells, a significant decrease in the expression of the homing receptors CCR5, CXCR4, and S1P1 was found on miR-155-KO T cells, suggesting that the loss of miR-155 could impair the migration of donor T cells to aGVHD target organs resulting in less lethality. Here, we further investigate the impact of miR-155 expression in T cell migration. Materials and Methods: Lethally irradiated BALB/c or B6D2F1 recipients were infused with T cell depleted WT bone marrow (BM) cells (5x10^6) and GFP expressing miR-155 KO or GFP-B6 WT T cells (1x10^6). Recipients were sacrificed at day 7, 14 and 21 post-transplant, organs harvested and donor T cell infiltration evaluated via confocal microscopy. Transwell migration assays towards CCR5 ligands macrophage inflammatory protein-1a (MIP-1a) (100ng/mL) and RANTES (100ng/mL) was performed utilizing WT or miR-155-KO T cells activated using irradiated BALB/c splenocytes as allogeneic stimulators at a stimulator: responder ratio of 1:5. Lower chambers with medium only served as a control for spontaneous migration. CCR5 ligand-dependent migration was calculated according to the formula: Migration Index (MI) = number of cells CCR5 ligands / number of cells medium only. Results: On days 7, 14 and 21 post transplant, recipient mice were sacrificed, and tissues harvested in order to study the kinetics of miR-155 KO T cell migration following allogeneic hematopoietic stem cell transplant. There was a dramatic decrease in T cell infiltration of peripheral organs (PeyerÕs patches, liver, lung and skin) in recipients of miR-155-KO T cells as compared to WT T cells as evidenced by confocal microscopy of GFP labeled donor cells, Figure 1. We reasoned that these effects could be due to the modulation of CCR5 and other chemokine receptors by miR-155. There was a significant decrease in CCR5 mRNA and protein expression in miR-155-KO versus WT donor T cells obtained from recipient mice at the time of aGVHD. To demonstrate the functional significance of decreased CCR5 expression in miR-155 KO donor T cells, we performed in vitro transwell migration assays to CCR5 ligands RANTES and MIP-1a. To our knowledge, we are the first to show that allo-activated miR-155 KO T cells show significantly reduced migration towards CCR5 ligands, as demonstrated by the average MI of 1.08, when compared to the average MI of WT T cells of 4.79, p=0.004, Figure 2. There were lower percentages of CCR5 positive T cells and decreased mean fluorescent intensity in the miR-155 KO T cells after allogeneic stimulation when compared to WT T cells, both in the CD4+ and CD8+ populations, confirming lower CCR5 expression in miR-155 KO T cells after in vitro allogeneic stimulation. To further elucidate the mechanism of miR-155 mediated modulation of CCR5 expression, we focused on long non-coding RNA (lncRNA) LincR-Ccr2-5′AS located in the vicinity of several chemokine receptor encoding genes including CCR1, CCR2 and CCR5, known to be important for migration of Th2 cells. We found that LincR-Ccr2-5′AS has 3 potential miR-155 binding sites and so set out to determine if miR-155 negatively regulates the expression of this lncRNA, thereby influencing chemokine receptor expression as well as T cell migration. We isolated T cells from B6D2F1 recipients 21 days post-transplant, and showed a significant decrease in CCR5 mRNA expression in miR-155 KO versus WT donor T cells but no significant difference in the levels of LincR-Ccr2-5′AS. However, this result does not exclude the possibility that miR-155 might influence the activity rather than the levels of LincR-Ccr2-5′AS, which we hope to determine in future experiments. Conclusion: Our data suggest that miR-155 may exert its modulating effects in aGVHD by affecting T cell migration. Experiments are currently underway to determine the role of miR-155 in modulating T cell migration through other chemokine receptors such as CXCR4, as well as S1P1 and ATP receptor P2X7R. Reference 1. Ranganathan P, Heaphy CE, Costinean S, et al. Regulation of acute graft-versus-host disease by microRNA-155. Blood. 2012 May 17;119(20):4786-97. Disclosures No relevant conflicts of interest to declare.

scholarly journals Form and pattern of MUC1 expression on T cells activated in vivo or in vitro suggests a function in T-cell migration

Immunology ◽  
2003 ◽  
Vol 108 (1) ◽  
pp. 32-41 ◽  
Author(s):  
Isabel Correa ◽  
Tim Plunkett ◽  
Anda Vlad ◽  
Arron Mungul ◽  
Jessica Candelora-Kettel ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Muc1 Expression ◽  
T Cell Migration

scholarly journals In Vivo Detection of Dendritic Cell Antigen Presentation to CD4+ T Cells

1997 ◽  
Vol 185 (12) ◽  
pp. 2133-2141 ◽  
Author(s):  
Elizabeth Ingulli ◽  
Anna Mondino ◽  
Alexander Khoruts ◽  
Marc K. Jenkins
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lymph Nodes ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Delayed Type Hypersensitivity ◽  
Physical Interaction

Although lymphoid dendritic cells (DC) are thought to play an essential role in T cell activation, the initial physical interaction between antigen-bearing DC and antigen-specific T cells has never been directly observed in vivo under conditions where the specificity of the responding T cells for the relevant antigen could be unambiguously assessed. We used confocal microscopy to track the in vivo location of fluorescent dye-labeled DC and naive TCR transgenic CD4+ T cells specific for an OVA peptide–I-Ad complex after adoptive transfer into syngeneic recipients. DC that were not exposed to the OVA peptide, homed to the paracortical regions of the lymph nodes but did not interact with the OVA peptide-specific T cells. In contrast, the OVA peptide-specific T cells formed large clusters around paracortical DC that were pulsed in vitro with the OVA peptide before injection. Interactions were also observed between paracortical DC of the recipient and OVA peptide-specific T cells after administration of intact OVA. Injection of OVA peptide-pulsed DC caused the specific T cells to produce IL-2 in vivo, proliferate, and differentiate into effector cells capable of causing a delayed-type hypersensitivity reaction. Surprisingly, by 48 h after injection, OVA peptide-pulsed, but not unpulsed DC disappeared from the lymph nodes of mice that contained the transferred TCR transgenic population. These results demonstrate that antigen-bearing DC directly interact with naive antigen-specific T cells within the T cell–rich regions of lymph nodes. This interaction results in T cell activation and disappearance of the DC.

scholarly journals CD40L+ CD4+ memory T cells migrate in a CD62P-dependent fashion into reactive lymph nodes and license dendritic cells for T cell priming

2008 ◽  
Vol 205 (11) ◽  
pp. 2561-2574 ◽  
Author(s):  
Alfonso Martín-Fontecha ◽  
Dirk Baumjohann ◽  
Greta Guarda ◽  
Andrea Reboldi ◽  
Miroslav Hons ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Lymph Nodes ◽  
Memory T Cells ◽  
Effector Memory ◽  
High Endothelial Venules ◽  
T Cell Priming ◽  
Dc Maturation

There is growing evidence that the maturation state of dendritic cells (DCs) is a critical parameter determining the balance between tolerance and immunity. We report that mouse CD4+ effector memory T (TEM) cells, but not naive or central memory T cells, constitutively expressed CD40L at levels sufficient to induce DC maturation in vitro and in vivo in the absence of antigenic stimulation. CD4+ TEM cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules. Trafficking of CD4+ TEM cells into chronic reactive lymph nodes maintained resident DCs in a mature state and promoted naive T cell responses and experimental autoimmune encephalomyelitis (EAE) to antigens administered in the absence of adjuvants. Antibodies to CD62P, which blocked CD4+ TEM cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE. These results show that TEM cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

scholarly journals Efficient Expansion of Polyfunctional Virus-Specific T Cells from Human Lymph Nodes: Implications for Cellular Therapies

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3715-3715 ◽  
Author(s):  
Stuart P. Weisberg ◽  
Mark Chang ◽  
Pawel Muranski ◽  
Donna Farber
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
T Cell ◽  
Lymph Nodes ◽  
Granzyme B ◽  
Organ Donors ◽  
Intracellular Cytokine ◽  
Viral Antigens ◽  
Tnf Α

Abstract BACKGROUND: Adoptive transfer of in vitro expanded autologous and allogeneic virus specific T (VST) cells has been successfully used to prevent and treat EBV viral reactivation in transplant patients and aggressive EBV-driven cancers such as post-transplant lymphoproliferative disease (PTLD), nasopharyngeal carcinoma, and extranodal NK/T-cell lymphoma. Due to the easy accessibility of peripheral blood, VST cell products are universally generated from circulating T cells. However, the T cells in circulation represent only a minor fraction of T cells in the body with most residing in tissue sites, particularly lymph nodes. Recent animal data suggest that unique T cell populations that sustain memory responses to chronic viral infections exclusively reside in lymph nodes. The efficacy of using lymph node-derived T cells for adoptive cellular therapy has not been reported. AIMS: To assess the feasibility of generating VST cells from human lymph nodes using our clinically-compatible strategy and to test the ability of T cells derived from peripheral lymph nodes to expand in response to EBV-derived viral antigens and display functionality compared to T cells derived from blood. METHODS: Human blood and lymphoid tissues were obtained from brain dead organ donors at the time of organ procurement for transplantation through an approved protocol with LiveOnNY. Human blood was also obtained from healthy volunteers through an IRB approved protocol. Donors were cancer free, EBV seropositive, and negative for hepatitis B, C and HIV. Lymph nodes were isolated in sterile fashion, enzymatically and mechanically digested to a single cell suspension. Overlapping 15 mer peptide pools (pepmixes) of EBV latency viral antigens EBNA1 and LMP1 (JPT, Berlin, Germany) were used for expansion and restimulation. T cells were isolated by fluorescence activated cell sorting and stimulated with peptide pulsed irradiated mononuclear cells from blood (healthy donors) or spleen (organ donors), followed by 14-day culture in IL-7 and 15 (10 ng/mL) with addition of IL-2 (20 IU/mL) starting on day +3. Expanded T cells were then rested overnight and restimulated with individual pepmixes for 6 hours followed by surface marker and intracellular cytokine staining to evaluate differentiation state and function. RESULTS: T cells from lymph node, blood and spleen displayed comparable levels of in vitro expansion (Fig. 1A). Compared to blood, there was increased EBNA1 reactive cell frequency (TNF-α/IFN-ꝩ positive) in the lymph node derived T cell cultures (Fig. 1B). VST cells were predominately CD8 from blood (56 ± 15%) and lymph node (86 ± 3.8%) but not spleen (24 ± 6.4%). One donor in this cohort displayed significant reactivity for LMP1. Both blood and lymph node derived VST cells were uniformly positive for granzyme B and the degranulation marker CD107a (Fig. 1C). Remarkably, the lymph node derived VST cells displayed markedly enhanced polyfunctionality with robust secretion of IL-2, as well as increased surface expression of the co-stimulatory molecule CD28 with 33±3.6% displaying strong co-expression of both molecules compared to 8.1±2.7% of those derived from blood (Fig. 1D). CONCLUSION: These results suggest that it is feasible to generate highly-reactive EBV-specific T cells from human lymph node tissue using the methodology compatible with good manufacturing practice (GMP). In contrast to VST cells derived from peripheral blood, increased expression of CD28 and IL-2 on lymph node derived EBV reactive cells may indicate a superior capacity to survive, expand in vivo and eradicate EBV-driven disease upon adoptive transfer. Figure 1. Characterization of lymph node derived EBV reactive T cells. A) Expanded T cells from Blood (BL), iliac lymph node (iLN), mesenteric lymph node (MLN), and spleen (Spl), were restimulated with EBNA1 or LMP1 peptides for 6 hours, followed by surface and intracellular cytokine stain and flow cytometry. (A) Shown are the live cell counts in each culture per 100,000 cells plated; (B) the frequencies of VST cells (TNF-α/IFN-ꝩ positive) in each culture and (C) the frequency of cytotoxic CD107a / Granzyme B (GZMB) positive cells within the VST cell population. (D) Representative flow cytometry data from matched samples of an organ donor is shown to the left of compiled data showing subsets of the EBNA1 reactive cells defined by CD28 and IL-2 expression. (mean ± SEM, n = 2-4). *P < 0.05 t-test with comparison to blood. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

Protein kinase C: a regulator of cytoskeleton remodelling and T-cell migration

2014 ◽  
Vol 42 (6) ◽  
pp. 1490-1497 ◽  
Author(s):  
Aideen Long ◽  
Michael Freeley
Keyword(s):  
T Cells ◽  
Protein Kinase C ◽  
Cell Migration ◽  
T Cell ◽  
Protein Kinase ◽  
Immune Responses ◽  
Chemokine Receptors ◽  
T Cell Subsets ◽  
T Cell Migration ◽  
Kinase C

Protein kinase C (PKC) is a family of ten serine/threonine kinases that have diverse roles in the signalling pathways regulating cellular proliferation, differentiation, apoptosis and immune responses. Elucidating roles for individual PKC isoforms in the immune responses of T-cells have long been a challenging prospect, because these cells are known to express nine of these isoforms. A variety of approaches including the use of knockout mice, overexpression of kinase-inactive mutants, cell-permeable peptides, pharmacological inhibitors and siRNAs have shown that PKCs regulate the production of inflammatory cytokines and the cytotoxic responses of various T-cell subsets. Central to the T-cell immune response is a requirement to migrate to various organs and tissues in search of pathogens and micro-organisms. T-cell migration is guided by specific sets of chemokines and integrin ligands that activate their cognate chemokine receptors and integrins on T-cells, resulting in remodelling of the cytoskeleton and the dynamic protrusive/contractile forces necessary for cell adhesion and motility. In the present article, we review the role of PKC in T-cell migration, with an emphasis on studies that have defined their roles in cytoskeletal remodelling, cell polarity and intracellular trafficking downstream of chemokine receptors and integrins.

scholarly journals ACTIVITY OF HOST-DERIVED T CELLS WHICH DIFFERENTIATE IN NUDE MICE GRAFTED WITH CO-ISOGENIC OR ALLOGENEIC THYMUSES

1974 ◽  
Vol 139 (5) ◽  
pp. 1215-1227 ◽  
Author(s):  
Berenice Kindred ◽  
Francis Loor
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lymph Nodes ◽  
Nude Mice ◽  
Precursor Cells ◽  
Donor Strain ◽  
Skin Grafts ◽  
Host Type ◽  
Spleen And Lymph Nodes

If nude mice are grafted with a neonatal thymus, host type precursor cells develop within the graft thymus and after about 6 wk the T-cell population of the thymus, spleen, and lymph nodes is of host type. However, immunological responsiveness produced in nude mice in this manner is incomplete: (a) the ability to react to T-cell mitogens in vitro is greater than in untreated nudes but lower than in normal mice; (b) the response to T-cell dependent antigens is less than normal; and (c) the rejection of skin grafts is slower than in normal animals. Whether host precursor cells which differentiate in an allogeneic thymus are able to reject skin grafts from thymus donor strain appears to depend on the strain combination used.

scholarly journals Assessment of T cell chemotaxis to S1P

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Brandon Leon ◽  
Sarvesh Chelvanambi ◽  
Rabab Rabab ElMergawy ◽  
Moraima Noda ◽  
Bernhard Maier ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Lymph Nodes ◽  
Surface Expression ◽  
Supt1 Cell ◽  
Activated T Cells ◽  
Chemical Gradients ◽  
T Cell Migration ◽  
Transwell Filter

Introduction: Migration of leukocytes in response to chemical gradients, chemotaxis, is dependent on many factors, including cell type, surface markers, the chemoattractant, etc. Sphinogsine-1-phosphate (S1P) is a chemoattractant playing a large role in migrating activated T cells out of lymph nodes by binding to their S1P receptor, S1P1. The importance of the egress in T cells from lymph nodes is highlighted by pharmacological disruption of this migration can lead to immune dampening and thus control of multiple sclerosis, an autoimmune disease. In the case of human immunodeficiency virus (HIV), it has been shown that HIV downregulates S1P1 surface expression, effectively inhibiting chemotaxis. Our experiments attempt to study a particular HIV-encoded protein, Nef in S1P-elicited T cell migration, and to optimize the conditions for assessing T cell migration in response to S1P. Methods: In our Transwell migration assays, migration of serum-starved SupT1 cells was induced using various concentrations of S1P bound to delipidated bovine serum albumin (BSA). Before migration, cells were labeled using Calcein AM. Cells were allowed to migrate for 2-4 hours at 37°C in serum-free media. After migration, fluorescence intensity was measured using a CLARIOstar microplate reader. Results: S1P showed a direct dose-dependent response to SupT1 cell migration from 0 to 100 nM S1P. Optimization of the migration showed that both number of trans-migrated cells and those still present within the transwell filter were significant indicators of SupT1 migration. Conclusion: S1P’s chemoattractant ability is prevalent in the migration of SupT1 cells in concentrations lower than 125nM. Because we have inducible systems for HIV-Nef expression established in this cell line, these data are useful for testing the role of Nef in HIV-mediated T cell retention.

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