Kari Dugger

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 3x/week for 4weeks; exercise was performed as treadmill running (13.5m/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-beta and lesser amounts of IL-10 and IL-17; however, blocking TGF-beta 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.

β2‐Integrins in demyelinating disease: not adhering to the paradigm

The β2‐integrins are a subfamily of integrins expressed on leukocytes that play an essential role in leukocyte trafficking, activation, and many other functions. Studies in EAE, the animal model for multiple sclerosis, show differential requirements for β2‐integrins in this disease model, ranging from critical in the case of LFA‐1 (CD11a/CD18) to unimportant in the case of CD11d/CD18. Importantly, expression of β2‐integrins on T cell subsets provides some clues as to the function(s) these adhesion molecules play in disease development. For example, transferred EAE studies have shown that Mac‐1 (CD11b/CD18) expression on αβ T cells is critical for disease development, and the absence of LFA‐1 on Tregs in recipient mice results in exacerbated disease. In this review, we summarize recent findings regarding the role of β2‐integrins in demyelinating disease and new information about the role of β2‐integrins with respect to alterations in Treg numbers and function. In addition, we discuss the potential for targeting β2‐integrins in human demyelinating disease in light of the recent animal model studies.

Effector and suppressor roles for LFA-1 during the development of experimental autoimmune encephalomyelitis.

LFA-1 (CD11a/CD18) is a member of the beta(2)-integrin family of adhesion molecules important in leukocyte trafficking and activation. Although LFA-1 is thought to contribute to the development of experimental autoimmune encephalomyelitis (EAE) primarily through its functions on effector T cells, its importance on other leukocyte populations remains unexplored. To address this question, we performed both adoptive transfer EAE experiments involving CD11a(-/-) mice and trafficking studies using bioluminescent T cells expressing luciferase under the control of a CD2 promoter (T-lux cells). Transfer of encephalitogenic CD11a(-/-) T cells to wild type mice resulted in a significant reduction in overall EAE severity compared to control transfers. We also observed, using in vivo imaging techniques, that CD11a(-/-) T-lux cells readily infiltrated lymph nodes and the CNS of wild type recipients with kinetics comparable to CD11a(+/+) transfers, although their overall numbers in these organs were reduced. Surprisingly, transfer of encephalitogenic wild type T cells to CD11a(-/-) mice induced a severe and sometimes fatal EAE disease course, associated with massive T cell infiltration and proliferation in the CNS. These data indicate that LFA-1 expression on leukocytes in recipient mice plays an important immunomodulatory role in EAE. Thus, LFA-1 acts as a key regulatory adhesion molecule during the development of EAE, serving both pro- and anti-inflammatory roles in disease pathogenesis.

Imaging CD8+ T cell dynamics in vivo using a transgenic luciferase reporter.

After activation, populations of antigen-specific T cells flow between sites of antigen expression, local lymphoid structures and other lymphoid and non-lymphoid organs. In this study, we documented the in vivo dynamics of a CD8(+) T cell response to antigen delivered using herpes simplex virus amplicon vectors and revealed several unexpected features. First, the T cells localized to the site of vector injection, as well as the draining lymph node within 24-48 h. Second, the major site to which T cells later redistributed were intra-abdominal lymphoid organs, including milky spots, mesenteric and lumbar lymph nodes. We determined the relationship between bioluminescent signal and antigen-specific T cell numbers in various lymphoid organs, and concluded that bioluminescent signal is a valid surrogate measure of T cell abundance in superficial lymph nodes, but not in deeper structures such as the spleen.

Epithelial cells as immune effector cells: The role of CD40

Through the expression of inflammatory mediators and immune-related molecules, epithelial cells function as immune effector cells in a wide variety of tissues; the expression of the CD40 receptor on these cells contributes this role. Engagement of CD40 activates epithelial cells and results in their release of pro- and anti-inflammatory mediators as well as pro-fibrotic molecules. As such, epithelial CD40 has been implicated in the pathogenesis of inflammatory disorders, generation of self-tolerance, and rejection of allografts.

Bioluminescence-based visualization of CD4 T cell dynamics using a T lineage-specific luciferase transgenic model1

BackgroundRapid clonal expansion of T cells occurs in response to antigenic challenges. The kinetics of the T cell response has previously been described using tissue-based studies performed at defined time points. Luciferase bioluminescence has recently been utilized for non-invasive analysis of in vivo biologic processes in real-time.ResultsWe have created a novel transgenic mouse model (T-Lux) using a human CD2 mini-gene to direct luciferase expression specifically to the T cell compartment. T-Lux T cells demonstrated normal homing patterns within the intact mouse and following adoptive transfer. Bioluminescent signal correlated with T cell numbers in the whole body images as well as within specific organ regions of interest. Following transfer into lymphopenic (RAG2-/-) recipients, homeostatic proliferation of T-Lux T cells was visualized using bioluminescent imaging. Real-time bioluminescent analysis of CD4+ T cell antigen-specific responses enabled real-time comparison of the kinetics and magnitude of clonal expansion and contraction in the inductive lymph node and tissue site of antigen injection. T cell expansion was dose-dependent despite the presence of supraphysiologic numbers of OVA-specific OT-II transgenic TCR T-Lux T cells. CD4+ T cells subsequently underwent a rapid (3–4 day) contraction phase in the draining lymph node, with a delayed contraction in the antigen delivery site, with bioluminescent signal diminished below initial levels, representing TCR clonal frequency control.ConclusionThe T-Lux mouse provides a novel, efficient model for tracking in vivo aspects of the CD4+ T cell response to antigen, providing an attractive approach for studies directed at immunotherapy or vaccine design.

Gemcitabine treatment promotes immunosuppressive microenvironment in pancreatic tumors by supporting the infiltration, growth, and polarization of macrophages

Chemotherapy-induced immunosuppression poses an additional challenge to its limited efficacy in pancreatic cancer (PC). Here we investigated the effect of gemcitabine on macrophages, which are the first line of immune-defense mechanisms. We observed an increased presence of macrophages in orthotopic human pancreatic tumor xenografts from mice treated with gemcitabine as compared to those from vehicle only-treated mice. Conditioned media from gemcitabine-treated PC cells (Gem-CM) promoted growth, migration and invasion of RAW264.7 macrophage. In addition, Gem-CM also induced upregulation of M2-polarized macrophage markers, arginase-1 and TGF-β1. Cytokine profiling of gemcitabine-treated PC cells identified IL-8 as the most differentially-expressed cytokine. Incubation of Gem-CM with IL-8 neutralizing antibody diminished its ability to induce growth, migration and invasion of RAW264.7 macrophages, but did not abrogate their M2 polarization. Together, our findings identify IL-8 as an important mediator in the gemcitabine-induced infiltration of macrophages within the pancreatic tumor microenvironment and suggest the requirement of additional mechanism(s) for macrophage polarization.