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Dynamics of central and peripheral immunomodulation in a murine glioma model.

Kennedy BC, Maier LM, D'Amico R, Mandigo CE, Fontana EJ, Waziri A, Assanah MC, Canoll P, Anderson RC, Anderson DE, Bruce JN - BMC Immunol. (2009)

Bottom Line: An increase in TIL frequency was also observed at these final time points.These data provide insight into the kinetics of the immunosuppressive state associated with tumor growth in a murine model of human gliomas.Functional impairment of TAMs occurs relatively late in the course of GBM tumor growth, potentially providing a window of opportunity for therapeutic strategies directed towards preventing their functional impairment.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurosurgery, Gabriele Bartoli Brain Tumor Research Laboratory Columbia University, New York, NY, USA. bck2104@columbia.edu

ABSTRACT

Background: Immunosuppression by gliomas contributes to tumor progression and treatment resistance. It is not known when immunosuppression occurs during tumor development but it likely involves cross-talk among tumor cells, tumor-associated macrophages and microglia (TAMs), and peripheral as well as tumor-infiltrating lymphocytes (TILs).

Results: We have performed a kinetic study of this immunomodulation, assessing the dynamics of immune infiltration and function, within the central nervous system (CNS) and peripherally. PDGF-driven murine glioma cells were injected into the white matter of 13 mice. Four mice were sacrificed 13 days post-injection (dpi), four mice at 26 dpi, and five mice at 40 dpi. Using multiparameter flow cytometry, splenic T cells were assessed for FoxP3 expression to identify regulatory T cells (Tregs) and production of IFN-gamma and IL-10 after stimulation with PMA/ionomycin; within the CNS, CD4+ TILs were quantified, and TAMs were quantified and assessed for TNF-alpha and IL-10 production after stimulation with LPS. Peripheral changes associated with tumor development were noted prior to effects within the CNS. The percentage of FoxP3+ regulatory T cells (Tregs) increased by day 26, with elevated frequencies throughout the duration of the study. This early increase in Tregs was paralleled by an increase in IL-10 production from Tregs. At the final time points examined (tumor morbidity or 40 dpi), there was an increase in the frequency of TAMs with decreased capacity to secrete TNF-alpha. An increase in TIL frequency was also observed at these final time points.

Conclusion: These data provide insight into the kinetics of the immunosuppressive state associated with tumor growth in a murine model of human gliomas. Functional impairment of TAMs occurs relatively late in the course of GBM tumor growth, potentially providing a window of opportunity for therapeutic strategies directed towards preventing their functional impairment.

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Frequency of tumor-infiltrating lymphocytes (TILs) in ex vivo tumor specimens over the course of tumor development. Specimens were obtained prior to and at three time-points following tumor induction and stained for CD3, CD4, and CD45 expression. (A) An increase in the frequency of TILs as characterized by CD3 and CD45 expression was observed at the late time-point (0.3% vs. 2.0%, P = 0.047). (B) The percentage of CD4+ T cells among TILs was reduced at the late time point (76.5% vs. 47.5%, P = 0.0003). 13 mice were analyzed at 13 dpi (n = 4), 26 dpi (n = 4), and at 29 dpi upon exhibiting clinical tumor morbidity (n = 1) or at 40 dpi (n = 4). Differences between the means at each time point were tested using two-sided t tests with unequal variances. *, P < 0.05; **, P < 0.01, ***, P < 0.001.
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Figure 4: Frequency of tumor-infiltrating lymphocytes (TILs) in ex vivo tumor specimens over the course of tumor development. Specimens were obtained prior to and at three time-points following tumor induction and stained for CD3, CD4, and CD45 expression. (A) An increase in the frequency of TILs as characterized by CD3 and CD45 expression was observed at the late time-point (0.3% vs. 2.0%, P = 0.047). (B) The percentage of CD4+ T cells among TILs was reduced at the late time point (76.5% vs. 47.5%, P = 0.0003). 13 mice were analyzed at 13 dpi (n = 4), 26 dpi (n = 4), and at 29 dpi upon exhibiting clinical tumor morbidity (n = 1) or at 40 dpi (n = 4). Differences between the means at each time point were tested using two-sided t tests with unequal variances. *, P < 0.05; **, P < 0.01, ***, P < 0.001.

Mentions: TILs were defined as cells present within ex vivo tumor specimens that expressed both CD3 and CD45 (Figures 1 and 4). Between the early and intermediate time points, there were no observed changes in TIL frequency (0.2% vs. 0.3%) (Figure 4A). However, later in the course of disease, when comparing the intermediate and late time points, an increase in TIL frequency was observed (0.3% vs. 2.0%, P = 0.047) (Figure 4A). Among these CD3+CD45+ TILs, we observed a decrease between the early/intermediate and the late time point in the proportion of TILs that were CD4+ (76.5% vs. 47.5%, P = 0.0003), suggesting that alternate T cell subsets may be infiltrating the tumor microenvironment, such as CD8+ CTLs or NK T cells (Figure 4B).


Dynamics of central and peripheral immunomodulation in a murine glioma model.

Kennedy BC, Maier LM, D'Amico R, Mandigo CE, Fontana EJ, Waziri A, Assanah MC, Canoll P, Anderson RC, Anderson DE, Bruce JN - BMC Immunol. (2009)

Frequency of tumor-infiltrating lymphocytes (TILs) in ex vivo tumor specimens over the course of tumor development. Specimens were obtained prior to and at three time-points following tumor induction and stained for CD3, CD4, and CD45 expression. (A) An increase in the frequency of TILs as characterized by CD3 and CD45 expression was observed at the late time-point (0.3% vs. 2.0%, P = 0.047). (B) The percentage of CD4+ T cells among TILs was reduced at the late time point (76.5% vs. 47.5%, P = 0.0003). 13 mice were analyzed at 13 dpi (n = 4), 26 dpi (n = 4), and at 29 dpi upon exhibiting clinical tumor morbidity (n = 1) or at 40 dpi (n = 4). Differences between the means at each time point were tested using two-sided t tests with unequal variances. *, P < 0.05; **, P < 0.01, ***, P < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2654428&req=5

Figure 4: Frequency of tumor-infiltrating lymphocytes (TILs) in ex vivo tumor specimens over the course of tumor development. Specimens were obtained prior to and at three time-points following tumor induction and stained for CD3, CD4, and CD45 expression. (A) An increase in the frequency of TILs as characterized by CD3 and CD45 expression was observed at the late time-point (0.3% vs. 2.0%, P = 0.047). (B) The percentage of CD4+ T cells among TILs was reduced at the late time point (76.5% vs. 47.5%, P = 0.0003). 13 mice were analyzed at 13 dpi (n = 4), 26 dpi (n = 4), and at 29 dpi upon exhibiting clinical tumor morbidity (n = 1) or at 40 dpi (n = 4). Differences between the means at each time point were tested using two-sided t tests with unequal variances. *, P < 0.05; **, P < 0.01, ***, P < 0.001.
Mentions: TILs were defined as cells present within ex vivo tumor specimens that expressed both CD3 and CD45 (Figures 1 and 4). Between the early and intermediate time points, there were no observed changes in TIL frequency (0.2% vs. 0.3%) (Figure 4A). However, later in the course of disease, when comparing the intermediate and late time points, an increase in TIL frequency was observed (0.3% vs. 2.0%, P = 0.047) (Figure 4A). Among these CD3+CD45+ TILs, we observed a decrease between the early/intermediate and the late time point in the proportion of TILs that were CD4+ (76.5% vs. 47.5%, P = 0.0003), suggesting that alternate T cell subsets may be infiltrating the tumor microenvironment, such as CD8+ CTLs or NK T cells (Figure 4B).

Bottom Line: An increase in TIL frequency was also observed at these final time points.These data provide insight into the kinetics of the immunosuppressive state associated with tumor growth in a murine model of human gliomas.Functional impairment of TAMs occurs relatively late in the course of GBM tumor growth, potentially providing a window of opportunity for therapeutic strategies directed towards preventing their functional impairment.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurosurgery, Gabriele Bartoli Brain Tumor Research Laboratory Columbia University, New York, NY, USA. bck2104@columbia.edu

ABSTRACT

Background: Immunosuppression by gliomas contributes to tumor progression and treatment resistance. It is not known when immunosuppression occurs during tumor development but it likely involves cross-talk among tumor cells, tumor-associated macrophages and microglia (TAMs), and peripheral as well as tumor-infiltrating lymphocytes (TILs).

Results: We have performed a kinetic study of this immunomodulation, assessing the dynamics of immune infiltration and function, within the central nervous system (CNS) and peripherally. PDGF-driven murine glioma cells were injected into the white matter of 13 mice. Four mice were sacrificed 13 days post-injection (dpi), four mice at 26 dpi, and five mice at 40 dpi. Using multiparameter flow cytometry, splenic T cells were assessed for FoxP3 expression to identify regulatory T cells (Tregs) and production of IFN-gamma and IL-10 after stimulation with PMA/ionomycin; within the CNS, CD4+ TILs were quantified, and TAMs were quantified and assessed for TNF-alpha and IL-10 production after stimulation with LPS. Peripheral changes associated with tumor development were noted prior to effects within the CNS. The percentage of FoxP3+ regulatory T cells (Tregs) increased by day 26, with elevated frequencies throughout the duration of the study. This early increase in Tregs was paralleled by an increase in IL-10 production from Tregs. At the final time points examined (tumor morbidity or 40 dpi), there was an increase in the frequency of TAMs with decreased capacity to secrete TNF-alpha. An increase in TIL frequency was also observed at these final time points.

Conclusion: These data provide insight into the kinetics of the immunosuppressive state associated with tumor growth in a murine model of human gliomas. Functional impairment of TAMs occurs relatively late in the course of GBM tumor growth, potentially providing a window of opportunity for therapeutic strategies directed towards preventing their functional impairment.

Show MeSH
Related in: MedlinePlus