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Tumor-specific CD4+ T cells develop cytotoxic activity and eliminate virus-induced tumor cells in the absence of regulatory T cells.

Akhmetzyanova I, Zelinskyy G, Schimmer S, Brandau S, Altenhoff P, Sparwasser T, Dittmer U - Cancer Immunol. Immunother. (2012)

Bottom Line: We show here that there is an expansion of tumor-specific CD4(+) T cells producing cytokines and cytotoxic molecule granzyme B (GzmB) in the early phase of tumor growth.Importantly, we demonstrate that in vivo depletion of regulatory T cells (Tregs) and CD8(+) T cells in FBL-3-bearing DEREG transgenic mice augments IL-2 and GzmB production by CD4(+) T cells and increases FV-specific CD4(+) T-cell effector and cytotoxic responses leading to the complete tumor regression.Therefore, the capacity to reject tumor acquired by tumor-reactive CD4(+) T cells largely depends on the direct suppressive activity of Tregs.

View Article: PubMed Central - PubMed

Affiliation: Institute for Virology, University of Duisburg-Essen, Virchowstr 179, 45147, Essen, Germany. ilseyar.akhmetzyanova@uni-due.de

ABSTRACT
The important role of tumor-specific cytotoxic CD8(+) T cells is well defined in the immune control of the tumors, but the role of effector CD4(+) T cells is poorly understood. In the current research, we have used a murine retrovirus-induced tumor cell line of C57BL/6 mouse origin, namely FBL-3 cells, as a model to study basic mechanisms of immunological control and escape during tumor formation. This study shows that tumor-specific CD4(+) T cells are able to protect against virus-induced tumor cells. We show here that there is an expansion of tumor-specific CD4(+) T cells producing cytokines and cytotoxic molecule granzyme B (GzmB) in the early phase of tumor growth. Importantly, we demonstrate that in vivo depletion of regulatory T cells (Tregs) and CD8(+) T cells in FBL-3-bearing DEREG transgenic mice augments IL-2 and GzmB production by CD4(+) T cells and increases FV-specific CD4(+) T-cell effector and cytotoxic responses leading to the complete tumor regression. Therefore, the capacity to reject tumor acquired by tumor-reactive CD4(+) T cells largely depends on the direct suppressive activity of Tregs. We suggest that a cytotoxic CD4(+) T-cell immune response may be induced to enhance resistance against oncovirus-associated tumors.

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In vivo killing activity of CD4+ T cells after different treatment of mice: Mean percentages of killing (a) and representative histograms (b) of in vivo CTL assay are depicted. Tumor-bearing mice were depleted for their Tregs alone or additionally for their CD8+ T cells. Target cells from donor CD45.1 mice (CFSE+ and CFSE−) were co-transferred i.v. in the same amount into tumor-bearing mice. CFSE+ cells were loaded with the class II-restricted peptide recognized by CD4+ T cells, whereas CFSE− cells were used as a control population 2 h later, lymphocytes were isolated from the draining lymph nodes and analyzed by flow cytometry to determine the percentage of remaining target cells that are either CFSE+ or CFSE−. Each dot represents an individual mouse, and the mean percentages are indicated by a line. Differences between two groups are indicated (*P < 0.05). All experiments were repeated two times with comparable results
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Fig6: In vivo killing activity of CD4+ T cells after different treatment of mice: Mean percentages of killing (a) and representative histograms (b) of in vivo CTL assay are depicted. Tumor-bearing mice were depleted for their Tregs alone or additionally for their CD8+ T cells. Target cells from donor CD45.1 mice (CFSE+ and CFSE−) were co-transferred i.v. in the same amount into tumor-bearing mice. CFSE+ cells were loaded with the class II-restricted peptide recognized by CD4+ T cells, whereas CFSE− cells were used as a control population 2 h later, lymphocytes were isolated from the draining lymph nodes and analyzed by flow cytometry to determine the percentage of remaining target cells that are either CFSE+ or CFSE−. Each dot represents an individual mouse, and the mean percentages are indicated by a line. Differences between two groups are indicated (*P < 0.05). All experiments were repeated two times with comparable results

Mentions: To analyze whether the increased expression of GzmB correlated with improved tumor-specific lysis of target cells after Treg depletion, we performed a series of in vivo killing experiments. The in vivo killing activity was quantified in the drLN of each mouse during tumor rejection. In non-depleted animals, CD4+ T cells showed a modest in vivo killing activity not exceeding a mean of 13 % target cell lysis (Fig. 6a). Surprisingly, depletion of Tregs alone did not significantly improve the lysis of target cells. In contrast, simultaneous ablation of CD8+ T cells and Treg significantly enhances the killing of peptide-loaded cells (Fig. 6a, b), which correlated with the high frequency of GzmB-producing cells in this group of mice (Fig. 5c, d). Notably, in this group, tumor growth was completely rejected even in the absence of CD8+ T cells (Fig. 4e). To demonstrate that cytotoxic CD4+ T cells mediated the target cell killing in the group of CD8+ T cell plus Treg depleted mice, we additionally depleted the effector CD4+ T cells. This completely abrogated the MHC II-restricted killing activity (Fig. 6a). Collectively, these data suggest that CD4+ T cells can gain cytotoxic activity against tumor cells when CD8+ T cells are not active but this activity is tightly controlled by Tregs during tumor rejection.Fig. 6


Tumor-specific CD4+ T cells develop cytotoxic activity and eliminate virus-induced tumor cells in the absence of regulatory T cells.

Akhmetzyanova I, Zelinskyy G, Schimmer S, Brandau S, Altenhoff P, Sparwasser T, Dittmer U - Cancer Immunol. Immunother. (2012)

In vivo killing activity of CD4+ T cells after different treatment of mice: Mean percentages of killing (a) and representative histograms (b) of in vivo CTL assay are depicted. Tumor-bearing mice were depleted for their Tregs alone or additionally for their CD8+ T cells. Target cells from donor CD45.1 mice (CFSE+ and CFSE−) were co-transferred i.v. in the same amount into tumor-bearing mice. CFSE+ cells were loaded with the class II-restricted peptide recognized by CD4+ T cells, whereas CFSE− cells were used as a control population 2 h later, lymphocytes were isolated from the draining lymph nodes and analyzed by flow cytometry to determine the percentage of remaining target cells that are either CFSE+ or CFSE−. Each dot represents an individual mouse, and the mean percentages are indicated by a line. Differences between two groups are indicated (*P < 0.05). All experiments were repeated two times with comparable results
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3569596&req=5

Fig6: In vivo killing activity of CD4+ T cells after different treatment of mice: Mean percentages of killing (a) and representative histograms (b) of in vivo CTL assay are depicted. Tumor-bearing mice were depleted for their Tregs alone or additionally for their CD8+ T cells. Target cells from donor CD45.1 mice (CFSE+ and CFSE−) were co-transferred i.v. in the same amount into tumor-bearing mice. CFSE+ cells were loaded with the class II-restricted peptide recognized by CD4+ T cells, whereas CFSE− cells were used as a control population 2 h later, lymphocytes were isolated from the draining lymph nodes and analyzed by flow cytometry to determine the percentage of remaining target cells that are either CFSE+ or CFSE−. Each dot represents an individual mouse, and the mean percentages are indicated by a line. Differences between two groups are indicated (*P < 0.05). All experiments were repeated two times with comparable results
Mentions: To analyze whether the increased expression of GzmB correlated with improved tumor-specific lysis of target cells after Treg depletion, we performed a series of in vivo killing experiments. The in vivo killing activity was quantified in the drLN of each mouse during tumor rejection. In non-depleted animals, CD4+ T cells showed a modest in vivo killing activity not exceeding a mean of 13 % target cell lysis (Fig. 6a). Surprisingly, depletion of Tregs alone did not significantly improve the lysis of target cells. In contrast, simultaneous ablation of CD8+ T cells and Treg significantly enhances the killing of peptide-loaded cells (Fig. 6a, b), which correlated with the high frequency of GzmB-producing cells in this group of mice (Fig. 5c, d). Notably, in this group, tumor growth was completely rejected even in the absence of CD8+ T cells (Fig. 4e). To demonstrate that cytotoxic CD4+ T cells mediated the target cell killing in the group of CD8+ T cell plus Treg depleted mice, we additionally depleted the effector CD4+ T cells. This completely abrogated the MHC II-restricted killing activity (Fig. 6a). Collectively, these data suggest that CD4+ T cells can gain cytotoxic activity against tumor cells when CD8+ T cells are not active but this activity is tightly controlled by Tregs during tumor rejection.Fig. 6

Bottom Line: We show here that there is an expansion of tumor-specific CD4(+) T cells producing cytokines and cytotoxic molecule granzyme B (GzmB) in the early phase of tumor growth.Importantly, we demonstrate that in vivo depletion of regulatory T cells (Tregs) and CD8(+) T cells in FBL-3-bearing DEREG transgenic mice augments IL-2 and GzmB production by CD4(+) T cells and increases FV-specific CD4(+) T-cell effector and cytotoxic responses leading to the complete tumor regression.Therefore, the capacity to reject tumor acquired by tumor-reactive CD4(+) T cells largely depends on the direct suppressive activity of Tregs.

View Article: PubMed Central - PubMed

Affiliation: Institute for Virology, University of Duisburg-Essen, Virchowstr 179, 45147, Essen, Germany. ilseyar.akhmetzyanova@uni-due.de

ABSTRACT
The important role of tumor-specific cytotoxic CD8(+) T cells is well defined in the immune control of the tumors, but the role of effector CD4(+) T cells is poorly understood. In the current research, we have used a murine retrovirus-induced tumor cell line of C57BL/6 mouse origin, namely FBL-3 cells, as a model to study basic mechanisms of immunological control and escape during tumor formation. This study shows that tumor-specific CD4(+) T cells are able to protect against virus-induced tumor cells. We show here that there is an expansion of tumor-specific CD4(+) T cells producing cytokines and cytotoxic molecule granzyme B (GzmB) in the early phase of tumor growth. Importantly, we demonstrate that in vivo depletion of regulatory T cells (Tregs) and CD8(+) T cells in FBL-3-bearing DEREG transgenic mice augments IL-2 and GzmB production by CD4(+) T cells and increases FV-specific CD4(+) T-cell effector and cytotoxic responses leading to the complete tumor regression. Therefore, the capacity to reject tumor acquired by tumor-reactive CD4(+) T cells largely depends on the direct suppressive activity of Tregs. We suggest that a cytotoxic CD4(+) T-cell immune response may be induced to enhance resistance against oncovirus-associated tumors.

Show MeSH
Related in: MedlinePlus