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Human Th17 cells can be induced through head and neck cancer and have a functional impact on HNSCC development.

Kesselring R, Thiel A, Pries R, Trenkle T, Wollenberg B - Br. J. Cancer (2010)

Bottom Line: In addition, tumour tissue and tumour-draining lymph nodes are infiltrated by a huge number of Th17 cells representing an important fraction of the tumour-infiltrating lymphocytes (TILs).We further showed that Th17 cells can be induced and expanded in tumour microenvironment through cytokines produced by tumour cells and TILs, and in addition can be recruited to the tumour milieu through a CCR6/CCL20-dependent mechanism.We conclude that Th17 cells have a substantial impact on the carcinogenesis of HNSCCs and on their metastasis and could serve as a potential therapeutic target to modulate anti-tumour response in HNSCC.

View Article: PubMed Central - PubMed

Affiliation: Department for Otorhinolaryngology, University of Luebeck, Ratzeburger Allee 160, Luebeck 23538, Germany.

ABSTRACT

Background: The T helper 17 (Th17) cells recently identified as distinct T helper cell lineage are characterised by their production of the proinflammatory cytokine interleukin 17. Although much effort has been made in understanding the function of Th17 cells in the pathogenesis of different diseases, their influence in carcinogenesis remain largely unknown.

Methods: We studied the prevalence and induction of Th17 cells in head and neck squamous cell carcinoma (HNSCC) patients by flow cytometry. To determine the migration mechanism of Th17 cells into primary tumours and metastasis of HNSCC, we performed chemotaxis assays. We analysed the proliferation and the angiogenesis-related proteins of HNSCCs in the presence of Th17 cells with MTT-based proliferation assay and an angiogenesis protein array.

Results: In this study, we showed that the prevalence of Th17 cells is elevated in peripheral blood of HNSCC patients. In addition, tumour tissue and tumour-draining lymph nodes are infiltrated by a huge number of Th17 cells representing an important fraction of the tumour-infiltrating lymphocytes (TILs). We further showed that Th17 cells can be induced and expanded in tumour microenvironment through cytokines produced by tumour cells and TILs, and in addition can be recruited to the tumour milieu through a CCR6/CCL20-dependent mechanism. Furthermore, we showed that the proliferation and angiogenesis of HNSCC are impaired in the presence of Th17 cells.

Conclusion: We conclude that Th17 cells have a substantial impact on the carcinogenesis of HNSCCs and on their metastasis and could serve as a potential therapeutic target to modulate anti-tumour response in HNSCC.

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Related in: MedlinePlus

Induction of Th17 cells in HNSCC microenvironment. (A) Expression of the Th-17-promoting cytokines IL-1β, -6 and -23 in tumour cells and TILs ex vivo analysed by flow cytometry. The bars show the mean of the positive cells±s.d. (B) Induction of Th17 cells in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days (n=30) with P<0.05. The cells were stained with CD3-FITC, CD4-PerCP, IL-17-APC and analysed by flow cytometry. The data is expressed as the frequency of Th17 cells in the CD4+ T cell population. The box plots show the median (middle line), 25th and 75th percentiles (box), the extreme values (whiskers, which indicate the 90th and 10th percentile) and the outliers (circles). (C) Blocking of Th17-inducing cytokines with blocking antibodies diminishes the Th17 induction through HNSCC tumourmilieu. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of blocking antibodies (n=15). First column cytokines are not blocked, columns 2–4 block one of the cytokines IL-1β, -6, -23, columns 5–7 block two of the Th17 cell-inducing cytokines, column 8 block all three cytokines. The error bars show the s.d. (D) PGE(2) contributes to Th17 cell induction in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of a PGE(2) blocking antibody (n=15). Blocking of PGE(2) (column 3) leads to a decrease in Th17 induction in HNSCC milieu. Blocking of IL-1β, -6, -23 and PGE(2) leads to a significant downregulation of Th17 cells compared with the number of Th17 cells induced through tumour supernatants alone. The error bars show the s.d.
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fig3: Induction of Th17 cells in HNSCC microenvironment. (A) Expression of the Th-17-promoting cytokines IL-1β, -6 and -23 in tumour cells and TILs ex vivo analysed by flow cytometry. The bars show the mean of the positive cells±s.d. (B) Induction of Th17 cells in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days (n=30) with P<0.05. The cells were stained with CD3-FITC, CD4-PerCP, IL-17-APC and analysed by flow cytometry. The data is expressed as the frequency of Th17 cells in the CD4+ T cell population. The box plots show the median (middle line), 25th and 75th percentiles (box), the extreme values (whiskers, which indicate the 90th and 10th percentile) and the outliers (circles). (C) Blocking of Th17-inducing cytokines with blocking antibodies diminishes the Th17 induction through HNSCC tumourmilieu. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of blocking antibodies (n=15). First column cytokines are not blocked, columns 2–4 block one of the cytokines IL-1β, -6, -23, columns 5–7 block two of the Th17 cell-inducing cytokines, column 8 block all three cytokines. The error bars show the s.d. (D) PGE(2) contributes to Th17 cell induction in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of a PGE(2) blocking antibody (n=15). Blocking of PGE(2) (column 3) leads to a decrease in Th17 induction in HNSCC milieu. Blocking of IL-1β, -6, -23 and PGE(2) leads to a significant downregulation of Th17 cells compared with the number of Th17 cells induced through tumour supernatants alone. The error bars show the s.d.

Mentions: We investigated whether the increased prevalence of Th17 cells in tumours and TDLNs is caused by an induction of Th17 cells or by the migration of Th17 cells towards the tumour site. The differentiation of Th17 cells in human beings is initiated and regulated by the cytokines IL-1β and -6 and their maintenance by IL-23 (Romagnani et al, 2009). Thus, to examine the first hypothesis, we determined the existence of these cytokines in HNSCC tumour milieu. The cytokines in tumour supernatants were determined with ELISA and multiplex assays. We found that HNSCC cell lines secrete the cytokines IL-6 and -23, but not IL-1β. In tumour supernatants of freshly derived tumours and TDLNs, we could detect all three cytokines IL-1β, -6 and -23 (data not shown). To evaluate which type of cells from the single cell suspensions secrete the corresponding cytokine, we determined the cytokines also directly in ex vivo derived tumour cells and tumour-infiltrating cells by flow cytometry. The tumour cells secrete the cytokines IL-23 (99%) and IL-6 (99%). The IL-1β is secreted only by TILs (26%), which produce IL-23 (29%) and IL-6 (32%) as well (Figure 3A).


Human Th17 cells can be induced through head and neck cancer and have a functional impact on HNSCC development.

Kesselring R, Thiel A, Pries R, Trenkle T, Wollenberg B - Br. J. Cancer (2010)

Induction of Th17 cells in HNSCC microenvironment. (A) Expression of the Th-17-promoting cytokines IL-1β, -6 and -23 in tumour cells and TILs ex vivo analysed by flow cytometry. The bars show the mean of the positive cells±s.d. (B) Induction of Th17 cells in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days (n=30) with P<0.05. The cells were stained with CD3-FITC, CD4-PerCP, IL-17-APC and analysed by flow cytometry. The data is expressed as the frequency of Th17 cells in the CD4+ T cell population. The box plots show the median (middle line), 25th and 75th percentiles (box), the extreme values (whiskers, which indicate the 90th and 10th percentile) and the outliers (circles). (C) Blocking of Th17-inducing cytokines with blocking antibodies diminishes the Th17 induction through HNSCC tumourmilieu. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of blocking antibodies (n=15). First column cytokines are not blocked, columns 2–4 block one of the cytokines IL-1β, -6, -23, columns 5–7 block two of the Th17 cell-inducing cytokines, column 8 block all three cytokines. The error bars show the s.d. (D) PGE(2) contributes to Th17 cell induction in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of a PGE(2) blocking antibody (n=15). Blocking of PGE(2) (column 3) leads to a decrease in Th17 induction in HNSCC milieu. Blocking of IL-1β, -6, -23 and PGE(2) leads to a significant downregulation of Th17 cells compared with the number of Th17 cells induced through tumour supernatants alone. The error bars show the s.d.
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fig3: Induction of Th17 cells in HNSCC microenvironment. (A) Expression of the Th-17-promoting cytokines IL-1β, -6 and -23 in tumour cells and TILs ex vivo analysed by flow cytometry. The bars show the mean of the positive cells±s.d. (B) Induction of Th17 cells in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days (n=30) with P<0.05. The cells were stained with CD3-FITC, CD4-PerCP, IL-17-APC and analysed by flow cytometry. The data is expressed as the frequency of Th17 cells in the CD4+ T cell population. The box plots show the median (middle line), 25th and 75th percentiles (box), the extreme values (whiskers, which indicate the 90th and 10th percentile) and the outliers (circles). (C) Blocking of Th17-inducing cytokines with blocking antibodies diminishes the Th17 induction through HNSCC tumourmilieu. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of blocking antibodies (n=15). First column cytokines are not blocked, columns 2–4 block one of the cytokines IL-1β, -6, -23, columns 5–7 block two of the Th17 cell-inducing cytokines, column 8 block all three cytokines. The error bars show the s.d. (D) PGE(2) contributes to Th17 cell induction in HNSCC microenvironment. T cells were incubated with tumoursupernatants from single cell suspension of tumour tissue for 7 days in the presence of a PGE(2) blocking antibody (n=15). Blocking of PGE(2) (column 3) leads to a decrease in Th17 induction in HNSCC milieu. Blocking of IL-1β, -6, -23 and PGE(2) leads to a significant downregulation of Th17 cells compared with the number of Th17 cells induced through tumour supernatants alone. The error bars show the s.d.
Mentions: We investigated whether the increased prevalence of Th17 cells in tumours and TDLNs is caused by an induction of Th17 cells or by the migration of Th17 cells towards the tumour site. The differentiation of Th17 cells in human beings is initiated and regulated by the cytokines IL-1β and -6 and their maintenance by IL-23 (Romagnani et al, 2009). Thus, to examine the first hypothesis, we determined the existence of these cytokines in HNSCC tumour milieu. The cytokines in tumour supernatants were determined with ELISA and multiplex assays. We found that HNSCC cell lines secrete the cytokines IL-6 and -23, but not IL-1β. In tumour supernatants of freshly derived tumours and TDLNs, we could detect all three cytokines IL-1β, -6 and -23 (data not shown). To evaluate which type of cells from the single cell suspensions secrete the corresponding cytokine, we determined the cytokines also directly in ex vivo derived tumour cells and tumour-infiltrating cells by flow cytometry. The tumour cells secrete the cytokines IL-23 (99%) and IL-6 (99%). The IL-1β is secreted only by TILs (26%), which produce IL-23 (29%) and IL-6 (32%) as well (Figure 3A).

Bottom Line: In addition, tumour tissue and tumour-draining lymph nodes are infiltrated by a huge number of Th17 cells representing an important fraction of the tumour-infiltrating lymphocytes (TILs).We further showed that Th17 cells can be induced and expanded in tumour microenvironment through cytokines produced by tumour cells and TILs, and in addition can be recruited to the tumour milieu through a CCR6/CCL20-dependent mechanism.We conclude that Th17 cells have a substantial impact on the carcinogenesis of HNSCCs and on their metastasis and could serve as a potential therapeutic target to modulate anti-tumour response in HNSCC.

View Article: PubMed Central - PubMed

Affiliation: Department for Otorhinolaryngology, University of Luebeck, Ratzeburger Allee 160, Luebeck 23538, Germany.

ABSTRACT

Background: The T helper 17 (Th17) cells recently identified as distinct T helper cell lineage are characterised by their production of the proinflammatory cytokine interleukin 17. Although much effort has been made in understanding the function of Th17 cells in the pathogenesis of different diseases, their influence in carcinogenesis remain largely unknown.

Methods: We studied the prevalence and induction of Th17 cells in head and neck squamous cell carcinoma (HNSCC) patients by flow cytometry. To determine the migration mechanism of Th17 cells into primary tumours and metastasis of HNSCC, we performed chemotaxis assays. We analysed the proliferation and the angiogenesis-related proteins of HNSCCs in the presence of Th17 cells with MTT-based proliferation assay and an angiogenesis protein array.

Results: In this study, we showed that the prevalence of Th17 cells is elevated in peripheral blood of HNSCC patients. In addition, tumour tissue and tumour-draining lymph nodes are infiltrated by a huge number of Th17 cells representing an important fraction of the tumour-infiltrating lymphocytes (TILs). We further showed that Th17 cells can be induced and expanded in tumour microenvironment through cytokines produced by tumour cells and TILs, and in addition can be recruited to the tumour milieu through a CCR6/CCL20-dependent mechanism. Furthermore, we showed that the proliferation and angiogenesis of HNSCC are impaired in the presence of Th17 cells.

Conclusion: We conclude that Th17 cells have a substantial impact on the carcinogenesis of HNSCCs and on their metastasis and could serve as a potential therapeutic target to modulate anti-tumour response in HNSCC.

Show MeSH
Related in: MedlinePlus