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The chemokine CXCL16 and its receptor, CXCR6, as markers and promoters of inflammation-associated cancers.

Darash-Yahana M, Gillespie JW, Hewitt SM, Chen YY, Maeda S, Stein I, Singh SP, Bedolla RB, Peled A, Troyer DA, Pikarsky E, Karin M, Farber JM - PLoS ONE (2009)

Bottom Line: Screening for 37 chemokines in prostate cancer cell lines and xenografts revealed CXCL16, the ligand for the receptor CXCR6, as the most consistently expressed chemokine.Immunohistochemistry and/or immunofluorescence and confocal imaging of 121 human prostate specimens showed that CXCL16 and CXCR6 were co-expressed, both on prostate cancer cells and adjacent T cells.We studied expression of CXCL16 in an additional 461 specimens covering 12 tumor types, and found that CXCL16 was expressed in multiple human cancers associated with inflammation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Bethesda, MD, USA. meravd@hadassah.org.il

ABSTRACT
Clinical observations and mouse models have suggested that inflammation can be pro-tumorigenic. Since chemokines are critical in leukocyte trafficking, we hypothesized that chemokines play essential roles in inflammation-associated cancers. Screening for 37 chemokines in prostate cancer cell lines and xenografts revealed CXCL16, the ligand for the receptor CXCR6, as the most consistently expressed chemokine. Immunohistochemistry and/or immunofluorescence and confocal imaging of 121 human prostate specimens showed that CXCL16 and CXCR6 were co-expressed, both on prostate cancer cells and adjacent T cells. Expression levels of CXCL16 and CXCR6 on cancer cells correlated with poor prognostic features including high-stage and high-grade, and expression also correlated with post-inflammatory changes in the cancer stroma as revealed by loss of alpha-smooth muscle actin. Moreover, CXCL16 enhanced the growth of CXCR6-expressing cancer and primary CD4 T cells. We studied expression of CXCL16 in an additional 461 specimens covering 12 tumor types, and found that CXCL16 was expressed in multiple human cancers associated with inflammation. Our study is the first to describe the expression of CXCL16/CXCR6 on both cancer cells and adjacent T cells in humans, and to demonstrate correlations between CXCL16 and CXCR6 vs. poor both prognostic features and reactive changes in cancer stoma. Taken together, our data suggest that CXCL16 and CXCR6 may mark cancers arising in an inflammatory milieu and mediate pro-tumorigenic effects of inflammation through direct effects on cancer cell growth and by inducing the migration and proliferation of tumor-associated leukocytes.

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Related in: MedlinePlus

CXCL16 can mediate the proliferation and migration of CD3-activated primary CD4 T cells.(A) CD4+ T cells purified from elutriated lymphocytes from healthy donors were stimulated for 3 days with plate-bound anti-CD3 (OKT3, 10 µg/ml) with or without 5 µg/ml plate-bound CXCL16 (bL16). Treatments of anti-CD3-activated cells with PTX, anti-CXCR6 (aR6) and anti-CXCL16 (aL16) without bL16 were done as controls. Mouse IgG (mIgG) and rat IgG (rIgG) were used as controls for anti-CXCR6 antibody and anti-CXCL16 antibody, respectively. Bars show means±SEM from one representative experiment out of five, using five donors. Anti-CXCL16 was used in a total of four, and anti-CXCR6 in two experiments. ns, not significant and ***, p<0.001. (B) CXCR6 and CXCL16 mRNAs were measured by real-time RT-PCR in CD3-activated and non-activated CD4+ T cells after 3 days. Values were normalized by setting the non-activated sample with lowest expression equal to 1. Bars show means±SEM combined from duplicate wells from seven different donors. **, p<0.01 and ***, p<0.001 vs. non-activated cells. (C) Anti-CD3-activated or nonactivated CD4+ T cells were analyzed for migration to CXCL16, expressed as percentages of input cells migrating. Each bar represents the mean±SEM obtained from triplicate wells from a total of three experiments performed. ***, p<0.001 on cross bars are indicated for comparisons between activated cells - 0 vs. 50 ng/ml CXCL16 and 50 ng/ml vs. 500 ng/ml CXCL16.
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pone-0006695-g006: CXCL16 can mediate the proliferation and migration of CD3-activated primary CD4 T cells.(A) CD4+ T cells purified from elutriated lymphocytes from healthy donors were stimulated for 3 days with plate-bound anti-CD3 (OKT3, 10 µg/ml) with or without 5 µg/ml plate-bound CXCL16 (bL16). Treatments of anti-CD3-activated cells with PTX, anti-CXCR6 (aR6) and anti-CXCL16 (aL16) without bL16 were done as controls. Mouse IgG (mIgG) and rat IgG (rIgG) were used as controls for anti-CXCR6 antibody and anti-CXCL16 antibody, respectively. Bars show means±SEM from one representative experiment out of five, using five donors. Anti-CXCL16 was used in a total of four, and anti-CXCR6 in two experiments. ns, not significant and ***, p<0.001. (B) CXCR6 and CXCL16 mRNAs were measured by real-time RT-PCR in CD3-activated and non-activated CD4+ T cells after 3 days. Values were normalized by setting the non-activated sample with lowest expression equal to 1. Bars show means±SEM combined from duplicate wells from seven different donors. **, p<0.01 and ***, p<0.001 vs. non-activated cells. (C) Anti-CD3-activated or nonactivated CD4+ T cells were analyzed for migration to CXCL16, expressed as percentages of input cells migrating. Each bar represents the mean±SEM obtained from triplicate wells from a total of three experiments performed. ***, p<0.001 on cross bars are indicated for comparisons between activated cells - 0 vs. 50 ng/ml CXCL16 and 50 ng/ml vs. 500 ng/ml CXCL16.

Mentions: Moreover, when we examined the effect of plate-bound CXCL16 on the proliferation of CD3-activated primary CD4 T cells, we found a significant stimulatory effect on proliferation that was inhibited by treating with pertussis toxin or antibodies against CXCR6 or CXCL16 (Figure 6A). It is notable, given that CXCL16 is synthesized as a transmembrane chemokine, that although plate-bound CXCL16 was stimulatory, soluble CXCL16 had no effect (data not shown).


The chemokine CXCL16 and its receptor, CXCR6, as markers and promoters of inflammation-associated cancers.

Darash-Yahana M, Gillespie JW, Hewitt SM, Chen YY, Maeda S, Stein I, Singh SP, Bedolla RB, Peled A, Troyer DA, Pikarsky E, Karin M, Farber JM - PLoS ONE (2009)

CXCL16 can mediate the proliferation and migration of CD3-activated primary CD4 T cells.(A) CD4+ T cells purified from elutriated lymphocytes from healthy donors were stimulated for 3 days with plate-bound anti-CD3 (OKT3, 10 µg/ml) with or without 5 µg/ml plate-bound CXCL16 (bL16). Treatments of anti-CD3-activated cells with PTX, anti-CXCR6 (aR6) and anti-CXCL16 (aL16) without bL16 were done as controls. Mouse IgG (mIgG) and rat IgG (rIgG) were used as controls for anti-CXCR6 antibody and anti-CXCL16 antibody, respectively. Bars show means±SEM from one representative experiment out of five, using five donors. Anti-CXCL16 was used in a total of four, and anti-CXCR6 in two experiments. ns, not significant and ***, p<0.001. (B) CXCR6 and CXCL16 mRNAs were measured by real-time RT-PCR in CD3-activated and non-activated CD4+ T cells after 3 days. Values were normalized by setting the non-activated sample with lowest expression equal to 1. Bars show means±SEM combined from duplicate wells from seven different donors. **, p<0.01 and ***, p<0.001 vs. non-activated cells. (C) Anti-CD3-activated or nonactivated CD4+ T cells were analyzed for migration to CXCL16, expressed as percentages of input cells migrating. Each bar represents the mean±SEM obtained from triplicate wells from a total of three experiments performed. ***, p<0.001 on cross bars are indicated for comparisons between activated cells - 0 vs. 50 ng/ml CXCL16 and 50 ng/ml vs. 500 ng/ml CXCL16.
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pone-0006695-g006: CXCL16 can mediate the proliferation and migration of CD3-activated primary CD4 T cells.(A) CD4+ T cells purified from elutriated lymphocytes from healthy donors were stimulated for 3 days with plate-bound anti-CD3 (OKT3, 10 µg/ml) with or without 5 µg/ml plate-bound CXCL16 (bL16). Treatments of anti-CD3-activated cells with PTX, anti-CXCR6 (aR6) and anti-CXCL16 (aL16) without bL16 were done as controls. Mouse IgG (mIgG) and rat IgG (rIgG) were used as controls for anti-CXCR6 antibody and anti-CXCL16 antibody, respectively. Bars show means±SEM from one representative experiment out of five, using five donors. Anti-CXCL16 was used in a total of four, and anti-CXCR6 in two experiments. ns, not significant and ***, p<0.001. (B) CXCR6 and CXCL16 mRNAs were measured by real-time RT-PCR in CD3-activated and non-activated CD4+ T cells after 3 days. Values were normalized by setting the non-activated sample with lowest expression equal to 1. Bars show means±SEM combined from duplicate wells from seven different donors. **, p<0.01 and ***, p<0.001 vs. non-activated cells. (C) Anti-CD3-activated or nonactivated CD4+ T cells were analyzed for migration to CXCL16, expressed as percentages of input cells migrating. Each bar represents the mean±SEM obtained from triplicate wells from a total of three experiments performed. ***, p<0.001 on cross bars are indicated for comparisons between activated cells - 0 vs. 50 ng/ml CXCL16 and 50 ng/ml vs. 500 ng/ml CXCL16.
Mentions: Moreover, when we examined the effect of plate-bound CXCL16 on the proliferation of CD3-activated primary CD4 T cells, we found a significant stimulatory effect on proliferation that was inhibited by treating with pertussis toxin or antibodies against CXCR6 or CXCL16 (Figure 6A). It is notable, given that CXCL16 is synthesized as a transmembrane chemokine, that although plate-bound CXCL16 was stimulatory, soluble CXCL16 had no effect (data not shown).

Bottom Line: Screening for 37 chemokines in prostate cancer cell lines and xenografts revealed CXCL16, the ligand for the receptor CXCR6, as the most consistently expressed chemokine.Immunohistochemistry and/or immunofluorescence and confocal imaging of 121 human prostate specimens showed that CXCL16 and CXCR6 were co-expressed, both on prostate cancer cells and adjacent T cells.We studied expression of CXCL16 in an additional 461 specimens covering 12 tumor types, and found that CXCL16 was expressed in multiple human cancers associated with inflammation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Bethesda, MD, USA. meravd@hadassah.org.il

ABSTRACT
Clinical observations and mouse models have suggested that inflammation can be pro-tumorigenic. Since chemokines are critical in leukocyte trafficking, we hypothesized that chemokines play essential roles in inflammation-associated cancers. Screening for 37 chemokines in prostate cancer cell lines and xenografts revealed CXCL16, the ligand for the receptor CXCR6, as the most consistently expressed chemokine. Immunohistochemistry and/or immunofluorescence and confocal imaging of 121 human prostate specimens showed that CXCL16 and CXCR6 were co-expressed, both on prostate cancer cells and adjacent T cells. Expression levels of CXCL16 and CXCR6 on cancer cells correlated with poor prognostic features including high-stage and high-grade, and expression also correlated with post-inflammatory changes in the cancer stroma as revealed by loss of alpha-smooth muscle actin. Moreover, CXCL16 enhanced the growth of CXCR6-expressing cancer and primary CD4 T cells. We studied expression of CXCL16 in an additional 461 specimens covering 12 tumor types, and found that CXCL16 was expressed in multiple human cancers associated with inflammation. Our study is the first to describe the expression of CXCL16/CXCR6 on both cancer cells and adjacent T cells in humans, and to demonstrate correlations between CXCL16 and CXCR6 vs. poor both prognostic features and reactive changes in cancer stoma. Taken together, our data suggest that CXCL16 and CXCR6 may mark cancers arising in an inflammatory milieu and mediate pro-tumorigenic effects of inflammation through direct effects on cancer cell growth and by inducing the migration and proliferation of tumor-associated leukocytes.

Show MeSH
Related in: MedlinePlus