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CCL25-CCR9 interaction modulates ovarian cancer cell migration, metalloproteinase expression, and invasion.

Johnson EL, Singh R, Singh S, Johnson-Holiday CM, Grizzle WE, Partridge EE, Lillard JW - World J Surg Oncol (2010)

Bottom Line: OvCa tissue microarrays (TMA) was used to confirm CCR9 expression in clinical samples.Our results show significantly (p<0.001) higher expression of CCR9 by mucinous adenocarcinoma, papillary serous carcinoma, and endometriod ovarian carcinoma cases, than compared to non-neoplastic ovarian tissue.Expression of collagenases (MMP-1, -8, and -13), gelatinases (MMP-2 and -9), and stromelysins (MMP-3, -10, and -11) by OvCa cells were modulated by CCL25 in a CCR9-dependent fashion.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Biochemistry, & Immunology, Morehouse School of Medicine, 720 Westview Drive SW, Atlanta, GA 30310-1495, USA.

ABSTRACT

Background: Ovarian carcinoma (OvCa) is the most lethal gynecological malignancy among women and its poor prognosis is mainly due to metastasis. Chemokine receptor CCR9 is primarily expressed by a small subset of immune cells and its only natural ligand, CCL25, is largely expressed in the thymus, which involutes with age. Other than the thymus, CCL25 is expressed by the small bowel. Interactions between CCL25 and CCR9 have been implicated in leukocyte trafficking to the small bowel, a frequent metastatic site for OvCa cells. The current study shows OvCa tissue and cells significantly express CCR9, which interacts with CCL25 to support carcinoma cell migration and invasion.

Methods: RT-PCR and flow cytometry techniques were used to quantify the expression CCR9 by OvCa cells. OvCa tissue microarrays (TMA) was used to confirm CCR9 expression in clinical samples. The Aperio ScanScope scanning system was used to quantify immunohistochemical staining. Cell invasion and migration assays were performed using cell migration and matrigel invasion chambers. Matrix metalloproteinase (MMP) mRNAs were quantified by RT-PCR and active MMPs were quantified by ELISA.

Results: Our results show significantly (p<0.001) higher expression of CCR9 by mucinous adenocarcinoma, papillary serous carcinoma, and endometriod ovarian carcinoma cases, than compared to non-neoplastic ovarian tissue. Furthermore, CCR9 expression was significantly elevated in OvCa cell lines (OVCAR-3 and CAOV-3) in comparison to normal adult ovarian epithelial cell mRNA. OvCa cells showed higher migratory and invasive potential towards chemotactic gradients of CCL25, which was inhibited by anti-CCR9 antibodies. Expression of collagenases (MMP-1, -8, and -13), gelatinases (MMP-2 and -9), and stromelysins (MMP-3, -10, and -11) by OvCa cells were modulated by CCL25 in a CCR9-dependent fashion.

Conclusions: These results demonstrate both biological significance and clinical relevance of CCL25 and CCR9 interactions in OvCa cell metastasis.

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CCR9-mediated ovarian cancer cell migration and invasion. (A) OVCAR-3 and CAOV-3 cells were tested for their ability to migrate toward chemotactic gradients of CCL25. Cells were co-cultured with 1.0 μg/mL mouse anti-CCR9 antibody during migration assays using 100 ng/mL of CCL25. (B) OVCAR-3 and CAOV-3 cells were also tested for their ability to invade or translocate across Matrigel™ matrix in response to 100 ng/mL of CCL25. Cells were co-cultured with 1.0 μg/mL monoclonal antibodies against CCR9 during invasion assays using 100 ng/mL of CCL25. The number of cells ± SEM that migrated or invaded is shown with asterisk(s) that indicate significant differences (p < 0.01) between no additions and chemokine-induced cells.
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Figure 3: CCR9-mediated ovarian cancer cell migration and invasion. (A) OVCAR-3 and CAOV-3 cells were tested for their ability to migrate toward chemotactic gradients of CCL25. Cells were co-cultured with 1.0 μg/mL mouse anti-CCR9 antibody during migration assays using 100 ng/mL of CCL25. (B) OVCAR-3 and CAOV-3 cells were also tested for their ability to invade or translocate across Matrigel™ matrix in response to 100 ng/mL of CCL25. Cells were co-cultured with 1.0 μg/mL monoclonal antibodies against CCR9 during invasion assays using 100 ng/mL of CCL25. The number of cells ± SEM that migrated or invaded is shown with asterisk(s) that indicate significant differences (p < 0.01) between no additions and chemokine-induced cells.

Mentions: OvCa cell lines were tested for CCL25-dependent migration and invasion. CAOV-3 and OVCAR-3 cells significantly migrated to CCL25, compared to media without CCL25 (Figure 3). This CCL25-dependent chemotaxis was neutralized by anti-CCR9 antibody treatment, but not by the isotype control antibody. These findings demonstrated the functional expression of CCR9 by OvCa cells, which migrate to CCL25. CAOV-3 and OVCAR-3 differentially invaded Matrigel in response to CCL25. CAOV-3, but not OVCAR-3, cell lines significantly invaded through Matrigel in response to CCL25. As with migration responses, CCL25-mediated invasion was CCR9-dependent since cell lines treated with anti-CCR9 antibody behaved like controls. Interestingly, the differences in cell invasion did not correlate with CCR9 expression, because OVCAR-3 cell lines expressed significantly more CCR9 than CAOV-3 cells.


CCL25-CCR9 interaction modulates ovarian cancer cell migration, metalloproteinase expression, and invasion.

Johnson EL, Singh R, Singh S, Johnson-Holiday CM, Grizzle WE, Partridge EE, Lillard JW - World J Surg Oncol (2010)

CCR9-mediated ovarian cancer cell migration and invasion. (A) OVCAR-3 and CAOV-3 cells were tested for their ability to migrate toward chemotactic gradients of CCL25. Cells were co-cultured with 1.0 μg/mL mouse anti-CCR9 antibody during migration assays using 100 ng/mL of CCL25. (B) OVCAR-3 and CAOV-3 cells were also tested for their ability to invade or translocate across Matrigel™ matrix in response to 100 ng/mL of CCL25. Cells were co-cultured with 1.0 μg/mL monoclonal antibodies against CCR9 during invasion assays using 100 ng/mL of CCL25. The number of cells ± SEM that migrated or invaded is shown with asterisk(s) that indicate significant differences (p < 0.01) between no additions and chemokine-induced cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2927595&req=5

Figure 3: CCR9-mediated ovarian cancer cell migration and invasion. (A) OVCAR-3 and CAOV-3 cells were tested for their ability to migrate toward chemotactic gradients of CCL25. Cells were co-cultured with 1.0 μg/mL mouse anti-CCR9 antibody during migration assays using 100 ng/mL of CCL25. (B) OVCAR-3 and CAOV-3 cells were also tested for their ability to invade or translocate across Matrigel™ matrix in response to 100 ng/mL of CCL25. Cells were co-cultured with 1.0 μg/mL monoclonal antibodies against CCR9 during invasion assays using 100 ng/mL of CCL25. The number of cells ± SEM that migrated or invaded is shown with asterisk(s) that indicate significant differences (p < 0.01) between no additions and chemokine-induced cells.
Mentions: OvCa cell lines were tested for CCL25-dependent migration and invasion. CAOV-3 and OVCAR-3 cells significantly migrated to CCL25, compared to media without CCL25 (Figure 3). This CCL25-dependent chemotaxis was neutralized by anti-CCR9 antibody treatment, but not by the isotype control antibody. These findings demonstrated the functional expression of CCR9 by OvCa cells, which migrate to CCL25. CAOV-3 and OVCAR-3 differentially invaded Matrigel in response to CCL25. CAOV-3, but not OVCAR-3, cell lines significantly invaded through Matrigel in response to CCL25. As with migration responses, CCL25-mediated invasion was CCR9-dependent since cell lines treated with anti-CCR9 antibody behaved like controls. Interestingly, the differences in cell invasion did not correlate with CCR9 expression, because OVCAR-3 cell lines expressed significantly more CCR9 than CAOV-3 cells.

Bottom Line: OvCa tissue microarrays (TMA) was used to confirm CCR9 expression in clinical samples.Our results show significantly (p<0.001) higher expression of CCR9 by mucinous adenocarcinoma, papillary serous carcinoma, and endometriod ovarian carcinoma cases, than compared to non-neoplastic ovarian tissue.Expression of collagenases (MMP-1, -8, and -13), gelatinases (MMP-2 and -9), and stromelysins (MMP-3, -10, and -11) by OvCa cells were modulated by CCL25 in a CCR9-dependent fashion.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Biochemistry, & Immunology, Morehouse School of Medicine, 720 Westview Drive SW, Atlanta, GA 30310-1495, USA.

ABSTRACT

Background: Ovarian carcinoma (OvCa) is the most lethal gynecological malignancy among women and its poor prognosis is mainly due to metastasis. Chemokine receptor CCR9 is primarily expressed by a small subset of immune cells and its only natural ligand, CCL25, is largely expressed in the thymus, which involutes with age. Other than the thymus, CCL25 is expressed by the small bowel. Interactions between CCL25 and CCR9 have been implicated in leukocyte trafficking to the small bowel, a frequent metastatic site for OvCa cells. The current study shows OvCa tissue and cells significantly express CCR9, which interacts with CCL25 to support carcinoma cell migration and invasion.

Methods: RT-PCR and flow cytometry techniques were used to quantify the expression CCR9 by OvCa cells. OvCa tissue microarrays (TMA) was used to confirm CCR9 expression in clinical samples. The Aperio ScanScope scanning system was used to quantify immunohistochemical staining. Cell invasion and migration assays were performed using cell migration and matrigel invasion chambers. Matrix metalloproteinase (MMP) mRNAs were quantified by RT-PCR and active MMPs were quantified by ELISA.

Results: Our results show significantly (p<0.001) higher expression of CCR9 by mucinous adenocarcinoma, papillary serous carcinoma, and endometriod ovarian carcinoma cases, than compared to non-neoplastic ovarian tissue. Furthermore, CCR9 expression was significantly elevated in OvCa cell lines (OVCAR-3 and CAOV-3) in comparison to normal adult ovarian epithelial cell mRNA. OvCa cells showed higher migratory and invasive potential towards chemotactic gradients of CCL25, which was inhibited by anti-CCR9 antibodies. Expression of collagenases (MMP-1, -8, and -13), gelatinases (MMP-2 and -9), and stromelysins (MMP-3, -10, and -11) by OvCa cells were modulated by CCL25 in a CCR9-dependent fashion.

Conclusions: These results demonstrate both biological significance and clinical relevance of CCL25 and CCR9 interactions in OvCa cell metastasis.

Show MeSH
Related in: MedlinePlus