Expression of SDF-1 and CXCR4 transcript variants and CXCR7 in epithelial ovarian cancer

Abstract

Chemokine stromal cell-derived factor-1 (SDF-1) and its receptors, CXCR4 and CXCR7, have been implicated in epithelial ovarian cancer progression and metastasis. However, limited data are available on the expression levels of SDF-1 and CXCR4 variants and CXCR7 in human epithelial ovarian cancer. The present study aimed to characterize the expression pattern and levels of SDF-1, CXCR4 and CXCR7 in normal human ovaries and epithelial ovarian cancer. The expression of SDF-1 and CXCR4 transcript variants and CXCR7 was determined by quantitative polymerase chain reaction (qPCR). Plasma SDF-1α levels were determined by commercially available EIA kits and cancer antigen 125 (CA 125) levels were quantified by automated microparticle enzyme immunosorbent assay. High expression levels of SDF-1 transcript variant 1 were identified in ovarian cancer and control ovaries. By contrast, in both groups the expression levels of SDF-1 transcript variants 3 and 4 were extremely low. Furthermore, SDF-1 variant 1 levels were notably higher in epithelial ovarian cancer than in control ovaries, while data for the remaining transcripts were similar in both groups. CXCR4 transcript variant 2 and CXCR7 expression levels in normal and neoplastic ovaries were similar. In both groups, CXCR4 transcript variant 2 was not detected. Plasma SDF-1α levels were notably higher in females with epithelial ovarian cancer than in the control ovaries. Elevated levels of blood SDF-1α were found prior to surgery, 6 days after surgery and following completion of the first chemotherapy course. These increases were independent of the type of epithelial ovarian cancer. Our results suggest that the expression of SDF-1 and the genes controlling alternative splicing are elevated in epithelial ovarian cancer, leading to an increased formation of SDF-1 variant 1. Elevated plasma SDF-1α levels in epithelial ovarian cancer patients are not associated with the presence of tumors and/or metastases, however reflect a general response to the disease.

Keywords: CXCR4; CXCR7; SDF-1; epithelial ovarian cancer; plasma CA 125; plasma SDF-1α; transcript variants.

Figure 1

Ethidium bromide-stained 2% agarose gel demonstrating cDNA amplified with specific primer from RNA of (A) three ovarian cancers and (B) three control ovaries. Lane 1, size marker (O’Range Ruler 50-bp DNA Ladder; MBI Fermentas, Lithuania); lanes 2–4, SDF-1, transcript variant 1; lanes 5–7, SDF-1, transcript variant 2; lanes 8–10, SDF-1, transcript variant 3; lanes 11–13, SDF-1, transcript variant 4; lanes 14–16, CXCR4, transcript variant 1; lanes 17–19, CXCR4, transcript variant 2; lanes 20–22, CXCR7. In the control and neoplastic ovaries, no reaction product for CXCR4, transcript variant 1 was found. For the remaining genes, reaction products with expected size were observed. SDF-1, stromal cell-derived factor-1.

Figure 2

Quantitative polymerase chain reaction assay of SDF-1, transcript variants 1–4 mRNA expression in ovarian cancer (n=27) and in control ovaries (n=13). SDF-1, transcript variants 1–4 mRNA expression levels were correlated with the expression levels of the mitochondrial ribosomal protein L19 reference gene. Bars represent the mean ± SE. Statistically significant difference in relation to the control group (Mann-Whitney U test): ^****P<0.001. SDF-1, stromal cell-derived factor-1..

Figure 3

Quantitative polymerase chain reaction assay of CXCR7 and CXCR4, transcript variant 2 mRNA expression in ovarian cancer (n=27) and in control ovaries (n=13). The total number of cases for CXCR7 was six. Bars represent the relative expression of CXRX7 and CXRX4 variant 2 genes in relation to the mitochondrial ribosomal protein L19 reference gene. Results are expressed as the means ± SE. .In the control ovaries and epithelial ovarian cancer, CXCR4, transcript variant 1 mRNA expression was not found, while in placenta (lanes 1–3) the signal was well detected indicating proper structure of the primer.

Figure 4

Plasma SDF-1α concentrations (ng/ml) in females with ovarian cancer prior to and after surgery (day 6), and after completion of the first chemotherapy course. The data for 43 cases are shown. The control group includes 30 females. Bars represent the mean ± SE. Statistically significant difference in relation to the control group (Wilcoxon tests): ^*P<0.05, ^**P<0.01 and ^***P<0.001. SDF-1, stromal cell-derived factor-1.

Figure 5

Plasma SDF-1α concentrations (ng/ml) in females with serous and other ovarian cancer prior to and after surgery (day 6), and after completion of the first chemotherapy course. The cases of epithelial ovarian cancer consisted of 29 patients classified as serous and 14 cases classified as other. Bars represent the mean ± SE. SDF-1, stromal cell-derived factor-1.

Figure 6

Plasma SDF-1 α concentrations (ng/ml) in females with epithelial ovarian cancer prior to and after (day 6) surgery and after completion of the first chemotherapy course, in relation to FIGO classification. FIGO I + II, n=9; FIGO III + IV, n=34. Bars represent the mean ± SE. SDF-1, stromal-derived factor-1; FIGO, International Federation of Gynecology and Obstetrics.

Figure 7

Plasma SDF-1α (ng/ml) and CA 125 (U/ml) concentrations in patients in relation to FIGO classifications for staging ovarian cancer. Open bars represent FIGO I + II (n=9), black bars represent FIGO III + IV (n=34). Bars represent the mean ± SE. Statistically significant difference in relation to FIGO I + II group (Mann-Whitney test): ^****P<0.001. SDF-1, stromal cell-derived factor-1; CA 125, cancer antigen 125; FIGO, International Federation of Gynecology and Obstetrics.

Figure 8

Plasma SDF-1α (ng/ml) and CA 125 (U/ml) concentrations in ovarian cancer patients in relation to ascites. Open bars represent ascites (n=27), black bars represent without ascites (n=34). Bars represent the mean + SE. Statistically significant difference in relation to ascites group (Mann-Whitney test): ^****P<0.001. SDF-1, stromal cell-derived factor-1; CA 125, cancer antigen 125.