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. 2015 Apr;9(4):1772-1778.
doi: 10.3892/ol.2015.2958. Epub 2015 Feb 12.

Cancer and embryo expression protein 65 promotes cancer cell growth and metastasis

Affiliations

Cancer and embryo expression protein 65 promotes cancer cell growth and metastasis

Genglin Jin et al. Oncol Lett. 2015 Apr.

Abstract

Cancer and embryo expression protein 65 (CEP65) is a centrosomal protein that is expressed at relatively high levels in embryonic tissue and different cancerous tissues, but its role in tumorigenesis remains unknown. In the present study, CEP65 was stably expressed in AGS gastric cancer cells. CEP65 was found to promote cell growth in the MTT assay and to enhance cell migration and invasion in Transwell chamber assays. To validate results from the in vitro experiments, CEP65 was stably expressed in BICR-H1 breast cancer cells through adenovirus-mediated transduction. By inoculating BICR-H1 cells on chick chorioallantoic membrane (CAM), it was found that CEP65 promotes cell growth on the CAM and increases cell metastasis to the lungs of the chicken. By utilizing a xenograft severe combined immunodeficiency mouse model, CEP65 was also found to accelerate BICR-H1 cell growth and metastasis to the lungs. Furthermore, it was shown that CEP65 increases matrix metalloproteinase (MMP)2 activity in zymographic assays, however, microarray screening and reverse transcription polymerase chain reaction validation revealed that CEP65 had no effect on the expression levels of MMP2 or MMP9, but decreased the expression levels of metastasis-associated genes, TIMP2, RAP and VTN. Taken together, the results of the present study demonstrated the oncogenic function of CEP65 in promoting cancer cell growth and metastasis.

Keywords: cancer and embryo expression protein 65; growth; invasion; metastasis; migration.

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Figures

Figure 1
Figure 1
CEP65 promotes AGS cell growth and invasiveness in vitro. (A) Expression of CEP65 in transfected AGS cells, as detected by RT-PCR and western blot analysis. (B) CEP65 promotes the growth of AGS cells. An MTT assay was performed and proliferation curves were generated by measuring the optical density (OD) at 490 nm at the indicated time-points after plating. (C) CEP65 decreases the adhesion of the AGS cells to the Matrigel. Data represent the results from three independent determinations. (D) CEP65 promotes the migration of the AGS cells. Cells were allowed to migrate for 12 h after plating. (E) CEP65 promotes the invasion of the AGS cells. Cells were allowed to migrate for 16 h after plating. *P<0.05 and **P<0.01 vs. AGS; #P<0.05 and ##P<0.01 vs. AGS-M. CEP65, cancer and embryo expression protein 65; GAPDH, glyceraldehyde 3-phosphate; AGS-M, AGS mock-transfected cells.
Figure 2
Figure 2
CEP65 promotes BICR-H1 cells growth and metastasis in vivo. (A) Expression of CEP65 in adenovirus-infected BICR-H1 cells, as detected by western blot analysis. (B) CEP65 promotes tumor formation on chick chorioallantoic membranes: Left image, macroscopic image of tumors; right image, graph summarizing the weight of the tumors. (C) CEP65 promotes BICR-H1 metastasis to the lungs of the chicken: Left, metastatic foci in the chicken lungs were detected using fluorescence microscopy; right, graph summarizing the numbers of metastatic foci. (D) CEP65 promotes tumor formation in SCID mice: Left, macroscopic image of tumors; graph summarizing the weight of the tumors. (E) CEP65 promotes BICR-H1 metastasis to the lungs of SCID mice, as detected by hematoxylin and eosin staining. *P<0.05 vs. pAd. CEP65, cancer and embryo expression protein 65; SCID, severe combined immunodeficiency.
Figure 3
Figure 3
CEP65 promotes MMP2 activity and decreases expression levels of metastasis-related genes. (A) CEP65 promotes MMP2 activity. Serum-free conditioned media from indicated cells were subjected to zymographic assay. Following removal of the SDS, the gelatin-containing gel was incubated in developing buffer and then visualized by Coomassie blue staining (upper panel). Relative activities of MMP2 and MMP9 were calculated (lower panel). (B) Effect of CEP65 on gene expression, as detected by reverse transcription polymerase chain reaction. **P<0.01 vs. AGS; ##P<0.01 vs. AGS-M. CEP65, cancer and embryo expression protein 65; MMP, matrix metalloproteinase; GAPDH, glyceraldehyde 3-phosphate; AGS-M, AGS mock-transfected cells.

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