The Mechanism Of Apelin-13 Promoting Breast Cancer MCF-7 Cells Proliferation And Invasion

Apelin, which was ligand of G-protein-coupled receptor APJ, was a kind of natural endogenous active peptide. Apelin-13 was the main subtype in many subtypes and was widely expressed in various tissues and organs including breast, lung, heart, kidney. It has reported that apelin overexpression aggravated the progression of tumors by promoting tumour cells proliferation and neovascularization in many malignant tumors including hepatocellular, lung, ovarian, oral. However, the effect and molecular mechanism of apelin-13 was still unclear. Therefore, we performed a series of experiments to study the effect of apelin-13 on breast cancer.MTT assay and Brdu flow cytometry assay has proved that apelin-13 could promote MCF-7 cell proliferation in a concentration-dependent manner. In the process of cell proliferation, the G1 phase activation is the critical process. Cyclin D1 is the responsible molecular for cell cycle progression in the transition from G0/G1 to S phase. The over-expression of cyclin D1 was found in breast cancer. Therefore, the expression of cyclin D1 at protein and transcription level was detected by means of ELISA and RT-PCR. The result demonstrated apelin-13 could up-regulate cyclin D1 in a concentration-dependent manner.Normal cells morphology changes, prone to cross the surrounding dense connective tissue they turned cancerous. As a consequence, the invasion and metastasis of cancer cells are an important part in tumor progression. Matrix metalloproteinases MMP-1 is the major collagenases of degradation of extracellular matrix. Excessive MMP-1 expression is closely related to the treatment and prognosis of tumor. We have detected the effect of apelin-13 on MCF-7 cell invasion and MMP-1 expression using Transwell invasion assay, ELISA and RT-PCR. Transwell assay demonstrated that the cells that invaded through thepolycarbonate membrane were obviously increased with apelin-13 concentration enrichment. ELISA and RT-PCR results showed that apelin-13 raised MMP-1 expression at the protein level and transcription level. These results suggested that apelin-13 could promote MCF-7 cells invasion.Whether apelin-13 promoted breast cancer MCF-7 cell proliferation and invasion via ERK1/2 signal pathways, we measured the ERK1/2 phosphorylation by western blot. The result showed that apelin-13 increased ERK1/2 phosphorylation in a concentration dependent manner. After blocking ERK1/2 signal pathway, the effect of apelin-13 on ERK1/2 activation could be significantly inhibited. To further investigate the possible relationship of the effect of apelin-13 on the MCF-7 cells proliferation and invasion and ERK1/2 activation, we have detected MCF-7 cells proliferation and invasion by PD98059 prestimulated for 1h, then adding 10μM apelin-13. The result showed that compared with apelin-13 group, MCF-7 cells proliferation and invasion was significantly inhibited in PD98059 and apelin-13 group, which may be mediated by ERK1/2.Recent studies found that amplified in breast cancer 1(AIB1) is a potential carcinogenic factor with high expression in multiple tumors, particularly breast cancer. So we detected AIB1 m RNA expression using RT-PCR. The result showed that apelin increased AIB1 m RNA expression and PD98059 could suppress apelin-13 induced AIB1 m RNA high expression.The results above showed that apelin-13 increased the expression of cyclin D1 and MMP-1 and induced MCF-7 cells proliferation and migration via activated ERK1/2/AIB1 signaling pathways, which provided new ideas and methods for clinical diagnosis, treatment and prognostic assessment of breast cancer.