| Background and ObjectiveOvarian cancer is one of the most common malignancy of the female reproductive system. It is lack of early obvious symptoms and highly sensitive screening method which is make it difficult to diagnose and detective early. Because 70% of patients diagnosed at advanced stage, so the prognosis is poor,5-year survival rate is only about 30% which the mortality rate is at the head of gynecological malignancies.In the development of tumor, the growth factor-mediated intracellular signal transduction pathways, has been a hot spot in the field of cancer research at home and abroad. Hepatocyte growth factor (HGF) is currently the most widely known one biologically active growth factor in a number of factors. HGF is a polypeptide growth factor, after the combine with c-Met receptor, causing a series of intracellular signal transduction via phosphorylation of tyrosine residues, lead to some downstream expression and biological effects of specific molecules. Poly ADP ribose polymerase-1(PARP-1) is a highly conserved ribozyme in most eukaryotic cells of which is play an important role in a variety of biological effects, such as maintaining the integrity of DNA, gene transcription, cell cycle, chromosome function, genome stability and cell death and so on. The study found that a variety of mechanisms growth factors, oncogenes and tumor suppressor genes regulating the expression of PARP-1 cells. Currently, there is no coverage at home and abroad on HGF and PARP-1 expression in ovarian cancer and the role of PARP-1 in HGF-induced ovarian cancer cell invasion and metastasis. This study was designed to investigate the effects of HGF on SKOV-3 cells, PARP-1 expression levels, and HGF/PARP-1 pathway in the regulation of SKOV-3 cell invasion and metastasis, which provide a new theoretical basis for the study of invasion and metastasis and its targeted therapies.Methods1. Cell culture:Select logarithmic growth phase cells. Cell groups:(1) control group: Cultured with RPIM-1640 medium which is contain 10% fetal bovine serum without HGF; (2) HGF stimulation group:adding an appropriate amount of recombinant human HGF, so that the final concentration of the culture medium HGF were 10 ng/mL,20 ng/mL,40 ng/mL,60 ng/mL,80 ng/mL; stimulation time was 6h,24h,48h.2. SKOV-3 cells incubated with increasing concentrations of HGF to examine their invasive abilities by Transwell test.3. Relative expression of PARP-1 after HGF treatment in SKOV-3 cells analyzed by RT-PCR.4. Relative expression of PARP-1 after HGF treatment in SKOV-3 cells analyzed by western blot.5. SKOV3 cells were transfected with either negative control siRNA (NC-siRNA group) or PARP-1 siRNA(PARPl-siRNA group), and divided into different groups depengding on different factors:blank control group; HGF group; PARP-siRNA group; HGF+PARP-siRNA group; NC-siRNA group; HGF+NC-siRNA group.6. RT-PCR was used to measure the expression of PARP-1 in different groups.7. western blotting was used to measure the expression of PARP-1 in different groups.8. Transwell test examine their invasive abilities.9. ELISA was used to measure the invasion of cells and the expression of MMP-2.Results1. HGF promotes cell invasion in a concentration-and-time-dependent manners in the SKOV-3 cell line.2. The expression of PARP-1 in SKOV-3 cells were increased after administered with HGF 40ng/ml for 24 hours.3. The expression of PARP-1 in PARP1-siRNA group were lower compared with the NC-siRNA group (P<0.05) 。4. The transfection significantly reduced the impact of HGF on SKOV-3 cell invasion and the expression of MMP-2.ConclusionHGF promotes the invasion and metastasis of Ovarian Cancer; and this effect may be related to the increased expression of MMP-2 which is mediated by PARP-1. |
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