The Effect Of Inhibitors Of Signal Transduction Pathway PD98059and LY294002on Proliferation, Invasiveness Of Ovarian Carcinoma Cell Interfered By SDF-1

Background and ObjectiveStromal cell-derived factorl is a chemotactic protein secreted by bone marrowstromal cells and other related mesenchymal cells and epithelial cells, system namedCXCL12. Chemokine receptor4is the main receptor of SDF-1. SDF-1can speciallycombine with CXCR4, then creat SDF-1/CXCR4biology axle. This axle participatein many pathological and physiological processes. Many researches have certificateSDF-1/CXCR4is related closely with tumor proliferation, invasiveness, metastasis. Thestructure and function mechanism is the basement to bring pathological andphysiological function into play. Dspite it was reported that SDF-1、CXCR4arehigher expressed in ovarian carcinoma issue and cell, and find SDF-1can promote theproliferation, invasive, metastasize capacity of ovarian carcinoma, but the mechanismof SDF-1/CXCR4acting on proliferation, invasive, metastasize capacity of ovariancarcinoma is still unclear. The active of signal transduction pathway is an early affairof cancer happening. Phosphatidylinositol3kinase/protein kinase B(PI3K/AKT)signal transduction pathway is an emotion inter-cell sigal pathway.Mitogell activated proteinkinase/extracellular regulated protein kinases(MEK/ERK) is an emotion exter-cell signal transduction factor. Our experiment mainly study thepromoting effect of SDF-1to proliferation, invasive capacity of ovarian carcimomaweather can be blocked by MEK inhibitor PD98059, AKT inhibitor LY294002, thento explore the relation between SDF-1/CXCR4bilogy axle and MEK/ERK,PI3K/AKT signal pathway.Methods1. Culturing ovarian cancer cell lines SKOV3by12%fetal bovine serum inRPMI-1640culture medium, placed in5%CO2,37℃, the cells were incubated in ahumidified incubator.2. The expression of P-AKT and P-ERK were detected by Immunocytochemistry.3. Using monotetrazoliμm(MTT) assay to detect the difference of opticaldensity(OD492) when were added100ng/ml SDF-1、neutralizing antibody of CXCR4、inhibitors of CXCR4AMD3100、the inhibitors of signal transduction pathwayLY294002、PD98059to SKOV3cells.4. Using transwell invasion trial to detect the number of cell-penetrating when wereadded100ng/ml SDF-1、neutralizing antibody of CXCR4、inhibitors of CXCR4AMD3100、 the inhibitors of signal transduction pathway LY294002、PD98059to SKOV3cells.5. We use SPSS16.0statistical software to handle all data statistics. Ordinal ranksum test was used to compare the qualitative variable. x±s,one-way ANOVA andLSD-t are used to compare the quantitative data.=0.05, P<0.05was considersignificant.Result1. P-ERK protein mainly expressed in cytoplasm and cell nucleus of SKOV3,especially in cell nucleus. P-AKT mainly expressed in cytoplasm of ovariancarcinoma, finding brown and yellow particulate under microscope. According to thepositive cell and the degree of dyeing, with the acting time longer, the colour ofparticulate is changing from light to heavy,and the difference is signifficant（P-ERK:χ2=24.011,P=0.000; P-AKT: χ2=16.408, P=0.003）. Besides, when the acting time is30min and60min, the dyeing level of brown and yellow particulate is similar, nosignificant difference（P-AKT: χ2=4.835, P=0.089; P-ERK: χ2=1.725，P=0.422), so the best acting time is30min. When the acting time is30min, the dyeing level ofP-AKT, P-ERK rise with the growth of SDF-1concentration, higher concertration,higher express level, and the difference is signifficant（P-ERK: χ2=8.467, P=0.015；P-AKT: χ2=8.345, P=0.015）.2. Compared with no serum group, the OD492of SDF-1group is obviously elevated（d=-0.12378，P=0.007）, but when added with10μg/mlCXCR4neutralizingantibody (d=0.12956, P=0.005)、1μg/mlCXCR4inhibitors AMD3100（d=0.13900,P=0.003）, compared with SDF-1group, there are no obvious elevation of OD492.Besides, when added with50μmol/L LY294002(d=0.10978, P=0.025)、50μmol/LPD98059（d=0.11594, P=0.014）, compared with SDF-1group, there are no obviouselevation of OD492too.3. Compared with no serum group（23.1±10.64007on average）, the number ofcell-penetrating of SDF-1group（70.2±9.22316on average） is obviously increasedand the difference is signifficant （d=-47.1,P=0.000）, but when added with10μg/mlCXCR4neutralizing antibody(27.7±4.73873on average)、1μg/mlCXCR4inhibitors AMD3100（30.3±6.25478on average）, compared with SDF-1group, thereare no obvious increasement of the number of cell-penetrating. Besides, when addedwith50μmol/L PD98059（20.7±4.34741on average）、50μmol/L LY294002（24.3±3.59166on average）, compared with SDF-1group, there are no obviousincreasement of the number of cell-penetrating too.ConclusionSDF-1can promote the proliferation,invasive,metastasize capacity of ovariancarcinoma, but this function can be blocked and neutralized MEK inhibitor PD98059,AKT inhibitor LY294002. Therefore, SDF-1/CXCR4may play its functionsthrough Ras/ERK and PI3K/Akt signaling pathway.