Optimal Concentration Of SPIO-ShRNA Transfected Into Ovarian Cancer Cells And The Effect Of Magnetic Field On The Transfection Efficiency In Vitro

Part1Objective:To explore the effects of SPIO-ShRNA dual functional molecular probe of different concentration on morphology and EGFR expression of ovarian cancer SKOV3 cells in vitro.Methods:The dual functional molecular probes at an iron concentration of 5mg/L,15mg/L,30mg/L,45mg/L,75mg/L, 100mg/L were transfected into SKOV3 cells. The transfection rate of probe was observed by fluorescence microscope. The distribution and content of iron particles in SKOV3 cells were determined by Prussian blue staining, atomic adsorption spectrometer and electron microscopy. Cell viability was observed by CCK-8.The apoptosis was detected by flow cytometry. The expression of protein within the cells was detected by Western-blot. The changes of signal intensity were measured by magnetic resonance imaging (MRI).Results:SPIO-ShRNA dual functional molecular probe was uptake in a concentration-dependence manner within a certain rang (5mg/L-30mg/L). When concentration of the probe was 45mg/L, the labeling rate of the cell closed to 100%,cell viability and EGFR protein expression were significantly decreased (all P<0.01), the signal intensity of T2*WI was decreased obviously (P<0.01).Conclusion:The dual functional molecular probe can effectively transfect and specifically inhibit the expression of SKOV3 cell lines at the iron concentration of 45mg/L, and it also can be detected by magnetic resonance imaging, dual functional molecular probe has been initially confirmed the role of diagnosis and treatment.Part2Objective:To explore the transfection rate of SPIO-shRNA dual functional molecular probe into ovarian carcinoma SKOV3 cells in external magnetic fieldMethods:Dual functional molecular probe at an iron concentration of 45mg/L was transfected into SKOV3 cells, probe and magnetic fields existed at the same time as the experimental group,only probe transfected cells as negative control group and normal cultured SKOV3 without any transfection as blank control group.The transfection rate was detected by flow cytometry. Cell viability was observed by CCK-8 assay.EGFR expression level in SKOV3 cells was determined by real-time quantitative PCR method and Western blot.The signal intensity was measured by magnetic resonance imaging (MRI).Results:The transfection rate of the experimental group was 79.20±3.31%.It was obviously higher than that of negative control group (P=0.001). Compared with the negative control group,the cell viability of experimental group decreased obviously(P=0.011). Protein and mRNA expression level of EGFR in experimental group were significantly decreased (both P<0.01). The signal intensity on T2*WI in the intervenion group also significantly decreased (P=0.0004).Conclusion.The external magnetic field can improve the SPIO-shRNA dual functional molecular probe transfection efficiency.