Optimal concentration of superparamagnetic iron oxide-short hairpin RNA dual functional molecular probe transfected into ovarian cancer cells in vitro / 北京大学学报(医学版)

ABSTRACT

Objective:To explore the effects of superparamagnetic iron oxide-short hairpin RNA ( SPIO-ShRNA) dual functional molecular probes of different concentrations on morphology and biological beha -vior of ovarian cancer SKOV3 cells in vitro.Methods:The dual functional molecular probes at an iron concentration of 5, 15, 30, 45, 75, and 100 mg/L were transfected into SKOV3 cells.The transfection rate of the 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 cell counting kit-8 ( CCK-8 ) .The apoptosis was detected by flow cytometry .The expression of protein within the cells was detected by Western blot .The changes of the signal intensity were measured by magnetic resonance imaging (MRI).Results: The SPIO-ShRNA dual functional molecular probe was uptaken in aconcentration-dependence manner within a certain range (5-30 mg/L) .When the concentration of the probe was 45 mg/L, the labeling rate of the cell was close to 100%;With the increase of the concentration of probe , the cell survival rate decreased gradual-ly.The cell survival rate of each experimental group were 94.626%±1.050%, 93.373%±1.180%, 91.700%±3.122%, 75.100%±4.362%, 72.983%±3.233%, 71.010%±2.910%,5, 15, 30mg/L cell survival rate was not significantly decreased , the difference was not statistically significant (P=0.226, P=0.068, P=0.475);When the concentration of the probe was greater than or equal to 45 mg/L,the survival rate decreased obviously ( P<0.001);Group of 45 mg/L protein expression rate was 68.905%± 3.510%, When the concentration of the probe was greater than or equal to 45 mg/L, the inhibition rate of the protein expression level of epidermal growth factor receptor was obviously higher than those of 5, 15, and 30 mg/L groups, the difference was statistically significant (P<0.001, P=0.001, P=0.003, all P<0.01);the MRI displayed that the signal intensity was decreased with increasing concentrations of the probe.The signal intensity of 45 mg/L group was 165.55 ±4.92, compared with the blank control group (same volume of phosphate buffer saline ), normal group(unlabeled ovarian cancer SKOV3 cells), 5, 15, and 30 mg/L groups , the signal intensity of 45 mg/L group decreased significantly (all P<0.001).Con-clusionThe dual functional molecular probe can effectively transfect and specifically inhibit the expression of SKOV3 cell lines at the iron concentration of 45 mg/L, and can also be detected by MRI .The role of diagnosis and treatment of the dual functional molecular probe has been initially confirmed .