Biological effects of upregulated expression of transfected FOLR1 gene on SKOV3 cell lines acted by cisplatin

Abstract

To explore biological effects of up-regulated expression of transfected FOLR1 gene on SKOV3 cell lines following action by cisplatin (DDP) . Three groups of cells originated from the same SKOV3 cell line were used in this research, including the SKOV3 cell line(blank control), the cell line transfected with lentiviral pWPI plasmid(no-load control) and the cell line transfected with FOLR1 gene via lentiviral pWPI plasmid(experimental group). Next, the mRNA and protein expression of FOLR1 gene in the three groups were detected by reverse transcription (RT) -PCR and western blot, respectively. Methyl thiazolyl tetrazolium (MTT) assay was used to analysis cells growth curve and identify their sensitivity to cisplatin, and their half inhibition concentration(IC50) values were calculated. Based on the IC50 value(3.6 µg/ml) in the experimental group, different levels of cisplatin concentration (0.5×IC50, 1×IC50, 2×IC50, respectively) were administered to the three groups of cells, and the inhibition rates, apoptosis rates as well as apoptosis proportion of each group after 24, 48, 72 hours were further recorded.Finally, the residual cisplatin concentrations in the three group cells acted successively by 1×IC50 cisplatin for 48 hours were measured by high performance liquid chromatography (HPLC) . P value less than 0.05 were defined as statistically significant. RT-PCR and western blot detection showed that stable mRNA and protein expression of the FOLR1 gene in the experimental group while the other two groups were not. MTT assay demonstrated that higher cell growth rate, sensitivity to cisplatin(IC50 = 3.6 µg/ml) and inhibition rate in the experimental group compared with those in the other two groups (P < 0.05) , which showed no significance in intergroup comparison(P > 0.05). Flow cytometry showed apoptosis rates among three groups increased with higher cisplatin concentrations and longer action duration in dosage-time dependent manner (P < 0.05) , and the proportion of S phase cells increased with higher cisplatin concentration in dosage-dependent manner (P < 0.05) ;for the same concentration and duration, the experimental group showed significantly different apoptosis rates and S phase cells compared with the other two groups, which demonstrated no significance in intergroup comparison (P > 0.05) . After action by cisplatin(3.6 µg/ml) for 48 hours, HPLC showed significantly higher residual cisplatin concentration (2.60 ± 0.21) µg/10(6) cell counts in experimental group than those in no-load control group (1.49 ± 0.12) µg/10(6) cell counts and blank control group(1.54 ± 0.11) µg/10(6) cell counts, respectively (P < 0.05) , and the comparison within the latter two groups showed no significance (P > 0.05). Up-regulated expression of the transfected FOLR1 gene in SKOV3 cells may be associated with higher sensitivity to cisplatin, residual cisplatin concentration and higher proportion of S phase cells, and tended to inhibit cancer cells growth and induce apoptosis.