Recomeinant Caspase3 Gene Transferred Into Cervical Cancer Cells By Ultrasound-Mediated Liposome: Experimental Study

Objective: To investigate the feasibility of recombinant caspase3 gene transferred into cervical carcinoma cell Ca Ski by ultrasound-mediated liposome and the apoptosis-inducing effect of recombinant caspase3 gene in cervical carcinoma cell line.Methods: In order to screen out the optimal ultrasonic intensity and time without conspicuous harm to Ca Ski cells, cells in 24 well culture plates were exposed to different ultrasonic intensity of 0.65w/cm² and 0.8w/cm². The exposure time was 10, 15, 20, 25, 30, 35 and 40s respectively. The selected ultrasonic condition was used to experiment. The Ca Ski cells were randomly divided into five groups as follows:â… control group,â…¡pure ultrasound,â…¢plasmid with ultrasound and liposome,â…£plasmid plus liposome, andâ…¤pure plasmid. (1)The cell growth inhibitory effect of each group was observed by MTT assay. (2)Morphological changes of apoptosis in each group were detected by Hoest33258. (3) Cell cycle distribution was analyzed using Flow cytometry (FCM). (4) Gene expression of each group was detected by RT-PCR.Results: The optimal ultrasonic conditions were intensity of 0.8w/cm2 when the Ca Ski cells exposed about 25s. (1)The results of MTT showed that the significant cell growth inhibition exist in pure ultrasound, plasmid with ultrasound and liposome, plasmid plus liposome groups. The cell inhibitory rate in the plasmid with ultrasound group was significantly higher than other groups (P<0.05). (2)Hoest33258 fluorescence dyeing point out that plasmid R-Caspase3 can induce cell apoptotic obviously so that the cell nucleus cracked and dyeing intensely. The cell inhibitory rate in the five groups respectively was (0.7±0.57)%, (3.8±1.32)%, (14.2±1.55)%, (11.5±1.78)%, (0.4±0.53)%. The result of plasmid with ultrasound and liposome group was significantly different with other groups (P<0.05). (3) The results of FCM showed that the distributional percentage of cell cycle in plasmid with ultrasound and liposome, plasmid plus liposome groups changed, which mainly were a decrease in the proportion of G₀/G₁ phase and an increase in the proportion of S and G₂/M phase in cell cycle. The increase in the proportion of S phase was the most significant in the group of plasmid with ultrasound and liposome (P<0.05). The change of cell cycle in pure ultrasound group mainly was an increase in the proportion of G₂/M phase and a decrease in the proportion of G0/G1 phase. (4) The results of RT-PCR suggested that plasmid with ultrasound and liposome, plasmid plus liposome groups can express gene of R-Caspase3 and the gray intensity of plasmid with ultrasound and liposome group was significantly higher than other groups(P<0.05).Conclusions: R-caspase3 gene can transfer into Ca Ski cells efficiently by ultrasound-mediated liposome; Recombinant Caspase3 gene can express in cervical cancer cell and lead cells apoptotic. It is very important for the gene therapy of cancer cells.