Abstract:

OBJECTIVE: To verify the lethal effects on cervical cancer cells by ultrasonic irradiation combined with lentivirus vector microbubble of double suicide gene，to lay the foundation of targeted gene therapy in cervical cancer. METHODS：After constructing lentivirus vector carrying the double suicide gene system (pLenti6-KDRP-CD/TK-EGFP)，transfecting it into the 293T cells，computing virus titer and mingling with microbubble (sonovue)，we then examined the encapsulating rate and drug loading capacity. The cervical cancer cells (HeLa) were divided to four groups：the blank group；the lentivirus vector group；the lentivirus vector microbubble group；ultrasonic irradiation+lentivirus vector microbubble group：(radiated 30 s by 0.25，0.5，1.0，2.0 W/cm2) 300 kHz. We examined the effect on proliferation of cervical cancer cells by MTT assay and apoptosis by flow cytometry. RESULTS：The virus titer，encapsulating rate and drug loadings capacity were assessed and the expression of EGFP were examined under the fluorescence microscope. MTT assay showed that proliferation of cervical cancer cells was obviously inhibited in the lentivirus vector group，the lentivirus vector microbubble group and 0.25 W/cm2+ the lentivirus vector microbubble group (P<0.05). When the intensity of ultrasound was higher than 0.5 W/cm2，the inhibition effect was more significant (P<0.01). With progressively lighter intensity of ultrasound，inhibition rates of cervical cancer cells were gradually increased until 2.0 W/cm2，when the inhibition rates tended to be steady. After 24 h，the inhibition effect decreased obviously (P<0.05). After 48 h，there was no significant change for this effect (P>0.05). The apoptosis rates of cervical cancer cells were obviously increased in the lentivirus vector group and lentivirus vector microbubble group (P<0.05). When combining with ultrasonic irradiation，the apoptosis rate of cervical cancer cells was more significant (P<0.01). CONCLUSION： Microbubble carrying lentivirus vector of double suicide gene could kill cervical cancer cells (HeLa). The lethal effect on cervical cancer cells was obviously increased by combining with ultrasonic irradiation.

Key words:  double suicide gene, ultrasound microbubble, lentivirus vector, proliferation, apoptosis