ABSTRACT
Objective To study the antiviral effect and mechanism of camptothecin against HSV-1 in vitro. Methods Adding different concentration of camptothecin to the Vero cells infected by HSV-1, the measure of cytopathologic effects (CPE) was taken to evaluate the result of antiviral effect and mechanism of camptothecin. The median toxic concentration (TC50), median inhibitory concentration (IC50), and therapeutic index (TI) of camptothecin were calculated, and the affection of camptothecin on HSV-1 mRNA expression was examined through the method of RT-qPCR. Results TC50 of camptothecin was 2 153.44 nmol/L, IC50 was 43.01 nmol/L, and TI was 50.07. The results of RT-qPCR showed that camptothecin can significantly inhibit the gene transcription of HSV-1, UL12, UL42, UL54, and TK, and its inhibitory effect was in dose-dependence relationship with the concentration of camptothecin. Conclusion Camptothecin has an obvious antiviral effect on HSV-1 and can inhibit the transcription of HSV-1 genes such as UL12, UL42, UL54, and TK.
ABSTRACT
Human cytomegalovirus (HCMV) is a DNA virus and serious opportunistic pathogen for both newborn and immunocompromised individuals.To research technique for gene silence and antiviral agents, ribozyme M1GS-T6 was constructed from external guide sequences(EGSs)that consist of a sequence complementary to HCMV UL54 gene RNA and M1 RNA, the catalytic RNA subunit of RNase P from Escherichia coli. The results showed that M1GS can efficiently cleave the mRNA sequence encoding UL54 protein in vitro.
ABSTRACT
AIM: To study the interrelation between the structure and the function of artificial ribozyme M_1GS. METHODS: Ribozyme M_1GS-T_7, which targeted the mRNA segment of HCMV UL54 gene, was constructed. The secondary structure of M_1GS was simulated under different temperatures (20 ℃, 37 ℃ or 55 ℃, at which the secondary structure of M_1GS was relatively stable) and the interrelation between the secondary structure and the cleavage activity of M_1GS was analyzed under different temperature in vitro. To investigate the interrelation between the structure and the function of ribozyme M_1GS further, mM_1GS-T_7 was designed, in which some mutation sites was added, according to the result of temperature change experiment and the simulated secondary structure, showing that were the same structures at 37 ℃ as that of M_1GS-T_7 at 55 ℃. RESULTS: In temperature change experiment, the cleavage activity of M_1GS-T_7 was highest at 55 ℃. The result of mutant experiment showed that the mutant type was more active than M_1GS-T_7 at 37 ℃. CONCLUSION: The cleavage activity of M_1GS, which has some certain secondary structure, is higher than others. There is some interrelation between the structure and the function of M_1GS.