鸦胆子素D对高危型HPV 16感染细胞的增殖抑制作用及对HPV16 E6、HPV 16 E7 mRNA表达的影响

Inhibiting effect of bruceine D on proliferation of high-risk human papillomavirus 16 infected cells and the effect on HPV16 E6 and HPV16 E7 mRNA expressions

目的探讨鸦胆子素D对高危型人乳头瘤病毒(HPV)16感染细胞的增殖抑制作用及对HPV16 E6、HPV16 E7mRNA表达的影响。方法以人宫颈鳞状上皮永生化细胞株(Ect1/E6E7)作为HPV16型感染的体外实验模型,而宫颈癌细胞株Caski作为阳性对照,不同浓度的鸦胆子素D分别体外作用于细胞24 h、48 h、72 h,采用噻唑篮(MTT)法测定细胞体外增殖活性,采用Real-time PCR法检测HPV16 E6、HPV16 E7基因mRNA的表达。结果与同期对照组相比,观察组不同浓度的鸦胆子素D作用的细胞生长抑制率(CGIR)均明显增高,另外,与同一浓度作用24 h相比,作用48 h、72 h的CGIR均明显增高,差异均有统计学意义(均P<0.05)。即鸦胆子素D的浓度越高及作用时间越长,对Ect1/E6E7细胞生长的抑制作用越强。Ect1/E6E7细胞中,观察组不同浓度的HPV16 E6、HPV16 E7 mRNA的表达比较差异无统计学意义,两组比较差异亦无统计学意义(均P>0.05)。而随着鸦胆子素D溶液的浓度上升,Caski细胞中HPV16 E6、HPV16 E7 mRNA的表达升高,两组比较差异有统计学意义(均P<0.05)。结论鸦胆子素D可抑制HPV16 E6、HPV16 E7 mRNA表达和HPV16细胞的增殖。

Objective To explore the inhibiting effect of bruceine D on proliferation of high-risk human papillomavirus （HPV） 16 infected cells and the effect on HPV E6 and HPV E7 mRNA expressions. Methods An immortalized squamous cell line of human cervical squamous epithelium （Eetl/E6E7） was used as an in vitro experimental model of HPV16 infection, and cervical cancer cell line Caski was used as positive control group. These cells were cultivated with different concentrations of hruceine D in vitro for 24, 48, and 72 hours, respectively. The proliferation activity of cells in vitro was measured with thiazolylblue tetrazolium bromide （MTT） assay, HPV16 E6 and HPV16 E7 mRNA expressions were detected by real-time PCR. Results Compared with control group, cell growth inhibition rate （CGIR） in observation group with different concentrations of bruceine D significantly increased. In addition, compared with the same concentration at 24 hours, CGIRs at 48 and 72 hours were significantly higher, and the differences were statistically significant （all P〈0. 05 ） . The higher the concentration of hrucea D and the longer the treatment time was, the stronger inhibitory effect on the growth of Eetl/E6E7 ceils was. There was no statistically significant difference in HPV16 E6 and HPV16 E7 mRNA expressions among Ectl/E6E7 cells treated with different concentrations in observation group, and there was no statistically significant difference between the two groups （ all P〉0. 05 ） . With the increase of bruceine D solution concentration, HPV16 E6 and HPV16 E7 mRNA expressions of Caski ceils increased, there were statistically significant differences between the two groups （all P〈0. 05 ） . Conclusion Bruceine D can inhibit HPV16 E6 and HPV16 E7 mRNA expressions and the proliferation of HPV16 infected cells.