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纸质出版日期:2018
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冉雪梦, 王世欣, 张立岩, 等. 清热解毒方体外抗人乳头瘤病毒感染及作用机制[J]. 中国实验方剂学杂志, 2018,24(24):171-176.
RAN Xue-meng, WANG Shi-xing, ZHANG Li-yan, et al. Effect of Qingre Jiedu Prescription in Resisting Human Papllimavirus Infection and Its Mechanism[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(24): 171-176.
冉雪梦, 王世欣, 张立岩, 等. 清热解毒方体外抗人乳头瘤病毒感染及作用机制[J]. 中国实验方剂学杂志, 2018,24(24):171-176. DOI: 10.13422/j.cnki.syfjx.20182423.
RAN Xue-meng, WANG Shi-xing, ZHANG Li-yan, et al. Effect of Qingre Jiedu Prescription in Resisting Human Papllimavirus Infection and Its Mechanism[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(24): 171-176. DOI: 10.13422/j.cnki.syfjx.20182423.
目的:采用高危型人乳头瘤状病毒16(human papillomavirus 16,HPV16)体外假病毒感染模型评价清热解毒方体外抗HPV16感染活性及作用方式,并探讨清热解毒方的作用机制。方法:首先,将HPV16假病毒加入到293T/17细胞中,建立HPV假病毒体外荧光感染模型,以此模型评价清热解毒方(20~1 000 g·L-1)体外抗HPV16作用,探讨清热解毒方抗HPV感染的量效关系;其次,采用HPV假病毒模型,通过预处理细胞、与病毒同时作用于细胞、病毒处理后作用于细胞的不同处理方法,探讨清热解毒方(20 g·L-1)抗HPV的作用方式;最后,从清热解毒方作用后HPV转化的HeLa细胞中提取致癌蛋白E7
细胞外信号调节激酶(ERK)以及核转录因子-κB(NF-κB),用蛋白免疫印迹法(Western blot)检测E7,ERK,NF-κB蛋白表达,探讨清热解毒方(3.33~20 g·L-1)在肿瘤相关蛋白水平上的作用机制。结果:与病毒组比较,清热解毒方可以明显抑制HPV16假病毒的感染(P<0.05),且呈现明显的浓度依赖关系;且清热解毒方能够通过保护细胞、抑制病毒吸附并抑制病毒感染后过程的多种方式发挥其抗HPV16作用,其中保护细胞的效果更显著(P<0.01),说明清热解毒方能够显著预防HPV16感染;与空白组比较,清热解毒方能够明显抑制E7致癌蛋白的表达,并抑制ERK,NF-κB蛋白的表达(P<0.05,P<0.01),达到抗肿瘤的作用。结论:清热解毒方具有较好的抑制HPV病毒的效果,该研究为清热解毒方清除HPV病毒和防治宫颈癌提供了创新性的理论依据。
Objective: To assess the activity and action modes of Qingre Jiedu prescription in resisting human papillomavirus 16 (HPV16) in vitro by establishing the cell model infected by HPV16 pseudovirus particles
in order to explore the mechanism of action of Qingre Jiedu prescription on transformed HeLa cell HPV. Method: Firstly
HPV16 pseudovirus particles were added to 293T/17 cells to establish an HPV pseudovirus infection model
so as to detect fluorescence intensity in vitro. Then this model was used to evaluate the effect of Qingre Jiedu prescription in resisting HPV16 infection in vitro and its dose-effect relationship of Qingre Jiedu prescription (20~1 000 g·L-1). Secondly
the HPV pseudovirus infection model was used to explore the action modes of Qingre Jiedu prescription (20 g·L-1) against HPV under such different conditions as pre-treating the cells
acting the cells simultaneously with virus and acting the cells after virus treatment. Finally
the mechanism of Qingre Jiedu prescription (3.33~20 g·L-1) was explored by Western blot. In brief
the oncogenic protein E7
extracellular signal-regulated kinase (ERK) and nuclear factor-κB (NF-κB) were extracted from HeLa cells HPV transformed after the action of Qingre Jiedu prescription and detected by Western blot. These proteins were compared with the corresponding protein expression levels in the blank group. Result: The results of experiment on HPV16 pseudovirus infection model showed that Qingre Jiedu prescription could significantly inhibit the infection of HPV16 pseudovirus in cells (P<0.05)
in an obvious concentration-dependent manner. The action modes of Qingre Jiedu prescription in the HPV16 pseudovirus infection model in vitro showed that Qingre Jiedu prescription could protect the cells
inhibit the absorption and post-virus infection process. And the drug can significantly prevent HPV16 infection because it could protect cells from the HPV16 infection (P<0.01). These results show that Qingre Jiedu prescription could protect cells from the HPV16 infection
inhibit the absorption and post-virus infection process. In addition
Qingre Jiedu prescription could obviously inhibit the expressions of oncoprotein E7
and ERK and NF-κB protein
so as to achieve the antitumor effect(P<0.05
P<0.01). Conclusion: Qingre Jiedu prescription has a better effect in inhibiting HPV virus
and the HPV16 pseudovirus infection experiment provides the innovative theory basis for the application of Qingre Jiedu prescription in anti-HPV and cervical cancer therapy.
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