摘要
目的:通过体外细胞实验探讨槲皮素抑制肠道病毒71型(Enterovirus 71,EV71)感染的能力,并对其抗病毒机制进行探讨,为手足口病的治疗及抗病毒中药开发提供基础研究数据。方法:TCID50法滴定病毒;MTT法行细胞毒性及槲皮素抗病毒活性实验,计算CC50、EC50及SIs;Western blot检测病毒蛋白;针对不同实验组Western blot检测TLR7、Caspase3及病毒蛋白VP1水平;实时荧光定量PCR检测实验组白细胞介素6(interleukin-6,IL-6)及白细胞介素8(interleukin-8,IL-8)等细胞因子水平,以此进行抗病毒机制研究。结果:槲皮素50%细胞毒性浓度(CC50)值>100μM,细胞毒性小,有效抑制50%RD细胞感染EV71的EC50为15μM,计算SI至少大于6,同时Western blot结果显示槲皮素处理的细胞中病毒VP1蛋白受到显著抑制,且存在剂量依赖效应;针对机制研究发现,EV71病毒感染RD细胞后上调TLR-7表达水平,而槲皮素通过抑制下游的IL-6及Caspase3表达水平发挥抗病毒作用。结论:槲皮素体外对EV71有较好的抗病毒作用,具有潜在的临床应用前景,其抗病毒机制可能通过抑制IL-6及下游的细胞凋亡来实现。
Objective:To explore the capability of quercetin inhibiting EV71 infection via in vitro cell assay,and discuss its antiviral mechanism,therefore to provide the data of basic researches for the therapy of hand-footand-mouth disease and the development of antiviral TCM.Methods:TCID50 method was used to titrate the virus;MTT method was applied to perform cytotoxicity test and antiviral activity of quercetin,to calculate CC50,EC50 and SIs;Western blot was adopted to detect viral proteins,and it was used to measure the levels of TLR7,Caspase 3 and VP1 of viral protein in different experimental groups;and real-time fluorescent quantitative PCR to detect the levels of IL-6 and IL-8 in the exprimental group,therefore to perform antiviral mechanism study.Results:When 50%cytotoxic concentration of quercetin>100μM,it shows low cytotoxicity,it could effectively inhibit 50%RD cells infected with EV71,when EC50 was 15μM,SI is greater than 6 at least,meanwhile,Western blot results indicated that viral VP1 protein in cells processed by quercetin was notably restrained,and it existed in dose-dependent effect;the mechanism study revealed that EV71 up regulated the expressions of TLR-7 in RD cells,while quercetin developed its antiviral effects by inhibiting the expressions of IL-6 and Caspase3.Conclusion:Quercetin has a good antiviral effect on EV71 in vitro,it possesses potential clinical application prospect,and its antiviral mechanism might be realized by inhibiting the levels of IL-6 and the apoptosis of the downstream cells.
作者
张亚婷
陈静
张娟丽
徐丁
ZHANG Yating;CHEN Jing;ZHANG Juanli;XU Ding(Lanzhou University Second Hospital,Lanzhou 730000,China)
出处
《西部中医药》
2022年第8期30-34,共5页
Western Journal of Traditional Chinese Medicine
基金
甘肃省中医药管理局科研项目(GZK-2015-73)。
