The Chinese herbal prescription JZ-1 induces autophagy to protect against herpes simplex Virus-2 in human vaginal epithelial cells by inhibiting the PI3K/Akt/mTOR pathway
The Chinese herbal prescription JZ-1 induces autophagy to protect against herpes simplex Virus-2 in human vaginal epithelial cells by inhibiting the PI3K/Akt/mTOR pathway
复制标题
中药方JZ-1通过抑制PI3K/Akt/mTOR通路诱导自噬，从而预防人阴道上皮细胞中的单纯疱疹病毒2
DOI：
10.1016/j.jep.2020.112611
复制
发表时间：
2020-05-23
影响因子：
5.4
通讯作者：
Chen, Zhuo
中科院分区：
文献类型：
--
作者：
Shao, Qingqing;Liu, Tong;Chen, Zhuo
民族药理学相关性：中药方洁泽-1（JZ-1）是基于一黄汤的修改，其首先由清代著名医生单父在《傅清竹女科》中描述为治疗白带疾病。JZ-1作为院内制剂，多年来在同济医院用于治疗各种女性下生殖道感染性疾病，包括宫颈炎、阴道炎、生殖器疱疹和尖锐湿疣。生殖器疱疹是由单纯疱疹病毒2型（HSV-2）引起的一种性传播疾病，是一种常见的性传播疾病，也是一种常见的性传播疾病。目前的疗法可以缓解患者的症状，但不能治愈或预防病毒的传播。本研究旨在从自噬诱导的角度探讨JZ-1抗HSV-2感染的作用及其机制，为抗病毒药物的研究提供新的思路和依据。材料与方法：采用MTT法和western blotting法评价JZ-1的抗病毒活性。采用Western blot和免疫荧光分析、透射电镜观察和慢病毒重组载体mRFP-GFP-LC 3B实验等方法检测VK 2/E6 E7细胞的自噬通量。为探讨JZ-1调节自噬的机制，采用Western blotting和实时定量PCR（qRT-PCR）方法检测PI 3 K/Akt/mTOR通路蛋白的表达，并检测应用PI 3 K抑制剂后该通路中关键分子的变化。结果：HSV-2感染可抑制VK 2/E6 E7细胞的自噬。进一步的研究表明，HSV-2感染诱导的PI 3 K/Akt/mTOR通路的激活可能导致自噬流受阻，抑制自噬体和自溶酶体的形成。JZ-1在VK 2/E6 E7细胞中表现出显著的抗病毒活性，表现为细胞活力增加，病毒蛋白表达减少，即最早病毒特异性感染细胞多肽5（ICP 5）和糖蛋白D（gD）。我们发现JZ-1处理抑制PI 3 K/Akt/mTOR通路蛋白的上调并促进自噬以对抗HSV-2感染，而PI 3 K抑制剂预处理阻止JZ-1诱导的增强的自噬。结论：JZ-1对HSV-2感染具有保护作用，其机制可能是通过抑制PI 3 K/Akt/mTOR信号通路诱导自噬而实现的。
Ethnophamacological relevance: The Chinese herbal prescription JieZe-1 (JZ-1) is based on the modification of Yihuang Tang, which was first described in Fu Qingzhu Nvke by the famous Qing Dynasty doctor Shan Fu as a treatment for leukorrheal diseases. As an in-hospital preparation, JZ-1 has been used in Tongji Hospital for many years to treat various infectious diseases of the lower female genital tract, including cervicitis, vaginitis, genital herpes and condyloma acuminatum. Our previous studies have shown that JZ-1 has curative effects on Candida albicans, Trichomonas vaginalis and Ureaplasma urealyticum infections.Aim of the study: Genital herpes is among the most common sexually transmitted diseases (STDs) worldwide and is mainly caused by herpes simplex virus type-2 (HSV-2). Current therapies can relieve symptoms in patients but do not cure or prevent the spread of the virus. This study was designed to investigate the effect of JZ-1 on HSV-2 infection and its mechanism, which is based on autophagy induction, to provide new ideas and a basis for the study of antiviral drugs.Materials and methods: Evaluation of the antiviral activity of JZ-1 was conducted by MTT assay and western blotting. Then, Western blot and immunofluorescence analyses, observations through transmission electron microscopy and experiments with the recombinant lentivirus vector mRFP-GFP-LC3B were used to monitor autophagic flux in VK2/E6E7 cells. To explore the mechanism by which JZ-1 regulates autophagy, western blotting and real-time quantitative PCR (qRT-PCR) were used to determine the expression of phosphoinositide 3kinase (PI3K)/Akt/mTOR pathway proteins and to detect changes in critical molecules in the pathway after the application of a PI3K inhibitor. Additionally, the mRNA expression levels of inflammatory cytokines, namely, IL6, IFN-alpha, IFN-beta and TNF-alpha, were measured with qRT-PCR.Results: HSV-2 infection inhibited autophagy in the VK2/E6E7 cells. Further study revealed that the activation of the PI3K/Akt/mTOR pathway induced by HSV-2 infection may result in the blocked autophagic flux and inhibited autophagosome and autolysosome formation. JZ-1 exhibited significant antiviral activity in the VK2/ E6E7 cells, which showed increased cell vitality and reduced viral protein expression, namely, earliest virusspecific infected cell polypeptides 5 (ICP5) and glycoprotein D (gD). We found that JZ-1 treatment inhibited the upregulation of the PI3K/Akt/mTOR pathway proteins and promoted autophagy to combat HSV-2 infection, while PI3K inhibitor pretreatment prevented the enhanced autophagy induced by JZ-1. Moreover, JZ-1 attenuated the increase in inflammatory cytokines that had been induced HSV-2 infection.Conclusion: Our results showed that JZ-1 protects against HSV-2 infection, and this beneficial effect may be mediated by inducing autophagy via inhibition of the PI3K/Akt/mTOR signaling axis.