The Chinese Herbal Prescription JieZe-1 Inhibits Membrane Fusion and the Toll-like Receptor Signaling Pathway in a Genital Herpes Mouse Model.

Chinese herbal prescription JieZe-1 is effective for genital herpes with no visible adverse effects clinically. It showed an excellent anti-HSV-2 effect in vitro. However, its mechanism of anti-HSV-2 effect in vivo remains unclear. This study was designed to evaluate the anti-HSV-2 effect of JieZe-1 and berberine in a genital herpes mouse model and explore the underlying mechanism. The fingerprint of JieZe-1 was determined by high-performance liquid chromatography. First, we optimized a mouse model of genital herpes. Next, the weight, symptom score, morphological changes, viral load, membrane fusion proteins, critical proteins of the Toll-like receptor signaling pathway, cytokines, and immune cells of vaginal tissue in mice at different time points were measured. Finally, we treated the genital herpes mouse model with JieZe-1 gel (2.5, 1.5, and 0.5 g/ml) and tested the above experimental indexes at 12 h and on the 9th day after modeling. JieZe-1 improved the symptoms, weight, and histopathological damage of genital herpes mice, promoted the keratin repair of tissues, and protected organelles to maintain the typical morphology of cells. It downregulated the expression of membrane fusion proteins, critical proteins of the Toll-like receptor signaling pathway, cytokines, and immune cells. The vaginal, vulvar, and spinal cord viral load and vaginal virus shedding were also significantly reduced. In summary, JieZe-1 shows significant anti-HSV-2 efficacy in vivo. The mechanism is related to the inhibition of membrane fusion, the Toll-like receptor signaling pathway, inflammatory cytokines, and cellular immunity. However, berberine, the main component of JieZe-1 monarch medicine, showed no efficacy at a concentration of 891.8 µM (0.3 mg/ml).

Zearalenone Blocks Autophagy Flow and Induces Cell Apoptosis During Embryo Implantation in Gilts.

Zearalenone (ZEA) has been proved to be toxic, particularly to the reproductive system of gilts. The effect of ZEA on gilts during embryo implantation window period is of particular interests. Here, we observed window stage dysontogenesis of gilts treated with ZEA. In endometrial tissues and cells, autophagosomes increased significantly and mitochondria were damaged with increasing ZEA concentration. Addition of autophagy inhibitor confirmed that ZEA blocks the autophagic flow in the fusion of autophagosomes and lysosomes. In conclusion, ZEA exposure during embryo implantation results in endometrium inflammation by activating autophagy while blocking autophagy flow at the same time, leading to the significant accumulation of autophagosomes. The aforementioned effects of ZEA induce the apoptosis of primary endometrial cells through the caspase3 pathway, which would break the uterus environment balance and finally lead to embryo implantation failure and dysontogenesis in gilts.

Antiviral effect of Chinese herbal prescription JieZe-1 on adhesion and penetration of VK2/E6E7 with herpes simplex viruses type 2.

ETHNOPHARMACOLOGICAL RELEVANCE: The Chinese Herbal Prescription JieZe-1(JZ-1), added and subtracted from Yihuang Decoction, a famous formula in the 12th year of Kangxi in Qing Dynasty, has a clear effect on Genital Herpes (GH) and no obvious adverse reactions occur clinically. JZ-1 also has preventive and therapeutic effects on Trichomonas vaginitis, Candida albicans vaginitis and GH in vitro and in vivo experiments. AIM OF STUDY: The effect and mechanism of JZ-1 on anti-herpes simplex virus type 2(HSV-2) in vitro focusing on adhesion and penetration stages were investigated. MATERIALS AND METHODS: A model of HSV-2 infection of VK2/E6E7 was developed. In order to explore JZ-1's anti-HSV-2 effect in vitro, cell morphology, ultrastructural pathology, cell viability and expression of viral glycoprotein D (gD) were assessed at 6 h, 12 h, 18 h, and 24 h of JZ-1 treatment. Then we measured the exact time required for adhesion and penetration of HSV-2 into VK2/E6E7 among a series of times at room temperature and under temperature control techniques. We treated VK2/E6E7 with JZ-1, penciclovir, or berberine and explored the mechanism of JZ-1 in blocking HSV-2 adhesion and penetration of host cells by assessing the cell ultrastructural pathology, viability, viral proteins gB, gD, VP16, ICP5, and ICP4 and host cell proteins HVEM, Nectin-1, and Nectin-2. RESULTS: HSV-2 can fully adhere and penetrate into VK/E6E7 within 5 mins at room temperature while it takes 60mins under temperature control techniques. JZ-1 and penciclovir showed significant anti-HSV-2 effects, with improved host cell morphologies and increased host cell viabilities observed after treatment for 24 h. The anti-HSV-2 effect of JZ-1 can be detected after treatment for 6 h while that of penciclovir was not obvious until treatment for 12 h. JZ-1 showed distinct effect on HSV-2 adhesion and penetration stages by significantly reducing the expression of viral proteins gB, gD, VP16, ICP5, and ICP4, improving cell morphology and increasing cell viability. However, these effects were not exerted via downregulated expression of membrane fusion-related proteins such as HVEM, Nectin-1, or Nectin-2. The specific anti-HSV-2 mechanism of JZ-1 need to be further explored. CONCLUSION: The anti-HSV-2 effect of JZ-1 was superior to that of penciclovir and berberine in vitro, and was mainly mediated by enhancing host cell defense and blocking adhesion and penetration of HSV-2.

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.

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 3-kinase (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, IL-6, IFN-α, IFN-ß and TNF-α, 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 virus-specific 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.