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COVID-19 is a disease caused by SARS-CoV-2, which has led to more than 4 million deaths worldwide. As a result, there is a worldwide effort to develop specific drugs for targeting COVID-19. Papain-like protease (PLpro) is an attractive drug target because it has multiple essential functions involved in processing viral proteins, including viral genome replication and removal of post-translational ubiquitination modifications. Here, we established two assays for screening PLpro inhibitors according to protease and anti-ISGylation activities, respectively. Application of the two screening techniques to the library of clinically approved drugs led to the discovery of tanshinone IIA sulfonate sodium and chloroxine with their IC50 values of lower than 10 μM. These two compounds were found to directly interact with PLpro and their molecular mechanisms of binding were illustrated by docking and molecular dynamics simulations. The results highlight the usefulness of the two developed screening techniques for locating PLpro inhibitors.

Ethnopharmacological relevance Zhuanggu Guanjie Pill (ZGGJP), a modern Chinese medicine formula, is composed of 12 herbs and has been used to treat osteoporosis in China for almost 30 years. However, no in vivo study of the influences of ZGGJP on the cytochrome P450 (CYP) activities have been reported. Aim of the study The aim of this study was to evaluate the effects of ZGGJP on the activities and the mRNA expression of CYP enzymes (CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A) and their corresponding nuclear receptor levels in rats. Materials and methods After 7 days oral treatment of ZGGJP at low- and high-dose, cocktail solution was given to rats. Blood samples were collected at series of time points. The plasma concentrations of probe drugs and their corresponding metabolites were determined by UPLC-MS/MS. The influence of ZGGJP on the activities of seven CYPs were evaluated the metabolic ratios (Cmax and AUC0-t) for metabolites/probe drugs. In addition, the effects of ZGGJP on the mRNA expression of CYPs and their corresponding nuclear receptors in rat liver were evaluated by real-time PCR. Results ZGGJP showed significant inductive effects on CYP1A2 and CYP2B6 of both male and female rats. The influence of ZGGJP on CYP2C9 and CYP3A showed gender difference. ZGGJP could induce the activities of CYP2C9 and CYP3A in female rats, but have no influence on the activities in male rats. ZGGJP had no effects on CYP2D6, CYP2C19 and CYP2E1. The mRNA expression results of CYPs were in accordance with the pharmacokinetic results. The mRNA expression levels of constitutive androstane receptor (CAR) and vitamin D receptor (VDR) were increased significantly in female rats at high dosage, but no significant changes were observed in male rats. Conclusion ZGGJP had inductive effects on CYP1A2 and CYP2B6 in both male and female rats. The results showed that ZGGJP could induce the activities of CYP2C9 and CYP3A in female rats, but had no effect in male rats. This may suggest that the influence of ZGGJP on CYP2C9 and CYP3A exhibit gender difference. The inductive effects of ZGGJP on the activities of CYPs, exhibiting gender difference, may be regulated by CAR and VDR. Therefore, co-administration of ZGGJP with other drugs, especially using CYP2C9 and CYP3A substrates in females, may need dose adjustment to avoid herb-drug interaction.

SARS‐CoV‐2 caused the emerging epidemic of coronavirus disease in 2019 (COVID‐19). To date, there are more than 82.9 million confirmed cases worldwide, there is no clinically effective drug against SARS‐CoV‐2 infection. The conserved properties of the membrane fusion domain of the spike (S) protein across SARS‐CoV‐2 make it a promising target to develop pan‐CoV therapeutics. Herein, two clinically approved drugs, Itraconazole (ITZ) and Estradiol benzoate (EB), are found to inhibit viral entry by targeting the six‐helix (6‐HB) fusion core of SARS‐CoV‐2 S protein. Further studies shed light on the mechanism that ITZ and EB can interact with the heptad repeat 1 (HR1) region of the spike protein, to present anti‐SARS‐CoV‐2 infections in vitro, indicating they are novel potential therapeutic remedies for COVID‐19 treatment. Furthermore, ITZ shows broad‐spectrum activity targeting 6‐HB in the S2 subunit of SARS‐CoV and MERS‐CoV S protein, inspiring that ITZ have the potential for development as a pan‐coronavirus fusion inhibitor., An effective drug against COVID‐19 is urgently needed while the current SARS‐CoV‐2 pandemic. Here Itraconazole and Estradiol benzoate are identified to inhibit SARS‐CoV‐2 infection via blocking the spike protein S2 subunit‐mediated membrane fusion and entry into the host cell by targeting the highly conserved six‐helical bundle (6‐HB) fusion core. Furthermore, Itraconazole is confirmed as a potential broad‐spectrum coronavirus fusion inhibitor.

Antimicrobial peptides (AMPs) as components of innate immunity system have been isolated from fish and other species. In this study, the crude proteins extracted with gradient ammonium sulfate precipitation technique from the processing by-products of African catfish Clarias gariepinus (C. gariepinus) were purified by size-exclusion chromatography and all the four obtained fractions, Clarias antimicrobial peptides I(CAP-I), CAP-II, CAP-III and CAP-IV, showed antimicrobial activity. Among of these fractions, CAP-IV showed the highest antimicrobial activity against Staphylococcus aureus, Aeromonas sobria, Aeromonas hydrophila, Escherichia coli by agar diffusion plate test and the diameter of inhibition zone was 8.34, 9.27, 6.76, 6.13 mm, respectively. The molecular weight of main peptides of CAP-IV was around 4.1 KD by SDS-PAGE analysis. CAP-IV showed antimicrobial activity against both gram-negative and gram-positive bacterial pathogens at minimum inhibitory concentrations (MICs) ranging from 105 to 420 μg/mL. The antimicrobial activity of CAP-IV was stable at wide pH range, 3–11 and was also heat-stable when temperature was below 80 °C. Freeze-thawing treatment also only had slight effects on the antimicrobial activity of CAP-IV. Besides, CAP-IV was not sensitive to the hydrolysis by pepsin and trypsin, except for protease K. These results suggest that CAP-IV isolated from C. gariepinus is potential to be developed as a new antimicrobial peptide and may partially explain the high disease resistance of African catfish C. gariepinus.

Stilbenoids including resveratrol contain the basic structural unit of 1,2-diphenylethylene. Naturally occurring stilbenoids have broad structural features due to oligomerization and modifications and some have demonstrated potent biological activities. In an effort to identify bioactive stilbenoids, we screened a group of dimeric and oligomeric stilbenoids against HSV-1 and HSV-2 infection. Several trimeric and tetrameric derivatives showed anti-herpetic activity at single digit micromolar concentrations. HSV-1 and HSV-2 replication requires for NF-κB and MAPK activation. The compounds showed no inhibitory activity against NF-κB and Erk/MAPK activation, instead those compounds promoted rapid and transient release of reactive oxygen species (ROS). Addition of N-acetylcysteine (NAC), a scavenger of ROS, reversed the inhibitory effect of those compounds against HSV replication. In addition to the identification of resveratrol derivatives with potent anti-HSV activity, our results uncover a mechanism of polyphenol-mediated anti-HSV response, linking anti-herpetic activity of oligomeric stilbenoids to innate immunity.

Highlights ►Houttuynia cordata Thunb. exhibits strong activity against herpes simplex virus. ►H. cordata blocks HSV-2 infection through inhibition of NF-κB, not MAPK. ► Flavonoids like quercetin, quercitrin and isoquercitrin as major active components., Houttuynia cordata Thunb. is a medicinal plant widely used in folk medicine in several Asian countries. It has been reported that a water extract of H. cordata exhibits activity against herpes simplex virus (HSV) and the virus of severe acute respiratory syndrome (SARS), although the mechanisms are not fully understood yet. Previous studies have demonstrated absolute requirement of NF-κB activation for efficient replication of HSV-1 and HSV-2 and inhibition of NF-κB activation has been shown to suppress HSV infection. Here we show that a hot water extract of H. cordata (HCWE) inhibits HSV-2 infection through inhibition of NF-κB activation. The IC50 was estimated at 50 μg/ml of lyophilized HCWE powder. At 150 and 450 μg/ml, HCWE blocked infectious HSV-2 production by more than 3 and 4 logs, respectively. The inhibitory activity was concomitant with an inhibition of NF-κB activation by HSV-2 infection. Although activation of NF-κB and Erk MAPK has been implicated for HSV replication and growth, HCWE showed no effect on HSV-2-induced Erk activation. Furthermore, we show that treatment with quercetin, quercitrin or isoquercitrin, major water extractable flavonoids from H. cordata, significantly blocked HSV-2 infection. These results together demonstrated that H. cordata blocks HSV-2 infection through inhibition of NF-κB activation.

In this paper, acetic acid was used for extracting crude antimicrobial proteins/peptides from by-products of healthy Clarias gariepinus reared at high stocking density. For the extraction process, three variables, namely the concentration of acetic acid, extraction time, and the ratio of liquid to solid were optimized by using orthogonal design L9(33) with the yield and antibacterial activity of the crude proteins/peptides extracts as test indices. The yields of extracted crude proteins were analyzed by range analysis. The results showed that the yield of crude proteins/peptides was different among nine groups, the highest was group 7, producing 1.5842 g (dialyzed in 1,000 MWCO and lyophilized) per 100 g raw material. All the nine extracted samples had similar inhibitory activity against Aeromonas hydrophila, Escherichia coli, Staphylococcus aureus, Citrobacter sp., and Microbacterium sp. except that groups 7 and 9 exhibited higher inhibitory activity against A. hydrophila and E. coli; moreover, the inhibitory activity of group 7 was higher than group 9 against the two bacteria strains. Nine groups did not display obvious different patterns on the SDS-PAGE gel. Based on these results above, the final conclusion was that group 7, namely 10 % acetic acid, 12 h extraction, and 5:1 liquid to solid ratio was the best combination for isolating the crude antimicrobial proteins/peptides from by-products of C. gariepinus.

An epizootic was discovered in grass carp (Ctenopharyngodon idellus) in earthen ponds in Ninghe, Tianjin Province, northern China, from May to June 2013. The cumulative mortality was 30% within 25 days, and the diseased fish presented with hemorrhaging in the abdominal wall, ascites in the abdomen and pale liver. Strains named PA131207 and PA131208 were isolated from the moribund fish, and were identified as Pseudomonas aeruginosa according to both biochemical tests and phylogenetic analysis derived from the 16S rRNA gene. The pathogenicity of PA131207 was confirmed by infectivity experiments and histopathological examination. Antimicrobial susceptibility tests showed that PA131207 was resistant to 9 of 15 antimicrobial agents tested.

OBJECTIVE To seek effective drugs inhibiting herpes simplex virus (HSV-2) with the signal pathway required by virus replication as the target spot. METHOD HSV-2-induced Vero cytopathic effect was observed, and MTT method was adopted to detect call activity, in order to assess the antiviral capacity of freeze dried powder of aqueous extracts of Saururus chinensis (AESC). Western blot was used to check the effect of AESC on signal pathway induced by HSV-2 virus in HeLa cells. RESULT AESC obviously inhibits the pathway activation of CPE induced by HSV-2 infection and NF-kappaB required for virus replication. The inhibition ratio of AESC freeze dried powder at 0.10, 0.03, 0.01 and 0.003 g x L(-1) were (70.68 +/- 3.39)%, (61.74 +/- 2.13)%, (39.31 +/- 1.10)% and (18.54 +/- 3.44)%, respectively. The IC50 was determined at (0.023 +/- 0.004) g x L(-1). The inhibition concentration of the positive control acyclovir was 0.001 g x L(-1) (5.0 x 10(-6) mol x L(-1)). The best administration time was from 2 h before infection to 6 h after infection. Western blot also showed that AESC can notably inhibit HSV-2-induced NF-kappaB nuclear transfer. CONCLUSION AESC can inhibit HSV-2 virus replication, which is related to the pathway activation of NF-kappaB required for virus replication.

Syndecan-4 (synd4) is a heparan sulfate proteoglycan, involved in repair following tissue damage, through modulating neovascularization and inflammation. In acute myocardial infarction its myocardial expression is up-regulated in a time-dependent manner, and in synd4-deficient mice severe cardiac dysfunction and abnormal remodeling are observed following induction of myocardial infarction. Here we explored the therapeutic potential of sustained synd4 over-expression in the context of myocardial infarction. Adenovirus containing the synd4 gene (Ad-synd4), or corresponding control adenovirus (Ad-null), was administered intramyocardially in rats immediately after induction of myocardial infarction. Cardiac function was ascertained by echocardiography, hemodynamic assessment and brain natriuretic peptide level 28 days post-intervention. Hearts were excised for molecular and histological analyses at predetermined time points. We observed reduced mortality and improved cardiac function post-myocardial infarction in the Ad-synd4 as compared to the Ad-null group, with associated attenuation of cardiac remodeling, less myocyte loss and reduced fibrosis. Additionally, the Ad-synd4 group exhibited endothelial cell activation and increased angiogenesis and arteriogenesis in the myocardium. The Ad-synd4 group also showed evidence of reduced myocardial inflammation as compared with the Ad-null group, with reduced inflammatory cell (CD45+) and myofibroblast (α-SMA+) infiltration as well as suppressed collagen III deposition and iNOS expression. Our results suggest that sustained synd4 over-expression in the myocardium is of therapeutic benefit following experimental myocardial infarction, through inducing neovascularization, suppressing tissue inflammation and fibrosis, with resultant improvements in cardiac function and remodeling.