[Construction of a replicative expression vector based on the porcine circovirus 2 replicon].

The antigen gene expression level of a DNA vaccine is the key factor influencing the efficacy of the DNA vaccine. Accordingly, one of the ways to improve the antigen gene expression level of a DNA vaccine is to utilize a plasmid vector that is replicable in eukaryotic cells. A replicative DNA vaccine vector pCMVori was constructed based on the non-replicative pcDNA3.1 and the replicon of porcine circovirus 2 (PCV2) in this study. An EGFP gene was cloned into pCMVori and the control plasmid pcDNA3.1. The two recombinant vectors were transfected into PK-15 cell, and the plasmid DNA and RNA were extracted from the transfected cells. Real-time PCR was used to determine the plasmid replication efficiency of the two plasmids using plasmid before and after Bcl Ⅰ digestion as templates, and the transcription level of the Rep gene in PCV2 replicon was detected by RT-PCR. The average fluorescence intensity of cells transfected with the two plasmids was analyzed with software Image J, and the transcription level of EGFP was determined by means of real-time RT-PCR. The results showed that the replication efficiency of pCMVori in PK-15 cells incubated for 48 h was 136%, and the transcriptions of Rep and Rep' were verified by RT-PCR. The average fluorescence intensity of the cells transfected with pCMVori-EGFP was 39.14% higher than that of pcDNA3.1-EGFP, and the transcription level of EGFP in the former was also 40% higher than that in the latter. In conclusion, the DNA vaccine vector pCMVori constructed in this study can independently replicate in eukaryotic cells. As a result, the expression level of cloned target gene was elevated, providing a basis for developing the pCMVori-based DNA vaccine.

Potential pharmacological mechanisms of four active compounds of Macleaya cordata extract against enteritis based on network pharmacology and molecular docking technology.

Background: Macleaya cordata extract (MCE) is effective in the treatment of enteritis, but its mechanism has not been fully elucidated. Therefore, this study combined network pharmacology and molecular docking technologies to investigate the potential pharmacological mechanism of MCE in the treatment of enteritis. Methods: The information of active compounds in MCE was accessed through the literature. Furthermore, PubChem, PharmMapper, UniProt, and GeneCards databases were used to analyze the targets of MCE and enteritis. The intersection of drug and disease targets was imported into the STRING database, and the analysis results were imported into Cytoscape 3.7.1 software to construct a protein-protein interaction (PPI) network and to screen core targets. The Metascape database was used for conducting Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. AutoDock Tools software was used for the molecular docking of active compounds with the core targets. Results: MCE has four active compounds, namely, sanguinarine, chelerythrine, protopine, and allocryptopine, and a total of 269 targets after de-duplication. Furthermore, a total of 1,237 targets were associated with enteritis, 70 of which were obtained by aiding the drug-disease intersection with the aforementioned four active compound targets of MCE. Five core targets including mitogen-activated protein kinase 1 (MAPK1) and AKT serine/threonine kinase 1 (AKT1) were obtained using the PPI network, which are considered the potential targets for the four active compounds of MCE in the treatment of enteritis. The GO enrichment analysis involved 749 biological processes, 47 cellular components, and 64 molecular functions. The KEGG pathway enrichment analysis revealed 142 pathways involved in the treatment of enteritis by the four active compounds of MCE, among which PI3K-Akt and MAPK signaling pathways were the most important pathways. The results of molecular docking showed that the four active compounds demonstrated good binding properties at the five core targets. Conclusion: The pharmacological effects of the four active compounds of MCE in the treatment of enteritis involve acting on signaling pathways such as PI3K-Akt and MAPK through key targets such as AKT1 and MAPK1, thus providing new indications for further research to verify its mechanisms.

Programmed-response cross-linked nanocarrier for multidrug-resistant ovarian cancer treatment.

The application of numerous chemotherapeutic drugs has been limited due to poor solubility, adverse side effects, and even multidrug resistance in patients. Polymeric micelles with reversibly cross-linked structures provide a promising solution to these issues. Herein, we optimized and synthesized programable-released disulfide cross-linked micelle (PDCM) based on our previous well-defined dendrimers to deliver the antitumor drug betulinic acid (BA) and paclitaxel (PDCM@PTX) and evaluated the therapeutic efficacy of multidrug-resistant (MDR) simulative orthotopic intraperitoneal ovarian cancer mice models. Comprehensive results demonstrated that PDCM@PTX formed stable nanoparticles able to improve the pharmacokinetic profile and circulation time of PTX, allowing for increased tumor penetration. Furthermore, in the tumor microenvironment, the programable-switches (ester bonds and disulfide cross-linking) of PDCM@PTX were cleaved by the high concentration of glutathione (tumor microenvironment) and esterase (intracellular) present in the tumor, allowing for in situ release of PTX and BA, resulting in intensive therapeutic efficacy in MDR ovarian cancer.

Effects of Schisandra chinensis Polysaccharide-Conjugated Selenium Nanoparticles on Intestinal Injury in Mice.

Schisandra chinensis polysaccharide (SCP) is an experimental therapeutic for the treatment of intestinal injury. Selenium nanoparticle modification can improve the bioactivity of polysaccharides. In this study, SCP was firstly extracted and purified by a DEAE-52 column, then SCP-Selenium nanoparticles (SCP-Se NPs) were prepared, and the procedure was optimized. Thereafter, the obtained SCP-Se NPs were characterized by transmission electron microscope, X-ray diffraction, energy-dispersive X-ray spectroscopy, and Fourier transform infrared spectroscopy. The influence of different storage environments on the stability of colloidal SCP-Se NPs was also investigated. Finally, the therapeutic effects of SCP-Se NPs on LPS-induced intestinal inflammatory injuries in mice were evaluated. Results showed that the optimized SCP-Se NPs were amorphous, uniform, spherical particles with a diameter of 121 nm, and the colloidal solution was stable at 4 °C for at least 14 d. Moreover, SCP-Se NPs could more effectively alleviate LPS-induced diarrhea, intestinal tissue injury, and tight junction destruction and decrease the elevated expression levels of TNF-α, IL-1ß, and IL-6 compared with SCP. These results demonstrate that SCP-Se NPs may alleviate LPS-induced enteritis through their anti-inflammatory effects, indicating that SCP-Se NPs can serve as a good candidate for preventing and treating enteritis in the livestock and poultry industry.

Enhancement of intestinal mucosal immunity and immune response to the foot-and-mouth disease vaccine by oral administration of danggui buxue decoction.

This study investigated the effect of Danggui Buxue decoction (DBD) on the immunity of an O-type foot-and-mouth disease (FMD) vaccine and intestinal mucosal immunity. SPF KM mice were continuously and orally administered DBD for 5 d and then inoculated with an O-type FMD vaccine. The contents of a specific IgG antibody and its isotypes IgG1, IgG2a, IgG2b, and IgG3 in serum and SIgA in duodenal mucosa were determined by ELISA at 1 and 3 W after the 2nd immunization. qRT-PCR was used to detect mRNA expression levels of IL-4, IL-10, IFN-γ, and IL-33 in the spleen, and mRNA expression levels of J-chain, pIgR, BAFF, APRIL, IL-10, IFN-γ and IL-33 in the duodenum. The results showed that compared with the control group, oral administration of DBD significantly increased levels of the anti-FMD virus (FMDV)-specific antibodies IgG, IgG1, and IgG2a in the serum of O-type FMD vaccine-immunized mice 1 W after the 2nd immunization (P < 0.05), upregulated mRNA expression levels of spleen lymphocyte cytokines IL-4 and IL-33 (P < 0.05), promoted the secretion of SIgA in duodenal mucosa (P < 0.05). The mRNA expression levels of J-chain, pIgR, BAFF, APRIL, IL-10, and IL-33 in duodenal tissues were upregulated (P < 0.05). This study indicates that DBD has a good promotion effect on the O-type FMD vaccine and the potential to be an oral immune booster.

Accelerating supramolecular aggregation by molecular sliding.

Diffusion-based translocation along DNA or RNA molecules is essential for genome regulatory proteins to execute their biological functions. The reduced dimensionality of the searching process makes the proteins bind specific target sites at a "faster-than-diffusion-controlled rate". We herein report a photoresponsive slider-track diffusion system capable of self-assembly rate acceleration, which consists of (-)-camphorsulfonic acid, 4-(4'-n-octoxylphenylazo)benzenesulfonic acid, and isotactic poly(2-vinylpyridine). The protonated pyridine rings act as the footholds for anionic azo sliders to diffusively bind and slide along polycationic tracks via electrostatic interactions. Ultraviolet light triggers the trans to cis isomerization and aggregation of azo sliders, which can be monitored by multiple spectroscopic methods without labeling. The presence of vinyl polymer track increases the aggregation rate of cis azobenzene up to ∼20 times, depending on the stereoregularity of the polymer chain, the acid/base ratio and the addition of salt. This system has a feature of simplicity, monitorability, controllability, and could find applications in designing molecular machines with desired functionalities.

Efficient breeding of low glutelin content rice germplasm by simultaneous editing multiple glutelin genes via CRISPR/Cas9.

Chronic kidney disease (CKD) and phenylketonuria (PKU) patients need to eat rice with low glutelin content. Therefore, breeding low glutelin content rice varieties with high yield and delicious taste is one of the major goals of rice breeders due to the high demand for the product. In this study, we designed three sgRNAs targeting nine glutelin genes and generated nine T-DNA-free homozygous editing lines with reduced glutelin content compared with the wild-type due to simultaneous mutation(s) in 5-7 glutelin genes. The glutelin content of two lines is even significantly lower than that of the low glutelin content cultivar, LGC-1. Compared to the wild-type, these low glutelin lines showed similar agronomic traits, including yield components and viscosity properties, and can be used as new varieties or parental materials for further breeding.

Self-Reporting Activated Ester-Amine Reaction for Enantioselective Multi-Channel Visual Detection of Chiral Amines.

Chiral recognition is of importance not only in living systems but also in estimating the optical purity of enantiomeric drugs and fabricating advanced materials. Herein we report a novel self-reporting activated ester-amine reaction that can provide multi-channel visual detection of organic amines. It relies on the reaction extent dependent cis-transoid to cis-cisoid helical transition of the polyphenylacetylene backbone and the thus triggered fluorescence. Owing to the high selectivity, this visual process can recognize structurally diverse achiral amines and quantitatively check the impurity content. It also shows an outstanding enantioselectivity towards various chiral amines and can be applied to determine enantiomeric composition. The multiple responses in absorption, circular dichroism, photoluminescence, and circularly polarized luminescence make the helical transition of the polymer backbone a potential detection mode for high-throughput screening of chiral chemicals.

Development of a Simple Single-Acupoint Electroacupuncture Frame and Evaluation of the Acupuncture Effect in Rabbits.

To reduce the circulation path of the output current of traditional electroacupuncture (TEA) process in the body, a simple single-acupoint electroacupuncture (SEA) frame was designed and the acupuncture effect of SEA was evaluated through Hou-san-li (ST-36) and Qian-san-li (LI-10) acupoints. Forty-two healthy New Zealand rabbits were randomly divided into seven groups and underwent acupuncture for 20 min in an awake state. Blood samples aseptically collected from the ear vein 3 h before acupuncture and 0, 3, 6, 9, 12 and 24 h after acupuncture were used for the detection of aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase MB (CK-MB) and motilin (MTL) in serum. The simple SEA frame was developed successfully, and the acupuncture results showed that the serum AST and ALT levels were significantly higher at 3 h after TEA with high frequency (p < 0.01) compared with the control group. Regarding serum CK-MB levels, no significant differences were found after SEA or TEA stimulation (p > 0.05). Serum MTL levels were significantly increased at 0 h after SEA and TEA (p < 0.05), but there were no significant differences at other time points after SEA and TEA treatment (p > 0.05). SEA not only maintains the effect of TEA but also shortens the circulation loop of the electroacupuncture (EA) current in the body, which effectively avoids body injury.

Effects of a novel anti-biofilm peptide CRAMP combined with antibiotics on the formation of Pseudomonas aeruginosa biofilms.

The remarkable ability of Pseudomonas aeruginosa to form biofilms renders antibiotic treatments inefficient and therefore causing a wide variety of chronic infections. The quorum sensing (QS) system in P. aeruginosa plays a role in the regulation of genes controlling virulence factors and biofilm formation, which may be an essential target for pharmacological intervention. The present study aimed to investigate the synergistic activity of sub-MIC concentrations of CRAMP (a cathelicidin-related antimicrobial peptide) with fourteen antibiotics against P. aeroginusa biofilms. Finally, CRAMP's best synergistic activity combined with colistin at 1/4 MIC was screened by the checkerboard method and the calculation of the synergetic coefficient. It was confirmed by experiments on 6-well plates, displaying the most significant biofilm formation inhibition % (91.05%, calculated by OD value of biofilm biomass) and the best bactericidal activity of biofilms (2.77-log10 decrease). These data correlate with the confocal laser scanning microscopy (CLSM) images obtained for the biofilm. The combination also down-regulated the expression of QS regulated genes, resulting in inhibitory effects on QS-regulated virulence phenotypes (pyocyanin and rhamnolipid). These results indicate that a proposed method of combination therapy of CRAMP with colistin has the potential to serve as a more effective therapy for P. aeruginosa biofilm infection.

The inhibitory effect of phosphorylated Codonopsis pilosula polysaccharide on autophagosomes formation contributes to the inhibition of duck hepatitis A virus replication.

Duck hepatitis A virus type 1 (DHAV) infection causes duck viral hepatitis and results in enormous loss to poultry farming industry. We reported that phosphorylated Codonopsis pilosula polysaccharide (pCPPS) inhibited DHAV genome replication. Here we further explored its underlying antiviral mechanisms. Autophagosomes formation is essential for the genome replication of picornaviruses. In this study, Western blot, confocal microscopy observation, and ELISA methods were performed to analyze polysaccharides' effects on autophagy by the in vitro and in vivo experiments. Results obtained from in vitro and in vivo experiments showed that Codonopsis pilosula polysaccharide did not play a role in regulating autophagy and had no therapeutic effects on infected ducklings. However, pCPPS treatment downregulated LC3-II expression level activated by DHAV and rapamycin, indicating the inhibition of autophagosomes formation. The interdiction of autophagosomes formation resulted in the inhibition of DHAV genome replication. Further study showed that pCPPS treatment reduced the concentration of phosphatidylinositol-3-phosphate (PI3P), an important component of membrane, in cells and serum, and consequently, autophagosomes formation was downregulated. In vivo experiments also verified the therapeutic effect of pCPPS. Phosphorylated Codonopsis pilosula polysaccharide treatment increased the infected ducklings' survival rate and alleviated hepatic injury. Our studies verified the effects of pCPPS against DHAV infection in duck embryo hepatocytes and ducklings and confirmed that phosphorylated modification enhanced the bioactivities of polysaccharides. The results also stated pCPPS's antiviral mechanisms, provided fundamental basis for the development of new anti-DHAV agents.

Sequential Targeting in Crosslinking Nanotheranostics for Tackling the Multibarriers of Brain Tumors.

The efficacy of therapeutics for brain tumors is seriously hampered by multiple barriers to drug delivery, including severe destabilizing effects in the blood circulation, the blood-brain barrier/blood-brain tumor barrier (BBB/BBTB), and limited tumor uptake. Here, a sequential targeting in crosslinking (STICK) nanodelivery strategy is presented to circumvent these important physiological barriers to improve drug delivery to brain tumors. STICK nanoparticles (STICK-NPs) can sequentially target BBB/BBTB and brain tumor cells with surface maltobionic acid (MA) and 4-carboxyphenylboronic acid (CBA), respectively, and simultaneously enhance nanoparticle stability with pH-responsive crosslinkages formed by MA and CBA in situ. STICK-NPs exhibit prolonged circulation time (17-fold higher area under curve) than the free agent, allowing increased opportunities to transpass the BBB/BBTB via glucose-transporter-mediated transcytosis by MA. The tumor acidic environment then triggers the transformation of the STICK-NPs into smaller nanoparticles and reveals a secondary CBA targeting moiety for deep tumor penetration and enhanced uptake in tumor cells. STICK-NPs significantly inhibit tumor growth and prolong the survival time with limited toxicity in mice with aggressive and chemoresistant diffuse intrinsic pontine glioma. This formulation tackles multiple physiological barriers on-demand with a simple and smart STICK design. Therefore, these features allow STICK-NPs to unleash the potential of brain tumor therapeutics to improve their treatment efficacy.

Assessment of the hepatocyte protective effects of gypenoside and its phosphorylated derivative against DHAV-1 infection on duck embryonic hepatocytes.

BACKGROUND: Duck viral hepatitis (DVH) is an acute disease of young ducklings with no effective veterinary drugs for treatment. Gynostemma pentaphyllum is a well-known traditional Chinese medicine that plays an important role in the treatment of various diseases. Gypenoside (GP), one of the main ingredients of Gynostemma pentaphyllum, was reported with good hepatoprotective effects. However, its low solubility limits its application in the clinics. To improve its solubility and bioactivity, a phosphorylated derivative of gypenoside (pGP) was prepared by the sodium trimetaphosphate-sodium tripolyphosphate (STMP-STPP) method. An infrared spectroscopy method was applied to analyse the structures of GP and pGP. Then, a methyl thiazolyl tetrazolium (MTT) colorimetric assay was applied to study the hepatocyte protective efficacy of these two drugs against duck hepatitis A virus type 1 (DHAV-1) infection, and qPCR, TUNEL labelling and flow cytometry methods were used to study the relevant hepatocyte protective in vitro. RESULTS: The infrared spectroscopy detection results showed that the phosphorylation modification of GP was successful. The MTT colorimetric assay results showed that both GP and pGP possessed good hepatocyte protective efficacy in vitro, and pGP performed better than GP when the drug was added before or after virus inoculation. Furthermore, the qPCR results revealed that both drugs could effectively inhibit the adsorption (when adding GP and pGP pre-virus inoculation), replication and release of DHAV-1, and the viral inhibition rate of pGP was greater than that of GP. The subsequent TUNEL labelling and flow cytometry assays showed that both GP and pGP could significantly inhibit duck embryo hepatocyte apoptosis induced by DHAV-1, and the inhibition effect of pGP was much stronger than that of GP. CONCLUSIONS: GP exerts good hepatocyte protective efficacy not only by inhibiting the proliferation of DHAV-1 but also by inhibiting duck embryonic hepatocyte apoptosis induced by DHAV-1, and phosphorylation modification significantly improves the antiviral and the anti-apoptotic effects of GP. Therefore, pGP has the potential to be developed into a novel drug against DHAV-1 infection.

Evaluation of the Therapeutic Effect of a Flavonoid Prescription against Rabbit Hemorrhagic Disease In Vivo.

Rabbit hemorrhagic disease (RHD) is an acute, high fatal contagious disease induced by rabbit hemorrhagic disease virus (RHDV) with acute severe hepatic injury and causes huge economic loss worldwide. In order to develop an effective and reliable drug to treat this disease in clinic, a prescription formulated with baicalin, linarin, icariin, and notoginsenoside R1 (BLIN) according to the theory of syndrome differentiation and treatment in traditional Chinese veterinary medicine was applied to investigate its curative effects against RHD in vivo. The preliminary study results showed that BLIN prescription exerted good curative effect on RHD therapy. To further validate the curative effect and to investigate the possible related curative mechanisms of this drug, the survival rates, the plasma biochemical indexes of hepatic function, the plasma evaluation indexes of oxidative injury, and the RHDV gene expression levels were detected and then the correlation among these indexes was also analyzed. These results showed that BLIN prescription could significantly increase the survival rate, reduce the hepatic injury severity, alleviate the oxidative injury, and decrease the RHDV gene expression level in rabbits infected with RHDV. All these results indicate that BLIN prescription possesses outstanding curative effect against RHD, and the curative mechanism may be related to its antioxidant and anti-RHDV activities. Therefore, this prescription can be expected to be exploited into a new candidate for RHD therapy in clinic.

PI3KC3-dependent autophagosomes formation pathway is of crucial importance to anti-DHAV activity of Chrysanthemum indicum polysaccharide.

We previously reported that Chrysanthemum indicum polysaccharide (CIPS) effectively inhibited the replication of duck hepatitis A virus (DHAV). However, the inhibition mechanisms are still unclear. Autophagy plays important role in virus genomic replication. Therefore, in present study, the effect of autophagy on DHAV genome replication as well as the influence of CIPS on autophagy were studied. qPCR, western blot, and ELISA methods were applied to observe the autophagy and analyze the inhibition mechanisms of CIPS on DHAV. Results showed that DHAV infection increased the expression level of LC3-II and interdicted the degradation of p62. Treating with rapamycin benefited DHAV gene expression level. What's more, DHAV infection and rapamycin treatment also promoted the expression of PI3KC3 and increased the concentration of PI3P. However, CIPS treatment significantly downregulated the expressions of LC3-II and PI3KC3 induced by DHAV and rapamycin, and consequently inhibited autophagosomes formation. As a result, DHAV replication was inhibited.

Baicalin Inhibits Biofilm Formation and the Quorum-Sensing System by Regulating the MsrA Drug Efflux Pump in Staphylococcus saprophyticus.

Staphylococcus saprophyticus (S. saprophyticus) is one of the main pathogens that cause serious infection due to its acquisition of antibiotic resistance. The efflux pump decreases antibiotic abundance, and biofilm compromises the penetration of antibiotics. It has been reported that baicalin is a potential agent to inhibit efflux pumps, biofilm formation, and quorum-sensing systems. The purpose of this study was to investigate whether baicalin can inhibit S. saprophyticus biofilm formation and the quorum-sensing system by inhibiting the MsrA efflux pump. First, the mechanism of baicalin inhibiting efflux was investigated by the ethidium bromide (EtBr) efflux assay, measurement of ATP content, and pyruvate kinase (PK) activities. These results revealed that baicalin significantly reduced the efflux of EtBr, the ATP content, and the activity of PK. Moreover, its role in biofilm formation and the agr system was studied by crystal violet staining, confocal laser scanning microscopy, scanning electron microscopy, and real-time polymerase chain reaction. These results showed that baicalin decreased biofilm formation, inhibited bacterial aggregation, and downregulated mRNA transcription levels of the quorum-sensing system regulators agrA, agrC, RNAIII, and sarA. Correlation analysis indicated that there was a strong positive correlation between the efflux pump and biofilm formation and the agr system. We demonstrate for the first time that baicalin inhibits biofilm formation and the agr quorum-sensing system by inhibiting the efflux pump in S. saprophyticus. Therefore, baicalin is a potential therapeutic agent for S. saprophyticus biofilm-associated infections.

In vitro synergistic effect of baicalin with azithromycin against Staphylococcus saprophyticus isolated from francolins with ophthalmia.

Francolins ophthalmia is often caused by resistant conditional pathogenic bacteria. Conditional pathogenic Staphylococcus saprophyticus is a potential reservoir of macrolides antibiotics resistance gene. Baicalin has been reported as a potential agent to synergistically inhibit the replication of Staphylococcus. The objective of this study was to isolate the pathogen of the francolins ophthalmia, identify the antibiotic resistance profile of isolated S. saprophyticus, and investigate the effect of baicalin combined with azithromycin (Azm) against azithromycin resistant S. saprophyticus (ARSS). The ARSS was isolated and identified from francolins suffered from ophthalmia by phenotypic and molecular biology methods. The antibiotic resistance profile was identified by Kirby-Bauer method. Then the minimal inhibitory concentration (MIC) of Azm in absence and presence of a sub-inhibitory concentration baicalin/verapamil was determined to assess the effect that baicalin combined with Azm against ARSS. ARSS was isolated and identified from francolins experienced ophthalmia. The isolated ARSS was resistant to 11 among the 13 antibiotics that were tested. The synergistic effect of baicalin and Azm was noticed with a reduction rate varied from 2 to 128-fold. It appears from this study that S. saprophyticus can cause francolins ophthalmia and baicalin may be used as a natural agent resistance inhibitor for ARSS.

Phosphorylation of Icariin Can Alleviate the Oxidative Stress Caused by the Duck Hepatitis Virus A through Mitogen-Activated Protein Kinases Signaling Pathways.

The duck virus hepatitis (DVH) caused by the duck hepatitis virus A (DHAV) has produced extensive economic losses to the duck industry. The currently licensed commercial vaccine has shown some defects and does not completely prevent the DVH. Accordingly, a new alternative treatment for this disease is urgently needed. Previous studies have shown that icariin (ICA) and its phosphorylated derivative (pICA) possessed good anti-DHAV effects through direct and indirect antiviral pathways, such as antioxidative stress. But the antioxidant activity showed some differences between ICA and pICA. The aim of this study is to prove that ICA and pICA attenuate oxidative stress caused by DHAV in vitro and in vivo, and to investigate their mechanism of action to explain their differences in antioxidant activities. In vivo, the dynamic deaths, oxidative evaluation indexes and hepatic pathological change scores were detected. When was added the hinokitiol which showed the pro-oxidative effect as an intervention method, pICA still possessed more treatment effect than ICA. The strong correlation between mortality and oxidative stress proves that ICA and pICA alleviate oxidative stress caused by DHAV. This was also demonstrated by the addition of hydrogen peroxide (H2O2) as an intervention method in vitro. pICA can be more effective than ICA to improve duck embryonic hepatocytes (DEHs) viability and reduce the virulence of DHAV. The strong correlation between TCID50 and oxidative stress demonstrates that ICA and pICA can achieve anti-DHAV effects by inhibiting oxidative stress. In addition, the superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of ICA and pICA showed significant difference. pICA could significantly inhibit the phosphorylation of p38, extra cellular signal regulated Kinase (ERK 1/2) and c-Jun N-terminal kinase (JNK), which were related to mitogen-activated protein kinases (MAPKs) signaling pathways. Ultimately, compared to ICA, pICA exhibited more antioxidant activity that could regulate oxidative stress-related indicators, and inhibited the phosphorylation of MAPKs signaling pathway.

RAW REHMANNIA RADIX POLYSACCHARIDE CAN EFFECTIVELY RELEASE PEROXIDATIVE INJURY INDUCED BY DUCK HEPATITIS A VIRUS.

BACKGROUND: Duck viral hepatitis (DVH), caused by duck hepatitis A virus (DHAV), is a fatal contagious infectious disease which spreads rapidly with high morbidity and high mortality, and there is no effective clinical drug against DVH. MATERIALS AND METHODS: Raw Rehmannia Radix Polysaccharide (RRRP), Lycii Fructus polysaccharides and Astragalus Radix polysaccharides were experimented in vitro and in vivo. Mortality rate, livers change, liver lesion scoring, peroxidative injury evaluation indexes in vitro and in vivo, and hepatic injury evaluation indexes of optimal one were detected and observed in this experiment. RESULTS: RRRP could reduce mortality with the protection rate about 20.0% compared with that of the viral control (VC) group, finding that RRRP was the most effective against DHAV. The average liver scoring of the VC, blank control (BC), RRRP groups were 3.5, 0, 2.1. Significant difference (P<0.05) appeared between any two groups, demonstrating that it can alleviate liver pathological change. RRRP could make the hepatic injury evaluation indexes similar to BC group while the levels of the VC group were higher than other two groups in general. The levels of SOD, GSH-Px, CAT of RRRP group showed significant higher than that of VC group while the levels of NOS and MDA showed the opposite tendency, thus, RRRP could release peroxidative injury. CONCLUSION: RRRP was the most effective against duck hepatitis A virus (DHAV). RRRP could reduce mortality, alleviate liver pathological change, down-regulate liver lesion score, release peroxidative injury and hepatic injury. The antiviral and peroxidative injury releasing activity of RRRP for DHAV provided a platform to test novel drug strategies for hepatitis A virus in human beings.

A flavone-polysaccharide based prescription attenuates the mitochondrial dysfunction induced by duck hepatitis A virus type 1.

The principal target organ of duck hepatitis A virus type 1 (DHAV-1) is duckling liver, which is an energy-intensive organ and plays important roles in body's energy metabolism and conversion. As the "power house" of the hepatocytes, mitochondria provide more than 90% of the energy. However, mitochondria are much vulnerable to the oxidative stress for their rich in polyunsaturated fatty acids. Although previous researches have demonstrated that DHAV-1 could induce the oxidative stress in the serum of the infected ducklings, no related study on the mitochondria during the pathological process of DVH has been reported by far. To address this issue, we examined the HE stained tissue pathological slices, detected the hepatic SOD, CAT and GPX activities and MDA contents and analyzed the ATP content, mitochondrial ultrastructure and the mitochondrial SOD, GPX activities and MDA content in the liver tissues. The results showed that the hepatic redox status was significantly disturbed so that causing the mitochondrial dysfunction, ATP depletion and mitochondrial oxidative stress during the process of the DHAV-1 infection, and a prescription formulated with Hypericum japonicum flavone, Radix Rehmanniae Recens polysaccharide and Salvia plebeia flavone (HRS), which had been demonstrated with good anti-oxidative activity in serum, could effectively alleviate the hepatic injury and the oxidative stress in liver tissue induced by DHAV-1 thus alleviating the mitochondrial injury and oxidative stress. In a word, this research discovers the oxidative stress induced mitochondrial dysfunction and oxidative stress during the DVH pathological process and demonstrates HRS exerts good anti-oxidative activity in liver tissue to protect mitochondria against reactive oxygen species (ROS).