Objective:To investigate the role of Platycoside E(PE)in a mouse model of bleomycin(BLM)-induced pulmonary fibrosis by targeting the JAK/STAT signaling pathway to suppress macrophage M2 polarization.Methods:Forty BALB/c mice were randomly assigned to five experimental groups:blank control group,model group,pirfenidone(PDF)experimental group,PE high-dose group and PE low-dose experimental group,each with eight mice.After adaptive feeding,except for blank control group,all mice were used in the model of BLM nasal drip-induced pulmonary fibrosis.HE and Masson staining were employed to examine pathological alterations of lung tissue in mice;ELISA to detect concentrations of IL-10,IL-4,IL-17A and TNF-α in mice serum;expressions of CD206 and CD11b in lung tissue were detected by immunofluorescence.Western blot to detect protein expressions of JAK1,p-JAK1,STAT6 and p-STAT6 in lung tissues.Results:Compared with blank control group,tissues in model group showed distorted structure and thickened alveolar walls,fibrotic foci were formed,and alveolar inflammatory fraction and collagen volume fraction were significantly increased(P<0.01).ELISA showed that concentrations of IL-4,IL-10,IL-6 and TNF-α in serum were significantly reduced compared to those of model group.Immunofluorescence showed that fluorescence intensity of CD11b and CD206 treated by PE were decreased significantly.Western blot showed that expressions of JAK1,p-JAK,STAT6 and p-STAT6 proteins were significantly elevated in lung tissues of model mice.Following PE treatment,levels of the above proteins were diminished.Conclusion:PE can effectively improve lung fibrosis induced by BLM in mice,the mechanism may be related to the inhibition of JAK/STAT pathway to block macrophage M2 polarization.

Objective:To investigate the role of Platycoside E(PE)in a mouse model of bleomycin(BLM)-induced pulmonary fibrosis by targeting the JAK/STAT signaling pathway to suppress macrophage M2 polarization.Methods:Forty BALB/c mice were randomly assigned to five experimental groups:blank control group,model group,pirfenidone(PDF)experimental group,PE high-dose group and PE low-dose experimental group,each with eight mice.After adaptive feeding,except for blank control group,all mice were used in the model of BLM nasal drip-induced pulmonary fibrosis.HE and Masson staining were employed to examine pathological alterations of lung tissue in mice;ELISA to detect concentrations of IL-10,IL-4,IL-17A and TNF-α in mice serum;expressions of CD206 and CD11b in lung tissue were detected by immunofluorescence.Western blot to detect protein expressions of JAK1,p-JAK1,STAT6 and p-STAT6 in lung tissues.Results:Compared with blank control group,tissues in model group showed distorted structure and thickened alveolar walls,fibrotic foci were formed,and alveolar inflammatory fraction and collagen volume fraction were significantly increased(P<0.01).ELISA showed that concentrations of IL-4,IL-10,IL-6 and TNF-α in serum were significantly reduced compared to those of model group.Immunofluorescence showed that fluorescence intensity of CD11b and CD206 treated by PE were decreased significantly.Western blot showed that expressions of JAK1,p-JAK,STAT6 and p-STAT6 proteins were significantly elevated in lung tissues of model mice.Following PE treatment,levels of the above proteins were diminished.Conclusion:PE can effectively improve lung fibrosis induced by BLM in mice,the mechanism may be related to the inhibition of JAK/STAT pathway to block macrophage M2 polarization.

AIM:This study examined the inhibitory effect of peiminine on the human colonic adenocarcino-ma cell line SW480 and explored the underlying mechanisms.METHODS:SW480 and human normal colonic epithelial CCD-841CoN cells were treated with different concentrations of peiminine and subjected to the CCK-8 assay to select the optimal treatment time and concentration of the compound.SW480 cell migration and invasion were evaluated by the wound-healing and Transwell assays.Cell cycle progression was analyzed by flow cytometry.The expression levels of cell cycle-related proteins were examined by Western blot.SW480 xenograft tumor model was established in nude mice to ex-amine the effect of peiminine on tumor growth and the expression of cell cycle-related proteins in vivo.RESULTS:Peimi-nine(110 mg/L)significantly inhibited the proliferation of SW480 cells compared with the control group(P<0.01),caused cell cycle arrest at G1 phase,and significantly downregulated the expression of cyclin dependent kinase 2(CDK2),CDK4,CDK6,cyclin D1,p-Rb/Rb,E2F1,E2F3,and E2F4(P<0.05).Peiminine inhibited SW480 xenograft tumor growth,prolonged the survival of model mice,and affected the expression of CDK2,CDK4,CDK6,and cyclin D1 in tu-mor tissues.CONCLUSION:Peiminine promotes G1 phase arrest by down-regulating the expression of CDK2,CDK4,CDK6,and cyclin D1,thereby inhibiting the proliferation of SW480 cells.

AIM:To explore the effect and mechanism of action of the aqueous extract of Fritillaria ussuriensis(FU-AE)against nonalcoholic fatty liver disease(NAFLD).METHODS:The association between Fritillaria ussuriensis Maxir.(FU)and NAFLD was analyzed by network pharmacology.A mouse model of NAFLD was induced in mice by high fat diet(HFD)+10%fructose drinking water,and three doses of Fritillaria ussuriensis aqueous extract were given to the mice for intervention.Colorimetric assay was used for detection of aspartate aminotransferase(AST),alanine aminotrans-ferase(ALT),triglyceride(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C),and low-density lipoprotein cholesterol(LDL-C)levels in the serum of experimental mice.Hematoxylin and eosin staining was used to as-sess the pathological and histological changes in the liver of mice and to clarify the anti-NAFLD effect of aqueous extracts of Fritillaria ussuriensis.Liver tissue proteins were extracted,and expression of proteins related to the AMP-activated pro-tein kinase(AMPK)/acetyl-CoA carboxylase(ACC)pathway was detected by Western blot to clarify the mechanism of an-ti-NAFLD action of Fritillaria ussuriensis.The microbial composition of cecum contents was explored using 16S rRNA se-quencing to reveal the modulatory effect of the aqueous extract of Fritillaria ussuriensis on the structure of intestinal flora in mice with nonalcoholic fatty liver disease.RESULTS:Aqueous extract of Fritillaria ussuriensis(high dose)ameliorated exogenous adipocyte infiltration in the liver of mice with NAFLD(P<0.05).AST,ALT,TG,TC and LDL-C levels were significantly decreased(P<0.05)and HDL-C levels were significantly increased(P<0.05)in the high-dose group.Aque-ous extract of Fritillaria ussuriensis(high dose)significantly increased expression of phosphorylated AMPKα,AMPKα,and phosphorylated ACC in the livers of the model mice(P<0.05),significantly reduced expression of ACC(P<0.05),and significantly increased the relative abundance of the potentially beneficial bacteria Faecalibaculum rodentium,Lacto-bacillus johnsonii,Akkermansia muciniphila(P<0.05).CONCLUSION:Aqueous extract of Fritillaria ussuriensis may ameliorate NAFLD in mice by activating the AMPK/ACC pathway and modulating the structure of intestinal flora.

The E protein of dengue virus type Ⅱ was presented on ferritin of Helicobacter pylori to construct a novel dengue nanoparticle vaccine candidate,and the immunological indexes of the vac-cine were evaluated,aiming to provide new ideas for the development of dengue vaccine.The re-combinant plasmid of E-Ferritin was optimized and synthesized,and then transfected into HEK-293F cells.The recombinant protein was expressed,identified,purified and analyzed.Mice were im-munized with E-Ferritin nanoparticle vaccine by intramuscular injection on the hind limbs on the day 0,14 and 28.ELISA,neutralization test,flow cytometry and lymphocyte proliferation test were used to detect the levels of specific antibodies,neutralizing antibodies,CD3+,CD4+and CD8+T lymphocytes in spleen cells and the proliferation of spleen lymphocytes after specific stimulation.The target protein with a size of about 69 kDa was expressed in the cells with a single band.The purified protein concentration was 0.407 g/L,and the purity was 82.32％.The results from transmission electron microscopy showed that E-Ferritin protein could be recombined into a particle structure with a particle size of about 50 nm.The results of mouse immune experiments showed that E-Ferritin protein had good immunogenicity.The average specific antibody titer of E-ferritin protein in serum was 1∶92 160 after immunization 42 d.The main subclass of antibody was IgGl.The results of flow cytometry showed that E-Ferritin as an immunogen could induce higher levels of CD4+and CD8+T lymphocyte immune response.In lymphocyte proliferation test,the level of specific stimulation in the vaccine group was significantly higher than that in the non-specific stimulation group.In conclusion,the dengue virus envelope protein ferritin nanoparticle vaccine constructed in this study has good immunogenicity,which can provide reference for the de-velopment of new dengue vaccine candidates.

The codon sequence of the S1 protein of the SARS-CoV-2 Beta variant was optimized ac-cording to the preference of CHO cells and cloned into pSN expression vector to construct the re-combinant plasmid pSN-Beta-sl.Recombinant protein was expressed in CHO cells,identified using SDS-PAGE and Western blot,and purified through affinity chromatography.BALB/c mice were immunized with purified recombinant protein Beta-S1 combined with aluminum hydroxide adju-vant.Specific IgG antibody and its subtypes in serum and the cross-neutralization antibody activity against SARS-CoV-2 variants were evaluated.The results showed that the recombinant plasmid pSN-Beta-S1 was successfully constructed,and the recombinant protein Beta-S1 was produced u-sing the CHO cell expression system.The purified recombinant protein had a single band of about 120 kDa with the purity exceeding 85％and can bind to RBD pAb and strep-tag mAb.The recom-binant S1 protein showed good immunogenicity in BALB/c mice.The titers of specific IgG antibodies against RBD protein and S1 protein reached 1∶66 260 and 1∶133 120 on average 21 d after the third immunization,the antibody subtypes were mainly inclined to IgG1.Serum neutrali-zing antibody titer was 1∶629 for wild type,1∶1 720 for Beta,1∶374 for Delta,1∶77 for Omi-cron BA1 and 1∶101 for Omicron BA2.In this study,S1 recombinant protein of SARS-CoV-2 Beta variant was successfully expressed in CHO cell expression system and produced good immunoge-nicity and cross-neutralizing activity in BALB/c mice.These results provide a reference for the fur-ther development of broad-spectrum SARS-CoV-2 vaccine.

The SYBR Green Ⅰ real-time fluorescence quantitative PCR detection method was used to determine the prevalence of Japanese encephalitis virus(JEV)in mosquito vectors and cattle se-rum samples in Xinjiang.The E gene fragment of the JEV strain was amplified by PCR,cloned into a pEASY-Blunt vector,produced as a recombinant plasmid,and its sensitivity,specificity and re-producibility were verified.Between 2021 and 2022,serum samples were taken in the regions of Hami,Altay,Ili,Aksu,and Kashi in order to monitor the prevalence of JEV in livestock in Xin-jiang.The positive rate was discovered and evaluated using the established detection method.The established detection method showed a good linear relationship,and the detection interval was 4.03X102-4.03×109 copies/pL.The correlation coefficient was 0.995,the slope was-3.431,and the extreme value of the lower limit of sensitivity was 4.03 × 102 copies/pL.This method has no specific amplification for Zika virus(ZIKV)and Dengue virus(DENV).The intra group coefficient of variation of reproducibility was 0.53％-1.27％,and the inter group coefficient of variation was 0.48％-1.43％.Using this method to detect serum samples from livestock in Xinjiang from 2021 to 2022,the total positive rate was 3.28％,with positive detection rates in horses,cows,and sheep being 2.35％,6.77％,and 3.74％respectively,the virus was identified as Type Ⅰ JEV.A SYBR Green Ⅰ real-time PCR method for the detection of genotype 1 JEV was established.JEV was de-tected in the serum of horses,cattle and sheep in Xinjiang,with a total positive rate of 3.11％.

Objective To construct a recombinant poxvirus vector vaccine, rVTTδTK-RBD, and to evaluate its safety and immunogenicity. Methods The receptor-binding domain (RBD) gene was synthesized with reference to the gene sequence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and was inserted into the polyclonal site of the self-constructed recombinant plasmid pSTKE, to construct the recombinant poxvirus shuttle vector pSTKE-RBD. This was then transfected into BHK-21 cells pre-infected with the vaccinia virus Tiantan strain (VTT). The recombinant poxvirus rVTTδTK-RBD was successfully obtained after several rounds of fluorescence phage screening. The effect of rVTTδTK-RBD on the body mass of BALB/c mice was detected after immunizing mice by intra-nasal vaccination. The levels of specific and neutralizing antibodies produced by rVTTδTK-RBD on BALB/c mice were analyzed after immunizing mice intramuscularly. The effect of rVTTδTK-RBD on T cell subsets in BALB/c mice was detected by flow cytometry. Results Through homologous recombination, enhanced green fluorescent protein (EGFP) screening marker, and multiple rounds of fluorescent phosphorescence phage screening, a recombinant poxvirus rVTTδTK-RBD, expressing RBD with deletions in the thymidine kinase (TK) gene, was successfully obtained, which was validated by PCR. The in vivo experiments on BALB/c mice showed that rVTTδTK-RBD was highly immunogenic against SARS-CoV-2 and significantly reduced toxicity to the body compared to the parental strain VTT. Conclusion The recombinant poxvirus vaccine rVTTδTK-RBD against SARS-CoV-2 is successfully constructed and obtained, with its safety and immunogenicity confirmed through various experiments.
Animals ; Mice ; SARS-CoV-2/genetics* ; COVID-19 ; Vaccines, Synthetic/genetics* ; Genes, Reporter ; Bacteriophages ; Mice, Inbred BALB C

As a potential vectored vaccine,Newcastle disease virus(NDV)has been subject to various studies for vaccine development,while relatively little research has outlined the immunomodulatory effect of the virus in antigen presentation.To elucidate the key inhibitory factor in regulating the interaction of infected dendritic cells(DCs)and T cells,DCs were pretreated with the NDV vaccine strain LaSota as an inhibitor and stimulated with lipopolysaccharide(LPS)for further detection by enzyme-linked immunosorbent assay(ELISA),flow cytometry,immunoblotting,and quantitative real-time polymerase chain reaction(qRT-PCR).The results revealed that NDV infection resulted in the inhibition of interleukin(IL)-12p40 in DCs through a p38 mitogen-activated protein kinase(MAPK)-dependent manner,thus inhibiting the synthesis of IL-12p70,leading to the reduction in T cell proliferation and the secretion of interferon-γ(IFN-γ),tumor necrosis factor-α(TNF-α),and IL-6 induced by DCs.Consequently,downregulated cytokines accelerated the infection and viral transmission from DCs to T cells.Furthermore,several other strains of NDV also exhibited inhibitory activity.The current study reveals that NDV can modulate the intensity of the innate?adaptive immune cell crosstalk critically toward viral invasion improvement,highlighting a novel mechanism of virus-induced immunosuppression and providing new perspectives on the improvement of NDV-vectored vaccine.

Exosomes (EXOs) are formed by intracellular multivesicular bodies and carry a variety of biomacromolecules such as lipids, proteins, encoding and non-coding RNAs, and mitochondrial DNA. EXOs can be released in vivo by different cell types, including hepatocytes, hepatic stellate cells, and immune cells and play the role of intercellular communication. More and more studies have shown that EXOs are involved in the development, progression, and prognosis of chronic hepatitis B (CHB) and hepatocellular carcinoma (HCC) caused by hepatitis B virus (HBV) infection and are expected to become potential biomarkers for the early diagnosis and prognostic evaluation of HBV-related HCC. This article reviews the role of EXOs in the host infection process of HBV and the importance of EXOs in the development, progression, and prognosis of CHB and HCC, in order to provide new ideas for the basic and clinical research in this field.