Purpose To explore the application value of spectral CT vascular optimization technique for laparoscopic right hemicolectomy.Materials and Methods Ninety patients who underwent contrast-enhanced CT with spectral CT before scheduled laparoscopic right hemicolectomy from March to November 2022 in Shengjing Hospital of China Medical University were retrospectively analyzed.The raw images were reconstructed to obtain 120 kVp conventional images and spectral-based virtual monochromatic images(VMIs)at 40,50,60,70 keV.The CT value and image noise of gastrocolic trunk of Henle(GTH),right colic vein(RCV)and ileocolic vein(ICV)were measured.The CT value,image noise,signal noise ratio and contrast noise ratio of VMIs and conventional image were analyzed.The image quality and venous anatomical information of each group were evaluated subjectively with three-dimensional reconstruction technique.Results The CT value[(405.55±103.06)Hu,(406.61±103.42)Hu,402.35(337.95,477.45)Hu],image noise[19.90(12.10,34.55)Hu,21.10(10.95,44.30)Hu,19.70(11.25,44.80)Hu],signal noise ratio[19.46(10.60,36.31),20.16(8.73,37.42),18.49(9.99,36.24)]and contrast noise ratio[53.36(35.92,70.79),52.34(33.73,81.03),56.23(34.69,79.97)]of GTH,RCV,ICV of VMIs at 40 keV were significantly higher than those of conventional image and other VMIs(F/H=268.53,23.17,33.09,198.77,272.51,28.93,25.48,189.62,318.24,29.42,20.13,212.46,all P<0.001).Subjective scores showed that the image quality and vascular visualization of GTH,RCV and ICV at 40 keV were the best.Conclusion Spectral CT can improve the imaging quality and vascular visualization of GTH,RCV and ICV.VMIs at 40 keV has the optimal performance.

To construct a subgenomic replication subsystem of foot-and-mouth disease virus(FM-DV)carrying fluorescent protein(UnaG),the full-length cDNA plasmid of FMDV was employed as the template.A few of structural and non-structural protein genes of FMDV were removed by double-enzyme digestion.By substituting reporter gene sequences expressing the green fluorescent protein UnaG for specific structural protein sequences of FMDV,FMDV-UnaG replicators were successfully created.After PCR and sequencing,linearized FMDV-UnaG replicons were transfected into BSR/T7 cells expressing T7 RNA polymerase,and the fluorescence signal was observed through fluorescence microscopy and laser confocal technique.The results demonstrated that the constructed FMDV-UnaG replicons could effectively express UnaG protein,and the protein colo-calized with FMDV 3A protein.Additionally,Western blot and RT-qPCR also detected that the replicator RNA could express the non-structural proteins of the virus and replicate autonomously in BSR/T7 cells,respectively.In conclusion,the successful construction of FMDV-UnaG sub-genomic replicators offers a favorable tool for further research on the replication and translation mechanism of FMDV and the development of vaccine vectors.

The objective of this study was to evaluate the protective effect and possible related mechanisms of Sophora subprostrate polysaccharide(SSP)on intestinal epithelial cell injury in-duced by Tert-Butyl hydroperoxide(TBHP).The optimal dose of TBHP and the safe concentra-tion range of SSP were determined using the MTT method.In this study,IPEC-J2 cells were divid-ed into five groups:the control group,the model group,the SSPL group,the SSPM group and the SSPH group,and the cell morphology,cell survival rate and LDH release rate were observed and measured.The content of intracellular reactive ROS was observed and determined by DCFH-DA staining.The content of MDA in the supernatant and the antioxidant index of cells were determined by the reagent kit.Transcriptome technology was employed to analyze the potential mechanisms by which SSP mitigates oxidative damage in IPEC-J2 cells.The results showed that treatment with 625 μmol/L TBHP for 2 h significantly reduced the activity of IPEC-J2 cells,markedly increased LDH release(P＜0.05),inhibited CAT superoxide SOD and glutathione GPX activities(P＜0.05),and significantly elevated MDA and ROS levels(P＜0.05).Compared to the model group,after SSP treatment,intracellular ROS levels were significantly reduced(P＜0.05),while CAT,SOD,and GPX activities were significantly increased(P＜0.05),and MDA content and LDH re-lease were significantly decreased(P＜0.05)in a dose-dependent manner.Transcriptome analysis revealed that TBHP treatment significantly altered the transcriptional profiles of IPEC-J2 cells,while SSP treatment could restore the transcriptional profiles of the damaged cells to a certain ex-tent.Gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)indicated that the differentially expressed genes between the CC and TBHP groups were significantly enriched in oxidative phosphorylation,ribosome,and other pathways.Meanwhile,the differentially expressed genes between the SSP and TBHP groups were mainly enriched in oxidative phosphorylation,ap-optosis,glyoxylate and dicarboxylate metabolism,and other pathways.These results suggest that TBHP may disrupt normal oxidative respiration in IPEC-J2 cells by affecting oxidative phospho-rylation and interfering with metabolism pathways involving glycine,serine,and threonine,leading to oxidative damage in intestinal epithelial cells.Conversely,SSP treatment may potentially restore oxidative phosphorylation processes,alleviate lysosomal damage,reduce cell apoptosis,and miti-gate oxidative damage in intestinal epithelial cells through modulation of oxidative phosphoryla-tion,apoptosis,and lysosomal pathways.This discovery provides a theoretical basis for the clinical application of SSP in alleviating oxidative damage in the porcine intestinal tract.

To construct a subgenomic replication subsystem of foot-and-mouth disease virus(FM-DV)carrying fluorescent protein(UnaG),the full-length cDNA plasmid of FMDV was employed as the template.A few of structural and non-structural protein genes of FMDV were removed by double-enzyme digestion.By substituting reporter gene sequences expressing the green fluorescent protein UnaG for specific structural protein sequences of FMDV,FMDV-UnaG replicators were successfully created.After PCR and sequencing,linearized FMDV-UnaG replicons were transfected into BSR/T7 cells expressing T7 RNA polymerase,and the fluorescence signal was observed through fluorescence microscopy and laser confocal technique.The results demonstrated that the constructed FMDV-UnaG replicons could effectively express UnaG protein,and the protein colo-calized with FMDV 3A protein.Additionally,Western blot and RT-qPCR also detected that the replicator RNA could express the non-structural proteins of the virus and replicate autonomously in BSR/T7 cells,respectively.In conclusion,the successful construction of FMDV-UnaG sub-genomic replicators offers a favorable tool for further research on the replication and translation mechanism of FMDV and the development of vaccine vectors.

The objective of this study was to evaluate the protective effect and possible related mechanisms of Sophora subprostrate polysaccharide(SSP)on intestinal epithelial cell injury in-duced by Tert-Butyl hydroperoxide(TBHP).The optimal dose of TBHP and the safe concentra-tion range of SSP were determined using the MTT method.In this study,IPEC-J2 cells were divid-ed into five groups:the control group,the model group,the SSPL group,the SSPM group and the SSPH group,and the cell morphology,cell survival rate and LDH release rate were observed and measured.The content of intracellular reactive ROS was observed and determined by DCFH-DA staining.The content of MDA in the supernatant and the antioxidant index of cells were determined by the reagent kit.Transcriptome technology was employed to analyze the potential mechanisms by which SSP mitigates oxidative damage in IPEC-J2 cells.The results showed that treatment with 625 μmol/L TBHP for 2 h significantly reduced the activity of IPEC-J2 cells,markedly increased LDH release(P＜0.05),inhibited CAT superoxide SOD and glutathione GPX activities(P＜0.05),and significantly elevated MDA and ROS levels(P＜0.05).Compared to the model group,after SSP treatment,intracellular ROS levels were significantly reduced(P＜0.05),while CAT,SOD,and GPX activities were significantly increased(P＜0.05),and MDA content and LDH re-lease were significantly decreased(P＜0.05)in a dose-dependent manner.Transcriptome analysis revealed that TBHP treatment significantly altered the transcriptional profiles of IPEC-J2 cells,while SSP treatment could restore the transcriptional profiles of the damaged cells to a certain ex-tent.Gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)indicated that the differentially expressed genes between the CC and TBHP groups were significantly enriched in oxidative phosphorylation,ribosome,and other pathways.Meanwhile,the differentially expressed genes between the SSP and TBHP groups were mainly enriched in oxidative phosphorylation,ap-optosis,glyoxylate and dicarboxylate metabolism,and other pathways.These results suggest that TBHP may disrupt normal oxidative respiration in IPEC-J2 cells by affecting oxidative phospho-rylation and interfering with metabolism pathways involving glycine,serine,and threonine,leading to oxidative damage in intestinal epithelial cells.Conversely,SSP treatment may potentially restore oxidative phosphorylation processes,alleviate lysosomal damage,reduce cell apoptosis,and miti-gate oxidative damage in intestinal epithelial cells through modulation of oxidative phosphoryla-tion,apoptosis,and lysosomal pathways.This discovery provides a theoretical basis for the clinical application of SSP in alleviating oxidative damage in the porcine intestinal tract.

Foot-and-mouth disease (FMD) is one of the major animal infectious diseases in the world. All cloven-hoofed animals are susceptible to FMD. Vaccination is still the first choice for the prevention and control of FMD. mRNA vaccines can be rapidly designed, synthesized, and produced on a large scale in vitro, and they can induce effective protective immune responses, demonstrating the advantages of rapid development, easy preparation, and low biosafety risks. The design of untranslated regions is a key to enhancing the expression and efficacy of mRNA vaccines. In order to generate an efficient FMD mRNA vaccine, we designed three FMD P12A3C expression vectors with different untranslated regions and synthesized corresponding mRNAs. By comparing expression efficiency of these vectors and their mRNAs at different time points and in different cell lines, we found that the mRNA P12A3C-UTR3 had the best expression and universality. This study laid a foundation for the development of mRNA vaccines against FMD and provided a theoretical basis for the optimal sequence design of efficient mRNA.
Foot-and-Mouth Disease Virus/genetics* ; Animals ; RNA, Messenger/biosynthesis* ; Foot-and-Mouth Disease/immunology* ; Antigens, Viral/biosynthesis* ; Viral Vaccines/biosynthesis* ; Genetic Vectors/genetics* ; Cell Line ; Vaccines, DNA/immunology*

Bio-mineralization has emerged as a promising strategy to enhance vaccine immunogenicity. This study optimized the calcium phosphate (CaP) mineralization process of foot-and-mouth disease virus-like particles (FMD VLPs) to achieve high mineralization efficiency and scalability. Key parameters, including concentrations of Ca²⁺, HPO₄^2-, NaCl, and VLPs, as well as stirring speed, were systematically optimized. Stability of the scaled-up reaction system and immunogenicity of the mineralized vaccine were evaluated. Optimal conditions [25.50 mmol/L Ca(NO₃)₂, 15 mmol/L Na₂HPO₄, 300 mmol/L NaCl, 0.75 mg/mL VLPs, and 1 500 r/min] yielded CaP-mineralized VLPs (VLPs-CaP) with high mineralization efficiency, uniform morphology, and a favorable particle size. Scaling up the reaction by 25 folds maintained consistent mineralization efficiency and particle characteristics. Immunization in mice demonstrated that VLPs-CaP induced higher titers of specific antibodies and neutralizing antibodies than unmineralized VLPs (P < 0.05). Higher IgG2a/IgG1 ratio and enhanced IFN-γ secretion (P < 0.05) further indicated robust cellular immune responses. We establish a stable and scalable protocol for VLPs-CaP, providing a theoretical and technical foundation for developing high-efficacy VLPs-CaP vaccines.
Vaccines, Virus-Like Particle/immunology* ; Immunogenicity, Vaccine ; Calcium Phosphates/chemistry* ; Foot-and-Mouth Disease Virus ; Biomineralization ; Particle Size ; Animals ; Mice ; Antibodies, Neutralizing/blood* ; Antibodies, Viral/blood* ; Immunity, Cellular

Vaccination is a crucial strategy for the prevention and control of infectious diseases. Virus-like particles (VLPs), composed of structural proteins, have garnered significant attention as a novel type of vaccine due to their excellent safety and immunogenicity. However, similar to most vaccine antigens, VLPs exhibit insufficient thermal stability, which not only restricts the widespread application of vaccines but also increases the risk of vaccine inactivation. This study aims to enhance the stability and shelf life of VLPs derived from type A foot-and-mouth disease virus (FMDV) by employing vacuum freeze-drying technology. The optimal lyoprotectant formulation was determined through single-factor and combinatorial screening. Subsequently, the correlation between the immunogenicity of the freeze-dried vaccine and the content of FMDV VLPs was evaluated via a mouse model. The stability of FMDV VLPs before and after freeze-drying was further assessed by storing them at 4, 25, and 37 ℃ for varying time periods. Results indicated that the lyoprotectant formulation No.1, composed of 7.5% trehalose, 0.1% Tween 80, 50 mmol/L glycine, 1% sodium glutamate, and 3% polyvinylpyrrolidone (PVP), effectively preserved the content of FMDV VLPs during the vacuum freeze-drying process. The immunization trial in mice revealed that the levels of specific antibodies, immunoglobulin G1 (IgG1), interleukin-4 (IL-4), and neutralizing antibodies induced by freeze-dried FMDV VLPs were comparable to those induced by non-freeze-dried FMDV VLPs. The heat treatment results showed that the storage periods of freeze-dried FMDV VLPs at 4, 25, and 37 ℃ were significantly longer than those of non-freeze-dried FMDV VLPs. In conclusion, the selected lyoprotectant formulation effectively improved the stability of FMDV VLPs vaccines. This study provides valuable insights for enhancing the stability of novel subunit vaccines.
Freeze Drying/methods* ; Animals ; Foot-and-Mouth Disease Virus/immunology* ; Mice ; Vaccines, Virus-Like Particle/chemistry* ; Foot-and-Mouth Disease/immunology* ; Vacuum ; Drug Stability ; Mice, Inbred BALB C ; Viral Vaccines/immunology*

Pseudorabies (PR) is an infectious disease caused by the pseudorabies virus (PRV), affecting various domesticated and wild animals. Since pigs are the only natural hosts of PRV, PR poses a serious threat to the pig farming industry. Currently, PR is primarily prevented through vaccination with inactivated vaccines or genetically modified attenuated live vaccines. Developing safe and effective genetically engineered vaccines would facilitate the eradication and control of PR. In this study, the PRV vaccine strain Bartha-K61 was used as the reference strain. The gB protein was expressed via the baculovirus-insect cell expression system. Non-denaturing gel electrophoresis confirmed that the gB protein could form a trimeric structure. The purified gB protein was used to immunize mice, and the immune effect was evaluated by a challenge test. The results showed that the gB antigen induced a strong immune response in mice, with the serum-neutralizing antibody titer above 1:70. The lymphocyte stimulation index reached more than 1.29, and the level of (interferon gamma, IFN-γ) release was higher than 100 pg/mL. After immunization, mice were challenged with the virus at a dose of 10⁴ TCID₅₀/mL, 200 μL per mouse, and the clinical protection rate was 100%. Immunohistochemistry, histopathological section, and tissue viral load results showed that the pathological damage and viral load in the gB-immunized group were significantly lower than those in the PBS group. In summary, the gB protein obtained in this study induced strong humoral and cellular immune responses in mice, laying a foundation for developing a recombinant gB protein subunit vaccine.
Animals ; Mice ; Baculoviridae/metabolism* ; Viral Envelope Proteins/biosynthesis* ; Herpesvirus 1, Suid/genetics* ; Pseudorabies/immunology* ; Swine ; Pseudorabies Vaccines/genetics* ; Antibodies, Viral/blood* ; Insecta/cytology* ; Mice, Inbred BALB C ; Female ; Viral Vaccines/immunology*

Porcine reproductive and respiratory syndrome (PRRS), caused by the porcine reproductive and respiratory syndrome virus (PRRSV), is one of the major diseases threatening the swine industry. This study aims to rationally design and optimize natural antimicrobial peptides to identify antiviral candidates with potent inhibitory activity against PRRSV, thereby establishing a foundation for the development of novel preventive and therapeutic agents targeting PRRS. In this study, with cecropin D (CD) as the parent peptide, three derivatives (CD-2, CD-3, and CD-4) were designed through amino acid substitutions. CD and derived peptides were obtained by solid-phase peptide synthesis. MS and reversed-phase (RP)-HPLC were employed for sequence identification, purification, and purity analysis. The secondary structures of the peptides were investigated by circular dichroism spectroscopy. CellTiter 96^® AQueous one solution cell proliferation assay was used to evaluate the cytotoxicity of the peptides. The inhibitory activities and mechanisms of the peptides against PRRSV were studied by Western blotting, RT-qPCR, and indirect immunofluorescence assay. The MS and RP-HPLC results showed that CD and derived peptides were successfully synthesized, with the purity reaching up to 95%. Circular dichroism analysis revealed that the CD derivatives exhibited more stable and abundant α-helices in a cell membrane-mimicking environment. The MTS assay indicated that all tested peptides at 100 μg/mL had negligible cytotoxicity. The experimental results of the action phase of the peptide against PRRSV demonstrated that the derived peptides significantly enhanced antiviral activities at the viral entry stage compared with the parent peptide. This enhancement was attributed to the introduction of lysine, tryptophan, and phenylalanine, which increased the hydrophobicity and positive charge of the peptides. These findings provide a theoretical basis for the application and structural optimization of antiviral peptides and may offer a new strategy for preventing and controlling PRRSV.
Porcine respiratory and reproductive syndrome virus/physiology* ; Animals ; Swine ; Antiviral Agents/chemistry* ; Porcine Reproductive and Respiratory Syndrome/virology* ; Virus Internalization/drug effects* ; Antimicrobial Peptides/chemistry*