It is believed that the basic mechanism of cough associated with interstitial lung disease is collaterals deficiency with latent wind： the deficiency of the lung organs and lung collaterals is the basis of its pathogenesis， and latent wind in lung collaterals is the key mechanism of the cough which is difficult to cure. Treatment is based on the principle of supplementing deficiency and treating wind， dispelling the pathogens and unblocking the collaterals. Supplementing deficiency should supplement lungs， boost kidneys， and strengthen spleens to consolidate the root and banking up the origin， and regulate and tonify the lung collaterals； dispelling the pathogens should treat the internal and external winds at the same time， and taking into account the combined pathogens of phlegm， stasis and dampness to clear the stagnation of the lung collaterals.

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*

A 50-year-old male patient diagnosed with extensive stage small cell lung cancer was treated with pamiparib in combination with temozolomide.Five days later,the patient developed fever with fatigue.After 10 days,the patient stopped taking the drug due to worsening symptoms and was diagnosed with chemotherapy-induced myelosuppression(grade 4).The clinicist evaluated the patient's condition and assessed the association of adverse reactions using the Naranjo's evaluation scale,and concluded that myelosuppression may be induced by the combination of pamiparib and temozolomide.After symptomatic treatment,the patient's myelosuppression recovered completely.This article discusses the correlation between myelosuppression and the combination of the two drugs,provides treatment measures for this situation,briefly describes the risk factors of myelosuppression,treatment and prevention,and guides medical personnel to adjust the treatment plan in time according to different individuals in the process of using similar programs,and strengthens the monitoring and education of adverse drug reactions,so as to provide references for safe drug use.

Tongue diagnosis is an important method of TCM diagnosis and treatment.Tongue is the key link of auxiliary diagnosis of tongue feature extraction and processing,and also is the bottleneck of intelligent tongue diagnosis in traditional Chinese medicine.Using image processing,artificial intelligence technology to the tongue as a quantitative and identify characteristics of traditional Chinese medicine,looking for both conforms to the original thinking of TCM,and TCM tongue diagnosis method of accurately,has become a common concern of traditional Chinese medicine and computer field.From the mobile terminal tongue as auxiliary diagnostic system of traditional Chinese medicine tongue acquisition basic attribute,tongue diagnosis and image information building,tongue like features are required for accurate extraction and so on related key technology is analyzed,and build overall architecture,so as to provide technical reference for the tongue like intelligent diagnosis,promote the development of technology of tongue diagnosis in traditional Chinese medicine modernization.

Fecal microbiota transplantation(FMT)is a therapeutic approach that targets intestinal microorganisms by transplanting fecal microorganisms from healthy individuals into the gastrointestinal tract of diseased individuals,thus restoring the recipient's disordered gastrointestinal microbiota by restructuring the intestinal flora.However,the mechanism of action and adverse effects of FMT in different diseases have not yet been clarified,thus limiting its wide clinical application.Its use still relies on in-depth preclinical studies;however,highly inconsistent or incomplete experimental details provided in current reports,coupled with a lack of authoritative standards and recommendations,seriously affect the interpretation of the study findings and replication of the experimental procedures,as well as hindering the clinical translation of the result.We therefore review and discuss the key steps of recipient selection and graft sample collection,storage,graft material preparation,and grafting route,with the aim of improving the utilization of experimental animals,consumables,and labor,and providing method ological recommendations and references to achieve replicability and standardization of preclinical FMT studies.

The variable domain of heavy-chain antibody (VHH) has been developed widely in drug therapy, diagnosis, and research. Escherichia coli is the most popular expression system for VHH production, whereas low bioactivity occurs sometimes. Mammalian cells are one of the most ideal hosts for VHH expression at present. To improve the yield of VHH in Expi293F cells, we optimized the signal peptide (SP) and codons of VHH. Firstly, the fusion protein VHH1-Fc was used to screen SPs. The SP IFN-α2 showed the highest secretion as quantified by enzyme-linked immunosorbent assay (ELISA). Subsequently, codon optimization by improving GC3 and GC content doubled the yield of VHH1 and kept its binding activity to Senecavirus A (SVA). Finally, the mean yields of other 5 VHHs that fused with SP IFN-α2 and codon-optimized were over 191.6 mg/L, and these VHHs had high recovery and high purity in the culture supernatant. This study confirms that SP IFN-α2 and codon optimization could produce VHHs in Expi293F cells efficiently, which provides a reference for the large-scale production of VHHs.
Codon/genetics* ; Protein Sorting Signals/genetics* ; Escherichia coli/metabolism* ; Humans ; Recombinant Fusion Proteins/biosynthesis* ; Interferon-alpha/metabolism* ; Immunoglobulin Heavy Chains/immunology* ; Cell Line ; Immunoglobulin Variable Region/immunology*