Immunogenicity of peptide-based vaccine composed of epitopes from Echinococcus granulosus rEg.P29.

Echinococcus granulosus is one of the main causes of economic loss in the livestock industry because of its food-borne transmission. Cutting off the transmission route is a valid prevention method, and vaccines are the most effective means of controlling and eliminating infectious diseases. However, no human-related vaccine has been yet marketed. As a genetic engineering vaccine, recombinant protein P29 of E. granulosus (rEg.P29) could provide protection against deadly challenges. In this study, we generated peptide vaccines (rEg.P29T , rEg.P29B , and rEg.P29T+B ) based on rEg.P29 and an immunized model was established by subcutaneous immunization. Further evaluation showed that peptide vaccine immunization in mice induced T helper type 1 (Th1)-mediated cellular immune responses, leading to high levels of rEg.P29 or rEg.P29B -specific antibodies. In addition, rEg.P29T+B immunization can induce a higher antibody and cytokine production level than single-epitope vaccines, and immune memory is also longer. Collectively, these results suggest that rEg.P29T+B has the potential to be developed as an efficient subunit vaccine for use in areas where E. granulosus is endemic.

A subunit vaccine based on Brucella rBP26 induces Th1 immune responses and M1 macrophage activation.

Brucellosis is a global zoonotic infection caused by Brucella bacteria, which poses a significant burden on society. While transmission prevention is currently the most effective method, the absence of a licenced vaccine for humans necessitates the urgent development of a safe and effective vaccine. Recombinant protein-based subunit vaccines are considered promising options, and in this study, the Brucella BP26 protein is expressed using prokaryotic expression systems. The immune responses are evaluated using the well-established adjuvant CpG-ODN. The results demonstrate that rBP26 supplemented with a CpG adjuvant induces M1 macrophage polarization and stimulates cellular immune responses mediated by Th1 cells and CD8 + T cells. Additionally, it generates high levels of rBP26-specific antibodies in immunized mice. Furthermore, rBP26 immunization activates, proliferates, and produces cytokines in T lymphocytes while also maintaining immune memory for an extended period of time. These findings shed light on the potential biological function of rBP26, which is crucial for understanding brucellosis pathogenesis. Moreover, rBP26 holds promise as an effective subunit vaccine candidate for use in endemic areas.

[A neonatal intelligent regulation system based on the combination of mild hypothermia mattress and hyperbaric oxygen chamber: introduction to a patent]. / 基于亚低温床垫与高压氧舱结合的新生儿智能调控系统.

Neonatal hypoxic-ischemic encephalopathy (HIE) is a common disease that affects brain function in neonates. At present, mild hypothermia and hyperbaric oxygen therapy are the main methods for the treatment of neonatal HIE; however, they are independent of each other and cannot be combined for synchronous treatment, without monitoring of brain function-related physiological information. In addition, parameter setting of hyperbaric oxygen chamber and mild hypothermia mattress relies on the experience of the medical practitioner, and the parameters remain unchanged throughout the medical process. This article proposes a new device for the treatment of neonatal HIE, which has the modules of hyperbaric oxygen chamber and mild hypothermic mattress, so that neonates can receive the treatment of hyperbaric oxygen chamber and/or mild hypothermic mattress based on their conditions. Meanwhile, it can realize the real-time monitoring of various physiological information, including amplitude-integrated electroencephalogram, electrocardiogram, and near-infrared spectrum, which can monitor brain function, heart rate, rhythm, myocardial blood supply, hemoglobin concentration in brain tissue, and blood oxygen saturation. In combination with an intelligent control algorithm, the device can intelligently regulate parameters according to the physiological information of neonates and give recommendations for subsequent treatment.

Baseline T-lymphocyte and cytokine indices in sheep peripheral blood.

BACKGROUND: Sheep are an important livestock species worldwide and an essential large-animal model for animal husbandry and veterinary research. Understanding fundamental immune indicators, especially T-lymphocyte parameters, is necessary for research on sheep diseases and vaccines, to better understand the immune response to bacteria and viruses for reducing the use of antibiotics and improving the welfare of sheep. We randomly selected 36 sheep of similar ages to analyze cell-related immune indicators in peripheral blood mononuclear cells (PBMCs). The proportions of CD4+ and CD8+ T cells in PBMCs were detected by flow cytometry. We used Concanavalin A (Con A) and Phorbol-12-myristate-13-acetate (PMA)/Ionomycin to stimulate PBMCs, and measured the expression of IFN-γ, IL-4, and IL-17A using enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunospot assay (ELISpot). Simultaneously, PMA/Ionomycin/brefeldin A (BFA) was added to PBMCs, then the expression of IFN-γ, IL-4, and IL-17A was detected by flow cytometry after 4 h of culturing. In addition, we observed the proliferation of PBMCs stimulated with Con A for 3, 4, and 5 days. RESULTS: The proportions of CD4+ T lymphocytes (18.70 ± 4.21%) and CD8+ T lymphocytes (8.70 ± 3.65%) were generally consistent among individuals, with a CD4/CD8 ratio of 2.40 ± 0.79. PBMCs produced high levels of IFN-γ, IL-4, and IL-17A after stimulation with PMA/Ionomycin and Con A. Furthermore, PMA/Ionomycin stimulation of PBMC yielded significantly higher cytokine levels than Con A stimulation. Flow cytometry showed that the level of IFN-γ (51.49 ± 11.54%) in CD8+ T lymphocytes was significantly (p < 0.001) higher than that in CD4+ T lymphocytes (14.29 ± 3.26%); IL-4 (16.13 ± 6.81%) in CD4+ T lymphocytes was significantly (p < 0.001) higher than that in CD8+ T lymphocytes (1.84 ± 1.33%), There was no difference in IL-17A between CD4+ (2.83 ± 0.98%) and CD8+ T lymphocytes (1.34 ± 0.67%). The proliferation of total lymphocytes, CD4+ T lymphocytes, and CD8+ T lymphocytes continued to increase between days 3 and 5; however, there were no significant differences in proliferation between the cell types during the stimulation period. CONCLUSIONS: Evaluating primary sheep immune indicators, especially T lymphocytes, is significant for studying cellular immunity. This study provided valuable data and theoretical support for assessing the immune response of sheep to pathogens and improving sheep welfare.

Identification of a dominant murine T-cell epitope in recombinant protein P29 from Echinococcus granulosus

Echinococcus granulosus causes echinococcosis, an important zoonotic disease worldwide and a major public health issue. Vaccination is an economical and practical approach for controlling E. granulosus. We have previously revealed that a recombinant protein P29 (rEg.P29) is a good vaccine candidate against E. granulosus. However, T cell immunogenic epitopes have not been identified. In the present study, we use rEg.P29-immunized mice as models to screen immunogenic epitopes for the construction of a novel multi-epitope vaccine. We search for immunodominant epitopes from an overlapping peptide library to screen the peptides of rEg.P29. Our results confirm that rEg.P29 immunization in mice elicits the activation of T cells and induces cellular immune responses. Further analyses show that a T cell epitope within amino acids 86­100 of rEg.P29 elicits significant antigen-specific IFN-γ production in CD4+ and CD8+ T cells and promotes specific T-cell activation and proliferation. Collectively, these results provide a reference for the construction of a novel vaccine against broad E. granulosus genotypes based on epitopes of rEg.P29.

Novel Plant Growth-Promoting Bacteria Isolated from Bauxite Residue: The Application for Revegetation.

Microbial inoculation with appropriate inorganic-organic amendments is a promising strategy for ecological rehabilitation at bauxite residue disposal areas. Nevertheless, research on screening suitable plant growth-promoting bacteria with tolerance to highly sodic-alkalinity is very limited in the literature. In this study, novel plant growth-promoting bacteria isolated from bauxite residue were used to investigate their potential for revegetation. Under high saline-alkalinity stress, inoculation of Z18 and Z28 increased the activity of antioxidative enzymes, whilst improving chlorophyll and carotenoid contents in ryegrass. Inoculation of the selected strains greatly reduced damage to organelles in ryegrass as observed by transmission electron microscopy. Based on 90-day soil incubation, inoculated strains improved physicochemical properties of bauxite residue and improved plant growth. These findings suggest that Z18 and Z28 may be selected as potential strains for vegetation establishment, aiding microbial remediation at bauxite disposal areas.

Improvements on physical conditions of bauxite residue following application of organic materials.

Soil formation and ecological rehabilitation is the most promising strategy to eliminate environmental risks of bauxite residue disposal areas. Its poor physical structure is nevertheless a major limitation to plant growth. Organic materials were demonstrated as effective ameliorants to improve the physical conditions of bauxite residue. In this study, three different organic materials including straw (5% W/W), humic acid (5% W/W), and humic acid-acrylamide polymer (0.2% and 0.4%, W/W) were selected to evaluate their effects on physical conditions of bauxite residue pretreated by phosphogypsum following a 120-day incubation experiment. The proportion of 2-1 mm macro-aggregates, mean weight diameter (MWD) and geometric mean diameter (GWD) increased following organic materials addition, which indicated that organic materials could enhance aggregate stability. Compared with straw, and humic acid, humic acid-acrylamide polymer application had improved effects on the formation of water-stable aggregates in the residues. Furthermore, organic materials increased the total porosity, total pore volume and average pore diameter, and reduced the micropore content according to nitrogen gas adsorption (NA) and mercury intrusion porosimetry (MIP) analysis, whilst enhancing water retention of the residues based on water characteristic curves. Compared with traditional organic wastes, humic acid-acrylamide polymer could be regarded as a candidate according to the comprehensive consideration of the additive amount and the effects on physical conditions of bauxite residue. These findings could provide a novel application to both Ca-contained acid solid waste and high-molecular polymers on ecological rehabilitation at disposal areas.

Screening, construction, and serological identification of the diagnostic antigen molecule EG-06283 for the diagnosis of Echinococcus granulosus.

Several strategies exist to prevent and control echinococcosis, a global parasitic disease. However, most treatments are ineffective and adverse effects are common. Therefore, we aimed to screen protoscolex antigen molecules of Echinococcus granulosus to identify a diagnostic biomarker for hydatid disease. Published E. granulosus transcriptome sequencing data were analyzed to screen for antigen molecules that are highly expressed in protoscoleces but not in oncospheres. The membrane protein EG-06283 (annotated as Frizzled-4) was selected from 16 antigens, and its gene fragment was subjected to codon optimization and synthesis. rEG-06283 expression was induced in the pET-24a/EG-06283/BL21 strain; subsequently, the protein was purified and subcutaneously injected into ICR mice at weeks 0, 2, 4, and 6. Blood sampling occurred periodically to quantify serum immunoglobulin G (IgG) levels via enzyme-linked immunosorbent assays (ELISA). Immunogenicity was determined by western blot assays using sera from normal mice and mice with secondary hydatid infections. The antigen's immune reactivity and diagnostic value were validated using sera of patients with hydatid disease. ELISA results confirmed that the antigen molecule induced specific IgG production in mice, resulting in significantly higher levels than those in the adjuvant and control groups (P < 0.05). The western blot results indicated that the protein was recognized by antibodies in the sera of mice with hydatid infection and the antisera of immunized mice. Quantification of protein levels in the sera of patients with hydatid disease significantly differed from levels in healthy participants (P < 0.05). These results indicate that rEG-06283 is a potential diagnostic antigen for E. granulosus infections.

Evaluation of the anticancer and anti-metastasis effects of novel synthetic sodium channel blockers in prostate cancer cells in vitro and in vivo.

BACKGROUND: Voltage-gated sodium channels (VGSCs) are involved in several cellular processes related to cancer cell growth and metastasis, including adhesion, proliferation, apoptosis, migration, and invasion. We here in investigated the effects of S0154 and S0161, two novel synthetic sodium channel blockers (SCBs), on human prostate cancer cells (PC3, DU145, and LnCaP) and a prostate cancer xenograft model. METHODS: The MTT assay was used to assess the anticancer effects of SCBs in PC3, DU145, and LnCaP cells. Sodium indicator and glucose uptake assays were used to determine the effects of S0154 and S0161 in PC3 cells. The impact of these SCBs on the proliferation, cell cycle, apoptosis, migration, and invasion of PC3 cells were determined using a CFDA-SE cell proliferation assay, cell cycle assay, annexin V-FITC apoptosis assay, transwell cell invasion assay, and wound-healing assay, respectively. The protein expression levels of Nav1.6, Nav1.7, CDK1, cyclin B1, MMP2, MMP9 in PC3 cells were analysis by Western blotting. The in vivo anticancer activity was evaluated using a PC3 xenograft model in nude mice. RESULTS: S0154 and S0161 both showed anticancer and anti-metastatic effects against prostate cancer cells and significantly inhibited cell viability, with IC50 values in the range of 10.51-26.60 µmol/L (S0154) and 5.07-11.92 µmol/L (S0161). Both compounds also increased the intracellular level of sodium, inhibited the protein expression of two α subunits of VGSCs (Nav1.6 and Nav1.7), and caused G2/M phase cell cycle arrest, with no or minor effects on cell apoptosis. Concentrations of 5 and 10 µmol/L of S0154 and S0161 significantly decreased the glucose uptake of PC3 cells. The compounds also inhibited the proliferation of PC3 cells and decreased their invasion in transwell assays. Furthermore, S0161 exerted antitumor activity in an in vivo PC3 xenograft model in nude mice, inhibiting the growth of the tumors by about 51% compared to the control group. CONCLUSIONS: These results suggest that S0154 and S0161 have anticancer and anti-metastasis effects in prostate cancer cells both in vitro and in vivo, supporting their further development as potential therapeutic agents for prostate cancer.

The Feasibility and Safety of Interventional Occlusion Treatment of Intracristal Ventricular Septal Defects: Clinical Report of 56 Cases.

BACKGROUND: To investigate the feasibility and safety of the O eccentric shape occluder in the interventional occlusion treatment of intracristal ventricular septal defect (IVSD). METHODS: A retrospective analysis of the clinical data of 56 IVSD patients treated by interventional occlusion at our center, as well as recording of their intraoperative and postoperative status, was performed. RESULTS: Of the 56 patients, a total of 48 patients underwent successful occlusion during the first surgical attempt. Four patients were transferred to the Surgery Department after occlusion when the largest occluder failed because of large defects. Two patients exhibited aortic valve regurgitation; 1 patient had mild regurgitation, which was not worsened after 6 months of follow-up. One patient had severe aortic regurgitation, and 2 days after the operation, the patient underwent a second operation. One patient exhibited a residual shunt, which was above the occluder; after 1 year of follow-up, re-occlusion was successful and eliminated the shunt. One patient developed complete right bundle branch block. CONCLUSION: Most IVSD interventional occlusion treatments are feasible and safe.

Evaluation of normal swallowing functions by using dynamic high-density surface electromyography maps.

BACKGROUND: Swallowing is a continuous process with substantive interdependencies among different muscles, and it plays a significant role in our daily life. The aim of this study was to propose a novel technique based on high-density surface electromyography (HD sEMG) for the evaluation of normal swallowing functions. METHODS: A total of 96 electrodes were placed on the front neck to acquire myoelectric signals from 12 healthy subjects while they were performing different swallowing tasks. HD sEMG energy maps were constructed based on the root mean square values to visualize muscular activities during swallowing. The effects of different volumes, viscosities, and head postures on the normal swallowing process were systemically investigated by using the energy maps. RESULTS: The results showed that the HD sEMG energy maps could provide detailed spatial and temporal properties of the muscle electrical activity, and visualize the muscle contractions that closely related to the swallowing function. The energy maps also showed that the swallowing time and effort was also explicitly affected by the volume and viscosity of the bolus. The concentration of the muscular activities shifted to the opposite side when the subjects turned their head to either side. CONCLUSIONS: The proposed method could provide an alternative method to physiologically evaluate the dynamic characteristics of normal swallowing and had the advantage of providing a full picture of how different muscle activities cooperate in time and location. The findings from this study suggested that the HD sEMG technique might be a useful tool for fast screening and objective assessment of swallowing disorders or dysphagia.

FastICA peel-off for ECG interference removal from surface EMG.

BACKGROUND: Multi-channel recording of surface electromyographyic (EMG) signals is very likely to be contaminated by electrocardiographic (ECG) interference, specifically when the surface electrode is placed on muscles close to the heart. METHODS: A novel fast independent component analysis (FastICA) based peel-off method is presented to remove ECG interference contaminating multi-channel surface EMG signals. Although demonstrating spatial variability in waveform shape, the ECG interference in different channels shares the same firing instants. Utilizing the firing information estimated from FastICA, ECG interference can be separated from surface EMG by a "peel off" processing. The performance of the method was quantified with synthetic signals by combining a series of experimentally recorded "clean" surface EMG and "pure" ECG interference. RESULTS: It was demonstrated that the new method can remove ECG interference efficiently with little distortion to surface EMG amplitude and frequency. The proposed method was also validated using experimental surface EMG signals contaminated by ECG interference. CONCLUSIONS: The proposed FastICA peel-off method can be used as a new and practical solution to eliminating ECG interference from multichannel EMG recordings.

Studies of the mechanism of an antibacterial peptide (cecropinA-magainin) on methicillin-resistant Staphylococcus aureus membranes.

As bacterial resistance becomes increasingly common, a new hybrid peptide, cecropinA-magainin (KWALSKEGPGKFLGKKKKF), has been developed that can kill a broad spectrum of bacteria without damaging human cells. The mechanism of antibacterial toxicity for the hybrid peptides is unknown. Herein, we investigate the localization of the hybrid peptide in methicillin-resistant Staphylococcus aureus (MRSA). The minimum inhibitory concentration was 64 µg/mL. The hybrid peptides could enhance the hydrophobicity of MRSA. Dye leakage experiments showed that the hybrid peptides caused dye leakage from liposomes. The hybrid peptides influenced the permeability of the outer membrane and plasma membrane of MRSA. After cecropinA-magainin treatment of MRSA, the membrane ultrastructure was damaged and the concentration of K+ increased. Ultimately, the peptide destroyed the integrity of the bacterial cell membrane, allowing the dye propidium iodide to enter the cytoplasm. Therefore, the hybrid antibacterial peptide can kill MRSA.

Barrier layer induced channeling effect of As ion implantation in HgCdTe and its influences on electrical properties of p-n junctions.

The HgCdTe layers (xCd∼0.285 and 0.225) were grown by molecular beam epitaxy and liquid phase epitaxy, respectively, followed by the deposition of CdTe and ZnS films as barrier layers by thermal evaporation. Then, the p-on-n photodiodes were fabricated by AS ion implantation, Hg overpressure annealing, passivation, and metallization. The secondary ion mass spectrometry and transmission electron microscopy results indicate that the evaporated CdTe layer with a column structure induces the channeling effect of As ion implantation causing the device performance degradation. This effect could be suppressed by depositing a CdTe film with a layered structure through E-beam evaporation. Finally, the current-voltage (I-V) and capacitance-voltage (C-V) characteristics of these p-n junctions were estimated and analyzed.

[Cloning, Expression and Immungenicity Analysis of Antigen Eg-01883 Screened from Protoscoleces of Echinococcus granulosus].

Objective: To screen for the Echinococcus granulosus 01883（Eg-01883） specifically expressed at the protoscolex period, clone and express this molecule as well as analyse its immunogenicity. Methods: Eg-01883, which is highly expressed at the protoscolex period but not in oncosphere， was screened by analysing the published mRNA sequences of E. granuolosus. Total RNA of E. granuolosus was extracted, Eg-01883 was cloned by RT-PCR, and the recombinant plasmid pET28a-Eg-01883 was constructed. Expression of the recombinant protein rEg-01883 was induced by isopropyl-ß-D-thiogalactoside （IPTG）. ICR mice were randomized into 3 groups （n=12 in each group）. Mice in the immunization group received subcutaneous injections of 10 µg rEg-01883 in 100 µl PBS emulsified in Freund's adjuvant at multiple sites, followed by immune enhancement after 2 weeks. Mice in the adjuvant group were injected with PBS and adjuvant. Mice in the control group received no treatment. Blood was obtained through caudal vein before immunization, and at 1, 2, and 4 weeks after the first immunization, and through the eyeball at 6 weeks after immunization. Serum levels of IgG, IFN-γ and IL-4 were determined by ELISA. The immunogenicity of rEg-01883 was identified by Western blotting. Results: Eg-01883 was screened, cloned, expressed and purified to obtain the recombinant protein rEg-01883, which mainly existed as the inclusion body. ELISA results showed that immunization with rEg-01883 induced production of specific IgG antibody. The serum IgG level in the immunization group increased from 1 week after the first immunization, peaked at 6 weeks（2.344±0.153）, which was significantly higher than those of the adjuvant group（0.206 1±0.006） and the control group （0.241±0.01） （P<0.01）. At 6 weeks after the first immunization, the serum levels of IFN-γ （43.23 pg/ml） and IL-4（24.88 pg/ml） in the immunization group were significantly higher than those in the adjuvant group（21.77 pg/ml, 13.27 pg/ml） and the control group（17.40 pg/ml, 12.25 pg/ml）（P<0.05）. Western blot showed that the recombinant protein rEg-01883 could be recognized by His-Tag antibodies, serum of immunized mice, and serum of mice with secondary infection. Conclusion: The recombinant protein rEg-01883 shows good immunogenicity in ICR mice.

A Secure and Interpretable AI for Smart Healthcare System: A Case Study on Epilepsy Diagnosis Using EEG Signals.

The efficient patient-independent and interpretable framework for electroencephalogram (EEG) epileptic seizure detection (ESD) has informative challenges due to the complex pattern of EEG nature. Automated detection of ES is crucial, while Explainable Artificial Intelligence (XAI) is urgently needed to justify the model detection of epileptic seizures in clinical applications. Therefore, this study implements an XAI-based computer-aided ES detection system (XAI-CAESDs), comprising three major modules, including of feature engineering module, a seizure detection module, and an explainable decision-making process module in a smart healthcare system. To ensure the privacy and security of biomedical EEG data, the blockchain is employed. Initially, the Butterworth filter eliminates various artifacts, and the Dual-Tree Complex Wavelet Transform (DTCWT) decomposes EEG signals, extracting real and imaginary eigenvalue features using frequency domain (FD), time domain (TD) linear feature, and Fractal Dimension (FD) of non-linear features. The best features are selected by using Correlation Coefficients (CC) and Distance Correlation (DC). The selected features are fed into the Stacking Ensemble Classifiers (SEC) for EEG ES detection. Further, the Shapley Additive Explanations (SHAP) method of XAI is implemented to facilitate the interpretation of predictions made by the proposed approach, enabling medical experts to make accurate and understandable decisions. The proposed Stacking Ensemble Classifiers (SEC) in XAI-CAESDs have demonstrated 2% best average accuracy, recall, specificity, and F1-score using the University of California, Irvine, Bonn University, and Boston Children's Hospital-MIT EEG data sets. The proposed framework enhances decision-making and the diagnosis process using biomedical EEG signals and ensures data security in smart healthcare systems.

Progress with polo-like kinase (PLK) inhibitors: a patent review (2018-present).

INTRODUCTION: Polo-like kinases (PLKs) have five isoforms, all of which play crucial roles in cell cycle and cell proliferation, offering opportunities for drug design and treatment of cancers and other related diseases. Notably, PLK1 and PLK4 have been extensively investigated as cancer drug targets. One distinctive feature of PLKs is the presence of a unique polo-box domain (PBD), which regulates kinase activity and subcellular localization. This provides possibilities for specifically targeting PLKs. AREA COVERED: This article provides an overview of the roles of PLKs in various cancers and related diseases, as well as the drug development involving PLKs, with a particular focus on PLK1 and PLK4. It summarizes the PLK1 and PLK4 inhibitors that have been disclosed in patents or literature (from 2018 - present), which were sourced from SciFinder and WIPO database. EXPERT OPINION: After two decades of drug development on PLKs, several drugs progressed into clinical trials for the treatment of many cancers; however, none of them has been approved yet. Further elucidating the mechanisms of PLKs and identifying and developing highly selective ATP-competitive inhibitors, highly potent drug-like PBD inhibitors, degraders, etc. may provide new opportunities for cancer therapy and the treatment for several nononcologic diseases. PLKs inhibition-based combination therapies can be another helpful strategy.

Omics-based investigation of pathological liver injury induced by Echinococcus multilocularis infection in mice.

BACKGROUND: Alveolar echinococcosis (AE) can cause severe liver injury and be fatal if left untreated. Currently, there are no effective therapeutic options for AE-induced liver injury. Therefore, by exploring the changes of gene proteins in mice with damaged liver, we attempted to identify the key molecules of liver damage, and provide data that will enable the development of drugs targeting hepatic AE. METHODS: BALB/c mice were inoculated with protoscoleces via the hepatic portal vein. Three months later, B-ultrasound examination and Hematoxylin-eosin (H&E) staining were used to confirm liver damage in mice. RNA sequencing and Liquid chromatography-mass spectrometry (LC-MS) were used to screen differentially expressed molecules associated with liver damage through bioinformatics, and Quantitative Real-Time PCR (qRT-PCR) was used to verify their expression. RESULTS: B-ultrasound examination showed liver lesions in the infected group, and H&E staining showed liver inflammation, fibrosis and liver necrosis. RNA sequencing and LC-MS results showed changes in the levels of more than 1000 genes and proteins, with upregulation of immune and inflammation pathways. By contrast, the downregulated genes and proteins were mostly involved in various metabolic reactions. Correlation analysis was conducted between the transcriptome data and proteome data. The results revealed 240 differentially expressed genes, of which 192 were upregulated, and 48 were downregulated. Many of these genes were involved in metabolic reactions, such as Catalase (Cat), fatty acid synthase (Fasn), and IL-16 genes, which may have relevance to liver injury. The results of qRT-PCR were consistent with those of bioinformatics analysis. CONCLUSIONS: The mechanisms of liver injury in mice infected with Echinococcus multilocularis are complex, involving abnormal metabolism, oxidative stress, inflammatory response, and many other factors. This study provides the data for preliminary exploration for the development of targeted therapies against AE.

Phytochemical determination and mechanistic investigation of Polygala tenuifolia root (Yuanzhi) extract for bronchitis: UPLC-MS/MS analysis, network pharmacology and in vitro/in vivo evaluation.

ETHNOPHARMACOLOGICAL RELEVANCE: Bronchitis is a respiratory disease characterized by a productive cough. Polygala tenuifolia Willd., commonly known as Yuan zhi, is a traditional Chinese herbal medicine used for relieving cough and removing phlegm. Despite its historical use, studies are lacking on the effectiveness of P. tenuifolia in treating bronchitis. Furthermore, the molecular mechanisms underlying the action of its bioactive compounds remain unknown. AIM OF THE STUDY: This study aims to identify the main bioactive compounds responsible for the effects of P. tenuifolia liquid extract (PLE) in treating bronchitis and to elucidate the associated molecular mechanisms. MATERIALS AND METHODS: The main chemical compounds in PLE were identified and determined using ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The antitussive, expectorant and anti-inflammatory activities of PLE were evaluated in an ammonia-induced mouse cough model, a tracheal phenol red excretion mouse model, and a xylene-induced ear swelling mouse model, respectively. A network pharmacology analysis was conducted to investigate the associated gene targets, gene ontology, and KEGG pathways related to the main bioactives in PLE targeting bronchitis. PLE and its five bioactive compounds were assessed for their potential anti-inflammatory activities in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Western blot analysis was conducted to elucidate the associated molecular mechanisms. RESULTS: Thirty-seven compounds in PLE were identified, and twelve main compounds were further quantified in PLE using UPLC-MS/MS. PLE oral gavage administrations (0.6 and 0.12 mg/kg) for 7 days markedly reduced cough frequency, prolonged latency period of cough, reduced phlegm and inflammation in mice. The network pharmacology analysis identified 57 gene targets of PLE against bronchitis. The PI3K/AKT and MAPK signalling pathways were the top two modulated pathways. In RAW264.7 cells, PLE (12.5-50 µg/mL) significantly reduced cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), interleukin (IL)-1ß, IL-6 and tumor necrosis factor (TNF)-α. PLE downregulated LPS-elevated protein targets in both PI3K/AKT and MAPK signaling pathways. In PLE, tenuifolin, polygalaxanthone ⅠⅠⅠ, polygalasaponin ⅩⅩⅤⅢ, tenuifoliside B, and 3,6'-Disinapoyl sucrose, were identified as the top five core components responsible for treating bronchitis. These compounds were also found to modulate the protein targets in the PI3K/AKT and MAPK signalling pathways. CONCLUSIONS: This study demonstrated the potential therapeutic effects of PLE on bronchitis by reducing cough, phlegm and inflammation. The anti-inflammatory action and molecular mechanisms of the 5 main bioactive compounds in PLE were partly validated through the in vitro assays. The findings provide valuable insights into the mechanisms underlying the traditional use of PLE for bronchitis.

[Position analysis of three recombinant proteins of Echinococcus granulosus protoscolex with two-dimensional electrophoresis].

OBJECTIVE: To obtain specific antibodies of the three recombinant antigens obtained previously, rEgZW-5, rEg14-3-3 and rEgP-29, for identifying the corresponding proteins in two-dimensional electrophoretogram of Echinococcus granulosus protoscolex. METHODS: The distribution of proteins from E. granulosus protoscoleces was judged by SDS-PAGE previously. Two-dimensional electrophoresis was used to separate proteins from E. granulosus protoscoleces, and the result was scanned and analyzed by the PDquest software to get the information about the quantity of proteins as well as their isoelectric point (IP) and relative molecular mass (MA,). Rabbits were immunized with the 3 recombinant antigens and antibodies were purified from antisera. Western blotting was used to identify the protein as marker in two-dimensional electrophoretogram of protoscolex. RESULTS: SDS-PAGE displayed that the proteins separated from Echinococcus granulosus protoscoleces mainly distributed in the M, region of 18,000-90,000. 240 proteins were obtained by two-dimensional electrophoresis with M, 15,790-117,050 and IP 4.0-9.5, and 85.8% (206/241) of the proteins showed the IP ranged from 5 to 9. Western blotting showed that the specific antibody of rEg14-3-3 identified the 14-3-3 protein in two-dimensional electrophoretogram of protoscolex with Mr 33 000 and IP 4.86, the specific antibody of rEgZW-5 identified the ZW-5 protein with Mr 23,000 and IP 4.98, and the specific antibody of rEg P-29 identified the P-29 protein with Mr 29,000 and IP 5.65. CONCLUSION: The antibodies against the three recombinant proteins from Echinococcus granulosus protoscoleces can identify corresponding proteins in the two-dimensional electrophoregrams.