Analysis of the components of Mycobacterium tuberculosis heat-resistant antigen (Mtb-HAg) and its regulation of γδ T-cell function.

BACKGROUND: Mycobacterium tuberculosis heat-resistant antigen (Mtb-HAg) is a peptide antigen released from the mycobacterial cytoplasm into the supernatant of Mycobacterium tuberculosis (Mtb) attenuated H37Ra strain after autoclaving at 121 °C for 20 min. Mtb-HAg can specifically induce γδ T-cell proliferation in vitro. However, the exact composition of Mtb-HAg and the protein antigens that are responsible for its function are currently unknown. METHODS: Mtb-HAg extracted from the Mtb H37Ra strain was subjected to LC‒MS mass spectrometry. Twelve of the identified protein fractions were recombinantly expressed in Escherichia coli by genetic engineering technology using pET-28a as a plasmid and purified by Ni-NTA agarose resin to stimulate peripheral blood mononuclear cells (PBMCs) from different healthy individuals. The proliferation of γδ T cells and major γδ T-cell subset types as well as the production of TNF-α and IFN-γ were determined by flow cytometry. Their proliferating γδ T cells were isolated and purified using MACS separation columns, and Mtb H37Ra-infected THP-1 was co-cultured with isolated and purified γδ T cells to quantify Mycobacterium viability by counting CFUs. RESULTS: In this study, Mtb-HAg from the attenuated Mtb H37Ra strain was analysed by LC‒MS mass spectrometry, and a total of 564 proteins were identified. Analysis of the identified protein fractions revealed that the major protein components included heat shock proteins and Mtb-specific antigenic proteins. Recombinant expression of 10 of these proteins in by Escherichia coli genetic engineering technology was used to successfully stimulate PBMCs from different healthy individuals, but 2 of the proteins, EsxJ and EsxA, were not expressed. Flow cytometry results showed that, compared with the IL-2 control, HspX, GroEL1, and GroES specifically induced γδ T-cell expansion, with Vγ2δ2 T cells as the main subset, and the secretion of the antimicrobial cytokines TNF-α and IFN-γ. In contrast, HtpG, DnaK, GroEL2, HbhA, Mpt63, EsxB, and EsxN were unable to promote γδ T-cell proliferation and the secretion of TNF-α and IFN-γ. None of the above recombinant proteins were able to induce the secretion of TNF-α and IFN-γ by αß T cells. In addition, TNF-α, IFN-γ-producing γδ T cells inhibit the growth of intracellular Mtb. CONCLUSION: Activated γδ T cells induced by Mtb-HAg components HspX, GroES, GroEL1 to produce TNF-α, IFN-γ modulate macrophages to inhibit intracellular Mtb growth. These data lay the foundation for subsequent studies on the mechanism by which Mtb-HAg induces γδ T-cell proliferation in vitro, as well as the development of preventive and therapeutic vaccines and rapid diagnostic reagents.

Immune responses induced by Mycobacterium tuberculosis heat-resistant antigen (Mtb-HAg) upon co-administration with Bacillus Calmette-Guérin in mice.

OBJECTIVES: To preliminarily assess the immunogenicity of Mtb-HAg in mice and the synergistic effect provided by HAg when co-immunised with BCG. METHODS: Mice were randomly grouped for different immunisations and then spleens were aseptically removed and lymphocytes were extracted for immediate detection of cytokines transcript levels and stimulation index(SI), cytokine secretion and multifunctional antigen-specific T cells were detected after incubation for different times. RESULTS: HAg extracted from active Mtb is a group of mixed polypeptides with molecular weights of (10-14) kDa. It can significantly stimulate lymphocytes proliferation and increase SI. Injection of HAg alone and in combination with BCG induced significantly higher numbers of multifunctional antigen-specific T cells including CD4+ IFN-γ+, CD4+ IL-2+, CD8+ IFN-γ+, and CD8+ IL-2+ cells than that in BCG-treated mice. Co-immunisation induced the secretion of higher levels of IFN-γ, TNF-α, IL-2 and IL-4 and increased their mRNA expression levels. Significant increases in the transcription levels of IL-10, IL-12 and IL-17 were observed in the co-immunised group with the assistance of HAg. CONCLUSION: We demonstrated that HAg has favourable immunogenicity, triggers a stronger Th1-type immune response and proposed the hypothesis that HAg can be used as a BCG booster to further enhance the benefits of BCG.

An analysis of neutrophil-to-lymphocyte ratios and monocyte-to-lymphocyte ratios with six-month prognosis after cerebral contusions.

Background and purpose: Neutrophil-to-lymphocyte ratio (NLR) and monocyte-to-lymphocyte ratio (MLR) have been identified as potential prognostic markers in various conditions, including cancer, cardiovascular disease, and stroke. This study aims to investigate the dynamic changes of NLR and MLR following cerebral contusion and their associations with six-month outcomes. Methods: Retrospective data were collected from January 2016 to April 2020, including patients diagnosed with cerebral contusion and discharged from two teaching-oriented tertiary hospitals in Southern China. Patient demographics, clinical manifestations, laboratory test results (neutrophil, monocyte, and lymphocyte counts) obtained at admission, 24 hours, and one week after cerebral contusion, as well as outcomes, were analyzed. An unfavorable outcome was defined as a Glasgow Outcome Score (GOS) of 0-3 at six months. Logistic regression analysis was performed to identify independent predictors of prognosis, while receiver characteristic curve analysis was used to determine the optimal cutoff values for NLR and MLR. Results: A total of 552 patients (mean age 47.40, SD 17.09) were included, with 73.19% being male. Higher NLR at one-week post-cerebral contusion (adjusted OR = 4.19, 95%CI, 1.16 - 15.16, P = 0.029) and higher MLR at admission and at 24 h (5.80, 1.40 - 24.02, P = 0.015; 9.06, 1.45 - 56.54, P = 0.018, respectively) were significantly associated with a 6-month unfavorable prognosis after adjustment for other risk factors by multiple logistic regression. The NLR at admission and 24 hours, as well as the MLR at one week, were not significant predictors for a 6-month unfavorable prognosis. Based on receiver operating characteristic curve analysis, the optimal thresholds of NLR at 1 week and MLR at admission after cerebral contusion that best discriminated a unfavorable outcome at 6-month were 6.39 (81.60% sensitivity and 70.73% specificity) and 0.76 (55.47% sensitivity and 78.26% specificity), respectively. Conclusion: NLR measured one week after cerebral contusion and MLR measured at admission may serve as predictive markers for a 6-month unfavorable prognosis. These ratios hold potential as parameters for risk stratification in patients with cerebral contusion, complementing established biomarkers in diagnosis and treatment. However, further prospective studies with larger cohorts are needed to validate these findings.

A dynamic nomogram for predicting intraoperative brain bulge during decompressive craniectomy in patients with traumatic brain injury: a retrospective study.

OBJECTIVE: The aim of this paper is to investigate the risk factors associated with intraoperative brain bulge (IOBB), especially the computed tomography (CT) value of the diseased lateral transverse sinus, and to develop a reliable predictive model to alert neurosurgeons to the possibility of IOBB. METHODS: A retrospective analysis was performed on 937 patients undergoing traumatic decompressive craniectomy. A total of 644 patients from Fuzong Clinical Medical College of Fujian Medical University were included in the development cohort, and 293 patients from the First Affiliated Hospital of Shantou University Medical College were included in the external validation cohort. Univariate and multifactorial logistic regression analyses identified independent risk factors associated with IOBB. The logistic regression models consisted of independent risk factors, and receiver operating characteristic curves, calibration, and decision curve analyses were used to assess the performance of the models. Various machine learning models were used to compare with the logistic regression model and analyze the importance of the factors, which were eventually jointly developed into a dynamic nomogram for predicting IOBB and published online in the form of a simple calculator. RESULTS: IOBB occurred in 93/644 (14.4%) patients in the developmental cohort and 47/293 (16.0%) in the validation cohort. Univariate and multifactorial regression analyses showed that age, subdural hematoma, contralateral fracture, brain contusion, and CT value of the diseased lateral transverse sinus were associated with IOBB. A logistic regression model (full model) consisting of the above risk factors had excellent predictive power in both the development cohort [area under the curve (AUC)=0.930] and the validation cohort (AUC=0.913). Among the four machine learning models, the AdaBoost model showed the best predictive value (AUC=0.998). Factors in the AdaBoost model were ranked by importance and combined with the full model to create a dynamic nomogram for clinical application, which was published online as a practical and easy-to-use calculator. CONCLUSIONS: The CT value of the diseased lateral transverse is an independent risk factor and a reliable predictor of IOBB. The online dynamic nomogram formed by combining logistic regression analysis models and machine learning models can more accurately predict the possibility of IOBBs in patients undergoing traumatic decompressive craniectomy.

Transcriptional analysis of human peripheral blood mononuclear cells stimulated by Mycobacterium tuberculosis antigen.

Background: Mycobacterium tuberculosis antigen (Mtb-Ag) is a polypeptide component with a molecular weight of 10-14 kDa that is obtained from the supernatant of the H37Ra strain after heat treatment. It stimulates the activation and proliferation of γδT cells in the blood to produce an immune response against tuberculosis. Mtb-Ag is therefore crucial for classifying and detecting the central genes and key pathways involved in TB initiation and progression. Methods: In this study, we performed high-throughput RNA sequencing of peripheral blood mononuclear cells (PBMC) from Mtb-Ag-stimulated and control samples to identify differentially expressed genes and used them for gene ontology (GO) and a Kyoto Encyclopedia of Genomes (KEGG) enrichment analysis. Meanwhile, we used PPI protein interaction network and Cytoscape analysis to identify key genes and qRT-PCR to verify differential gene expression. Single-gene enrichment analysis (GSEA) was used further to elucidate the potential biological functions of key genes. Analysis of immune cell infiltration and correlation of key genes with immune cells after Mtb-Ag-stimulated using R language. Results: We identified 597 differentially expressed genes in Mtb-Ag stimulated PBMCs. KEGG and GSEA enrichment analyzed the cellular pathways related to immune function, and DEGs were found to be primarily involved in the TNF signaling pathway, the IL-17 signaling pathway, the JAK-STAT signaling pathway, cytokine-cytokine receptor interactions, and the NF-κB signaling pathway. Wayne analysis using GSEA, KEGG, and the protein-protein interaction (PPI) network showed that 34 genes, including PTGS2, IL-1ß, IL-6, TNF and IFN-γ et al., were co-expressed in the five pathways and all were up-regulated by Mtb-Ag stimulation. Twenty-four DEGs were identified using qRT-PCR, including fourteen up-regulated genes (SERPINB7, IL20, IFNG, CSF2, PTGS2, TNF-α, IL36G, IL6, IL10, IL1A, CXCL1, CXCL8, IL4, and CXCL3) and ten down-regulated genes (RTN1, CSF1R CD14, C5AR1, CXCL16, PLXNB2, OLIG1, EEPD1, ENG, and CCR1). These findings were consistent with the RNA-Seq results. Conclusion: The transcriptomic features associated with Mtb-Ag provide the scientific basis for exploring the intracellular immune mechanisms against Mtb. However, more studies on these DEGs in pathways associated with Mtb-Ag stimulation are needed to elucidate the underlying pathologic mechanisms of Mtb-Ag during Mtb infection.

Low serum calcium is a novel predictor of unfavorable prognosis after traumatic brain injury.

Background: Accurate and convenient serological markers for prognosis after traumatic brain injury (TBI) are still lacking. We aimed to explore the predictive value of serum calcium for prognosing outcomes within 6 months after TBI. Methods: In this multicenter retrospective study, 1255 and 719 patients were included in development and validation cohorts, respectively, and their 6-month prognoses were recorded. Serum calcium was measured through routine blood tests within 24 h of hospital admission. Two multivariate predictive models with or without serum calcium for prognosis were developed. Receiver operating characteristics and calibration curves were applied to estimate their performance. Results: The patients with lower serum calcium levels had a higher frequency of unfavorable 6-month prognosis than those without. Lower serum calcium level at admission was associated with an unfavorable 6-month prognosis in a wide spectrum of patients with TBI. Lower serum calcium level and our prognostic model including calcium performed well in predicting the 6-month unfavorable outcome. The calcium nomogram maintained excellent performance in discrimination and calibration in the external validation cohort. Conclusions: Lower serum calcium level upon admission is an independent risk factor for an unfavorable 6-month prognosis after TBI. Integrating serum calcium into a multivariate predictive model improves the performance for predicting 6-month unfavorable outcomes.

High-throughput sequencing-based Detection of Japanese encephalitis virus and its effect on micro ribonucleic acid.

it was to explore the mechanism of Japanese encephalitis virus (JEV) and micro ribonucleic acid (miRNA) under high-throughput sequencing. 20 experimental mice, with good growth status and no disease infection, were selected. The cells used in the experiment included mouse microglial cell line (BV2), mouse neuroblastoma cell line (NA), and mouse brain endothelial cell line (bEnd.3). JEV titration was performed with JEV-infected cells, ribonucleic acid (RNA) in the cells was extracted, and finally the miRNA high-throughput sequencing data was analyzed. Agarose gel electrophoresis showed that the 28S and 18S electrophoresis bands were bright. Among the miRNAs detected in mouse brain tissues, 2986 were down-regulated and 1251 were up-regulated. Among miRNAs detected in NA cells, 4238 the decreasing expression and 2356 were expressed increasingly. In reducing miRNA expression, 1 multiplicity of infection (MOI) of P3 strain infection was more significant than 0.1 MOI. 10 miRNAs with significantly decreasing expression were miR-466d-3p, miR-381-3p, miR-540-3p, miR-466a-3p, miR-467a-3p, miR-574-5p, miR-199a-5p, miR-467a-5p, miR-674-5p, and miR-376b-3p. These were all obviously down-regulated in JEV-infected BV2, NA, and bEnd.3 neurons. High-throughput sequencing of JEV-infected mouse brain tissues and mouse neuronal cells found that JEV infection led to down-regulation of overall miRNA expression in host cells.

A dynamic nomogram for predicting unfavorable prognosis after aneurysmal subarachnoid hemorrhage.

OBJECTIVE: The aim of this study was to examine the predictive value of the multiplication of neutrophil and monocyte counts (MNM) in peripheral blood, and develop a new predictive model for the prognosis of patients with aneurysmal subarachnoid hemorrhage (aSAH). METHODS: This is a retrospective analysis that included 2 separate cohorts of patients undergoing endovascular coiling for aSAH. The training cohort consisted of 687 patients in the First Affiliated Hospital of Shantou University Medical College; the validation cohort consisted of 299 patients from Sun Yat-sen University's Affiliated Jieyang People's Hospital. The training cohort was used to develop 2 models to predict unfavorable prognosis (modified Rankin scale of 3-6 at 3 months): one was based on traditional factors (e.g., age, modified Fisher grade, NIHSS score, and blood glucose), and another model that included traditional factors as well as MNM on admission. RESULTS: In the training cohort, MNM upon admission was independently associated with unfavorable prognosis (odds ratio after adjustment, 1.06; 95% confidence interval [CI], 1.03-1.10). In the validation cohort, the basic model that included only traditional factors had 70.99% sensitivity, 84.36% specificity, and 0.859 (95% CI, 0.817-0.901) area under the receiver operating characteristic curve (AUC). Adding MNM increased model sensitivity (from 70.99% to 76.48%), specificity (from 84.36% to 88.63%), and overall performance (AUC 0.859 [95% CI, 0.817-0.901] to 0.879 [95% CI, 0.841-0.917]). INTERPRETATION: MNM upon admission is associated with unfavorable prognosis in patients undergoing endovascular embolization for aSAH. The nomogram including MNM is a user-friendly tool to help clinicians quickly predict the outcome of patients with aSAH.

Association between plasma metal elements and platelet dysfunction in trauma-induced coagulopathy rat model.

BACKGROUND: Disorders of metal elements and platelet dysfunction are common in patients with trauma-induced coagulopathy (TIC). AIM: The aim of this study was to explore the potential role of plasma metal elements in platelet dysfunction in TIC. METHODS: Thirty Sprague-Dawley rats were divided into control, hemorrhage shock (HS) and multiple injury (MI) groups. At timepoints of 0.5 and 3 h after trauma and being documented as HS 0.5 h, HS3 h, MI 0.5 h or MI3 h, blood samples were harvested for inductively coupled plasma mass spectrometer, conventional coagulation function and thromboelastograph. RESULTS: The plasma zinc (Zn), vanadium (V) and cadmium (Ca) decreased initially in HS 0.5 h and recovered slightly in HS3 h, whereas their plasma concentrations continued to decrease from beginning till MI3 h (p < 0.05). In HS, plasma Ca, V and nickel were negatively correlated to the time taken to reach the initial formation (R), whereas R was positively correlated to plasms Zn, V, Ca and selenium in MI (p < 0.05). In MI, plasma Ca was positively correlated to maximum amplitude, and plasma V was positively correlated to platelet count (p < 0.05). CONCLUSION: The plasma concentrations of Zn, V and Ca appeared to contribute to platelet dysfunction in HS 0.5 h, HS3 h, MI 0.5 h and MI3 h, which were trauma type sensitive.

Lidocaine inhibits influenza a virus replication by up-regulating IFNα4 via TBK1-IRF7 and JNK-AP1 signaling pathways.

Influenza A viruses (IAV), significant respiratory pathogenic agents, cause seasonal epidemics and global pandemics in intra- and interannual cycles. Despite effective therapies targeting viral proteins, the continuous generation of drug-resistant IAV strains is challenging. Therefore, exploring novel host-specific antiviral treatment strategies is urgently needed. Here, we found that lidocaine, widely used for local anesthesia and sedation, significantly inhibited H1N1(PR8) replication in macrophages. Interestingly, its antiviral effect did not depend on the inhibition of voltage-gated sodium channels (VGSC), the main target of lidocaine for anesthesia. Lidocaine significantly upregulated early IFN-I, interferon α4 (IFNα4) mRNA, and protein levels, but not those of early IFNß in mouse RAW 264.7 cell line and human THP-1 derived macrophages. Knocking out IFNα4 by CRISPR-Cas9 partly reversed lidocaine's inhibition of PR8 replication in macrophages. Mechanistically, lidocaine upregulated IFNα4 by activating TANK-binding kinase 1 (TBK1)-IRF7 and JNK-AP1 signaling pathways. These findings indicate that lidocaine has an incredible antiviral potential by enhancing IFN-I signaling in macrophages. In conclusion, our results indicate the potential auxiliary role of lidocaine for anti-influenza A virus therapy and even for anti-SARS-CoV-2 virus therapy, especially in the absence of a specific medicine.

Artificial Intelligence-Based Diagnostic Model for Detecting Keratoconus Using Videos of Corneal Force Deformation.

Purpose: To develop a novel method based on biomechanical parameters calculated from raw corneal dynamic deformation videos to quickly and accurately diagnose keratoconus using machine learning. Methods: The keratoconus group was included according to Rabinowitz's criteria, and the normal group included corneal refractive surgery candidates. Independent biomechanical parameters were calculated from dynamic corneal deformation videos. A novel neural network model was trained to diagnose keratoconus. Tenfold cross-validation was performed, and the sample set was divided into a training set for training, a validation set for parameter validation, and a testing set for performance evaluation. External validation was performed to evaluate the model's generalizability. Results: A novel intelligent diagnostic model for keratoconus based on a five-layer feedforward network was constructed by calculating four biomechanical characteristics, including time of the first applanation, deformation amplitude at the highest concavity, central corneal thickness, and radius at the highest concavity. The model was able to diagnose keratoconus with 99.6% accuracy, 99.3% sensitivity, 100% specificity, and 100% precision in the sample set (n = 276), and it achieved an accuracy of 98.7%, sensitivity of 97.4%, specificity of 100%, and precision of 100% in the external validation set (n = 78). Conclusions: In the absence of corneal topographic examination, rapid and accurate diagnosis of keratoconus is possible with the aid of machine learning. Our study provides a new potential approach and sheds light on the diagnosis of keratoconus from a purely corneal biomechanical perspective. Translational Relevance: Our findings could help improve the diagnosis of keratoconus based on corneal biomechanical properties.

TGF-ß1 Protects Trauma-injured Murine Cortical Neurons by Upregulating L-type Calcium Channel Cav1.2 via the p38 Pathway.

Traumatic brain injury (TBI) is a leading cause of disability and death in adolescents, and there is a lack of effective methods of treatment. The neuroprotective effects exerted by TGF-ß1 can ameliorate a range of neuronal lesions in multiple central nervous system diseases. In this study, we used an in-vitro TBI model of mechanical injury on murine primary cortical neurons and the neuro-2a cell line to investigate the neuroprotective role played by TGF-ß1 in cortical neurons in TBI. Our results showed that TGF-ß1 significantly increased neuronal viability and inhibited apoptosis for 24 h after trauma. The expression of Cav1.2, an L-type calcium channel (LTCC) isoform, decreased significantly after trauma injury, and this change was reversed by TGF-ß1. Nimodipine, a classic LTCC blocker, abolished the protective effect of TGF-ß1 on trauma-induced neuronal apoptosis. The knockdown of Cav1.2 in differentiated neuro-2a cells significantly inhibited the anti-apoptosis effect of TGF-ß1 exerted on injured neuro-2a cells. Moreover, TGF-ß1 rescued and enhanced the trauma-suppressed neuro-2a intracellular Ca2+ concentration, while the effect of TGF-ß1 was partially inhibited by nimodipine. TGF-ß1 significantly upregulated the expression of Cav1.2 by activating the p38 MAPK pathway and by inhibiting trauma-induced neuronal apoptosis. In conclusion, TGF-ß1 increased trauma-injured murine cortical neuronal activity and inhibited apoptosis by upregulating Cav1.2 channels via activating the p38 MAPK pathway. Therefore, the TGF-ß1/p38 MAPK/Cav 1.2 pathway has the potential to be used as a novel therapeutic target for TBI.

A Clinical Predictive Nomogram for Traumatic Brain Parenchyma Hematoma Progression.

INTRODUCTION: Acute traumatic intraparenchymal hematoma (tICH) expansion is a major cause of clinical deterioration after brain contusion. Here, an accurate prediction tool for acute tICH expansion is proposed. METHODS: A multicenter hospital-based study for multivariable prediction model was conducted among patients (889 patients in a development dataset and 264 individuals in an external validation dataset) with initial and follow-up computed tomography (CT) imaging for tICH volume evaluation. Semi-automated software was employed to assess tICH expansion. Two multivariate predictive models for acute tICH expansion were developed and externally validated. RESULTS: A total of 198 (22.27%) individuals had remarkable acute tICH expansion. The novel Traumatic Parenchymatous Hematoma Expansion Aid (TPHEA) model retained several variables, including age, coagulopathy, baseline tICH volume, time to baseline CT time, subdural hemorrhage, a novel imaging marker of multihematoma fuzzy sign, and an inflammatory index of monocyte-to-lymphocyte ratio. Compared with multihematoma fuzzy sign, monocyte-to-lymphocyte ratio, and the basic model, the TPHEA model exhibited optimal discrimination, calibration, and clinical net benefits for patients with acute tICH expansion. A TPHEA nomogram was subsequently introduced from this model to facilitate clinical application. In an external dataset, this device showed good predicting performance for acute tICH expansion. CONCLUSIONS: The main predictive factors in the TPHEA nomogram are the monocyte-to-lymphocyte ratio, baseline tICH volume, and multihematoma fuzzy sign. This user-friendly tool can estimate acute tICH expansion and optimize personalized treatments for individuals with brain contusion.

Neutrophil to lymphocyte ratio predicts early growth of traumatic intracerebral haemorrhage.

OBJECTIVE: The neutrophil to lymphocyte ratio (NLR) has been proposed to capture the inflammatory status of patients with various conditions involving the brain. This retrospective study aimed to explore the association between the NLR and the early growth of traumatic intracerebral haemorrhage (tICH) in patients with traumatic brain injury (TBI). METHODS: A multicentre, observational cohort study was conducted. Patients with cerebral contusion undergoing baseline computed tomography for haematoma volume analysis within 6 h after primary injury and follow-up visits within 48 h were included. Routine blood tests were performed upon admission, and early growth of tICH was assessed. Prediction accuracies of the NLR for the early growth of tICH and subsequent surgical intervention in patients were analysed. RESULTS: There were a total of 1077 patients who met the criteria included in the study cohort. Univariate analysis results showed that multiple risk factors were associated with the early growth of tICH and included in the following multivariate analysis models. The multivariate logistic regression analysis results revealed that the NLR was highly associated with the early growth of tICH (p < 0.001) while considering other risk factors in the same model. The prediction accuracy of the NLR for the early growth of tICH in patients is 82%. INTERPRETATION: The NLR is easily calculated and might predict the early growth of tICH for patients suffering from TBI.

Development and external validation of a novel multihematoma fuzzy sign on computed tomography for predicting traumatic intraparenchymal hematoma expansion.

Acute traumatic intraparenchymal hematoma (tICH) expansion is a devastating neurological complication that is associated with poor outcome after cerebral contusion. This study aimed to develop and validate a novel noncontrast computed tomography (CT) (NCCT) multihematoma fuzzy sign to predict acute tICH expansion. In this multicenter, prospective cohort study, multihematoma fuzzy signs on baseline CT were found in 212 (43.89%) of total 482 patients. Patients with the multihematoma fuzzy sign had a higher frequency of tICH expansion than those without (90.79% (138) vs. 46.71% (71)). The presence of multihematoma fuzzy sign was associated with increased risk for acute tICH expansion in entire cohort (odds ratio [OR]: 16.15; 95% confidence interval (CI) 8.85-29.47; P < 0.001) and in the cohort after propensity-score matching (OR: 9.37; 95% CI 4.52-19.43; P < 0.001). Receiver operating characteristic analysis indicated a better discriminative ability of the presence of multihematoma fuzzy sign for acute tICH expansion (AUC = 0.79; 95% CI 0.76-0.83), as was also observed in an external validation cohort (AUC = 0.76; 95% CI 0.67-0.84). The novel NCCT marker of multihematoma fuzzy sign could be easily identified on baseline CT and is an easy-to-use predictive tool for tICH expansion in the early stage of cerebral contusion.

Development of a multivalent enterovirus subunit vaccine based on immunoinformatic design principles for the prevention of HFMD.

Hand, foot and mouth disease (HFMD) is mainly caused by EV-A71 and CV-A16. An increasing number of cases have been found to be caused by CV-A10, CV-A6, CV-B3 and the outbreaks are becoming increasingly more complex, often accompanied by the prevalence of a variety of enteroviruses. Based on the principle of synthetic peptide vaccines, we applied immune-informatics to design a highly efficient and safe multivalent epitope-based vaccine against EV-A71, CV-A16, CV-A10, CV-A6 and CV-B3. By screening B-cells, HTL and CTL cell antigen epitopes with high conservativity and immunogenicity that have no toxic effect on the host, further analysis confirmed that the vaccine built was IFN-γ inductive and IL-4 non-inductive HTL cell epitopes and had population coverage corresponding to MHC molecular alleles associated with T-cell phenotype. The multivalent enterovirus vaccine was constructed to connect the 50 s ribosomal protein L7/L12 adjuvant and candidate epitopes sequentially through appropriate linkers. Then, the antigenic, allergen and physical properties of the vaccine were evaluated, followed by a secondary structure analysis and tertiary structure modeling, disulfide engineering, refinement and validation. Moreover, the conformational B cell epitope of the vaccine was analyzed. The stability of the TLR4/MD2/Vaccine complex and details at atomic level were investigated by docking and molecular dynamics simulation. Finally, in silico immune simulation and in vivo immune experiments were done. This study provides a high cost-effective method of designing a multivalent enterovirus vaccine protect against a wide range of enterovirus pathogens.

The Monocyte-to-Lymphocyte Ratio at Hospital Admission Is a Novel Predictor for Acute Traumatic Intraparenchymal Hemorrhage Expansion after Cerebral Contusion.

PURPOSE: To explore the potential of monocyte-to-lymphocyte ratio (MLR) at hospital admission for predicting acute traumatic intraparenchymal hematoma (tICH) expansion in patients with cerebral contusion. Patients and Methods. This multicenter, observational study included patients with available at-hospital admission (baseline) and follow-up computed tomography for volumetric analysis (retrospective development cohort: 1146 patients; prospective validation cohort: 207 patients). Semiautomated software assessed tICH expansion (defined as ≥33% or 5 mL absolute growth). MLR was acquired from routine blood tests upon admission. We constructed two predictive models: basic combined model of clinical and imaging variables and MLR combined model of both MLR and other variables in the basic model. Receiver operating characteristic (ROC) analysis and decision curve analysis (DCA) were used to estimate the performance of MLR for predicting acute tICH expansion. RESULTS: MLR was significantly larger in patients with acute tICH expansion compared to those without acute tICH expansion (mean [SD], 1.08 [1.05] vs. 0.59 [0.37], P < 0.001). A nonlinear positive relationship between MLR and the incidence of acute tICH expansion was observed. Multivariate logistic regression indicated MLR as an independent risk factor for acute tICH expansion (odds ratio (OR), 5.88; 95% confidence interval (CI), 4.02-8.61). The power of the multivariate model for predicting acute tICH expansion was substantially improved with the inclusion of MLR (AUC 0.86 vs. AUC 0.74, P < 0.001), as was also observed in an external validation cohort (AUC 0.83 vs. AUC 0.71, P < 0.001). The net benefit of MLR model was higher between threshold probabilities of 20-100% in DCA. For clinical application, a nomogram derived from the multivariate model with MLR was introduced. In addition, MLR was positively associated with 6-month unfavorable outcome. CONCLUSION: MLR is a novel predictor for traumatic parenchymatous hematoma expansion. A nomogram derived from the MLR model may provide an easy-to-use tool for predicting acute tICH expansion and promoting the individualized treatment of patients with hemorrhagic cerebral contusion. MLR is associated with long-term outcome after cerebral contusion.

Dynamic Changes in the Microbiome and Mucosal Immune Microenvironment of the Lower Respiratory Tract by Influenza Virus Infection.

Influenza is a major public health concern, and the high mortality rate is largely attributed to secondary bacterial infections. There are several mechanisms through which the virus increases host susceptibility to bacterial colonization, but the micro-environment in lower respiratory tract (LRT) of host, infected with influenza virus, is unclear. To this end, we analyzed the LRT microbiome, transcriptome of lung and metabolome of bronchoalveolar lavage fluid (BALF) in mice inoculated intra-nasally with H1N1 to simulate human influenza, and we observed significant changes in the composition of microbial community and species diversity in the acute (7 days post inoculation or dpi), convalescent (14 dpi) and the recovery (28 dpi) periods. The dominant bacterial class shifted from Alphaproteobacteria to Gammaproteobacteria and Actinobacteria in the infected mice, with a significant increase in the relative abundance of anaerobes and facultative anaerobes like Streptococcus and Staphylococcus. The dysbiosis in the LRT of infected mice was not normalized even in the recovery phase of the infection. In addition, the infected lung transcriptome showed significant differences in the expression levels of genes associated with bacterial infection and immune responses. Finally, the influenza virus infection also resulted in significant changes in the metabolome of the BALF. These alterations in the microbiome, transcriptome, and metabolome of infected lungs were not only appeared at the acute period, but also observed at the recovery period. Furthermore, the infection of influenza virus induced a long-term effect in LRT micro-environmental homeostasis, which may give a chance for the invasion of potential pathogens.

Differential responses of stressful elements to predatory exposure in behavior-lateralized mice.

BACKGROUND: Predatory stress as a psychological stressor can elicit the activation of the hypothalamic-pituitary-adrenal (HPA) axis, which is involved in the dialogue of the neuroimmunoendocrine network. The brain has been proven to regulate the activity of the HPA axis by way of lateralization. In the present study, we probed the pivotal elements of the HPA circuitry including CRH, GR and a multifunctional cytokine in behavior-lateralized mice to determine their changes when the animals were subjected to predator exposure. METHODS: Behavior-lateralized mice were classified into left-pawed and right-pawed mice through a paw-preference test. Thereafter, mice in the acute stress group received a single 60-min cat exposure, and mice in the chronic group received daily 60-min cat exposure for 14 consecutive days. The plasma CS and TNF-α were determined by ELISA, the hypothalamic CRH mRNA and hippocampal GR mRNA were detected by real-time PCR, and the hippocampal GR protein was detected by western blot analysis. RESULTS: The results revealed that the levels of plasma CS were significantly elevated after chronic predatory exposure in both right-pawed and left-pawed mice; the right-pawed mice exhibited a higher plasma CS level than the left-pawed mice. Similarly, the acute or chronic cat exposure could induce the release of plasma TNF-α, and the left-pawed mice tended to show a higher level after the acute stress. Chronic stress significantly upregulated the expression of hypothalamic CRH mRNA in both left-pawed and right-pawed mice. Normally, the left-pawed mice exhibited a higher GR expression in the hippocampus than the right-pawed mice. After the cat exposure, the expression of GR in both left-pawed and right-pawed mice was revealed to be greatly downregulated. CONCLUSION: Our findings indicate that predatory stress can invoke a differential response of stressful elements in behavior-lateralized mice. Some of these responses shaped by behavioral lateralization might be helpful for facilitating adaption to various stimuli.

Oxymatrine Inhibits Influenza A Virus Replication and Inflammation via TLR4, p38 MAPK and NF-κB Pathways.

Oxymatrine (OMT) is a strong immunosuppressive agent that has been used in the clinic for many years. In the present study, by using plaque inhibition, luciferase reporter plasmids, qRT-PCR, western blotting, and ELISA assays, we have investigated the effect and mechanism of OMT on influenza A virus (IAV) replication and IAV-induced inflammation in vitro and in vivo. The results showed that OMT had excellent anti-IAV activity on eight IAV strains in vitro. OMT could significantly decrease the promoter activity of TLR3, TLR4, TLR7, MyD88, and TRAF6 genes, inhibit IAV-induced activations of Akt, ERK1/2, p38 MAPK, and NF-κB pathways, and suppress the expressions of inflammatory cytokines and MMP-2/-9. Activators of TLR4, p38 MAPK and NF-κB pathways could significantly antagonize the anti-IAV activity of OMT in vitro, including IAV replication and IAV-induced cytopathogenic effect (CPE). Furthermore, OMT could reduce the loss of body weight, significantly increase the survival rate of IAV-infected mice, decrease the lung index, pulmonary inflammation and lung viral titter, and improve pulmonary histopathological changes. In conclusion, OMT possesses anti-IAV and anti-inflammatory activities, the mechanism of action may be linked to its ability to inhibit IAV-induced activations of TLR4, p38 MAPK, and NF-κB pathways.