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.

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.

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.

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.

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.

NS1 of H7N9 Influenza A Virus Induces NO-Mediated Cellular Senescence in Neuro2a Cells.

BACKGROUND/AIMS: The novel avian H7N9 influenza A virus has been detected in brain tissues and associated with central nervous system (CNS) symptoms in infected human and mice. Roles of its virulence factor, NS1 protein in influenza virus infected neuron has yet to be explored. METHODS: Nitric oxide (NO) release and inducible nitric oxide synthase (iNOS) expression in H7N9/NS1-expressed Neuro2a cells were detected by Griess test and western blotting. Cell proliferation rate of H7N9/NS1-expressing cells was recorded by Cell Counting Kit-8. Effects of H7N9/NS1 on cellular senescence were investigated by senescence-associated ß-galactosidase (SA-ß-gal) staining, immunofluorescent staining of phosphorylated heterochromatin protein 1γ (pHP1γ) and qPCR analysis of IL-6 and IL-8. RESULTS: H7N9/NS1 in Neuro2a cells and primary cultured mouse cortical neurons increased the expression of iNOS and boosted NO release. Neuro2a cells constitutively expressing NS1 displayed a reduced proliferative ability, enhanced SA-ß-gal staining, increased level of IL-6 and IL-8 and a typical punctuate structure of pHP1γ in nuclei. In addition, p38 MAPK was elevated in NS1-expressing Neuro2a cells. Reduced iNOS expression and subdued cellular senescence effect was found in p38 MAPK inhibitor-treated NS1-expressing Neuro2a cells. CONCLUSION: Our results suggest that H7N9/NS1 protein increases the iNOS expression and NO release in Neuro2a cells, which can induce cell growth arrest and cellular senescence.

Cytokine cascades induced by mechanical trauma injury alter voltage-gated sodium channel activity in intact cortical neurons.

BACKGROUND: Traumatic brain injury (TBI) triggers both immediate (primary) and long-term (secondary) tissue damages. Secondary damages can last from hours to days or even a lifetime. Secondary damages implicate several mechanisms, including influence of inflammatory mediators, mainly cytokines, on excitability of ion channels. However, studies should further explore the effects of inflammatory cytokines on voltage-gated sodium channels (VGSCs) and excitability in distal intact neurons. METHODS: Mixed cultures of mouse cortical astrocytes and neurons were subjected to mechanical injury (trauma) to mimic TBI in vitro. Expression of various cytokines in these cultures were measured by real-time polymerase chain reaction and enzyme-linked immunosorbent assay. A trauma-conditioned medium with or without brain-derived neurotrophic factor (BDNF) was added to mouse primary cortical neurons for 6 and 24 h to mimic combined effects of multiple inflammatory cytokines on VGSCs. Spike behaviors of distal intact neurons were examined by whole-cell patch-clamp recordings. RESULTS: Mechanical injury in mixed cortical neuron-astrocyte cultures significantly increased expression levels of multiple cytokines, including interleukin (IL)-1ß, IL-6, tumor necrosis factor-α, monocyte chemoattractant protein-1, chemokine (C-C motif) ligand-5, IL-10, and transforming growth factor-ß1, at 6 and 24 h after injury. Incubation in trauma-conditioned medium increased functional VGSCs in neuronal membranes and Na+ currents. Enhanced VGSCs were almost completely abolished by BDNF, and reinforcement of Na+ currents was also reduced in a dose-dependent manner. BDNF (30 ng/mL) also significantly reversed reduced neuronal cell viability, which was induced by medium conditioned at 6 h. At 6 and 24 h, trauma-conditioned medium significantly increased spike frequency but not spike threshold. CONCLUSIONS: In TBI, the combined effect of inflammatory cytokines is directly involved in VGSC, Na+ current, and excitability dysfunction in distal intact neurons. BDNF may partly exert neuroprotective effects by maintaining balance of VGSC function in distal intact neurons.

Emodin Inhibition of Influenza A Virus Replication and Influenza Viral Pneumonia via the Nrf2, TLR4, p38/JNK and NF-kappaB Pathways.

Lasting activations of toll-like receptors (TLRs), MAPK and NF-κB pathways can support influenza A virus (IAV) infection and promote pneumonia. In this study, we have investigated the effect and mechanism of action of emodin on IAV infection using qRT-PCR, western blotting, ELISA, Nrf2 luciferase reporter, siRNA and plaque inhibition assays. The results showed that emodin could significantly inhibit IAV (ST169, H1N1) replication, reduce IAV-induced expressions of TLR2/3/4/7, MyD88 and TRAF6, decrease IAV-induced phosphorylations of p38/JNK MAPK and nuclear translocation of NF-κB p65. Emodin also activated the Nrf2 pathway, decreased ROS levels, increased GSH levelss and GSH/GSSG ratio, and upregulated the activities of SOD, GR, CAT and GSH-Px after IAV infection. Suppression of Nrf2 via siRNA markedly blocked the inhibitory effects of emodin on IAV-induced activations of TLR4, p38/JNK, and NF-κB pathways and on IAV-induced production of IL-1ß, IL-6 and expression of IAV M2 protein. Emodin also dramatically increased the survival rate of mice, reduced lung edema, pulmonary viral titer and inflammatory cytokines, and improved lung histopathological changes. In conclusion, emodin can inhibit IAV replication and influenza viral pneumonia, at least in part, by activating Nrf2 signaling and inhibiting IAV-induced activations of the TLR4, p38/JNK MAPK and NF-κB pathways.

The Novel H7N9 Influenza A Virus NS1 Induces p53-Mediated Apoptosis of A549 Cells.

BACKGROUND: H7N9, emerged as an avian influenza virus outbreak in Eastern China in early 2013, and represented another major threat to global health. Roles of its NS1 protein, an essential viral factor, in regulating apoptosis remain unknown. METHODS: Apoptotic effect and features of H7N9/NS1 in the human A549 alveolar basal epithelial cell line were examined by caspase 3/7 activity assay and western blotting of apoptotic associated proteins. Effects of H7N9/NS1on mitochondrial membrane potential were investigated by flow cytometry. RESULTS: The expression of H7N9/NS1 in A549 cells activated caspase 3/7 and increased the protein levels of cleaved caspase 7 and cleaved poly (ADP-ribose) polymerase (PARP). H7N9/NS1-expressing A549 cells displayed a decrease in mitochondrial membrane potential. In addition, H7N9/NS1 increased the protein levels of total p53, p53 phosphorylated at Ser46 and Ser37, activated caspase 9, and the Bax/Bcl-2 ratio. CONCLUSION: Our results suggest that H7N9/NS1 protein causes the accumulation of p53 by increasing phosphorylation levels of p53 and the induction of mitochondrial dysfunction, which may contribute to H7N9/NS1-induced apoptosis in A549 cells.

LPS-Primed Release of HMGB-1 from Cortical Astrocytes is Modulated Through PI3K/AKT Pathway.

Studies have shown that LPS-preconditioned tolerant state could protect against brain injury to subsequent challenges. We hypothesized astrocytes were directly involved in the readjustment to confer neuroprotective effects with LPS pretreatment. High-mobility group box 1(HMGB-1) from LPS-preconditioned astrocytes, presumably serving as a positive regulator, might contribute to the favorable preconditioned effects. Furthermore, a potential cellular pathway (PI3K/AKT pathway), has been proposed for the active regulation of LPS-primed reactive astrocytes to secrete HMGB-1. In the present study, we used a low concentration of LPS to directly prime the astrocytes in vitro, and the subsequent astrocytic reactions, including cytokine secretion, the expression of transcription factors, and the release of HMGB-1 were examined after the blockade of the PI3K pathway. The data showed that LPS preconditioning could reduce some capacity of astrocytes to subsequent challenge in vitro. PI3K/AKT pathway was partially involved in the modulation of the release HMGB-1 from reactive astrocytes. These findings offer direct evidence supporting the flexible roles of astrocytes in mediating LPS-primed neuroprotection, and highlight additional targets for future attempts to modify the protective effects of astrocytes through LPS preconditioning.

Tumor necrosis factor-α enhances voltage-gated Na⁺ currents in primary culture of mouse cortical neurons.

BACKGROUND: Previous studies showed that TNF-α could activate voltage-gated Na(+) channels (VGSCs) in the peripheral nervous system (PNS). Since TNF-α is implicated in many central nervous system (CNS) diseases, we examined potential effects of TNF-α on VGSCs in the CNS. METHODS: Effects of TNF-α (1-1000 pg/mL, for 4-48 h) on VGSC currents were examined using whole-cell voltage clamp and current clamp techniques in primary culture of mouse cortical neurons. Expression of Nav1.1, Nav1.2, Nav1.3, and Nav1.6 were examined at both the mRNA and protein levels, prior to and after TNF-α exposure. RESULTS: TNF-α increased Na(+) currents by accelerating the activation of VGSCs. The threshold for action potential (AP) was decreased and firing rate were increased. VGSCs were up-regulated at both the mRNA and protein levels. The observed effects of TNF-α on Na(+) currents were inhibited by pre-incubation with the NF-κB inhibitor BAY 11-7082 (1 µM) or the p38 mitogen-activated protein kinases (MAPK) inhibitor SB203580 (1 µM). CONCLUSIONS: TNF-α increases Na(+) currents by accelerating the channel activation as well as increasing the expression of VGSCs in a mechanism dependent upon NF-κB and p38 MAPK signal pathways in CNS neurons.

Chloroquine enhances replication of influenza A virus A/WSN/33 (H1N1) in dose-, time-, and MOI-dependent manners in human lung epithelial cells A549.

Anti-malaria drug, chloroquine, has been reported to be effective against influenza A virus (IAV) in vitro and used in in-vivo experiments and clinical trial for prevention or treatment of influenza. In this study, it has been shown by immunofluorescence, hemagglutination, and plaque assays that chloroquine enhanced A/WSN/33 (H1N1) replication with pronounced cytopathic effect in dose-, time-, and MOI-dependent manners in human lung epithelial cells A549. Time-of-addition assay showed that inhibitory effect on virus replication by chloroquine pre-treatment was indistinctive, and virus productions were enhanced when the drug was applied after viral adsorption. The effectiveness of chloroquine as an anti-influenza drug is questioned, and caution in its use is recommended.

Inhibition of Tanshinone IIA, salvianolic acid A and salvianolic acid B on Areca nut extract-induced oral submucous fibrosis in vitro.

Salvia miltiorrhiza Bunge has been reported to possess excellent antifibrotic activity. In this study, we have investigated the effect and mechanism of tanshinone IIA (Tan-IIA), salvianolic acid A (Sal-A) and salvianolic acid B (Sal-B), the important active compounds of Salvia miltiorrhiza Bunge, on areca nut extract (ANE)-induced oral submucous fibrosis (OSF) in vitro. Through human procollagen gene promoter luciferase reporter plasmid assay, hydroxyproline assay, gelatin zymography assay, qRT-PCR, ELISA and Western blot assay, the influence of these three compounds on ANE-stimulated cell viability, collagen accumulation, procollagen gene transcription, MMP-2/-9 activity, MMP-1/-13 and TIMP-1/-2 expression, cytokine secretion and the activation of PI3K/AKT, ERK/JNK/p38 MAPK and TGF-ß/Smads pathways were detected. The results showed that Tan-IIA, Sal-A and Sal-B could significantly inhibit the ANE-stimulated abnormal viability and collagen accumulation of mice oral mucosal fibroblasts (MOMFs), inhibit the transcription of procollagen gene COL1A1 and COL3A1, increase MMP-2/-9 activity, decrease TIMP-1/-2 expression and inhibit the transcription and release of CTGF, TGF-ß1, IL-6 and TNF-α; Tan-IIA, Sal-A and Sal-B also inhibited the ANE-induced activation of AKT and ERK MAPK pathways in MOMFs and the activation of TGF-ß/Smads pathway in HaCaT cells. In conclusion, Tan-IIA, Sal-A and Sal-B possess excellent antifibrotic activity in vitro and can possibly be used to promote the rehabilitation of OSF patients.

Chronic haloperidol increases voltage-gated Na+ currents in mouse cortical neurons.

Typical antipsychotics are characterized by extrapyramidal syndrome (EPS). Previous studies demonstrated that typical antipsychotics could inhibit neuronal voltage-gated sodium channel (VGSC). However, EPS typically emerge only upon prolonged exposure. As a result, we examined effects of haloperidol, a prototype typical antipsychotic, on neuronal VGSC upon incubation for varying duration. Briefly, VGSC currents were activated and recorded using a whole-cell patch-clamp technique in primary culture of mouse cortical neurons. VGSC activity was inhibited by acute haloperidol exposure (for minutes), but enhanced in a time- and concentration-dependent manner by chronic haloperidol exposure (for hours). The effects of chronic haloperidol were associated with increased expression of VGSC subunits as well as corresponding electrophysiological channel properties. In summary, we found enhanced VGSC currents upon chronic haloperidol exposure in cortical neurons in contrast to inhibition by acute haloperidol exposure. Such a results may contribute to EPS of typical antipsychotics.

Panax notoginseng saponins inhibit areca nut extract-induced oral submucous fibrosis in vitro.

BACKGROUND: Oral submucous fibrosis (OSF) is a premalignant and fibrosing disease, which is closely associated with the habit of chewing areca nut. Panax notoginseng Buck F. H. Chen is an often used antifibrotic and antitumor agent. To treat areca nut-induced OSF, we have developed a chewable tablet, in which one of the major medicines is total Panax notoginseng saponins (PNS). In this study, we have investigated the antifibrotic effect and mechanism of PNS on areca nut-induced OSF in vitro. METHODS: Through human procollagen gene promoter luciferase reporter plasmid, hydroxyproline assay, gelatin zymography, qRT-PCR, ELISA, and Western blot, the influences of PNS on areca nut extract (ANE)-induced cell growth, collagen accumulation, procollagen gene transcription, MMP-2/-9 activity, MMP-1/-13 and TIMP-1/-2 expression, cytokine secretion, and the activation of PI3K/AKT, ERK/JNK/p38 MAPK, and TGFß/Smads pathways were detected. RESULTS: Panax notoginseng saponins could inhibit the ANE-induced abnormal growth and collagen accumulation of oral mucosal fibroblasts in a concentration-dependent manner. PNS (25 µg/ml) could significantly inhibit the ANE-induced expression of Col1A1 and Col3A1, augment the ANE-induced decrease of MMP-2/-9 activity, inhibit the ANE-induced increase of TIMP-1/-2 expression, and decrease the ANE-induced transcription and release of CTGF, TGFß1, IL-6, and TNFα. PNS (25 µg/ml) also significantly inhibited the ANE-induced activation of AKT and ERK/JNK/p38 MAPK pathways in oral mucosal fibroblasts and the ANE-induced activation of TGFß/smad pathway in HaCaT cells. CONCLUSION: Panax notoginseng saponins possess excellent anti-OSF activity, and its mechanism may be related to its ability to inhibit the ANE-induced activation of PI3K/AKT, ERK/JNK/p38 MAPK, and TGFß/smad pathways.

RNA N6-methyladenosine methylation in influenza A virus infection.

Influenza A virus (IAV) is a negative-sense single-stranded RNA virus that causes acute lung injury and acute respiratory distress syndrome, posing a serious threat to both animal and human health. N6-methyladenosine (m6A), a prevalent and abundant post-transcriptional methylation of RNA in eukaryotes, plays a crucial regulatory role in IAV infection by altering viral RNA and cellular transcripts to affect viral infection and the host immune response. This review focuses on the molecular mechanisms underlying m6A modification and its regulatory function in the context of IAV infection and the host immune response. This will provide a better understanding of virus-host interactions and offer insights into potential anti-IAV strategies.

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.

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.

Transforming growth factor-ß1 protects mechanically injured cortical murine neurons by reducing trauma-induced autophagy and apoptosis.

Transforming growth factor ß1 (TGF-ß1) has a neuroprotective function in traumatic brain injury (TBI) through its anti-inflammatory and immunomodulatory properties. However, the precise mechanisms underlying the neuroprotective actions of TGF-ß1 on the cortex require further investigation. In this study, we were aimed to investigate the regulatory function of TGF-ß1 on neuronal autophagy and apoptosis using an in vitro primary cortical neuron trauma-injury model. LDH activity was assayed to measure cell viability, and intracellular [Ca2+] was measured using Fluo-4-AM in an in vitro primary cortical neuron trauma-injury model. RNA-sequencing (RNAseq), immunofluorescent staining, transmission electron microscopy (TEM), western blot and CTSD activity detection were employed. We observed significant enrichment of DEGs related to autophagy, apoptosis, and the lysosome pathway in trauma-injured cortical neurons. TEM confirmed the presence of autophagosomes as well as autophagolysosomes. Western blot revealed upregulation of autophagy-related protein light chain 3 (LC3-II/LC3-I), sequestosome 1 (SQSTM1/p62), along with apoptosis-related protein cleaved-caspase 3 in trauma-injured primary cortical neurons. Furthermore, trauma-injured cortical neurons showed an upregulation of lysosomal marker protein (LAMP1) and lysosomal enzyme mature cathepsin D (mCTSD), but a decrease in the activity of CTSD enzyme. These results indicated that apoptosis was up-regulated in trauma- injured cortical neurons at 24 h, accompanied by lysosomal dysfunction and impaired autophagic flux. Notably, TGF-ß1 significantly reversed these changes. Our results suggested that TGF-ß1 exerted neuroprotective effects on trauma- injured cortical neurons by reducing lysosomal dysfunction, decreasing the accumulation of autophagosomes and autophagolysosomes, and enhancing autophagic flux.