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OBJECTIVE: To systematically synthesize research evidence on barriers and facilitators to pressure injury prevention in hospital settings. METHODS: A systematic literature review of quantitative, qualitative, and mixed methods research was undertaken using PubMed, MEDLINE, Embase, CINAHL, and Cochrane Library. Studies that reported barriers or/and facilitators to pressure injury prevention in the acute care settings and published in English from 2008 to 2022 were included. Studies were excluded if they were conducted in residential care facilities and nursing homes, or other long-term community care settings. Two authors independently screened articles against the inclusion and exclusion criteria. Quality appraisal was conducted by two authors by using the Mixed Methods Appraisal Tool. Reported results were mapped to the Theoretical Domains Framework to identify the barriers and facilitators to pressure injury prevention. RESULTS: A total of 78 studies were included. There were 65 quantitative studies, 11 qualitative studies, and two mix-methods studies. The most salient Theoretical Domains Framework domains identified in this review were "Knowledge", "Skills", "Environmental Context and Resources", "Optimism", "Social/Professional Role and Identity", and "Social influences". CONCLUSION: The barriers and facilitators to pressure injury prevention in hospital settings identified in this systematic review were diverse, and included issues at both individual and organizational level. Healthcare organizations can address the barriers and facilitators from the influential Theoretical Domains Framework domains. Future research is required to investigate the effectiveness of behaviour change interventions that specifically target these barriers and facilitators to pressure injury prevention.
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Lesões por Esmagamento , Úlcera por Pressão , Humanos , Úlcera por Pressão/prevenção & controle , Casas de Saúde , Hospitais , Pesquisa QualitativaRESUMO
As an effective targeted therapy for advanced hepatocellular carcinoma (HCC), sorafenib resistance has been frequently reported in recent years, with the activation of autophagy by cancer cells under drug stress being one of the crucial reasons. Sorafenib treatment could enhance autophagy in HCC cells and autophagy is also considered as an important mechanisms of drug resistance. Therefore, the inhibition of autophagy is a potential way to improve the sensitivity and eliminate drug resistance to restore their efficacy. To determine whether autophagy is involved in sorafenib resistance and investigate its role in the regulation of HepG2 cells' (an HCC cell line) chemosensitivity to sorafenib, we simultaneously treated HepG2 with sorafenib and 3-Methyladenine (3-MA) (a common autophagy inhibitor). First, by performing cell counting kit 8 cell viability assay, Hoechst 33342 apoptosis staining, and Annexin V-fluorescein isothiocyanate/propidium iodide apoptosis kit detection, we found that both sorafenib and 3-MA effectively inhibitted the proliferative activity of HepG2 cells and induced their apoptosis to a certain extent. This effect was significantly enhanced after these two drugs were combined, which was also confirmed by the increased expression of apoptosis-related proteins. Subsequently, by using AAV-GFP-LC3 transfection methods and transmission electron microscopy, we found that both the number and activity of autophagosomes in HepG2 cells in sorafenib and 3-MA group were significantly reduced, suggesting that autophagy activity was inhibited, and this result was consistent with the expression results of autophagy-related proteins. Therefore, we conclude that 3-MA may attenuate the acquired drug resistance of sorafenib by counteracting its induction of autophagy activity, thus enhancing its sensitivity to advanced HCC therapy.
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Adenina/análogos & derivados , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Sorafenibe/farmacologia , Adenina/administração & dosagem , Adenina/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Carcinoma Hepatocelular/patologia , Sinergismo Farmacológico , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologia , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Sorafenibe/administração & dosagemRESUMO
Background: Family partial lipodystrophy (FPLD) is a rare autosomal dominant disease characterized by disorders of variable body fat loss associated with metabolic complications. FPLD6 has only been reported in a limited number of cases. Here, we reported a Chinese FPLD6 patient with compound heterozygous mutations in the lipase E, hormone-sensitive type (LIPE) gene. Case presentation: A 20-year-old female patient presented with hypertriglyceridemia, diabetes mellitus, hepatomegaly, and hepatic steatosis. Subcutaneous fat was significantly diminished in her face, abdomen, and limbs. The patient was assessed by detailed clinical and biochemical examinations. A liver biopsy showed severe lipodystrophy. In addition, there were retinal changes, peripheral nerve damage, and renal tubular injury. Sequencing was performed on extracted DNA. Genetic analysis revealed that the patient had compound heterozygous mutations in the LIPE gene: c.2497_250ldel (p.Glu833LysfsTer22) and c.2705del (p.Ser902ThrfsTer27) heterozygous mutations. Verification revealed that this mutation was inherited from her father and mother, respectively, and that they formed newly discovered compound heterozygous mutations occurring in the LIPE gene, causing FPLD6. Conclusion: We reported the first case of FPLD6 in China. Gene analysis demonstrated compound heterozygous mutations in LIPE in this patient. Our case emphasizes the importance of genetic testing in young patients with severe metabolic syndromes.
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Background: Lemierre's syndrome is a rare and serious complication of pharyngitis with an estimated annual incidence of 1 in 100,000 people worldwide. It is characterized by septic thrombophlebitis of the internal jugular vein with metastatic infection, usually after oropharyngeal infection. Rare cases of Lemierre's syndrome have been reported to be caused by odontogenic infection. Case report: A 33-year-old male visited our hospital with symptoms of fever and sore throat for 16 days. The other symptoms included pain in his left neck and shoulder. In addition, metabolic syndrome was diagnosed based on waist circumference, diabetes, and hyperlipidemia. Fusobacterium necrophorum bacteria was detected using the metagenomic next-generation sequencing (mNGS) technique. The enhanced computerized tomography (CT) scan showed thrombosis of the left proximal jugular vein and brachiocephalic vein. Based on these observations, Lemierre's syndrome was diagnosed. The etiology was that the fillings in the root canal tooth were lost with no blood or pain about 2 weeks before the onset. The patient recovered after treatment with antibiotics and blood purification. Conclusion: Lemierre's syndrome should be evaluated for patients with fever, sore throat, and neck pain. If the loss of fillings from root canal therapy occurs, especially for those with metabolic syndrome, we should be aware of the possibility of this disease. Furthermore, the mNGS test can be used as a crucial supplementary diagnostic tool for patients with undetermined fever.
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Vaccines represent a significant milestone in the history of human medical science and serve as the primary means for controlling infectious diseases. In recent years, the geographical distribution of Japanese encephalitis viruses (JEV) of various genotypes has become increasingly complex, which provides a rationale for the development of safer and more effective vaccines. The advent of subunit and nucleic acid vaccines, especially propelled by advancements in genetic engineering since the 1980s, has accelerated the application of novel adjuvants. These novel vaccine adjuvants have diversified into toll-like receptor (TLR) agonists, complex adjuvants, nanoparticles and so on. However, the efficacy of adjuvant combinations can vary depending on the host system, disease model, or vaccine formulation, sometimes resulting in competitive or counteractive effects. In our previous study, we constructed a pJME-LC3 chimeric DNA vaccine aimed at inducing an immune response through autophagy induction. Building on this, we investigated the impact of the TLR7/8 agonist imiquimod (IMQ) and the TLR9 agonist CpG ODN 1826 as adjuvants on the immunogenicity of the Japanese encephalitis chimeric DNA vaccine. Our findings indicate that the combination of the pJME-LC3 vaccine with IMQ and CpG ODN 1826 adjuvants enhanced the innate immune response, promoting the maturation and activation of antigen-presenting cells in the early immune response. Furthermore, it played a regulatory and optimizing role in subsequent antigen-specific immune responses, resulting in effective cellular and humoral immunity and providing prolonged immune protection. The synergistic effect of IMQ and CpG ODN 1826 as adjuvants offers a novel approach for the development of Japanese encephalitis nucleic acid vaccines.
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Adjuvantes Imunológicos , Vírus da Encefalite Japonesa (Espécie) , Encefalite Japonesa , Imiquimode , Vacinas contra Encefalite Japonesa , Oligodesoxirribonucleotídeos , Receptor 7 Toll-Like , Receptor Toll-Like 9 , Vacinas de DNA , Encefalite Japonesa/prevenção & controle , Encefalite Japonesa/imunologia , Oligodesoxirribonucleotídeos/farmacologia , Oligodesoxirribonucleotídeos/administração & dosagem , Animais , Adjuvantes Imunológicos/farmacologia , Vírus da Encefalite Japonesa (Espécie)/imunologia , Vacinas de DNA/imunologia , Receptor Toll-Like 9/agonistas , Camundongos , Vacinas contra Encefalite Japonesa/imunologia , Receptor 7 Toll-Like/agonistas , Feminino , Receptor 8 Toll-Like/agonistas , Humanos , Adjuvantes de Vacinas/farmacologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
We investigated the cellular immune responses elicited by a plasmid DNA vaccine encoding prM-E protein from the Japanese encephalitis (JE) virus (JEV) with or without various forms of intercellular adhesion molecule (ICAM)-1 gene to maximize the immune responses evoked by the JE DNA vaccine. We observed that co-immunization with the construct containing murine ICAM-1 gene (pICAM-1) resulted in a significant increase in the percentage of CD4(+)T cells, high level of JEV-specific cytotoxic T lymphocyte response, and high production of T helper 1 (Th1)-type cytokines in splenic T cells. Furthermore, the co-expression of ICAM-1 and DNA immunogens was found to be more effective in generating T cell-mediated immune responses than those induced by immunization with pJME in combination with pICAM-1. Our results suggested that ICAM-1 enhanced T cell receptor signaling and activated Th1 immune responses in the JEV model system by increasing the induction of CD4(+)Th1 cell subset and activating dendritic cells.
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Células Dendríticas/imunologia , Molécula 1 de Adesão Intercelular/administração & dosagem , Molécula 1 de Adesão Intercelular/imunologia , Vacinas contra Encefalite Japonesa/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Anticorpos Antivirais/imunologia , Células CHO , Linhagem Celular , Cricetinae , Vírus da Encefalite Japonesa (Espécie)/genética , Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/imunologia , Encefalite Japonesa/prevenção & controle , Feminino , Imunidade Celular/imunologia , Molécula 1 de Adesão Intercelular/genética , Ativação Linfocitária/imunologia , Teste de Cultura Mista de Linfócitos , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos/genética , Plasmídeos/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T Citotóxicos/imunologia , Células Th1/imunologia , Vacinas de DNA/imunologia , Proteínas Virais/administração & dosagem , Proteínas Virais/imunologiaRESUMO
Non-SMC condensin I complex subunit D2 (NCAPD2) is overexpressed in some malignant tumors. However, there are few studies on the function of NCAPD2 in pan-cancer. We used the Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), Human Protein Atlas (HPA), and UALCAN to analyze NCAPD2 expression and promoter methylation levels in 33 tumors and normal samples. We performed immunohistochemistry (IHC) on liver cancer and corresponding normal tissues to examine NCAPD2 protein expression in LIHC. Kaplan-Meier survival and univariate regression analyses were performed to explore the pan-cancer clinical significance of NCAPD2. Moreover, correlative analysis between NCAPD2 expression and clinical characteristics, immune cell infiltration, immune checkpoints, immune regulators, tumor mutation burden (TMB), microsatellite instability (MSI), ribonucleic acid (RNA) methylation regulators, and drug sensitivity was conducted using data from TCGA. We also investigated the effects of NCAPD2 expression on immunotherapy efficacy and prognosis. Gene set enrichment analysis (GSEA) was conducted using NCAPD2. Bioinformatic analysis showed that NCAPD2 was overexpressed in most tumors and correlated with the clinical characteristics of some cancers. IHC results demonstrated that NCAPD2 protein expression was higher in LIHC than in normal liver. NCAPD2 expression was linked with T stage, clinical stage, and histologic grade in LIHC. Overexpression of NCAPD2 resulted in poor overall survival, and disease-specific survival in adrenocortical carcinoma, kidney renal papillary cell carcinoma, brain lower grade glioma, liver hepatocellular carcinoma, lung adenocarcinoma, mesothelioma, pancreatic adenocarcinoma, sarcoma, skin cutaneous melanoma, and uterine corpus endometrial carcinoma. NCAPD2 was considered an independent biomarker by Cox regression in LIHC. The time ROC curve demonstrated that the survival rate of 1-, 3-, and 5-year OS and DSS in LIHC was above 0.6. The expression of NCAPD2 was significantly correlated with immune cell infiltration, immune checkpoints, TMB, MSI, and RNA methylation regulators in several tumors. NCAPD2 had a high predictive value for immunotherapy efficiency in certain tumors. In our study, drugs sensitive to NCAPD2 protein were screened by sensitivity analysis. GSEA analysis showed that NCAPD2 mainly participated in the G2M checkpoint, mitotic spindle, and KRAS-signaling. NCAPD2 may act as a prognostic molecular marker in most cancers.
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Adenocarcinoma , Neoplasias do Córtex Suprarrenal , Carcinoma Hepatocelular , Carcinoma de Células Renais , Neoplasias Renais , Neoplasias Pulmonares , Melanoma , Neoplasias Pancreáticas , Neoplasias Cutâneas , Humanos , Prognóstico , Proteínas de Ligação a Poli-ADP-Ribose , Proteínas Cromossômicas não Histona , Melanoma Maligno CutâneoRESUMO
Glucocorticoids are used as a first-line treatment for severe alcoholic hepatitis, and albumin reduces both the number of hospitalizations and mortality in patients with decompensated cirrhosis. However, for acute-on-chronic liver failure (ACLF), there is no definitive evidence that glucocorticoid therapy is beneficial. In this case report, we describe a male patient who developed into ACLF based on alcoholic cirrhosis, whose symptoms and clinical indicators continued to deteriorate after initial symptomatic treatment. The patient's condition gradually improved after low-dose glucocorticoid therapy, and long-term albumin supplementation resulted in a satisfactory outcome.
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Insuficiência Hepática Crônica Agudizada , Insuficiência Hepática Crônica Agudizada/diagnóstico , Insuficiência Hepática Crônica Agudizada/tratamento farmacológico , Insuficiência Hepática Crônica Agudizada/etiologia , Albuminas , Glucocorticoides/uso terapêutico , Humanos , Cirrose Hepática/complicações , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática Alcoólica/complicações , Cirrose Hepática Alcoólica/tratamento farmacológico , MasculinoRESUMO
Toxic epidermal necrolysis (TEN) and Stevens-Johnson syndrome (SJS) are acute inflammatory skin adverse reactions characterized by epidermal exfoliation and multi-site mucositis and are considered medical emergencies. The risk factors for SJS/TEN include immune disorders, malignancy, and genetic susceptibility. In most cases, medication is considered to be the leading cause of TEN. In addition, several studies suggest that infections, such as the herpes simplex virus, human immunodeficiency virus (HIV), Mycoplasma pneumoniae, streptococcus, and meningococcus infections, can trigger the occurrence of SJS/TEN. In this rare case, we share our experience managing TEN in a hepatitis A virus infection with an acute-on-chronic liver failure patient. A 38-year-old man was infected with hepatitis A virus on the basis of liver cirrhosis and progressed to acute-on-chronic liver failure. As the infection progressed, the target-like skin lesions accompanied by mucosal involvement worsened. The condition of the patient progressively worsened with a severe generalized rash, bullae, and epidermal detachment accompanied by severe erosive mucosal lesions. His skin detachment area gradually involved 30% of the body surface area (BSA), and the disease progressed to TEN. The intravenous infusion of corticosteroids alleviated the patient's hypersensitivity, and the patient obtained lasting remission without severe adverse reactions and complications.
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For decades, it has been widely believed that the blood-brain barrier (BBB) provides an immune privileged environment in the central nervous system (CNS) by blocking peripheral immune cells and humoral immune factors. This view has been revised in recent years, with increasing evidence revealing that the peripheral immune system plays a critical role in regulating CNS homeostasis and disease. Neurodegenerative diseases are characterized by progressive dysfunction and the loss of neurons in the CNS. An increasing number of studies have focused on the role of the connection between the peripheral immune system and the CNS in neurodegenerative diseases. On the one hand, peripherally released cytokines can cross the BBB, cause direct neurotoxicity and contribute to the activation of microglia and astrocytes. On the other hand, peripheral immune cells can also infiltrate the brain and participate in the progression of neuroinflammatory and neurodegenerative diseases. Neurodegenerative diseases have a high morbidity and disability rate, yet there are no effective therapies to stop or reverse their progression. In recent years, neuroinflammation has received much attention as a therapeutic target for many neurodegenerative diseases. In this review, we highlight the emerging role of the peripheral and central immune systems in neurodegenerative diseases, as well as their interactions. A better understanding of the emerging role of the immune systems may improve therapeutic strategies for neurodegenerative diseases.
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Background: The outbreak of coronavirus disease (COVID-19) poses a great threat to global public health. At present, the number of newly confirmed COVID-19 cases and deaths is increasing worldwide. The strategy of comprehensive and scientific detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) through quantitative real-time polymerase chain reaction (qRT-PCR) for special populations and environments provides great support for the prevention and control of this pandemic in China. Our study focused on determining the factors associated with the length of time from symptom onset to the first positive nucleic acid test of throat swabs in COVID-19 patients, evaluating the effect of early positive nucleic acid detection on the disease severity and its significance in prognosis, and predicting the factors associated with the time from positive SARS-CoV-2 RNA test to negative conversion (negative conversion of SARS-CoV-2 virus) in COVID-19 patients. Methods: This study included 116 hospitalized patients with COVID-19 from January 30, 2020 to March 4, 2020 in Wuhan, China. Throat swab samples were collected for qRT-PCR testing of SARS-CoV-2 RNA, and all patients included in this study were positive for this test. Results: The multivariate Cox proportional hazards model showed that disease severity (HR = 0.572; 95% CI 0.348-0.942; p = 0.028) was a protective factor for the time from symptom onset to positive nucleic acid detection. Meanwhile, the time from symptom onset to positive nucleic acid detection (HR = 1.010; 95% CI 1.005-1.020; p = 0.0282) was an independent risk factor for the delay in negative conversion time of SARS-CoV-2 virus. However, the severity of the disease (HR=1.120; 95% CI 0.771-1.640; p = 0.544) had no correlation with the negative conversion time of SARS-CoV-2 virus. Conclusions: Patients with more severe disease had a shorter time from symptom onset to a positive nucleic acid test. Prolonged time from symptom onset to positive nucleic acid test was an independent risk factor for the delay in negative conversion time of SARS-CoV-2 virus, and the severity of the disease had no correlation with negative conversion time of SARS-CoV-2 virus.
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A polysaccharide from Plumula Nelumbinis (PNP), was isolated and purified. PNP had a molecular weight of 450â¯kDa and consisted five monosaccharides, including rhamnose, galacturonic acid, xylose, galactose, and arabinose. The methylation and nuclear magnetic resonance (NMR) analysis revealed that the main glycosidic linkage types of PNP were â5)-α-L-Araf-(1â, â3)-ß-D-Galp-(1â, ß-D-Xylp-(â1, â3,4)-ß-D-Rhap-(1â, â4)-ß-D-GalpA-(1â. In the range of 25-1200⯵g/mL, PNP had no cytotoxicity to RAW264.7 cells. PNP could protect RAW264.7 cell from oxidative damage by reducing the production of ROS and MDA and the secretion of LDH, enhancing the activity of SOD, CAT, and GSH-Px, and increasing the content of GSH. Anti-inflammatory activity experiments showed that PNP inhibited the expression of NO, TNF-α, INF-γ, IL-1ß, and IL-6. PNP could inhibit the activation of MAPK/NF-κB cell pathways. PNP could be used as a potential natural antioxidant and anti-inflammatory substance in functional foods and pharmaceuticals.
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Antioxidantes , Polissacarídeos , Animais , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Camundongos , Monossacarídeos/análise , Polissacarídeos/química , Células RAW 264.7RESUMO
Ferroptosis is a newly discovered type of cell death mediated by iron-dependent lipid peroxide. The disturbance of iron metabolism, imbalance of the amino acid antioxidant system, and lipid peroxide accumulation are considered distinct fingerprints of ferroptosis. The dysregulation of ferroptosis has been intensively studied in recent years due to its participation in various diseases, including cancer, kidney injury, and neurodegenerative diseases. Notably, increasing evidence indicates that ferroptosis plays different roles in a wide spectrum of liver diseases. On the one hand, inhibiting ferroptosis may counteract the pathophysiological progression of several liver diseases, such as alcoholic liver injury, nonalcoholic steatosis hepatitis and fibrosis. On the other hand, inducing ferroptosis may restrict the emergence of secondary resistance to current medicines, such as sorafenib, for hepatocellular carcinoma (HCC) therapy. Here, we summarize the biological characteristics and regulatory signalling pathways of ferroptosis involved in liver disease. The current available medical agents targeting ferroptosis, including inducers or inhibitors applied in liver diseases, are also reviewed. This work aims to provide new insight into the emerging role of pathogenesis and therapeutic approaches for liver diseases.
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OBJECTIVES: To retrospectively evaluate the clinical and immunological characteristics of patients who died of COVID-19 and to identify patients at high risk of death at an early stage and reduce their mortality. RESULTS: Total white blood cell count, neutrophil count and C-reactive protein were significantly higher in patients who died of COVID-19 than those who recovered from it (p < 0.05), but the total lymphocyte count, CD4 + T cells, CD8 + T cells, B cells and natural killer cells were significantly lower when compared in the same groups. Multiple logistic regression analysis showed that increased D-dimer, decreased CD4 + T cells and increased neutrophils were risk factors for mortality. Further multiple COX regression demonstrated that neutrophil ≥ 5.27 × 109/L increased the risk of death in COVID-19 patients after adjustment for age and gender. However, CD4 + T cells ≥ 260/µL appeared to reduce the risk of death. CONCLUSION: SARS-CoV-2 infection led to a significant decrease of lymphocytes, and decreased CD4 + T cell count was a risk factor for COVID-19 patients to develop severe disease and death. METHODS: This study included 190 hospitalized COVID-19 patients from January 30, 2020 to March 4, 2020 in Wuhan, China, of whom 85 died and 105 recovered. Two researchers independently collected the clinical and laboratory data from electronic medical records.
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COVID-19/sangue , COVID-19/imunologia , Linfócitos/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/imunologia , Proteína C-Reativa/análise , Proteína C-Reativa/imunologia , Linfócitos T CD4-Positivos/imunologia , COVID-19/diagnóstico , COVID-19/mortalidade , Feminino , Produtos de Degradação da Fibrina e do Fibrinogênio/análise , Produtos de Degradação da Fibrina e do Fibrinogênio/imunologia , Humanos , Células Matadoras Naturais/imunologia , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Neutrófilos/imunologia , Prognóstico , Estudos Retrospectivos , Fatores de Risco , SARS-CoV-2/isolamento & purificaçãoRESUMO
Striking difference in density between the oxide and the steel results in difficulty in preparing oxide dispersion strengthened steel with large size parts or materials. In this research, Al2O3 and TiO2 particles were initially milled with the 20 steel, and then the mixture was heated to a molten state to form a master alloy, which was used as a raw material for further preparation of the object steel. It was found that homogeneous distribution of the oxide particles was obtained in the mass production of the steel. Moreover, the obtained 45 carbon structural steel presents fine microstructures, together with improved mechanical properties, especially the impact ductility. This should be attributable to the transformation from the introduced micro-size oxide particles to the nano ones, which act as heterogeneous nucleants that play an important role in grain refinement and dispersion strengthening for the steel, during the remelting of the master alloy.
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OBJECTIVE: To investigate the immune responses elicited by pJME with or without various forms of granulocyte-macrophage colony-stimulating factor (GM-CSF) gene. METHODS: The changes of the T lymphocyte subsets and the levels of Th cell intracellular cytokines IFN-gamma and IL-4 were evaluated by flow cytometric analysis. The cytotoxic T lymphocyte kill activity was assessed by lactate dehydrogenase activity release test. An 80% plaque reduction neutralization test was performed to titrate the neutralization antibody before and after viral challenge. RESULTS: We demonstrated that simultaneous administration of pJME plus plasmid-ecoded GM-CSF (pGM-CSF) activated Th1 immune responses similar to those found by injecting pGM-CSF i.m. into mice 3 days before pJME vaccination, and enhancement of Th2 immunity predominated when the pGM-CSF was injected 3 days after pJME vaccination. Furthermore, the immunization with DNA vaccine encoding precursor membrane envelope/GM-CSF fusion protein was more effective in generating immune responses than that induced by immunization with pJME alone or in combination with pGM-CSF. CONCLUSIONS: These observations support the potential of GM-CSF DNA adjuvant for the Th1/Th2 balance and the enhancement of immune responses by showing that the timing of the administration of pGM-CSF and the application of different forms of GM-CSF gene influence the outcome of the resultant immune responses.
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Vírus da Encefalite Japonesa (Espécie)/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Vacinas de DNA/imunologia , Proteínas Virais/imunologia , Animais , Testes Imunológicos de Citotoxicidade , Vírus da Encefalite Japonesa (Espécie)/genética , Feminino , Injeções Intramusculares , Interferon gama/biossíntese , Interleucina-4/biossíntese , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Neutralização , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Linfócitos T/imunologia , Vacinas de DNA/administração & dosagem , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Proteínas Virais/genéticaRESUMO
For decades, an on-going concerted effort has been made to develop a universal DNA vaccine to combat the looming threat of a potential outbreak of the emerging Japanese encephalitis virus (JEV) infection. However, effective strategies are urgently required to counter poor immunogenicity and insufficient long-term protection. Recent reports have confirmed the critical role of autophagy in antigen presentation, long-term immune memory and immune responses against JEV. In this study, JEV prM and E protein with strong immunogenicity were fused with microtubule-associated protein 1 light chain 3 (LC3) encoding gene to construct an autophagy-mediated pJME-LC3 DNA vaccine. Researches indicated significant increase of autophagosomes or LC3 ⠡ expression in pJME-LC3 transfected cells. Furthermore, prME-LC3 fused protein was observed co-localized with GFP-LC3 to autophagosomes, which means it was successfully targeted to autophagosomes. After immunizing with pJME-LC3, mice were detected highest proportion of CD3+CD8+ T lymphocytes, CD8+ effector memory T cells (TEMs) and JEV specific cytotoxic T lymphocyte (CTL) activity to eliminate JEV. pJME-LC3 also enhanced IgG2a antibody in serum and cytokines IFN-γ, IL-12 produced by splenocytes, thus skew toward Th1 type immune response by activating the JAK2/STAT1 signaling pathway and upregulating expression of transcription factor T-bet. Notably, mice immunized with pJME-LC3 showed highest survival rate and long-lasting neutralizing antibody when challenged with virulent JEV, which were consistent with augment in percentage of CD4+ central memory T cells (TCMs). In brief, our studies suggested that autophagy can be used as a optimization strategy to enhance JEV specific immune response and long-term immune memory. Our attempt will contribute towards future efforts to develop an efficacious JEV vaccine.
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Autofagia/imunologia , Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/imunologia , Encefalite Japonesa/prevenção & controle , Vacinas contra Encefalite Japonesa/imunologia , Vacinas de DNA/imunologia , Animais , Anticorpos Antivirais/imunologia , Antígenos Virais/genética , Antígenos Virais/imunologia , Células CHO , Linhagem Celular , Cricetulus , Citocinas/metabolismo , Modelos Animais de Doenças , Encefalite Japonesa/metabolismo , Feminino , Imunização , Imunogenicidade da Vacina , Imunomodulação , Vacinas contra Encefalite Japonesa/administração & dosagem , Camundongos , Proteínas Recombinantes de Fusão , Vacinas de DNA/administração & dosagemRESUMO
Plasmid-encoded granulocyte-macrophage colony-stimulating factor (GMCSF) is an adjuvant for genetic vaccines; however, how GM-CSF enhances immunogenicity remains to be elucidated. In the present study, it was demonstrated that injection of a plasmid encoding the premembrane (prM) and envelope (E) protein of Japanese encephalitis virus and mouse GM-CSF (pJME/GM-CSF) into mouse muscle recruited large and multifocal conglomerates of macrophages and granulocytes, predominantly neutrophils. During the peak of the infiltration, an appreciable number of immature dendritic cells (DCs) appeared, although no T and B-cells was detected. pJME/GM-CSF increased the number of splenic DCs and the expression of major histocompatibility complex class II (MHCII) on splenic DC, and enhanced the antigenic capture, processing and presentation functions of splenic DCs, and the cell-mediated immunity induced by the vaccine. These findings suggested that the immune-enhancing effect by pJME/GM-CSF was associated with infiltrate size and the appearance of integrin αx (CD11c)+cells. Chitosan-pJME/GM-CSF nanoparticles, prepared by coacervation via intramuscular injection, outperformed standard pJME/GM-CSF administrations in DC recruitment, antigen processing and presentation, and vaccine enhancement. This revealed that muscular injection of chitosanpJME/GM-CSF nanoparticles may enhance the immunoadjuvant properties of GM-CSF.
Assuntos
Encefalite Japonesa/prevenção & controle , Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Plasmídeos/administração & dosagem , Vacinas de DNA/imunologia , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/química , Animais , Antígeno CD11c/genética , Antígeno CD11c/imunologia , Movimento Celular/efeitos dos fármacos , Quitosana/química , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/virologia , Vírus da Encefalite Japonesa (Espécie)/química , Vírus da Encefalite Japonesa (Espécie)/efeitos dos fármacos , Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/imunologia , Encefalite Japonesa/virologia , Feminino , Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Injeções Intramusculares , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/virologia , Camundongos , Camundongos Endogâmicos BALB C , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/imunologia , Nanopartículas/química , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/virologia , Plasmídeos/química , Plasmídeos/imunologia , Baço/efeitos dos fármacos , Baço/imunologia , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Proteínas do Envelope Viral/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genéticaRESUMO
This study aimed to investigate the immune adjuvant effect and mechanism induced by chitosan nanoparticles carrying pJME/GM-CSF. In this study, plasmid DNA (pJME/GM-CSF) was encapsulated in chitosan to prepare chitosan-pJME/GM-CSF nanoparticles using a complex coacervation process. Immunohistochemistry was used to detect the type of infiltrating cells at the site of intramuscular injection. The phenotype and functional changes of splenic DCs were measured by flow cytometry after different immunogens were injected intramuscularly. The killing activity of CTLs was assessed using the lactate dehydrogenase (LDH) release assay. The preparation of chitosan-pJME/GM-CSF nanoparticles matched the expected theoretical results. Our results also found that, after pJME/GM-CSF injection, the incoming cells were a mixture of macrophages, neutrophils, and immature DCs. Meanwhile, pJME/GM-CSF increased the expression of MHC class II molecules on splenic DCs, and enhanced their Ag capture and presentation functions. Cell-mediated immunity was induced by the vaccine. Furthermore, chitosan-pJME/GM-CSF nanoparticles outperformed the administration of standard pJME/GM-CSF in terms of DC recruitment, antigen processing and presentation, and vaccine enhancement. These findings reveal that chitosan could be used as delivery vector for DNA vaccine intramuscular immunizations, and enhance pJME/GM-CSF-induced cellular immune responses.
Assuntos
Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/imunologia , Encefalite Japonesa/prevenção & controle , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Imunidade Celular , Vacinas contra Encefalite Japonesa/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Quitosana/administração & dosagem , Quitosana/imunologia , Células Dendríticas/imunologia , Células Dendríticas/virologia , Vírus da Encefalite Japonesa (Espécie)/genética , Encefalite Japonesa/virologia , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Vacinas contra Encefalite Japonesa/administração & dosagem , Vacinas contra Encefalite Japonesa/genética , Camundongos , Camundongos Endogâmicos BALB C , Nanopartículas/administração & dosagem , Baço/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/virologia , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Vacinas de DNA/imunologiaRESUMO
Hepatitis B virus X protein (HBx) is a multifunctional protein, and it activates multiple signal transduction pathways in multiple types of cells and regulates the process of cell apoptosis. In the present study, we mainly investigated the correlation between HBx and renal tubular epithelial cell apoptosis in hepatitis B virus-associated glomerulonephritis (HBVGN) and the possible signaling mechanism. Cell apoptosis in nephridial tissues of patients with HBVGN were determined by the TUNEL method. HBx, p-STAT3 and STAT3 levels in nephridial tissues were determined by immunohistochemical assay, and a correlation analysis between HBx expression levels and apoptosis index in nephridial tissues was conducted. The activation of the JAK2/STAT3 signaling pathway in HK-2 cells and the expression of the apoptosis-related proteins Bax and Bcl-2 were determined by western blot analysis following transfection with the HBx eukaryotic expression vector. Cellular proliferation activity was determined by the CCK8 method, and cell apoptosis was determined with HO33342 staining using transmission electron microscopy and Annexin V/PI double staining flow cytometry. The results revealed that the apoptosis index in nephridial tissues of patients with HBVGN was significantly higher when compared to that of the control group, and p-STAT3 expression levels in HBVGN nephridial tissues were significantly increased. In the control group, no HBx expression was observed in the nephridial tissues, whereas HBx expression was found in the nephridial tissues of 86% of the patients with HBVGN. The HBx expression levels had a linear correlation with the apoptosis index in the nephridial tissues. After target gene HBx infection, expression levels of both p-JAK2 and p-STAT3 in human proximal HK-2 cells were significantly increased, and the Bax/Bcl-2 ratio was also significantly increased. At the same time, cellular proliferation of HK-2 cells was significantly inhibited, and the rate of apoptosis was increased. After incubation with AG490, the JAK2/STAT3 signaling pathway was partially blocked, which caused a decrease in the Bax/Bcl-2 ratio and reduced cell apoptosis caused by HBx. In conclusion, HBx upregulates the Bax/Bcl-2 ratio by activating the JAK2/STAT3 signaling pathway to cause renal tubular epithelial cell apoptosis, and it is possibly involved in the pathogenic mechanism of nephridial tissue damage caused by HBV.