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Interface residues at sites of protein-protein interaction (PPI) are the focus for affinity optimisation. However, protein hydrophobic cores (HCs) play critical roles and shape the protein surface. We hypothesise that manipulating protein HCs can enhance PPI interaction affinities. A cell stress molecule, major histocompatibility complex class I chain-related protein A (MICA), binds to the natural killer group 2D (NKG2D) homodimer to form three molecule interactions. MICA was used as a study subject to support our hypothesis. We redesigned MICA HCs by directed mutagenesis and isolated high-affinity variants through a newly designed partial-denature panning (PDP) method. A few mutations in MICA HCs increased the NKG2D-MICA interaction affinity by 325-5613-fold. Crystal structures of the NKG2D-MICA variant complexes indicated that mutagenesis of MICA HCs stabilised helical elements for decreasing intermolecular interactive free energy (ΔG) of the NKG2D-MICA heterotrimer. The repacking of MICA HC mutants maintained overall surface residues and the authentic binding specificity of MICA. In conclusion, this study provides a new method for MICA redesign and affinity optimisation through HC manipulation without mutating PPI interface residues. Our study introduces a novel approach to protein manipulation, potentially expanding the toolkit for protein affinity optimisation.
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Antígenos de Histocompatibilidad Clase I , Interacciones Hidrofóbicas e Hidrofílicas , Humanos , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase I/química , Antígenos de Histocompatibilidad Clase I/genética , Modelos Moleculares , Mutación , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Subfamilia K de Receptores Similares a Lectina de Células NK/química , Subfamilia K de Receptores Similares a Lectina de Células NK/genética , Unión ProteicaRESUMEN
Toxoplasmosis is a zoonosis caused by Toxoplasma gondii (T. gondii). The current treatment for toxoplasmosis remains constrained due to the absence of pharmaceutical interventions. Thus, the pursuit of more efficient targets is of great importance. Lipid metabolism in T. gondii, including fatty acid metabolism, phospholipid metabolism, and neutral lipid metabolism, assumes a crucial function in T. gondii because those pathways are largely involved in the formation of the membranous structure and cellular processes such as division, invasion, egress, replication, and apoptosis. The inhibitors of T. gondii's lipid metabolism can directly lead to the disturbance of various lipid component levels and serious destruction of membrane structure, ultimately leading to the death of the parasites. In this review, the specific lipid metabolism pathways, correlative enzymes, and inhibitors of lipid metabolism of T. gondii are elaborated in detail to generate novel ideas for the development of anti-T. gondii drugs that target the parasites' lipid metabolism.
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Toxoplasma , Toxoplasmosis , Animales , Metabolismo de los Lípidos , Apoptosis , Zoonosis , Toxoplasmosis/tratamiento farmacológicoRESUMEN
Introduction: The presence of soluble human programmed cell death-ligand 1 (shPD-L1) in the blood of patients with cancer has been reported to be negatively correlated with disease prognosis. However, little information exists about the mechanisms underlying high levels of shPD-L1 for promoting disease progression. Methods: In this study, we first analyzed the correlations between shPD-L1 and apoptosis of T cells in patients with cancer, then tested the effect of shPD-L1 on T-cell functions and the production of regulatory T cells. Results: We found that the apoptosis of human peripheral PD-1+CD4+ T cells was significantly elevated in patients with cancer compared with healthy donors and was positively correlated with circulating PD-L1 levels in patients with cancer. In vitro, monomeric shPD-L1 significantly inhibited the proliferation, cytokine secretion, and cancer cell-killing activity of peripheral blood mononuclear cells (PBMCs) activated by either agonist antibodies or HATac (high-affinity T cell activation core)-NYE (NY-ESO-1 antigen). It also promoted CD4+ T cells to express forkhead family transcription factor 3 (FoxP3) for the conversion of induced T regulatory cells, which was more significant than that mediated by soluble human PD-L1 fusion protein (shPD-L1-Fc). Discussion: These results confirm that soluble PD-L1 could be a candidate for inhibiting the functions of activated T cells, promoting peripheral tolerance to tumor cells, and implicating in system tumor immune escape in addition to the tumor microenvironment. This is an important mechanism explaining the negative correlation between peripheral blood PD-L1 levels and cancer prognosis. Therefore, understanding the roles of hPD-L1 in peripheral blood will be helpful for the development of precision immunotherapy programs in treating various tumors.
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Antígeno B7-H1 , Neoplasias , Humanos , Antígeno B7-H1/metabolismo , Leucocitos Mononucleares/metabolismo , Ligandos , Apoptosis , Microambiente TumoralRESUMEN
BACKGROUND: Decompressive craniectomy (DC) and intracranial pressure (ICP) monitoring are common approaches to reduce the death rate of Traumatic brain injury (TBI) patients, but the outcomes of these patients are unfavorable, particularly those who receive bilateral DC. The authors discuss their experience using ICP and other potential methods to improve the outcomes of TBI patients who receive bilateral DC. METHODS: Data from TBI patients receiving bilateral DC from Jan. 2008 to Jan. 2022 were collected via a retrospective chart review. Included patients who received unplanned contralateral DC after initial surgery were identified as unplanned secondary surgery (USS) patients. Patients' demographics and baseline medical status; pre-, intra-, and postoperative events; and follow-up visit outcome data were analyzed. RESULTS: A total of 151 TBI patients were included. Patients who underwent USS experienced more severe outcomes as assessed using the 3-month modified Rankin Scale score (P = 0.024). In bilateral DC TBI patients, USS were associated with worsen outcomes, moreover, ICP monitoring was able to lower their death rate and was associated with a lower USS incidence. In USS patients, ICP monitoring was not associated with improved outcomes but was able to lower their mortality rate (2/19, 10.5%, vs. 10/25, 40.0%; P = 0.042). CONCLUSION: The avoidance of USS may be associated with improved outcomes of TBI patients who underwent bilateral DC. ICP monitoring was a potential approach to lower USS rate in TBI patients, but its specific benefits were uncertain.
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Lesiones Traumáticas del Encéfalo , Lesiones Encefálicas , Craniectomía Descompresiva , Humanos , Craniectomía Descompresiva/métodos , Presión Intracraneal , Estudios Retrospectivos , Resultado del Tratamiento , Lesiones Traumáticas del Encéfalo/cirugíaRESUMEN
OBJECTIVE: To explore feasibility of protecting ulnar nerve by ultrasound in treating children with supracondylar fracture of humerus by closed reduction and intercross needle fixation. METHODS: From January 2018 to December 2019, 63 children with supracondylar fractures of humerus were divided into two groups(ultrasound group and X-ray group) depend on the different ways of guidance. Therer were 32 children in ultrasound group treated with closed reduction and Kirschner wire fixation guided by ultrasound, including 20 males and 12 females, aged from 3 to 11 years old with an average of (6.06±2.02) years old. There were 31 children in X-ray group treated with closed reduction and Kirschner wire fixation guided by X-ray, including 17 males and 14 females, aged from 2 to 10 years old with an average of (5.61±1.96) years old. Operation time, X-ray times, fracture healing time, ulnar nerve injury and postoperaqtive Flynn clinical function assessment at 1 year between two groups were recorded and compared. RESULTS: All patients were followed up. The follow-up time of ultrasound group ranged from 9 to 12 months with an average of (11.53±0.76) months, and X-ray group ranged from 10 to 13 months with an average of (11.51±0.72) months. There was no significant difference in operation time, follow-up time and fracture healing time between two groups(P>0.05). The number of intraoperative electrodialysis in ultrasound group was (3.06±1.24) times, and that in X-ray group was (21.65±5.58) times, which was significantly higher than that in ultrasound group(P<0.01). No iatrogenic ulnar nerve injury occurred in ultrasound group, and 2 cases of ulnar nerve injury occurred in X-ray group, the incidence of ulnar nerve injury in ultrasound group was lower than that in X-ray group, but the difference was not statistically significant(P>0.05). At 1 year after operation, Flynn clinical function assessment results in ultrasound group was excellent in 27 cases, 4 cases good and 1 case fair, in X-ray group 23 cases got excellent result, 6 cases good, 1 fair and 1 poor, there was no significant difference between two groups(P>0.05 ). CONCLUSION: Ultrasound guided and X-ray guided treatment of supracondylar fractures of humerus in children have similar effect of opertaion time and fracture healing, while ultrasound guidance could clearly detected the position of ulnar nerve, and avoid occurrence of iatrogenic ulnar nerve injury caused by ulnar puncture, which is a safe and effective treatment method.
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Fracturas del Húmero , Niño , Preescolar , Estudios de Factibilidad , Femenino , Fijación Interna de Fracturas/métodos , Humanos , Fracturas del Húmero/diagnóstico por imagen , Fracturas del Húmero/cirugía , Húmero/diagnóstico por imagen , Húmero/cirugía , Masculino , Nervio CubitalRESUMEN
The transportation industry has entered a new stage from quantity expanding to structure optimization, quality and efficiency improvement, and from respective governance to integrative development. This indicates that the traditional corridor mode allocation dominated by quantity equilibrium can no longer meet the requirements of the new stage. In this paper, we propose a multi-entity programming model based on the economic equilibrium between supply and demand. It not only ensures the economic equilibrium in the market, but also maximizes the social benefits of the whole system, thereby realizing the sustainable development of the transportation system. Also, the Globalsearch algorithm and intlinprog algorithm are designed to solve the problem. The actual case of Beijing-Shanghai corridor shows that the model and algorithms are effective, providing decision support for the optimal allocation of regional transport network resources.
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Algoritmos , Transportes , Beijing , China , Modelos TeóricosRESUMEN
BACKGROUND: In patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC), virus-specific cytotoxic T lymphocytes (CTLs) fail to eliminate HCC cells expressing HBV antigens. As the expression of viral antigen in HBV-associated HCC may decrease to allow tumor to escape immune attacks, we hypothesized that an HBV surface antigen (HBsAg)-specific affinity-improved-T-cell receptor (TCR) will enable T cells to target HCC more effectively than corresponding wild-type-TCR. We also postulated that TCR promiscuity can be exploited to efficiently capture HBV variants that can hinder CTL-based therapeutics. METHODS: We applied flexi-panning to isolate affinity-improved TCRs binding to a variant antigen, the human leukocyte antigen (HLA)-A*02:01-restricted nonapeptide HBs371-379-ILSPFLPLL, from libraries constructed with a TCR cloned using the decapeptide HBs370-379-SIVSPFIPLL. The potency and safety of the affinity-improved-TCR engineered T-cells (Ai-TCR-T) were verified with potentially cross-reactive human and HBV-variant peptides, tumor and normal cells, and xenograft mouse models. RESULTS: Ai-TCR-T cells retained cognate HBV antigen specificity and recognized a wide range of HBV genotypic variants with improved sensitivity and cytotoxicity. Cell infusions produced complete elimination of HCC without recurrence in the xenograft mouse models. Elevated accumulation of CD8+ Ai-TCR-T cells in tumors correlated with tumor shrinkage. CONCLUSION: The in vitro and in vivo studies demonstrated that HBsAg-specific Ai-TCR-T cells had safety profiles similar to those of their wild-type counterparts and significantly enhanced potency. This study presents an approach to develop new therapeutic strategies for HBV-related HCC.
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Carcinoma Hepatocelular/virología , Virus de la Hepatitis B/patogenicidad , Neoplasias Hepáticas/virología , Linfocitos T/metabolismo , Ingeniería de Tejidos/métodos , Animales , Carcinoma Hepatocelular/inmunología , Carcinoma Hepatocelular/patología , Humanos , Neoplasias Hepáticas/patología , Masculino , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Giant cell glioblastoma (gcGBM) is a rare histological variant of GBM, accounting for about 1% of all GBM. The prognosis is poor generally though gcGBM does slightly better than the other IDH-wild-type GBM. Because of the rarity of the cases, there has been no comprehensive molecular analysis of gcGBM. Previously, single-gene study identified genetic changes in TP53, PTEN and TERT promoter mutation in gcGBM. In this report, we performed whole-exome sequencing (WES) to identify somatically acquired mutations and copy number variations (CNVs) in 10 gcGBM genomes. We also examined TERT promoter mutation and MGMT methylation in our cohort. On top of the reported mutations, WES revealed ATRX, PIK3R1, RB1 and SETD2 as the recurrent mutations in gcGBM. Notably, one tumor harbored a mutation in MutS homolog 6 (MSH6) that is a key mismatch repair (MMR) gene. This tumor demonstrated hypermutation phenotype and showed an increased number of somatic mutations. TERT promoter mutation and MGMT methylation were observed in 20% and 40% of our samples, respectively. In conclusion, we described relevant mutation profiling for developing future targeted therapies in gcGBM.
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Glioblastoma/genética , Adolescente , Adulto , Anciano , Neoplasias Encefálicas/genética , Fosfatidilinositol 3-Quinasa Clase Ia/genética , Estudios de Cohortes , Variaciones en el Número de Copia de ADN/genética , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Femenino , Glioblastoma/metabolismo , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Masculino , Persona de Mediana Edad , Mutación , Pronóstico , Proteínas de Unión a Retinoblastoma/genética , Telomerasa/genética , Proteínas Supresoras de Tumor/genética , Ubiquitina-Proteína Ligasas/genética , Secuenciación del Exoma/métodos , Proteína Nuclear Ligada al Cromosoma X/genéticaRESUMEN
Cytokine-activated T cells (CATs) can be easily expanded and are widely applied to cancer immunotherapy. However, the good efficacy of CATs is rarely reported in clinical applications because CATs have no or very low antigen specificity. The low-efficacy problem can be resolved using T cell antigen receptor-engineered CAT (TCR-CAT). Herein, we demonstrate that NY-ESO-1157-165 HLA-A*02:01-specific high-affinity TCR (HAT)-transduced CATs can specifically kill cancer cells with good efficacy. With low micromolar range dissociation equilibrium constants, HAT-transduced CATs showed good specificity with no off-target killing. Furthermore, the high-affinity TCR-CATs delivered significantly better activation and cytotoxicity than the equivalent TCR-engineered T cells (TCR-Ts) in terms of interferon-γ and granzyme B production and in vitro cancer cell killing ability. TCR-CAT may be a very good alternative to the expensive TCR-T, which is considered an effective personalized cyto-immunotherapy.
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Citocinas/metabolismo , Inmunoterapia Adoptiva/métodos , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T/inmunología , Línea Celular Tumoral , Citotoxicidad Inmunológica , Ingeniería Genética , Antígeno HLA-A2/metabolismo , Humanos , Activación de Linfocitos , Receptores de Antígenos de Linfocitos T/genéticaRESUMEN
Tumor cells can escape immune surveillance through the programmed cell death protein 1 (PD-1) axis suppressing T cells. However, we recently demonstrated that high-affinity variants of soluble human programmed death-ligand 1 (shPD-L1) could diminish the suppression. We propose that in comparison to the wild-type shPD-L1, the further affinity enhancement will confer the molecule with opposite characteristics that augment T-cell activation and immunotherapeutic drug potential. In this study, a new shPD-L1 variant, L3C7c, has been generated to demonstrate â¼167 fold greater affinity than wild-type hPD-L1. The L3C7c-Fc fusion protein demonstrated completely opposite effects of conventional PD-1 axis by promoting redirected T-cell proliferation, activation and cytotoxicity in vitro, as being slightly better than that of anti-PD1-Ab (Pembrolizumab). Moreover, L3C7c-Fc was more effective than Pembrolizumab in enhancing redirected T cells' ability to suppress Mel624 melanoma growth in vivo. As a downsized L3C7c-Fc variant, L3C7v-Fc improved the anti-tumor efficacy in vivo when combined with dendritic cell vaccines. In conclusion, our studies demonstrate that high-affinity hPD-L1 variants could be developed as the next generation reagents for tumor immunotherapy based on the blockade of the PD-1 axis.
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Antígeno B7-H1/genética , Antígeno B7-H1/inmunología , Melanoma/inmunología , Melanoma/terapia , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales Humanizados/farmacología , Vacunas contra el Cáncer/inmunología , Línea Celular , Línea Celular Tumoral , Femenino , Humanos , Inmunoterapia/métodos , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Ratones SCID , Receptor de Muerte Celular Programada 1/genética , Receptor de Muerte Celular Programada 1/inmunologíaRESUMEN
OBJECTIVE: Hematologic inflammatory markers are simple, inexpensive prognostic markers for various conditions. The prognostic significance of the neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), monocyte-to-lymphocyte ratio (MLR), and red blood cell distribution width (RDW) has been shown in a variety of tumors. We evaluated the prognostic value of these markers in glioma. METHODS: We performed a retrospective medical record review of 219 patients with glioma from January 2012 to January 2017, evaluating the effect of NLR, PLR, MLR, and RDW on prognosis. Correlations among these hematologic inflammatory markers were also examined. RESULTS: The patients were divided into high and low groups using the cutoff points from the receiver operating characteristic curves. High NLR was associated with a higher tumor grade (P = 0.000). Kaplan-Meier survival analyses showed that the high NLR, PLR, and MLR groups experienced inferior median overall survival (OS) compared with the low NLR, PLR, and MLR groups (11 vs. 32 months; P = 0.000; 12 vs. 21 months; P = 0.001; and 12 vs. 22 months; P = 0.006, respectively). No significant difference was found in the median OS between the high and low RDW groups (15 vs. 23 months; P = 0.184). Multivariate analysis demonstrated that NLR was an independent predictor of OS (hazard ratio, 1.758; P = 0.008). CONCLUSIONS: A high preoperative NLR, PLR, and MLR was predictive of a poor prognosis for patients with glioma. NLR was an independent prognostic factor for OS in patients with glioma.
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Plaquetas/fisiología , Neoplasias Encefálicas/mortalidad , Glioma/mortalidad , Leucocitos Mononucleares/fisiología , Linfocitos/fisiología , Anciano , Biomarcadores de Tumor , Encefalitis/mortalidad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neutrófilos/fisiología , Recuento de Plaquetas , Cuidados Preoperatorios , Pronóstico , Estudios RetrospectivosRESUMEN
The inhibitory checkpoint molecule programmed death (PD)-1 plays a vital role in maintaining immune homeostasis upon binding to its ligands, PD-L1 and PD-L2. Several recent studies have demonstrated that soluble PD-1 (sPD-1) can block the interaction between membrane PD-1 and PD-L1 to enhance the antitumor capability of T cells. However, the affinity of natural sPD-1 binding to PD-L1 is too low to permit therapeutic applications. Here, a PD-1 variant with approximately 3000-fold and 70-fold affinity increase to bind PD-L1 and PD-L2, respectively, was generated through directed molecular evolution and phage display technology. Structural analysis showed that mutations at amino acid positions 124 and 132 of PD-1 played major roles in enhancing the affinity of PD-1 binding to its ligands. The high-affinity PD-1 mutant could compete with the binding of antibodies specific to PD-L1 or PD-L2 on cancer cells or dendritic cells, and it could enhance the proliferation and IFN-γ release of activated lymphocytes. These features potentially qualify the high-affinity PD-1 variant as a unique candidate for the development of a new class of PD-1 immune-checkpoint blockade therapeutics.
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Proteína 2 Ligando de Muerte Celular Programada 1/metabolismo , Receptor de Muerte Celular Programada 1/metabolismo , Aminoácidos/metabolismo , Proliferación Celular/fisiología , Técnicas de Visualización de Superficie Celular/métodos , Células Dendríticas/metabolismo , Humanos , Interferón gamma/metabolismo , Ligandos , Activación de Linfocitos/fisiología , Unión Proteica/fisiología , Linfocitos T/metabolismoRESUMEN
Recently, bi-functional molecules that can redirect immune effectors to tumour cells have emerged as potentially robust mediators of tumour regression in clinical trials. Two modalities in particular, bi-specific antibodies for T-cell redirection and activation (BiTe) and immune-mobilizing monoclonal T-cell receptors against cancer (ImmTAC), are being evaluated in efficacy studies as 'off-the-shelf' reagents. Optimal therapy will require an understanding and means to address regulatory mechanisms of limiting efficacy. In light of this, we evaluated the impact of induced regulatory T (iTreg) cells on the efficacy of tumour cell killing redirected by ImmTAC and demonstrated down-regulation of T-cell proliferation and expression of CD25, CD107a, Granzyme B and Perforin by ImmTAC-redirected T cells. Significant recovery of ImmTAC potency, however, could be achieved when combined with an anti-programmed cell death protein 1 monoclonal antibody. Furthermore, we found that among lung cancer patients failing to respond to ImmTAC therapy, there was a significantly higher fraction of Treg cells in the peripheral blood mononuclear cells of lung cancer patients than in healthy donors. These results provide in vitro evidence for an iTreg cell-mediated immunosuppression of ImmTAC-redirected T-cell responses. Whilst immune checkpoint blockade can reverse the Treg cell suppression, it forms a rational basis for a combination of the blockade with ImmTAC in clinical trials.
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Anticuerpos Monoclonales/farmacología , Citotoxicidad Inmunológica , Terapia de Inmunosupresión , Activación de Linfocitos/inmunología , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Citometría de Flujo , Humanos , Neoplasias/inmunología , Neoplasias/metabolismo , Neoplasias/patología , Neoplasias/terapia , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/metabolismoRESUMEN
The activated T cells can be suppressed by programed death-1 (PD-1) axis through low affinity interaction between PD-1 and PD-ligand 1 (PD-L1) in solution or on antigen presenting cells. In clinic, the concentration of soluble PD-L1 in peripheral blood negatively correlates with cancer prognosis. However, there is little information about the relation between the affinity of PD-1/PD-L1 interaction and the suppressive capacity of PD-1 axis. In this study, we analyzed inhibitory roles of high affinity soluble human PD-L1 (hPD-L1) variants, which were generated with directed molecular evolution. Resultant two clones L3C7-hPD-L1 and L3B3-hPD-L1 showed over 20 folds greater affinity than that of native hPD-L1. We found that L3B3-hPD-L1 and L3C7-hPD-L1 could compete with an anti-PD-1 antibody (EH12.1) for binding to hPD-1. More importantly, although native soluble hPD-L1 can induce suppressive effects on activated T cells, we found L3B3-hPD-L1 and L3C7-hPD-L1 attenuated the strength of PD-1 axis for suppressing the proliferation and interferon γ (IFN-γ) secretion of PBMC. In conclusion, our data provide direct evidence in which immune checkpoint receptor-ligand interactive strength can alter the the suppressive function, in particular, the suppressive capacity of PD-1 axis could be decreased with enhanced affinity of soluble PD-L1 and PD-1 interaction. Our study might provide a new direction for manipulating immune checkpoints.
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Dual-specificity tyrosine-regulated kinase 2 (DYRK2), a protein kinase that phosphorylates its substrates on serine/threonine, is expressed in numerous human tumors, but little is known about its role in the pathophysiology of glioma. In this study, we made an effort to explore the expression and function in human glioma. Western blot and immunohistochemistry analysis were performed to investigate the expression of DYRK2 protein in glioma tissues in 84 patients. Wound healing and transwell assay were carried out to determine the cell migration ability. We showed that the level of DYRK2 was significantly decreased in high-grade glioma tissues compared with low-grade tissues. In addition, the expression level of DYRK2 was positively correlated with glioma pathological grade and E-cadherin expression. Kaplane-Meier analysis revealed that low expression of DYRK2 was related to poor prognosis of glioma patients. Furthermore, wound healing and transwell assay revealed that DYRK2 could suppress cell migration and affect the expression levels of E-cadherin and vimentin through PI3K/AKT/GSK3ß signaling pathway. Taken together, our results implied that DYRK2 could serve as a promising prognostic biomarker as well as a potential therapeutical target of glioma.
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Biomarcadores de Tumor/biosíntesis , Neoplasias Encefálicas/metabolismo , Movimiento Celular/fisiología , Glioma/metabolismo , Proteínas Serina-Treonina Quinasas/biosíntesis , Proteínas Tirosina Quinasas/biosíntesis , Adulto , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Femenino , Glioma/patología , Humanos , Masculino , Persona de Mediana Edad , Clasificación del Tumor/métodos , Quinasas DyrKRESUMEN
p53-induced protein with a RING-H2 domain (Pirh2), also known as Rchy1, is an ubiquitin E3 ligase that regulates the turnover and functionality of several proteins involved in cell proliferation and differentiation, cell cycle checkpoints, and cell death. However, it remains unclear whether aberrant expression of Pirh2 is involved in the development of glioma, a major type of primary brain tumor in adults. Western blot and immunohistochemical analyses showed that Pirh2 was highly expressed in glioma specimens, compared with normal brain tissues. High Pirh2 expression was positively correlated with higher tumor grade, as well as Ki-67 expression. Kaplan-Meier analysis revealed that patients with high Pirh2 expression had worsened prognosis, compared with those with low Pirh2 expression. Moreover, to determine whether Pirh2 could regulate malignant behavior of glioma cells, we transfected glioma cells with interfering RNA targeting Pirh2 to specifically silence Pirh2 expression. Mechanistically, our results indicated that knockdown of Pirh2 induced the apoptosis of glioma cells. In addition, depletion of Pirh2 diminished the expression of PCNA and cyclin D1 and led to cell cycle arrest at G1 phase. Taken together, these findings for the first time suggest that Pirh2 might play an important role in the regulation of glioma proliferation and apoptosis and thus serve as a promising therapeutic target in the treatment of glioma.
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Neoplasias Encefálicas/metabolismo , Regulación Neoplásica de la Expresión Génica , Glioma/metabolismo , Ubiquitina-Proteína Ligasas/biosíntesis , Adulto , Biomarcadores de Tumor/biosíntesis , Biomarcadores de Tumor/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Femenino , Glioma/genética , Glioma/patología , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Tasa de Supervivencia/tendencias , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Vps4, vacuolar protein sorting 4, belongs to ATPases Associated with diverse cellular Activities (AAA) protein family which is made up of Vps4A and Vps4B. Previous studies demonstrated that Vps4A plays vital roles in diverse aspects such as virus budding, the efficient transport of H-Ras to the PM (plasma membrane) and the involvement in the MVB (multivesiculate bodies) pathway. Interestingly, Vps4A is also expressed in the brain. However, the distribution and function of Vps4A in ICH diseases remain unclear. In this study, we show that Vps4A may be involved in neuronal apoptosis during pathophysiological processes of intracerebral hemorrhage (ICH). Based on the results of Western blot and immunohistochemistry, we found a remarkable up-regulation of Vps4A expression surrounding the hematoma after ICH. Double labeled immunofluorescence showed that Vps4A was co-expressed with NeuN but rarely with astrocytes and microglia. Morever, we detected that neuronal apoptosis marker active caspase-3 had co-localizations with Vps4A. Additionaly, Vps4A knockdown in vitro specifically leads to decreasing neuronal apoptosis coupled with increased Akt phosphorylation. All datas suggested that Vps4A was involved in promoting neuronal apoptosis via inhibiting Akt phosphorylation after ICH.
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ATPasas Asociadas con Actividades Celulares Diversas/biosíntesis , Apoptosis/efectos de los fármacos , Hemorragia Cerebral/metabolismo , ATPasas de Translocación de Protón Vacuolares/biosíntesis , Animales , Antígenos Nucleares/metabolismo , Conducta Animal/efectos de los fármacos , Caspasa 3/metabolismo , Hemorragia Cerebral/patología , Hemorragia Cerebral/psicología , Femenino , Técnicas de Silenciamiento del Gen , Masculino , Proteínas del Tejido Nervioso/metabolismo , Proteína Oncogénica v-akt/metabolismo , Fosforilación , Embarazo , Cultivo Primario de Células , Ratas , Ratas Sprague-Dawley , Regulación hacia Arriba/efectos de los fármacosRESUMEN
The proliferation and differentiation of neural stem cells (NSCs) is important for neural regeneration after cerebral injury. Here, for the first time, we show that phosphorylated (p)-ser847-nNOS (NP847), rather than nNOS, may play a major role in NSC proliferation after traumatic brain injury (TBI). Western blot results demonstrated that the expression of NP847 and Sox2 in the hippocampus is up-regulated after TBI, and they both peak 3 days after brain injury. In addition, an immunofluorescence experiment indicated that NP847 and Sox2 partly co-localize in the nuclei of NSCs after TBI. Further immunoprecipitation experiments found that NP847 and Sox2 can directly interact with each other in NSCs. Moreover, in an OGD model of NSCs, NP847 expression is decreased, which is followed by the down-regulation of Sox2. Interestingly, in this study, we did not observe changes in the expression of nNOS in the OGD model. Further research data suggest that the NP847-Sox2 complex may play a major role in NSCs through the Shh/Gli signaling pathway in a CaMKII-dependent manner after brain injury.
RESUMEN
OBJECTIVE: Impulse oscillometry (IOS) is a new method for determination of breathing mechanics, which features convenient operation, good repeatability and wider range analysis. As there is no standardized normal value in China at present, this study will provide a normal value of lung function determination by impulse oscillometry for children in Chengdu area. METHOD: Totally 549 children were chosen at random from Chengdu area, with 292 boys and 257 girls who were 4 to 14 years old. The subjects were assigned into 10 age groups according to their chronological age with one year difference between every two adjacent groups. The respiratory total impedance (Zrs), viscosity resistance (Rrs) and elastic resistance (Xrs) at various oscillation frequency were measured by the Master Screen IOS which was manufactured by German Jaeger Company. The measured data were treated with the linear stepwise multiple regression, and established the prediction equation. At the same time, paired comparison was carried out with the measured data and equation obtained from this study, Lechtenboerger equation and prediction equation obtained from Guangzhou area. RESULT: The total impedance and airway resistance were negatively correlated with the children's height and age. Zrs (male) = -0.756 + 189.586/height, r = -0.782, P < 0.001; Zrs (female) = -0.497 + 152.468/height, r = -0.726, P < 0.001. Rrs became the same in trend; while Xrs were proportional to the height, e.g. the values increased as the height increased. The difference of the airway resistance (R(5)-R(20)) was negatively correlated with the children's height: R(5)-R(20) (male) = 0.601 - 0.0034 x height, r = -0.677, P < 0.001; R(5)-R(20) (female) = 0.549 - 0.0031 x height, r = -0.658, P < 0.001. Among the relationships with many impulse oscillometry parameters, height ranked at first place; age at second. The multiple regression equation of IOS primary index was established. Both the measured data and the correlation coefficient of the study obtained equation were greater than the coefficient correlation of the Lechtenboerger equation, but had no significant difference compared with that of prediction equation in Guangzhou area. CONCLUSION: The normal value in impulse oscillometry in children in Chengdu area is different from the predicted parameters in other countries. The equation obtained from this study seems to be more suitable for the children in its local area. It is recommended to apply the predicted value from the corresponding population in the determination of the lung function by impulse oscillometry.