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The eukaryotic translation initiation factor eIF4E can regulate cellular translation via phosphorylation on serine 209. In a recent study, by two rounds of TMT relative quantitative proteomics, we found that phosphorylated eIF4E (p-eIF4E) favors the translation of selected mRNAs, and the encoded proteins are mainly involved in ECM-receptor, focal adhesion, and PI3K-Akt signaling. The current paper is focused on the relationship between p-eIF4E and the downstream host cell proteins, and their presumed effect on efficient entry of PEDV. We found that the depletion of membrane-residential factor TSPAN3, CD63, and ITGB2 significantly inhibited viral invasion of PEDV, and reduced the entry of pseudotyped particles PEDV-pp, SARS-CoV-pp, and SARS-CoV-2-pp. The specific antibodies of TSPAN3, CD63, and ITGB2 blocked the adsorption of PEDV into host cells. Moreover, we detected that eIF4E phosphorylation was increased at 1 h after PEDV infection, in accordance with the expression of TSPAN3, CD63, and ITGB2. Similar trends appeared in the intestines of piglets in the early stage of PEDV challenge. Compared with Vero cells, S209A-Vero cells in which eIF4E cannot be phosphorylated showed a decrease of invading PEDV virions. MNK kinase inhibitor blocked PEDV invasion, as well as reduced the accumulation of TSPAN3, CD63, and ITGB2. Further study showed that the ERK-MNK pathway was responsible for the regulation of PEDV-induced early phosphorylation of eIF4E. This paper demonstrates for the first time the connections among p-eIF4E stimulation and membrane-residential host factors. Our findings also enrich the understanding of the biological function of phosphorylated eIF4E during the viral life cycle.IMPORTANCEThe eukaryotic translation initiation factor eIF4E can regulate cellular translation via phosphorylation. In our previous study, several host factors susceptible to a high level of p-eIF4E were found to be conducive to viral infection by coronavirus PEDV. The current paper is focused on cell membrane-residential factors, which are involved in signal pathways that are sensitive to phosphorylated eIF4E. We found that the ERK-MNK pathway was activated, which resulted in the stimulation of phosphorylation of eIF4E in early PEDV infection. Phospho-eIF4E promoted the viral invasion of PEDV by upregulating the expression of host factors TSPAN3, CD63, and ITGB2 at the translation level rather than at the transcription level. Moreover, TSPAN3, CD63, or ITGB2 facilitates the efficient entry of coronavirus SARS-CoV, SARS-CoV-2, and HCoV-OC43. Our findings broaden our insights into the dynamic phosphorylation of eIF4E during the viral life cycle, and provide further evidence that phosphorylated eIF4E regulates selective translation of host mRNA.
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Membrana Celular , Fator de Iniciação 4E em Eucariotos , Vírus da Diarreia Epidêmica Suína , Biossíntese de Proteínas , Internalização do Vírus , Animais , Membrana Celular/química , Membrana Celular/genética , Membrana Celular/metabolismo , Membrana Celular/virologia , Chlorocebus aethiops , Fator de Iniciação 4E em Eucariotos/química , Fator de Iniciação 4E em Eucariotos/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Cadeias beta de Integrinas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Vírus da Diarreia Epidêmica Suína/fisiologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteômica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Suínos , Tetraspaninas/metabolismo , Células VeroRESUMO
Inflammatory bowel disease (IBD) is a chronic systemic inflammatory condition regarded as a major risk factor for colitis-associated cancer. However, the underlying mechanisms of IBD remain unclear. First, five GSE data sets available in GEO were used to perform 'batch correction' and Robust Rank Aggregation (RRA) to identify differentially expressed genes (DEGs). Candidate molecules were identified using CytoHubba, and their diagnostic effectiveness was predicted. The CIBERSORT algorithm evaluated the immune cell infiltration in the intestinal epithelial tissues of patients with IBD and controls. Immune cell infiltration in the IBD and control groups was determined using the least absolute shrinkage selection operator algorithm and Cox regression analysis. Finally, a total of 51 DEGs were screened, and nine hub genes were identified using CytoHubba and Cytoscape. GSE87466 and GSE193677 were used as extra data set to validate the expression of the nine hub genes. CD4-naïve T cells, gamma-delta T cells, M1 macrophages and resting dendritic cells (DCs) are the main immune cell infiltrates in patients with IBD. Signal transducer and activator of transcription 1, CCR5 and integrin subunit beta 2 (ITGB2) were significantly upregulated in the IBD mouse model, and suppression of ITGB2 expression alleviated IBD inflammation in mice. Additionally, the expression of ITGB2 was upregulated in IBD-associated colorectal cancer (CRC). The silence of ITGB2 suppressed cell proliferation and tumour growth in vitro and in vivo. ITGB2 resting DCs may provide a therapeutic strategy for IBD, and ITGB2 may be a potential diagnostic marker for IBD-associated CRC.
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Biologia Computacional , Doenças Inflamatórias Intestinais , Humanos , Animais , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/imunologia , Doenças Inflamatórias Intestinais/patologia , Biologia Computacional/métodos , Camundongos , Perfilação da Expressão Gênica , Modelos Animais de Doenças , Antígenos CD18/genética , Antígenos CD18/metabolismo , Mapas de Interação de Proteínas , Receptores CCR5/genética , Receptores CCR5/metabolismoRESUMO
BACKGROUND: Sevoflurane (Sevo) preconditioning and postconditioning play a protective role against injury induced by hepatic ischemia/reperfusion (I/R). At the same time, the involvement of macrophage infiltration in this process and the precise mechanisms are unclear. Here, we designed this research to elucidate the protective effects of Sevo against hepatic I/R injury and the molecules involved. METHODS: The alleviating effect of Sevo on the liver injury was analyzed by liver function analysis, hematoxylin and eosin staining, Masson trichrome staining, terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling, western blot analysis and an enzyme-linked immunosorbent assay. An in vitro cell model was developed using alpha mouse liver 12 (AML12) cells, and the cell model was treated with oxygen-glucose deprivation and reoxygenation and Sevo. Multiple bioinformatics databases were used to screen transcriptional regulators related to hepatic I/R injury and the targets of Krueppel-like factor 5 (KLF5). KLF5 expression was artificially upregulated alone or with integrin beta-2 (ITGB2) knockdown to substantiate their involvement in Sevo-mediated hepatoprotection. RESULTS: Sevo protected the liver against I/R injury by reducing cell apoptosis and inflammatory response. KLF5 was upregulated in liver tissues following I/R injury, whereas KLF5 overexpression aggravated macrophage infiltration and liver injury induced by I/R injury. KLF5 bound to the promoter of ITGB2 to enhance ITGB2 transcription. Knockdown of ITGB2 reversed the aggravation of injury caused by KLF5 overexpression in mice and AML12 cells. CONCLUSIONS: Sevo blocked KLF5-mediated transcriptional activation of ITGB2, thereby inhibiting macrophage infiltration in hepatic I/R injury.
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Cadeias beta de Integrinas , Fatores de Transcrição Kruppel-Like , Fígado , Macrófagos , Traumatismo por Reperfusão , Sevoflurano , Animais , Camundongos , Apoptose , Antígenos CD18/metabolismo , Antígenos CD18/genética , Linhagem Celular , Modelos Animais de Doenças , Regulação da Expressão Gênica , Fatores de Transcrição Kruppel-Like/efeitos dos fármacos , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Fígado/metabolismo , Fígado/patologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/genética , Sevoflurano/farmacologia , Ativação Transcricional , Cadeias beta de Integrinas/efeitos dos fármacos , Cadeias beta de Integrinas/genética , Cadeias beta de Integrinas/metabolismoRESUMO
In the realm of glioma treatment, our groundbreaking research has uncovered the pivotal role of Integrin Beta 2 (ITGB2) in non-apoptotic cell death and its profound implications for immunotherapy efficacy. Gliomas, known for their aggressive and infiltrative nature, demand innovative therapeutic strategies for improved patient outcomes. Our study bridges a critical gap by examining the interplay between non-apoptotic cell death and immunotherapy response in gliomas. Through comprehensive analysis of ten diverse glioma datasets, we developed a unique death enrichment score and identified ITGB2 as a significant risk marker. This study demonstrates that ITGB2 can predict immune activity, mutation characteristics, and drug response in glioma patients. We reveal that ITGB2 not only mediates glioma proliferation and migration but also crucially influences immunotherapy responses by modulating the interaction between gliomas and macrophages by single-cell sequencing analysis (iTalk and ICELLNET). Employing a variety of molecular and cellular methodologies, including in vitro models, our findings highlight ITGB2 as a potent marker in glioma biology, particularly impacting macrophage migration and polarization. We present compelling evidence of ITGB2's dual role in regulating tumor cell behavior and shaping the immune landscape, thereby influencing therapeutic outcomes. The study underlines the potential of ITGB2-targeted strategies in enhancing the efficacy of immunotherapy and opens new avenues for personalized treatment approaches in glioma management. In conclusion, this research marks a significant stride in understanding glioma pathology and therapy, positioning ITGB2 as a key biomarker and a promising target in the quest for effective glioma treatments.
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BACKGROUND: Myocardial infarction (MI) is one of the most serious cardiovascular diseases, characterized by a rapid and irreversible decline in myocardial function. Early detection of patients with MI and prolonging the optimal therapeutic window of acute myocardial infarction (AMI) are particularly important. This study aimed to identify the diagnostic biomarkers and novel therapeutic targets for acute myocardial infarction. METHOD: We generated the AMI mouse models by ligating the proximal left anterior descending coronary artery. Six time points-Sham, AMI 10-min, 1-h, 6-h, 24-h, and 72-h-were chosen to examine the molecular changes that occur during the AMI process. RNA-seq and DIA-MS were performed on the infarcted left ventricular tissues of AMI mice at each time point. Co-expression pattern genes were screened from myocardial infarction samples at different time points by time-series analysis. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were used to examine these genes. Using the Interactive Gene/Protein Retrieval Tool (STRING) database, the protein-protein interaction network (PPI) was constructed and the hub genes were identified. In order to evaluate the diagnostic value of hub genes, a receiver operating characteristic (ROC) curve was constructed. An independent data set, GSE163772, confirmed the diagnostic value of hub genes further. RESULT: We obtained the expression profiles at different time points after the occurrence of heart failure through high-throughput sequencing, and found 167 genes with similar expression patterns through time series analysis. The immune response and immune-related pathways had the greatest enrichment of these genes. Among them, Itgb2 Syk, Tlr4, Tlr2, Itgax, and Lcp2 may play key roles as hub genes. Combined with the results of proteomic analysis, it was found that the expression of Coro1a in both omics increased with time. The results of external validation showed that TLR2, ITGAX, and LCP2 had good predictive ability for AMI diagnosis. CONCLUSION: Itgb2, Syk, Tlr4, Tlr2, Itgax, Lcp2 and Coro1a are considered to be the seven key genes significantly associated with AMI. Our results may provide potential targets for the prevention of adverse ventricular remodeling and the treatment of AMI.
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Infarto do Miocárdio , Receptor 2 Toll-Like , Humanos , Animais , Camundongos , Receptor 2 Toll-Like/genética , Proteômica , Receptor 4 Toll-Like/genética , Transcriptoma , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Biomarcadores/metabolismoRESUMO
BACKGROUNDS: HLA-B*58:01 allele was strongly associated with allopurinol induced severe cutaneous adverse drug reaction (SCAR). However, HLA-B genotype is not sufficient to predict the occurrence of allopurinol-induced SCAR. OBJECTIVE: To discover DNA methylation markers for allopurinol-induced SCAR which may improve the prediction accuracy of genetic testing. STUDY DESIGN: The study was designed as a retrospective case-control clinical study in multicenter hospitals across Taiwan, Mainland China, Malaysia and Canada. 125 cases of allopurinol-induced SCAR patients and 139 cases of allopurinol tolerant controls were enrolled in this study during 2005 to 2021. RESULTS: The results of genome-wide DNA methylation assay of 62 patients revealed that ITGB2 showed strong discriminative ability of allopurinol-induced SCAR in both HLA-B*58:01 positive and negative patients with AUC value of 0.9364 (95% CI 0.8682-1.000). In validation study, significant hypermethylation of ITGB2 were further validated in allopurinol-induced SCAR patients compared to tolerant controls, especially in those without HLA-B*58:01(AUC value of 0.8814 (95% CI 0.7121-1.000)). Additionally, the methylation levels of 2 sites on ITGB2 were associated with SCAR phenotypes. Combination of HLA-B*58:01 genotyping and ITGB2 methylation status could improve the prediction accuracy of allopurinol-induced SCAR with the AUC value up to 0.9387 (95% CI 0.9089-0.9684), while the AUC value of HLA-B*58:01 genotyping alone was 0.8557 (95% CI 0.8030-0.9083). CONCLUSIONS: Our study uncovers differentially methylated genes between allopurinol-induced SCAR patients and tolerant controls with positive or negative HLA-B*58:01 allele and provides the novel epigenetic marker that improves the prediction accuracy of genetic testing for prevention of allopurinol-induced SCAR.
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Hipersensibilidade a Drogas , Síndrome de Stevens-Johnson , Humanos , Alopurinol/efeitos adversos , Estudos Retrospectivos , Metilação de DNA , Hipersensibilidade a Drogas/epidemiologia , Antígenos HLA-B/genética , Síndrome de Stevens-Johnson/tratamento farmacológico , Síndrome de Stevens-Johnson/genéticaRESUMO
Leukocyte adhesion deficiency (LAD) is an immunodeficiency caused by defects in the adhesion of leukocytes (especially neutrophils) to the blood vessel wall. As a result, patients with LAD suffer from severe bacterial infections and impaired wound healing, accompanied by neutrophilia. In LAD-I, characterized directly after birth by delayed separation of the umbilical cord, mutations are found in ITGB2, the gene that encodes the ß subunit (CD18) of the ß2 integrins. In the rare LAD-II disease, the fucosylation of selectin ligands is disturbed, caused by mutations in SLC35C1, the gene that encodes a GDP-fucose transporter of the Golgi system. LAD-II patients lack the H and Lewis Lea and Leb blood group antigens. Finally, in LAD-III, the conformational activation of the hematopoietically expressed ß integrins is disturbed, leading to leukocyte and platelet dysfunction. This last syndrome is caused by mutations in FERMT3, encoding the kindlin-3 protein in all blood cells, involved in the regulation of ß integrin conformation. This article contains an update of the mutations that we consider to be relevant for the various forms of LAD.
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Síndrome da Aderência Leucocítica Deficitária , Humanos , Adesão Celular/genética , Síndrome da Aderência Leucocítica Deficitária/genética , Antígenos CD18/genética , Antígenos CD18/metabolismo , Leucócitos , MutaçãoRESUMO
Electroporation is a widely used method for delivering CRISPR components into cells; however, it presents challenges when applied to difficult-to-transfect cells like adult buffalo fibroblasts. In this study, the ITGB2 gene (encoding the CD18 protein), plays vital for cellular adhesion and immune responses, was selected for editing experiments. To optimize electroporation conditions, we investigated parameters such as electric field strength, pulse duration, plasmid DNA amount, cuvette type, and cell type. The best transfection rates were obtained in a 4 mm gap cuvette with a single 20-millisecond pulse of 300 V using a 10 µg of all-in-one CRISPR plasmid for 106 cells in 100 µL of electroporation buffer. Increasing DNA quantity enhanced transfection rates but compromised cell viability. The 4 mm cuvette gap had high transfection rates than the 2 mm gap, and newborn cells exhibited higher transfection rates than adult cells. We achieved transfection rates of 10-12% with a cell viability of 25-30% for adult fibroblast cells. Subsequently, successfully edited the ITGB2 gene with a 30% editing efficiency, confirmed through various analysis methods, including T7E1 assay, TIDE and ICE analysis, and TA cloning. In conclusion, electroporation conditions reported here can edit buffalo gene(s) for various biotechnological research applications.
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Búfalos , Edição de Genes , Animais , Edição de Genes/métodos , Búfalos/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Eletroporação , Transfecção , Fibroblastos , DNA , Sistemas CRISPR-Cas/genéticaRESUMO
PURPOSE: Glioma is the most common primary tumor in the brain, accounting for 81% of intracranial malignancies. Nowadays, cancer immunotherapy has become a novel and revolutionary treatment for patients with advanced, highly aggressive tumors. However, to date, there are no effective biomarkers to reflect the response of glioma patients to immunotherapy. In this study, we aimed to assess the clinical predictive value of ITGB2 in patients with glioma. METHODS: The correlation between ITGB2 expression levels and glioma progression was explored and validated using data from CGGA, TCGA, GEO datasets, and patient samples from our hospital. Univariate and multivariate cox regression models were developed to determine the predictive role of ITGB2 on the prognosis of patients with glioma. The relationship between ITGB2 and immune activation was then analyzed. Finally, we predicted the immunotherapy response in both high and low ITGB2 expression subgroups. RESULTS: ITGB2 was significantly elevated in gliomas with higher malignancy and predicted poor prognosis. In multivariate analysis, the hazard ratio for ITGB2 expression (low versus high) was 0.71 with 95% CI (0.59-0.85) (P < 0.001). Furthermore, we found that ITGB2 stratified glioma patients into high and low ITGB2 expression subgroups, exhibiting different clinical outcomes and immune activation status. At last, we demonstrated that glioma patients with high ITGB2 expression levels had better immunotherapy response. CONCLUSIONS: This study demonstrated ITGB2 as a novel predictor for clinical prognosis and response to immunotherapy in gliomas. Assessing expression levels of ITGB2 is a promising method to discover patients that may benefit from immunotherapy.
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Biomarcadores Tumorais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Glioma/diagnóstico , Glioma/metabolismo , Integrina beta1/metabolismo , Neoplasias Encefálicas/etiologia , Neoplasias Encefálicas/terapia , Biologia Computacional/métodos , Bases de Dados Genéticas , Progressão da Doença , Perfilação da Expressão Gênica , Glioma/mortalidade , Glioma/terapia , Humanos , Integrina beta1/genética , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Gradação de Tumores , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Modelos de Riscos Proporcionais , Transcriptoma , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismoRESUMO
Leukocyte adhesion deficiency type I is a rare primary immunodeficiency disorder characterized by mutations in the ITGB2 gene encoding CD18. We present clinical and immunological features of 15 patients with leukocyte adhesion deficiency type 1 (LAD-1). Targeted next-generation sequencing was performed with either a primary immunodeficiency gene panel comprising 266 genes or a small LAD-panel consisting of five genes for genetic analysis. To measure the expression level of integrins on the leukocyte surface, flow cytometry analysis was performed. The median age of the patients at diagnosis was 3 (1-48) months. Eleven (73%) of the 15 patients had a LAD-1 diagnosis in their first 6 months and 14 (93%) patients had consanguineous parents. Delayed separation of the umbilical cord was present in 80% (n = 12) of the patients in our cohort, whereas omphalitis was observed in 53% (n = 8) of the patients. Leukocytosis with neutrophil predominance was observed in 73% (n = 11) patients. Nine distinct variants in the ITGB2 gene in 13 of the 15 patients with LAD-1 were characterized, two of which (c.305_306delAA and c.779_786dup) are novel homozygous mutations of ITGB2. Four unrelated patients from Syria had a novel c.305_306delAA mutation that might be a founder effect for patients of Syrian origin. Four (27%) patients underwent hematopoietic stem cell transplantation. Two patients died because of HSCT complications and the other two are alive and well. Early differential diagnosis of the patients is critical in the management of the disease and genetic evaluation provides a basis for family studies and genetic counseling.
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Antígenos CD18/genética , Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala , Síndrome da Aderência Leucocítica Deficitária , Mutação , Feminino , Humanos , Lactente , Recém-Nascido , Síndrome da Aderência Leucocítica Deficitária/diagnóstico , Síndrome da Aderência Leucocítica Deficitária/genética , Masculino , TurquiaRESUMO
BACKGROUND: Knowledge of stenosis in coronary arteries requires an understanding of the cellular and molecular processes that occur throughout the leukocyte rolling process. In this study, the roles of miR-125a-5p and miR-495-3p were investigated on the adhesion of endothelial cells (ECs) isolated from the human aorta. METHODS: Human primary endothelial cells were obtained from the aorta of people who had died of brain death. Whole blood was used to isolate the monocytes. The miR-125 and miR-495 were predicted and transfected into ECs using Poly Ethylene Imine (PEI). The expression levels of adhesion molecules and monocyte recruitment were identified by the RT-qPCR technique and Leukocyte-Endothelial Adhesion Assay kit, respectively. RESULTS: The ICAM-1, ICAM-2 and VCAM-1 expression levels decreased significantly in the miR-495/PEI-transfected ECs (P < 0.05) while in the miR-125/PEI-transfected ECs only the ICAM-2 and ITGB-2 expression levels decreased significantly (P < 0.05) as compared to the miR-synthetic/PEI-transfected ECs. Furthermore, the monocyte adhesion was decreased in the miR-125 and miR-mix/PEI-transfected ECs as compared to the miR-synthetic/PEI-transfected ECs (P = 0.01 and P = 0.04, respectively). CONCLUSION: According to the findings, the efficient relations between miR-125 and adhesion molecules may be responsible for the inhibition of monocyte rolling.
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Aorta/metabolismo , Moléculas de Adesão Celular/metabolismo , Adesão Celular , Células Endoteliais/metabolismo , MicroRNAs/metabolismo , Monócitos/metabolismo , Antígenos CD/genética , Antígenos CD/metabolismo , Aorta/citologia , Antígenos CD18/genética , Antígenos CD18/metabolismo , Moléculas de Adesão Celular/genética , Células Cultivadas , Regulação da Expressão Gênica , Humanos , Iminas/química , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Migração e Rolagem de Leucócitos , MicroRNAs/genética , Polietilenos/química , Transdução de Sinais , Transfecção , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
Long noncoding RNA (lncRNA) has been considered as potentially critical regulators in pancreatic ductal adenocarcinoma (PDAC). In this study, we prospectively investigate the effect and mechanism of lncRNA integrin subunit beta 2-anti-sense RNA 1 (ITGB2-AS1) on regulation of PDAC progression. The expression of ITGB2-AS1 and its target were analyzed by quantitative real-time polymerase chain reaction and in situ hybridization. 3-(4,5-Dimethylthiazol-z-yl)-2,5-diphenyltetrazolium bromide, flow cytometry, wound healing, and transwell assays were used to investigate the influence of ITGB2-AS1 on cell proliferation, cell cycle, migration, and invasion, respectively. The interaction between ITGB2-AS1 and its target was determined via luciferase activity assay and RNA immunoprecipitation. The subcutaneous xenotransplanted tumor model was established and employed to detect the tumorigenic function of ITGB2-AS1, which was evaluated by western blot analysis, immunohistochemistry, and hematoxylin and eosin staining. The results showed that ITGB2-AS1 was elevated in both PDAC tumor tissues and cell lines, predicting a poor prognosis in PDAC patients. Knocking down of ITGB2-AS1 suppressed PDAC cell proliferation, invasion, and migration but induced cell apoptosis in vitro. Moreover, ITGB2-AS1 could target and inhibit the expression of miR-4319 and miR-4319-targeted and -suppressed serine/threonine kinase RAF1. ITGB2-AS1 promoted PDAC progression via inhibition of miR-4319. Interference of ITGB2-AS1 could suppress in vivo tumorigenic ability of PDAC via downregulation of RAF1. In conclusion, ITGB2-AS1 promoted PDAC progression via sponging miR-4319 to upregulate RAF1, suggesting the potential therapeutic target ability of ITGB2-AS1 in PDAC.
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Yes-associated protein (YAP) is a transcriptional co-activator and a major effector of the Hippo pathway that promotes cell proliferation and stemness, while inhibiting apoptosis. YAP plays a central role in organ size control, and its deregulation strongly promotes cancer initiation and progression. However, the mechanisms by which YAP promotes cell invasion and metastasis are not fully understood. Here, we report that YAP induces leukocyte-specific integrin ß2 (ITGB2) expression in cancer cells, thereby promoting cell invasion through the endothelium in a manner mimicking leukocytes. Through independent biochemical purification and a functional screen, we further identified PR/SET domain 4 (PRDM4) as a transcription factor interacting with the WW domains of YAP to mediate ITGB2 expression and cell invasion. Consistently, ITGB2 and PRDM4 mRNA levels are significantly increased in metastatic prostate cancer. In addition, PRDM4 contributes to YAP-induced tumorigenesis possibly via mediating the expression of other YAP target genes. Our results demonstrate that YAP promotes cell invasion by inducing leukocyte-specific integrin expression, and identify PRDM4 as a novel transcription factor for YAP targets.
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Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Antígenos CD18/metabolismo , Proteínas de Ligação a DNA/metabolismo , Fosfoproteínas/fisiologia , Neoplasias da Próstata/patologia , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Humanos , Masculino , Camundongos , Invasividade Neoplásica , Fosfoproteínas/genética , Neoplasias da Próstata/metabolismo , Fatores de Transcrição/genética , Proteínas de Sinalização YAPRESUMO
The kynurenine pathway (KP) has a principal role in the metabolism of tryptophan. This pathway is also involved in the pathogenesis of cancer. We evaluated expression of two rate limiting enzymes from this pathway (IDO1 and TDO2) as well as three long non-coding RNAs (lncRNAs) that have been predicted to alter expression of IDO1 (ITGB2-AS1, HCP5 and MIR155HG) in 82 breast cancer tissues and their adjacent non-cancerous tissues (ANCTs). While IDO1 expression levels were not significantly different between malignant tissues and ANCTs (expression ratio = 0.56, P = .21), TDO2 was significantly down-regulated in malignant tissues compared with ANCTs (Expression ratio = 0.001, P < .001). Among lncRNAs, expression of HCP5 was significantly lower in malignant tissues compared with ANCTs (Expression ratio = 0.17, P < .001). However, expression of ITGB2-AS1 was higher in malignant tissues compared with ANCTs (Expression ratio = 3.38, P = .01). Expressions of genes were not associated with any of clinical or demographic data of patients. However, there were trends towards association between IDO1 expression and tumor size as well as estrogen receptor (ER) status (P values 0.09 and 0.08 respectively). Significant pairwise correlations were found between expression levels of genes especially in ANCTs. Notably, TDO2 expression levels were correlated with expression of all other genes in ANCTs but none of them in tumor tissues. Based on the area under curve (AUC) values, HCP5 and TDO2 had "fair" diagnostic power (AUC values of 0.73 and 0.72). Notably, combination of HCP5, ITGB2-AS1 and TDO2 genes increased the diagnostic power to the level of "good". The current investigation underscores the role of KP in breast cancer and potentiates some genes within this pathway as diagnostic markers in breast cancer.
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Neoplasias da Mama/genética , RNA Longo não Codificante/genética , Triptofano Oxigenase/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Neoplasias da Mama/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indóis/metabolismo , Cinurenina/metabolismo , Redes e Vias Metabólicas/genética , MicroRNAs/genética , Pessoa de Meia-Idade , Triptofano/metabolismoRESUMO
PURPOSE: We aimed to report the characteristics of leukocyte adhesion deficiency-I (LAD-I) and four novel mutations in the ITGB2 gene in a Chinese cohort. METHODS: Seven patients with LAD-I were reported in our study. Clinical manifestations and immunological phenotypes were reviewed. The expression of CD18 was detected by flow cytometry. Next-generation sequencing (NGS) and Sanger sequencing were performed to identify gene mutations. RESULTS: The mean onset age of all the patients was 1.3 months. Recurrent bacterial infections of the skin and lungs were the most common symptoms. Most patients (6/7) had delayed cord separation. The number of white blood cells (WBC) was increased significantly, except that two patients had a mild increase in the number of WBC during infection-free periods. The expression of CD18 was very low in all patients. Homozygous or compound heterozygous mutations in the ITGB2 gene were identified in each patient. Four mutations were novel, including c.1794dupC (p.N599Qfs*93), c.1788C>A (p.C596X), c.841-849del9, and c.741+1delG. Two patients had large deletions of the ITGB2 gene. Five patients were cured by hematopoietic stem cell transplantation (HSCT). CONCLUSIONS: This study reported the clinical and molecular characteristics of a Chinese patient cohort. It is helpful in understanding the current status of the disease in China.
Assuntos
Autoantígenos/genética , Síndrome da Aderência Leucocítica Deficitária/genética , Mutação/genética , Colágenos não Fibrilares/genética , Infecções Bacterianas , Antígenos CD18/metabolismo , China , Estudos de Coortes , Feminino , Transplante de Células-Tronco Hematopoéticas , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Masculino , Colágeno Tipo XVIIRESUMO
Systemic sclerosis (SSc), an autoimmune disease of connective tissue, is characterized by inflammation, fibrosis, and vessel endothelial damage. Products of Integrin subunit beta 2 (ITGB2) and selectin L (SELL) genes participate in several functional pathways of immune system. The aim of this investigation was to survey the transcript level of ITGB2 and SELL genes as well as methylation status of CpG sites in promoter region of differently expressed gene in PBMCs of SSc patients. PBMCs were isolated from whole blood of 50 SSc patients and 30 healthy controls. Total RNA and DNA contents of PBMCs were extracted. Gene expression was analyzed by real-time PCR using the SYBR Green PCR Master Mix. To investigate the methylation status of CpG sites, DNA samples were treated by bisulfite, amplified through nested PCR, and sequenced through Sanger difficult sequencing method. ITGB2 gene in PBMCs of SSc patients was overexpressed significantly in comparison to healthy controls. However, no altered SELL expression was observed. Three CpG sites of 12, 13 and 14 were significantly hypomethylated in patients group, despite overall methylation status of ITGB2 gene promoter revealed no significant difference between study groups. There was no statistically significant correlation between methylation status of ITGB2 promoter and the gene expression in patients. Regarding to lack of correlation of increased expression of ITGB2 with its promoter hypomethylation in SSc patients, our study suggests that upregulation of ITGB2 in PBMCs from SSc patients is probably due to another mechanism other than methylation alteration.
Assuntos
Antígenos CD18/genética , Metilação de DNA , Selectina L/genética , Regiões Promotoras Genéticas , Escleroderma Sistêmico/genética , Antígenos CD18/metabolismo , Estudos de Casos e Controles , Ilhas de CpG , Humanos , Selectina L/metabolismo , Leucócitos Mononucleares/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Escleroderma Sistêmico/diagnóstico , Escleroderma Sistêmico/metabolismo , Regulação para CimaRESUMO
In the previous study, we screened a novel lncRNA-ITGB2-AS1, which was down-regulated by bone morphogenetic protein 9 (BMP9) in breast cancer cell. Studying ITGB2-AS1 will lay the foundation for the exploring mechanism of the BMP9 inhibitory effect on breast cancer. The expression analysis related to ITGB2-AS1 in clinical samples was conducted on online websites. The overexpression plasmid or siRNA fragment was transfected into breast cancer cells to alter its gene expression. The MTT assay and flow cytometry were used to measure cell viability and cell cycle. Additionally, cell migration and invasion were detected by wound healing and transwell assay. The results of biological function experiments showed that ITGB2-AS1 could promote the migration and invasion of breast cancer. Furthermore, ITGB2-AS1 increased the mRNA and protein expression of ITGB2. Consistent with ITGB2-AS1, ITGB2 exerted the promotion effect on the migration and invasion of breast cancer and activated integrin-related FAK signaling. The OL plasmid expressing the truncation of ITGB2-AS1, which was complementary to ITGB2, was essential for activation of FAK signaling. In conclusion, LncRNA ITGB2-AS1 could promote the migration and invasion of breast cancer cells by up-regulating ITGB2.
Assuntos
Neoplasias da Mama/genética , Integrina beta3/genética , Invasividade Neoplásica/genética , RNA Longo não Codificante/genética , Neoplasias da Mama/patologia , Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica/patologia , RNA Mensageiro/genética , Ativação Transcricional/genética , TransfecçãoRESUMO
PURPOSE: Leukocyte adhesion deficiency type-I (LAD-I) is caused by mutations in the ITGB2 gene, encoding the ß2-subunit of ß2-integrin (CD18) which leads to markedly reduced expression of CD18 on leukocytes resulting into recurrent life threatening infections. Here we aim to identify the molecular defects underlying LAD-I in Indian patients and correlate with the clinical presentation. METHODS: Blood was collected from 30 patients and their parents for absolute neutrophil count, expression of CD18 and CD11 by flow cytometry and DNA extraction. PCR and DNA sequencing of the ITGB2 gene was done for mutation characterization. RESULTS: Phenotypically, 22 patients were LAD-I(0), 1 was LAD-I(-) and 7 were LAD-I(+) showing no expression and reduced expression of CD18 respectively. Nine novel mutations in 15 patients and 11 known mutations in 16 patients were detected. Prenatal diagnosis was performed for 5 families. CONCLUSION: In this study 30 patients were phenotypically and genotypically evaluated for a less known disease LAD-I. Unavailability of curative options to majority of the patients and high cost of supportive care emphasize the need to increase awareness about a suspicious case so that timely management can be given to the patient and prenatal diagnosis can be offered to their families.
Assuntos
Antígenos CD18/genética , Síndrome da Aderência Leucocítica Deficitária/genética , Mutação , Análise Mutacional de DNA , Feminino , Humanos , Índia , Lactente , Recém-Nascido , Síndrome da Aderência Leucocítica Deficitária/diagnóstico , Leucócitos/metabolismo , Leucócitos/patologia , Masculino , Diagnóstico Pré-Natal , População Branca/genéticaRESUMO
BACKGROUND: Blood DNA methylation was associated with coronary heart disease (CHD) risk in Caucasians. We investigated the association between DNA methylation in peripheral blood at the reported loci and CHD in the Chinese population. METHODS: The integrin subunit beta 2 (ITGB2) gene was identified in 196 CHD cases and 184 controls, and its methylation level was determined by mass spectrometry. Logistic regression was used to assess the association. RESULTS: Hypomethylation of ITGB2 was significantly associated with heart failure CHD and NYHA â &â ¡ CHD patients with minor to medium cardiac function impairment (ITGB2_CpG_11/cg08422803, OR per -10 % methylation = 1.15 and 1.16; p = 0.012 and 0.018 by Bonferroni correction, respectively). Hypomethylation of ITGB2_CpG_11/cg08422803 was a risk factor for CHD in people < 65 years and males (p < 0.05 after Bonferroni correction). The combination of ITGB2 methylation and conventional CHD risk factors could efficiently discriminate CHD, heart failure CHD, NYHA I&II CHD, and myocardial infarction CHD patients from controls (AUC = 0.78, 0.81, 0.80, and 0.81, respectively). CONCLUSION: Blood-based ITGB2 methylation has the potential as a biomarker for CHD. The combination of ITGB2 methylation and conventional CHD risk factors may improve the risk assessment and detection of CHD.
Assuntos
Doença das Coronárias , Insuficiência Cardíaca , Infarto do Miocárdio , Humanos , Masculino , Estudos de Casos e Controles , Doença das Coronárias/diagnóstico , Metilação de DNA , Insuficiência Cardíaca/genética , Infarto do Miocárdio/genética , Antígenos CD18RESUMO
PURPOSE: Uveal melanoma (UM) with BAP1 inactivating mutations has a high risk of metastasis, but the mechanism behind BAP1 deficiency driving UM metastasis is unknown. METHODS: We analyzed the single-cell RNA sequencing (scRNA-Seq) data comprised primary and metastatic UM with or without BAP1 mutations (MUTs) to reveal inter- and intra-tumor heterogeneity among different groups. Then, an immune-competent mouse liver metastatic model was used to explore the role of ITGB2-ICAM1 in BAP1-associated UM metastasis. RESULTS: Cluster 1 tumor cells expressed high levels of genes linked to tumor metastasis, such as GDF15, ATF3, and CDKN1A, all of which are associated with poor prognosis. The strength of communication between terminally exhausted CD8+ T cells and GDF15hiATF3hiCDKN1Ahi tumor cells was enhanced in BAP1-mutated UM, with CellChat analysis predicting strong ITGB2-ICAM1 signaling between them. High expression of either ITGB2 or ICAM1 was a worse prognostic indicator. Using an immune-competent mouse liver metastatic model, we indicated that inhibiting either ICAM1 or ITGB2 prevented liver metastasis in the BAP1-mutated group in vivo. The inhibitors primarily inhibited hypoxia- and ECM-related pathways indicated by changes in the expression of genes such as ADAM8, CAV2, ENO1, PGK1, LOXL2, ITGA5, and VCAN. etc. CONCLUSION: This study suggested that the ITGB2-ICAM1 axis may play a crucial role for BAP1-associated UM metastasis by preserving hypoxia- and ECM- related signatures, which provide a potential strategy for preventing UM metastasis in patients with BAP1 mutation.