RESUMO
Pilocytic astrocytoma (PA) is the most common brain tumor in children but is rare in adults, and hence poorly studied in this age group. We investigated 222 PA and report increased aneuploidy in older patients. Aneuploid genomes were identified in 45% of adult compared with 17% of pediatric PA. Gains were non-random, favoring chromosomes 5, 7, 6 and 11 in order of frequency, and preferentially affecting non-cerebellar PA and tumors with BRAF V600E mutations and not with KIAA1549-BRAF fusions or FGFR1 mutations. Aneuploid PA differentially expressed genes involved in CNS development, the unfolded protein response, and regulators of genomic stability and the cell cycle (MDM2, PLK2),whose correlated programs were overexpressed specifically in aneuploid PA compared to other glial tumors. Thus, convergence of pathways affecting the cell cycle and genomic stability may favor aneuploidy in PA, possibly representing an additional molecular driver in older patients with this brain tumor.
Assuntos
Aneuploidia , Astrocitoma/classificação , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/classificação , Adulto , Fatores Etários , Astrocitoma/genética , Astrocitoma/patologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Criança , Estudos de Coortes , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Mutação/genética , Estadiamento de Neoplasias , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-mdm2/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto JovemRESUMO
Pediatric glioblastomas (GBM) including diffuse intrinsic pontine gliomas (DIPG) are devastating brain tumors with no effective therapy. Here, we investigated clinical and biological impacts of histone H3.3 mutations. Forty-two DIPGs were tested for H3.3 mutations. Wild-type versus mutated (K27M-H3.3) subgroups were compared for HIST1H3B, IDH, ATRX and TP53 mutations, copy number alterations and clinical outcome. K27M-H3.3 occurred in 71 %, TP53 mutations in 77 % and ATRX mutations in 9 % of DIPGs. ATRX mutations were more frequent in older children (p < 0.0001). No G34V/R-H3.3, IDH1/2 or H3.1 mutations were identified. K27M-H3.3 DIPGs showed specific copy number changes, including all gains/amplifications of PDGFRA and MYC/PVT1 loci. Notably, all long-term survivors were H3.3 wild type and this group of patients had better overall survival. K27M-H3.3 mutation defines clinically and biologically distinct subgroups and is prevalent in DIPG, which will impact future therapeutic trial design. K27M- and G34V-H3.3 have location-based incidence (brainstem/cortex) and potentially play distinct roles in pediatric GBM pathogenesis. K27M-H3.3 is universally associated with short survival in DIPG, while patients wild-type for H3.3 show improved survival. Based on prognostic and therapeutic implications, our findings argue for H3.3-mutation testing at diagnosis, which should be rapidly integrated into the clinical decision-making algorithm, particularly in atypical DIPG.
Assuntos
Neoplasias do Tronco Encefálico/genética , Glioma/genética , Histonas/genética , Ponte/patologia , Adolescente , Neoplasias do Tronco Encefálico/mortalidade , Neoplasias do Tronco Encefálico/patologia , Criança , Pré-Escolar , Feminino , Perfilação da Expressão Gênica , Glioma/mortalidade , Glioma/patologia , Humanos , Lactente , Masculino , Mutação , Prognóstico , Taxa de SobrevidaRESUMO
Obesity has been shown to create stress in the endoplasmic reticulum (ER), and that initiates the activation of the unfolded protein response (UPR). This has been reported to cause insulin resistance in selective tissues through activation of the inositol-requiring enzyme 1α (IRE1α)-c-Jun NH(2)-terminal kinase (JNK) pathway, which results in the phosphorylation of the insulin receptor substrate-1 (IRS-1) at an inhibitory site and blocks insulin receptor signaling. In this study, we report that the Src homology domain-containing adaptor protein Nck1, previously shown to modulate the UPR, is of functional importance in obesity-induced ER stress signaling and inhibition of insulin actions. We have examined obese Nck1(-/-) and Nck1(+/+) mice for glucose tolerance, insulin sensitivity, and signaling as well as for ER stress markers and IRS-1 phosphorylation at Ser(307). Our findings show that obese Nck1-deficient mice display improved glucose disposal accompanied by enhanced insulin signaling in liver. This correlates with attenuated IRE1α and JNK activation and IRS-1 phosphorylation at Ser(307) compared with obese wild-type mice. Consistent with our in vivo data, we report that downregulation of Nck1 using siRNA in HepG2 cells results in decreased thapsigargin-induced IRE1α activation and signaling and IRS-1 phosphorylation at Ser(307), whereas it markedly enhances insulin signaling. Overall, in liver and in cultured cells, we show that depletion of Nck1 attenuates the UPR signal and its inhibitory action on insulin signaling. Taken all together, our findings implicate Nck1 in regulating the UPR, which secondary to obesity impairs glucose homeostasis and insulin actions.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Retículo Endoplasmático/fisiologia , Intolerância à Glucose/genética , Intolerância à Glucose/fisiopatologia , Insulina/fisiologia , Fígado/fisiologia , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Animais , Glicemia/metabolismo , Western Blotting , Glicogênio/biossíntese , Células HEK293 , Homeostase/fisiologia , Humanos , Resistência à Insulina/genética , Resistência à Insulina/fisiologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Camundongos Obesos , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Resposta a Proteínas não Dobradas/genética , Resposta a Proteínas não Dobradas/fisiologiaRESUMO
The scavenger receptor class B, type I (SR-BI) mediates cholesteryl esters (CE) selective uptake from low density lipoprotein (LDL) and high-density lipoprotein (HDL) particles. In a number of tissues expressing caveolin, SR-BI is localized in caveolae. We show using detergent-free sucrose gradients that SR-BI is found in membrane rafts devoid of caveolin-1 in the human hepatoma HepG2 cell. Perturbation of the structure of HepG2 cell membrane rafts with cholesterol oxidase or sphingomyelinase decreased LDL-CE association due to selective uptake by 60%, while HDL3-CE selective uptake was increased 2.3-fold by cholesterol oxidase but was not affected by sphingomyelinase. Sequestration of membrane cholesterol with filipin III decreased LDL-CE selective uptake by 25%, while it had no effect on HDL3-CE selective uptake. Extraction of cell membrane cholesterol with beta-cyclodextrin increased LDL- and HDL3-CE selective uptake by 1.6-fold and 3-fold, respectively. We found that CE-selective uptake from both HDL and LDL occurs by a pathway involving retro-endocytosis in HepG2 cells. An analysis of the effect of SR-BI level on the expression of critical lipid sensor and lipid binding proteins was conducted with stable transformants of HepG2 cell overexpressing SR-BI. We found that liver-type fatty acid binding protein expression level is higher in SR-BI-overexpressing cells and that caveolin-1 and sterol response element binding protein-2 levels are reduced. Thus, in this hepatic cell model, SR-BI is associated with membrane rafts devoid of caveolin and its expression affects intracellular lipid binding and lipid sensor proteins. SR-BI-dependent LDL- and HDL-CE selective uptake are affected differently by the integrity of membrane rafts, but both occur by a retroendocytic pathway in HepG2 cells.