Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 7 de 7
Filter
Add more filters










Database
Language
Publication year range
1.
J Biol Chem ; 292(29): 12077-12087, 2017 07 21.
Article in English | MEDLINE | ID: mdl-28576828

ABSTRACT

The NLRP3 inflammasome is a multiprotein complex that regulates the activation of caspase-1 leading to the maturation of the proinflammatory cytokines IL-1ß and IL-18 and promoting pyroptosis. Classically, the NLRP3 inflammasome in murine macrophages is activated by the recognition of pathogen-associated molecular patterns and by many structurally unrelated factors. Understanding the precise mechanism of NLRP3 activation by such a wide array of stimuli remains elusive, but several signaling events, including cytosolic efflux and influx of select ions, have been suggested. Accordingly, several studies have indicated a role of anion channels in NLRP3 inflammasome assembly, but their direct involvement has not been shown. Here, we report that the chloride intracellular channel proteins CLIC1 and CLIC4 participate in the regulation of the NLRP3 inflammasome. Confocal microscopy and cell fractionation experiments revealed that upon LPS stimulation of macrophages, CLIC1 and CLIC4 translocated into the nucleus and cellular membrane. In LPS/ATP-stimulated bone marrow-derived macrophages (BMDMs), CLIC1 or CLIC4 siRNA transfection impaired transcription of IL-1ß, ASC speck formation, and secretion of mature IL-1ß. Collectively, our results demonstrate that CLIC1 and CLIC4 participate both in the priming signal for IL-1ß and in NLRP3 activation.


Subject(s)
Chloride Channels/metabolism , Inflammasomes/drug effects , Interleukin-1beta/agonists , Macrophage Activation/drug effects , Macrophages/drug effects , Mitochondrial Proteins/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Active Transport, Cell Nucleus/drug effects , Animals , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Bone Marrow Cells/immunology , Bone Marrow Cells/metabolism , Cell Line , Cells, Cultured , Chloride Channels/antagonists & inhibitors , Chloride Channels/genetics , Inflammasomes/immunology , Inflammasomes/metabolism , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Lipopolysaccharides/toxicity , Macrophages/cytology , Macrophages/immunology , Macrophages/metabolism , Mice , Mice, 129 Strain , Mice, Inbred C57BL , Mice, Knockout , Mitochondrial Proteins/antagonists & inhibitors , Mitochondrial Proteins/genetics , Protein Transport/drug effects , Pyroptosis/drug effects , RAW 264.7 Cells , RNA Interference , Signal Transduction/drug effects
2.
Cell Metab ; 21(1): 65-80, 2015 Jan 06.
Article in English | MEDLINE | ID: mdl-25565206

ABSTRACT

Macrophages activated by the TLR4 agonist LPS undergo dramatic changes in their metabolic activity. We here show that LPS induces expression of the key metabolic regulator Pyruvate Kinase M2 (PKM2). Activation of PKM2 using two well-characterized small molecules, DASA-58 and TEPP-46, inhibited LPS-induced Hif-1α and IL-1ß, as well as the expression of a range of other Hif-1α-dependent genes. Activation of PKM2 attenuated an LPS-induced proinflammatory M1 macrophage phenotype while promoting traits typical of an M2 macrophage. We show that LPS-induced PKM2 enters into a complex with Hif-1α, which can directly bind to the IL-1ß promoter, an event that is inhibited by activation of PKM2. Both compounds inhibited LPS-induced glycolytic reprogramming and succinate production. Finally, activation of PKM2 by TEPP-46 in vivo inhibited LPS and Salmonella typhimurium-induced IL-1ß production, while boosting production of IL-10. PKM2 is therefore a critical determinant of macrophage activation by LPS, promoting the inflammatory response.


Subject(s)
Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Interleukin-1beta/metabolism , Macrophages/metabolism , Pyruvate Kinase/metabolism , Animals , Bone Marrow Cells/cytology , Cells, Cultured , Enzyme Activators/pharmacology , Gene Expression/drug effects , Glycolysis , Hypoxia-Inducible Factor 1, alpha Subunit/chemistry , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Interleukin-1beta/genetics , Lipopolysaccharides/toxicity , Macrophage Activation/drug effects , Macrophages/cytology , Mice , Mice, Inbred C57BL , Promoter Regions, Genetic , Protein Binding , Pyruvate Kinase/chemistry , Pyruvate Kinase/genetics , RNA, Messenger/metabolism , Salmonella typhimurium/physiology , Toll-Like Receptor 4/agonists , Toll-Like Receptor 4/metabolism
4.
Nucleic Acids Res ; 42(3): e17, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24357407

ABSTRACT

MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate gene expression at a post-transcriptional level. An miRNA may target many messenger RNA (mRNA) transcripts, and each transcript may be targeted by multiple miRNAs. Our understanding of miRNA regulation is evolving to consider modules of miRNAs that regulate groups of functionally related mRNAs. Here we expand the model of miRNA functional modules and use it to guide the integration of miRNA and mRNA expression and target prediction data. We present evidence of cooperativity between miRNA classes within this integrated miRNA-mRNA association matrix. We then apply bicluster analysis to uncover miRNA functional modules within this integrated data set and develop a novel application to visualize and query these results. We show that this wholly unsupervised approach can discover a network of miRNA-mRNA modules that are enriched for both biological processes and miRNA classes. We apply this method to investigate the interplay of miRNAs and mRNAs in integrated data sets derived from neuroblastoma and human immune cells. This study is the first to apply the technique of biclustering to model functional modules within an integrated miRNA-mRNA association matrix. Results provide evidence of an extensive modular miRNA functional network and enable characterization of miRNA function and dysregulation in disease.


Subject(s)
MicroRNAs/metabolism , Models, Genetic , RNA, Messenger/metabolism , Cluster Analysis , Computer Graphics , Gene Expression Profiling , Gene Expression Regulation , Humans , Immune System/metabolism , MicroRNAs/classification , Neuroblastoma/genetics , Neuroblastoma/metabolism , Software
5.
Pediatr Surg Int ; 29(2): 101-19, 2013 Feb.
Article in English | MEDLINE | ID: mdl-23274701

ABSTRACT

Neuroblastoma is a highly heterogeneous tumor accounting for 15 % of all pediatric cancer deaths. Clinical behavior ranges from the spontaneous regression of localized, asymptomatic tumors, as well as metastasized tumors in infants, to rapid progression and resistance to therapy. Genomic amplification of the MYCN oncogene has been used to predict outcome in neuroblastoma for over 30 years, however, recent methodological advances including miRNA and mRNA profiling, comparative genomic hybridization (array-CGH), and whole-genome sequencing have enabled the detailed analysis of the neuroblastoma genome, leading to the identification of new prognostic markers and better patient stratification. In this review, we will describe the main genetic factors responsible for these diverse clinical phenotypes in neuroblastoma, the chronology of their discovery, and the impact on patient prognosis.


Subject(s)
Epigenomics/methods , Neuroblastoma/genetics , Child , Chromosome Deletion , Chromosome Mapping/methods , Humans
6.
Nat Genet ; 44(11): 1199-206, 2012 Nov.
Article in English | MEDLINE | ID: mdl-23042116

ABSTRACT

LIN28B regulates developmental processes by modulating microRNAs (miRNAs) of the let-7 family. A role for LIN28B in cancer has been proposed but has not been established in vivo. Here, we report that LIN28B showed genomic aberrations and extensive overexpression in high-risk neuroblastoma compared to several other tumor entities and normal tissues. High LIN28B expression was an independent risk factor for adverse outcome in neuroblastoma. LIN28B signaled through repression of the let-7 miRNAs and consequently resulted in elevated MYCN protein expression in neuroblastoma cells. LIN28B-let-7-MYCN signaling blocked differentiation of normal neuroblasts and neuroblastoma cells. These findings were fully recapitulated in a mouse model in which LIN28B expression in the sympathetic adrenergic lineage induced development of neuroblastomas marked by low let-7 miRNA levels and high MYCN protein expression. Interference with this pathway might offer therapeutic perspectives.


Subject(s)
DNA-Binding Proteins/genetics , MicroRNAs , Neuroblastoma , Nuclear Proteins , Oncogene Proteins , Animals , Cell Differentiation , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Neoplastic , Gene Silencing , Humans , Mice , Mice, Transgenic , MicroRNAs/genetics , MicroRNAs/metabolism , N-Myc Proto-Oncogene Protein , Neuroblastoma/genetics , Neuroblastoma/metabolism , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Oncogene Proteins/genetics , Oncogene Proteins/metabolism , RNA-Binding Proteins , Signal Transduction
7.
PLoS One ; 7(5): e38129, 2012.
Article in English | MEDLINE | ID: mdl-22662276

ABSTRACT

BACKGROUND: Neuroblastoma is one of the most challenging malignancies of childhood, being associated with the highest death rate in paediatric oncology, underlining the need for novel therapeutic approaches. Typically, patients with high risk disease undergo an initial remission in response to treatment, followed by disease recurrence that has become refractory to further treatment. Here, we demonstrate the first silica nanoparticle-based targeted delivery of a tumor suppressive, pro-apoptotic microRNA, miR-34a, to neuroblastoma tumors in a murine orthotopic xenograft model. These tumors express high levels of the cell surface antigen disialoganglioside GD2 (GD(2)), providing a target for tumor-specific delivery. PRINCIPAL FINDINGS: Nanoparticles encapsulating miR-34a and conjugated to a GD(2) antibody facilitated tumor-specific delivery following systemic administration into tumor bearing mice, resulted in significantly decreased tumor growth, increased apoptosis and a reduction in vascularisation. We further demonstrate a novel, multi-step molecular mechanism by which miR-34a leads to increased levels of the tissue inhibitor metallopeptidase 2 precursor (TIMP2) protein, accounting for the highly reduced vascularisation noted in miR-34a-treated tumors. SIGNIFICANCE: These novel findings highlight the potential of anti-GD(2)-nanoparticle-mediated targeted delivery of miR-34a for both the treatment of GD(2)-expressing tumors, and as a basic discovery tool for elucidating biological effects of novel miRNAs on tumor growth.


Subject(s)
Gangliosides/immunology , MicroRNAs/administration & dosage , Nanoconjugates/administration & dosage , Neuroblastoma/drug therapy , Animals , Apoptosis/drug effects , Apoptosis/genetics , Cell Line, Tumor , Gangliosides/metabolism , Gene Expression , Gene Expression Profiling , Humans , Mice , Mice, SCID , MicroRNAs/chemistry , MicroRNAs/metabolism , N-Myc Proto-Oncogene Protein , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/genetics , Neuroblastoma/genetics , Neuroblastoma/metabolism , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Oncogene Proteins/genetics , Oncogene Proteins/metabolism , Xenograft Model Antitumor Assays
SELECTION OF CITATIONS
SEARCH DETAIL
...