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1.
Mucosal Immunol ; 14(1): 26-37, 2021 01.
Article in English | MEDLINE | ID: mdl-32457448

ABSTRACT

Type-2 immunity is characterised by interleukin (IL)-4, IL-5 and IL-13, eosinophilia, mucus production, IgE, and alternatively activated macrophages (AAM). However, despite the lack of neutrophil chemoattractants such as CXCL1, neutrophils, a feature of type-1 immunity, are observed in type-2 responses. Consequently, alternative mechanisms must exist to ensure that neutrophils can contribute to type-2 immune reactions without escalation of deleterious inflammation. We now demonstrate that type-2 immune-associated neutrophil infiltration is regulated by the mouse RNase A homologue, eosinophil-associated ribonuclease 11 (Ear11), which is secreted by AAM downstream of IL-25-stimulated ILC2. Transgenic overexpression of Ear11 resulted in tissue neutrophilia, whereas Ear11-deficient mice have fewer resting tissue neutrophils, whilst other type-2 immune responses are not impaired. Notably, administration of recombinant mouse Ear11 increases neutrophil motility and recruitment. Thus, Ear11 helps maintain tissue neutrophils at homoeostasis and during type-2 reactions when chemokine-producing classically activated macrophages are infrequently elicited.


Subject(s)
Immunity, Innate , Lymphocytes/physiology , Macrophage Activation/immunology , Macrophages/physiology , Neutrophil Infiltration/immunology , Neutrophils/physiology , Ribonucleases/biosynthesis , Animals , Dendritic Cells/immunology , Dendritic Cells/metabolism , Eosinophils/immunology , Eosinophils/metabolism , Immunomodulation , Immunophenotyping , Interleukin-13/biosynthesis , Lung/immunology , Lung/metabolism , Lung/pathology , Mice , Mice, Transgenic , Ribonucleases/genetics
2.
Sci Rep ; 7: 44899, 2017 03 21.
Article in English | MEDLINE | ID: mdl-28322325

ABSTRACT

Haemangioblastoma is a rare malignancy of the CNS where vascular proliferation causes lesions due to endothelial propagation. We found that conditionally expressing mutant Kras, using Rag1-Cre, gave rise to CNS haemangioblastoma in the cortex and cerebellum in mice that present with highly vascular tumours with stromal cells similar to human haemangioblastomas. The aberrant haemangioblastoma endothelial cells do not express mutant Kras but rather the mutant oncogene is expressed in CNS interstitial cells, including neuronal cells and progeny. This demonstrates a non-cell autonomous origin of this disease that is unexpectedly induced via Rag1-Cre expression in CNS interstitial cells. This is the first time that mutant RAS has been shown to stimulate non-cell autonomous proliferation in malignancy and suggests that mutant RAS can control endothelial cell proliferation in neo-vascularisation when expressed in certain cells.


Subject(s)
Cerebellar Neoplasms/genetics , Cerebellar Neoplasms/pathology , Genes, ras , Hemangioblastoma/genetics , Hemangioblastoma/pathology , Mutation , Animals , Cerebellar Neoplasms/mortality , Disease Models, Animal , Gene Expression , Genes, Reporter , Hemangioblastoma/mortality , Humans , Incidence , Mice , Mice, Transgenic
3.
J Gastroenterol ; 47(11): 1198-211, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22539101

ABSTRACT

BACKGROUND: Interleukin-25 (IL-25) is a potent activator of type-2 immune responses. Mucosal inflammation in ulcerative colitis is driven by type-2 cytokines. We have previously shown that a neutralizing anti-IL-25 antibody abrogated airways hyperreactivity in an experimental model of lung allergy. Therefore, we asked whether blocking IL-25 via neutralizing antibodies against the ligand or its receptor IL-17BR could protect against inflammation in an oxazolone-induced mouse model of colitis. METHODS: Neutralizing antibodies to IL-25 or IL-17BR were administered to mice with oxazolone-induced colitis, a model of ulcerative colitis. The disease onset was evaluated by weight loss and degree of colon ulceration. Also, lamina propria and mesenteric lymph node (MLN) infiltrates were assessed for mucosal inflammation and cultured in vitro to determine cytokine production. RESULTS: We found that in oxazolone colitis IL-25 production derives from intestinal epithelial cells and that IL-17BR(+) IL-13-producing natural killer T (NKT) cells and nuocytes drive the intestinal inflammation. Blocking IL-25 signalling considerably improved the clinical aspects of the disease, including weight loss and colon ulceration, and resulted in fewer nuocytes and NKT cells infiltrating the mucosa. The improved pathology correlated with a decrease in IL-13 production by lamina propria cells, a decrease in the production of other type-2 cytokines by MLN cells, and a decrease in blood eosinophilia and IgE. CONCLUSION: IL-25 plays a pro-inflammatory role in the oxazolone colitis model, and neutralizing antibodies to IL-25 or IL-17BR can slow the ongoing inflammation in this disease. Because this model mimics aspects of human ulcerative colitis, these antibodies may represent potential therapeutics for reducing gut inflammation in patients.


Subject(s)
Antibodies, Neutralizing/immunology , Colitis, Ulcerative/immunology , Interleukin-17/immunology , Receptors, Interleukin-17/immunology , Animals , Antibodies, Neutralizing/administration & dosage , Colitis, Ulcerative/pathology , Disease Models, Animal , Female , Inflammation/immunology , Interleukin-13/immunology , Intestinal Mucosa/immunology , Intestinal Mucosa/pathology , Mice , Mice, Inbred BALB C , Natural Killer T-Cells/immunology , Oxazolone/toxicity , Signal Transduction/immunology
4.
Nat Immunol ; 13(3): 229-36, 2012 Jan 22.
Article in English | MEDLINE | ID: mdl-22267218

ABSTRACT

Nuocytes are essential in innate type 2 immunity and contribute to the exacerbation of asthma responses. Here we found that nuocytes arose in the bone marrow and differentiated from common lymphoid progenitors, which indicates they are distinct, previously unknown members of the lymphoid lineage. Nuocytes required interleukin 7 (IL-7), IL-33 and Notch signaling for development in vitro. Pro-T cell progenitors at double-negative stage 1 (DN1) and DN2 maintained nuocyte potential in vitro, although the thymus was not essential for nuocyte development. Notably, the transcription factor RORα was critical for the development of nuocytes and their role in the expulsion of parasitic worms.


Subject(s)
Cell Differentiation , Leukocytes/immunology , Nuclear Receptor Subfamily 1, Group F, Member 1/immunology , Animals , Interleukin-7/immunology , Interleukin-7/metabolism , Leukocytes/cytology , Leukocytes/metabolism , Mice , Nippostrongylus/immunology , Nuclear Receptor Subfamily 1, Group F, Member 1/genetics , Nuclear Receptor Subfamily 1, Group F, Member 1/metabolism , Signal Transduction , Strongylida Infections/immunology , Thymocytes/immunology
5.
J Allergy Clin Immunol ; 129(1): 191-8.e1-4, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22079492

ABSTRACT

BACKGROUND: IL-4, IL-5, and IL-13 are thought to be central to the allergic asthmatic response. Previous work supposed that the essential source of these cytokines was CD4(+) T(H)2 cells. However, more recent studies have suggested that other innate production of type 2 cytokines might be as important. OBJECTIVES: Nuocytes are a novel population of IL-13-producing innate cells, which are critical for protective immunity in Nippostrongylus brasiliensis infection. Given this, we investigated the potential existence and functional importance of nuocytes in experimental allergic asthma. METHODS: We generated Il4(+/eGFP)Il13(+/Tomato) dual-reporter mice to study cytokine-producing cells during allergic inflammation. We adoptively transferred innate IL-13-producing cells to investigate their role in airways hyperreactivity (AHR). RESULTS: We show that allergen-induced nuocytes infiltrate the lung and are a major innate source of IL-13. CD4(+) T cells in the lung almost exclusively express only IL-13, whereas IL-4-producing T cells were restricted to the draining lymph nodes. Intranasal administration of IL-25 or IL-33 induced IL-13-producing nuocytes in the BAL fluid. Strikingly, adoptive transfer of wild-type nuocytes, but not Il13(-/-) nuocytes, into Il13(-/-) mice, which are normally resistant to IL-25-induced AHR, restored airways resistance and lung cell infiltration. CONCLUSIONS: These findings identify nuocytes as a novel cell type in allergic lung inflammation and an innate source of IL-13 that can directly induce AHR in the absence of IL-13-producing CD4(+) T cells. These data highlight nuocytes as an important new consideration in the development of future allergic asthma therapy.


Subject(s)
Asthma/immunology , Bronchial Hyperreactivity/immunology , Immunity, Innate , Interleukin-13/biosynthesis , Pneumonia/immunology , Administration, Intranasal , Animals , Asthma/genetics , Asthma/metabolism , Bronchial Hyperreactivity/genetics , Bronchial Hyperreactivity/metabolism , Cytokines/administration & dosage , Cytokines/genetics , Cytokines/immunology , Disease Models, Animal , Mice , Mice, Inbred BALB C , Mice, Transgenic , Pneumonia/genetics , Pneumonia/metabolism
6.
Genes Dev ; 25(9): 917-29, 2011 May 01.
Article in English | MEDLINE | ID: mdl-21536732

ABSTRACT

Removal of the assembly factor eukaryotic initiation factor 6 (eIF6) is critical for late cytoplasmic maturation of 60S ribosomal subunits. In mammalian cells, the current model posits that eIF6 release is triggered following phosphorylation of Ser 235 by activated protein kinase C. In contrast, genetic studies in yeast indicate a requirement for the ortholog of the SBDS (Shwachman-Bodian-Diamond syndrome) gene that is mutated in the inherited leukemia predisposition disorder Shwachman-Diamond syndrome (SDS). Here, by isolating late cytoplasmic 60S ribosomal subunits from Sbds-deleted mice, we show that SBDS and the GTPase elongation factor-like 1 (EFL1) directly catalyze eIF6 removal in mammalian cells by a mechanism that requires GTP binding and hydrolysis by EFL1 but not phosphorylation of eIF6 Ser 235. Functional analysis of disease-associated missense variants reveals that the essential role of SBDS is to tightly couple GTP hydrolysis by EFL1 on the ribosome to eIF6 release. Furthermore, complementary NMR spectroscopic studies suggest unanticipated mechanistic parallels between this late step in 60S maturation and aspects of bacterial ribosome disassembly. Our findings establish a direct role for SBDS and EFL1 in catalyzing the translational activation of ribosomes in all eukaryotes, and define SDS as a ribosomopathy caused by uncoupling GTP hydrolysis from eIF6 release.


Subject(s)
Eukaryotic Initiation Factors/metabolism , Guanosine Triphosphate/metabolism , Ribosomes/pathology , Animals , Bone Marrow Diseases/genetics , Bone Marrow Diseases/physiopathology , Catalysis , Cells, Cultured , Disease Models, Animal , Eukaryotic Initiation Factors/genetics , Exocrine Pancreatic Insufficiency/genetics , Exocrine Pancreatic Insufficiency/physiopathology , Humans , Hydrolysis , Lipomatosis , Liver/pathology , Mice , Mice, Inbred C57BL , Models, Molecular , Mutation , Peptide Initiation Factors/genetics , Peptide Initiation Factors/metabolism , Phosphorylation , Protein Binding , Protein Structure, Tertiary , Proteins/chemistry , Proteins/genetics , Proteins/metabolism , Ribosome Subunits, Large, Eukaryotic , Shwachman-Diamond Syndrome
7.
Cell Cycle ; 9(21): 4286-93, 2010 Nov 01.
Article in English | MEDLINE | ID: mdl-20980806

ABSTRACT

Myelodysplastic Syndromes (MDS) are a heterogeneous group of acquired clonal bone marrow disorders, characterised by ineffective hematopoiesis. The mechanisms underlying many of these blood disorders have remained elusive due to the difficulty in pinpointing specific gene mutations or haplo-insufficencies, which can occur within large deleted regions. However, there is an increasing interest in the classification of some of these diseases as ribosomopathies. Indeed, studies have implicated Ribosomal Protein (RP) S14 as a strong candidate for haploinsufficiency in 5q- syndrome, a particular form of MDS. Recently, two novel mouse models have provided evidence for the involvement of both RPS14 and the p53 pathway, and specific miRNAs in 5q- syndrome. In this review we will discuss: 5q- syndrome mouse models, the possible mechanisms underlying this blood disorder with respect to the candidate genes and comparisons with other ribosomopathies and the involvement of the p53 pathway in these diseases.


Subject(s)
Ribosomal Proteins/metabolism , Anemia, Macrocytic/genetics , Anemia, Macrocytic/metabolism , Animals , Antigens, Differentiation, B-Lymphocyte/genetics , Antigens, Differentiation, B-Lymphocyte/physiology , Chromosome Deletion , Chromosomes, Human, Pair 5/genetics , Chromosomes, Human, Pair 5/metabolism , Disease Models, Animal , Histocompatibility Antigens Class II/genetics , Histocompatibility Antigens Class II/physiology , Humans , Membrane Proteins/genetics , Membrane Proteins/physiology , Mice , MicroRNAs/metabolism , Neoplasm Proteins/genetics , Neoplasm Proteins/physiology , RNA Interference , Ribosomal Proteins/genetics , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism
9.
Nat Med ; 16(1): 59-66, 2010 Jan.
Article in English | MEDLINE | ID: mdl-19966810

ABSTRACT

The identification of the genes associated with chromosomal translocation breakpoints has fundamentally changed understanding of the molecular basis of hematological malignancies. By contrast, the study of chromosomal deletions has been hampered by the large number of genes deleted and the complexity of their analysis. We report the generation of a mouse model for human 5q- syndrome using large-scale chromosomal engineering. Haploinsufficiency of the Cd74-Nid67 interval (containing Rps14, encoding the ribosomal protein S14) caused macrocytic anemia, prominent erythroid dysplasia and monolobulated megakaryocytes in the bone marrow. These effects were associated with defective bone marrow progenitor development, the appearance of bone marrow cells expressing high amounts of the tumor suppressor p53 and increased bone marrow cell apoptosis. Notably, intercrossing with p53-deficient mice completely rescued the progenitor cell defect, restoring common myeloid progenitor and megakaryocytic-erythroid progenitor, granulocyte-monocyte progenitor and hematopoietic stem cell bone marrow populations. This mouse model suggests that a p53-dependent mechanism underlies the pathophysiology of the 5q- syndrome.


Subject(s)
Anemia, Macrocytic/genetics , Chromosome Deletion , Disease Models, Animal , Genes, p53/genetics , Myelodysplastic Syndromes/genetics , Animals , Apoptosis/genetics , Chromosomes, Mammalian/genetics , Hematopoietic Stem Cells/physiology , Humans , Mice , Synteny/genetics
10.
EMBO J ; 24(17): 3136-46, 2005 Sep 07.
Article in English | MEDLINE | ID: mdl-16096649

ABSTRACT

Chromosomal translocations are primary events in tumorigenesis. Those involving the mixed lineage leukaemia (MLL) gene are found in various guises and it is unclear whether MLL fusions can affect haematopoietic differentiation. We have used a model in which chromosomal translocations are generated in mice de novo by Cre-loxP-mediated recombination (translocator mice) to compare the functionally relevant haematopoietic cell contexts for Mll fusions, namely pluripotent stem cells, semicommitted progenitors or committed cells. Translocations between Mll and Enl or Af9 cause myeloid neoplasias, initiating in pluripotent stem cells or multipotent myeloid progenitors. However, while Mll-Enl translocations can also cause leukaemia from T-cell progenitors, no tumours arose with Mll-Af9 translocations in the T-cell compartment. Furthermore, Mll-Enl translocations in T-cell progenitors can cause lineage reassignment into myeloid tumours. Therefore, a permissive cellular environment is required for oncogenicity of Mll-associated translocations and Mll fusions can influence haematopoietic lineage commitment.


Subject(s)
Cell Transformation, Neoplastic/pathology , DNA-Binding Proteins/metabolism , Integrases/metabolism , Recombination, Genetic , Transcription Factors/metabolism , Translocation, Genetic/physiology , Animals , Bone Marrow Cells/metabolism , Bone Marrow Cells/pathology , Cell Lineage/physiology , Cell Transformation, Neoplastic/genetics , Cell Transformation, Neoplastic/metabolism , Cells, Cultured , DNA-Binding Proteins/genetics , Histone-Lysine N-Methyltransferase , Integrases/genetics , Leukemia, Lymphoid/metabolism , Leukemia, Lymphoid/pathology , Leukemia, Myeloid/metabolism , Leukemia, Myeloid/pathology , Mice , Multipotent Stem Cells/metabolism , Multipotent Stem Cells/pathology , Myeloid Progenitor Cells/metabolism , Myeloid Progenitor Cells/pathology , Myeloid-Lymphoid Leukemia Protein , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Pluripotent Stem Cells/metabolism , Pluripotent Stem Cells/pathology , Proto-Oncogenes/genetics , T-Lymphocytes/metabolism , T-Lymphocytes/pathology , Transcription Factors/genetics , Translocation, Genetic/genetics
11.
PLoS Biol ; 3(8): e242, 2005 Aug.
Article in English | MEDLINE | ID: mdl-15974803

ABSTRACT

The EWS-ERG fusion protein is found in human sarcomas with the chromosomal translocation t(21;22)(q22;q12), where the translocation is considered to be an initiating event in sarcoma formation within uncommitted mesenchymal cells, probably long-lived progenitors capable of self renewal. The fusion protein may not therefore have an oncogenic capability beyond these progenitors. To assess whether EWS-ERG can be a tumour initiator in cells other than mesenchymal cells, we have analysed Ews-ERG fusion protein function in a cellular environment not typical of that found in human cancers, namely, committed lymphoid cells. We have used Ews-ERG invertor mice having an inverted ERG cDNA cassette flanked by loxP sites knocked in the Ews intron 8, crossed with mice expressing Cre recombinase under the control of the Rag1 gene to give conditional, lymphoid-specific expression of the fusion protein. Clonal T cell neoplasias arose in these mice. This conditional Ews gene fusion model of tumourigenesis shows that Ews-ERG can cause haematopoietic tumours and the precursor cells are committed cells. Thus, Ews-ERG can function in cells that do not have to be pluripotent progenitors or mesenchymal cells.


Subject(s)
Leukemia, T-Cell/genetics , Lymphoma, T-Cell/genetics , Oncogene Proteins, Fusion/physiology , Transcription Factors/physiology , Animals , B-Lymphocytes/metabolism , Base Sequence , Bone Marrow/pathology , Cell Lineage , Genes, T-Cell Receptor beta , Humans , Integrases/metabolism , Leukemia, T-Cell/pathology , Lymphoma, T-Cell/pathology , Mice , Models, Animal , Molecular Sequence Data , Oncogene Fusion , Oncogene Proteins, Fusion/genetics , RNA, Messenger/metabolism , T-Lymphocytes/cytology , T-Lymphocytes/metabolism , Thymoma/genetics , Thymoma/immunology , Thymoma/pathology , Thymus Neoplasms/genetics , Thymus Neoplasms/immunology , Thymus Neoplasms/pathology , Transcription Factors/genetics , Viral Proteins/metabolism
12.
Nat Methods ; 2(1): 27-30, 2005 Jan.
Article in English | MEDLINE | ID: mdl-15782166

ABSTRACT

Knock-in models of tumor-specific chromosomal translocations can generate lethal mutations. To circumvent this, a new conditional gene fusion model has been developed (invertor mice) and exemplified with the Ews-ERG fusion oncogene. An ERG segment, flanked by loxP sites, was knocked in to an intron of the Ews gene but in an inverted transcription orientation and lineage-specific Ews-ERG fusion created by Cre-mediated inversion. This invertor method is a completely conditional approach, applicable to any gene fusion, to emulate effects of translocations found in human cancers.


Subject(s)
Chromosomes/ultrastructure , Genetic Techniques , Alleles , Animals , Base Sequence , Cell Separation , DNA, Complementary/metabolism , Flow Cytometry , Humans , Introns , Mice , Mice, Inbred C57BL , Models, Genetic , Molecular Sequence Data , Mutation , RNA/metabolism , Recombination, Genetic , Reverse Transcriptase Polymerase Chain Reaction , Transcription, Genetic , Translocation, Genetic
13.
Cancer Cell ; 3(5): 449-58, 2003 May.
Article in English | MEDLINE | ID: mdl-12781363

ABSTRACT

The etiology of human tumors often involves chromosomal translocations. Models that emulate translocations are essential to understanding the determinants of frank malignancy, those dictating the restriction of translocations to specific lineages, and as a basis for development of rational therapeutic methods. We demonstrate that developmentally regulated Cre-loxP-mediated interchromosomal recombination between the Mll gene, whose human counterpart is involved in a spectrum of leukemias, and the Enl gene creates reciprocal chromosomal translocations that cause myeloid tumors. There is a rapid onset and high penetrance of leukemogenesis in these translocator mice, and high proportions of cells carrying chromosomal translocations can be found in bone marrow as early as 12 days after birth. This de novo strategy is a direct recapitulation of naturally occurring human cancer-associated translocations.


Subject(s)
Chromosomes/ultrastructure , DNA-Binding Proteins/genetics , Genetic Techniques , Neoplasms/genetics , Oncogene Proteins, Fusion/genetics , Proto-Oncogenes , Transcription Factors , Translocation, Genetic , Alleles , Amino Acid Sequence , Animals , Base Sequence , Disease Models, Animal , Flow Cytometry , Genotype , Histone-Lysine N-Methyltransferase , Humans , In Situ Hybridization, Fluorescence , Leukocytes/metabolism , Mice , Models, Genetic , Molecular Sequence Data , Myeloid-Lymphoid Leukemia Protein , Phenotype , Recombination, Genetic , Time Factors
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