ABSTRACT
BACKGROUND AND AIMS: Persistent inflammation observed in inflammatory bowel disease may be the consequence of an increased or aberrant immune response to normal gut constituents or an overall immune dysregulation and imbalance. Cytokines play an important role in immune regulation and interleukin 18 (IL-18) is one such cytokine that has emerged as being instrumental in driving CD4+ T cell responses towards a Th1-type. IL-18 can also directly mediate inflammation, moderate interleukin 1 activity, and can act on cell types other than T cells. It has been reported recently that IL-18 mRNA and protein are upregulated in gut tissue from IBD patients. The aim of this study was to understand more about the role of IL-18 in contributing to the pathology of IBD and to assess whether blocking IL-18 activity may be of therapeutic benefit as a treatment regimen for IBD. METHODS: Mice with dextran sulphate sodium (DSS) induced colitis were treated with recombinant IL-18 binding protein (IL-18bp.Fc), a soluble protein that blocks IL-18 bioactivity. Histopathological analysis was performed and RNA from the large intestine was analysed using the RNase protection assay and gene arrays. RESULTS: IL-18 RNA levels increased very early in the colon during DSS colitis. Treatment of mice with IL-18bp.Fc inhibited IBD associated weight loss and significantly inhibited the intestinal inflammation induced by DSS. IL-18bp.Fc treatment also attenuated mRNA upregulation of multiple proinflammatory cytokine genes, chemokine genes, and matrix metalloprotease genes in the large intestine that are commonly elevated during IBD. CONCLUSIONS: IL-18bp treatment attenuated inflammation during DSS induced colitis in mice. Neutralising IL-18 activity may therefore be of benefit for ameliorating the inflammation associated with human intestinal diseases.
Subject(s)
Colitis, Ulcerative/chemically induced , Dextran Sulfate/adverse effects , Glycoproteins/pharmacology , Interleukin-18/antagonists & inhibitors , Animals , Colitis, Ulcerative/pathology , Cytokines/metabolism , Female , Immunohistochemistry/methods , Intercellular Signaling Peptides and Proteins , Interleukin-18/physiology , Lymph Nodes/physiology , Mesentery , Mice , Mice, Inbred C57BL , RNA/metabolism , Up-Regulation , Weight LossABSTRACT
The cellular receptor for murine thymic stromal lymphopoietin (TSLP) was detected in a variety of murine, but not human myelomonocytic cell lines by radioligand binding. cDNA clones encoding the receptor were isolated from a murine T helper cell cDNA library. TSLP receptor (TSLPR) is a member of the hematopoietin receptor family. Transfection of TSLPR cDNA resulted in only low affinity binding. Cotransfection of the interleukin 7 (IL-7)Ralpha chain cDNA resulted in conversion to high affinity binding. TSLP did not activate cells from IL-7Ralpha(-/)- mice, but did activate cells from gammac(-/)- mice. Thus, the functional TSLPR requires the IL-7Ralpha chain, but not the gammac chain for signaling.
Subject(s)
Hematopoiesis/drug effects , Lymphocytes/drug effects , Receptors, Cytokine/physiology , Receptors, Interleukin-7/physiology , Amino Acid Sequence , Animals , Base Sequence , Cell Line , Chromosome Mapping , Cloning, Molecular , Cytokines/pharmacology , Humans , Interleukin-7/pharmacology , Lymphocytes/physiology , Male , Mice , Molecular Sequence Data , RNA, Messenger/analysis , Receptors, Cytokine/chemistry , Receptors, Cytokine/genetics , Receptors, Interleukin-7/chemistry , Recombinant Proteins/chemistry , Signal TransductionABSTRACT
Using a bioassay consisting of the proliferation of a murine B cell line, a cDNA of a gene whose product supports the growth of that cell line was isolated from a thymic stromal cell line. This factor, termed thymic stromal lymphopoietin (TSLP), is a protein of 140 amino acids. The gene encoding TSLP was mapped to murine chromosome 18. Purified recombinant TSLP supported the growth of pre-B cell colonies in vitro, but had no myelopoietic activity. TSLP had comitogenic activity for fetal thymocytes, but was not as potent as interleukin 7 in lobe submersion cultures. Injection of TSLP into neonatal mice induced the expansion of B220(+)BP-1(+) pre-B cells.
Subject(s)
B-Lymphocytes/drug effects , Cytokines/pharmacology , Hematopoiesis/drug effects , Amino Acid Sequence , Animals , Base Sequence , Chromosome Mapping , Cloning, Molecular , Cytokines/chemistry , Cytokines/genetics , DNA, Complementary/isolation & purification , Female , Interleukin-7/pharmacology , Male , Mice , Mice, Inbred C57BL , Molecular Sequence Data , RNA, Messenger/analysis , Recombinant Proteins/chemistry , Recombinant Proteins/isolation & purification , Recombinant Proteins/pharmacology , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Two novel members of the interleukin-1 receptor (IL-1R) family, identified by homology searches of human genomic sequence data bases, are described. The genes have been named according to their structural features: TIGIRR-1 (three immunoglobulin domain-containing IL-1 receptor-related) and TIGIRR-2. TIGIRR-2 has recently been identified as causing mental retardation when mutated (Carrie, A., Jun, L., Bienvenu, T., Vinet, M. C., McDonell, N., Couvert, P., Zemni, R., Cardona, A., Van Buggenhout, G., Frints, S., Hamel, B., Moraine, C., Ropers, H. H., Strom, T., Howell, G. R., Whittaker, A., Ross, M. T., Kahn, A., Fryns, J. P., Beldjord, C., Marynen, P., and Chelly, J. (1999) Nat. Genet. 23, 25-31) and called IL1RAPL, a name we will also use henceforth. Neither receptor alone was able to mediate transcriptional activation of NF-kappaB in response to IL-1alpha, IL-1beta, or IL-18. In order to begin to elucidate the function of these and other orphan IL-1R family members, we have developed a functional assay utilizing a panel of chimeric receptors containing the extracellular and transmembrane domains of either type I IL-1R or IL-1R accessory protein (AcP) coupled to the cytoplasmic domains of all family members. Coexpression of each IL-1R chimera with each AcP chimera and an NF-kappaB-responsive reporter demonstrated that the cytoplasmic domains could be classified as IL-1R-like, AcP-like, or neither. Any IL-1R-like cytoplasmic domain could cooperate with any AcP-like cytoplasmic domain. The TIGIRR-1 and IL1RAPL cytoplasmic domains, however, were unable to signal as either IL-1R-like or AcP-like components, suggesting that they function as a new class of receptors within this family.
Subject(s)
Multigene Family , Receptors, Interleukin-1/metabolism , Amino Acid Sequence , Genomic Library , Humans , Interleukin-1/metabolism , Interleukin-1 Receptor Accessory Protein , Interleukin-18/metabolism , Molecular Sequence Data , NF-kappa B/metabolism , Proteins/genetics , Proteins/metabolism , Receptors, Interleukin-1/classification , Receptors, Interleukin-1/genetics , Recombinant Fusion Proteins/metabolism , Sequence Homology, Amino Acid , Signal TransductionABSTRACT
We report here the cloning and characterization of four new members of the interleukin-1 (IL-1) family (FIL1delta, FIL1epsilon, FIL1zeta, and FIL1eta, with FIL1 standing for "Family of IL-1"). The novel genes demonstrate significant sequence similarity to IL-1alpha, IL-1beta, IL-1ra, and IL-18, and in addition maintain a conserved exon-intron arrangement that is shared with the previously known members of the family. Protein structure modeling also suggests that the FIL1 genes are related to IL-1beta and IL-1ra. The novel genes form a cluster with the IL-1s on the long arm of human chromosome 2.
Subject(s)
Chromosomes, Human, Pair 2 , Interleukin-1/genetics , Multigene Family , Amino Acid Sequence , Animals , Base Sequence , Cell Line , Chromosome Mapping , Cloning, Molecular , Conserved Sequence , Exons , Female , Genomic Library , Humans , Interleukin-1/chemistry , Interleukin-1/metabolism , Introns , Male , Mice , Models, Molecular , Molecular Sequence Data , Open Reading Frames , Protein Conformation , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Structure, Secondary , Receptors, Interleukin-1/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Tumor Cells, CulturedABSTRACT
Murine MyD88, an RNA with homology both to the interleukin-1 receptor signaling domain and to 'death-domains', is rapidly upregulated during differentiation of the myeloleukemic cell line M1. We have cloned the human homologue of murine MyD88 and re-evaluated the murine sequence. The open reading frame for both species encodes a 296 amino acid protein, which for murine MyD88 is 53 amino acids longer than originally published. Human MyD88 cDNA is encoded by 5 exons, and maps to chromosome 3p21.3-p22 by fluorescence in situ hybridization (FISH). Overexpression of the death domain region leads to transcriptional activation of the IL-8 promoter.
Subject(s)
Antigens, Differentiation , Proteins/genetics , Receptors, Immunologic , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Animals , Base Sequence , COS Cells , Chromosome Mapping , Chromosomes, Human, Pair 3 , DNA, Complementary/genetics , Exons , Gene Expression , Humans , In Situ Hybridization, Fluorescence , Interleukin-8/genetics , Introns , Mice , Molecular Sequence Data , Myeloid Differentiation Factor 88 , Oligonucleotides, Antisense , Promoter Regions, Genetic , Proteins/chemistry , RNA, Messenger/genetics , Receptors, Interleukin-1/chemistry , Receptors, Interleukin-1/genetics , TransfectionABSTRACT
Through data base searches, we have discovered new proteins that share homology with the signaling domain of the type I interleukin-1 receptor (IL-1RI): human "randomly sequenced cDNA 786" (rsc786), murine MyD88, and two partial Drosophila open reading frames, MstProx and STSDm2245. Comparisons between these new proteins and known IL-1RI homologous proteins such as Toll, 18-Wheeler, and T1/ST2 revealed six clusters of amino acid similarity. We tested the hypothesis that sequence similarity between the signaling domain of IL-1RI and the three mammalian family members might indicate functional similarity. Chimeric IL-1RI receptors expressing the putative signaling domains of T1/ST2, MyD88, and rsc786 were assayed by three separate IL-1 responsive assays, NF-kappaB, phosphorylation of an epidermal growth factor receptor peptide, and an interleukin 8 promoter-controlled reporter construct, for their ability to transduce an IL-1-stimulated signal. All three assays were positive in response to the T1/ST2 chimera, while the MyD88 and rsc786 chimeras failed to respond. These data indicate that the sequence homology between IL-1RI and T1/ST2 indicates a functional homology as well.
Subject(s)
Membrane Proteins , Proteins/chemistry , Proteins/physiology , Receptors, Interleukin-1/chemistry , Receptors, Interleukin-1/physiology , Signal Transduction , Amino Acid Sequence , Animals , Base Sequence , Cell Line , Chlorocebus aethiops , Cloning, Molecular , Consensus Sequence , DNA Primers , Drosophila melanogaster/metabolism , ErbB Receptors/metabolism , Humans , Interleukin-1 Receptor-Like 1 Protein , Kinetics , Mammals , Mice , Molecular Sequence Data , NF-kappa B/metabolism , Phosphorylation , Protein Biosynthesis , Rats , Receptors, Cell Surface , Receptors, Interleukin , Receptors, Interleukin-1/biosynthesis , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/metabolism , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , TransfectionABSTRACT
A novel member of the interleukin-1 receptor family has been cloned by polymerase chain reaction using degenerate oligonucleotide primers derived from regions of sequence conservation, using as template a yeast artificial chromosome known to contain both interleukin-1 (IL-1) receptors and T1/ST2. The new receptor, called IL-1 receptor-related protein or IL-1Rrp, fails to bind any of the known IL-1 ligands. A chimeric receptor, in which the IL-1Rrp cytoplasmic domain is fused to the extracellular and transmembrane regions of the IL-1 receptor, responds to IL-1 following transfection into COS cells by activation of NFkappaB and induction of IL-8 promoter function.
