| Production of the rat type 1 insulin-like growth factor-binding protein by well differentiated H4EIIC3 hepatoma cells: identification, purification, and N-terminal amino acid analysis. | |
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MedLine Citation:
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PMID: 2164920 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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We recently identified a 32 K mol wt insulin-like growth factor (IGF)-binding protein (BP) which is markedly increased in the serum of streptozotocin-diabetic rats and recognized by antiserum against the human amniotic fluid IGFBP (hIGFBP-1). In the present study we sought to confirm that this protein represents the rat homolog of IGFBP-1 (rIGFBP-1), and that rIGFBP-1 may, therefore, play an important role in the regulation of IGF bioactivity in experimental diabetes. Since the abundance of related hepatic mRNA is high in diabetic rats, we asked whether well differentiated H4EIIC3 rat hepatoma cells produce rIGFBP-1 and provide sufficient amounts of this protein for purification and further characterization. Specific IGF-binding activity in hepatoma conditioned medium was detected initially by incubation with 125I-labeled recombinant human IGF-II and precipitation with polyethylene glycol. Ligand blotting demonstrated a 32 K BP, identical in size to the major low mol wt IGFBP found in diabetic rat serum. Affinity labeling and immunoprecipitation confirmed that this BP is related to human IGFBP-1 and is distinct from the fetal rat IGFBP, rIGFBP-2. Incorporation of [35S]methionine into 32 K BPs confirmed synthesis by hepatoma cells. For purification of BPs, conditioned medium was collected in roller culture, and BPs were purified by ammonium sulfate precipitation, Sephadex G-75 chromatography, and reverse phase HPLC. Partial amino acid sequencing of purified protein demonstrated 68% identity with the human IGFBP-1 and distinguished this BP from previously characterized rat IGFBPs. Purified protein bound both IGF-I and IGF-II with high affinity. We conclude that the 32 K IGFBP produced by H4EIIC3 hepatoma cells in culture represents the rat form of IGFBP-1 (rIGFBP-1). Regulation of rIGFBP-1 may play an important role in the modulation of IGF bioactivity in experimental animals with metabolic disease. The availability of purified rIGFBP-1 and identification of a cell line that produces this BP will greatly facilitate future studies of IGFBP-1 in the rat model. |
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Authors:
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T G Unterman; D T Oehler; M B Gotway; P W Morris |
Publication Detail:
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Type: Comparative Study; Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.; Research Support, U.S. Gov't, P.H.S. |
Journal Detail:
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Title: Endocrinology Volume: 127 ISSN: 0013-7227 ISO Abbreviation: Endocrinology Publication Date: 1990 Aug |
Date Detail:
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Created Date: 1990-08-30 Completed Date: 1990-08-30 Revised Date: 2007-11-14 |
Medline Journal Info:
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Nlm Unique ID: 0375040 Medline TA: Endocrinology Country: UNITED STATES |
Other Details:
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Languages: eng Pagination: 789-97 Citation Subset: AIM; IM |
Affiliation:
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Department of Medicine, University of Illinois College of Medicine, Chicago. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Amino Acid Sequence Animals Cell Differentiation Chromatography, Gel Chromatography, High Pressure Liquid Immunoblotting Liver Neoplasms, Experimental / metabolism* Molecular Sequence Data Rats Rats, Inbred BUF Receptors, Cell Surface / biosynthesis*, genetics, isolation & purification Receptors, Somatomedin Sequence Homology, Nucleic Acid Somatomedins / metabolism Tumor Cells, Cultured / cytology, metabolism* |
| Grant Support | |
ID/Acronym/Agency:
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1-S10-RR-03521/RR/NCRR NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Receptors, Cell Surface; 0/Receptors, Somatomedin; 0/Somatomedins |
| Comments/Corrections | |
Erratum In:
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Endocrinology 1990 Oct;127(4):1977 |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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