Developmental expression of rat insulin-like growth factor binding protein-2 by astrocytic glial cells in culture.
Brain astrocytes were established in primary culture from postnatal and adult rats to characterize the developmental expression of secreted proteins. Astrocytes cultured from 21-day rat brain, but not 1-day rat brain, secreted a distinct group of proteins with Mr of 35,000 as determined by analysis of [35S]methionine-labeled proteins using two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. N-terminal amino acid sequence analysis of this protein group showed 100% identity to rat insulin-like growth factor binding protein-2 (rIGFBP-2), the BRL-3A IGFBP purified from a fetal rat liver cell line. An antiserum was generated against this astrocyte 35,000 Mr protein, and immunoblot analysis revealed a dramatic increase in rIGFBP-2 secretion in astrocytes cultured from 14-day, 21-day, and adult rat brain compared to astrocytes from 1-day and 7-day rat brain. Similar analysis of neonatal rat brain neurons in culture failed to show immunoreactive rIGFBP-2 in cell lysates or secreted protein. Ligand Western blot analysis demonstrated [125I]IGF-II binding to a single protein band which comigrated with a prominant rIGFBP-2 immunoreactive species in nonreduced conditioned medium from 21-day astrocytes. In comparison [125I]IGF-II binding proteins were detected only at low levels in medium from astrocytes cultured from 1-day rat brain and were undetectable in neuron-conditioned media. Northern blot analysis using a rIGFBP-2 complementary DNA revealed 5-fold greater messenger RNA levels in astrocytes from 21-day rat brain compared with astrocytes from 1-day brain, whereas neonate neurons showed no transcripts. Thus, rIGFBP-2 exhibits a pattern of developmental and cell-specific expression in cultured rat brain cells.
Olson
J A
JA
Department of Pharmacology and Therapeutics, University of Florida College of Medicine, Gainesville 32610.
Shiverick
K T
KT
Ogilvie
S
S
Buhi
W C
WC
Raizada
M K
MK
D000375:Aging / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn / Q000378:metabolism; D001253:Astrocytes / Q000378:metabolism*; D001921:Brain / Q000166:cytology / Q000254:growth & development*; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D018971:Insulin-Like Growth Factor Binding Protein 2; D007335:Insulin-Like Growth Factor II / Q000378:metabolism; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains
D002352:Carrier Proteins; D018971:Insulin-Like Growth Factor Binding Protein 2; D012333:RNA, Messenger; D007335:Insulin-Like Growth Factor II