Hypoxia stimulates insulin-like growth factor binding protein 1 (IGFBP-1) gene expression in HepG2 cells: a possible model for IGFBP-1 expression in fetal hypoxia.
IGFBP-1 is elevated in fetuses with long-term, chronic hypoxia and intrauterine growth restriction. We investigated the hypothesis that hypoxia regulates IGFBP-1 in the human fetus in vivo and IGFBP-1 gene expression and protein in vitro. Umbilical artery IGFBP-1 levels (mean +/- SEM) from term babies with respiratory acidosis (acute hypoxia), normal babies, and those with mixed respiratory/metabolic acidosis (more profound and prolonged hypoxia) were measured using an immunoradiometric assay. IGFBP-1 levels were similar in normal (n = 12) and acutely hypoxic (n = 6) babies (189.1 +/- 71.8 vs. 175.8 +/- 45.9 ng /ml, respectively, P = 0.789). However, with more profound and prolonged hypoxia (n = 19), IGFBP-1 levels were markedly elevated (470.6 +/- 80.0 ng /ml, P = 0.044). To investigate IGFBP-1 regulation by hypoxia in vitro, HepG2 cells were incubated under hypoxia (pO2 = 2%) and normoxia (pO2 = 20%). IGFBP-1 protein and mRNA increased 8- and 12-fold, respectively, under hypoxic conditions. Hypoxia did not affect protein or mRNA levels of IGFBP-2 or -4. IGFBP-5 and -6 mRNAs, undetectable in control cells, were not induced by hypoxia, whereas minimally expressed IGFBP-3 mRNA increased twofold. Investigation into IGFBP-1 gene structure revealed three potential consensus sequences for the hypoxia response element (HRE) in the first intron. To investigate functionality, a 372-bp fragment of IGFBP-1 intron 1, containing putative HREs, was placed 5' to a heterologous hsp70 promoter in a plasmid using luciferase as a reporter gene. Under hypoxia, reporter gene activity increased up to 30-fold. Mutations in the middle HRE abolished reporter activity in response to hypoxia, suggesting that this HRE is functional in the IGFBP-1 hypoxia response. Cotransfection of HRE reporter genes with a constitutively expressing hypoxia-inducible factor 1 plasmid in HepG2 cells resulted in a fourfold induction of reporter activity, suggesting a role for hypoxia-inducible factor 1 in hypoxia induction of IGFBP-1 gene expression. These data support the hypothesis that hypoxia regulation of IGFBP-1 may be a mechanism operating in the human fetus to restrict insulin-like growth factor-mediated growth in utero under conditions of chronic hypoxia and limited substrate availability.
Department of Gynecology and Obstetrics, Stanford University Medical School, Stanford, CA, USA.
S I
SI
Tazuke
N M
NM
Mazure
J
J
Sugawara
G
G
Carland
G H
GH
Faessen
L F
LF
Suen
J C
JC
Irwin
D R
DR
Powell
A J
AJ
Giaccia
L C
LC
Giudice
D000818:Animals; D001483:Base Sequence; D015687:Cell Hypoxia / Q000235:genetics*; D002460:Cell Line; D016384:Consensus Sequence; D017931:DNA Primers / Q000235:genetics; D005314:Embryonic and Fetal Development / Q000235:genetics / Q000502:physiology; D005260:Female; D005312:Fetal Blood / Q000378:metabolism; D005317:Fetal Growth Retardation / Q000209:etiology / Q000235:genetics / Q000503:physiopathology; D005311:Fetal Hypoxia / Q000150:complications / Q000235:genetics* / Q000503:physiopathology; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D007231:Infant, Newborn; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000097:blood / Q000235:genetics* / Q000502:physiology; D051379:Mice; D008954:Models, Biological; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D014162:Transfection
D017931:DNA Primers; D018970:Insulin-Like Growth Factor Binding Protein 1; D012333:RNA, Messenger