期刊
ENDOCRINOLOGY
卷 150, 期 1, 页码 220-231出版社
ENDOCRINE SOC
DOI: 10.1210/en.2008-0657
关键词
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资金
- NSERC
- National Institutes of Health [2RO1HL60679]
- National Science Foundation Research [IOB 0110864]
- NATIONAL HEART, LUNG, AND BLOOD INSTITUTE [R01HL060679] Funding Source: NIH RePORTER
Fetal growth restriction is often caused by uteroplacental insufficiency that leads to fetal hypoxia and nutrient deprivation. Elevated IGF binding protein (IGFBP)-1 expression associated with fetal growth restriction has been documented. In this study we tested the hypothesis that hypoxia and nutrient deprivation induce IGFBP-1 phosphorylation and increase its biological potency in inhibiting IGF actions. HepG2 cells were subjected to hypoxia and leucine deprivation to mimic the deprivation of metabolic substrates. The total IGFBP-1 levels measured by ELISA were approximately 2- to 2.5-fold higher in hypoxia and leucine deprivation-treated cells compared with the controls. Two-dimensional immunoblotting showed that whereas the nonphosphorylated isoform is the predominant IGFBP-1 in the controls, the highly phosphorylated isoforms were dominant in hypoxia and leucine deprivation-treated cells. Liquid chromatography-tandem mass spectrometry analysis revealed four serine phosphorylation sites: three known sites (pSer 101, pSer 119, and pSer 169); and a novel site (pSer 98). Liquid chromatography-mass spectrometry was used to estimate the changes of phosphorylation upon treatment. Biacore analysis indicated that the highly phosphorylated IGFBP-1 isoforms found in hypoxia and leucine deprivation-treated cells had greater affinity for IGF-I [dissociation constant 5.83E (times 10 to the power)-10 M and 6.40E-09 M] relative to the IGFBP-1 from the controls (dissociation constant similar to 1.54E-07 M). Furthermore, the highly phosphorylated IGFBP-1 had a stronger effect in inhibiting IGF-I-stimulated cell proliferation. These findings suggest that IGFBP-1 phosphorylation may be a novel mechanism of fetal adaptive response to hypoxia and nutrient restriction. (Endocrinology 150: 220-231, 2009)
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