Exposure of decidualized HIESC to low oxygen tension and leucine deprivation results in increased IGFBP-1 phosphorylation and reduced IGF-I bioactivity

Abstract

Phosphorylation of decidual IGFBP-1 enhances binding of IGF-I, limiting the bioavailability of this growth factor which may contribute to reduced placental and fetal growth. The mechanisms regulating decidual IGFBP-1 phosphorylation are incompletely understood. Using decidualized human immortalized endometrial stromal cells we tested the hypothesis that low oxygen tension or reduced leucine availability, believed to be common in placental insufficiency, increase the phosphorylation of decidual IGFBP-1. Multiple reaction monitoring-MS (MRM-MS) was used to quantify IGFBP-1 phosphorylation. MRM-MS validated the novel phosphorylation of IGFBP-1 at Ser58, however this site was unaffected by low oxygen tension/leucine deprivation. In contrast, significantly elevated phosphorylation was detected for pSer119, pSer98/pSer101 and pSer169/pSer174 sites. Immunoblotting and dual-immunofluorescence using phosphosite-specific IGFBP-1 antibodies further demonstrated increased IGFBP-1 phosphorylation in HIESC under both treatments which concomitantly reduced IGF-I bioactivity. These data support the hypothesis that down regulation of IGF-I signaling links decidual IGFBP-1 hyperphosphorylation to restricted fetal growth in placental insufficiency.

Increased Insulin-like Growth Factor Binding Protein-1 Phosphorylation in Decidualized Stromal Mesenchymal Cells in Human Intrauterine Growth Restriction Placentas

“…12 However, the cellular localization of site-specific decidual IGFBP-1 phosphoisoforms is unknown and it remains to be established if the abundance of IGFBP-1 phosphorylation at distinct amino acid residues in the decidua is altered in IUGR pregnancies. Our recent findings of marked site-specific IGFBP-1 hyperphosphorylation in response to hypoxia and/or nutrient deprivation in cultured decidualized human immortalized endometrial stromal cells (HIESCs), 18 is consistent with the possibility that decidual IGFBP-1 phosphorylation at Ser101, Ser119, and Ser169 (individually or in combination) is modulated in IUGR due to placental insufficiency.…”

“…Another limitation of the study is that we examined the status of IGFBP-1 phosphorylation singly at Ser101, 119, and 169, with our existing phospho-site antibodies. Considering that in our in vitro study, multiple reaction monitoring mass spectrometry (MRM-MS) allowed us to discover dually phosphorylated sites in decidualized HIESCs, it is therefore likely that combined pSer98+pSer101 and/or pSer174+ pSer169 residues such as in HIESCs, 18 may be affected clinically in IUGR with more severe consequences to fetal growth in vivo. Furthermore, IGFBP-1 is one of the few validated markers that can selectively identify decidualized cells from among the mixed cell Representative dual immunofluorescence IHC images of merged channels of phosphorylated IGFBP-1 (pSer169; pseudo-red), vimentin (pseudo-green), and nuclei/DAPI (blue; A and E).…”

Novel roles of mechanistic target of rapamycin signaling in regulating fetal growth†

“…Hypoxia and leucine deprivation markedly increased IGFBP-1 phosphorylation and decreased IGF-I bioactivity in cultured human endometrial stromal cells decidualized in vitro [146]. Moreover, IGFBP-1 phosphorylation is increased in decidualized stromal mesenchymal cells in human IUGR [147].…”

Niche Modulation of IGF-1R Signaling: Its Role in Stem Cell Pluripotency, Cancer Reprogramming, and Therapeutic Applications

“…Furthermore, the secretion of IGF-1 from microglia increases under hypoxia, which eventually enhances VEGFR expression in endothelial cells to promote retinal angiogenesis (Yin et al, 2017 ). IGFBP-1 phosphorylation is also increased by hypoxia, resulting in limited IGF-1 bioavailability (Shehab et al, 2017 ). Moreover, hypoxia increases the secretion of IGFBP-3 from cardiomyocytes, which reduces cellular survival through the reduction of IGF-1 bioavailability (Chang R.L.…”