Ovarian granulosa cell-derived insulin-like growth factor (IGF) binding proteins

Release of low molecular weight, high-affinity IGF-selective species

Eli Y. Adashi, Carol E. Resnick, Eleuterio R. Hernandez, Arye Hurwitz, Ronald (Ron) Rosenfeld

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

The ovarian granulosa cell has previously been shown to be a site of insulin-like growth factor (IGF) I production, reception, and action. It is the objective of this study to characterize in greater detail the soluble IGF binding activity released by this cell type. To this end, use was made of granulosa cells from immature diethylstilbestrol-treated rats. Serum-free media conditioned for 72 h by cultured untreated cells acquired polyethylene glycol (PEG)-precipitable [125I]IGF-I binding activity. The latter proved cell density-dependent, displaying a minimal inoculum requirement of ≤ 3 × 105 cells/culture. The daily elaboration of IGF-I binding activity appeared constant throughout the 72 h experimental period, the overall time-dependent accumulation of binding activity (over the same time period) proving virtually additive. Scatchard analysis of detailed competition studies with IGF-I suggests that the latter ligand binds to granulosa cell-derived IGF binding protein(s) (IGFBPs) with an ap0parent affinity of 3 × 10-10 M. Qualitatively similar results were obtained when using [125I]IGF-II suggesting that the IGFBPs in question are not IGF-I-selective. In fact, specificity studies using either [125I]IGF-I or [125I]GF-II revealed a rank order of competitive potencies compatible with that observed in other tissues so studied (IGF-II > IGF-I ≫ insulsin). The proteinacious nature of the acid-stable IGF binding activity under study was indicated by its sensitivity to relatively low concentrations of cycloheximide, its apparent deactivation following repeated cycles of freezing and thawing, and its virtual elimination when subjected to boiling or trypsin treatment. Cycloheximide-induced blockade of protein biosynthesis also revealed that the IGF binding activity is subject to measurable turnover thereby suggesting that its accumulation represents the balance struck between synthetic and degradative processes. Western ligand blotting of sodium dodecyl sulfate-polyacrylamide gel electrophoresis-fractionated media revealed a non-glycosylated major band doublet of 28-29 kDa. A single minor IGFBP species represented by a 23 kDa band was also appreciated in some but not all experiments. Taken together, these findings document the ability of ovarian granulosa cells to secrete a heterogenous mix of low molecular weight, high-affinity IGF-selective binding species. As such, these observations are in keeping with the concept of a complete intraovarian IGF system replete with ligands, receptors, and soluble binding activity.

Original languageEnglish (US)
Pages (from-to)175-184
Number of pages10
JournalMolecular and Cellular Endocrinology
Volume74
Issue number3
DOIs
StatePublished - Dec 21 1990
Externally publishedYes

Fingerprint

Insulin-Like Growth Factor Binding Proteins
Granulosa Cells
Somatomedins
Insulin-Like Growth Factor I
Molecular Weight
Molecular weight
Insulin-Like Growth Factor II
Cycloheximide
Ligands
Diethylstilbestrol
Thawing
Biosynthesis
Serum-Free Culture Media
Protein Biosynthesis
Electrophoresis
Cell culture
Freezing
Sodium Dodecyl Sulfate
Trypsin
Boiling liquids

Keywords

  • (Rat)
  • Insulin-like growth factor binding protein
  • Ovarian granulosa cell

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Ovarian granulosa cell-derived insulin-like growth factor (IGF) binding proteins : Release of low molecular weight, high-affinity IGF-selective species. / Adashi, Eli Y.; Resnick, Carol E.; Hernandez, Eleuterio R.; Hurwitz, Arye; Rosenfeld, Ronald (Ron).

In: Molecular and Cellular Endocrinology, Vol. 74, No. 3, 21.12.1990, p. 175-184.

Research output: Contribution to journalArticle

Adashi, Eli Y. ; Resnick, Carol E. ; Hernandez, Eleuterio R. ; Hurwitz, Arye ; Rosenfeld, Ronald (Ron). / Ovarian granulosa cell-derived insulin-like growth factor (IGF) binding proteins : Release of low molecular weight, high-affinity IGF-selective species. In: Molecular and Cellular Endocrinology. 1990 ; Vol. 74, No. 3. pp. 175-184.
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