Most of the circulating insulin-like growth factors-I and -II are present in the 150 kDa complex during human pregnancy

Abstract

ABSTRACT The aim of this study was to assess the molecular size distribution of insulin-like growth factors (IGFs) complexed to IGF-binding proteins (IGFBPs) in serum of non-pregnant and pregnant women. Sera were fractionated on a size-exclusion column at pH 7·4 to resolve different IGF–IGFBP complexes from unbound IGFs. Each fraction was further chromatographed on a size-exclusion column under acid conditions to dissociate IGFs from binding proteins prior to measurement of the IGF content of the complexes. The IGF-containing fractions from the acid column were assayed specifically for IGF-I and IGF-II. Serum pooled from pregnant women contained more IGF-I and IGF-II than serum from non-pregnant women. In both groups most of the IGF-I and IGF-II was found in a large (150 kDa) complex in serum and the remainder was present in complexes eluting in the 40–50 kDa region at pH 7·4. Some free IGF-I was also detected. Serum fractions collected by size-exclusion chromatography at pH 7·4 were also analysed by Western-ligand blotting to characterize the IGFBPs in the two main IGFBP size classes. In serum pooled from non-pregnant women, the 150 kDa IGF–IGFBP complexes contained a 40–50 kDa IGFBP doublet following Western-ligand blot analysis. This IGFBP co-migrated with a pure IGFBP-3 standard. IGFBPs of 34, 28 and 24 kDa all contributed to the 40–50 kDa IGF–IGFBP complexes of the pH 7·4 chromatograph. In serum pooled from pregnant women, the 40–50 kDa IGFBP-3 doublet and 34 kDa IGFBP were not evident in any fractions from the pH 7·4 column. Thus, although the amounts of IGF-I and IGF-II in 150 kDa IGF–IGFBP complexes were increased during late pregnancy, IGFBP-3 measured by Western-ligand blot analysis of this complex was greatly diminished. The large amount of IGF-I and IGF-II in 150 kDa complexes is strong evidence for the presence of IGFBP-3 in serum during pregnancy, because IGFs and IGFBP-3 are normally present in equimolar amounts in this complex. We suggest that during pregnancy in women, the IGFBP-3 in the 150 kDa complex becomes unstable and this may explain the failure of the Western-ligand blot to detect IGFBP-3. Journal of Endocrinology (1991) 131, 491–497