The composition and distribution of Insulin-like Growth Factors (IGFs) and IGF-Binding Proteins (IGFBPs) in the serum of growth hormone receptor-deficient patients: Effects of IGF-I therapy on IGFBP-3

We have previously reported that adult GH receptor-deficient (GHRD) patients treated subcutaneously with recombinant human insulin-like growth factor (IGF)-I have increased serum IGF-I levels and decreased IGF-II levels, whereas IGF-binding protein-3 (IGFBP-3) levels were unchanged. To further investigate the effects of IGF-I administration upon the IGF-IGFBP axis in GHRD, we have examined: 1) the molecular distribution of IGF-I and IGF-II among the IGFBPs; 2) the composition and distribution of the IGFBPs, in particular IGFBP-3; and 3) the acid labile subunit (ALS). Serum samples from adult GHRD patients who were treated sc with recombinant human IGF-I (40 μg/kg, sc, twice a day) or from normal Ecuadorian adults were incubated with [¹²⁵I]IGF-II and subjected to neutral size-exclusion chromatography. The fractions were then subjected to Western ligand blot, Western immunoblot, IGFBP-3 RIA, and IGF RIAs. Serum of healthy adults incorporated [¹²⁵I]IGF-II into the 150- and 44-kilodalton (kDa) IGFBP region. The 150-kDa IGFBP region contained most of the circulating IGFBP-3, whereas the 44-kDa IGFBP region contained mainly IGFBP-1, 2, and 4. The 150-kDa region also contained a unique 28-kDa immunoreactive form of IGFBP-3, which was not detectable by Western ligand blot. Endogenous IGF-I and IGF-II were distributed equally in the 150- and 44-kDa IGFBP regions. Sera from GHRD patients mainly incorporated [¹²⁵I]IGF-II into the 44-kDa IGFBP region. Similar to control sera, the 150-kDa IGFBP region contained IGFBP-3, albeit at lower concentrations. The 44-kDa IGFBP region contained all IGFBPs including 50% of the total immunoreactive IGFBP-3. The two immunoreactive forms of IGFBP 3 (40- to 45-kDa doublet and 28-kDa band) were present in both IGFBP regions. The IGF size-distribution study revealed that the 150-kDa IGFBP region carried half of the circulating endogenous IGF-I, but only 30% of the IGF-II. Concentrations of the ALS were consistently low. Administration of IGF-I to GHRD patients was unable to increase concentrations of the molecular forms of IGFBP-3, correct the aberrant distribution of IGFs among the IGFBPs, or increase serum concentrations of ALS. In conclusion, we have found two forms of IGFBP-3 associated with IGF and ALS, which are capable of forming the ternary 150-kDa complex in healthy adult serum. The ratio of these two forms of IGFBP-3 and their distribution in serum was different between GHRD and control patients. These data may provide an explanation for the altered IGF distribution also observed in GHRD patients. Furthermore, we have found that the administration of IGF-I to patients with GHRD failed to elevate total IGF levels or to increase IGFBP-3 or ALS levels in the circulation.