Insulinlike growth factor action and production in adipocytes and endothelial cells from human adipose tissue

Primary cultures of microvascular endothelial cells and isolated adipocytes were prepared from human omental adipose tissue to study the potentially overlapping roles of insulin and insulinlike growth factors (IGFs) in human adipose tissue. To determine whether adipocytes contain type I IGF-receptors, binding experiments were carried out with ¹²⁵I-labeled IGF-I. At 16°C, saturation of specific binding to adipocytes was reached after 30 min and was 0.7% per 10⁶ cells. At 37°C, chloroquine produced an increase in cell-associated ¹²⁵I-IGF-I, suggesting that IGF-I is internalized and degraded in a manner analogous to insulin. In competition experiments, IGF-I competed for binding more effectively than rat IGF-II or insulin. The concentrations of IGF-I, rat IGF-II, and insulin necessary to displace 50% of ¹²⁵I-IGF-I binding were 2.5, 15, and 90 nM, respectively. In addition, a monoclonal antibody (α-IR₃) that has been shown to block the type I IGF receptor was used in competition binding experiments. The antibody also inhibited binding of ¹²⁵I-IGF-I to adipocytes. The biological effects of insulin and IGF-I were examined by studying adipocyte lipoprotein lipase (LPL). Insulin stimulated [¹⁴C]glucose incorporation into cellular lipid in a dose-dependent manner, with 50% effective concentration (EC₅₀) of 0.3 nM. However, an increase in LPL activity was observed only at a high insulin concentration, with an EC₅₀ of ~30 nM. In contrast, IGF-I stimulated a progressive increase in LPL, with an EC₅₀ of 3.2 nM. In addition, α-IR₃ blocked the stimulatory effect of IGF-I on adipocyte LPL. To study the possible local production of IGFs on adipose tissue, adipose-derived human microvascular endothelial cells were cultured. The conditioned medium from these cells contained significant quantities of immunoreactive IGF. After passing the conditioned medium through a Deltapak C₄ 100-Å column and eluting with an acetonitrile gradient, IGF was separated from binding protein, and most of the immunoreactive IGF in endothelial cell-conditioned medium consisted of IGF binding protein, with smaller contributions from IGF-II and IGF-I. Thus, in human adipose tissue, IGFs and IGF binding protein are produced by microvascular endothelial cells. In addition, type I IGF receptors are present on adipocytes, and LPL is stimulated through type I receptors.