Characterization of insulin and IGF-1 receptor binding in equine liver and lamellar tissue: implications for endocrinopathic laminitis

Although it is well established that equine laminitis can be triggered by extreme hyperinsulinemia, the mechanism of insulin action is not known. High concentrations of insulin lead to separation of the weight-bearing apparatus from the hoof wall and are associated with an increased cycle of cell death and proliferation in the lamellae. Gene expression and immunohistochemistry studies have indicated that the lamellae are sparsely populated with insulin receptors, whereas IGF-1 receptors (IGF-1R) are abundant, suggesting that the action of insulin may be mediated by insulin binding to the IGF-1R. To investigate this possibility, cell membrane fragments containing IGF-1R were extracted from the livers of 6 horses and the lamellae of >50 horses euthanized for nonresearch purposes at an abattoir. Radioligand-binding studies using ¹²⁵I-IGF-1 and ¹²⁵I-insulin confirmed an abundance of high-affinity IGF-1R in the liver (K_D 0.11 nM, B_max 223 fmol/mg protein) and lamellae (K_D 0.16 nM, B_max 243 fmol/mg protein). However, the affinity of insulin for binding to the lamellar IGF-1R (K_i 934 nM) was >5,800 fold less than that of IGF-1, suggesting that insulin is unlikely to bind to equine IGF-1R at physiological concentrations. Although insulin receptors could be detected in the liver (K_D 0.48 nM, B_max 123 fmol/mg protein), they were barely detectable in lamellae (estimated B_max 14 fmol/mg protein). There was no evidence to support the presence of insulin/IGF-1 hybrid receptors in either tissue. These findings suggest that insulin does not act directly through IGF-1 receptors and that an alternative theory is required to explain the mechanism of insulin action in laminitis.