Effect of the Insulin-like Growth Factor (IGF) Axis on the Transport Properties of Endothelial and Epithelial Cells In Vitro

The overall objective of this research consists of two main parts: (1) provide evidence that autocrine production of IGF-I modulates tight junction permeability and (2) demonstrate the ability of IGFBPs to regulate IGF-I delivery across cell layers. To meet the first objective, parental and IGF-I secreting bovine mammary epithelial cells were tested for cell layer permeability, tight and adherens junction proteins, IGF-IR, and a downstream signaling components of IGF-IR. In comparison with parental cells, IGF-I secreting cells had high levels of IGF-IRs, but low levels of the junction components E-cadherin, b-catenin, and occludin. The differences in parental and IGF-I secreting cells was not due to extracellular stimuli since inclusion of IGF-I, IGFBP-3, or co-culture with SV40-IGF-I cells did not alter the barrier properties of parental cells, suggesting that intracrine signaling may alter cell connectivity. The second objective focused on exogenous rather than endogenous IGF-I and the role of IGFBPs and IGF-IRs in ligand transcytosis. Bovine aortic endothelial cells (BAECs) cultured on surfaces optimized to minimize paracellular transport were utilized to investigate the kinetics involved in the transport of insulin-like growth factor-I from the apical side of confluent monolayers to the basolateral side. Binding competitors were used to determine the role of the cell surface insulin-like growth factor-I receptor (IGF-IR) and cell surface insulin-like growth factor binding proteins (IGFBPs) in this transport process. Although IGFBPs initially retard delivery of IGF-I, using a computation model, this report shows that pulse durations of less than 6 hrs resulted in enhanced delivery of IGF-I in the presence of IGFBPs, above that for delivery in the absence of IGFBPs. In addition, the model was utilized to identify key parameters to target when developing engineered growth factors for the treatment of diseases. It is shown that the sorting factions and internalization rates are reasonable targets for the design of engineered growth factors. Since the sorting fractions are dictated by binding affinities in the acidic environment of the endosomes, it may be beneficial to design and analog of IGF-I that is more resistant to changes in pH, similar to those develop from epidermal growth factor.