Transport of insulin-like growth factor-I across endothelial cell monolayers and its binding to the subendothelial matrix

J. Grulich-Henn; J. Ritter; S. Mesewinkel; U. Heinrich; M. Bettendorf; K. T. Preissner

doi:10.1055/s-2002-23488

Experimental and Clinical Endocrinology & Diabetes, Table of Contents

Exp Clin Endocrinol Diabetes 2002; 110(2): 67-73
DOI: 10.1055/s-2002-23488

Articles

Transport of insulin-like growth factor-I across endothelial cell monolayers and its binding to the subendothelial matrix

J. Grulich-Henn ¹ , J. Ritter ¹ , S. Mesewinkel ¹ , U. Heinrich ¹ , M. Bettendorf ¹ , K. T. Preissner ²

¹ Department of Pediatrics, University of Heidelberg, Germany
² Institute for Biochemistry, University of Giessen, Germany

Abstract

Summary

Cultured human umbilical vein endothelial cells (HUVEC) were used as a model to study transendothelial IGF-I transport, and its deposition into the extracellular matrix (ECM). Specific binding of ¹²⁵I-IGF-I to HUVEC monolayers was demonstrated, which was inhibited by aIR-3, a specific antibody directed against the IGF-I receptor. ECM-associated ¹²⁵I-IGF-I was approximately 10% of cell-bound IGF-I at 22 °C, and increased 4.5-fold at 37 °C, indicating that endothelial metabolism is required for the transport. However, neither monensin and cytochalasin B, both of which block endocytosis, nor aIR-3 did inhibit transport of ¹²⁵I-IGF-I into the ECM. In order to characterize IGF-I binding to the subendothelial ECM, HUVEC were removed nonenzymatically by treatment with Triton X-100 and ammonia. Specific, saturable binding of ¹²⁵I-IGF-I to the isolated ECM was observed, which was protease-sensitive. Antibodies directed against vitronectin inhibited IGF-I binding to the matrix by 35%, while antibodies directed against other ECM proteins had no significant influence on IGF-I binding. Using radioimmunoassays the IGF binding protein-2 was detected in the ECM, while IGFBP-1 and IGFBP-3 were below the detection limits. In order to evaluate functional aspects of IGF-I binding to the matrix, HUVEC were incubated under serum-free conditions in the absence and presence of IGF-I. Under serum-free conditions 48% of cells rounded up and started to detach after 2 hours incubation, while only 23% of the cells started to detach in the presence of IGF-I. These data indicate that IGF-I is transported via a paracelluar route across endothelial cells, and becomes bound to the subendothelial ECM. Vitronection seems to be involved in binding of IGF-I to the ECM. ECM-associated IGF-I might play a role in endothelial cell survival and stability.

Key words:

Insulin-like growth factor-I - Endothelial cells - Extracelluar matrix - Vitronectin

Full Text

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Dr. J. Grulich-Henn

University Children's Hospital Heidelberg

Im Neuenheimer Feld 150

D-69120 Heidelberg

Germany

Phone: + 49-6221-562311

Email: Juergen_Grulich-Henn@med.uni-heidelberg.de