Production of IGF-binding proteins by vascular endothelial cells

We recently characterized three FGF-binding proteins (FGF-BPs) which are soluble forms of the extracellular domains of the high affinity FGF receptors (Hanneken, A. M., W. Ying, N. Ling, and A. Baird. Proc. Natl. Acad. Sci. USA. 1994. 91:9170-9174). These proteins circulate in blood and have been proposed to modulate the biological activity of the FGF family of proteins. Immunohistochemical studies now demonstrate that these soluble, truncated FGF receptors are also present in the basement membranes of retinal vascular endothelial cells. These immunoreactive proteins can be detected with antibodies raised to the extracellular domain of FGFR-1 but not with antibodies raised to either the juxtamembrane domain or the cytoplasmic domain of FGFR-1. Western blotting of human retinal extracts with the antibody raised to the extracellular domain of FGFR-1 detects specific, low molecular mass proteins at 85 kD and 55 kD, corresponding in size to the FGF-BPs, which are not detected with antibodies against the cytoplasmic domain of the receptor. The interaction of this receptor with the extracellular matrix is not dependent on the presence of FGF-2. Immunoreactive receptors are still detected in vascular basement membranes after the removal of FGF-2 with heparitinase. In addition, the recombinant extracellular domain of FGFR-1 continues to bind to corneal endothelial cell matrix after endogenous FGF-2 has been removed with 2 M NaCl. Acid treatment, which has been shown to disrupt protein interactions with the extracellular matrix, leads to a significant reduction in the presence of the matrix form of the FGF receptor. This loss can be restored with exogenous incubations of the recombinant extracellular domain of FGFR-1. This report is the first demonstration that a truncated form of a high affinity growth factor receptor can be localized to the extracellular matrix. These findings add to the list of binding proteins associated with the extracellular matrix (IGFBP-5) and suggest a potentially new regulatory mechanism for controlling the biological availability of FGF, and other peptide growth factors, in the extracellular matrix.

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Distribution of Chimeric IGF Binding Protein (IGFBP)-3 and IGFBP-4 in the Rat Heart: Importance of C-Terminal Basic Region

Endocrinology ◽

10.1210/endo.142.9.8353 ◽

2001 ◽

Vol 142 (9) ◽

pp. 3749-3755 ◽

Cited By ~ 7

Author(s):

K. L. Knudtson ◽

M. Boes ◽

A. Sandra ◽

B. L. Dake ◽

B. A. Booth ◽

...

Keyword(s):

Endothelial Cells ◽

Cardiac Muscle ◽

Rat Heart ◽

Binding Proteins ◽

Binding Protein ◽

Homologous Region ◽

Microvascular Endothelium ◽

Igf Binding Proteins ◽

Igf Binding ◽

Igf Binding Protein

Abstract IGF binding proteins-3 and -4, whether given in the perfused rat heart or given iv in the intact animal, cross the microvascular endothelium of the heart and distribute in subendothelial tissues. IGF binding protein-3, like IGF-I/II, localizes in cardiac muscle, with lesser concentrations in CT elements. In contrast, IGFBP-4 preferentially localizes in CT. In this study, chimeric IGF binding proteins were prepared in which a basic 20-amino-acid C-terminal region of IGF binding protein-3 was switched with the homologous region of IGF binding protein-4, and vice-versa, to create IGF binding protein-34 and IGF binding protein-43. Perfused IGF binding protein-34 behaved like IGF binding protein-4, localizing in connective tissue elements, whereas IGF binding protein-43 now localized in cardiac muscle at concentrations identical to perfused IGF binding protein-3. To determine whether these small mutations altered the affinity of the chimera for cells, the ability of 125I-IGF binding protein-34 and 125I-IGF binding protein-43 to bind to microvascular endothelial cells was determined and compared with IGF binding protein-3. IGF binding protein-34 retained 15% of the binding capacity of IGF binding protein-3, whereas IGF binding protein-43 bound to microvessel endothelial cells with higher affinity and greater total binding than that of IGF binding protein-3. We conclude that small changes in the C-terminal basic domain of IGF binding protein-3 and the corresponding region of IGF binding protein-4 can alter their affinity for cultured cells and influence their tissue distribution in the rat heart.

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