A reevaluation of the response of human umbilical vein endothelial cells to certain growth factors

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Abstract
Human umbilical vein endothelial cells (HUV-EC) were isolated and maintained in pure culture on a fibronectin matrix with hypothalamic derived endothelial cell growth factor included in the culture medium. HUV-EC maintained under these conditons displayed only slight alterations in morphological appearance and continued to produce Factor VIII antigen. The cells showed an unaltered growth response to serum and added growth factors up to passage 16 (>30 population doublings). We found that epidermal growth factor (EGF) was potently mitogenic for HUV-EC, but only in the presence of endothelial cell growth factor. Also in contrast to a previous report, we were unable to demonstrate a potentiation of this response by human α-thrombin. Because of these discrepancies, we performed studies to determine if they might be explained by a difference in the interaction of our HUV-EC with EGF. In studies utilizing 125I-EGF as tracer probe, we determined that our HUV-EC have an EGF receptor number of 13,000 sites/cell with an apparent Kd∼︁4.0 × 10−9 M. In addition, receptor-bound 125I-EGF was rapidly internalized and degraded presumably by a lysosomally mediated pathway since degradation was complete to the amino acid level. These results are in agreement with those previously published and thus do not provide a basis on which to resolve discrepancies regarding the growth response of HUV-EC to various growth factors.