Downregulation of c-kit expression in human endothelial cells by inflammatory stimuli

Abstract

In recent studies we have shown that the expression of stem cell factor (SCF) in human endothelial cells is regulated by inflammatory processes. Gram-negative bacteria, interleukin-1 (IL-1), and lipopolysaccharide were able to upregulate the expression of SCF in human umbilical vein endothelial cells (HUVEC) (Blood 83:2836, 1994). Interestingly enough c-kit, the receptor of SCF, is coexpressed on HUVEC, suggesting an autoregulatory mechanism. To investigate the relation of c-kit and inflammatory processes we stimulated HUVEC with IL-1α and we established an in vitro model of inflammation. Binding experiments with 126I-SCF were performed to study the c-kit receptor expression on HUVEC. Scatchard analysis revealed both high-affinity receptors (K(d) ≃0.36 nmol/L) end low-affinity receptors (K(d) ≃2.9 nmol/L). Exposure to IL-1α led to a significant 50% reduction of c-kit high- affinity receptors, whereas the number of low-affinity receptors was not affected, in comparison to a control group of untreated HUVEC. Furthermore, using Northern blot analysis we studied the regulation c-kit mRNA expression in HUVEC after stimulation with IL-1α. Kinetic experiments showed a time- dependent downregulation of c-kit specific transcripts. In addition, we cocultured HUVEC with diverse bacterial strains. Experiments were performed over time with 1 x 108 bacteria/mL. Our data showed that, in contrary to the previously reported upregulation of SCF mRNA expression, stimulation with Yersinia enterocolitica or with Neisseria meningitidis led to a significant time-dependent downregulation of c-kit mRNA within 3 hours. These data indicate that inflammatory stimuli such as IL-1 or living bacteria activate a mechanism that downregulates c-kit receptor expression in human endothelial cells during the state of inflammation.