NFATc1 Mediates Vascular Endothelial Growth Factor-induced Proliferation of Human Pulmonary Valve Endothelial Cells

Abstract

Mice deficient for the transcription factor NFATc1 fail to form pulmonary and aortic valves, a defect reminiscent of some types of congenital human heart disease. We examined the mechanisms by which NFATc1 is activated and translocated to the nucleus in human pulmonary valve endothelial cells to gain a better understanding of its potential role(s) in post-natal valvular repair as well as valve development. Herein we demonstrate that activation of NFATc1 in human pulmonary valve endothelial cells is specific to vascular endothelial growth factor (VEGF) signaling through VEGF receptor 2. VEGF-induced NFATc1 nuclear translocation was inhibited by either cyclosporin A or a calcineurin-specific peptide inhibitor; these findings suggest that VEGF stimulates NFATc1 nuclear import in human pulmonary valve endothelial cells by a calcineurin-dependent mechanism. Importantly, both cyclosporin A and the calcineurin-specific peptide inhibitor reduced VEGF-induced human pulmonary valve endothelial cell proliferation, indicating a functional role for NFATc1 in endothelial growth. In contrast, VEGF-induced proliferation of human dermal microvascular and human umbilical vein endothelial cells was not sensitive to cyclosporin A. Finally, NFATc1 was detected in the endothelium of human pulmonary valve leaflets by immunohistochemistry. These results suggest VEGF-induced NFATc1 activation may be an important mechanism in cardiac valve maintenance and function by enhancing endothelial proliferation.

Vascular endothelial growth factor receptor signaling is required for cardiac valve formation in zebrafish

“…In summary, based on results presented here, as well as previous studies (Dor et al, 2001;Johnson et al, 2003), we speculate that by inducing nuclear translocation of NFATc1, zebrafish VEGF-R(s) regulate genes involved in proliferation, migration, and differentiation of endocardial cells in the valveforming region of the developing heart valve in a precisely regulated spatiotemporal manner. This speculation is consistent with recent findings reported by Chang and colleagues (Chang et al, 2004).…”

“…The role of VEGF in EMT is likely to be complex, and depend on spatial and temporal regulation given that, in two other studies of mouse embryonic valve development, VEGF was found to be a positive regulator of EMT (Enciso et al, 2003;Hallaq et al, 2004). In our laboratory, we showed a functional link between VEGF and the transcription factor NFATc1 in human post-natal valve endothelial cells (HPVEC) (Johnson et al, 2003). This prompted us to test directly the role of VEGF-R in cardiac valve development.…”

“…We demonstrated previously that VEGF-induced proliferation of HPVEC is mediated by VEGF-R2-induced NFATc1 translocation into the nucleus (Johnson et al, 2003). Therefore, we tested whether AAC 789 and PTK 787 could block VEGF-induced NFATc1 nuclear translocation in HPVEC.…”

“…HPVEC were isolated from human pulmonary valve leaflets as described (Johnson et al, 2003). To detect VEGF-induced nuclear-localization of NFATc1, HPVECs were fixed in 4% paraformaldehyde, permeabilized with 0.5% Triton X-100, and incubated with mouse anti-human NFATc1 monoclonal antibody (7A6 from Santa Cruz Biotechnology, Inc.) diluted 1:500 followed by FITC-conjugated anti-mouse IgG diluted 1:200.…”

Dual roles of Sema6D in cardiac morphogenesis through region-specific association of its receptor, Plexin-A1, with off-track and vascular endothelial growth factor receptor type 2

“…2B). This indicates that VEGFR2 is dispensable for Sema6D signaling during this stage of cardiac development, and rather suggests that the Plexin-A1-VEGFR2 complex may mediate the Sema6D activity at the later stage, particularly in endocardiac cushion formation, in which VEGF is known to be critically involved (Dor et al 2001;Johnson et al 2003).…”