Circulating Endothelial Cells Expressing the Angiogenic Transcription Factor Krüppel-Like Factor 4 are Decreased in Patients with Coronary Artery Disease.

OBJECTIVE: The zinc finger transcription factor KLF4 is known to control diverse EC functions. METHODS: The functional role of KLF4 for angiogenesis and its association with CAD was examined in HUVECs and human CECs. RESULTS: In two different angiogenesis assays, siRNA-mediated KLF4 downregulation impaired HUVEC sprouting and network formation. Conversely, KLF4 overexpression increased HUVEC sprouting and network formation. Similar findings were observed after incubation of HUVECs with CdM from KLF4 cDNA-transfected cells, suggesting a role of paracrine factors for mediating angiogenic KLF4 effects. In this regard, VEGF expression was increased in KLF4-overexpressing HUVECs, whereas its expression was reduced in HUVECs transfected with KLF4 siRNA. To examine the relevance of our in vitro findings for human endothelial dysfunction, we analyzed the expression of KLF4 in CECs of patients with stable CAD. Flow cytometry analyses revealed decreased numbers of KLF4-positive CECs in peripheral blood from CAD patients compared to healthy controls. CONCLUSIONS: Our findings suggest that KLF4 may represent a potential biomarker for EC dysfunction. In the future, (therapeutic) modulation of KLF4 may be useful in regulating EC function during vascular disease processes.

In vitro and in vivo effects of human monocytes and their subsets on new vessel formation.

OBJECTIVE: Human monocytes can be divided into CD16(-) monocytes and CD16(+) monocytes. Studies in mice suggested differential effects of monocyte subsets during new vessel formation. METHODS: The functional role of human monocyte subsets in neovascularization processes was investigated. For in vivo experiments, nude mice underwent unilateral hindlimb ischemia surgery before being injected with either total monocytes, CD16(-) monocytes or CD16(+) monocytes isolated from healthy individuals. RESULTS: In vitro, cytokine array analysis demonstrated that monocytes release numerous angiogenic cytokines, some of which were differentially expressed in monocyte subsets. Sprout length was enhanced in EC spheroids being cultured in conditioned medium obtained from total monocytes and, to a lesser extent, also in supernatants of CD16(-) monocytes. Laser Doppler perfusion imaging up to day 28 after surgery revealed a trend toward improved revascularization in mice treated with monocytes, but no significant differences between monocyte subsets. Histological analyses four weeks after surgery showed an increased arteriole size in mice having received CD16(+) monocytes, whereas the number of capillaries did not significantly differ between groups. CONCLUSIONS: Our findings suggest additive and differential effects of monocyte subsets during neovascularization processes, possibly due to an altered secretion of angiogenic factors and their paracrine capacity to stimulate new vessel formation.