Further characterization of a new in vitro angiogenesis model under serum free culture conditions; suppression of endothelial cell differentiation by serum.
We studied the regulation of the extracellular matrix in the recently established murine vascular endothelial cell clones, F-2 or F-2C. F-2 cells constitutively show a cobblestone growth pattern under serum supplemented culture conditions, whereas F-2C cells undergo spontaneous histodifferentiation to form tubular structures in chemically defined media. We reported that the tubulogenesis induced by F-2C might relate to the heavy deposition of a 'basement membrane analog' as a subendothelial matrix (SEM). We further characterized the regulation of extracellular matrix (ECM) metabolism in these cell clones, in terms of gelatinase expression, ECM degradation and the effects of serum. F-2C cells in culture medium containing 1% serum did not undergo tubulogenesis but presented cobblestone growth. Zymography analysis showed that both F-2 and F-2C cells express two gelatinase (72 and 92 kDa). However, F-2 cells mainly expressed the former and faintly the latter, whereas F-2C mainly expressed the latter. Proteolysis studies showed that the spent media conditioned by F-2C cells partially cleaved type IV collagen and completely degraded type V collagen. The cleavage of type V collagen was suppressed by the addition of serum, whereas that of type IV collagen was not. The proteolysis of laminin and fibronectin by the conditioned medium was not observed. Serum-supplemented F-2 or F-2C cultures markedly suppressed SEM deposition. These results indicated that F-2C cells under serum free culture conditions not only present a simple and useful in vitro model with which to study the dynamic processes of proteolysis and ECM metabolism during the sequential phases of angiogenesis, but is also useful for analyzing the serum effects on angiogenesis (AG).
Chen
C S
CS
Department of Dermatology, Faculty of Medicine, Kyoto University, Japan.
Toda
K
K
Fujii
K
K
Imamura
S
S
D000818:Animals; D001790:Blood Physiological Phenomena; D002454:Cell Differentiation / Q000502:physiology; D002999:Clone Cells; D016895:Culture Media, Serum-Free; D004730:Endothelium, Vascular / Q000166:cytology*; D005109:Extracellular Matrix / Q000378:metabolism; D018093:Gelatinases / Q000378:metabolism; D051379:Mice; D018919:Neovascularization, Physiologic / Q000502:physiology*
D016895:Culture Media, Serum-Free; D018093:Gelatinases