Canfield A E, Schor A M
CRC Department of Medical Oncology, Christie Hospital NHS Trust, Manchester, UK.
J Cell Sci. 1995 Feb;108 ( Pt 2):797-809. doi: 10.1242/jcs.108.2.797.
Cultured endothelial cells undergo a reversible transition from a resting (cobblestone) phenotype to an angiogenic (sprouting) phenotype. This transition mimics the early events of angiogenesis. We have previously reported that the addition of exogenous xylosides inhibits endothelial cel sprouting and modifies the extracellular matrix (ECM) synthesised by the cells. We have now investigated whether endothelial sprouting is mediated by the nature of the extracellular matrix in contact with the cells. Accordingly, cell-free matrices deposited by bovine aortic endothelial cells (BAEC) were isolated. These matrices were produced under conditions in which the formation of the sprouting phenotype was permitted (controls) or inhibited (by the addition of exogenous xylosides). BAEC were then plated on these matrices and grown under conditions which promote sprouting. Sprouting proceeded normally on control matrices, whereas it was inhibited when the cells were grown on matrices deposited in the presence of xylosides. The composition of the permissive and inhibitory matrices was then analysed. Inhibitory matrices contained reduced levels of tenascin and increased levels of thrombospondin-1 by comparison to the permissive matrices. In contrast, no differences were detected in the relative levels of laminin. The roles of tenascin and thrombospondin-1 in endothelial sprouting were confirmed using specific antibodies. Immunolocalisation studies revealed the presence of both proteins in sprouting cells. Antibodies to tenascin inhibited the formation of sprouting cells on permissive matrices and on gelatin-coated dishes without affecting cell growth. Tenascin synthesis was increased when sprouting cells were present in the cultures. Antibodies to thrombospondin-1 stimulated sprouting on inhibitory matrices. These results suggest that the transition from a resting to a sprouting phenotype is promoted by tenascin and inhibited by thrombospondin-1.
培养的内皮细胞会经历从静止（鹅卵石样）表型到血管生成（发芽）表型的可逆转变。这种转变模拟了血管生成的早期事件。我们之前报道过，添加外源性木糖苷会抑制内皮细胞发芽，并改变细胞合成的细胞外基质（ECM）。我们现在研究了内皮细胞发芽是否由与细胞接触的细胞外基质的性质介导。因此，分离了牛主动脉内皮细胞（BAEC）沉积的无细胞基质。这些基质是在允许发芽表型形成的条件下（对照）或被抑制的条件下（通过添加外源性木糖苷）产生的。然后将BAEC接种在这些基质上，并在促进发芽的条件下培养。在对照基质上发芽正常进行，而当细胞在存在木糖苷的情况下沉积的基质上生长时，发芽受到抑制。然后分析了允许性和抑制性基质的组成。与允许性基质相比，抑制性基质中腱生蛋白水平降低，血小板反应蛋白-1水平升高。相比之下，层粘连蛋白的相对水平未检测到差异。使用特异性抗体证实了腱生蛋白和血小板反应蛋白-1在内皮细胞发芽中的作用。免疫定位研究揭示了这两种蛋白在发芽细胞中的存在。针对腱生蛋白的抗体抑制了允许性基质和明胶包被培养皿上发芽细胞的形成，而不影响细胞生长。当培养物中存在发芽细胞时，腱生蛋白的合成增加。针对血小板反应蛋白-1的抗体刺激了抑制性基质上的发芽。这些结果表明，从静止表型到发芽表型的转变由腱生蛋白促进，而由血小板反应蛋白-1抑制。
