Kibbey M C, Corcoran M L, Wahl L M, Kleinman H K
Laboratory of Developmental Biology, National Institutes of Health, Bethesda, Maryland 20892.
J Cell Physiol. 1994 Jul;160(1):185-93. doi: 10.1002/jcp.1041600121.
Angiogenesis has been investigated in vivo using subcutaneously injected reconstituted basement membrane (Matrigel) supplemented with angiogenic factors. Previously we found that the laminin-derived synthetic peptide containing SIKVAV (ser-ile-lys-val-ala-val) promoted angiogenesis in vivo. In parallel studies, it was observed that new vessel formation in response to this peptide occurred several days after basic fibroblast growth factor-induced angiogenesis. Since this delay suggested that SIKVAV-induced angiogenesis may be secondary to other events, we investigated here earlier time points to determine if both indirect and direct mechanisms of angiogenesis are involved. We found that neutrophils are continuously recruited to the SIKVAV-containing plugs between 4 hours to 3 days following the initial injection. By day 7, columns of endothelial cells begin to migrate into the plug and form small blood vessels. In contrast, neutropenic mice had a 62% reduction in SIKVAV-induced angiogenesis when compared to control mice. Freshly isolated neutrophils also degraded laminin, the major component of the basement membrane Matrigel. These cells also produced factors in response to SIKVAV peptide which induced proliferation of human umbilical vein endothelial cells relative to a control peptide. In vitro experiments utilizing human neutrophils demonstrated that these cells migrate to the SIKVAV peptide and possess a specific cell surface SIKVAV binding protein of approximately 56 kD. These data suggest that neutrophils are induced to migrate to the Matrigel plugs, at least in part, by SIKVAV peptide, where they may release their own angiogenic factors and degrade the matrix, thus physically facilitating cell migration and liberating additional angiogenic matrix fragments and/or cytokines.
已使用补充有血管生成因子的皮下注射重组基底膜（基质胶）在体内研究血管生成。此前我们发现，含有SIKVAV（丝氨酸 - 异亮氨酸 - 赖氨酸 - 缬氨酸 - 丙氨酸 - 缬氨酸）的层粘连蛋白衍生合成肽在体内促进血管生成。在平行研究中，观察到对该肽的新血管形成发生在碱性成纤维细胞生长因子诱导的血管生成几天后。由于这种延迟表明SIKVAV诱导的血管生成可能是其他事件的继发结果，我们在此研究了更早的时间点，以确定血管生成的间接和直接机制是否都参与其中。我们发现，在初次注射后的4小时至3天内，中性粒细胞持续募集到含有SIKVAV的栓子处。到第7天，内皮细胞柱开始迁移到栓子中并形成小血管。相比之下，与对照小鼠相比，中性粒细胞减少的小鼠中SIKVAV诱导的血管生成减少了62％。新鲜分离的中性粒细胞也降解层粘连蛋白，即基质胶基底膜的主要成分。这些细胞还产生响应SIKVAV肽的因子，相对于对照肽，该因子诱导人脐静脉内皮细胞增殖。利用人中性粒细胞的体外实验表明，这些细胞迁移至SIKVAV肽并拥有一种约56 kD的特异性细胞表面SIKVAV结合蛋白。这些数据表明，中性粒细胞至少部分地被SIKVAV肽诱导迁移至基质胶栓子处，在那里它们可能释放自身的血管生成因子并降解基质，从而在物理上促进细胞迁移并释放额外的血管生成基质片段和/或细胞因子。
