Bikfalvi A, Dupuy E, Inyang A L, Fayein N, Leseche G, Courtois Y, Tobelem G
INSERM U 150, Hôpital Lariboisière, Paris, France.
Exp Cell Res. 1989 Mar;181(1):75-84. doi: 10.1016/0014-4827(89)90183-3.
The binding, internalization, and degradation of basic fibroblast growth factor (bFGF) in human omental microvascular endothelial cells (HOME cells) were investigated. Binding studies of bFGF in human endothelial cells have not yet been reported. Basic FGF bound to HOME cells (KD of 42.0 +/- 3.8 pM and 70,526 +/- 6121 binding sites/cell for the high-affinity sites, KD of 0.933 +/- 0.27 nM and 630,252 +/- 172,459 sites/cell for low-affinity binding sites). The number of low-affinity binding sites was found to be variable. Washing the cells with 2 M phosphate-buffered saline removed completely 125I-bFGF bound to low-affinity binding sites but decreased also the high-affinity binding. The majority of the surface-bound 125I-bFGF was removed by washing the cells with acetic acid buffer at pH 3. At 37 degrees C, 30% of the cell-associated 125I-bFGF became resistant to the acidic wash after 90 min, suggesting that this fraction of bound 125I-bFGF was internalized. At this temperature, degradation of the internalized ligand was followed after 1 h by the appearance of three major bands of 15,000, 10,000, and 8,000 Da and was inhibited by chloroquine. These results demonstrated two classes of binding sites for bFGF in HOME cells; the number of high-affinity binding sites being larger than the number reported for bovine capillary endothelial cells. The intracellular processing of bFGF in HOME cells seems to be different from that of heparin binding growth factor-1 in murine lung capillary endothelial cells and of eye-derived growth factor-1 in Chinese hamster fibroblasts.
研究了碱性成纤维细胞生长因子（bFGF）在人网膜微血管内皮细胞（HOME细胞）中的结合、内化和降解情况。尚未见关于bFGF在人内皮细胞中结合研究的报道。碱性成纤维细胞生长因子与HOME细胞结合（高亲和力位点的解离常数KD为42.0±3.8 pM，每个细胞有70,526±6121个结合位点；低亲和力结合位点的KD为0.933±0.27 nM，每个细胞有630,252±172,459个位点）。发现低亲和力结合位点的数量是可变的。用2M磷酸盐缓冲盐水洗涤细胞可完全去除与低亲和力结合位点结合的125I-bFGF，但也会降低高亲和力结合。用pH3的醋酸缓冲液洗涤细胞可去除大部分表面结合的125I-bFGF。在37℃下，90分钟后30%与细胞相关的125I-bFGF对酸性洗涤具有抗性，这表明这部分结合的125I-bFGF被内化。在此温度下，1小时后内化配体开始降解，出现分子量为15,000、10,000和8,000 Da的三条主要条带，且降解受到氯喹的抑制。这些结果表明HOME细胞中存在两类bFGF结合位点；高亲和力结合位点的数量多于报道的牛毛细血管内皮细胞中的数量。HOME细胞中bFGF的细胞内加工似乎不同于小鼠肺毛细血管内皮细胞中肝素结合生长因子-1以及中国仓鼠成纤维细胞中眼源性生长因子-1的加工。
