BINDING AND INTERNALIZATION OF HEPARIN BY VASCULAR SMOOTH-MUSCLE CELLS
BINDING AND INTERNALIZATION OF HEPARIN BY VASCULAR SMOOTH-MUSCLE CELLS
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DOI：
10.1002/jcp.1041240104
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发表时间：
1985-01-01
影响因子：
5.6
通讯作者：
KARNOVSKY, MJ
中科院分区：
文献类型：
--
作者：
CASTELLOT, JJ;WONG, K;KARNOVSKY, MJ
肝素在体内外均能特异性抑制血管平滑肌细胞增殖。检查肝素通过平滑肌细胞的结合和内化模式。在这些研究中，合成了保留其抗增殖能力的放射性标记和荧光素化（FITC）肝素探针。3 H-肝素通过特异性高亲和力结合位点（Kd = 10-9 M，100，000个结合位点/细胞）与这些细胞结合。约80%的结合到细胞表面的肝素在2小时内脱落到培养基中。留在细胞表面上的肝素以双相动力学内化; 50%的结合物质在2小时内被内化。在这种最初的快速摄取之后，速率显著减慢，剩余的肝素需要1-2天才能内化。使用视频图像增强荧光显微镜监测FITC肝素的结合和摄取。当平滑肌细胞暴露于FITC肝素在4.0 ℃。C，观察到弥漫性表面染色模式。将细胞加热至37度后。在37 ℃下15分钟后，在2分钟内观察到强荧光囊泡叠加在扩散表面染色上。C，细胞内可见大量大的点状小泡。2 h后，这些囊泡集中在核周区域。当考虑沿着特异性高亲和力结合位点的存在和3 H-肝素的初始快速摄取时，这种摄取模式表明肝素通过受体介导和其他内吞途径进入平滑肌细胞。
Heparin evidently specifically inhibits the proliferation of vascular smooth muscle cells in vivo and in vitro. The binding and mode of internalization of heparin by smooth muscle cells is examined. For these studies, radiolabeled and fluoresceinated (FITC) heparin probes were synthesized that retained their antiproliferative capacity. Binding of 3H-heparin to these cells occurs via specific, high-affinity binding sites (Kd = 10-9 M, 100,000 binding sites/cell). Approximately 80% of the heparin bound to the cell surface was shed into the culture medium within 2 h. Heparin that was left on the cell surface was internalized with biphasic kinetics; .apprx. 50% of the bound material was internalized within 2 h. After this initial rapid uptake, the rate slowed substantially, with the remaining heparin requiring 1-2 days to be internalized. Binding and uptake of FITC heparin was monitored using video image intensification fluorescence microscopy. When smooth muscle cells were exposed to FITC heparin at 4.degree. C, a diffuse surface staining pattern was observed. After warming the cells to 37.degree. C, intensely fluorescent vesicles were seen superimposed over the diffuse surface staining within 2 min. After 15 min at 37.degree. C, numerous large punctate vesicles were seen inside the cell. After 2 h, these vesicles had concentrated in the perinuclear region. This pattern of uptake, when considered along with the presence of specific, high-affinity binding sites and the initial rapid uptake of 3H-heparin, suggests that heparin enters smooth muscle cells by both receptor-mediated and other endocytic pathways.