Ganglioside-dependent adhesion events of human neuroblastoma cells regulated by the RGDS-dependent fibronectin receptor and proteoglycans.
Ganglioside-dependent adhesion events of human neuroblastoma cells regulated by the RGDS-dependent fibronectin receptor and proteoglycans.
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人神经母细胞瘤细胞的神经节苷脂依赖性粘附事件受 RGDS 依赖性纤连蛋白受体和蛋白聚糖调节。
DOI：
10.1016/0014-4827(88)90189-9
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发表时间：
1988
影响因子：
3.7
通讯作者：
Culp,LA
中科院分区：
文献类型：
--
作者：
Mugnai,G;Lewandowska,K;Choi,HU;Rosenberg,LC;Culp,LA
人神经母细胞瘤细胞(Platt和La-N1)在霍乱毒素B(CTB)提供的神经节苷脂GM1结合底物上黏附和延伸轴突。这些黏附反应类似于血浆纤维连接蛋白(PFN)，需要一种或多种细胞表面蛋白的调节[G.Mugnai和L.A.卡尔普(1987)Exp。单元格Res.169,328]。现在已经测试了两个PFN受体分子参与神经节苷脂GM1介导的CTB反应。为了检测细胞FN与其糖蛋白受体整合素的结合作用，在培养液中加入含有ArgGlyAspSer(RGDS)序列的可溶肽。它不抑制CTB的附着，但完全抑制突起的形成；相反，RGDS对PFN的附着或突起形成的抑制作用最小。一旦形成，CTB上的轴突就会对这种多肽产生抵抗力。为了检测细胞表面硫酸乙酰肝素蛋白多糖(HS-PG)的作用，采用了两种方法。首先，使用HS结合蛋白血小板因子-4(PF4)稀释底物上的CTB或PFN，或者将其添加到培养基中。用PF4稀释底物配基对CTB和PFN的附着均无影响。然而，CTB上的轴突形成很容易被抑制，而PFN上的轴突形成则被部分抑制；PF4的作用远远大于非结合白蛋白的类似稀释液。当Pf4加入细胞培养液中时，两种底物上的附着以及突起在Pfn上的生长都没有受到影响，这表明Pf4‘S抑制作用作为底物结合组分和介质携带组分是不同的。而在低浓度(1μg/ml)的培养液中，PF4对CTB的轴突形成有明显的抑制作用。第二种方法是将牛软骨硫酸皮胶蛋白多糖(DS-PG)加入到底物或培养基中，通过酶联免疫吸附试验证明DS-PG能与PFN结合，也能与CTB结合，或在底物或培养液中加入软骨硫酸软骨素/硫酸角蛋白多糖(CS/KS-PG)。在低浓度时，DS-PG促进CTB上的轴突形成，而在较高浓度下，DS-PG则完全抑制附着和轴突的形成。DS-PG可部分抑制PFN的贴壁，但对贴壁细胞突起的形成无影响。神经母细胞瘤细胞在一定程度上附着在只涂有DS-PG的底物上，这表明PGs的“受体”允许稳定的相互作用。这些数据表明，纤维连接蛋白的两种受体，整合素和硫酸乙酰肝素蛋白多糖，以顺式导向的方式与细胞表面的GM1相互作用，调节这些神经细胞对神经节苷脂结合底物的黏附反应。讨论了这种相互作用的两种可供选择的机制。
Human neuroblastoma cells (Platt and La-N1) adhere and extend neurites on a ganglioside GM1-binding substratum provided by cholera toxin B (CTB). These adhesive responses, similar to those on plasma fibronectin (pFN), require the mediation of one or more cell-surface proteins [G. Mugnai and L. A. Culp (1987)Exp. Cell Res.169, 328]. The involvement of two pFN receptor molecules in ganglioside GM1-mediated responses on CTB have now been tested. In order to test the role of cellular FN binding to its glycoprotein receptor integrin, a soluble peptide containing the ArgGlyAspSer (RGDS) sequence was added to the medium. It did not inhibit attachment on CTB but completely inhibited formation of neurites; in contrast, theRGDSpeptide minimally inhibited attachment or neurite formation on pFN. Once formed, neurites on CTB became resistant to the peptide. In order to test the role of cell-surface heparan sulfate proteoglycan (HS-PG), two approaches were used. First, the HS-binding protein platelet factor-4 (PF4) was used to dilute CTB or pFN on the substratum or, alternatively, added to the medium. Diluting the substratum ligand with PF4 had no effects on attachment on either CTB or pFN. However, neurite formation on CTB was readily inhibited and on pFN partially inhibited; the effects of PF4 were far greater than a similar dilution with nonbinding albumin. When PF4 was added to the medium of cells, attachment on either substratum was unaffected as was neurite outgrowth on pFN, revealing differences in PF4's inhibition as the substratum-bound or medium-borne component. In contrast, PF4 in the medium at low concentrations (1μg/ml) was highly inhibitory for neurite formation on CTB. The second approach utilized the addition of bovine cartilage dermatan sulfate proteoglycan (DS-PG), shown to bind to pFN as well as tosubstratum-boundCTB by ELISA, or cartilage chondroitin sulfate/keratan sulfate proteoglycan (CS/KS-PG) to the substratum or to the medium. At low concentrations, DS-PG but not CS/KS-PG actually stimulated neurite formation on CTB while at higher concentrations DS-PG completely inhibited attachment and neurite formation. While DS-PG partially inhibited attachment on pFN, it had no effect on neurite formation of the attached cells. Neuroblastoma cells adhered to some extent to substrata coated only with DS-PG, indicating “receptors” for PGs that permit stable interaction. These data provide evidence that two receptors for the fibronectins, integrin and heparan sulfate proteoglycan, interact with cell-surface GM1 in a “cis-oriented” manner to regulate adhesive responses of these neural cells to ganglioside-binding substrata. Two alternative mechanisms of this interaction are discussed.