Regulation of cell motility by glioma motility factor in gliomas : its biological significance in glioma invasion

恶性胶质瘤侵袭的分子机制尚不清楚。为了阐明肿瘤侵袭的生物信号，我们已经专注于胶质瘤细胞的细胞粘附和运动，并成功地纯化了胶质瘤运动因子（GMFs），其刺激生产细胞的趋化性和趋化动力学迁移。在本研究中，我们研究了GMF在胶质瘤侵袭中的可能作用，并分析了GMF的结构，以研究胶质瘤细胞运动的调节机制。胶质瘤细胞的侵袭性通过体外化学侵袭试验和体内肿瘤植入模型来评估。通过用赖氨酰内肽酶消化GMF进行GMF的序列分析。由于GMF与纤维连接蛋白（FN）具有部分同源性，因此用几种能识别FN不同结构域的抗体对GMF进行了免疫印迹分析，发现GMF有两种，即GMF-Ⅰ和GMF-Ⅱ，分子量分别为145 K和165 K。GMF具有FN的细胞结合和肝素结合结构域，并且还表达艾德（extra-domain）区，这在癌胚FN中可见，在GMF-I和-II之间以不同的方式表达。仅表达ED-B结构域的GMF-I显示出比同时表达ED-A和ED-B结构域的GMF-II强5至10倍的迁移活性。GMF诱导的胶质瘤细胞迁移与这些细胞的体外侵袭性相关，并且GMF加入胶质瘤细胞增强了细胞的侵袭性。此外，胶质瘤细胞与GMF共存时，其迁移能力明显高于胶质瘤细胞。这些结果表明，GMFs在胶质瘤侵袭中起重要作用，GMFs ED区的表达差异可能调节胶质瘤细胞的运动性。

The molecular mechanism of tumor invasion in malignant gliomas remains unknown. In order to clarify a biosignal of the tumor invasion, we have focused on cell adhesion and motility of glioma cells and succeeded in purification of glioma motility factors (GMFs) which stimulate both chemotactic and chemokinetic migration of the producer cells. In the present study, we have investigated a possible role of GMF in glioma invasion and analyzed the structure of GMF to examine a regulatory mechanism in glioma cell motility. Invasiveness of glioma cells was assessed by in vitro chemoinvasion assay and an in vivo tumor-implanted model. Sequence analysis of GMF was performed by digestion of the GMF with lysylendopeptidase. Since the GMF showed a partial homology to fibronectin (FN), immunoblot anayses of the GMF were carried out using several antibodies which recognized various domains of FN.There were two species of GMF,ie, GMF-I and GMF-II with a molecular mass of 145 K and 165 K,respectively. The GMFs had cell binding and heparin binding domains of FN and also expressed ED (extra-domain)-regions, which were seen in the oncofetal FN,in a different way between GMF-I and -II.GMF-I expressing only ED-B domain showed migration activity 5 to 10 times stronger than GMF-II expressing both ED-A and ED-B domains. The GMF-induced migration of glioma cells correlated well with in vitro invasiveness of these cells and addition of GMF to glioma cells enhanced the invasiveness of the cells. In addition, coexistance of GMF with glioma cells which were implanted in rat brains promoted the migration of the cells much more than glioma cell only. These results suggest that GMFs paly an important role in glioma invasion and a difference of expression in ED-regions of GMFs may regulate a cell motility in glioma cells.