Popow A, Nowak D, Malicka-Błaszkiewicz M
Faculty of Biotechnology, Department of Cell Pathology, University of Wrocław, Wrocław, Poland.
J Physiol Pharmacol. 2006 Nov;57 Suppl 7:111-23.
Our studies were focused on the isolation and characterization of highly motile fraction of cells from hepatoma Morris 5123 population. Cells that underwent several migration cycles through Matrigel - coated filters were successfully cultured. The invasion index was determined by means of Matrigel invasion assay. Statically significant increase in the value of invasion factor for selected cells variant in comparison to the parental population was observed. The considerable changes in the cell shape were followed by the reorganization of the actin cytoskeleton structure including a dense subcortical congestion in the distribution of beta -actin isoform. The visualization of this protein in tumor cells was performed by immunostaining and scanning fluorescent confocal microscopy. The results were confirmed by densitometry analysis of Western blots. In addition, the increased state of actin polymerization in the cytoplasmic fraction of selected cells was determined as measured by filamentous to monomeric (F:G) actin ratio. Concluding, the selected fraction of hepatoma Morris 5123 cells with higher invasion capacity was characterized by rounded shape, remarkable increase of beta -actin level, its submembrane concentration as well as with the increased state of actin polymerization with respect to parental cells population.
我们的研究集中于从肝癌Morris 5123细胞群体中分离出高迁移性的细胞组分并对其进行表征。成功培养了通过基质胶包被的滤膜经历多个迁移周期的细胞。侵袭指数通过基质胶侵袭试验来测定。与亲代群体相比,观察到所选细胞变体的侵袭因子值有统计学意义的显著增加。细胞形状的显著变化伴随着肌动蛋白细胞骨架结构的重组,包括β-肌动蛋白异构体分布中的致密皮质下聚集。通过免疫染色和扫描荧光共聚焦显微镜对肿瘤细胞中的这种蛋白质进行可视化。结果通过蛋白质印迹的光密度分析得到证实。此外,通过丝状肌动蛋白与单体肌动蛋白(F:G)比率测定,所选细胞细胞质组分中肌动蛋白聚合状态增加。总之,所选具有较高侵袭能力的肝癌Morris 5123细胞组分的特征是圆形形态、β-肌动蛋白水平显著增加、其在细胞膜下的聚集以及相对于亲代细胞群体肌动蛋白聚合状态增加。