Investigation of MMP-2 and -9 in a highly invasive A431 tumor cell sub-line selected from a Boyden chamber assay
Kao, W.-T.; Lin, C.-Y.; Lee, L.-T.; Lee, P.-P.H.; Hung, C.-C.; Lin, Y.-S.; Chen, S.-H.; Ke, F.-C.; Hwang, J.-J.; Lee, M.-T.
Anticancer Research 28(4b): 2109-2120
2008
ISSN/ISBN: 0250-7005 PMID: 18751383 Document Number: 620509
In this study, highly invasive tumor cell lines (designated A431-I, -II and -III) derived from parental A431 tumor cells (A431-P) were isolated by three successive passages through a Boyden chamber with matrigel-coated membrane support. The invasive potential and the activity of secreted MMP-9 of each sub-line increased significantly compared to the A431-P (A431-III > A431-II > A431-I) as evidenced by the in vitro invasion assay, gelatin zymography and immunoblotting analyses. RT-PCR results also revealed the elevated expression of MMP-9 in A431-III. We further characterized the A431-III sub-line and found these cells exhibited a greater potential for attachment and spreading on fibronectin-coated substratum and for migration. The A431-III cells displayed multiple cytoplasmic extensions with focal contacts (vinculin-positive staining) during cell spreading within 30 min. We also noticed an increase in FAK phosphorylation, but no significant change in FAK protein level in the A431-III sub-line compared to those of A431-P cells. Together, these results demonstrate that the greater invasion potential exhibited by the highly invasive A431-III subline is likely attributed to an increased ability for attachment, spreading and migration, as well as increased MMP activity. Thus, A431-P and the highly invasive A431-III sub-line could be an excellent model for studying the mechanism of cancer metastasis.