Studies On GD1a Functions With Special Reference To Regulation Of MMP-9 And Lipid Raft Arrangement

Sialic acid-containing glycosphingolipids (gangliosides) have been implicated in the regulation of various biological phenomena such as cell growth, differentiation, apoptosis and metastasis. Previously, it has been demonstrated that GD1a suppresses the metastasis of FBJ cells. To elucidate the mechanism, research has been performed to clarify which molecules possibly related to tumor metastasis are regulated by GD1a, using GD1a-lacking FBJ-LL and FBJ-M5 cells, and GD1a-enriched FBJ-S1 and FBJ 5-30 cells. The present study has shown that MMP-9 expression as revealed by RT-PCR is regulated inversely by GD1a in such that MMP-9 was rich in LL and M5 cells, while less in S1 and 5-30 cells. This was further confirmed by exogenous addition of GD1a to M5 or LL cells, where both cell lines displayed a decrease in MMP-9 production in both dose and time dependent manners. Treatment of 5-30 cells with D-PDMP, an inhibitor of glucosylceramide synthesis, resulted in an increased MMP-9 expression. In a cell line derived from FBJ S1 cell where st3gal2 (sialyltransferase for the synthesis of GD1a) was knocked down by siRNA, enhanced synthesis of MMP-9 was observed. The activity of MMP-9 in conditioned medium of FBJ cells was analyzed by gelatin zymography, and the results were consistent with MMP-9 mRNA expression pattern. Taken together it is evident that MMP-9 of FBJ-cells is under the control of ganglioside GD1a. To elucidate the mechanism by which GD1a controls MMP-9 expression, several signaling pathway inhibitors were used. The present study demonstrated that GD1a-suppressed MMP-9 expression was not through MAPK-Erk pathway, thus far known as a major pathway that regulates MMP-9 expression. PI3K pathway is possibly involved in suppression of MMP-9 expression by GD1a in FBJ cell lines.