Hyaluronan induces migration of multidrug resistant murine lymphoma cell lines through Tiam1 activation.

GARCIA MG; ALANIZ LD; CORDO RUSSO R; SACCODOSSI N; ALVAREZ E; HAJOS SE

San Diego, CA, USA

Congreso; 99º Reunión Anual de la AACR (American Association for Cancer Research).; 2008

Migration and invasion of tumor cells are processes regulated by both adhesion mechanisms and proteolytic interactions with extracellular matrix components, such as hyaluronan (HA). In lymphoid tumors, the migratory mechanisms related with cytoskeletal organization seem to be more important than the proteolytic activity involved in cellular invasion. Tiam1, the guanine nucleotide exchange factor for Rac in vivo, has been found to be involved in cytoskeletal reorganization during tumor invasion. Therefore, the aim of this work was to determine the role of HA and Tiam1 in cellular migration. For this purpose we used murine lymphoma cell lines resistant to doxorubicin (LBR-D160), vincristine (LBR-V160), as well as a sensitive line (LBR-) that presented dissimilar invasive behavior in vivo. We observed that the three cell lines presented different migratory capacity towards HA gradient in vitro, being LBR-D160 the one that showed a significant increase in HA-dependent migration. However, the proteolytic activity of metalloproteinases (MMPs) determined by gelatin zymogram analysis revealed similar MMP-2 and MMP-9 activity in the three cell lines, suggesting that other mechanisms may be involved in such process. Western blot analyses of membrane extracts demonstrated that Tiam1 expression was higher in LBR-D160 than in the other cell lines. Moreover, treatment with HA induced an increase in its expression and activity, since translocation to the plasma membrane was observed. Confocal microscopy of the cell lines transfected with Tiam1-GFP, also showed translocation of Tiam1 to the membrane after HA treatment, demonstrating that HA would be able to induce Tiam1 activation. We also analyzed the role of PI3K/Akt pathway in Tiam1 activation. We observed by western blot that HA treatment activated this signaling pathway, increasing translocation of PI3K to the plasma membrane and inducing Akt phosphorylation. Inhibiton of this pathway by wortmannin modified the migratory capacity and also downregulated Tiam1 activity in these cell lines. In conclusion, we suggest that HA induces the migration of these lymphoma cell lines through Tiam1 activation and this effect is mediated by PI3K/Akt. Our data also demonstrated that differences in cell migration mediated by cytoskeletal reorganization seem to be more relevant in these tumor cells than proteolytic activity.in vivo, has been found to be involved in cytoskeletal reorganization during tumor invasion. Therefore, the aim of this work was to determine the role of HA and Tiam1 in cellular migration. For this purpose we used murine lymphoma cell lines resistant to doxorubicin (LBR-D160), vincristine (LBR-V160), as well as a sensitive line (LBR-) that presented dissimilar invasive behavior in vivo. We observed that the three cell lines presented different migratory capacity towards HA gradient in vitro, being LBR-D160 the one that showed a significant increase in HA-dependent migration. However, the proteolytic activity of metalloproteinases (MMPs) determined by gelatin zymogram analysis revealed similar MMP-2 and MMP-9 activity in the three cell lines, suggesting that other mechanisms may be involved in such process. Western blot analyses of membrane extracts demonstrated that Tiam1 expression was higher in LBR-D160 than in the other cell lines. Moreover, treatment with HA induced an increase in its expression and activity, since translocation to the plasma membrane was observed. Confocal microscopy of the cell lines transfected with Tiam1-GFP, also showed translocation of Tiam1 to the membrane after HA treatment, demonstrating that HA would be able to induce Tiam1 activation. We also analyzed the role of PI3K/Akt pathway in Tiam1 activation. We observed by western blot that HA treatment activated this signaling pathway, increasing translocation of PI3K to the plasma membrane and inducing Akt phosphorylation. Inhibiton of this pathway by wortmannin modified the migratory capacity and also downregulated Tiam1 activity in these cell lines. In conclusion, we suggest that HA induces the migration of these lymphoma cell lines through Tiam1 activation and this effect is mediated by PI3K/Akt. Our data also demonstrated that differences in cell migration mediated by cytoskeletal reorganization seem to be more relevant in these tumor cells than proteolytic activity.