RESULTS: We applied a loss-of-function genetic approach employing lymphatic endothelial conditional mutations in
heparan sulfate biosynthesis to study the effects on
tumor-lymphatic trafficking and
lymph node metastasis. Lymphatic endothelial deficiency in N-deacetylase/N-
sulfotransferase-1 (Ndst1), a key
enzyme involved in sulfating nascent
heparan sulfate chains, resulted in altered
lymph node metastasis in
tumor-bearing gene targeted mice. This occurred in mice harboring either a pan-endothelial Ndst1 mutation or an inducible lymphatic-endothelial specific mutation in Ndst1. In addition to a marked reduction in
tumor metastases to the regional lymph nodes in mutant mice, specific immuno-localization of CCL21, a
heparin-binding
chemokine known to regulate leukocyte and possibly
tumor-cell traffic, showed a marked reduction in its ability to associate with
tumor cells in mutant lymph nodes. In vitro modified chemotaxis studies targeting
heparan sulfate biosynthesis in lymphatic endothelial cells revealed that
heparan sulfate secreted by lymphatic endothelium is required for CCL21-dependent directional migration of murine as well as human lung
carcinoma cells toward the targeted lymphatic endothelium. Lymphatic
heparan sulfate was also required for binding of CCL21 to its receptor CCR7 on
tumor cells as well as the activation of migration signaling pathways in
tumor cells exposed to lymphatic
conditioned medium. Finally, lymphatic cell-surface
heparan sulfate facilitated receptor-dependent binding and concentration of CCL21 on the lymphatic endothelium, thereby serving as a mechanism to generate lymphatic
chemokine gradients.
CONCLUSIONS: