Methods to study lymphatic vessel integrins

The lymphatic system plays a key role in the drainage of fluids and proteins from tissues and in the trafficking of immune cells throughout the body. Comprised of a network of capillaries, collecting vessels, and lymph nodes, the lymphatic system plays a role in the metastasis of tumor cells to distant parts of the body. Tumors induce lymphangiogenesis, the growth of new lymphatic vessels, in the peritumoral space and also within tumors and lymph nodes. Tumor lymphangiogenesis has been shown to play a role in promoting tumor metastasis. As mediators of lymphatic endothelial cell adhesion, migration, and survival, integrins play key roles in the regulation of lymphangiogenesis. Recent studies indicate that select integrins promote lymphangiogenesis during development and disease and that inhibitors or loss of expression of these integrins can block lymphangiogenesis. In this report, we describe methods to isolate and culture murine and human lymphatic endothelial cells as well as methods to analyze the expression of integrins on these cells. We also show how to assess integrin-mediated adhesion, migration, and tube formation in vitro. We demonstrate how to evaluate integrin function during lymphangiogenesis in a variety of animal models in vivo. Additionally, we show how to study lymphangiogenesis using intravital microscopy.