抄録
Biological and physico-chemical mechanisms of lymphatic metastasis of tumor cells and tumor-mediated lymphangiogenesis remain still unclear. And while many angiogenic molecules are widely studied, only few lymphangiogenic molecules except for VEGF-C/D are examined. To address molecular biology of lymphatic endothelial cells (LECs), firstly we should establish the cell lines isolated from small-sized lymph vessels. Thus we have attempted to develop a novel method for the isolation and culture of anatomically-defined LECs of rat thoracic ducts and afferent lymph vessels of iliac lymph nodes. Lymph vessels were dissected using a dissecting microscope. Then LECs were isolated enzymatically by intraluminal perfusion of trypsin. The perfusates were centrifuged and the cells were collected and harvested in hypoxic environment. The colonies were characterized with immunohistochemistry. Proliferation assay was also done for 96 hours. Using this technique, the LECs were able to isolate successfully and show VEGFR3. The LECs from the afferent vessels demonstrated higher proliferation activity compared with the LECs isolated from the thoracic ducts. Wound assay activity with VEGF-C stimulation is observed clearly both in the LECs from thoracic ducts and afferent vessels. Unlike casual techniques with the antigenic selection, the advantages of this method has enabled us to define directly and easily lymphatic endothelial cells, and then to offer suitable cultured lymphatic endothelial cells to study the mechanisms of lymphangiogenesis and lymphatic metastasis of tumor cells. [Jpn J Physiol 55 Suppl:S92 (2005)]
