Abstract
Objective:The objective of the present study was to establish a rat lymphatic endothelial cell line and then to investigate the morphological and immunohistochemical properties of the cells. Methods:The lymphatic endothelial cells of rat thoracic ducts were isolated enzymatically by trypsin digestion and were cultured in endothelium growth medium (EGM)-2 in an atmosphere of low oxygen (5% O2, 5% CO2, and 90% N2) or high oxygen (21% O2, 5% CO2, and 74% N2). Results:The number of the cells cultured in the low-oxygen atmosphere was significantly larger than that obtained in the high-oxygen atmosphere. The cultured cells in the low-oxygen atmosphere showed a monolayer with uniform cobblestone appearance, suggesting the morphological properties of endothelial cells. Factor VIII-related antigen and cell surface carbohydrates (i.e., D-galactose α and D-N-acetylgalactosamine α) were found on the lymphatic cultured cells. The phagocytosis of 1,1-diocadecyl1-3,3,3′,3′-tetramethylindocarbocyanine perchlorate-labeled acetylated low-density lipoprotein also was observed in the cultured cells. The cytoskeleton protein F-actin was located on the plasma membrane of the cultured cells as circumferential thin bundles and in the cytoplasma as filamentous bundles. Conclusions:The present study indicates that the choice of EGM-2 as a culture medium and the hypoxic atmosphere (∼5%) enabled us to establish rat lymphatic endothelial cell line. Microcirculation(2003) 10,127–131. doi:10.1038/sj.mn.7800181