Use of transwell system to analyze T lymphocyte migration across lymphatic endothelium

Abstract

Trafficking of lymphocytes through lymphatics and secondarylymphoid organs is crucial for immunity. While the process ofT lymphocyte migration across high endothelial venules (HEV)into lymph nodes (LN) is well characterized, relativelylittle is known about the mechanisms that regulate migrationof lymphocytes from tissues into afferent lymphatics. Severalgroups have attempted to develop in vitro methods toinvestigate lymphatic migration. Culture of freshly isolatedlymphatic endothelial cells has proven very difficult, andultimately no cell line produced has been shown to retain alymphatic lineage after multiple rounds of passage.TheC3H/HeJ mouse endothelial line SVEC4-10 is an SV-40transformant of endothelial cells isolated from the axillaryLN, and has been variously reported as lymphatic, HEV, orblood microvascular endothelium. We have characterized thiscell line via flow cytometry, RT-PCR, and gene arrayanalysis, and have determined that it represents a lymphaticendothelial cell lineage. We have also shown