An improved and reliable method for isolation of microvascular endothelial cells from human omentum.

OBJECTIVES: Despite an increasing research demand for human microvascular endothelial cells, isolation of primary endothelial cells from human tissue remains difficult. The omentum, a highly vascular visceral adipose tissue, could provide an excellent source of these cells.

METHODS: A reliable method to isolate HOMECs has been developed. It consists of initial enzymatic digestion (to deplete cell contaminants), followed by further digestion, selective filtration, and immunoselection using Dynabeads coated with CD31 antibody. Cultures were characterized for expression of endothelial cell markers and their ability to undergo VEGF-dependent in vitro tube structure formation.

RESULTS: Omental-derived cultures of microvascular endothelial cells were achieved with <5% contamination of other cell types. The endothelial origin of cells was confirmed by the constitutive expression of a range of vascular endothelial markers (CD31, CD105, vWF) and internalization of DiI-AcLDL. Furthermore, cultures were negative for lymphatic endothelial markers, underwent in vitro angiogenesis, and exhibited typical endothelial morphology.

CONCLUSIONS: This isolation method produces homogeneous HOMEC cultures that can be maintained in vitro for at least six passages without loss of cellular features characterizing endothelial cells.