Human microvascular endothelial cells: Coordinate induction of morphologic differentiation and twofold extension of life span

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Abstract
Human microvascular endothelial cells (HMVEC) from adult adipose tissue were cultured in MCDB 131 medium supplemented with 10% fetal bovine serum. Under these conditions, HMVEC from seven different donors had finite proliferative life spans ranging from 14.5 to 23.5 population doublings (PD), with a mean life span of 19 PD. Addition of 10% conditioned medium from activated human leukocyte cultures (BM Condimed®) extended the life span of HMVEC to 31 to 41 PD, with a mean life span of 37 PD. At the end of lifespan, HMVEC cultures both with and without BM Condimed had very low labeling indices (0 to 5% [3H]thymidine labeled nuclei) and consisted of enlarged cells. However, the morphologies of the two types of HMVEC cultures were very different. Untreated HMVEC were polygonal endothelial cells that formed cobblestonelike monolayers with no cell overlapping. In contrast, BM Condimed-treated HMVEC were more elongated, less regularly shaped cells that were not strictly inhibited from overlapping. When old, these cells accumulated numerous vacuoles. The BM Condimed-treated HMVEC expressed Factor VIII antigen, which confirms their identity as endothelial cells. These cells reverted rapidly to the polygonal morphology of untreated HMVEC when they were removed from BM Condimed. Likewise, their proliferative capacity was not extended further once BM condimed was removed. These results suggest that HMVEC can exist in two distinct morphologic states in which the cells have different finite proliferative life spans.