Actin cytoskeletal isoforms in human endothelial cells in vitro: alteration with cell passage

The microfilamentous actin component of the cytoskeleton is crucial to endothelial angiogenesis and vascular permeability. Differences in actin cytoskeletal profiles in cultured human endothelial cells were explored: when first isolated, both primary human umbilical vein endothelial cells (HUVEC) and primary human placental microvascular endothelial cells (HPMEC) expressed F-actin, but not beta-actin or alpha-smooth muscle actin. A similar endothelial actin profile was observed in cryo-sections of freshly delivered term umbilical cord and placenta. In subsequent cell culture, although the actin cytoskeleton of HUVEC remained unchanged, the actin profiles of HPMEC altered after the second passage with the induction of alpha-smooth muscle actin expression, which was intercellularly heterogeneous and increased to 20% at P4. This behaviour occurred in HPMEC monolayers cultured on a variety of extracellular matrices. Comparisons with a spontaneously immortalized human microvascular cell-line, HGTEN 21, revealed that in prolonged passage, both alpha-smooth muscle actin and beta-actin were expressed, whereas HPMEC at P4 showed a lower level of beta-actin expression. Therefore, in comparison with large vessels, microvascular cells are more likely to dedifferentiate. This may reflect the ability of microvascular cells to remodel according to changing requirements for new vessel formation. In conclusion, passage of human microvascular endothelial cells, but not of larger vessel endothelial cells, alters the expression of actin isoforms. This may be important in relation to comparisons of in vitro and in vivo vascular permeability; higher passage microvascular endothelial cells should thus be used with caution in such studies.