Phenotypic stability and variation in cells of the porcine aorta: Collagen and elastin production

Abstract

The extracellular matrix of the developing vasculature varies in composition as a function of time and position. Cellular models of vascular biology and pathology depend on the assumption that stable phenotypic characteristics of vascular cells can be propagated through several generations of in vitro cultivation. We show that the positional and developmental heterogeneity of matrix phenotypes in the porcine aorta are expressed by explanted vascular smooth muscle cell (SMC) and adventitial cell populations for a limited number of passages. Elastin was expressed most highly by thoracic SMC while interstitial collagen production was usually maximal in abdominal segments. Parallel gradients of collagen types I, III and V, detected by specific ELISA assays, were expressed in early-passage SMC. Adventitial cell populations from the abdominal aorta of the neonatal pig accumulated significant levels of collagen, while these fibroblasts produced less than 10% of the elastin made by SMC. All cell populations expressed alpha-smooth muscle actin in vitro. Gradients of collagen and elastin expression were evident for no more than three passages, and direct outgrowth of cells without limited digestion of the matrix further reduced phenotypic stability. Variation and decline of the elastin phenotype could be due to hypermethylation of regulatory sequences in the elastin gene or trans-acting factors, but elastin production was dose-dependently stimulated to a similar extent (100%; 10 microM 5-azacytidine) in all segmental SMC populations at early (p1) and late (p3) passage. These data indicated that faithful reflection of in vivo SMC behavior was limited to a few population doublings, at least under standard culture conditions. Modification of the cellular environment by reducing serum factors, changing matrix, or adding mechanical stimulation may increase phenotypic stability.