A systematic analysis of 40 random genes in cultured vascular smooth muscle subtypes reveals a heterogeneity of gene expression and identifies the tight junction gene zonula occludens 2 as a marker of epithelioid "pup" smooth muscle cells and a participant in carotid neointimal formation

An accumulation of evidence suggests that vascular smooth muscle is compose d of cell subpopulations with distinct patterns of gene expression. Much of this evidence has come from serendipitous discoveries of genes marking phe notypically distinct aortic cultures derived from 12-day-old and 3-month-ol d rats. To identify more systematic differences, we isolated 40 genes at ra ndom from libraries of these 2 cultures and examined message expression pat terns. To determine consistency of differential expression, we measured mRN A levels in 4 sets of cultures in 6 phenotypically distinct aortic cell. cl ones and in balloon injured rat carotid arteries to determine the relevance of these differences in vitro to in vivo biology. The following 5 consiste ntly differentially expressed genes were identified in vitro: zonula occlud ens 2 (ZO-2); peroxisome proliferator-activated receptor delta (PPAR delta) ; secreted protein, acidic and rich in cysteine (SPARC); alpha 1(I)collagen ; and A2, an uncharacterized gene. We examined these 5 clones during caroti d artery injury and an inconsistently differentially expressed clone Krox-2 4 because, as an early response transcription factor, it could be involved in the injury response. PPAR delta, A2, and Krox-24 mRNAs were upregulated during the day after injury. ZO-2 and alpha 1(I)collagen messages were modu lated for up to a month, whereas SPARC message showed no consistent change. An analysis of ZO-2 and other tight junction genes indicates that tight ju nctions may play a role in smooth muscle biology. These data suggest that a systematic analysis of these libraries is likely to identify a very large number of differentially expressed genes. ZO-2 is particularly intriguing b oth because of this tight junction gene's pattern of prolonged over-express ion after injury and because of its potential role in determining the disti nctive epithelioid phenotype of smooth muscle cells identified in rat and o ther species.