The expression of apolipoprotein E in cultured neonatal rabbit aortic smooth muscle cells was examined. Northern blot analysis determined that there was a single RNA transcript of approximately 1.2 kb. Moreover, a polyclonal antibody against rabbit apolipoprotein E was prepared in a goat and used in immunoprecipitations to demonstrate that the cultured cells secreted apolipoprotein E into the media. A double band typical of apolipoprotein E migrated at apparent molecular masses of 37 and 39 kDa. Analysis of steady-state levels of apolipoprotein E mRNA demonstrated that expression increased as cell seeding density increased. When examined as a function of time in culture, there were two peaks of expression evident 1 day and 8 days after seeding. The addition of beta VLDL (beta-very low density lipoprotein) to smooth muscle cells increased both [3H]thymidine incorporation into DNA as well as cell number and these increases were accompanied by a decrease in the levels of apolipoprotein E mRNA in cells treated with the lipoprotein for 1 and 7 days. After incubation of the cultures with beta VLDL for 1 week, the cells were radiolabeled with [35S]methionine and the media was subjected to immunoprecipitation with anti-apolipoprotein E. The data revealed that the amount of apolipoprotein E secreted into the media decreased in the presence of beta VLDL. In summary, these results show that apolipoprotein E expression in aortic smooth muscle cells is regulated by cell density, time in culture, cell proliferative state, and beta VLDL addition. These observations may have relevance to the conditions that are known to accompany the development of the atherosclerotic lesion.