P19arf controls proliferation in murine pulmonary artery smooth muscle cells

P19 arf is a tumor suppressor that functions, in part, by sequestering MDM2 thus allowing an increase in p53 expression. To determine whether p19 was important in controlling proliferation in pulmonary artery smooth muscle cells (PA SMC) in adult mice, we examined the effect of p19 deletion on PA SMC growth curves over time. We harvested the pulmonary artery from wild type (WT) and p19‐null mice (in which the p19 gene had been replaced with GFP [courtesy Dr. Zindy]) and examined proliferation within the first 35 days after digestion. Cells were plated at 100,000 per well, allowed to grow for 7 days, harvested, counted, and then re‐plated at 100,000. This was repeated through 35 days. No differences in cell number between p19 WT, heterozygous (Hz), or null cells were seen during the first 14 days. After 14 days, the p19 −/− PA SMC grew significantly faster than WT or Hz cells over each seven‐day period (10‐fold increase by day 28); over‐expression of p19 using a retrovirus prevented this proliferative advantage. Coincident with this increased growth was an up‐regulation of GFP expression indicating activation of the p19 promoter. The expression of Bmi‐1, a known inhibitor of p19 transcription, increased in WT PA SMC by day 21 whereas it was difficult to detect in p19 −/− cells. Conclusion: Loss of p19 leads to increased proliferation of PA SMC. This occurs despite a down‐regulation of the proto‐oncogene, Bmi‐1. Supported by NIH/NHLBI award, RO1 HL70273‐01