A microarray analysis of age-related vascular smooth muscle cell polyploidy

Polyploidy is characterized by a greater than diploid content of DNA in a cell. Increases in the number of polyploid cells have been associated both with normal aging and with pathological conditions, including hypertension, in humans and rodents. We hypothesized that aortic vascular smooth muscle polyploidy can serve as a biomarker for aging and that the DNA content per se might affect expression of selective gene-specific transcripts. Our results demonstrated that approximately 60% of the vascular smooth muscle cells (VSMC) in the thoracic aorta of old Brown Norway rats are tetraploid compared to 8% in their younger counterparts. We found that tetraploidy increases exponentially over the life span of the animal. Aortas from caloric-restricted rats, which have a prolonged life span, display a consistently lower rate of tetraploidization compared to age-matched rats fed ad libitum. Microarray analysis and RT-PCR were performed with RNA from sorted diploid (2N) and tetraploid (4N) VSMC isolated from old rats to identify differentially expressed RNAs. Among those RNAs which show a reduction in tetraploid cells are those encoding the inflammation-associated proteins: insulin-like growth factor binding protein 6, macrophage inflammatory protein 2 precursor, macrophage galactose N-acetyl-galactoseamine specific lectin, complement component C4, developmentally regulated intestinal protein, and decorin. Conversely, the expression of p21 is elevated. Microarray analysis was also performed using RNA extracted from unfractionated aortic VSMC of young and old rats. Transcript levels of phospholipase A₂ group IIA, synuclein-γ, and fibronectin were found to be increased in the aging aorta whereas tropoelastin was decreased. Among these gene-specific transcripts, phospholipase A₂ and synuclein-γ were found to be upregulated in the 2N versus 4N cells of old aortas. This is of interest because the 2N cells have a higher proliferation potential and transgenic overexpression of phospholipase A2 was shown to increase VSMC atherogenesis. Both of these events are associated with aging. Based on these investigations, we speculate that increased tetraploidy could be an adaptive response of the aging aorta that imparts an anti-proliferative and anti-inflammatory capacity.