We compared the distribution of DNA synthesis over the arterial tree of young normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR) with marginally elevated blood pressure. Six-week-old male SHR and WKY rats were therefore infused with 5-bromo-2'-deoxyuridine (BrdUrd) for 2 days and the nuclear incorporation of the thymidine analogue in the media of various arteries was determined by immunohistochemistry. In WKY rats, 2.5% of the arterial smooth muscle nuclei in elastic, muscular and resistance arteries incorporated BrdUrd. In SHR, DNA synthesis was more marked in large arteries than in resistance arteries. It was in addition significantly larger in the aorta, superior mesenteric, renal and femoral arteries of the SHR than in those of the WKY rats. However, nuclear incorporation of BrdUrd in vivo did not differ between SHR and WKY rats in aortic endothelium, carotid arterial smooth muscle, nor in mesenteric or renal resistance arteries. Between 6 and 20 weeks of age, the number of nuclear profiles per media cross-section did not increase in large arteries of WKY rats and SHR. During this period of time, however, carotid artery and thoracic aorta weight and DNA content increased. SHR large arteries gained more DNA than those of WKY rats. These data indicate that DNA synthesis is uniformly distributed over the arterial system in young WKY rats and that DNA synthesis is elevated in the smooth muscle of large arteries of 6-week-old SHR but not in their resistance arteries.
We evaluated effects of endothelium removal and of endothelium-derived vasoactive agents on DNA synthesis and contractility in the isolated arterial wall. The experiments were performed on renal artery segments that had been 1) isolated from adult rats, 2) suspended in tissue culture for 3 days in the continuous presence of fetal calf serum, and 3) exposed for the last 24 h to 5-bromo-2'-deoxyuridine. Nuclear incorporation of this thymidine analogue was visualized by immunohistochemistry and used as an index of DNA synthesis. Tissue culture did not alter relaxing responses to guanosine 3',5'-cyclic monophosphate (cGMP)-generating agents but promoted relaxing responses to adenosine 3',5'-cyclic monophosphate (cAMP)-generating agents. It stimulated DNA synthesis in the endothelium, media, and adventitia. Mechanical removal of endothelium increased intra-arterial DNA synthesis. This was most prominent in the media. In preparations with endothelium, indomethacin and methylene blue did not enhance DNA synthesis. In segments that had been denuded of endothelium, atrial natriuretic factor, forskolin, iloprost, and prostaglandin E2, but not isobutylmethylxanthine or sodium nitroprusside, significantly reduced intra-arterial DNA synthesis. These data indicate that endothelium removal promotes the mitogenic response of the arterial wall to exogenous growth factors. This cannot be attributed to inhibitory influences of endothelium-derived relaxing factor or prostaglandins released by the endothelium under basal conditions.
To evaluate long-term effects of contractile and mitogenic stimuli on the contractile reactivity of arterial smooth muscle, we measured the incorporation of the thymidine analogue 5-bromo-2'-deoxyuridine (BrdUrd) and mechanical responses in arterial segments that had been maintained in tissue culture. The experiments were performed on renal arteries that had been isolated from adult rats, chemically sympathectomized, mechanically denuded from endothelium and mounted under distension. Exposure of arterial segments for up to 3 weeks to culture medium supplemented with fetal calf serum resulted in the following consecutive changes: a strong acute contraction, selective pharmacological changes that included decreased contractile responses to phenylephrine and vasopressin and increased relaxing responses to isoproterenol, increased incorporation of BrdUrd, a progressive fall in contractile responses to all vasoconstrictor stimuli, and an increase in excitability. Serum-free medium resulted in a much smaller acute arterial contraction, induced less incorporation of BrdUrd, accelerated the occurrence of hyperexcitability, but did not affect early pharmacological changes or the subsequent fall in overall arterial contractility with tissue culture. Dialysis of the serum or addition of ketanserin abolished the contractile effect of serum but did not affect the incorporation of BrdUrd or the loss of contractility with tissue culture. Addition of serotonin to serum-free culture medium mimicked the contractile response to serum but not the stimulation of BrdUrd incorporation. These data indicate that tissue culture alters the properties of the arterial wall, that contraction does not underlie the proliferative response of arterial smooth muscle to serum-derived mitogens in vitro, and that stimulation of DNA synthesis does in itself not lead to selective changes in arterial contractility.
