A standardised method of culturing aortic explants, suitable for the study of factors affecting the phenotypic modulation, migration and proliferation of aortic smooth muscle cells.
The study of factors affecting phenotypic change and growth of aortic smooth muscle cells (SMC) typically involves either the isolation of SMC by enzymatic dissociation or observation of outgrowth of cells from primary explants of vascular tissue. Explants provide a system in which the growth of cells can be investigated without dissociating them totally from their normal environment and avoids some of the problems of variability associated with enzymatic digestion. We describe here a standardised method for the preparation of medial explants of arterial tissue using a McIlwain tissue chopper, which is both fast and reproducible. Measurement was made of the percentage of explants showing outgrowth and of the distance migrated by cells at various times after plating explants singly into wells of a 96-well plate. Using this method, by 12 days after explanting, more than 95% of explants from normal rabbit aorta had shown outgrowth, in contrast to only 50% of explants prepared using a scalpel blade. Explants from atherosclerotic rabbit aorta showed a shorter lag phase before outgrowth commenced than explants from normal rabbit aorta of a similar age, but the subsequent rate of growth was the same. In contrast, when explants of normal rabbit aorta were grown in hyperlipidic rabbit serum, the lag phase was the same as for normal serum, but the subsequent rate of growth was greater. Explants from normal rabbit aorta treated with heparin showed an increased lag phase but reduced rate of growth. Treatment with heparinase decreased the lag phase and increased the rate of growth as did elastase.
McMurray
H F
HF
Department of Pathology, University of Cambridge, U.K.
Parrott
D P
DP
Bowyer
D E
DE
D000818:Animals; D001011:Aorta / Q000166:cytology* / Q000502:physiology; D001161:Arteriosclerosis / Q000473:pathology / Q000503:physiopathology; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D006493:Heparin / Q000494:pharmacology; D019763:Heparin Lyase; D006801:Humans; D008055:Lipids / Q000097:blood; D008297:Male; D008722:Methods; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000502:physiology; D010196:Pancreatic Elastase / Q000494:pharmacology; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011817:Rabbits
D008055:Lipids; D006493:Heparin; D010196:Pancreatic Elastase; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase