Cyclin D1 overexpression permits the reproducible detection of senescent human vascular smooth muscle cells.
The senescence of mitotic cells is hypothesized to play a causal role in organismal aging. Cultures of normal human cells become senescent in vitro as a result of a continuous decline in the mitotic fraction from cell turnover. However, one potential barrier to the evaluation of the frequency and distribution of senescent cells in tissues is the absence of a panel of robust markers for the senescent state. In parallel with an analysis of the growth kinetics of human vascular smooth muscle cells, we have undertaken transcriptomic comparisons of early- and late-passage cultures of human vascular smooth muscle cells to identify potential markers that can distinguish between senescent and growth-competent cells. A wide range of genes are upregulated at senescence in human vascular smooth muscle cells. In particular, we have identified a 12-fold upregulation of expression in the cyclin D1 message, which is reflected in a concomitant upregulation at the protein level. Quantitative cytochemical analysis of senescent and growing vascular smooth muscle cells indicates that cyclin D1 reactivity is a considerably better marker of replicative senescence than senescence-associated beta-galactosidase activity. We have applied this new marker (in combination with Ki67, COMET, and TUNEL staining) to the study of human vascular smooth muscle cells treated with resveratrol, a putative anti-aging molecule known to have significant effects on cell growth.
School of Pharmacy and Biomolecular Sciences, Cockcroft Building, University of Brighton, Brighton, East Sussex, BN2 4GJ, UK.
Dominick G A
DG
Burton
Angela N
AN
Sheerin
Elizabeth L
EL
Ostler
Kaye
K
Smith
Peter J
PJ
Giles
Jill
J
Lowe
William
W
Rhys-Williams
David G
DG
Kipling
Richard G A
RG
Faragher
D000375:Aging / Q000502:physiology; D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology*; D020552:Comet Assay; D056741:Cyclin D; D016213:Cyclins / Q000096:biosynthesis*; D006801:Humans; D020287:In Situ Nick-End Labeling; D019394:Ki-67 Antigen / Q000096:biosynthesis; D008938:Mitosis / Q000502:physiology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D014158:Transcription, Genetic / Q000502:physiology*; D015854:Up-Regulation / Q000502:physiology; D001616:beta-Galactosidase / Q000096:biosynthesis
D015415:Biomarkers; D056741:Cyclin D; D016213:Cyclins; D019394:Ki-67 Antigen; D012333:RNA, Messenger; D001616:beta-Galactosidase