Deregulated expression of the c-myc oncogene abolishes inhibition of proliferation of rat vascular smooth muscle cells by serum reduction, interferon-gamma, heparin, and cyclic nucleotide analogues and induces apoptosis.
We have investigated the requirement for c-myc downregulation in the growth arrest of vascular smooth muscle cells (VSMCs). Rat VSMCs were infected with a retrovirus vector directing constitutive expression of either the complete human c-Myc protein (VSM-myc cells) or the c-Myc deletion mutant D106-143, which is inactive in cotransformation and autosuppression assays (VSM-D106-143 myc cells). Clones of transfected VSM-myc cells were isolated that constitutively expressed a range of levels of c-Myc protein from that observed in normal proliferating VSMCs to approximately seven times normal. The growth rates of these clones and their responses to growth inhibitors were then assessed. VSM-myc clones possessed a shorter mean intermitotic time than normal cells, which was inversely correlated (P < .05) with the level of c-Myc protein expressed. VSM-myc cells also expressed lower levels of alpha-smooth muscle actin mRNA and protein and exhibited an altered morphology. The proliferation of normal VSMCs and VSM-D106-143 myc cells was inhibited by serum reduction (0.5% fetal calf serum) and also by treatment with interferon-gamma (100 IU/mL), heparin (50 micrograms/mL), 8-bromo-cAMP (0.1 mmol/L), or 8-bromo-cGMP (0.1 mmol/L). In contrast, proliferation of VSM-myc cells was not inhibited by any of these agents, even if present at 10-fold higher concentrations. However, approximately 75% of VSM-myc cells expressing levels of c-Myc protein seen in normal proliferating VSMCs underwent apoptosis after 4 days of serum reduction or treatment with interferon-gamma. The results show that constitutive c-myc expression induces continuous cell proliferation, reduction in alpha-smooth muscle actin expression and apoptosis in VSMCs. We conclude that downregulation of c-myc is a prerequisite for growth arrest and subsequent survival of VSMCs. Conversely, deregulated c-myc expression may be important in the proliferation and death of VSMCs--characteristics of the pathogenesis of atherosclerosis.
Bennett
M R
MR
Department of Cardiology, University of Wales College of Medicine, Heath Park, Cardiff, UK.
Evan
G I
GI
Newby
A C
AC
D000199:Actins / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001769:Blood / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology*; D016259:Genes, myc; D006493:Heparin / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000378:metabolism; D009712:Nucleotides, Cyclic / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D012192:Retroviridae Infections / Q000235:genetics / Q000473:pathology
D000199:Actins; D009712:Nucleotides, Cyclic; D007371:Interferon-gamma; D006493:Heparin