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Inhibitory effects of antisense oligodeoxynucleotides targeting c-myc mRNA on smooth muscle cell proliferation and
1Cardiology Branch, National Heart, Lung, and Blood Institute, Bethesda, MD 20892.
Abstract:
Smooth muscle cell (SMC) proliferation and migration play pivotal roles in restenosis following angioplasty. c-myc is an immediate early response gene induced by various mitogens, and several lines of evidence derived from experiments using transformed or hematopoietic cell lines, or transgenic mice, suggest its protein product plays a role in numerous signaling transduction pathways, including those modulating cell division. We therefore reasoned that a strategy employing oligodeoxynucleotides (ODNs) complementary to c-myc mRNA (antisense ODNs) might be potent inhibitors of SMC proliferation and, perhaps, of SMC migration. To evaluate this concept, we tested several antisense ODNs targeted to c-myc mRNA (15- or 18-mer ODNs complementary to different c-myc mRNA sequences) by introducing them individually into the medium of cultured rat aortic SMCs. Phosphoroamidate-modified ODNs were employed to retard degradation. Antisense ODNs inhibited, in a concentration-dependent manner, SMC proliferation and SMC migration. Maximal inhibitory effect was 50% for proliferation and > 90% for migration. These effects were associated with decreased SMC expression of c-myc-encoded protein by Western immunoblotting and immunocytochemical staining. ODNs with the same nucleotides but a scrambled sequence caused no effect. These results indicate that the c-myc gene product is involved in the signal transduction pathways mediating SMC proliferation and migration in the in vitro model we employed. The results also suggest a potential role of antisense strategies designed to inhibit c-myc expression for the prevention of coronary restenosis.
Insights
Antisense oligodeoxynucleotides (ODNs) targeting the c-myc gene significantly inhibited smooth muscle cell proliferation and migration in vitro. This suggests a potential therapeutic strategy for preventing restenosis after angioplasty.
Area of Science:
- Molecular Biology
- Cell Biology
- Cardiovascular Research
Background:
- Smooth muscle cell (SMC) proliferation and migration are key factors in restenosis post-angioplasty.
- The c-myc gene, an immediate early response gene, is implicated in cell division and signaling pathways.
Purpose of the Study:
- To investigate the potential of antisense oligodeoxynucleotides (ODNs) to inhibit SMC proliferation and migration.
- To evaluate the role of c-myc in SMC signaling pathways relevant to restenosis.
Main Methods:
- Cultured rat aortic SMCs were treated with phosphoroamidate-modified antisense ODNs targeting c-myc mRNA.
- Inhibition of proliferation and migration was assessed in a dose-dependent manner.
- c-myc protein expression was analyzed using Western immunoblotting and immunocytochemistry.
Main Results:
- Antisense ODNs significantly inhibited SMC proliferation (up to 50%) and migration (over 90%) in a concentration-dependent manner.
- Reduced c-myc protein levels were observed in SMCs treated with antisense ODNs.
- Scrambled sequence ODNs did not produce any inhibitory effects, confirming target specificity.
Conclusions:
- The c-myc gene product is involved in the signaling pathways that mediate SMC proliferation and migration.
- Antisense strategies targeting c-myc expression show promise for preventing coronary restenosis.