Shi Y, Hutchinson H G, Hall D J, Zalewski A
Division of Cardiology, Thomas Jefferson University, Philadelphia, PA 19107.
Circulation. 1993 Sep;88(3):1190-5. doi: 10.1161/01.cir.88.3.1190.
Proliferation of smooth muscle cells (SMCs) plays an important role in vascular pathobiology, being involved in the development of coronary restenosis and atherosclerosis. The activation of nuclear proto-oncogenes appears to be a final common pathway onto which various mitogenic signals coverage. Accordingly, we attempted to determine whether the activation of the c-myc nuclear proto-oncogene is essential for human SMC proliferation and explored the possibility of inhibiting their growth using antisense oligonucleotides directed against c-myc messenger RNA (mRNA).
Proliferation of human SMCs was associated with an increase in c-myc mRNA expression after growth stimulation. Using 15-mer phosphorothioate oligonucleotides (oligomers), we tested their growth-inhibitory effect in SMCs in vitro. Antisense oligomers directed against the translation initiation region of the human c-myc gene exhibited a significant antiproliferative effect, whereas sense and mismatched oligomers did not inhibit the growth. The growth-inhibitory effect of c-myc antisense oligomers was dose dependent and preventable by an excess of sense oligomers. Furthermore, growth inhibition of SMCs treated with c-myc antisense oligomers was associated with a marked decrease in the c-myc mRNA level. Phosphorothioate oligomers remained stable in medium containing 20% serum and were detectable in SMCs as early as 1 hour after cell exposure. Intact oligomers rapidly accumulated intracellularly and persisted within human SMCs for at least 16 hours.
c-myc antisense oligomers reduced c-myc expression and produced a significant growth inhibition of human SMCs, indicating an important role of c-myc gene activation in the process of SMC proliferation. Furthermore, extracellular stability and rapid cellular uptake provide the basis for future studies assessing the therapeutic role of the c-myc antisense approach in reducing SMC proliferation in the process of vascular restenosis.
平滑肌细胞（SMC）的增殖在血管病理生物学中起重要作用，参与冠状动脉再狭窄和动脉粥样硬化的发展。核原癌基因的激活似乎是各种促有丝分裂信号汇聚的最终共同途径。因此，我们试图确定c-myc核原癌基因的激活对人SMC增殖是否至关重要，并探索使用针对c-myc信使核糖核酸（mRNA）的反义寡核苷酸抑制其生长的可能性。
生长刺激后人SMC的增殖与c-myc mRNA表达增加有关。我们使用15聚体硫代磷酸酯寡核苷酸（寡聚物）在体外测试了它们对SMC的生长抑制作用。针对人c-myc基因翻译起始区的反义寡聚物表现出显著的抗增殖作用，而正义和错配寡聚物则不抑制生长。c-myc反义寡聚物的生长抑制作用呈剂量依赖性，且可被过量的正义寡聚物阻断。此外，用c-myc反义寡聚物处理的SMC的生长抑制与c-myc mRNA水平的显著降低有关。硫代磷酸酯寡聚物在含有20%血清的培养基中保持稳定，细胞暴露后1小时即可在SMC中检测到。完整的寡聚物迅速在细胞内积累，并在人SMC中持续至少16小时。
c-myc反义寡聚物降低了c-myc表达，并对人SMC产生了显著的生长抑制作用，表明c-myc基因激活在SMC增殖过程中起重要作用。此外，细胞外稳定性和快速的细胞摄取为未来评估c-myc反义方法在减少血管再狭窄过程中SMC增殖的治疗作用的研究提供了基础。
