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Effect of antisense mitogen-activated protein kinase oligonucleotides on rat vascular smooth muscle cell
1Laboratory of Molecular Pharmacology, Hu-nan Medical University, Changsha, China.
Aim:
To study the preventive effect of down-regulating mitogen-activated protein kinase (MAPK) on vascular smooth muscle cell (VSMC) proliferation.
Methods:
Cultured rat VSMC was pretreated with a phosphorothioate-protected 17-mer antisense MAPK oligodeoxynucleotides (ODN) directed against the initiation of translation sites of the p42- and p44-MAPK isoforms by liposomal transfection. A 17-mer sense and a random sequence MAPK ODN were used as control. After liposomal transfection, cells were exposed to epidermal growth factor (EGF) 1 nmol.L-1 for 10 min and then harvested in lysis buffer. MAPK activity was measured by Western blot and P-81 phosphocellulose filter papers method by using [gamma-32P] ATP and myelin basic protein as substrate. DNA synthesis was measured by [3H]thymidine incorporation.
Results:
Antisense ODN 0.2 mumol.L-1 reduced EGF-induced MAPK activities by 84%, and inhibited VSMC [3H]thymidine incorporation stimulated by EGF.
Conclusion:
A 17-mer MAPK antisense oligonucleotide directed against the initiation of translation sites of the p44- and p42-MAPK inhibited EGF-stimulated rat VSMC proliferation.
