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Ribozyme Oligonucleotides Against Transforming Growth Factor-&bgr; Inhibited Neointimal Formation After Vascular Injury in Rat Model: Potential Application of Ribozyme Strategy to Treat Cardiovascular Disease

BackgroundBecause the mechanisms of atherosclerosis or restenosis after angioplasty have been postulated to involve an increase in transforming growth factor (TGF)-&bgr;, a selective decrease in TGF-&bgr; may have therapeutic value. Thus, we used the ribozyme strategy to actively cleave the targeted gene to selectively inhibit TGF-&bgr; expression. Methods and ResultsWe constructed ribozyme oligonucleotides (ONs) targeted to the sequence of the TGF-&bgr; gene that shows 100% homology among the human, rat, and mouse species. The specificity of ribozyme against TGF-&bgr; gene was confirmed by selective inhibition of TGF-&bgr; mRNA in cultured vascular smooth muscle cells as well as balloon-injured blood vessels in vivo. Importantly, the marked decrease in TGF-&bgr; resulted in significant inhibition of neointimal formation after vascular injury in a rat carotid artery model (P <0.01), whereas DNA-based control ONs and mismatched ribozyme ONs did not have any inhibitory effect on neointimal formation. Inhibition of neointimal formation was accompanied by (1) a reduction in collagen synthesis and mRNA expression of collagen I and III and (2) a significant decrease in DNA synthesis as assessed by proliferating cell nuclear antigen staining. Moreover, we modified ribozyme ONs containing phosphorothioate DNA and RNA targeted to the TGF-&bgr; gene. Of importance, modified ribozyme ONs showed a further reduction in TGF-&bgr; expression. ConclusionsOverall, this study provides the first evidence that selective blockade of TGF-&bgr; resulted in inhibition of neointimal formation, accompanied by a reduction in collagen synthesis and DNA synthesis in a rat model. We anticipate that modification of ribozyme ON pharmacokinetics will facilitate the potential clinical utility of the ribozyme strategy.