USE OF A HAMMERHEAD RIBOZYME WITH CATIONIC LIPOSOMES TO REDUCE LEUKOCYTE TYPE 12-LIPOXYGENASE EXPRESSION IN VASCULAR SMOOTH-MUSCLE

Chemically synthesized hammerhead-type ribozymes targeted against the porcine leukocyte-type 12-lipoxygenase (LO) have been developed and st udied. One chimeric ribozyme consists of DNA in the non-enzymatic port ions, and RNA in the enzymatic core as well as two phosphorothioate in ternucleotide linkages at 3' terminus. The second ribozyme consists of ribonucleotide sequences generated by in vitro transcription. In this chapter we describe methodologies to first analyze the ribozyme catal ytic activity in vitro by studying cleavage of target RNA in vitro. Th e subsequent sections will describe how to target the catalytic ribozy me and deliver it to porcine vascular smooth muscle cells (PVSMC) by a liposome-mediated method. Finally ways to evaluate its activity to in hibit expression of the 12-LO mRNA will be presented. These results de monstrate the feasibility of using ribozymes as novel candidates for t herapeutic agents to block specific gene expression in vascular cells.