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 studied. One chimeric ribozyme consists of DNA in the non-enzymatic portions, and RNA in the enzymatic core as well as two phosphorothioate internucleotide 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 catalytic activity in vitro by studying cleavage of target RNA in vitro. The subsequent sections will describe how to target the catalytic ribozyme and deliver it to porcine vascular smooth muscle cells (PVSMC) by a liposome-mediated method. Finally ways to evaluate its activity to inhibit expression of the 12-LO mRNA will be presented. These results demonstrate the feasibility of using ribozymes as novel candidates for therapeutic agents to block specific gene expression in vascular cells.
Gu
J L
JL
Department of Diabetes, Endocrinology and Metabolism, City of Hope Medical Center, Duarte, CA 91010, USA.
Nadler
J
J
Rossi
J
J
D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000187:drug effects*; D001483:Base Sequence; D005786:Gene Expression Regulation / Q000187:drug effects; D007962:Leukocytes / Q000737:chemistry / Q000201:enzymology*; D008081:Liposomes / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D016859:Lipoxygenase Inhibitors; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D016337:RNA, Catalytic / Q000138:chemical synthesis / Q000494:pharmacology*; D013552:Swine
D008081:Liposomes; D016859:Lipoxygenase Inhibitors; D016337:RNA, Catalytic; D001092:Arachidonate 12-Lipoxygenase