Characterization in vitro and in vivo of hammerhead ribozymes directed against murine tumor necrosis factoralpha.

Abstract:

:A hammerhead ribozyme directed against murine TNFalpha (mTNFalpha) mRNA has been constructed. In vitro studies showed that this ribozyme was released from the parent molecule by flanking cis-acting hammerhead and hairpin ribozymes. This same anti-mTNFalpha ribozyme specifically cleaved both synthetically derived substrate RNA and mTNFalpha mRNA within a pool of total cellular RNA. Endogenous delivery of this anti-mTNFalpha ribozyme via the self-cleaving cassette reduced mTNFalpha mRNA and protein levels in lipopolysaccharide (LPS)-stimulated, stably transfected murine macrophage RAW 264.7 cells. When complexed to liposomes and exogenously delivered to mouse peritoneal macrophages, the same ribozyme, with and without the cis-acting ribozymes, reduced mTNFalpha protein levels. However, an irrelevant ribozyme delivered in an identical fashion was also effective at reducing mTNFalpha protein levels. These data suggest that anti-mTNFalpha ribozymes can be constructed which efficiently cleave mTNFalpha mRNA, but irrelevant RNA/liposome complexes also effectively limit TNFalpha mRNA expression and can mimic functional ribozyme activity under in vitro conditions.

authors

MacKay SL,Tannahill CL,Auffenberg T,Ksontini R,Copeland EM 3rd,Moldawer LL

doi

10.1006/bbrc.1999.0927

subject

Has Abstract

pub_date

1999-07-05 00:00:00

pages

390-7

issue

2

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(99)90927-0

journal_volume

260

pub_type

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