In vitro biological activity of anti-C II TA hammerhead ribozyme--a novel approach for autoimmune diseases.

Abstract

This study investigated the feasibility of using an hammerhead ribozyme against C II TA, a major regulator of MHC II antigens, to repress the expression of MHC II molecules on Hela cells. A hammerhead ribozyme (Rz464) specific to 463-465 GUC triplet of C II TA and its target gene were transcribed, then mixed up and incubated in vitro. The cleavage products were analyzed by PAGE and silver-staining. Rz464 was then inserted into the pIRES2-EGFP vector (pRz464). Stable transfectants of Hela with pRz464 were tested for class II MHC induction by recombinant human interferon-gamma (IFN-gamma). mRNA of C II TA was measured by RT-PCR. Our results showed that Rz464 could exclusively cleave C II TA RNA. When induced with IFN-gamma, the expression of HLA-DR, -DP, -DQ on pRz464+ Hela was induced, and the mRNA content of C II TA decreased too. It is concluded that Rz464 could inhibit C II TA and thus the family of genes was regulated by C II TA:MHC II molecules. These results provided insight into the future application of Rz464 as a new nucleic acid drug against auto-immune diseases.