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Repression of allo-cell transplant rejection through CIITA ribonuclease P+ hepatocyte.

Abstract：

AIM:Allo-cell transplant rejection and autoimmune responses were associated with the presence of class II major histocompatibility complex (MHC II) molecules on cells. This paper studied the effect of Ribonuclease P (RNase P) against CIITA, which was a major regulator of MHCII molecules, on repressing the expression of MHCII molecules on hepatocyte. METHODS:M1-RNA is the catalytic RNA subunit of RNase P from Escherichia coli. It were constructed that M1-RNA with guide sequences (GS) recognizing the 452, 3408 site of CIITA by PCR from pTK117 plasmid, then were cloned into the EcoRI/BglII or EcoR//SalIsite of vector psNAV (psNAV-M1-452-GS, psNAV-M1-3408-GS) respectively. The target mould plate (3176-3560) of CIITA was obtained from Raji cell by RT-PCR, and then inserted into the XhoI/EcoRIof pGEM-7zf(+) plasmid (pGEM-3176). These recombinant plasmids were screened out by sequence analysis. psNAV-M1-452-GS, psNAV-M1-3408-GS and its target RNA pGEM-3176 were transcribed and then mixed up and incubated in vitro. It showed that M1-3408-GS could exclusively cleave target RNA that formed a base pair with the GS. Stable transfectants of hepatocyte cell line with psNAV-M1-3408-GS were tested for expression of class II MHC through FCM, for mRNA abundance of MHCII, Ii and CIITA by RT-PCR., for the level of IL-2 mRNA on T cell by mixed lymphocyte reaction. RESULTS:When induced with recombinant human interferon-gamma (IFN-gamma), the expression of HLA-DR, -DP, -DQ on psNAV-M1-3408-GS(+) hepatocyte was reduced 83.27 %, 88.93 %, 58.82 % respectively, the mRNA contents of CIITA, HLA-DR, -DP, -DQ and Ii decreased significantly. While T cell expressed less IL-2 mRNA in the case of psNAV-M1-3408-GS(+) hepatocyte. CONCLUSION:The Ribonuclease P against CIITA-M1-3408-GS could effectively induce antigen-specific tolerance through cleaving CIITA. These results provided insight into the future application of M1-3408-GS as a new nucleic acid drug against allo-transplantation rejection and autoimmune diseases.

journal_name

World J Gastroenterol

journal_title

World journal of gastroenterology

authors

Guo R,Zou P,Fan HH,Gao F,Shang QX,Cao YL,Lu HZ

doi

10.3748/wjg.v9.i5.1077

keywords:

subject

Has Abstract

pub_date

2003-05-01 00:00:00

pages

1077-81

issue

eissn

1007-9327

issn

2219-2840

journal_volume

pub_type

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Repression of allo-cell transplant rejection through CIITA ribonuclease P+ hepatocyte.