Inhibitory effects of anti-CII TA M1-RNA on IFN-gamma induced major histocompatibility complex class II antigens expression on cultured human chondrocytes.
Major histocompatibility complex class II (MHC-II) trans-activator (CII TA) has been shown to be required for constitutive and IFN-gamma-induced MHC-II transcription. This study investigated the inhibitory effect of anti-CII TA M1-RNA on expression of MHC-II in chondrocytes in response to IFN-gamma. M1-RNAs with different guide sequence (GS) recognizing 452 or 3408 sites in CII TA (M1-452-GS and M1-3408-GS, respectively) were cloned into pUC19 vector. Target mRNA (3176-3560) in CII TA was obtained from Raji cell and inserted into pGEM-7zf(+) plasmid. The recombinant M1-RNAs and their target mRNA were incubated in a cell-free condition. It showed that only M1-3408-GS could cleave the target mRNA exclusively. M1-3408-GS was also cloned into psNAV vector (named pA3408). Chondrocytes was stably transfected with pA3408 and expressions of classical MHC-II (HLA-DR, -DP, -DQ) were analyzed by Flow Cytometry. The level of CII TA mRNA was measured by RT-PCR. Peripheral blood mono-nucleated cells (PBMNCs) were stimulated by pA3408-positive chondrocytes in mixed lymphocyte reaction, and proliferation of PBMNCs and IL-2 mRNA were detected. The expression of HLA-DR and HLA-DP on pA3408-positive chondrocytes in response to IFN-gamma decreased 73.00%+/-5.24%, 88.47%+/-2.02%, respectively (P<0.05); So did the content of CII TA mRNA (70.11%+/-5.79%, P<0.05). Proliferation of PBMNCs and production of IL-2 mRNA were both inhibited by pA3408 in mixed lymphocyte reaction. This is the first description that anti-CII TA M1-RNA could prevent IFN-gamma-induced CII TA transcription and results in a decreased MHC-II expression in chondrocytes.
He
Fei
F
Department of Cardiology, Guangdong Provincial People's Hospital, Guangzhou, Guangdong Province, 510080, China.
Guo
Rong
R
Du
Xin
X
Lu
Ze-Sheng
ZS
Weng
Jian-Yu
JY
Lin
Wei
W
D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000276:immunology*; D029968:Escherichia coli Proteins / Q000235:genetics*; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism; D000949:Histocompatibility Antigens Class II / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007371:Interferon-gamma / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D007959:Lymphocyte Culture Test, Mixed; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000378:metabolism; D043262:Ribonuclease P / Q000235:genetics*; D015534:Trans-Activators / Q000037:antagonists & inhibitors* / Q000235:genetics
D029968:Escherichia coli Proteins; D015395:Histocompatibility Antigens Class I; D000949:Histocompatibility Antigens Class II; C083728:MHC class II transactivator protein; D009687:Nuclear Proteins; D012333:RNA, Messenger; D015534:Trans-Activators; D007371:Interferon-gamma; D043262:Ribonuclease P; C470865:ribonuclease P, E coli