Abstract

OBJECTIVE
Lentiviral-mediated shRNA against RelB was used to produce tolerogenic dendritic cells from murine bone marrow derived dendritic cells (BMDCs).
METHOD
RelB expression in the BMDCs was silenced by lentivirus carrying RelB shRNA. The apoptosis rate and surface markers of DCs were assessed by flow cytometry. IL-12,IL-10,TGF-β1 secreted by DCs and DNA binding capacity of NF-κB subunits in the nucleus were measured by ELISA, independently. MLR was used to analyze the capacity of DCs to inhibit immune response.
RESULTS
RelB expression was significantly inhibited in DCs following lentiviral mediated delivery of RelB specific shRNA. The RelB shRNA-DC produced lower IL-12 and higher IL-10 than mature dendritic cells (mDCs) and silencing control DCs. There was no difference in the apoptosis rate between shRNA RelB-DCs and mDCs. The expression levels of co-stimulatory molecules (CD80, CD86 and CD83) and MHC-II class molecule were lower in the RelB shRNA-DCs than in the mDCs and silencing control DCs. In addition, RelB shRNA also inhibited the RelB DNA binding capacity but had no effect on other NF-κB subunits. The shRNA RelB-DCs can significantly inhibit mixed lymphocyte reaction (MLR) and down-regulate Th1 cytokines and prompt the production of Th2 cytokines.
CONCLUSION
Our results indicate RelB shRNA transfection of DCs can induce the immature status, and produce tolerogenic DCs.

Links

Publisher Full Text

Authors

Qiu T, Zhu HC, Liu XH, Dong WC, Weng XD, Hu CH, Kuang YL, Gao RH, Dan C, Tao T

Institution

Department of Urology, Renmin Hospital of Wuhan University, Wuhan University, Wuhan 430060, China.

Source

International immunopharmacology 12:3 2012 Mar pg 501-9

MeSH

Animals
Apoptosis
Blotting, Western
Bone Marrow Cells
DNA-Binding Proteins
Dendritic Cells
Flow Cytometry
Gene Silencing
Genetic Vectors
Interleukin-10
Interleukin-12
Lentivirus
Lymphocyte Culture Test, Mixed
Membrane Proteins
Mice
Mice, Inbred C57BL
NF-kappa B
RNA, Small Interfering
Real-Time Polymerase Chain Reaction
Th1 Cells
Th2 Cells
Transcription Factor RelB

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22266276