[The analysis in the expression of Toll-like receptor 4 mRNA and the effect of dexamethasone on mice dendritic cells by real-time PCR].
OBJECTIVE
To establish a SYBR Green I quantitative real-time PCR method for detecting the expression of the TLR4 mRNA of mice and to monitor the dynamics for TLR4 mRNA level of mice bone marrow-derived dendritic cells (DC) in the process of the maturation and the effect of dexamethasone (DEX) on TLR4 expression.


METHODS
(1) DC from mice bone marrow were induced by cytokines and separated by magnetic beads. (2) The combined plasmid pUCm-T/TLR4 and pUCm-T/beta-actin for the standard materials in the real-time PCR was reconstructed. (3) The dynamics for the express TLR4 mRNA of DC cultivated in vitro for 4, 6, 8, 10, 12 days was detected respectively. (4) The TLR4 mRNA expression between DC treated with or without DEX was detected and compared.


RESULTS
(1) The DC with the purity over 90% were fully separated in success. (2) A SYBR Green I quantitative real-time PCR method for analyzing the TLR4 mRNA expression level of the mice was successful established. (3) TLR4 mRNA level was stable early during the culture of DC and then rise obviously at last.


CONCLUSIONS
There exists a close relationship between the level of TLR4 mRNA and the period of maturation of DC and the expression of TLR4 could be increased by DEX.
Zhang
Han
H
Department of Immunology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Lu
Ze
Z
Li
Dong-mei
DM
He
Xiu-juan
XJ
Wu
Xiang-ling
XL
Hu
Yong-xiu
YX
D000818:Animals; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology*; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*; D051197:Toll-Like Receptor 4 / Q000235:genetics*
D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D003907:Dexamethasone