Quantification of expression of leukotriene B4 inducing tumor necrosis factor-? and interleukin-1? at mRNA level in synovial membrane cells of rheumatoid arthritis by real-time quantitative PCR
- VernacularTitle:类风湿关节炎滑膜细胞中白三烯B4诱导肿瘤坏死因子-?和白细胞介素-1? mRNA表达的测定
- Author:
Zhankun CHEN
;
Houshan LV
- Publication Type:Journal Article
- Keywords:
Leukotriene B4;
Tumor necrosis factor-alpha;
Interleukin-1;
Arthritis, rheumatoid;
Polymerase chain reaction
- From:
Journal of Peking University(Health Sciences)
2003;0(05):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To investigate quantification of expression of LTB4 inducing IL-1? and TNF-? at mRNA level in synovial membrane cells of rheumatoid arthritis. Methods: Primary cultured synovial cells from RA patients were treated with exogenous LTB4,MK-886(inhibitor of 5-lipoxygenase activating protein) and Bestatin(inhibitor of leukotriene A4 hydrolase) in the presence of LIT respectively, expressions of TNF-? and IL-1? were detected at mRNA level by Real-time Quantitative PCR. Results: Expressions of basic TNF-?(TNF-?/GAPDH) and IL-?(IL-?/GAPDH) at mRNA level in primary cultured synovial cells were 0.02?0.00 and 0.16?0.01 respectively. LTB4(10~ -9 mol/L-10~ -8 mol/L) was shown to induce dose-dependent increase of mRNA expression of TNF-?.(7-15 times) and IL-1? (1 time) , endogenous product of LTB4 by LIT significantly increased mRNA expressions of TNF-? (145 times) and IL-1?(12 times) respectively. LIT-treated synoviocytes with addition of MK-886 (5-LOX exciting protein FLAP inhibitor) (1-10 ?mol/L) were inhibited to secrete LTB4 dose-dependently, following the markedly down-regulated expressions of TNF-? (15%-66%) and IL-1? (41%-71%) at mRNA level . Bestatin(100 mg/L) could also remarkably diminish LTB4-induced mRNA expressions of TNF-?(86%) and IL-1?(79%). Conclusion: LTB4 of synovial membrance cells in rheumatoid arthritis could induce expressions of TNF-? and IL-1? at mRNA level, and their expression at mRNA level had been quantified successfully. It is a beneficial help to quantify all kinds of cytokines in methodology.
