|
KMID : 1148120120020020047
|
|
Journal of Advanced Spine Surgery
2012 Volume.2 No. 2 p.47 ~ p.59
|
|
|
Steroid Treatment Can Inhibit Intranuclear Nuclear Factor-κB Pathway in TNF-α Stimulated Human Disc Cells
|
|
Kim Young-Yul
Park Sang-Eun
Quan Meiling
Lin Zhenhua
Hong Myung-Wha
Rhyu Kee-Won
|
|
|
Abstract
|
|
|
Purpose: The main purpose of this study was the investigation of the effect of Dexamethasone (DEXA) treatment on
TNF-α stimulated intervertebral disc cells through the action mechanism of NF-κB in the cytoplasm and nucleus.
Materials and Methods: We separated cultured human intervertebral disc cells passed three times into four groups. A: control group, B: TNF-α treatment group, C: DEXA treatment group, D: TNF-α and DEXA treatment simultaneously. After extraction of cytoplasmic and nuclear protein from the 4 groups at time points including 10 minutes, 1 hour, and 2 hours, we measured the protein expression levels of p50, p65, p52, and p100 by western blot analysis. Also, we observed the expression of p50, p52, p65 in each group at the 1 hour time point by immunofluorescence analysis.
Results: Western blot analysis demonstrated that cytoplasmic levels of p50 and p65 at1 hour in groups B and D were decreased, groups C showed no significant change as compared to the control group. Nuclear levels of p50 at 1 hour in groups B (10.99 fold) and D (7.24 fold) were increased, and group D had decreased expression compared to group B. Nuclear levels of p50 expression at 2 hours in groups B (12.33 fold) was increased compared to the levels measured at 1 hour. Levels of nuclear p50 in group D at 2 hour time points showed no significant change as compared to the group D at 1 hour time points. In the nucleus, the level of p65 at 1 hour had the same pattern as the p50 expression, however, group B (4.13 fold) and D (4.13 fold) expression levels at 2 hours were decreased compared to the group B (7.49 fold) and D (6.79 fold) at 1 hour. In the cytoplasm, the expression of p100 in groups B and D were decreased after 1hour, and other groups had the same trend as that observed for the control group. Nuclear p100 expression levels were observed in, groups B, C, and D after 2 hours. The cytoplasmic levels of p52 at 10min, 1 hour, and 2 hours were same. The nuclear levels of p52 in group D at 1 hour had no expression and decreased at 2 hours (0.08 fold). The results of the immunofluorescence analysis and the western blot analysis at the 1 hour time point is quite consistent.
Conclusion: Transcriptional mediation of NF-κB was mainly focused on the classical pathway but several protein levels were influenced by the alternative pathway following stimulation with TNF-α in disc cells. The effect of DEXA on NF-κB transcription signaling was observed through the delayed expression of involved proteins and inhibited the translocation of p50, p52, p65 to prevent the expression of corresponding genes.
|
|
|
KEYWORD
|
|
|
Human disc cell, NF-κB, TNF-α, Dexamethasone
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
|
|
|