Alternation and modulation of tumor necrosis factor-alpha and protein kinase C in celiac macrophage of mouse after serious scalding

Yong WANG; Wenhua HUANG; Daizhi PENG; Xin ZHOU; Jing LIU

ABSTRACT

BACKGROUND:

Serious scalding leads to dysfunction of each aspect in immune system, and activated macrophage can secret many bioactive transmitters. The relationship between macrophage dysfunction and signal conduction after scalding is unclear at present.

OBJECTIVE:

To observe the alternation of tumor necrosis factor- alpha(TNF-α) at different time points after scalding and the activity of nuclear factor κB(NF-κB) and alternation of protein kinase C (PKC) after the application of specific modulator H-7 to explore whether PKC participates in the modulation of TNF-α in macrophage on signal conduction level for the clarification of some mechanisms of macrophage dysfunction.

DESIGN:

A randomized controlled experimental study by employing experimental animals as subjects

SETTING:

An institute of burn research of a military medical university MATERIALS The study was conducted in the Laboratory (state) of the Institute of Burn Research, Third Military Medical University of Chinese PLA between January and December 1999. Experimental animals were 32 healthy clean inbreeding Kunming white mice.

METHODS:

15% Ⅲ scalding was created in mice for the establishment of routine scalding model. Mice were randomly divided into 6 groups according to different time points before or after scalding, I.e. 0(normal control group), 2, 6, 12, 24, or 48 hours group. Celiac macrophages were collected for the detection of TNF-α content by radioimmunoassay, NF-κB activity by electrophoretic mobility shift analysis (EMSA), and membrane or plasma PKC activity by isotope analysis.MAIN OUTCOME

MEASURES:

① TNF-α content; ② NF-κB activity; ③Membrane or plasma PKC activity

RESULTS:

After scalding, macrophage excessively secreted TNF-α and reached its peak of (1 085.65 ± 122.99) ng/L at 12 hours, which was significantly higher than that of control group( t = 14.92, P ＜ 0.01 ).Compared with control group, membrane PKC activity increased after scalding, which significantly heightened to(231.80 ± 31.66) nmol/min · g at 2hours( t = 7. 930, P ＜ 0.01 ), slightly decreased to close to normal level of (174.29±16.80) nmol/min· gat 6hours(t=2.531, P ＜0.05), and rapidly elevated at 12 hours [512. 10 ±33.42) nmol/min · g] and 24 hours [ (454.70 ± 21.06) nmol/min · g] to reach its peak of(530.49 ± 28.54)nmol/min. G at 48 hours( t = 29.42, 28.03, 30. 19, P ＜ 0. 01 ). Correlation analysis of the alternation between TNF-α and membrane PKC indicated a significant positive correlation( r = 0. 796 4, P ＜ 0. 05) . As indicated by EMSA image, NF-κB activity significantly elevated after scalding. Twelve hours after scalding was set as modulation point, NF-κB activity was significantly inhibited by the application of H-7.

CONCLUSION:

The secretion of TNF-α and the activities of PKC and NF-κB are significantly activated in celiac macrophage after scalding, and PKC-NF-κB signal pathway participates in the modulation of TNF-α expression, which provide experimental data for the modulation of immune function and rehabilitative intervention during scalding.serious scalding.