[Influence of nuclear factor-kappaB activation on the expression of cytokines in monocytes stimulated by burn serum].

OBJECTIVE

To investigate the effects of NF-kappaB activation on the expression of cytokines in monocytes stimulated by burn serum, so as to explore the mechanism in monocyte activation by burn serum.

METHODS

Peripheral blood monocytes (PBMCs) isolated from healthy volunteers were employed as the target cells. The cells were stimulated by serum from healthy volunteers (control), by serum from burn patients (burn serum), and by burn serum with addition of PDTC (pyrrolidine dithiocarbamate). Activation of monocytic NF-kappaB before stimulation and at 0.5, 1.0, 2.0 and 4.0 poststimulation hours (PSH) was assessed with electrophoretic mobility shift assay (EMSA). Expression of TNF-alpha and IL-8 mRNA at 1.0, 2.0, 4.0, 6.0 PSHs was assayed with in situ hybridization (ISH). Meanwhile, the contents of TNF-alpha and IL-8 in the supernatants were assayed by enzyme linked immunosorbent assay (ELISA).

RESULTS

The monocytic NF-kappaB activity in burn serum group increased significantly and reached the peak level at 1 PSH [(30.2 +/- 3.5) x 10(4) integration gray scale value] after the PBMCs were stimulated by burn serum, and it was obviously higher than that in control group [(4.4 +/- 0.8) x 10(4) integration gray scale value], (P < 0.01). It gradually decreased and returned to the pre-stimulation state at 2 PSH. The monocytic NF-kappaB activity in PDTC group decreased to [(6.8 +/- 0.9) x 10(4) integration gray scale value at 1 PSH] after the stimulation. The expression of TNF-alpha mRNA of the monocytes and the TNF-alpha level in the supernatant of the cultured PBMCs reached peak level at 1 PSHs after being stimulated by burn serum, and they were obviously higher than those in control group (P < 0.01). While the expression of IL-8 mRNA and the IL-8 level in the supernatant of the cultured PBMCs reached peak level at the 4 PSHs after being stimulated by burn serum, which were obviously higher than those in control group (P < 0.01) too. In addition, the synthesis and release of TNF-alpha (peaked at 1 PSH: 0.52 +/- 0.06 microg/L) and IL-8 (peaked at 4 PSH: 239 +/- 20 ng/L) in the supernatant of PBMCs in PDTC group were evidently higher than those in control group [(0.13 +/- 0.07) microg/L, < 156 ng/L] (P < 0.01).

CONCLUSION

Burn serum can induce the activation of NF-kappaB which lead to the synthesis and release of cytokines from PBMC. This result indicates that the activation of NF-kappaB plays important role in the secretion of cytokines from PBMCs induced by the burn serum.