Cytokines and neutrophils as important mediators of platelet-activating factor-induced kinin B1 receptor expression

Abstract

PAF injection into the rat paw is accompanied by the concomitant activation of NF-kappaB and neutrophil influx, which appears to be relevant to the up-regulation of kinin B1 receptors. Herein, we analyse the role of TNF-alpha and IL-1beta production for PAF-induced B1 receptor upregulation in the rat paw. Additionally, we evaluate how cytokine production and neutrophil migration fit into the temporal sequence of events leading to PAF-induced B1 receptor upregulation. In our experiments, treatment with PAF resulted in a marked increase of B1 receptor-mediated paw oedema and in situ production of TNF-alpha at 1 h and IL-1beta at 3 and 6 h later. B1 receptor-mediated paw oedema was significantly inhibited by anti-TNF-alpha antibody and by interleukin-1 receptor antagonist (IRA). TNF-alpha was necessary for the local PAF-induced IL-1beta production. NF-kappaB blocker PDTC prevented the production of both TNF-alpha and IL-1beta, indicating that cytokine production is NF-kappaB dependent. Depletion of neutrophils with an anti-PMN antibody prevented IL-1beta, but not TNF-alpha, production. Although both TNF-alpha and IL-1beta are relevant to functional B1 receptor upregulation, PAF-induced increase in B1 receptor mRNA was markedly suppressed by anti-TNF-alpha and, to a lesser extent, by IRA. B1 receptor mRNA expression was also prevented by the anti-PMN antibody. In conclusion, the activation of the TNF-alpha/neutrophil axis by PAF seems to be sufficient for B1 receptor mRNA production. However, the TNF-alpha/neutrophil axis is also necessary for IL-1beta production. These two processes might lead to the appearance of functional kinin B1 upregulation receptors in vivo after PAF treatment.

Figure 1

Effect of Abs anti-TNF-α or IRA on DABK-induced paw oedema. The anti-murine Abs anti-TNF-α (50 ng paw⁻¹; a), recombinant human IRA (100 μg paw⁻¹; b) or anti-murine Abs anti-IL-1β (50 ng paw⁻¹; c) were coadministered with PAF (10 nmol paw⁻¹) 6 h before the injection of DABK. Values represent the differences between volume (in ml) of vehicle-injected (0.1 ml of PBS solution) and drug-injected paws. Each point represents the mean±s.e.m. of four to six animals. In some cases, error bars are hidden within the symbols. Significantly different from control values. ^**P<0.01.

Figure 2

Effect of Abs anti-TNF-α, IRA or Abs anti-PMN on kinin B₁ receptor mRNA expression induced by PAF. The anti-murine anti-TNF-α (50 ng paw⁻¹) or recombinant human IRA (100 μg paw⁻¹) was coadministered with PAF injection (10 nmol paw⁻¹). The anti-rat anti-PMN (34 μg kg⁻¹, i.p.) was administered systemically 30 min before PAF injection (10 nmol paw⁻¹). Bottom, Graphic representation of B₁/β-actin signals ratio. Lane 1, PBS; lane 2, PAF 3h; lane 3, anti-TNF-α; lane 4, IRA; lane 5, anti-PMN. Number of replicates=3. Significantly different from PBS ^**P<0.01 or PAF-injected paws ^#P<0.05, ^##P<0.01 values.

Figure 3

Time-dependent effect of PAF treatment on TNF-α levels and effect of PDTC on TNF-α production. (a) Effect of PAF injection (10 nmol paw⁻¹, 1–3 h) on TNF-α levels in the rat paw. (b) Effect of PDTC (100 mg kg⁻¹, i.p.) administered 30 min before PAF treatment on TNF-α production. Each column represents the mean±s.e.m. of four to six animals. Significantly different from PBS ^**P<0.01 or PAF-injected paws (^##) values.

Figure 4

Time-dependent effect of PAF treatment on IL-1β levels and effect of PDTC, Abs anti-TNF-α or anti-PMN on IL-1β production. (a) Effect of PAF injection (10 nmol paw⁻¹, 1–12 h) on IL-1β levels in the rat paw. (b) Effect of PDTC (100 mg kg⁻¹, i.p.), anti-PMN (34 μg kg⁻¹, i.p.; administered 30 min before PAF) or anti-murine anti-TNF-α (50 ng paw⁻¹; coadministered with PAF) on IL-1β production (3 h after PAF injection). Each column represents the mean±s.e.m. of four to six animals. Significantly different from PBS ^**P<0.01 or PAF-injected paws (^##) values.

Figure 5

Effect of Abs anti-TNF-α or IRA on increase of MPO levels induced by PAF. (a) Effect of anti-murine anti-TNF-α (50 ng paw⁻¹; a) or recombinant human IRA (100 μg paw⁻¹; b) both co-administered with PAF (10 nmol paw⁻¹, 6 h prior) on MPO levels in the rat paw. Each column represents the mean±s.e.m. of four to six animals. Significantly different from PBS values ^**P<0.01.

Figure 6

Possible sequence of events leading to B₁ receptor expression after local PAF administration in vivo.