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. 2011 May 19:8:54.
doi: 10.1186/1742-2094-8-54.

Inhibition of interleukin-6 trans-signaling in the brain facilitates recovery from lipopolysaccharide-induced sickness behavior

Michael D Burton  1 , Nathan L Sparkman, Rodney W Johnson
Affiliations

Inhibition of interleukin-6 trans-signaling in the brain facilitates recovery from lipopolysaccharide-induced sickness behavior

Michael D Burton et al. J Neuroinflammation. .

Abstract

Background: Interleukin (IL)-6 is produced in the brain during peripheral infection and plays an important but poorly understood role in sickness behavior. Therefore, this study investigated the capacity of soluble gp130 (sgp130), a natural inhibitor of the IL-6 trans-signaling pathway to regulate IL-6 production in microglia and neurons in vitro and its effects on lipopolysaccharide (LPS)-induced sickness behavior in vivo.

Methods: A murine microglia (BV.2) and neuronal cell line (Neuro.2A) were used to study the effects of stimulating and inhibiting the IL-6 signaling pathway in vitro. In vivo, adult (3-6 mo) BALB/c mice received an intracerebroventricular (ICV) injection of sgp130 followed by an intraperitoneal (i.p.) injection of LPS, and sickness behavior and markers of neuroinflammation were measured.

Results: Soluble gp130 attenuated IL-6- and LPS-stimulated IL-6 receptor (IL-6R) activation along with IL-6 protein release in both microglial (BV.2) and neuronal (Neuro.2A) cell types in vitro. Moreover, in vivo experiments showed that sgp130 facilitated recovery from LPS-induced sickness, and this sgp130-associated recovery was paralleled by reduced IL-6 receptor signaling, mRNA, and protein levels of IL-6 in the hippocampus.

Conclusions: Taken together, the results show that sgp130 may exert an anti-inflammatory effect on microglia and neurons by inhibiting IL-6 binding. These data indicate that sgp130 inhibits the LPS-induced IL-6 trans-signal and show IL-6 and its receptor are involved in maintaining sickness behavior.

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Figures

Figure 1
Figure 1
Differential expression of IL-6R, gp130, and TLR-4 on microglia and neurons. A) Two-color dot plot; cells were incubated with anti-CD126 PE and anti-CD130 APC and expression of the cell surface markers were assessed on microglia and neurons. B) Single-parameter histograms; cells were incubated with anti-TLR-4 and compared with isotype controls.
Figure 2
Figure 2
IL-6 and STAT3 expression in BV.2 microglial and Neuro.2A neuronal cells. IL-6 receptor activation in BV.2 and Neuro.2A cells was verified by STAT3 phosphorylation 20 min after treatment with 10 and 50 ng/mL of IL-6. Results are an average of 5 independent experiments. Means with different letters are significantly different from one another (P < 0.05).
Figure 3
Figure 3
IL-6 trans-signaling in BV.2 microglia and Neuro.2A cells. BV.2 and Neuro2A cells were pre-treated for 1 h with 25 ng/mL sIL-6R and A) IL-6-induced STAT3 phosphorylation and B) LPS-induced IL-6 protein secretion were measured at 20 min and 3 h, respectively. Results are an average of 5 independent experiments. Means with different letters are significantly different from one another (P < 0.05).
Figure 4
Figure 4
sgp130 attenuation of LPS-induced STAT3 phosphorylation in Neuro.2A and BV.2 cells. BV.2 microglia and Neuro.2A neuronal cells were pre-treated for 1 h with 100 ng sgp130 and LPS-induced A) STAT3 phosphorylation and B) IL-6 protein secretion were measured. Results are an average of 5 independent experiments. Means with different letters are significantly different from each other (P < 0.05).
Figure 5
Figure 5
sgp130 facilitates recovery from LPS-induced sickness behavior. Mice were injected ICV with vehicle or 100 ng sgp130 and i.p. with sterile saline or LPS. Social exploratory baseline behavior was measured before LPS injection and at 2, 4, 6, 8, and 24 h post-injection. Bars represent the mean ± SEM (n = 11-12) Means with * are statistically different (P < 0.05) from saline controls.
Figure 6
Figure 6
sgp130 reduces IL-6 receptor activation, gene expression, and protein secretion in vivo. Hippocampal tissue was collected 8 h after ICV sgp130 and i.p. LPS and assayed for A) STAT3 phosphorylation, B) mRNA levels of IL-6¸IL-1β, and TNF-α, C) and IL-6 protein. sgp130 reduced LPS-induced STAT3 phosphorylation, mRNA levels of IL-6, but not IL-1β or TNF-α, and IL-6 protein in the hippocampus. Bars represent the mean ± SEM (n = 8-9) Means with different letters are statistically different from each other (P < 0.05).
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