Central inhibition of interleukin-6 trans-signaling during peripheral infection reduced neuroinflammation and sickness in aged mice

Abstract

During systemic infection, inflammatory cytokines such as interleukin (IL)-6 are produced in excess in the brain of aged mice and induce severe behavioral deficits. However, no studies have examined how pro-inflammatory IL-6 trans-signaling is involved in the exaggerated production of IL-6 in the aged brain, nor the extent to which IL-6 trans-signaling affects other markers of neuroinflammation, adhesion molecules, and behavior. Therefore, this study investigated in aged mice the presence of IL-6 signaling subunits in microglia; the central effects of soluble gp130 (sgp130)-a natural inhibitor of the IL-6 trans-signaling pathway-on IL-6 production in microglia; and the effects of sgp130 given intracerebroventricularly (ICV) on neuroinflammation and sickness behavior caused by i.p. injection of lipopolysaccharide (LPS). Here we show that microglia isolated from aged mice have higher expression of IL-6 receptor (IL-6R) compared to microglia from adults; and the level of mRNA for ADAM17, the enzyme responsible for shedding membrane-bound IL-6R in trans-signaling, is higher in the hippocampus of aged mice compared to adults. Additionally, we show in aged mice that peripheral LPS challenge elicits a hyperactive IL-6 response in microglia, and selective blockade of trans-signaling by ICV injection of sgp130 mitigates this. The sgp130-associated inhibition of IL-6 was paralleled by amelioration of exaggerated and protracted sickness behavior in aged mice. Taken together, the results show that microglia are important regulators of the IL-6 trans-signaling response in the aged brain and sgp130 exerts an anti-inflammatory effect by inhibiting the pro-inflammatory arm of IL-6 signaling.

Figure 1. Differential expression of IL-6 receptor and gp130 on microglia isolated from adult and aged mice

Average percentage of cells that were A) CD11b⁺/IL-6R⁺ B) and CD11b⁺/gp130⁺. Cell surface markers were assessed for CD11b, CD45, and IL-6R or gp130 expression; compared with isotype and unstained controls. Bars represent the means ± SEM (n = 8). Means marked with a * are significantly different from each other (P < 0.05).

Figure 2. Hippocampal gene expression of IL-6 receptor sheddases ADAM17 and ADAM10 from adult and aged mice

Hippocampal tissue was collected and assayed for A) ADAM17 and B) ADAM10. Aged animals had a significant upregulation of baseline levels of ADAM17 gene expression. Bars represent the mean ± SEM (n = 8–9) Means with * are statistically different from each other (P<0.05).

Figure 3. sgp130-mediated LPS-induced sickness behavior in adult and aged animals

Mice were injected ICV with vehicle or sgp130 (100ng) and i.p. with sterile saline or LPS (10 μg). Spontaneous locomotor baseline behavior was measured before injections (0) and at 4, 6, 8, and 24 h post-injection. Bars represent the mean ± SEM (n = 10–11) Means with * are statistically different (P<0.05) from saline controls.

Figure 4. sgp130 ameliorates LPS-induced microglial IL-6 production in the aged, while unaltering MHC-II expression

Adult and aged mice were injected ICV with vehicle or sgp130 (100ng) and i.p. with sterile saline or LPS (10 μg) and microglia were isolated by percoll density gradient 6 h later. Cells were subjected to the BD Cytofix/Cytoperm^™ fixation/permeabilization protocol and stained with anti-CD11b-APC, anti-CD45-FITC, and anti-IL-6-PE or MHC-II-PE. Bars represent the means ± SEM (n =7–8). Means with different letters are statistically different (P<0.05) from each other.