Systemic Administration of α7-Nicotinic Acetylcholine Receptor Ligands Does Not Improve Renal Injury or Behavior in Mice With Advanced Systemic Lupus Erythematosus

Abstract

There is a critical need for safe treatment options to control inflammation in patients with systemic lupus erythematosus (SLE) since the inflammation contributes to morbidity and mortality in advanced disease. Endogenous neuroimmune mechanisms like the cholinergic anti-inflammatory pathway can be targeted to modulate inflammation, but the ability to manipulate such pathways and reduce inflammation and end organ damage has not been fully explored in SLE. Positive allosteric modulators (PAM) are pharmacological agents that inhibit desensitization of the nicotinic acetylcholine receptor (α7-nAChR), the main anti-inflammatory feature within the cholinergic anti-inflammatory pathway, and may augment α7-dependent cholinergic tone to generate therapeutic benefits in SLE. In the current study, we hypothesize that activating the cholinergic anti-inflammatory pathway at the level of the α7-nAChR with systemic administration of a partial agonist, GTS-21, and a PAM, PNU-120596, would reduce inflammation, eliminating the associated end organ damage in a mouse model of SLE with advanced disease. Further, we hypothesize that systemic α7 ligands will have central effects and improve behavioral deficits in SLE mice. Female control (NZW) and SLE mice (NZBWF1) were administered GTS-21 or PNU-120596 subcutaneously via minipumps for 2 weeks. We found that the increased plasma dsDNA autoantibodies, splenic and renal inflammation, renal injury and hypertension usually observed in SLE mice with advanced disease at 35 weeks of age were not altered by GTS-21 or PNU-120596. The anxiety-like behavior presented in SLE mice was also not improved by GTS-21 or PNU-120596. Although no significant beneficial effects of α7 ligands were observed in SLE mice at this advanced stage, we predict that targeting this receptor earlier in the pathogenesis of the disease may prove to be efficacious and should be addressed in future studies.

Keywords: SLE; behavior; cholinergic anti-inflammatory pathway; positive allosteric modulators; renal inflammation; renal injury.

Figure 1

Plasma levels of dsDNA autoantibodies (activity units) in control and SLE mice treated with either vehicle, GTS-21, or PNU-120596. Neither GTS-21 nor PNU-120596 had a significant effect on albumin excretion rate in control or SLE mice (n = 14–20 per group). Data were analyzed using a two-way ANOVA and the results of this analysis are listed on the graph.

Figure 2

Splenic (A), renal cortical (B), and medullary (C) TNF-α (presented as percent difference from controls) in control and SLE mice treated with either vehicle, GTS-21, or PNU-120596. Neither GTS-21 nor PNU-120596 had a significant effect on renal or splenic inflammation in control or SLE mice (n = 9–13 per group). Data were analyzed using a two-way ANOVA and the results of this analysis are listed on the graph. Representative blots are included above each figure. The molecular weight of TNF-α is 26 kDa.

Figure 3

(A) Albumin excretion rate (mg/day) in control and SLE mice treated with either vehicle, GTS-21, or PNU-120596. Neither GTS-21 nor PNU-120596 had a significant effect on albumin excretion rate in control or SLE mice (n = 13–19 per/group) Data were analyzed using a two-way ANOVA and the results of this analysis are listed on the graph. Note: the y-axis is presented as a log scale. (B) Mean arterial pressure (mmHg) in control and SLE mice treated with either vehicle, GTS-21, or PNU-120596. Neither GTS-21 nor PNU-120596 had a significant effect on mean arterial pressure in conscious and freely moving control or SLE mice (n = 3–6 per group). Data were analyzed using a two-way ANOVA and the results of this analysis are listed on the graph.

Figure 4

Open field test in control and SLE mice pre-treatment (A,C,E,G) and post-treatment with GTS-21 and PNU-120596 (B,D,F,H). The pre-treatment data is a comparison between all control and all SLE mice at baseline. A t-test was used to determine significant differences between SLE and control mice pretreatment. SLE mice had lower locomotion (A), spent a larger amount of time sleeping (C), and traveled less during the test than controls (G) (all p < 0.05). A two-way ANOVA with repeated measures was used to determine differences between all groups post-treatment (B,D,F,H; n = 3–5 per group). The results of each ANOVA appears on the graph.

Figure 5

Light-dark box test in control and SLE mice pre-treatment (A,C) and post-treatment with GTS-21 and PNU-120596 (B,D). The pre-treatment data is a comparison between all control and all SLE mice at baseline. A t-test was used to determine significant differences between SLE and control mice pretreatment and there were no differences (A,C). A two-way ANOVA with repeated measures was used to determine differences between all groups post treatment (B,D; n = 3–5 per group). The results of each ANOVA appears on the graph.

Figure 6

Tail suspension test in control and SLE mice pre-treatment (A) and post-treatment with GTS-21 and PNU-120596 (B). The pre-treatment data is a comparison between all control and all SLE mice at baseline. A t-test was used to determine significant differences between SLE and control mice pretreatment. There was not a difference in immobility during the suspension test between SLE mice and controls. A two-way ANOVA with repeated measures was used to determine differences between all groups post treatment (B; n = 3-5 per group). The results of each ANOVA appears on the graph.