Hepatic Mitochondrial Dysfunction and Immune Response in a Murine Model of Peanut Allergy

Abstract

Background: Evidence suggests a relevant role for liver and mitochondrial dysfunction in allergic disease. However, the role of hepatic mitochondrial function in food allergy is largely unknown. We aimed to investigate hepatic mitochondrial dysfunction in a murine model of peanut allergy.

Methods: Three-week-old C3H/HeOuJ mice were sensitized by the oral route with peanut-extract (PNT). We investigated: 1. the occurrence of effective sensitization to PNT by analysing acute allergic skin response, anaphylactic symptoms score, body temperature, serum mucosal mast cell protease-1 (mMCP-1) and anti-PNT immunoglobulin E (IgE) levels; 2. hepatic involvement by analysing interleukin (IL)-4, IL-5, IL-13, IL-10 and IFN-γ mRNA expression; 3. hepatic mitochondrial oxidation rates and efficiency by polarography, and hydrogen peroxide (H₂O₂) yield, aconitase and superoxide dysmutase activities by spectrophotometry.

Results: Sensitization to PNT was demonstrated by acute allergic skin response, anaphylactic symptoms score, body temperature decrease, serum mMCP-1 and anti-peanut IgE levels. Liver involvement was demonstrated by a significant increase of hepatic Th2 cytokines (IL-4, IL-5 and IL-13) mRNA expression. Mitochondrial dysfunction was demonstrated by lower state 3 respiration rate in the presence of succinate, decreased fatty acid oxidation in the presence of palmitoyl-carnitine, increased yield of ROS proven by the inactivation of aconitase enzyme and higher H₂O₂ mitochondrial release.

Conclusions: We provide evidence of hepatic mitochondrial dysfunction in a murine model of peanut allergy. These data could open the way to the identification of new mitochondrial targets for innovative preventive and therapeutic strategies against food allergy.

Keywords: Th2 cytokines; food allergy; mitochondrial function; oxidative stress.

Figure 1

Schematic overview of the experimental design. Three-week-old female C3H/HeOuJ mice (n = 6 per group) were sensitized orally every 7 days for 4 weeks using a blunt needle with peanut extract (PNT) + cholera toxin (CT) as adjuvant. Controls mice receive CT only. On day 34, mice received intradermal injection of PNT in the ear pinnae acute allergic skin response was measured. After 24 h, mice were challenged by gavage with PNT and anaphylaxis score and body temperature were determined. On the next day mice were sacrificed, liver and blood samples were collected.

Figure 2

Proofs of effective sensitization to PNT. Acute allergic ear swelling response 1 h after intradermal injection of PNT in the ear pinnae (A), anaphylactic symptoms score (B), body temperature decrease (C), serum levels of mMCP-1 (D) and anti-peanut IgE (E). Data were analyzed with independent sample t-test. CTRL = control mice; PNT = mice sensitized with peanut extract. * vs. CTRL, p < 0.05; ** vs. CTRL, p < 0.0001. Data represent pooled values from two separate experiments.

Figure 3

Proofs of hepatic involvement in Th2 response in mice sensitized with PNT: increase in IL-4 (A), IL-5 (B) and IL-13 (C) mRNA liver expression detected by Real Time PCR. No significant difference in IL-10 (D) and IFN-γ (E) mRNA liver expression was observed between two groups. Data are reported as means ± SEM. Data were analysed with independent sample t-test. CTRL = control mice; PNT = mice sensitized with peanut extract. * vs. CTRL, p < 0.05; ** vs. CTRL, p < 0.0001.

Figure 4

Evidence of hepatic mitochondrial dysfunction in mice sensitized by PNT. Hepatic mitochondrial state 3 and 4 respiration rates in the presence of succinate (A) or palmitoyl-carnitine (B) were evaluated, and the relative respiratory control ratios (RCRs) were shown (upper panels). Oxygen consumption in the presence of oligomycin or uncoupled by FCCP (C), and degree of coupling values (D) were also shown. The mitochondrial H₂O₂ yield (E), the SOD activity (F), the basal and total activity of the aconitase (G) and its ratio (H) were reported. Data are reported as means ± SEM from duplicate analyses. Data were analyzed with independent sample t-test. CTRL = control mice; PNT = mice sensitized with peanut extract. * vs. CTRL, p < 0.05; ** vs. CTRL, p < 0.0001.