Glycated bovine serum albumin for use in feeding trials with animal models - In vitro methodology and characterization of a glycated substrate for modifying feed pellets.

This study investigated different methods to produce Nε-carboxymethyl-lysine (CML)-enriched bovine serum albumin (BSA) as alternatives to the classical approach using glyoxylic acid (GA) and sodium cyanoborohydride (NaBH3CN) which results in toxic hydrogen cyanide (HCN). The reaction of GA (6 mmol/L) and NaBH3CN (21 mmol/L) to produce CML remained the most effective with CML yields of 24-35%, followed by 13-24% using 300 mmol/L glyoxal (GO). GA promoted specific modification of lysine to CML, and fewer structural modifications of the BSA molecule compared with GO, as evidenced by fluorescence and proteomic analyses. GO promoted greater arginine modification compared with GA (76 vs 23%). Despite structural changes to BSA with GO, murine fecal clearance of CML was similar to literature values. Hence, BSA glycation with 300 mmol/L glyoxal is a suitable alternative to GA and NaBH3CN for generating CML-enriched protein free of HCN, but a CML-only fortification model remains to be described.

Treatment of gastric eosinophilia by epicutaneous immunotherapy in piglets sensitized to peanuts.

BACKGROUND: Eosinophilic gastrointestinal disorders (EGIDs) are hypersensitivity disorders frequently triggered by food allergy and manifested by mucosal eosinophilic infiltration at any level of the gastrointestinal tract. This study established a model of gastric eosinophilia in peanut-sensitized piglets to evaluate the efficacy of epicutaneous immunotherapy (EPIT) for its treatment. METHODS: Experiments were carried out in piglets first sensitized by three intra-peritoneal injections of peanut protein extract (PPE) with adjuvant, and then given PPE orally for 10 days, a sequence leading to gastric eosinophilia assessed by endoscopy. For 3 months, eight piglets received active EPIT, using Viaskin® loaded with PPE, applied daily on the ear, while eight received placebo EPIT (Placebo). Piglets were exposed to a second 10-day period of PPE orally. Lesions were scored by endoscopy on the last day of PPE exposure. After killing, all parts of the digestive tract were analysed by a pathologist unaware of the piglets' status. IgE response was measured, and mechanistic parameters were analysed in the spleen. RESULTS: After sensitization, a significant increase of total IgE was observed in sensitized compared to naive animals (61.1 ± 13.3 vs 27.8 ± 6 ng/mL, P < .01). Following oral intake of PPE, sensitized piglets developed moderate gastritis compared to naive piglets (1.5 vs 1.0, median score). After 3 months of immunotherapy, median IgE was significantly reduced in EPIT vs placebo piglets (61.4 ± 16.3 vs 105.9 ± 25.6 ng/mL, P < .01). Active EPIT significantly reduced gastric mucosal lesions induced by PPE oral intake (macroscopic score 0 [0-2] vs 2 [1-3], P < .01, respectively, active vs placebo) and gastric mucosa eosinophils counts (239 eosinophils/mm2 [59-645] vs 2554 eosinophils/mm2 [462-8057], P < .01, respectively active vs placebo). GATA-3, IL-5 and eotaxin mRNA expression decreased significantly after EPIT (P < .05). CONCLUSIONS: This study describes a large animal model of gastric eosinophil in peanut-sensitized piglets. Utilizing this model, we demonstrated the efficacy of EPIT in treating peanut-induced EGIDs.

Dose-dependent effects of endotoxins on allergen sensitization and challenge in the mouse.

BACKGROUND: Levels of endotoxins greatly differ according to environmental settings. OBJECTIVE: To study the effect of lipopolysaccharide (LPS) at increasing doses (0.1-1000 ng) on allergen sensitization and challenge in the mouse. METHODS: Mice were sensitized systemically and challenged locally with ovalbumin (OVA) in the presence or absence of LPS. Inflammation was assessed by determining total and differential cell counts and T-helper type 2 (Th)2 cytokine (IL-4 and IL-5) levels in bronchoalveolar lavage fluid (BALF). Total and OVA-specific IgE levels were quantified in serum. Airway hyper-responsiveness (AHR) was assessed by whole-body barometric plethysmography. RESULTS: Administered prior to sensitization, LPS at 100 or 1000 ng dose-dependently decreased allergen- induced total and OVA-specific IgE, airway eosinophilia and Th2 cytokines in BALF, without changing AHR. Administered during OVA challenge, LPS at 1 ng (an infra-clinical dose) or 100 ng (a dose triggering neutrophilia) enhanced airway eosinophilia, without affecting IgE levels or AHR. CONCLUSION: Our data clearly demonstrate that exposure to LPS influences allergen-induced IgE production and airway eosinophilia in a time and dose-dependent manner, preventing IgE production and development of eosinophilia when administered during allergen sensitization at high doses, and inducing exacerbation of eosinophilia when administered upon allergen challenge at low doses, including infra-clinical doses.