A Defined Diet Combined with Sonicated Inoculum Provides a High Incidence, Moderate Severity Form of Experimental...

Myelin oligodendrocyte glycoprotein 35-55 (MOG_35-55)-peptide induced experimental autoimmune encephalomyelitis (EAE) is a model for inflammation of the brain and spinal cord. However, its severity and incidence vary within and between laboratories. Severe scores can lead to premature termination and are both unnecessary for readouts and detrimental to animal welfare. Ideally, the model would have high incidence, moderate severity, and low interindividual variability to fulfill the "Refine" aspect of the 3R concept. Nevertheless, most efforts to increase incidence also increase the severity. When the effects of potential therapies are tested, moderate severity is sufficient to detect useful drug effects as long as variation is low. Low variation can also reduce group sizes, which supports the "Reduce" aspect of 3R approaches in disease modeling. We set out to reduce variation and control severity by assessing the effects of mouse age, dietary fiber, antigen emulsion, and the dose of MOG and pertussis toxin on incidence, variability, and severity in the MOG-EAE model.

We compared 14- and 33-week-old female C57BL/6 mice and varied the diet and inoculum in two studies. We measured disease signs in vivo as well as gene expression in the brain and spinal cord and histology by immunofluorescence. Ordinary one-way ANOVA was used for multiple comparisons.

The most reliable induction conditions were with a low-fermentative/fiber diet (AIN 93M) combined with a sonicated emulsion of the MOG_35-55-peptide. High-dose pertussis toxin increased EAE severity and incidence in 14-week-old mice (25% survival) while being more moderate in mature mice (100% survival). Varying all parameters suggests that duration of prefeeding defined diet, emulsion quality, and mouse maturity were factors that increase uniformity of response allowing incidence to reach 100% without excess severity. Microglia and astrocyte-associated markers were upregulated proportionally to score consistent with known EAE pathology.

A defined fiber/high-sugar diet with sonicated inoculum provides for a moderate severity, high incidence, and less variable EAE. The resulting uniformity in animal response and associated cytokine patterns, and the strong link to a defined diet, suggest that this may be a more clinically translatable protocol for the induction of EAE. This is consistent with reported effects of low-fermentable diets on immune modulation in human patients with autoimmune diseases.