Martorelli Mariella, Dengler Matthias, Laux Julian, Fischer Tina, Vaiceliunaite Agne, Hahn Ulrike, Weinstein Thilo, Cruces Santiago, Pokoj Christina, de Oliveira da Cunha Luciano, Wohlbold Lara, Koch Pierre, Laufer Stefan, Burnet Michael, Maier Florian
Synovo GmbH, Paul-Ehrlich-Straße 15, 72076 Tübingen, Germany.
Department of Pharmaceutical/Medicinal Chemistry, Eberhard Karls University Tübingen, Auf der Morgenstelle 8, 72076 Tübingen, Germany.
ACS Pharmacol Transl Sci. 2024 Nov 5;7(12):3827-3845. doi: 10.1021/acsptsci.4c00189. eCollection 2024 Dec 13.
Myelin oligodendrocyte glycoprotein 35-55 (MOG)-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-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.
髓鞘少突胶质细胞糖蛋白35 - 55（MOG）肽诱导的实验性自身免疫性脑脊髓炎（EAE）是一种脑和脊髓炎症模型。然而，其严重程度和发病率在不同实验室之间以及同一实验室内部存在差异。严重的评分可能导致实验过早终止，这对于实验结果的读取既无必要，又对动物福利有害。理想情况下，该模型应具有高发病率、中等严重程度和低个体间变异性，以符合3R概念中的“优化”方面。然而，大多数提高发病率的努力也会增加严重程度。在测试潜在疗法的效果时，只要变异性低，中等严重程度就足以检测到有效的药物作用。低变异性还可以减少样本量，这支持了疾病建模中3R方法的“减少”方面。我们通过评估小鼠年龄、膳食纤维、抗原乳剂以及MOG和百日咳毒素剂量对MOG - EAE模型发病率、变异性和严重程度的影响，来降低变异性并控制严重程度。
我们比较了14周龄和33周龄的雌性C57BL/6小鼠，并在两项研究中改变了饮食和接种物。我们在体内测量了疾病体征，并通过免疫荧光测量了脑和脊髓中的基因表达以及组织学情况。采用普通单因素方差分析进行多重比较。
最可靠的诱导条件是低发酵/纤维饮食（AIN 93M）与MOG肽的超声乳化液相结合。高剂量百日咳毒素增加了14周龄小鼠的EAE严重程度和发病率（存活率25%），而在成熟小鼠中则较为温和（存活率100%）。对所有参数的变化进行分析表明，预喂特定饮食的持续时间、乳化液质量和小鼠成熟度是增加反应一致性的因素，可使发病率达到100%且不过度严重。小胶质细胞和星形胶质细胞相关标志物的上调与评分成比例，这与已知的EAE病理学一致。
特定的纤维/高糖饮食与超声接种物相结合可产生中等严重程度、高发病率且变异性较小的EAE。动物反应和相关细胞因子模式的一致性，以及与特定饮食的紧密联系，表明这可能是一种更具临床可转化性的EAE诱导方案。这与低发酵饮食对自身免疫性疾病人类患者免疫调节的报道效果一致。
