Changes in the intestinal microbiome and disturbances in intestinal structure in rats with different severity of experimental allergic encephalomyelitis

Multiple sclerosis (MS) is a chronic autoimmune disorder of the central nervous system, characterized by inflammatory demyelination and neurodegeneration. Increasing attention has been directed to the “microbiota-gut-brain” axis, as intestinal barrier dysfunction and alterations in gut microbiota composition have been reported to precede the clinical onset of MS, although their association with disease severity remains uncertain. Experimental autoimmune encephalomyelitis (EAE) in rodents is a well-established model that reproduces the key immunological and morphological features of MS. However, comparative investigations addressing both the taxonomic structure of the microbiota and intestinal morphological changes across different severities of EAE remain scarce. To compare the taxonomic composition of the gut microbiota and structural alterations of the intestinal epithelium in rats with different severities of experimental autoimmune encephalomyelitis (EAE). The study was conducted on 39 female Wistar rats (intact, n=9; EAE-induced, n=30). EAE was induced by a single subcutaneous injection of spinal cord homogenate in complete Freund’s adjuvant. Morphological and ultrastructural changes in the intestinal epithelium were assessed using histological techniques and electron microscopy. The taxonomic composition of the gut microbiota was analyzed by 16S rRNA gene sequencing. When comparing the intestinal microbiome of mildly and severely ill rats, significant differences were observed in the representation of 4 phyla: Bacillota, Bacteroidota, Actinomycetota and Cyanobacteriota. In the baseline microbiome of severely affected rats, the abundance of the class Epsilonproteobacteria (Pseudomonadota) was increased. Clinical symptoms developed in both groups against the background of a decreased number of Bacteroidota (family Prevotellaceae). During the recovery phase, the abundance of Bacteroidota was restored only in mildly affected and recovered animals, whereas in rats with severe EAE the proportion of Actinomycetota increased. At the genus level, distinct group-specific dynamics during EAE were identified for 9 genera (unclassified Clostridiaceae and Peptoscreptococaceae, Allobaculum, Turicibacter, Prevotella, Parabacteroides, Desulfovibrio and Bifidobacterium). Alongside more pronounced alterations in the gut microbiome, rats with severe EAE displayed significant morphological changes in the intestine. At the peak of the disease, leukocyte infiltration of the mucosa, reduction in goblet cells numbers, shortened and thickened small intestinal villi and decreased crypt depth in the colon were observed, all correlating with the severity of the disease. Ultrastructural analysis further revealed mitochondrial and microvilli damage in enterocytes and colonocytes, which was more pronounced in severely affected animals. These findings support the involvement of the “microbiota-gut-brain” axis in the development of autoimmune demyelinating CNS disorders. The observed association between intestinal morphological alterations, gut microbiota composition, and EAE severity highlights the potential of targeting intestinal inflammation and barrier function for therapeutic strategies in multiple sclerosis

experimental allergic encephalomyelitis, multiple sclerosis, gastrointestinal tract, intestinal microbiota, goblet cells, inflammation

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