Impact of Siponimod on Enteric and Central Nervous System Pathology in Late-Stage Experimental Autoimmune Encephalomyelitis.

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS). Although immune modulation and suppression are effective during relapsing-remitting MS, secondary progressive MS (SPMS) requires neuroregenerative therapeutic options that act on the CNS. The sphingosine-1-phosphate receptor modulator siponimod is the only approved drug for SPMS. In the pivotal trial, siponimod reduced disease progression and brain atrophy compared with placebo. The enteric nervous system (ENS) was recently identified as an additional autoimmune target in MS. We investigated the effects of siponimod on the ENS and CNS in the experimental autoimmune encephalomyelitis model of MS. Mice with late-stage disease were treated with siponimod, fingolimod, or sham. The clinical disease was monitored daily, and treatment success was verified using mass spectrometry and flow cytometry, which revealed peripheral lymphopenia in siponimod- and fingolimod-treated mice. We evaluated the mRNA expression, ultrastructure, and histopathology of the ENS and CNS. Single-cell RNA sequencing revealed an upregulation of proinflammatory genes in spinal cord astrocytes and ependymal cells in siponimod-treated mice. However, differences in CNS and ENS histopathology and ultrastructural pathology between the treatment groups were absent. Thus, our data suggest that siponimod and fingolimod act on the peripheral immune system and do not have pronounced direct neuroprotective effects

Murine Esophagus Expresses Glial-Derived Central Nervous System Antigens

Multiple sclerosis (MS) has been considered to specifically affect the central nervous system (CNS) for a long time. As autonomic dysfunction including dysphagia can occur as accompanying phenomena in patients, the enteric nervous system has been attracting increasing attention over the past years. The aim of this study was to identify glial and myelin markers as potential target structures for autoimmune processes in the esophagus. RT-PCR analysis revealed glial fibrillary acidic protein (GFAP), proteolipid protein (PLP), and myelin basic protein (MBP) expression, but an absence of myelin oligodendrocyte glycoprotein (MOG) in the murine esophagus. Selected immunohistochemistry for GFAP, PLP, and MBP including transgenic mice with cell-type specific expression of PLP and GFAP supported these results by detection of (1) GFAP, PLP, and MBP in Schwann cells in skeletal muscle and esophagus; (2) GFAP, PLP, but no MBP in perisynaptic Schwann cells of skeletal and esophageal motor endplates; (3) GFAP and PLP, but no MBP in glial cells surrounding esophageal myenteric neurons; and (4) PLP, but no GFAP and MBP in enteric glial cells forming a network in the esophagus. Our results pave the way for further investigations regarding the involvement of esophageal glial cells in the pathogenesis of dysphagia in MS

Neurochemical classification of serotonin-immunoreactive neurons co-innervating motor endplates in the mouse esophagus

Serotonin immunoreactivity was previously found in myenteric neurons co-innervating motor endplates in the mouse esophagus striated muscle and aninvolvement in motility control was suggested. However, it is not known ifother neuroactive substances are present in these neurons and to what extentthey co-localize. First, vasoactive intestinal peptide (VIP) was established as abona fide marker for putative inhibitory myenteric neurons by evaluating co-localization with neuronal nitric oxide synthase (nNOS) and neuropeptide Y(NPY). Then, co-localization of serotonin and VIP was tested in co-innervatingaxons on motor endplates, which were visualized withα-bungarotoxin (α-BT)by multilabel immunofluorescence. Myenteric ganglia were also surveyed forco-localization in neuronal perikarya and varicosities. nNOS, NPY, and VIPwere completely co-localized in enteric co-innervating nerve terminals onmotor endplates. After co-staining with VIP, we found (a) serotonin (5-HT)-positive nerve endings without VIP (44% of 5-HT-positively innervated end-plates), (b) 5-HT- and VIP-positive endings without co-localization (35%), and(c) 5-HT- and VIP-positive endings with co-localization (21%). About one-fifthof nerve terminals on motor endplates containing 5-HT originate from putativeinhibitory peptidegic nitrergic neurons. However, the majority represents a different population presumably subserving different functions