Investigating the Alleviation of Disease Progression in EAE (Experimental Autoimmune Encephalomyelitis) Model Mice by Dexmedetomidine Combined with Remimazolam via Endoplasmic Reticulum Stress-Induced Autophagy
YIN Hao1, WU Wen2*
This study was to investigate the neuroprotective effects and mechanisms of Dex (dexmedetomidine) combined with Rem (remimazolam) on LPS (lipopolysaccharide)-induced BV2 microglial cell injury and EAE (experimental autoimmune encephalomyelitis) model mice. In vitro, LPS (1 μg/mL) was used to stimulate BV2 cells to establish an inflammatory model. In vivo, an EAE model was established in C57BL/6 mice immunized with MOG35-55. Cell viability was detected by CCK8 assay; migration and invasion capabilities were evaluated by scratch wound healing and Transwell assays; apoptosis rate and inflammatory factors [IL-1β (interleukin-1β)/IL-6/TNF-α (tumor necrosis factor-α)] were analyzed by flow cytometry and ELISA; endoplasmic reticulum stress markers [Bip (immuno- globulin heavy chain-binding protein)/CHOP (CCAAT enhancer binding protein homologous protein)/IRE1α (inositol-requiring enzyme 1α)] and autophagy-related proteins [LC3II/I (microtubule-associated protein 1 light chain 3 II/I), PINK1 (PTEN-induced kinase 1), Parkin, p62 (sequestosome 1), Beclin1] were examined by confocal microscopy and Western blot; and spinal cord histopathology, demyelination, and apoptosis were assessed by HE (hematoxylineosin), LFB, and TUNEL staining. Dex monotherapy at 9 μmol/L significantly increased cell viability (P<0.001), and combination with Rem (90 μg/mL) further enhanced viability (P<0.01). The Dex+Rem group showed improved cell wound healing (P<0.01) and invasion capacity (P<0.05) compared with the Dex or Rem alone groups. Compared with the LPS group, the Dex+Rem group exhibited a reduced apoptosis rate (P<0.001), decreased release of inflammatory factors (IL-1β, IL-6, TNF-α; P<0.001), downregulated expression of endoplasmic reticulum stress proteins (Bip, CHOP, IRE1α; P<0.001), and activated autophagy flux (increased protein expression of LC3II/I, PINK1, Parkin, Beclin1, and decreased p62 expression; P<0.001). Tun (Tunicamycin) reversed all these processes. In vivo, compared with the Model group, the Dex+Rem group significantly improved the neurological function score in EAE mice (P<0.01), alleviated spinal cord neuronal apoptosis and demyelination, concurrently inhibited the expression of spinal cord endoplasmic reticulum stress proteins (Bip, CHOP, IRE1α), and promoted autophagy (increased LC3II/I and Beclin1 protein expression, decreased p62 expression; P<0.001). The combination of dexmedetomidine and remimazolam effectively mitigates LPS-induced cellular damage and delays the neuropathological progression in the EAE model by synergistically inhibiting endoplasmic reticulum stress, activating autophagy, and reducing inflammation. This combination provides a novel strategy for the clinical treatment of multiple sclerosis.
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