Betaine Attenuates Cuprizone-Induced Cortical Demyelination: The Role of the WNT4/mTOR Signalling Pathway in Regulating Astrogliosis and Synaptic Integrity.

Cortical grey matter pathology manifests early demyelinating diseases and is a primary driver of permanent neurological impairment. Our study investigated the neuroprotective effects of betaine against cuprizone (CPZ)-induced damage to the cerebral cortex in rats, focusing on its ability to modulate the upstream WNT4/mTOR signalling pathway. Cortical tissues were collected for biochemical assessments of oxidative and inflammatory markers, real-time PCR analysis for WNT4, GFAP, and mTOR gene expressions, histopathological examination, and immunohistochemical analysis of vimentin, S100, and synaptophysin. Exposure to CPZ resulted in severe cortical damage with profound oxidative stress. This oxidative status triggered a pronounced neuroinflammatory response, leading to elevated levels of IL-17A, IL-1β, and TNF-α, which in turn caused chronic, hyper-reactive astrogliosis. This was confirmed by the upregulation of GFAP gene expression and enhanced immunohistochemical staining of vimentin and S100β, resulting in extensive loss of cortical synaptic integrity (depletion of synaptophysin). Importantly, treatment with betaine served as a molecular brake, significantly downregulating the aberrant gene expressions of WNT4 and mTOR. This targeted downregulation effectively reduced the overactive glial response, leading to a significant reduction in astrogliosis (GFAP, vimentin, and S100β), alleviated the pro-inflammatory cytokines, restored endogenous antioxidant functions, and ultimately preserved cortical synaptophysin expression and cellular structure. Overall, these results demonstrate that betaine exerts a neuroprotective effect against CPZ-induced cortical injury, which is strongly associated with the attenuation of WNT4/mTOR signalling pathway hyperactivation, suppression of astrogliosis, and preservation of synaptic density. The preservation of cortical grey matter integrity is directly associated with alleviating cognitive deficits and long-term physical disabilities in multiple sclerosis patients, emphasising the translational relevance of targetable neuroprotective therapies.