Background Neuroinflammation and disrupted cholesterol metabolism in microglia are key contributors to Alzheimer’s disease (AD) pathogenesis. Liver X receptors (LXRα/β) regulate lipid metabolism and inflammation. Synthetic pan-LXR agonists, such as T0901317 and GW3965, exert neuroprotective effects by modulating lipid metabolism, making them promising therapeutic strategies for neurodegenerative disorders like Alzheimer’s Disease (AD). However, their clinical use is limited by hepatic side effects, including hypertriglyceridemia and steatosis. Purpose To overcome these limitations, we investigated 24(S)-saringosterol, a phytosterol from Sargassum fusiforme, and its potential dissociating effect as a LXR agonist on myeloid cells vs hepatocytes. Method Using primary cultures of myeloid cells (microglia, bone marrow-derived macrophages) and hepatocytes, we performed transcriptomic and lipidomic analyses to assess the impact of 24(S)-saringosterol on lipid metabolism and inflammatory pathways. Results 24(S)-saringosterol strongly activated LXR-regulated genes, upregulating cholesterol efflux transporter Abca1 in a dose-dependent manner. In myeloid cells, it reduced the expression of interferon-β pathway genes and promoted cholesterol efflux, mirroring GW3965’s anti-inflammatory effects. Notably, 24(S)-saringosterol downregulated cholesterol biosynthesis (Dhcr24) and influx (Ldlr) via Srebp2 in both cell types, contrasting with GW3965, which increased lipid synthesis genes via Srebp1. Conclusion These findings suggest 24(S)-saringosterol acts as a selective LXR agonist in a cell-specific manner, retaining beneficial effects while minimizing hepatic risks. This compound represents a promising candidate for AD and other metabolic or inflammatory disorders.

Proteins affect most physiological functions in cells and play a key role in maintaining cellular homeostasis. Protein function is tuned at multiple levels, including synthesis and degradation, intracellular localization, and posttranslational modifications (PTMs). PTMs are a key chemical modification for regulating protein biological activity, including protein-folding relevant shifts in electrical charge, protein-protein interactions, and hydrogen bond formation. Such chemical changes can also result in protein unfolding or denaturation. PTMs can target proteins across multiple cellular compartments, including the cell membrane, cytoplasm, nucleus, endoplasmic reticulum, and mitochondria, playing crucial roles in regulating cellular processes, and their dysregulation is linked to the development of numerous diseases. It is well-established that various autoimmune conditions, including multiple sclerosis (MS), are influenced by the PTMs of endogenous proteins. This review adopts an integrated perspective on PTMs, with a particular focus on the citrullination of myelin basic protein (MBP). It moves beyond the traditional view of MS solely as an autoimmune disease, highlighting the broader implications of this initial chemical event, including the role of PAD2, in early disease onset and progression.

This study examined the role of ST3GAL3 as a regulator of excitatory/inhibitory (E/I) synaptic homeostasis using a human iPSC-based model. Neurodevelopmental disorders (NDDs) are increasingly linked to disruptions in the E/I balance, yet the molecular determinants remain poorly defined. ST3GAL3, a sialyltransferase associated with both rare monogenic disorders, including intellectual disability and infantile epilepsy, and complex polygenic conditions, such as ADHD, represents a strong candidate gene for involvement in synaptic regulation. To investigate this, isogenic ST3GAL3 knockout (ST3GAL3 KO) and wildtype (WT) iPSC lines were generated through CRISPR/Cas9 editing and diRerentiated into cortical neurons using both directed and induced protocols. This dual strategy enabled robust comparisons across cellular contexts and minimised methodological bias. To this end, we conducted functional characterisation using microelectrode array (MEA) technology alongside transcriptomic profiling through RNA sequencing (RNAseq), directly comparing ST3GAL3 KO-derived neurons with their isogenic controls. Functional assays using MEA revealed aberrant bursting patterns, particularly prolonged burst durations and heightened variability in S3GAL3KO neurons. Complementary transcriptomic profiling performed via RNAseq demonstrated downregulation in ST3GAL3 KO lines of genes involved in cognition, memory, as well as glutamatergic and GABAergic synaptic plasticity and functionality, providing molecular evidence for widespread synaptic dysregulation. Together, these findings establish ST3GAL3 as a key regulator of E/I balance in the cortices, advancing current knowledge on the pathophysiological involvement of ST3GAL3 deficiencies in the development of NDDs.

Remyelination failure is a central obstacle in demyelinating diseases, yet the endogenous signals coordinating CNS repair remain poorly defined. Here we identify amphiregulin (AREG), an EGFR ligand produced by regulatory immune cells during peripheral tissue repair, as a coordinator of central nervous system remyelination. Systemic AAV delivery drives astrocytic AREG expression in the mouse brain and enhances myelin recovery following cuprizone-induced demyelination, with increased OPC density and reduced CD68hi microglia. Single-cell RNA sequencing shows that AREG reprograms CD68+ microglia toward a homeostatic, debris-clearing state and primes OPCs for differentiation through downregulation of developmental transcripts and upregulation of pro-differentiation genes. Cell–cell communication analysis nominates the microglia-to-oligodendrocyte Psap–GPR37 axis as a candidate effector, with GPR37 protein rescued by AREG. Recombinant AREG enhances the differentiation of human iPSC-derived OPCs. These findings establish astrocyte-delivered AREG as an endogenous coordinator of glial crosstalk during remyelination and identify EGFR-ligand signaling as an entry point for pro-remyelinating therapies.

Regulatory T cells are well known for their immunomodulatory role but also their regenerative function is increasingly recognized across multiple tissue types including skeletal muscle, lung and the central nervous system (CNS). In the CNS, Tregs have been shown to promote oligodendrocyte precursor cell differentiation and their interaction with microglia supports a pro-regenerative microenvironment in the brain. The suppressive function of Tregs is highly dependent on the epigenetic signature, specifically DNA methylation, of certain network of Treg-related genes including FOXP3, CTLA4, IKZF2, IKZF4 and TNFRSF18 which collectively function to maintain Treg cell lineage and stability. Whether a similar or the same epigenetic program also influences the regenerative capacity of Tregs remains unknown, and the methylation status of regenerative genes such as AREG, NT3, and osteopontin has never been characterized. Understanding the epigenetic regulation of Tregs in this respect is relevant for demyelinating disease such as multiple sclerosis (MS). MS is characterized by chronic neuroinflammation and neurodegeneration associated with remyelination failure. In MS, Treg suppressive function is compromised and epigenetic dysregulation at key Treg loci has been reported. However, the role of those loci in repair is unknown and the methylation status of regenerative genes in MS Tregs has not been studied. This review examines the evidence for Treg-mediated CNS repair, the epigenetic mechanisms controlling Treg identity, and highlights the epigenetic regulation of Treg regenerative genes as a critical gap in the field with potential implications for remyelination failure in MS.

Enhancing glutamate (Glu) uptake by positive allosteric modulation of excitatory amino acid transporter subtype 2 (EAAT2) is an attractive strategy to enable neuroprotection. However, while the EAAT field is rich in reports on inhibitors, enhancing EAAT2 protein dynamics is a much more difficult objective. A natural product approach reported that a spider venom HPLC fraction, number 10, showed neuroprotective effects. It was referred to as parawixin10, and later the structure of the key component responsible for the neuroprotective action was disclosed to be an N-acylamine, compound 2. We have resynthesized the N-acylamine 2 (parawixin10) and show here that this compound does not enhance Glu uptake in a wide range of radioligand binding assays, nor does it, in our hands, show any neuroprotective effect in a dose-dependent, statistically significant way.

Failure of remyelination drives neurodegeneration in demyelinating disorders such as multiple sclerosis (MS), with disrupted lipid handling and metabolic stress in oligodendrocyte precursor cells (OPCs) posing major barriers to repair. Here, we identify the dual ApoC-II mimetic- ApoC-III antagonist peptide D6PV as a metabolic modulator that directly enhances OPC differentiation and myelin repair. Across ex vivo and in vivo models of chemically induced demyelination, D6PV promotes oligodendrocyte maturation and restores myelin integrity independently of lipoprotein hydrolysis or modulation of lipid droplet-containing phagocytes. Guided by transcriptomics analyses, we find that D6PV stimulates mitochondrial oxidative phosphorylation and fatty acid β-oxidation, while suppressing inflammatory transcriptional programs, thereby driving OPCs toward a myelinating phenotype. Notably, D6PV does not alter peripheral immune composition or autoimmune-driven pathology in the experimental autoimmune encephalomyelitis model, indicating a central nervous system (CNS) cell-autonomous effect. These findings reveal a metabolism-linked pathway for remyelination and position D6PV as a promising therapeutic strategy to enhance CNS repair in demyelinating diseases. ### Competing Interest Statement The authors have declared no competing interest. Fondation Charcot, https://ror.org/04j8wah18, CHARCO24VS, CHARCOT23JB, CHARCOT24JB, CHARCOT25JB, CHARCOT25JH FWO Vlaanderen, 11PAO24N, 1SH6G24N, 1104425N, 1245724N, 12B1I24N, 1210026N FWO Vlaanderen, G0A7922, G0A7922, S01623N, G075823, G0A3B24 MS Liga Vlaanderen, MSLIGABOGJ GSKE, GSKE-BOGJ special research fund Hasselt University, 22DOC38BOF, 23INC06BOF

Air pollution has been implicated in various adverse health effects, including neurodevelopmental and neurodegenerative impairments. However, the long-term impact of early-life ultrafine particle (UFP) exposure on the brain remains poorly understood. Using a sequential exposure mouse model, we investigated how early-life ultrafine carbonaceous particles (UFPC) exposure programs neurobehavioural and molecular vulnerability upon adult re-exposure. Wild-type C57BL/6J mice were exposed to either HEPA-filtered air or UFPC during the prenatal (gestational days 8-9 and 16-17) and/or postnatal periods (postnatal days 4-7 and 10-13), followed by a 4-day re-exposure in adulthood (postnatal days 142-145). Behavioural assessments revealed hippocampus-dependent spatial memory deficits and anxiolytic-like behaviour following cumulative exposure. Brain proteomic analysis identified reduced protein levels of key modulators of synaptic signalling and neurovascular homeostasis (Erbb4 and Ddah1), accompanied by gene-specific promoter methylation changes and shortened telomere length, indicating persistent epigenetic reprogramming and accelerated cellular aging. We validated the epigenetic sensitivity of ERBB4 to prenatal air pollution in human cord blood from the ENVIRONAGE birth cohort. The integrative design, encompassing behavioural phenotyping and molecular profiling, offers a comprehensive systems-level perspective on the neurobiological effects of UFPC. Our findings suggest that developmental UFPC exposure induces increased susceptibility to re-exposure on behavioural, epigenetic, and proteomic outcomes. This work provides evidence for UFP as a potentially critical environmental determinant of brain health throughout life.

Abstract Alzheimer’s disease (AD) is characterized not only by amyloid-β plaques, tau neurofibrillary tangles and associated neuronal loss, but also by alterations in non-neuronal cell types essential for neuronal support. Oligodendrocytes and their myelin sheaths play a central role in maintaining axonal function, yet detailed molecular profiling of myelin dynamics in the human AD brain remains limited. Although neuroimaging studies increasingly highlight myelin degeneration in white matter as an important contributor to AD pathophysiology, the status of myelin within cortical grey matter is less well understood. Here, we performed a detailed histopathological characterization of myelin integrity and oligodendrocyte dynamics in both grey and white matter of the middle temporal gyrus (MTG), making use of post-mortem tissue from AD cases (n = 15) and age-, sex-, and APOE genotype–matched controls (n = 15). Strikingly, we identified a specific vulnerability of cortical grey matter myelin in AD, whereas white matter myelin appeared relatively preserved. This selective grey matter disruption was accompanied by a seemingly insufficient oligodendrocyte regenerative response, suggesting ongoing attempts at myelin repair, yet featured by a differentiation block. Importantly, the extent of myelin damage and OPC differentiation strongly correlated with proximity to tau pathology, linking cortical demyelination to neuronal and synaptic dysfunction within vulnerable AD regions. Together, our findings reveal cortical grey matter myelin disruption as a previously underrecognized, highly localized feature of AD pathology. By highlighting the tight intertwining of oligodendrocyte and myelin dynamics with tau-associated neurodegeneration, this work positions cortical myelin pathology as a potential new mechanistic and therapeutic avenue in AD.

Increasing evidence suggests that myelin dysfunction and oligodendrocyte pathology are active contributors to neurodegeneration. In Alzheimer's disease (AD), the link between tau aggregation and myelin integrity remains unclear, despite the preferential emergence of tau pathology in late-myelinating regions. Here, we propose a myelin-centered framework for tau pathology based on three mechanisms. First, vulnerability of late-myelinating oligodendrocytes may drive myelin breakdown, metabolic stress, and axonal dysfunction, promoting tau hyperphosphorylation. Second, microglial responses to myelin injury may become maladaptive, with lipid overload impairing tau clearance. Third, oligodendrocytes may act as conditional reservoirs facilitating tau propagation across myelinated networks. Together, these processes suggest that myelin loss may contribute to tau accumulation, clearance deficits, and spread, providing a framework for future experimental testing. This perspective highlights myelin biology as a source of new conceptual insights and therapeutic strategies in AD.

Stroke remains the second leading cause of death worldwide, highlighting the urgent need for novel treatment options. Phosphodiesterase 4 (PDE4) inhibition has been shown to reduce neuroinflammation and improve neurological outcomes in several neurodegenerative diseases, such as multiple sclerosis. Especially the PDE4B gene is known to contribute to the inflammatory reaction. Therefore, we investigated the effects of PDE4 and PDE4B inhibition in ischemic stroke. We used the distal middle cerebral artery occlusion (dMCAO) mouse model to assess inflammatory cell infiltration and lesion size, and complemented the data with in vitro studies on neutrophils. Our results show that prophylactic PDE4 and PDE4B inhibition reduced the lesion size and neutrophil infiltration in vivo, whereas post-stroke administration of these inhibitors did not show an effect. In vitro, neutrophil activation was decreased following PDE4 and PDE4B inhibition. Furthermore, spatial proteomics analysis of the ischemic brain identified C1QBP as a potential contributing factor to the beneficial effects of prophylactic PDE4B inhibition. Taken together, our research provides evidence for a potential role of prophylactic, but not acute PDE4 and PDE4B inhibition in ischemic stroke treatment, especially for patients at risk of recurrent stroke.

BACKGROUND AND PURPOSE:Liver X receptors (LXRs) are promising therapeutic targets for alleviating Alzheimer's disease (AD) symptoms. We assessed the impact of the semi-synthetic LXR agonist 22-ketositosterol on disease progression in an AD mouse model. EXPERIMENTAL APPROACH:From 5.5 months of age, APPswePS1ΔE9 (AD) mice and wild-type (WT) littermates received a regular or 22-ketositosterol-supplemented diet (0.017% w/w). Cognition was assessed with object location and recognition tasks and a spontaneous alternation Y-maze test. Amyloid β was quantified using immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA), microglia (Iba1, CD68) and astrocyte (GFAP) markers using IHC. Sterols were determined in food, serum, liver and cerebellum. KEY RESULTS:22-Ketositosterol activated both liver X receptors-α and -β and promoted cholesterol efflux in cell cultures. Diet supplementation with 22-ketositosterol prevented a decline in the performance of APPswePS1ΔE9 mice in the object location task but not in the other two tasks. Without affecting amyloid β deposition, 22-ketositosterol decreased microglia (Iba1, CD68) and astrocyte (GFAP) markers in the cortex and hippocampus of APPswePS1ΔE9, suggesting potential anti-inflammatory effects. No lipid accumulation was detected in the liver or serum upon 22-ketositosterol supplementation. CONCLUSIONS AND IMPLICATIONS:Diet supplementation with 22-ketositosterol prevented the decline in spatial memory of APPswePS1ΔE9 mice. Our data suggest therapeutic benefits of 22-ketositosterol possibly by enhancing cholesterol efflux and mitigating inflammatory responses, without inducing hepatosteatosis or hypertriglyceridemia.

Modulating the excitatory amino acid transporter 3 (EAAT3) can be considered a novel approach for the treatment of multiple sclerosis (MS). EAAT3 plays a crucial role in regulating oxidative stress and oligodendrocyte function through its ability to transport cysteine, the rate-limiting building block in the synthesis of the antioxidant glutathione. Therefore, EAAT3 activation is hypothesised to improve oligodendrocyte health and relieve its differentiation block in MS, improving remyelination capacity. Using a cuprizone-induced demyelination model, the effects of EAAT3 overexpression by viral transduction of oligodendrocytes and pharmacological inhibition of EAAT3 were examined. Surprisingly, EAAT3 overexpression significantly hampered remyelination, while EAAT3 inhibition prevented demyelination and improved functional remyelination as assessed by visual evoked potentials and post mortem myelin basic protein fluorescent staining. Next, cellular mechanisms underlying these results were investigated. Consistent with the in vivo findings, post mortem gene expression analysis of the corpus callosum of cuprizone treated animals revealed a trend towards upregulation of oligodendrocyte lineage genes in response to EAAT3 inhibition, supporting its role in oligodendrocyte health and myelination processes. In vitro studies using the human oligodendroglioma (HOG) cell line demonstrated the beneficial effects of EAAT3 inhibition on cellular morphology, indicating potential roles in promoting oligodendrocyte maturation and myelination. In contrast, EAAT3 overexpression appears to hamper these processes. These findings suggest that, contrary to our initial hypothesis, EAAT3 inhibition could improve oligodendrocyte function and myelination processes, highlighting its potential as a therapeutic target for demyelinating disorders. Future studies should address the exact molecular mechanism through which this effect is obtained.

Synaptic dysfunction is a hallmark of neurodevelopmental disorders (NDDs), often linked to genes involved in cytoskeletal regulation. While the role of these genes has been extensively studied in neurons, microglial functions such as phagocytosis are also dependent on cytoskeletal dynamics. We demonstrate that disturbance of actin cytoskeletal regulation in microglia, modeled by genetically impairing the scaffold protein Disrupted-in-Schizophrenia 1 (DISC1), which integrates actin-binding proteins, causes a shift in actin regulatory balance favoring filopodial versus lamellipodial actin organization. The resulting microglia-specific dysregulation of actin dynamics leads to excessive uptake of synaptic proteins. Genetically engineered DISC1-deficient mice show diminished hippocampal excitatory transmission and associated spatial memory deficits. Reintroducing wild-type microglia-like cells via bone marrow transplantation in adult DISC1-deficient mice restores the synaptic function of neurons and rescues cognitive performance. These findings reveal a pivotal role for microglial actin cytoskeletal remodeling in preserving synaptic integrity and cognitive health. Targeting microglial cytoskeletal dynamics may effectively address cognitive impairments associated with NDDs, even in adulthood.

Marine sterols from brown seaweeds, particularly fucosterol and its oxidized derivative saringosterol, have shown therapeutic potential for Alzheimer’s disease (AD) and cardiovascular diseases. Here, we aimed to elucidate the cellular and in vivo mechanisms underlying their beneficial effects. In human HepG2 hepatocytes and CCF-STTG1 astrocytoma cells, we assessed liver x receptor (LXRα /LXRβ) activation, sterol uptake, and effects on cholesterol metabolism using luciferase reporter assays, GC–MS sterol profiling, and 13C-acetate incorporation. In THP-1–derived macrophages, we evaluated sterol-induced cholesterol efflux using radiolabeled [3H]-cholesterol assays and characterized anti-inflammatory responses by quantifying lipopolysaccharide (LPS) -induced cytokine production. Wild-type C57BL/6J mice were fed diets enriched with either fucosterol (0.2% w/w) or saringosterol (0.02% w/w) for 7 days, after which sterol profiles in serum, liver, and brain were quantified by GC–MS. Hippocampal transcriptional responses were assessed by RNA sequencing. Both fucosterol and saringosterol were internalized by HepG2 and CCF-STTG1 cells and activated LXRα/β, but elicited distinct metabolic effects: fucosterol increased cholesterol synthesis and intracellular desmosterol, whereas saringosterol reduced both; only saringosterol suppressed LPS-induced interleukin (IL)-6 and tumor necrosis factor (TNF)-α production in macrophages, while both enhanced cholesterol efflux. In vivo, fucosterol somewhat elevated hepatic desmosterol and decreased 5α-cholestanol and circulating oxysterols, whereas saringosterol also increased hepatic desmosterol and elevated 7α-hydroxycholesterol in liver and brain as well as serum 27-hydroxycholesterol. Transcriptome analysis revealed that fucosterol primarily modulated synaptic signaling and hormonal pathways linked to neuronal plasticity, while saringosterol affected protein quality control and neurodegenerative pathways. These data are the first on the direct comparison of the cellular and in vivo effects of fucosterol and saringosterol, revealing shared LXR activation but divergent impacts on hepatic, brain and systemic cholesterol metabolism and expression of genes involved in neural pathways, indicating complementary neuroprotective effects with therapeutic potential for AD and related disorders.

Liver X Receptors (LXRα and LXRβ) are nuclear receptors that regulate various metabolic processes via transcriptional regulation, including lipid and cholesterol homeostasis. Recent evidence highlights the involvement of LXR activation in myelin synthesis and maintenance. Given the essential role of myelin in neuronal communication, its loss in disorders such as multiple sclerosis and Alzheimer's disease underlines the urgent need for effective remyelinating therapies. Restoring the functions of oligodendrocytes to stimulate remyelination offers an interesting approach to protect neurons and slow down neurodegeneration. LXRs have been suggested as potential therapeutic targets in demyelinating disorders as they can promote cholesterol turnover and reduce inflammation, creating a favorable environment for remyelination. Furthermore, activation of LXR directly enhances remyelination by inducing myelin genes. Since various literature and research describe the potential neuroprotective and (re)myelinating benefits of LXR, this review discusses the role of the LXR pathway in (re)myelinating strategies. It highlights the pharmacological compounds for LXR activation, as well as naturally occurring LXR agonists with potential therapeutic value for promoting remyelination.

Charcot-Marie-Tooth disease type 1A (CMT1A) is an inherited peripheral neuropathy caused by a duplication of the peripheral myelin protein 22 (PMP22) gene. It is primarily marked by Schwann cell dedifferentiation and demyelination, leading to motor and sensory deficits. Cyclic adenosine monophosphate (cAMP) is crucial for Schwann cell differentiation and maturation. Therefore, increasing cAMP by inhibiting its degraders, phosphodiesterases (PDE), is a potential therapeutic strategy for CMT1A. This study investigated the therapeutic potential of the specific PDE4D inhibitor Gebr32a using the C3-PMP22 mouse model for CMT1A and patient-induced Pluripotent Stem Cell (iPSC)-derived Schwann cells. C3-PMP22 mice, injected subcutaneously with Gebr32a twice a day for 10 weeks, showed significantly increased nerve conduction in sciatic nerves compared to vehicle-injected controls, indicating improved myelination. Additionally, Gebr32a-treated C3-PMP22 mice exhibited improved sensorimotor functions. Grip strength analysis revealed significantly increased strength in all limbs of Gebr32a-treated C3-PMP22 mice. Post-mortem histological and ultrastructural analysis confirmed enhanced myelination in the sciatic nerve of treated mice compared to controls. In primary mouse CMT1A Schwann cells, Gebr32a dose-dependently increased the expression of pro-myelinating genes such as oct6, Krox20, Mbp, Mpz, and Plp, while downregulating the dedifferentiation marker c-Jun and human PMP22. Similar effects on gene expression were observed in iPSC-derived Schwann cells from a CMT1A patient, highlighting the clinical relevance of our findings. In conclusion, inhibition of PDE4D with Gebr32a improves the functional and molecular outcomes in mouse and human models of CMT1A, highlighting its potential as a new therapeutic strategy for CMT1A disease management.

Inhibition of the cyclic‐AMP degrading enzyme phosphodiesterase type 4 (PDE4) in the brains of animal models is protective in Alzheimer's disease (AD). We show for the first time that enzymes from the subfamily PDE4D not only colocalize with beta‐amyloid (Aβ) plaques in a mouse model of AD but that Aβ directly associates with the catalytic machinery of the enzyme. Peptide mapping suggests that PDE4D is the preferential PDE4 subfamily for Aβ as it possesses a unique binding site. Intriguingly, exogenous addition of Aβ to cells overexpressing the PDE4D5 longform caused PDE4 activation and a decrease in cAMP. We suggest a novel mechanism where PDE4 longforms can be activated by Aβ, resulting in the attenuation of cAMP signalling to promote loss of cognitive function in AD.

Spinal cord injury (SCI) is a life-changing event that severely impacts the patient's quality of life. Modulating neuroinflammation, which exacerbates the primary injury, and stimulating neuro-regenerative repair mechanisms are key strategies to improve functional recovery. Cyclic adenosine monophosphate (cAMP) is a second messenger crucially involved in both processes. Following SCI, intracellular levels of cAMP are known to decrease over time. Therefore, preventing cAMP degradation represents a promising strategy to suppress inflammation while stimulating regeneration. Intracellular cAMP levels are controlled by its hydrolyzing enzymes phosphodiesterases (PDEs). The PDE4 family is most abundantly expressed in the central nervous system (CNS) and its inhibition has been shown to be therapeutically relevant for managing SCI pathology. Unfortunately, the use of full PDE4 inhibitors at therapeutic doses is associated with severe emetic side effects, hampering their translation toward clinical applications. Therefore, in this study, we evaluated the effect of inhibiting specific PDE4 subtypes (PDE4B and PDE4D) on inflammatory and regenerative processes following SCI, as inhibitors selective for these subtypes have been demonstrated to be well-tolerated. We reveal that administration of the PDE4D inhibitor Gebr32a, even when starting 2 dpi, but not the PDE4B inhibitor A33, improved functional as well as histopathological outcomes after SCI, comparable to results obtained with the full PDE4 inhibitor roflumilast. Furthermore, using a luminescent human iPSC-derived neurospheroid model, we show that PDE4D inhibition stabilizes neural viability by preventing apoptosis and stimulating neuronal differentiation. These findings strongly suggest that specific PDE4D inhibition offers a novel therapeutic approach for SCI.

Sphingosine-1-phosphate receptor (S1PR) modulators are clinically used to treat relapse-remitting multiple sclerosis (MS) and the early phase of progressive MS when inflammation still prevails. In the periphery, S1PR modulators prevent lymphocyte egress from lymph nodes, hence hampering neuroinflammation. Recent findings suggest a role for S1PR modulation in remyelination. As the Gi alpha-coupled S1P1 subtype is the most prominently expressed S1PR in oligodendrocyte precursor cells (OPCs), selective modulation (functional antagonism) of S1P1 may have direct effects on OPC functionality. We hypothesized that functional antagonism of S1P1 by ponesimod induces remyelination by boosting OPC differentiation. In the cuprizone mouse model of demyelination, we found ponesimod to decrease the latency time of visual evoked potentials compared to vehicle conditions, which is indicative of functional remyelination. In addition, the Y maze spontaneous alternations test revealed that ponesimod reversed cuprizone-induced working memory deficits. Myelin basic protein (MBP) immunohistochemistry and transmission electron microscopy of the corpus callosum revealed an increase in myelination upon ponesimod treatment. Moreover, treatment with ponesimod alone or in combination with A971432, an S1P5 monoselective modulator, significantly increased primary mouse OPC differentiation based on O4 immunocytochemistry. In conclusion, S1P1 functional antagonism by ponesimod increases remyelination in the cuprizone model of demyelination and significantly increases OPC differentiation in vitro. Ponesimod reverses a cuprizone-induced working memory deficit, restores the cuprizone-induced delay in latency time of the optic pathway, and enhances remyelination after cuprizone intoxication in vivo. Furthermore, ponesimod enhances differentiation of oligodendrocyte precursor cells into mature oligodendrocytes in vitro.image