Mechanical properties of the brain such as intracranial pressure or stiffness of the matrix play an important role in the brain’s normal physiology and pathophysiology. The physical properties are sensed by the cells through mechanoreceptors and translated into ion currents which activate multiple biochemical cascades allowing the cells to adapt and respond to changes in their microenvironment. Piezo1 is one of the first identified mechanoreceptors. It modulates various central nervous system functions such as axonal growth or activation of astrocytes. Piezo1 signaling was also shown to play a role in the pathophysiology of Alzheimer’s disease. Here, we explore the expression of the mechanoreceptor Piezo1 in human MO3.13 oligodendrocytes and human MS/non-MS patients’ brains and investigate its putative effects on oligodendrocyte proliferation, maturation, and migration. We found that Piezo1 is expressed in human oligodendrocytes and oligodendrocyte progenitor cells in the human brain and that its inhibition with GsMTx4 leads to an increment in proliferation and migration of MO3.13 oligodendrocytes. Activation of Piezo1 with Yoda-1 induced opposite effects. Further, we observed that expression of Piezo1 decreased with MO3.13 maturation in vitro. Differences in expression were also observed between healthy and multiple sclerosis brains. Remarkably, the data showed significantly lower expression of Piezo1 in the white matter in multiple sclerosis brains compared to its expression in the white matter in healthy controls. There were no differences in Piezo1 expression between the white matter plaque and healthy-appearing white matter in the multiple sclerosis brain. Taken together, we here show that Piezo1-induced signaling can be used to modulate oligodendrocyte function and that it may be an important player in the pathophysiology of multiple sclerosis.

BackgroundAdenosine deaminase (ADA) via two isoenzymes, ADA1 and ADA2, regulates intra- and extracellular adenosine concentrations by converting it to inosine. In the central nervous system (CNS), adenosine modulates the processes of neuroinflammation and demyelination that together play a critical role in the pathophysiology of multiple sclerosis (MS). Except for their catalytic activities, ADA isoenzymes display extra-enzymatic properties acting as an adhesion molecule or a growth factor.AimsThis study aimed to explore the distribution and activity of ADA1 and ADA2 in the plasma and the CSF of MS patients as well as in the human brain microvascular endothelial cells (HBMEC), human brain vascular pericytes and human astrocytes.Methods and resultsThe enzyme assay following reverse phase-high performance liquid chromatography (HPLC) analysis was used to detect the ADA1 and ADA2 activities and revealed an increased ratio of ADA1 to ADA2 in both the plasma and the CSF of MS patients. Plasma ADA1 activity was significantly induced in MS, while ADA2 was decreased in the CSF, but significance was not reached. The brain astrocytes, pericytes and endothelial cells revealed on their surface the activity of ADA1, with its basal level being five times higher in the endothelial cells than in the astrocytes or the pericytes. In turn, ADA2 activity was only observed in pericytes and endothelial cells. Stimulation of the cells with pro-inflammatory cytokines TNFα/IL17 for 18 h decreased intracellular nucleotide levels measured by HPLC only in pericytes. The treatment with TNFα/IL17 did not modulate cell-surface ATP and AMP hydrolysis nor adenosine deamination in pericytes or astrocytes. Whereas in endothelial cells it downregulated AMP hydrolysis and ADA2 activity and upregulated the ADA1, which reflects the ADA isoenzyme pattern observed here in the CSF of MS patients.ConclusionIn this study, we determined the impaired distribution of both ADA isoenzymes in the plasma and the CSF of patients with MS. The increased ADA1 to ADA2 ratio in the CSF and plasma may translate to unfavorable phenotype that triggers ADA1-mediated pro-inflammatory mechanisms and decreases ADA2-dependent neuroprotective and growth-promoting effects in MS.

Acidosis is one of the hallmarks of demyelinating central nervous system (CNS) lesions in multiple sclerosis (MS). Response to acidic pH is primarily mediated by a family of G protein-coupled proton-sensing receptors: OGR1, GPR4, and TDAG8. These receptors are inactive at alkaline pH, while at acidic pH they are maximally activated. Genome-wide association studies identified a locus within the TDAG8 gene to be associated with several autoimmune diseases including MS. Notably, we here found that TDAG8 expression is upregulated in MS plaques which prompted us to explore the expression and function of TDAG8 in the CNS in human MO3.13 oligodendrocytesin vitroandin vivoin the lipopolysaccharide-induced neuroinflammation model. We found that TDAG8 is upregulated in maturing oligodendrocytes and temporarily under acidic conditions. Acidic pH also induces oligodendrocyte branching, inhibits chemotaxis and affects the expression of oligodendrocyte maturation markers, PDGFRα and MBPin vitro. Even though myelination was not affected in the adult TDAG8-deficient mice, the expression of human and murine TDAG8 was strongly regulated upon inflammationin vivoin the brain andin vitroin lipopolysaccharide and pro-inflammatory cytokine-treated oligodendrocytes. Together these findings point toward a potential role of TDAG8 in oligodendrocyte biology, neuroinflammation and pathophysiology of MS and provide new directions for further scientific enquiry.

The EBI2 receptor is a major modulator of innate immunity and, together with its ligand, oxysterol 7α,25OHC, has been implicated in several neuroinflammatory and autoimmune diseases. 7alpha,25OHC is synthesised from cholesterol with CH25H and CYP7B1 enzymes and degraded with HSD3B7. The concentration of 7alpha,25OHC in the brain increases in the early phases of the murine model of multiple sclerosis, leading to an enhanced central nervous system (CNS) infiltration with EBI2-expressing lymphocytes. Here, we aimed to investigate whether the enzymes involved in the synthesis and degradation of 7alpha,25OHC are expressed directly in the mouse brain microvascular cells and whether systemic inflammation modulates their levels in these cells. Normal mouse brain capillaries were isolated and immunostained for EBI2, CH25H, CYP7B1 and HSD3B7. Subsequently, mice were challenged with lipopolysaccharide and the mRNA expression in whole brain homogenates was measured. Changes in the receptor and enzyme levels were quantified directly in endothelial cells (ECs), pericytes and astrocytes. The data indicated high levels of EBI2 in the brain microvascular ECs, pericytes and astrocytes with the highest co-localisation in pericytes. CH25H was detected in ECs and astrocytes with low levels in pericytes. CYP7B1 was moderately expressed in ECs, astrocytes and pericytes. The 7alpha,25OHC degrading enzyme HSD3B7 was the least detected in astrocytes. Moreover, the data indicated that systemic inflammation downregulated the mRNA levels of Ebi2 and upregulated Ch25h expression in the whole brain. Specifically in each cell type, EBI2 was not induced in ECs but increased in astrocytes and pericytes. CH25H levels increased in astrocytes and pericytes and CYP7B1 in ECs and astrocytes. The degrading enzyme, HSD3B7, was least affected by systemic inflammation. Taken together, we here demonstrate that EBI2 and the enzymes regulating its ligand levels are differentially expressed in mouse brain microvessels and are highly modulated by systemic inflammation. Upregulated concentration of 7alpha,25OHC in the brain during inflammation may lead to an increased migration of EBI2-expressing immune cells into the CNS during infection or neuroinflammatory disease. Modulation of the EBI2/oxysterol system in the brain or directly in the brain blood vessels may thus provide a new approach to treating neuroinflammatory diseases including multiple sclerosis.