The osmotic demyelination syndrome (ODS) is a non-primary inflammatory disorder of the central nervous system myelin that is often associated with a precipitous rise of serum sodium concentration. To investigate the physiopathology of ODS in vivo, we generated a novel murine model based on the abrupt correction of chronic hyponatremia. Accordingly, ODS mice developed impairments in brainstem auditory evoked potentials and in grip strength. At 24 hr post-correction, oligodendrocyte markers (APC and Cx47) were downregulated, prior to any detectable demyelination. Oligodendrocytopathy was temporally and spatially correlated with the loss of astrocyte markers (ALDH1L1 and Cx43), and both with the brain areas that will develop demyelination. Oligodendrocytopathy and astrocytopathy were confirmed at the ultrastructural level and culminated with necroptotic cell death, as demonstrated by pMLKL immunoreactivity. At 48 hr post-correction, ODS brains contained pathognomonic demyelinating lesions in the pons, mesencephalon, thalamus and cortical regions. These damages were accompanied by blood-brain barrier (BBB) leakages. Expression levels of IL-1β, FasL, TNFRSF6 and LIF factors were significantly upregulated in the ODS lesions. Quiescent microglial cells type A acquired an activated type B morphology within 24 hr post-correction, and reached type D at 48 hr. In conclusion, this murine model of ODS reproduces the CNS demyelination observed in human pathology and indicates ambiguous causes that is regional vulnerability of oligodendrocytes and astrocytes, while it discards BBB disruption as a primary cause of demyelination. This study also raises new queries about the glial heterogeneity in susceptible brain regions as well as about the early microglial activation associated with ODS.

Inhibition of CYP450-mediated retinoic acid (RA) metabolism by RA metabolism blocking agents increases endogenous retinoids and is an alternative to retinoid therapy. Currently available RA metabolism blocking agents (i.e., liarozole and talarozole) tend to have fewer adverse effects than traditional retinoids but lack target specificity. Substrate-based inhibitor DX314 has enhanced selectivity for RA-metabolizing enzyme CYP26B1 and may offer an improved treatment option for keratinization disorders such as congenital ichthyosis and Darier disease. In this study, we used RT-qPCR, RNA sequencing, pathway, upstream regulator, and histological analyses to demonstrate that DX314 can potentiate the effects of all-trans-RA in healthy and diseased reconstructed human epidermis. We unexpectedly discovered that DX314, but not all-trans-RA or previous RA metabolism blocking agents, appears to protect epidermal barrier integrity. In addition, DX314-induced keratinization and epidermal proliferation effects are observed in a rhino mice model. Altogether, the results indicate that DX314 inhibits all-trans-RA metabolism with minimal off-target activity and shows therapeutic similarity to topical retinoids in vitro and in vivo. Findings of a barrier-protecting effect require further mechanistic study but may lead to a unique strategy in barrier-reinforcing therapies. DX314 is a promising candidate compound for further study and development in the context of keratinization disorders.

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TMEM45A gene encodes an initially uncharacterized predicted transmembrane protein. We previously showed that this gene is highly expressed in keratinocytes where its expression correlates with keratinization, suggesting a role in normal epidermal physiology. To test this hypothesis, we generated TMEM45A knockout mice and found that these mice develop without any evident phenotype. The morphology of the epidermis assessed by histology and by labelling differentiation markers in immunofluorescence was not altered. Toluidine blue permeability assay showed that the epidermal barrier develops normally during embryonic development. We also showed that depletion of TMEM45A in human keratinocytes does not alter their potential to form in vitro 3D-reconstructed epidermis. Indeed, epidermis with normal morphogenesis were generated from TMEM45A-silenced keratinocytes. Their expression of differentiation markers quantified by RT-qPCR and evidenced by immunofluorescence labelling as well as their barrier function estimated by Lucifer yellow permeability were similar to the control epidermis. In summary, TMEM45A gene expression is dispensable for epidermal morphogenesis, keratinization and barrier formation. If this protein plays a role in the epidermis, its experimental depletion can possibly be compensated by other proteins in the two experimental models analyzed in this study.

Aim To model in vitro the contact between adult dental pulp stem cells (DPSCs) and lipoteichoic acid (LTA), a cell wall component expressed at the surface of most Gram‐positive bacteria. Methodology Human DPSCs obtained from impacted third molars were cultured and exposed to various concentrations of S. aureus LTA (0.1, 1.0 and 10 µg mL−1). The effects of LTA on DPSCs proliferation and apoptosis were investigated by MTT assay and flow cytometry. Mineralization of DPSCs was evaluated by alizarin red staining assay. Migration was investigated by microphotographs of wound‐healing and Transwell migration assays. Reverse transcription polymerase chain reaction was used to examine the effects of LTA on p65 NF‐κB translocation and TLR1, TLR2 or TLR6 regulation. Enzyme‐linked immunosorbent assay was used to investigate LTA‐stimulated DPSCs cytokine production. One‐way or two‐way ANOVA and Tukey post hoc multiple comparison were used for statistical analysis. Results DPSCs expressed TLR1, TLR2 and TLR6 involved in the recognition of various forms of LTA or lipoproteins. Exposure to LTA did not up‐ or down‐regulate the mRNAs of TLR1, TLR2 or TLR6 whilst LPS acted as a potent inducer of them [TLR1 (P ≤ 0.05), TLR2 (P ≤ 0.001) and TLR6 (P ≤ 0.001)]. Translocation of p65 NF‐κB to the nucleus was detected in LTA‐stimulated cells, but to a lesser extent than LPS‐stimulated DPSCs (P ≤ 0.001). The viability of cells exposed to LTA was greater than unstimulated cells, which was attributed to an increased proliferation and not to less cell death [LTA 1 μg mL−1 (P ≤ 0.001) and 10 μg mL−1 (P ≤ 0.01)]. For specific doses of LTA (1.0 µg mL−1), adhesion of DPSCs to collagen matrix was disturbed (P ≤ 0.05) and cells enhanced their horizontal mobility (P ≤ 0.001). LTA‐stimulated DPSCs released IL‐6 and IL‐8 in a dose‐dependent manner (P ≤ 0.0001). At all concentrations investigated, LTA did not influence osteogenic/odontoblastic differentiation. Conclusions Human DPSCs were able to sense the wall components of Gram‐positive bacteria likely through TLR2 signalling. Consequently, cells modestly proliferated, increased their migratory behaviour and contributed significantly to the local inflammatory response through cytokine release.