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Defective 3β-hydroxysterol-Δ7 -reductase (DHCR7) in the developmental disorder, Smith-Lemli-Opitz syndrome (SLOS), results in deficiency in cholesterol and accumulation of its precursor, 7-dehydrocholesterol (7-DHC). Here, we show that loss of DHCR7 causes accumulation of 7-DHC-derived oxysterol metabolites, premature neurogenesis, and perturbation of neuronal localization in developing murine or human cortical neural precursors, both in vitro and in vivo. We found that a major oxysterol, 3β,5α-dihydroxycholest-7-en-6-one (DHCEO), mediates these effects by initiating crosstalk between glucocorticoid receptor (GR) and neurotrophin receptor kinase TrkB. Either loss of DHCR7 or direct exposure to DHCEO causes hyperactivation of GR and TrkB and their downstream MEK-ERK-C/EBP signaling pathway in cortical neural precursors. Moreover, direct inhibition of GR activation with an antagonist or inhibition of DHCEO accumulation with antioxidants rescues the premature neurogenesis phenotype caused by the loss of DHCR7. These results suggest that GR could be a new therapeutic target against the neurological defects observed in SLOS.

We previously found that the widely used disinfectants, benzalkonium chlorides (BACs), alter cholesterol and lipid homeostasis in neuronal cell lines and in neonatal mouse brains. Here, we investigate the effects of BACs on neurospheres, an in vitro three-dimensional model of neurodevelopment. Neurospheres cultured from mouse embryonic neural progenitor cells (NPCs) were exposed to increasing concentrations (from 1 to 100 nM) of a short-chain BAC (BAC C12), a long-chain BAC (BAC C16), and AY9944 (a known DHCR7 inhibitor). We found that the sizes of neurospheres were decreased by both BACs but not by AY9944. Furthermore, we observed potent inhibition of cholesterol biosynthesis at the step of DHCR7 by BAC C12 but not by BAC C16, suggesting that cholesterol biosynthesis inhibition is not responsible for the observed reduction in neurosphere growth. By using immunostaining and cell cycle analysis, we found that both BACs induced apoptosis and decreased proliferation of NPCs. To explore the mechanisms underlying their effect on neurosphere growth, we carried out RNA sequencing on neurospheres exposed to each BAC at 50 nM for 24 h, which revealed the activation of the integrated stress response by both BACs. Overall, these results suggest that BACs affect neurodevelopment by inducing the integrated stress response in a manner independent of their effects on cholesterol biosynthesis.

© 2016 Elsevier Inc. Specific spontaneous heritable neurodegenerative diseases have been associated with lower serum and cerebrospinal fluid α-tocopherol (α-TOH) concentrations. Equine neuroaxonal dystrophy (eNAD) has similar histologic lesions to human ataxia with vitamin E deficiency caused by mutations in the α-TOH transfer protein gene (TTPA). Mutations in TTPA are not present with eNAD and the molecular basis remains unknown. Given the neuropathologic phenotypic similarity of the conditions, we assessed the molecular basis of eNAD by global transcriptome sequencing of the cervical spinal cord. Differential gene expression analysis identified 157 significantly (FDR

Drug-induced kidney injury, largely caused by proximal tubular intoxicants, limits development and clinical use of new and approved drugs. Assessing preclinical nephrotoxicity relies on animal models that are frequently insensitive; thus, potentially novel techniques — including human microphysiological systems, or “organs on chips” — are proposed to accelerate drug development and predict safety. Polymyxins are potent antibiotics against multidrug-resistant microorganisms; however, clinical use remains restricted because of high risk of nephrotoxicity and limited understanding of toxicological mechanisms. To mitigate risks, structural analogs of polymyxins (NAB739 and NAB741) are currently in clinical development. Using a microphysiological system to model human kidney proximal tubule, we exposed cells to polymyxin B (PMB) and observed significant increases of injury signals, including kidney injury molecule-1 KIM-1and a panel of injury-associated miRNAs (each P < 0.001). Surprisingly, transcriptional profiling identified cholesterol biosynthesis as the primary cellular pathway induced by PMB (P = 1.22 ×10–16), and effluent cholesterol concentrations were significantly increased after exposure (P < 0.01). Additionally, we observed no upregulation of the nuclear factor (erythroid derived-2)–like 2 pathway, despite this being a common pathway upregulated in response to proximal tubule toxicants. In contrast with PMB exposure, minimal changes in gene expression, injury biomarkers, and cholesterol concentrations were observed in response to NAB739 and NAB741. Our findings demonstrate the preclinical safety of NAB739 and NAB741 and reveal cholesterol biosynthesis as a potentially novel pathway for PMB-induced injury. To our knowledge, this is the first demonstration of a human-on-chip platform used for simultaneous safety testing of new chemical entities and defining unique toxicological pathway responses of an FDA-approved molecule.

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Smith-Lemli-Opitz Syndrome (SLOS) is a recessive human disease caused by defective cholesterol (CHOL) synthesis at the level of DHCR7 (7-dehydrocholesterol reductase), which normally catalyzes the conversion of 7-dehydrocholesterol (7DHC) to CHOL. Formation and abnormal accumulation of 7DHC and 7DHC-derived oxysterols occur in SLOS patients and in rats treated with the DHCR7 inhibitor AY9944. The rat SLOS model exhibits progressive and irreversible retinal dysfunction and degeneration, which is only partially ameliorated by dietary CHOL supplementation. We hypothesized that 7DHC-derived oxysterols are causally involved in this retinal degeneration, and that blocking or reducing their formation should minimize the phenotype. Here, using the SLOS rat model, we demonstrate that combined dietary supplementation with CHOL plus antioxidants (vitamins E and C, plus sodium selenite) provides better outcomes than dietary CHOL supplementation alone with regard to preservation of retinal structure and function and lowering 7DHC-derived oxysterol formation. These proof-of-principle findings provide a translational, pre-clinical framework for designing clinical trials using CHOL-antioxidant combination therapy as an improved therapeutic intervention over the current standard of care for the treatment of SLOS.