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Abstract Ceramides are lipids that play vital roles in complex lipid synthesis, membrane function, and cell signaling. Disrupted ceramide homeostasis is implicated in cell-death and several neurologic diseases. Ceramides are often analyzed in tissue, but this approach fails to resolve cell-type differences in ceramide homeostasis that are likely essential to understanding cell and non-cell autonomous contributions to neurodegeneration. We show that human iPSC-derived neurons and glia differ in their rate of ceramide synthesis, ceramide isoform composition, and responses to altered ceramide levels. RNA-sequencing of cells treated to increase or decrease ceramides revealed connections to inflammation, ER stress, and apoptosis. Moreover, introducing labeled sphinganine showed that glia readily synthesize ceramide de novo and that neurons are relatively more sensitive to ceramide toxicity. Our findings provide a framework for understanding neurologic diseases with sphingolipid alternations and insights into designing therapeutics that target ceramide for treating them.

Abstract While significant advances have been made in understanding renal pathophysiology, less is known about the role of glycosphingolipid (GSL) metabolism in driving organ dysfunction. Here, we used a small molecule inhibitor of glucosylceramide synthase to modulate GSL levels in three mouse models of distinct renal pathologies: Alport syndrome (Col4a3 KO), polycystic kidney disease (Nek8 jck ), and steroid-resistant nephrotic syndrome (Nphs2 cKO). At the tissue level, we identified a core immune-enriched transcriptional signature that was shared across models and enriched in human polycystic kidney disease. Single nuclei analysis identified robust transcriptional changes across multiple kidney cell types, including epithelial and immune lineages. To further explore the role of GSL modulation in macrophage biology, we performed in vitro studies with homeostatic and inflammatory bone marrow-derived macrophages. Cumulatively, this study provides a comprehensive overview of renal dysfunction and the effect of GSL modulation on kidney-derived cells in the setting of renal dysfunction.

Background: A salutary effect of treatments for Gaucher disease (GD) has been a reduction in the incidence of avascular osteonecrosis (AVN). However, there are reports of AVN in patients receiving enzyme replacement therapy (ERT) , and it is not known whether it is related to individual treatments, GBA genotypes, phenotypes, biomarkers of residual disease activity, or anti-drug antibodies. Prompted by development of AVN in several patients receiving ERT, we aimed to delineate the determinants of AVN in patients receiving ERT or eliglustat substrate reduction therapy (SRT) during 20 years in a tertiary referral center. Methods: Longitudinal follow-ups of 155 GD patients between 2001 and 2021 were analyzed for episodes of AVN on therapy, type of therapy, GBA1 genotype, spleen status, biomarkers, and other disease indicators. We applied mixed-effects logistic model to delineate the independent correlates of AVN while receiving treatment. Results: The patients received cumulative 1382 years of treatment. There were 16 episodes of AVN in 14 patients, with two episodes, each occurring in two patients. Heteroallelic p.Asn409Ser GD1 patients were 10 times (95% CI, 1.5–67.2) more likely than p.Asn409Ser homozygous patients to develop osteonecrosis during treatment. History of AVN prior to treatment initiation was associated with 4.8-fold increased risk of AVN on treatment (95% CI, 1.5–15.2). The risk of AVN among patients receiving velaglucerase ERT was 4.68 times higher compared to patients receiving imiglucerase ERT (95% CI, 1.67–13). No patient receiving eliglustat SRT suffered AVN. There was a significant correlation between GlcSph levels and AVN. Together, these biomarkers reliably predicted risk of AVN during therapy (ROC AUC 0.894, p

Background: Neuronopathic Gaucher disease (nGD) is a rare neurodegenerative disorder caused by biallelic mutations in GBA and buildup of glycosphingolipids in lysosomes. Neuronal injury and cell death are prominent pathological features; however, the role of GBA in individual cell types and involvement of microglia, blood-derived macrophages, and immune infiltrates in nGD pathophysiology remains enigmatic. Methods: Here, using single-cell resolution of mouse nGD brains, lipidomics, and newly generated biomarkers, we found induction of neuroinflammation pathways involving microglia, NK cells, astrocytes, and neurons. Results: Targeted rescue of Gba in microglia and neurons, respectively, in Gba-deficient, nGD mice reversed the buildup of glucosylceramide (GlcCer) and glucosylsphingosine (GlcSph), concomitant with amelioration of neuroinflammation, reduced serum neurofilament light chain (Nf-L), and improved survival. Serum GlcSph concentration was correlated with serum Nf-L and ApoE in nGD mouse models as well as in GD patients. Gba rescue in microglia/macrophage compartment prolonged survival, which was further enhanced upon treatment with brain-permeant inhibitor of glucosylceramide synthase, effects mediated via improved glycosphingolipid homeostasis, and reversal of neuroinflammation involving activation of microglia, brain macrophages, and NK cells. Conclusions: Together, our study delineates individual cellular effects of Gba deficiency in nGD brains, highlighting the central role of neuroinflammation driven by microglia activation. Brain-permeant small-molecule inhibitor of glucosylceramide synthase reduced the accumulation of bioactive glycosphingolipids, concomitant with amelioration of neuroinflammation involving microglia, NK cells, astrocytes, and neurons. Our findings advance nGD disease biology whilst identifying compelling biomarkers of nGD to improve patient management, enrich clinical trials, and illuminate therapeutic targets. Funding: Research grant from Sanofi; other support includes R01NS110354.

In Gaucher disease (GD), genetic deficiency of acid β-glucosidase leads to accumulation of its substrate glucosylceramide (GlcCer) and glucosylsphingosine (GlcSph). Lipid-laden cells, most prominently seen as macrophages engorged with GlcCer and GlcSph-laden lysosomes, trigger chronic metabolic inflammation and multisystemic phenotypes. Among the pleiotropic effects of inflammatory cascades, the induction of glucosylceramide synthase accentuates the primary metabolic defect. First-line therapies for adults with GD type 1 include Enzyme Replacement Therapy (ERT) and eliglustat Substrate Reduction Therapy (SRT). The ENCORE phase 3 clinical trial of eliglustat demonstrated non-inferiority compared to ERT. It is not known whether switching stable patients from long-term ERT to SRT results in the incremental reversal of the disease phenotype and its surrogate indicators. Herein, we report real-world experience from a single tertiary referral center of 38 adult GD type 1 patients, stable on long-term ERT (mean 13.3 years), who switched to eliglustat SRT (mean 3.1 years). After switch to SRT, there was significant reduction in spleen volume (P = 0.003) while liver volume, which was normal at baseline, remained unchanged. Platelet counts increased significantly (P = 0.026). Concomitantly, there was reduction of three validated biomarkers of Gaucher disease activity: plasma GlcSph decreased from 63.7 ng/ml (95% CI, 37.6-89.8) to 26.1 ng/ml (95% CI, 15.7-36.6) (P