BACKGROUND: Acid sphingomyelinase deficiency (ASMD) or Niemann-Pick disease types A, A/B, and B is a progressive, life-limiting, autosomal recessive disorder caused by sphingomyelin phosphodiesterase 1 (SMPD1) gene mutations. There is a need to increase the understanding of morbidity and mortality across children to adults diagnosed with ASMD.
METHODS: This observational retrospective survey analysed medical records of patients with ASMD with retrievable data from 27 hospitals in France, diagnosed/followed up between 1st January 1990 and 31st December 2020. Eligible records were abstracted to collect demographic, medical/developmental history, and mortality data. Survival outcomes were estimated from birth until death using Kaplan-Meier survival analyses; standardised mortality ratio (SMR) was also explored.
RESULTS: A total of 118 medical records of patients with ASMD (type B [n = 94], type A [n = 15], and type A/B [n = 9]) were assessed. The majority of patients were males (63.6%); the median [range] age at diagnosis was 8.0 [1.0-18.0] months (type A), 1.0 [0-3] year (type A/B), and 5.5 [0-73] years (type B). Overall, 30 patients were deceased at the study completion date; the median [range] age at death for patients with ASMD type A (n = 14) was 1 [0-3.6] year, type A/B (n = 6) was 8.5 [3.0-30.9] years, and type B (n = 10) was 57.6 [3.4-74.1] years. The median [95% confidence interval (CI)] survival age from birth in patients with ASMD type A and type A/B was 2.0 [1.8-2.7] years and 11.4 [5.5-18.5] years, respectively. Survival analysis in ASMD type B was explored using SMR [95% CI] analysis (3.5 [1.6-5.9]), which showed that age-specific deaths in the ASMD type B population were 3.5 times more frequent than those in the general French population. The causes of death were mostly severe progressive neurodegeneration (type A: 16.7%), cancer (type B: 16.7%), or unspecified (across groups: 33.3%).
CONCLUSIONS: This study illustrated a substantial burden of illness with high mortality rates in patients with ASMD, including adults with ASMD type B, in France.

Ceramides generated by the activity of the neutral sphingomyelinase 2 (nSMase2) play a pivotal role in stress responses in mammalian cells. Dysregulation of sphingolipid metabolism has been implicated in numerous inflammation-related pathologies. However, its influence on inflammatory cytokine-induced signaling is yet incompletely understood. Here, we used proximity labeling to explore the plasma membrane proximal protein network of nSMase2 and TNFα-induced changes thereof. We established Jurkat cells stably expressing nSMase2 C-terminally fused to the engineered ascorbate peroxidase 2 (APEX2). Removal of excess biotin phenol substantially improved streptavidin-based affinity purification of biotinylated proteins. Using our optimized protocol, we determined nSMase2-proximal biotinylated proteins and their changes within the first 5 min of TNFα stimulation by quantitative mass spectrometry. We observed significant dynamic changes in the nSMase2 microenvironment in response to TNFα stimulation consistent with rapid remodeling of protein networks. Our data confirmed known nSMase2 interactors and revealed that the recruitment of most proteins depended on nSMase2 enzymatic activity. We measured significant enrichment of proteins related to vesicle-mediated transport, including proteins of recycling endosomes, trans-Golgi network, and exocytic vesicles in the proximitome of enzymatically active nSMase2 within the first minutes of TNFα stimulation. Hence, the nSMase2 proximal network and its TNFα-induced changes provide a valuable resource for further investigations into the involvement of nSMase2 in the early signaling pathways triggered by TNFα.

BACKGROUND: This study aims to investigate the involvement of acid sphingomyelinase (ASM) in the pathology of dermatomyositis (DM), making it a potential therapeutic target for DM.
METHODS: Patients with DM and healthy controls (HCs) were included to assess the serum level and activity of ASM, and to explore the associations between ASM and clinical indicators. Subsequently, a myositis mouse model was established using ASM gene knockout and wild-type mice to study the significant role of ASM in the pathology and to assess the treatment effect of amitriptyline, an ASM inhibitor. Additionally, we investigated the potential treatment mechanism by targeting ASM both in vivo and in vitro.
RESULTS: A total of 58 DM patients along with 30 HCs were included. The ASM levels were found to be significantly higher in DM patients compared to HCs, with median (quartile) values of 2.63 (1.80-4.94) ng/mL and 1.64 (1.47-1.96) ng/mL respectively. The activity of ASM in the serum of DM patients was significantly higher than that in HCs. Furthermore, the serum levels of ASM showed correlations with disease activity and muscle enzyme levels. Knockout of ASM or treatment with amitriptyline improved the severity of the disease, rebalanced the CD4 T cell subsets Th17 and Treg, and reduced the production of their secreted cytokines. Subsequent investigations revealed that targeting ASM could regulate the expression of relevant transcription factors and key regulatory proteins.
CONCLUSIONS: ASM is involved in the pathology of DM by regulating the differentiation of naive CD4 + T cells and can be a potential treatment target.

Rituximab (RTX) resistance is a notable challenge in treating diffuse large B-cell lymphoma (DLBCL). β-Sitosterol (β-ST) is a plant sterol that has been found in a broad variety of fruits, spices, and medicinal plants. The antineoplastic properties of β-ST are established in various solid malignancies; however, its effect on DLBCL is uncharted. This study investigates the role of β-ST in DLBCL as well as the underlying mechanisms. Our findings indicated that β-ST impeded DLBCL cell proliferation in a concentration- and time-dependent manner. β-ST appeared to alter sphingolipid metabolism, facilitate acid sphingomyelinase (ASM) translocation to the plasma membrane, augment ceramide platforms through increased ceramide synthesis, and consequently induce apoptosis in DLBCL cells. Furthermore, we found that RTX initiated both apoptotic and survival pathways in vitro, with the former contingent on the transient activation of the ASM, and β-ST could amplify the anti-DLBCL efficacy of RTX by modulating ASM/Ceramide (Cer) signaling. Collectively, our findings elucidate the mechanistic role of β-ST in DLBCL and underscore its potential in amplifying the antineoplastic efficacy of RTX via ASM activation, proposing a potential avenue to improve the efficacy of RTX therapy.

Leptospirosis is a re-emerging infectious disease that presents a diagnostic enigma for clinicians with frequent misdiagnosis due to lack of rapid and accurate diagnostic tests, as the current methods are encumbered by inherent limitations. The development of a diagnostic sensor with a sample-in-result-out capability is pivotal for prompt diagnosis. Herein, we developed a microfluidic paper-based analytical device (spin-μPAD) featuring a sample-in-result-out fashion for the detection of Leptospira specific urinary biomarker, sph2 sphingomyelinase, crucial for noninvasive point-of-care testing. Fabrication of paper devices involved precise photolithography techniques, ensuring a high degree of reproducibility and replicability. By optimizing the device\'s configuration and protein components, a remarkable sensitivity and specificity was achieved for detecting leptospiral sph2 in urine, even at low concentrations down to 1.5 fg/mL, with an assay time of 15 min. Further, the spin-μPAD was validated with 20 clinical samples, suspected of leptospirosis including other febrile illnesses, and compared with gold standard microscopic agglutination test, culture, Lepto IgM ELISA, darkfield microscopy, and Leptocheck WB spot test. In contrast to commercial diagnostic tools, the spin-μPAD was noninvasive, rapid, easy to use, specific, sensitive, and cost-effective. The results highlight the potential of this innovative spin-μPAD for an efficient and dependable approach to noninvasive leptospirosis diagnosis, addressing critical needs in the realms of public health and clinical settings.

Extracellular vesicles (EVs) are shed from the plasma membrane, but the regulation and function of these EVs remain unclear. We found that oxidative stress induced by H2O2 in Hela cells stimulated filopodia formation and the secretion of EVs. EVs were small (150 nm) and labeled for CD44, indicating that they were derived from filopodia. Filopodia-derived small EVs (sEVs) were enriched with the sphingolipid ceramide, consistent with increased ceramide in the plasma membrane of filopodia. Ceramide was colocalized with neutral sphingomyelinase 2 (nSMase2) and acid sphingomyelinase (ASM), two sphingomyelinases generating ceramide at the plasma membrane. Inhibition of nSMase2 and ASM prevented oxidative stress-induced sEV shedding but only nSMase2 inhibition prevented filopodia formation. nSMase2 was S-palmitoylated and interacted with ASM in filopodia to generate ceramide for sEV shedding. sEVs contained nSMase2 and ASM and decreased the level of these two enzymes in oxidatively stressed Hela cells. A novel metabolic labeling technique for EVs showed that oxidative stress induced secretion of fluorescent sEVs labeled with NBD-ceramide. NBD-ceramide-labeled sEVs transported ceramide to mitochondria, ultimately inducing cell death in a proportion of neuronal (N2a) cells. In conclusion, using Hela cells we provide evidence that oxidative stress induces interaction of nSMase2 and ASM at filopodia, which leads to shedding of ceramide-rich sEVs that target mitochondria and propagate cell death.

BACKGROUND: Since the pathogenesis of depression is complex, antidepressant therapy remains unsatisfactory. Recent evidence suggests a link between depression and lipid metabolism. Saikosaponin (SS) exhibits antidepression and lipid-regulating effects in modern pharmacology. However, it is unknown whether lipid regulation is the key mechanism of the SS antidepressant effect and how it works.
OBJECTIVE: In this study, we investigated the relationship between the antidepressant activity of SS and the regulation of lipid metabolism and explored potential mechanisms.
METHODS: APOE-/- mice, in combination with the chronic unpredictable mild stress (CUMS) model, were used to study the relationship between SS antidepressant activity and lipid metabolism through behavioral, electrophysiological techniques, and non-targeted lipidomics. Western blot, primary cell culture technology, and laser speckle cerebral blood flow imaging were employed to elucidate potential mechanisms. GraphPad Prism was used for statistical analysis, and p < 0.05 was considered statistically significant.
RESULTS: APOE-/- mice exhibit more severe depressive-like behavior and dysregulation of sphingolipid metabolism in CUMS. SS alleviates depressive behavior and cortical sphingolipid metabolism disorder caused by CUMS, but has no effect on APOE-/- mice. SS alleviates the imbalance between ceramide (Cer) and sphingomyelin (SM) through acidic sphingomyelinase (AMSase). In addition, SS regulates neuronal glutamate release via sphingolipid metabolism, thereby alleviating the CUMS-induced inhibition of neurovascular coupling (regulates metabotropic glutamate receptor and IP3 receptor), which ameliorates the reduction of cerebral blood flow in depressed mice.
CONCLUSIONS: Our study highlights the role of lipid metabolism in the antidepressant activity of SS and explores its underlying mechanisms. This study provided new insights into the better understanding of the antidepressant mechanisms of phytomedicine while increasing the possibility of lipid metabolism as a therapeutic strategy for depression.

BACKGROUND: Traumatic brain injury (TBI) causes neuroinflammation and can lead to long-term neurological dysfunction, even in cases of mild TBI (mTBI). Despite the substantial burden of this disease, the management of TBI is precluded by an incomplete understanding of its cellular mechanisms. Sphingolipids (SPL) and their metabolites have emerged as key orchestrators of biological processes related to tissue injury, neuroinflammation, and inflammation resolution. No study so far has investigated comprehensive sphingolipid profile changes immediately following TBI in animal models or human cases. In this study, sphingolipid metabolite composition was examined during the acute phases in brain tissue and plasma of mice following mTBI.
METHODS: Wildtype mice were exposed to air-blast-mediated mTBI, with blast exposure set at 50-psi on the left cranium and 0-psi designated as Sham. Sphingolipid profile was analyzed in brain tissue and plasma during the acute phases of 1, 3, and 7 days post-TBI via liquid-chromatography-mass spectrometry. Simultaneously, gene expression of sphingolipid metabolic markers within brain tissue was analyzed using quantitative reverse transcription-polymerase chain reaction. Significance (P-values) was determined by non-parametric t-test (Mann-Whitney test) and by Tukey\'s correction for multiple comparisons.
RESULTS: In post-TBI brain tissue, there was a significant elevation of 1) acid sphingomyelinase (aSMase) at 1- and 3-days, 2) neutral sphingomyelinase (nSMase) at 7-days, 3) ceramide-1-phosphate levels at 1 day, and 4) monohexosylceramide (MHC) and sphingosine at 7-days. Among individual species, the study found an increase in C18:0 and a decrease in C24:1 ceramides (Cer) at 1 day; an increase in C20:0 MHC at 3 days; decrease in MHC C18:0 and increase in MHC C24:1, sphingomyelins (SM) C18:0, and C24:0 at 7 days. Moreover, many sphingolipid metabolic genes were elevated at 1 day, followed by a reduction at 3 days and an absence at 7-days post-TBI. In post-TBI plasma, there was 1) a significant reduction in Cer and MHC C22:0, and an increase in MHC C16:0 at 1 day; 2) a very significant increase in long-chain Cer C24:1 accompanied by significant decreases in Cer C24:0 and C22:0 in MHC and SM at 3 days; and 3) a significant increase of C22:0 in all classes of SPL (Cer, MHC and SM) as well as a decrease in Cer C24:1, MHC C24:1 and MHC C24:0 at 7 days.
CONCLUSIONS: Alterations in sphingolipid metabolite composition, particularly sphingomyelinases and short-chain ceramides, may contribute to the induction and regulation of neuroinflammatory events in the early stages of TBI, suggesting potential targets for novel diagnostic, prognostic, and therapeutic strategies in the future.

Multiple studies have confirmed that acid sphingomyelinase (ASM) activity is associated with depression. The discovery of direct inhibitors against ASM is of great significance for exploring antidepressants and their mechanisms of action. Herein, a series of novel phenylpyrazole analogues were rationally designed and synthesized. Among them, compound 46 exhibited potent inhibitory activity (IC50 = 0.87 μM) and good drug-like properties. In vivo studies demonstrated that compound 46 was involved in multiple antidepressant mechanisms of action, which were associated with a decline of ceramide, including increasing the Bcl-2/Bax ratio and BDNF expression, down-regulating caspase-3 and caspase-9, ameliorating oxidative stress, reducing the levels of proinflammatory cytokines such as TNF-α, IL-1β, and IL-6, and elevating 5-HT levels in the brains of mice, respectively. These meaningful results reveal for the first time that direct inhibitors exhibit remarkable antidepressant effects in the CUMS-induced mouse model through multiple mechanisms of antidepressant action.

Acid sphingomyelinase deficiency (ASMD) is a rare, lysosomal storage disease with limited evidence on its natural history. This retrospective, medical record abstraction study aimed to characterize the natural history of ASMD (types B and A/B) during childhood and adolescence. Recruiting sites were European centers (i.e., France, Germany, Italy, and the United Kingdom) from the ASCEND-Peds trial (NCT02292654); these sites were targeted because of the rarity of ASMD and specialized care provided at these centers. The study population comprised ASMD trial patients (before exposure to treatment) and ASMD non-trial participants who were managed at the same trial sites. Overall, 18 patients were included (11 trials; 7 non-trials; median [Q1; Q3] age at ASMD diagnosis: 2.5 [1.0; 4.0] years). Median follow-up duration was 10.0 years. Frequently reported medical conditions were hepatobiliary (17 [94.4%]) and blood and lymphatic system disorders (16 [88.9%]). Adenoidectomy (3 [16.7%]) was the most commonly reported surgical procedure; gastroenteritis (5 [27.8%]) was the most frequently reported infection, and epistaxis (6 [33.3%]) was the most commonly reported bleeding event. Abnormal spleen (16 [88.9%]) and liver (15 [83.3%]) size and respiratory function (8 [44.4%]) were commonly reported during physical examination. Overall, 11 (61.1%) patients were hospitalized; 6 (33.3%) patients had emergency room visits. Findings were consistent with published literature and support the current understanding of natural history of ASMD.