PLASMA LIPIDS AND AMINO ACIDS LINKED TO LOW SARS-CoV-2 SUSCEPTIBILITY

Background: The mechanisms driving SARS-CoV-2 susceptibility remain poorly understood, especially the factors determining why a subset of unvaccinated individuals remain uninfected despite high-risk exposures. Method(s): We studied an exceptional group of unvaccinated healthcare workers heavily exposed to SARS-CoV-2 ('nonsusceptible') from April to June 2020, who were compared against 'susceptible' individuals to SARS-CoV-2, including uninfected subjects who became infected during the follow-up, and hospitalized patients with different disease severity providing samples at early disease stages. We analyzed plasma samples using different mass spectrometry technique and obtained metabolites and lipids profiles. Result(s): We found that the metabolite profiles were predictive of the selected study groups and identified lipids profiles and metabolites linked to SARS-CoV-2 susceptibility and COVID-19 severity. More importantly, we showed that non-susceptible individuals exhibited unique metabolomics and lipidomic patterns characterized by upregulation of most lipids -especially ceramides and sphingomyelin-and amino acids related to tricarboxylic acid cycle and mitochondrial metabolism, which could be interpreted as markers of low susceptibility to SARS-CoV-2 infection. Lipids and metabolites pathways analysis revealed that metabolites related to energy production, mitochondrial and tissue dysfunction, and lipids involved in membrane structure and virus infectivity were key markers of SARS-CoV-2 susceptibility. Conclusion(s): Lipid and metabolic profiles differ in 'nonsusceptible' compared to individuals susceptible to SARS-CoV-2. Our study suggests that lipid profiles are relevant actors during SARS-CoV-2 pathogenesis and highlight certain lipids relevant to understand SARS-CoV-2 pathogenesis. (Figure Presented).

Effect of circulating ceramides on adiposity and insulin resistance in patients with type 2 diabetes: An observational cross-sectional study.

INTRODUCTION: Insulin resistance (IR) is one of the common chronic metabolic disorders in Africa and elsewhere. Accumulation of lipids in the body may be due to an imbalance in the metabolism of lipids, glucose and proteins. Ceramides are a sphingolipid class of lipids that are biologically active and vital in the production of more complex lipids. Circulating ceramides are thought to have a role in the development of obesity-related IR, although the precise involvement remains unclear. AIM: To investigate the impact of circulating ceramide on IR and body adiposity in people with and without type 2 diabetes mellitus (T2DM). METHODOLOGY: The study was observational and cross-sectional. There were a total of 84 volunteers with T2DM and 75 nondiabetics (control). The participants' ages, body mass indexes (BMI), waist circumferences, and blood pressure (BP) were among the clinical parameters assessed. Ceramide levels, fasting plasma glucose (FPG), lipids, basal insulin levels and glycated haemoglobin (HbA1c) were also measured. Additionally, the homeostatic model assessment for IR (HOMA-IR) and beta cell function (HOMA-ß) were computed. RESULTS: T2DM and control participants had different mean values for anthropometric parameters, BP, FPG, HbA1c, lipids, insulin, HOMA-IR, HOMA-ß and ceramide levels (p < .05 for all). HOMA-IR, HOMA-ß and cardiovascular risk were significant correlates with ceramide levels in the T2DM group (r = 0.24; -0.34; 0.24, p < .05, respectively). Further, FPG (OR = 1.83, p = .01) and ceramide (OR = 1.05, p = .01) levels were significant predictors of IR in the case group. CONCLUSION: Patients with T2DM exhibited high ceramide concentrations, which, when combined with high FPG, were associated with IR. The consequences of circulating ceramides in health and disease; however, merit further research.

COVID-19, Blood Lipid Changes, and Thrombosis.

Although there is increasing evidence that oxidative stress and inflammation induced by COVID-19 may contribute to increased risk and severity of thromboses, the underlying mechanism(s) remain to be understood. The purpose of this review is to highlight the role of blood lipids in association with thrombosis events observed in COVID-19 patients. Among different types of phospholipases A2 that target cell membrane phospholipids, there is increasing focus on the inflammatory secretory phospholipase A2 IIA (sPLA2-IIA), which is associated with the severity of COVID-19. Analysis indicates increased sPLA2-IIA levels together with eicosanoids in the sera of COVID patients. sPLA2 could metabolise phospholipids in platelets, erythrocytes, and endothelial cells to produce arachidonic acid (ARA) and lysophospholipids. Arachidonic acid in platelets is metabolised to prostaglandin H2 and thromboxane A2, known for their pro-coagulation and vasoconstrictive properties. Lysophospholipids, such as lysophosphatidylcholine, could be metabolised by autotaxin (ATX) and further converted to lysophosphatidic acid (LPA). Increased ATX has been found in the serum of patients with COVID-19, and LPA has recently been found to induce NETosis, a clotting mechanism triggered by the release of extracellular fibres from neutrophils and a key feature of the COVID-19 hypercoagulable state. PLA2 could also catalyse the formation of platelet activating factor (PAF) from membrane ether phospholipids. Many of the above lipid mediators are increased in the blood of patients with COVID-19. Together, findings from analyses of blood lipids in COVID-19 patients suggest an important role for metabolites of sPLA2-IIA in COVID-19-associated coagulopathy (CAC).

Sphingolipid metabolism and its relationship with cardiovascular, renal and metabolic diseases.

Sphingolipids (sphingomyelin, glycolipids, gangliosides) are located in cell membranes, plasma, and lipoproteins. In patients with cardiovascular, renal, and metabolic diseases, the profile of sphingolipids and their metabolites (ceramide, sphingosine, and sphingosine-1-phosphate) is modified, and these changes may explain the alterations in some cellular responses such as apoptosis. Furthermore, sphingosine and sphingosine-1-phosphate have been suggested to prevent COVID-19. This review also briefly mentions the techniques that allow us to study sphingolipids and their metabolites.

Los esfingolípidos (esfingomielina, glucolípidos y gangliósidos) se localizan en las membranas celulares, el plasma y las lipoproteínas. En pacientes con enfermedades cardiovasculares, renales y metabólicas, el perfil de los esfingolípidos y sus metabolitos (ceramida, esfingosina y esfingosina-1-fosfato) se modifica, y estos cambios pueden explicar las alteraciones en algunas respuestas celulares, como la apoptosis. Además, se ha sugerido que la esfingosina y la esfingosina-1-fosfato previenen la COVID-19. En esta revisión también se mencionan brevemente las técnicas que permiten el estudio de los esfingolípidos y sus metabolitos.

Irradiation accelerates SARS-CoV-2 infection by enhancing sphingolipid metabolism.

Cancer patients who receive radiotherapy have a high risk of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection, but the concrete reason remains unclear. Herein, we investigated the influence of irradiation on the vulnerability of cancer cells to SARS-CoV-2 using S pseudovirions and probed the underlying mechanism via RNA-seq and other molecular biology techniques. Owing to the enhancement of sphingolipid metabolism, irradiation accelerated pseudovirion infection. Mechanistically, irradiation induced the expression of acid sphingomyelinase (ASM), which catalyses the hydrolysis of sphingomyelin to ceramide, contributing to lipid raft formation and promoting SARS-CoV-2 invasion. Inhibition of lipid raft formation with methyl-ß-cyclodextrin (MßCD) or the tyrosine kinase inhibitor genistein and ASM suppression through small interfering RNA or amitriptyline (AMT) treatment abolished the enhancing effect of irradiation on viral infection. Animal experiments supported the finding that irradiation promoted SARS-CoV-2 S pseudovirion infection in A549 cell tumour-bearing BALB/c nude mice, whereas AMT treatment dramatically decreased viral infection. This study discloses the role of sphingolipid metabolism in irradiation-induced SARS-CoV-2 infection, thus providing a potential target for clinical intervention to protect patients receiving radiotherapy from COVID-19.

Antidepressant Use and Its Association with 28-Day Mortality in Inpatients with SARS-CoV-2: Support for the FIASMA Model against COVID-19.

To reduce Coronavirus Disease 2019 (COVID-19)-related mortality and morbidity, widely available oral COVID-19 treatments are urgently needed. Certain antidepressants, such as fluvoxamine or fluoxetine, may be beneficial against COVID-19. We included 388,945 adult inpatients who tested positive for SARS-CoV-2 at 36 AP-HP (Assistance Publique-Hôpitaux de Paris) hospitals from 2 May 2020 to 2 November 2021. We compared the prevalence of antidepressant use at admission in a 1:1 ratio matched analytic sample with and without COVID-19 (N = 82,586), and assessed its association with 28-day all-cause mortality in a 1:1 ratio matched analytic sample of COVID-19 inpatients with and without antidepressant use at admission (N = 1482). Antidepressant use was significantly less prevalent in inpatients with COVID-19 than in a matched control group of inpatients without COVID-19 (1.9% versus 4.8%; Odds Ratio (OR) = 0.38; 95%CI = 0.35-0.41, p < 0.001). Antidepressant use was significantly associated with reduced 28-day mortality among COVID-19 inpatients (12.8% versus 21.2%; OR = 0.55; 95%CI = 0.41-0.72, p < 0.001), particularly at daily doses of at least 40 mg fluoxetine equivalents. Antidepressants with high FIASMA (Functional Inhibitors of Acid Sphingomyelinase) activity seem to drive both associations. These treatments may reduce SARS-CoV-2 infections and COVID-19-related mortality in inpatients, and may be appropriate for prophylaxis and/or COVID-19 therapy for outpatients or inpatients.

Optimization of common hand cleansers for protection from viral spread and occupational hand dermatoses

Irritant contact dermatitis (ICD) affects over 20% of health care workers, who manage their condition by substituting soap with an emollient cleanser. It is not clear whether emollient cleansers have the same level of virus eliminating activity as soap. Therefore, we evaluated a range of emollient cleansers for virus eliminating activity against enveloped (coronavirus and herpes simplex virus) and nonenveloped viruses (adenovirus (Ad)). In accordance with European standards a range of cleansers were combined with viruses under different (‘clean’ and ‘dirty’ hand hygiene) conditions. Virus viability and architecture were determined by plaque /TCDI50 assays and transmission electron microscopy. Traditional soaps (natural fatty acid), synthetic soaps and emollient cleansers (e.g., ceramide-containing cleansers) exhibited significant antiviral activity in enveloped viruses. However, the antiviral activity of traditional soaps reduced drastically when combined with hard water. Moreover, nonenveloped viruses were less susceptible to both synthetic soaps and emollients cleansers. Interestingly, traditional soaps inhibited the viability of Ad at high concentrations, but only in soft water. Most emollient cleansers were effective at eliminating enveloped viruses, suggesting that they are an acceptable substitute for soap to control the spread of viruses, like SARS-CoV-2, and protect against ICD. Nonenveloped viruses showed resistance to most of the hand hygiene products tested, except for traditional soap. This suggests that hand washing alone may not be sufficient to control the spread of nonenveloped viruses. Taken together this suggests that different virus strains respond differently to soaps and cleansers, and that this should be considered in the guidance for hand hygiene.

Effect of Subacute Angiotensin II Elevations on Sphingolipid Levels and Lung Injury in Mice

Background: There is a paucity of therapies for acute lung injury (ALI) induced by respiratory viruses. A previously demonstrated key mechanism of ALI, particularly in the setting of severe acute respiratory syndrome coronavirus infections, has been ascribed to decreased cell surface angiotensin converting enzyme 2 (ACE2) leading to increased circulating levels of angiotensin II (Ang2). In turn, supraphysiological Ang2 levels trigger a cascade of events that culminates with endothelial injury in the systemic circulation via acid sphingomyelinase (ASMase) activation. ASMase has been implicated in several models of ALI, but its specific involvement in Ang2-induced ALI is unknown. ASMase hydrolyzes sphingomyelin to pro-apoptotic, edemagenic ceramide, which can be metabolized to endothelial-protective sphingosine-1-phosphate (S1P). Therefore, the ratio of ceramide/S1P can determine endothelial cell fate and lung vascular permeability. We hypothesized that ceramide levels are increased relative to S1P in mice with Ang2-induced ALI. Methods: Following a published protocol of Ang2-induced ALI (Wu et al, 2017), we delivered Ang2 via osmotic pumps (1 ug/kg/min, 7 days;Ang2-mice), using saline (sham) or untreated C57BL/6 mice as controls. We evaluated pulmonary function (FlexiVent);albumin, IgM (ELISA), and inflammatory cell abundance in bronchoalveolar lavage fluid (BALF);and lung parenchyma inflammation and fibrosis (Ashcroft score) on H/E-stained lungs. Sphingolipid levels in lungs and plasma were measured by tandem liquid chromatography/mass spectrometry. Results: Inspiratory capacity, lung compliance, and body weight all decreased in Ang2-mice (by 13-14%, p<0.05 each) compared to sham. Lung pressure-volume loops exhibited a right-shift in Ang2- vs. sham or untreated mice. There was no significant change in BALF albumin, IgM, or inflammatory cells, or in lung histology inflammation or fibrosis scores in Ang2-mice. Compared to sham, S1P levels were significantly increased in plasma and unlavaged lung in Ang2-mice, decreasing ceramide/S1P ratios (from 3.1 to 2.0, and 26 to 20, respectively, p<0.05 each). Conclusions: Sustained subacute systemic elevations of Ang2 increased lung stiffness, but did not cause severe ALI in mice. Lung and circulatory elevations of S1P but not ceramide may have protected against lung edema and inflammatory injury. Although the cause of increased lung stiffness in this model remains to be elucidated, it is notable that chronic (months) supraphysiological elevations of either Ang2 or S1P have been associated with lung fibrosis. In conclusion, a second-hit injury may be necessary to augment the susceptibility of murine lung to Ang2-induced endothelial damage and inflammation relevant to coronavirus.

Skeletal Muscle and COVID-19: The Potential Involvement of Bioactive Sphingolipids.

SARS-CoV-2 virus infection is the cause of the coronavirus disease 2019 (COVID-19), which is still spreading over the world. The manifestation of this disease can range from mild to severe and can be limited in time (weeks) or persist for months in about 30-50% of patients. COVID-19 is considered a multiple organ dysfunction syndrome and the musculoskeletal system manifestations are beginning to be considered of absolute importance in both COVID-19 patients and in patients recovering from the SARS-CoV-2 infection. Musculoskeletal manifestations of COVID-19 and other coronavirus infections include loss of muscle mass, muscle weakness, fatigue or myalgia, and muscle injury. The molecular mechanisms by which SARS-CoV-2 can cause damage to skeletal muscle (SkM) cells are not yet well understood. Sphingolipids (SLs) represent an important class of eukaryotic lipids with structural functions as well as bioactive molecules able to modulate crucial processes, including inflammation and viral infection. In the last two decades, several reports have highlighted the role of SLs in modulating SkM cell differentiation, regeneration, aging, response to insulin, and contraction. This review summarizes the consequences of SARS-CoV-2 infection on SkM and the potential involvement of SLs in the tissue responses to virus infection. In particular, we highlight the role of sphingosine 1-phosphate signaling in order to aid the prediction of novel targets for preventing and/or treating acute and long-term musculoskeletal manifestations of virus infection in COVID-19.

Fetal Effects of COVID-19 Infection: Targeted Metabolomic profiling of cord blood

Objective: The impact of maternal COVID-19 infection on fetal health remains to be determined. Using targeted metabolomic analysis of newborn umbilical cord blood, we aimed to evaluate the biological consequences of maternal infection on the fetus and develop metabolite biomarkers for the identification of newborn intrauterine exposure. Study Design: Cord blood serum samples from 23 COVID-19 cases (mother infected/ newborn negative) and 23 gestational age-matched controls were analyzed using nuclear magnetic spectroscopy and direct injection liquid chromatography mass spectrometry-mass spectrometry. Logistic regression models were developed using metabolites to predict intrauterine exposure with Area under the Receiver Operating Characteristics curve [AUC (95% CI)], sensitivity, and specificity. Metabolite set enrichment analysis was used to evaluate altered biochemical pathways to highlight biological mechanisms of COVID-19 intrauterine exposure. Results: There were no significant differences in gestational age at delivery between groups (p >0.05). All neonates tested negative for COVID-19 infection. Significant concentration differences (p-value < 0.05 or -log10=1.301) were observed in 19 metabolites between groups. The top metabolite model [cortisol and Ceramide (d18:1/20:0)] achieved an AUC (95% CI) = 0.839 (0.722 - 0.956) with a sensitivity of 91% and specificity of 69% (Table 1). Enrichment analysis revealed significantly (p< 0.05) altered metabolic pathway of steroidogenesis and gluconeogenesis (Figure 1). Cortisol is the stress hormone that increases glucose production through gluconeogenesis resulting in higher oxidative metabolism and energy generation. Ceramides are known to have anti-inflammatory properties. Elevated hypoxanthine has also been correlated with tissue hypoxia and inflammation. Conclusion: We found evidence of intrauterine stress, altered energy metabolism and inflammation in fetal life in cases of maternal COVID-19 infection but ultimately negative newborn culture. Elucidation of long-term consequences is imperative considering the large number of exposures in the population. [Formula presented] [Formula presented]

Activation of Sphingomyelinase-Ceramide-Pathway in COVID-19 Purposes Its Inhibition for Therapeutic Strategies.

Effective treatment strategies for severe coronavirus disease (COVID-19) remain scarce. Hydrolysis of membrane-embedded, inert sphingomyelin by stress responsive sphingomyelinases is a hallmark of adaptive responses and cellular repair. As demonstrated in experimental and observational clinical studies, the transient and stress-triggered release of a sphingomyelinase, SMPD1, into circulation and subsequent ceramide generation provides a promising target for FDA-approved drugs. Here, we report the activation of sphingomyelinase-ceramide pathway in 23 intensive care patients with severe COVID-19. We observed an increase of circulating activity of sphingomyelinase with subsequent derangement of sphingolipids in serum lipoproteins and from red blood cells (RBC). Consistent with increased ceramide levels derived from the inert membrane constituent sphingomyelin, increased activity of acid sphingomyelinase (ASM) accurately distinguished the patient cohort undergoing intensive care from healthy controls. Positive correlational analyses with biomarkers of severe clinical phenotype support the concept of an essential pathophysiological role of ASM in the course of SARS-CoV-2 infection as well as of a promising role for functional inhibition with anti-inflammatory agents in SARS-CoV-2 infection as also proposed in independent observational studies. We conclude that large-sized multicenter, interventional trials are now needed to evaluate the potential benefit of functional inhibition of this sphingomyelinase in critically ill patients with COVID-19.

The Manifold Roles of Sphingolipids in Viral Infections.

Sphingolipids are essential components of eukaryotic cells. In this review, we want to exemplarily illustrate what is known about the interactions of sphingolipids with various viruses at different steps of their replication cycles. This includes structural interactions during entry at the plasma membrane or endosomal membranes, early interactions leading to sphingolipid-mediated signal transduction, interactions with internal membranes and lipids during replication, and interactions during virus assembly and budding. Targeted interventions in sphingolipid metabolism - as far as they can be tolerated by cells and organisms - may open novel possibilities to support antiviral therapies. Human immunodeficiency virus type 1 (HIV-1) infections have intensively been studied, but for other viral infections, such as influenza A virus (IAV), measles virus (MV), hepatitis C virus (HCV), dengue virus, Ebola virus, and severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), investigations are still in their beginnings. As many inhibitors of sphingolipid metabolism are already in clinical use against other diseases, repurposing studies for applications in some viral infections appear to be a promising approach.

Sphingolipids as Modulators of SARS-CoV-2 Infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the COVID-19 pandemic with severe consequences for afflicted individuals and the society as a whole. The biology and infectivity of the virus has been intensively studied in order to gain a better understanding of the molecular basis of virus-host cell interactions during infection. It is known that SARS-CoV-2 binds to angiotensin-converting enzyme 2 (ACE2) via its spike protein. Priming of the virus by specific proteases leads to viral entry via endocytosis and to the subsequent steps in the life cycle of SARS-CoV-2. Sphingosine and ceramide belong to the sphingolipid family and are abundantly present in cell membranes. These lipids were recently shown to interfere with the uptake of virus particles of SARS-CoV-2 into epithelial cell lines and primary human nasal cells in culture. The mechanisms of action were partly different, as sphingosine blocked, whilst ceramide facilitated viral entry. Acid sphingomyelinase (ASM) is vital for the generation of ceramide and functional inhibition of ASM by drugs like amitriptyline reduced SARS-CoV-2 entry into the epithelial cells. Recent data indicates that serum level of sphingosine-1-phosphate (S1P) is a prognostic factor for COVID-2 severity. Further, stimulation of sphingosine-1-phosphate receptor 1 (S1PR1) might also constrain the hyper-inflammatory conditions linked to SARS-CoV-2. Here, we review recent exciting findings regarding sphingolipids in the uptake of SARS-CoV-2 and in the course of COVID-19 disease. More studies are required on the mechanisms of action and the potential use of antidepressant drugs and sphingolipid modifiers in SARS-CoV-2 infections and in the treatment of the more serious and fatal consequences of the disease.

Sphingolipidomics in Translational Sepsis Research-Biomedical Considerations and Perspectives.

Scientific Background: Sphingolipids are a highly diverse group of lipids with respect to physicochemical properties controlling either structure, distribution, or function, all of them regulating cellular response in health and disease. Mass spectrometry, on the other hand, is an analytical technique characterizing ionized molecules or fragments thereof by mass-to-charge ratios, which has been prosperingly developed for rapid and reliable qualitative and quantitative identification of lipid species. Parallel to best performance of in-depth chromatographical separation of lipid classes, preconditions of precise quantitation of unique molecular species by preprocessing of biological samples have to be fulfilled. As a consequence, "lipid profiles" across model systems and human individuals, esp. complex (clinical) samples, have become eminent over the last couple of years due to sensitivity, specificity, and discriminatory capability. Therefore, it is significance to consider the entire experimental strategy from sample collection and preparation, data acquisition, analysis, and interpretation. Areas Covered: In this review, we outline considerations with clinical (i.e., human) samples with special emphasis on sample handling, specific physicochemical properties, target measurements, and resulting profiling of sphingolipids in biomedicine and translational research to maximize sensitivity and specificity as well as to provide robust and reproducible results. A brief commentary is also provided regarding new insights of "clinical sphingolipidomics" in translational sepsis research. Expert Opinion: The role of mass spectrometry of sphingolipids and related species ("sphingolipidomics") to investigate cellular and compartment-specific response to stress, e.g., in generalized infection and sepsis, is on the rise and the ability to integrate multiple datasets from diverse classes of biomolecules by mass spectrometry measurements and metabolomics will be crucial to fostering our understanding of human health as well as response to disease and treatment.

Ceramide and Related Molecules in Viral Infections.

Ceramide is a lipid messenger at the heart of sphingolipid metabolism. In concert with its metabolizing enzymes, particularly sphingomyelinases, it has key roles in regulating the physical properties of biological membranes, including the formation of membrane microdomains. Thus, ceramide and its related molecules have been attributed significant roles in nearly all steps of the viral life cycle: they may serve directly as receptors or co-receptors for viral entry, form microdomains that cluster entry receptors and/or enable them to adopt the required conformation or regulate their cell surface expression. Sphingolipids can regulate all forms of viral uptake, often through sphingomyelinase activation, and mediate endosomal escape and intracellular trafficking. Ceramide can be key for the formation of viral replication sites. Sphingomyelinases often mediate the release of new virions from infected cells. Moreover, sphingolipids can contribute to viral-induced apoptosis and morbidity in viral diseases, as well as virus immune evasion. Alpha-galactosylceramide, in particular, also plays a significant role in immune modulation in response to viral infections. This review will discuss the roles of ceramide and its related molecules in the different steps of the viral life cycle. We will also discuss how novel strategies could exploit these for therapeutic benefit.

The Anti-Infectious Role of Sphingosine in Microbial Diseases.

Sphingolipids are important structural membrane components and, together with cholesterol, are often organized in lipid rafts, where they act as signaling molecules in many cellular functions. They play crucial roles in regulating pathobiological processes, such as cancer, inflammation, and infectious diseases. The bioactive metabolites ceramide, sphingosine-1-phosphate, and sphingosine have been shown to be involved in the pathogenesis of several microbes. In contrast to ceramide, which often promotes bacterial and viral infections (for instance, by mediating adhesion and internalization), sphingosine, which is released from ceramide by the activity of ceramidases, kills many bacterial, viral, and fungal pathogens. In particular, sphingosine is an important natural component of the defense against bacterial pathogens in the respiratory tract. Pathologically reduced sphingosine levels in cystic fibrosis airway epithelial cells are normalized by inhalation of sphingosine, and coating plastic implants with sphingosine prevents bacterial infections. Pretreatment of cells with exogenous sphingosine also prevents the viral spike protein of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) from interacting with host cell receptors and inhibits the propagation of herpes simplex virus type 1 (HSV-1) in macrophages. Recent examinations reveal that the bactericidal effect of sphingosine might be due to bacterial membrane permeabilization and the subsequent death of the bacteria.

Inhibition of acid sphingomyelinase by ambroxol prevents SARS-CoV-2 entry into epithelial cells.

The acid sphingomyelinase/ceramide system has been shown to be important for cellular infection with at least some viruses, for instance, rhinovirus or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Functional inhibition of the acid sphingomyelinase using tricyclic antidepressants prevented infection of epithelial cells, for instance with SARS-CoV-2. The structure of ambroxol, that is, trans-4-[(2,4-dibromanilin-6-yl)-methyamino]-cyclohexanol, a mucolytic drug applied by inhalation, suggests that the drug might inhibit the acid sphingomyelinase and thereby infection with SARS-CoV-2. To test this, we used vesicular stomatitis virus pseudoviral particles presenting SARS-CoV-2 spike protein on their surface (pp-VSV-SARS-CoV-2 spike), a bona fide system for mimicking SARS-CoV-2 entry into cells. Viral uptake and formation of ceramide localization were determined by fluorescence microscopy, activity of the acid sphingomyelinase by consumption of [14C]sphingomyelin and ceramide was quantified by a kinase method. We found that entry of pp-VSV-SARS-CoV-2 spike required activation of acid sphingomyelinase and release of ceramide, events that were all prevented by pretreatment with ambroxol. We also obtained nasal epithelial cells from human volunteers prior to and after inhalation of ambroxol. Inhalation of ambroxol reduced acid sphingomyelinase activity in nasal epithelial cells and prevented pp-VSV-SARS-CoV-2 spike-induced acid sphingomyelinase activation, ceramide release, and entry of pp-VSV-SARS-CoV-2 spike ex vivo. The addition of purified acid sphingomyelinase or C16 ceramide restored entry of pp-VSV-SARS-CoV-2 spike into ambroxol-treated epithelial cells. We propose that ambroxol might be suitable for clinical studies to prevent coronavirus disease 2019.

Pharmacological Inhibition of Acid Sphingomyelinase Prevents Uptake of SARS-CoV-2 by Epithelial Cells.

The acid sphingomyelinase/ceramide system plays an important role in bacterial and viral infections. Here, we report that either pharmacological inhibition of acid sphingomyelinase with amitriptyline, imipramine, fluoxetine, sertraline, escitalopram, or maprotiline or genetic downregulation of the enzyme prevents infection of cultured cells or freshy isolated human nasal epithelial cells with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or vesicular stomatitis virus (VSV) pseudoviral particles (pp-VSV) presenting SARS-CoV-2 spike protein (pp-VSV-SARS-CoV-2 spike), a bona fide system mimicking SARS-CoV-2 infection. Infection activates acid sphingomyelinase and triggers a release of ceramide on the cell surface. Neutralization or consumption of surface ceramide reduces infection with pp-VSV-SARS-CoV-2 spike. Treating volunteers with a low dose of amitriptyline prevents infection of freshly isolated nasal epithelial cells with pp-VSV-SARS-CoV-2 spike. The data justify clinical studies investigating whether amitriptyline, a safe drug used clinically for almost 60 years, or other antidepressants that functionally block acid sphingomyelinase prevent SARS-CoV-2 infection.