TY - GEN A1 - Naser, Eyad A1 - Kadow, Stephanie A1 - Schumacher, Fabian A1 - Mohamed, Zainelabdeen H. A1 - Kappe, Christian A1 - Hessler, Gabriele A1 - Pollmeier, Barbara A1 - Kleuser, Burkhard A1 - Arenz, Christoph A1 - Becker, Katrin Anne A1 - Gulbins, Erich A1 - Carpinteiro, Alexander T1 - Characterization of the small molecule ARC39 BT - a direct and specific inhibitor of acid sphingomyelinase in vitro[S] T2 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Inhibition of acid sphingomyelinase (ASM), a lysosomal enzyme that catalyzes the hydrolysis of sphingomyelin into ceramide and phosphorylcholine, may serve as an investigational tool or a therapeutic intervention to control many diseases. Specific ASM inhibitors are currently not sufficiently characterized. Here, we found that 1-aminodecylidene bis-phosphonic acid (ARC39) specifically and efficiently (>90%) inhibits both lysosomal and secretory ASM in vitro. Results from investigating sphingomyelin phosphodiesterase 1 (SMPD1/Smpd1) mRNA and ASM protein levels suggested that ARC39 directly inhibits ASM's catalytic activity in cultured cells, a mechanism that differs from that of functional inhibitors of ASM. We further provide evidence that ARC39 dose- and time-dependently inhibits lysosomal ASM in intact cells, and we show that ARC39 also reduces platelet- and ASM-promoted adhesion of tumor cells. The observed toxicity of ARC39 is low at concentrations relevant for ASM inhibition in vitro, and it does not strongly alter the lysosomal compartment or induce phospholipidosis in vitro. When applied intraperitoneally in vivo, even subtoxic high doses administered short-term induced sphingomyelin accumulation only locally in the peritoneal lavage without significant accumulation in plasma, liver, spleen, or brain. These findings require further investigation with other possible chemical modifications. In conclusion, our results indicate that ARC39 potently and selectively inhibits ASM in vitro and highlight the need for developing compounds that can reach tissue concentrations sufficient for ASM inhibition in vivo. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1407 KW - sphingolipids KW - sphingomyelin KW - cerami-des KW - lipid metabolism KW - enzymology KW - lysosome KW - lysosomal hydrolases KW - acid ceramidase KW - bisphosphonates KW - functional inhibitors of acid sphin-gomyelinase KW - 1-aminodecylidene bis-phosphonic acid Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-516635 SN - 1866-8372 IS - 6 ER - TY - JOUR A1 - Naser, Eyad A1 - Kadow, Stephanie A1 - Schumacher, Fabian A1 - Mohamed, Zainelabdeen H. A1 - Kappe, Christian A1 - Hessler, Gabriele A1 - Pollmeier, Barbara A1 - Kleuser, Burkhard A1 - Arenz, Christoph A1 - Becker, Katrin Anne A1 - Gulbins, Erich A1 - Carpinteiro, Alexander T1 - Characterization of the small molecule ARC39 BT - a direct and specific inhibitor of acid sphingomyelinase in vitro[S] JF - Journal of Lipid Research N2 - Inhibition of acid sphingomyelinase (ASM), a lysosomal enzyme that catalyzes the hydrolysis of sphingomyelin into ceramide and phosphorylcholine, may serve as an investigational tool or a therapeutic intervention to control many diseases. Specific ASM inhibitors are currently not sufficiently characterized. Here, we found that 1-aminodecylidene bis-phosphonic acid (ARC39) specifically and efficiently (>90%) inhibits both lysosomal and secretory ASM in vitro. Results from investigating sphingomyelin phosphodiesterase 1 (SMPD1/Smpd1) mRNA and ASM protein levels suggested that ARC39 directly inhibits ASM's catalytic activity in cultured cells, a mechanism that differs from that of functional inhibitors of ASM. We further provide evidence that ARC39 dose- and time-dependently inhibits lysosomal ASM in intact cells, and we show that ARC39 also reduces platelet- and ASM-promoted adhesion of tumor cells. The observed toxicity of ARC39 is low at concentrations relevant for ASM inhibition in vitro, and it does not strongly alter the lysosomal compartment or induce phospholipidosis in vitro. When applied intraperitoneally in vivo, even subtoxic high doses administered short-term induced sphingomyelin accumulation only locally in the peritoneal lavage without significant accumulation in plasma, liver, spleen, or brain. These findings require further investigation with other possible chemical modifications. In conclusion, our results indicate that ARC39 potently and selectively inhibits ASM in vitro and highlight the need for developing compounds that can reach tissue concentrations sufficient for ASM inhibition in vivo. KW - sphingolipids KW - sphingomyelin KW - cerami-des KW - lipid metabolism KW - enzymology KW - lysosome KW - lysosomal hydrolases KW - acid ceramidase KW - bisphosphonates KW - functional inhibitors of acid sphin-gomyelinase KW - 1-aminodecylidene bis-phosphonic acid Y1 - 2021 U6 - https://doi.org/10.1194/jlr.RA120000682 SN - 1539-7262 SN - 0022-2275 VL - 61 IS - 6 SP - 896 EP - 910 PB - American Society for Biochemistry and Molecular Biology CY - Bethesda ER - TY - JOUR A1 - Fayyaz, Susann A1 - Japtok, Lukasz A1 - Schumacher, Fabian A1 - Wigger, Dominik A1 - Schulz, Tim Julius A1 - Haubold, Kathrin A1 - Gulbins, Erich A1 - Völler, Heinz A1 - Kleuser, Burkhard T1 - Lysophosphatidic acid inhibits insulin signaling in primary rat hepatocytes via the LPA(3) receptor subtype and is increased in obesity JF - Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry and pharmacology N2 - Background/Aims: Obesity is a main risk factor for the development of hepatic insulin resistance and it is accompanied by adipocyte hypertrophy and an elevated expression of different adipokines such as autotaxin (ATX). ATX converts lysophosphatidylcholine to lysophosphatidic acid (LPA) and acts as the main producer of extracellular LPA. This bioactive lipid regulates a broad range of physiological and pathological responses by activation of LPA receptors (LPA1-6). Methods: The activation of phosphatidylinositide 3-kinases (PI3K) signaling (Akt and GSK-3ß) was analyzed via western blotting in primary rat hepatocytes. Incorporation of glucose into glycogen was measured by using radio labeled glucose. Real-time PCR analysis and pharmacological modulation of LPA receptors were performed. Human plasma LPA levels of obese (BMI > 30, n = 18) and normal weight individuals (BMI 18.5-25, n = 14) were analyzed by liquid chromatography tandem-mass spectrometry (LC-MS/MS). Results: Pretreatment of primary hepatocytes with LPA resulted in an inhibition of insulin-mediated Gck expression, PI3K activation and glycogen synthesis. Pharmacological approaches revealed that the LPA3-receptor subtype is responsible for the inhibitory effect of LPA on insulin signaling. Moreover, human plasma LPA concentrations (16: 0 LPA) of obese participants (BMI > 30) are significantly elevated in comparison to normal weight individuals (BMI 18.5-25). Conclusion: LPA is able to interrupt insulin signaling in primary rat hepatocytes via the LPA3 receptor subtype. Moreover, the bioactive lipid LPA (16: 0) is increased in obesity. KW - Lysophosphatidic acid KW - Insulin signaling KW - Adipose tissue KW - Autotaxin KW - Hepatic insulin resistance KW - LPA(3) receptor subtype Y1 - 2017 U6 - https://doi.org/10.1159/000480470 SN - 1015-8987 SN - 1421-9778 VL - 43 SP - 445 EP - 456 PB - Karger CY - Basel ER - TY - JOUR A1 - Neuber, Corinna A1 - Schumacher, Fabian A1 - Gulbins, Erich A1 - Kleuser, Burkhard T1 - Mass Spectrometric Determination of Fatty Aldehydes Exemplified by Monitoring the Oxidative Degradation of (2E)-Hexadecenal in HepG2 Cell Lysates JF - Lipidomics N2 - Within the last few decades, liquid chromatography-mass spectrometry (LC-MS) has become a preferred method for manifold issues in analytical biosciences, given its high selectivity and sensitivity. However, the analysis of fatty aldehydes, which are important components of cell metabolism, remains challenging. Usually, chemical derivatization prior to MS detection is required to enhance ionization efficiency. In this regard, the coupling of fatty aldehydes to hydrazines like 2,4-dinitrophenylhydrazine (DNPH) is a common approach. Additionally, hydrazones readily react with fatty aldehydes to form stable derivatives, which can be easily separated using high-performance liquid chromatography (HPLC) and subsequently detected by MS. Here, we exemplarily present the quantification of the long-chain fatty aldehyde (2E)-hexadecenal, a break-down product of the bioactive lipid sphingosine 1-phosphate (S1P), after derivatization with 2-diphenylacetyl-1,3-indandione-1-hydrazone (DAIH) via isotope-dilution HPLC-electrospray ionization-quadrupole/time-of-flight (ESI-QTOF) MS. Moreover, we show that the addition of N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC hydrochloride) as a coupling agent allows for simultaneous determination of fatty aldehydes and fatty acids as DAIH derivatives. Taking advantage of this, we describe in detail how to monitor the degradation of (2E)-hexadecenal and the concurrent formation of its oxidation product (2E)-hexadecenoic acid in lysates of human hepatoblastoma (HepG2) cells within this chapter. KW - (2E)-hexadecenal KW - (2E)-hexadecenoic acid KW - Sphingosine 1-phosphate KW - Derivatization KW - DAIH KW - EDC KW - Isotope-dilution KW - HPLC-ESI-QTOF Y1 - 2017 SN - 978-1-4939-6946-3 SN - 978-1-4939-6944-9 U6 - https://doi.org/10.1007/978-1-4939-6946-3_10 SN - 0893-2336 SN - 1940-6045 VL - 125 SP - 147 EP - 158 PB - Humana Press CY - Totowa ER - TY - JOUR A1 - Hoehn, Richard S. A1 - Jernigan, Peter L. A1 - Japtok, Lukasz A1 - Chang, Alex L. A1 - Midura, Emily F. A1 - Caldwell, Charles C. A1 - Kleuser, Burkhard A1 - Lentsch, Alex B. A1 - Edwards, Michael J. A1 - Gulbins, Erich A1 - Pritts, Timothy A. T1 - Acid sphingomyelinase inhibition in stored erythrocytes reduces transfusion-associated lung inflammation JF - Annals of surgery : a monthly review of surgical science and practice N2 - Objective: We aimed to identify the role of the enzyme acid sphingomyelinase in the aging of stored units of packed red blood cells (pRBCs) and subsequent lung inflammation after transfusion. Summary Background Data: Large volume pRBC transfusions are associated with multiple adverse clinical sequelae, including lung inflammation. Microparticles are formed in stored pRBCs over time and have been shown to contribute to lung inflammation after transfusion. Methods: Human and murine pRBCs were stored with or without amitriptyline, a functional inhibitor of acid sphingomyelinase, or obtained from acid sphingomyelinase-deficient mice, and lung inflammation was studied in mice receiving transfusions of pRBCs and microparticles isolated from these units. Results: Acid sphingomyelinase activity in pRBCs was associated with the formation of ceramide and the release of microparticles. Treatment of pRBCs with amitriptyline inhibited acid sphingomyelinase activity, ceramide accumulation, and microparticle production during pRBC storage. Transfusion of aged pRBCs or microparticles isolated from aged blood into mice caused lung inflammation. This was attenuated after transfusion of pRBCs treated with amitriptyline or from acid sphingomyelinase-deficient mice. Conclusions: Acid sphingomyelinase inhibition in stored pRBCs offers a novel mechanism for improving the quality of stored blood. KW - acid sphingomyelinase KW - blood banking KW - ceramide KW - lung inflammation KW - microparticle Y1 - 2017 U6 - https://doi.org/10.1097/SLA.0000000000001648 SN - 0003-4932 SN - 1528-1140 VL - 265 IS - 1 SP - 218 EP - 226 PB - Lippincott Williams & Wilkins CY - Philadelphia ER - TY - JOUR A1 - Folkesson, Maggie A1 - Vorkapic, Emina A1 - Gulbins, Erich A1 - Japtok, Lukasz A1 - Kleuser, Burkhard A1 - Welander, Martin A1 - Länne, Toste A1 - Wågsäter, Dick T1 - Inflammatory cells, ceramides, and expression of proteases in perivascular adipose tissue adjacent to human abdominal aortic aneurysms JF - Journal of vascular surgery N2 - Background: Abdominal aortic aneurysm (AAA) is a deadly irreversible weakening and distension of the abdominal aortic wall. The pathogenesis of AAA remains poorly understood. Investigation into the physical and molecular characteristics of perivascular adipose tissue (PVAT) adjacent to AAA has not been done before and is the purpose of this study. Methods and Results: Human aortae, periaortic PVAT, and fat surrounding peripheral arteries were collected from patients undergoing elective surgical repair of AAA. Control aortas were obtained from recently deceased healthy organ donors with no known arterial disease. Aorta and PVAT was found in AAA to larger extent compared with control aortas. Immunohistochemistry revealed neutrophils, macrophages, mast cells, and T-cells surrounding necrotic adipocytes. Gene expression analysis showed that neutrophils, mast cells, and T-cells were found to be increased in PVAT compared with AAA as well as cathepsin K and S. The concentration of ceramides in PVAT was determined using mass spectrometry and correlated with content of T-cells in the PVAT. Conclusions: Our results suggest a role for abnormal necrotic, inflamed, proteolytic adipose tissue to the adjacent aneurysmal aortic wall in ongoing vascular damage. Y1 - 2016 U6 - https://doi.org/10.1016/j.jvs.2015.12.056 SN - 0741-5214 VL - 65 IS - 4 SP - 1171 EP - 1179 PB - Elsevier CY - New York ER - TY - JOUR A1 - Zoicas, Iulia A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Reichel, Martin A1 - Gulbins, Erich A1 - Fejtova, Anna A1 - Kornhuber, Johannes A1 - Rhein, Cosima T1 - The forebrain-specific overexpression of acid sphingomyelinase induces depressive-like symptoms in mice JF - Cells N2 - Human and murine studies identified the lysosomal enzyme acid sphingomyelinase (ASM) as a target for antidepressant therapy and revealed its role in the pathophysiology of major depression. In this study, we generated a mouse model with overexpression of Asm (Asm-tg(fb)) that is restricted to the forebrain to rule out any systemic effects of Asm overexpression on depressive-like symptoms. The increase in Asm activity was higher in male Asm-tg(fb) mice than in female Asm-tg(fb) mice due to the breeding strategy, which allows for the generation of wild-type littermates as appropriate controls. Asm overexpression in the forebrain of male mice resulted in a depressive-like phenotype, whereas in female mice, Asm overexpression resulted in a social anxiogenic-like phenotype. Ceramides in male Asm-tg(fb) mice were elevated specifically in the dorsal hippocampus. mRNA expression analyses indicated that the increase in Asm activity affected other ceramide-generating pathways, which might help to balance ceramide levels in cortical brain regions. This forebrain-specific mouse model offers a novel tool for dissecting the molecular mechanisms that play a role in the pathophysiology of major depression. KW - Smpd1 KW - acid sphingomyelinase KW - forebrain KW - depressive-like behavior KW - anxiety-like behavior KW - ceramide Y1 - 2020 VL - 9 IS - 5 PB - MDPI CY - Basel ER - TY - GEN A1 - Zoicas, Iulia A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Reichel, Martin A1 - Gulbins, Erich A1 - Fejtova, Anna A1 - Kornhuber, Johannes A1 - Rhein, Cosima T1 - The forebrain-specific overexpression of acid sphingomyelinase induces depressive-like symptoms in mice T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Human and murine studies identified the lysosomal enzyme acid sphingomyelinase (ASM) as a target for antidepressant therapy and revealed its role in the pathophysiology of major depression. In this study, we generated a mouse model with overexpression of Asm (Asm-tg(fb)) that is restricted to the forebrain to rule out any systemic effects of Asm overexpression on depressive-like symptoms. The increase in Asm activity was higher in male Asm-tg(fb) mice than in female Asm-tg(fb) mice due to the breeding strategy, which allows for the generation of wild-type littermates as appropriate controls. Asm overexpression in the forebrain of male mice resulted in a depressive-like phenotype, whereas in female mice, Asm overexpression resulted in a social anxiogenic-like phenotype. Ceramides in male Asm-tg(fb) mice were elevated specifically in the dorsal hippocampus. mRNA expression analyses indicated that the increase in Asm activity affected other ceramide-generating pathways, which might help to balance ceramide levels in cortical brain regions. This forebrain-specific mouse model offers a novel tool for dissecting the molecular mechanisms that play a role in the pathophysiology of major depression. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1186 KW - Smpd1 KW - acid sphingomyelinase KW - forebrain KW - depressive-like behavior KW - anxiety-like behavior KW - ceramide Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-524368 SN - 1866-8372 IS - 5 ER - TY - JOUR A1 - Becker, Katrin Anne A1 - Riethmueller, Joachim A1 - Seitz, Aaron P. A1 - Gardner, Aaron A1 - Boudreau, Ryan A1 - Kamler, Markus A1 - Kleuser, Burkhard A1 - Schuchman, Edward A1 - Caldwell, Charles C. A1 - Edwards, Michael J. A1 - Grassme, Heike A1 - Brodlie, Malcolm A1 - Gulbins, Erich T1 - Sphingolipids as targets for inhalation treatment of cystic fibrosis JF - Advanced drug delivery reviews N2 - Studies over the past several years have demonstrated the important role of sphingolipids in cystic fibrosis (CF), chronic obstructive pulmonary disease and acute lung injury. Ceramide is increased in airway epithelial cells and alveolar macrophages of CF mice and humans, while sphingosine is dramatically decreased. This increase in ceramide results in chronic inflammation, increased death of epithelial cells, release of DNA into the bronchial lumen and thereby an impairment of mucociliary clearance; while the lack of sphingosine in airway epithelial cells causes high infection susceptibility in CF mice and possibly patients. The increase in ceramide mediates an ectopic expression of beta 1-integrins in the luminal membrane of CF epithelial cells, which results, via an unknown mechanism, in a down-regulation of acid ceramidase. It is predominantly this down-regulation of acid ceramidase that results in the imbalance of ceramide and sphingosine in CF cells. Correction of ceramide and sphingosine levels can be achieved by inhalation of functional acid sphingomyelinase inhibitors, recombinant acid ceramidase or by normalization of beta 1-integrin expression and subsequent re-expression of endogenous acid ceramidase. These treatments correct pulmonary inflammation and prevent or treat, respectively, acute and chronic pulmonary infections in CF mice with Staphylococcus aureus and mucoid or non-mucoid Pseudomonas aeruginosa. Inhalation of sphingosine corrects sphingosine levels only and seems to mainly act against the infection. Many antidepressants are functional inhibitors of the acid sphingomyelinase and were designed for systemic treatment of major depression. These drugs could be repurposed to treat CF by inhalation. KW - Ceramide KW - Acid sphingomyelinase KW - Cystic fibrosis KW - COPD KW - Inhalation Y1 - 2018 U6 - https://doi.org/10.1016/j.addr.2018.04.015 SN - 0169-409X SN - 1872-8294 VL - 133 SP - 66 EP - 75 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Beckmann, Nadine A1 - Kadow, Stephanie A1 - Schumacher, Fabian A1 - Goethert, Joachim R. A1 - Kesper, Stefanie A1 - Draeger, Annette A1 - Schulz-Schaeffer, Walter J. A1 - Wang, Jiang A1 - Becker, Jan U. A1 - Kramer, Melanie A1 - Kuehn, Claudine A1 - Kleuser, Burkhard A1 - Becker, Katrin Anne A1 - Gulbins, Erich A1 - Carpinteiro, Alexander T1 - Pathological manifestations of Farber disease in a new mouse model JF - Biological chemistry N2 - Farber disease (FD) is a rare lysosomal storage disorder resulting from acid ceramidase deficiency and subsequent ceramide accumulation. No treatments are clinically available and affected patients have a severely shortened lifespan. Due to the low incidence, the pathogenesis of FD is still poorly understood. Here, we report a novel acid ceramidase mutant mouse model that enables the study of pathogenic mechanisms of FD and ceramide accumulation. Asah1(tmEx1) mice were generated by deletion of the acid ceramidase signal peptide sequence. The effects on lysosomal targeting and activity of the enzyme were assessed. Ceramide and sphingomyelin levels were quantified by liquid chromatography tandem-mass spectrometry (LC-MS/MS) and disease manifestations in several organ systems were analyzed by histology and biochemistry. We show that deletion of the signal peptide sequence disrupts lysosomal targeting and enzyme activity, resulting in ceramide and sphingomyelin accumulation. The affected mice fail to thrive and die early. Histiocytic infiltrations were observed in many tissues, as well as lung inflammation, liver fibrosis, muscular disease manifestations and mild kidney injury. Our new mouse model mirrors human FD and thus offers further insights into the pathogenesis of this disease. In the future, it may also facilitate the development of urgently needed therapies. KW - acid ceramidase KW - ceramide KW - Farber disease KW - lysosomal storage disorders Y1 - 2018 U6 - https://doi.org/10.1515/hsz-2018-0170 SN - 1431-6730 SN - 1437-4315 VL - 399 IS - 10 SP - 1183 EP - 1202 PB - De Gruyter CY - Berlin ER -