TY - JOUR A1 - Gerecke, Christian A1 - Scholtka, Bettina A1 - Loewenstein, Yvonne A1 - Fait, Isabel A1 - Gottschalk, Uwe A1 - Rogoll, Dorothee A1 - Melcher, Ralph A1 - Kleuser, Burkhard T1 - Hypermethylation of ITGA4, TFPI2 and VIMENTIN promoters is increased in inflamed colon tissue: putative risk markers for colitis-associated cancer JF - Journal of cancer research and clinical oncology : official organ of the Deutsche Krebsgesellschaft N2 - Epigenetic silencing of tumor suppressor genes is involved in early transforming events and has a high impact on colorectal carcinogenesis. Likewise, colon cancers that derive from chronically inflamed bowel diseases frequently exhibit epigenetic changes. But there is little data about epigenetic aberrations causing colorectal cancer in chronically inflamed tissue. The aim of the present study was to evaluate the aberrant gain of methylation in the gene promoters of VIM, TFPI2 and ITGA4 as putative early markers in the development from inflamed tissue via precancerous lesions toward colorectal cancer. Initial screening of different cancer cell lines by using methylation-specific PCR revealed a putative colon cancer-specific methylation pattern. Additionally, a demethylation assay was performed to investigate the methylation-dependent gene silencing of ITGA4. The candidate markers were analyzed in colonic tissue specimens from patients with colorectal cancer (n = 15), adenomas (n = 76), serrated lesions (n = 13), chronic inflammation (n = 10) and normal mucosal samples (n = 9). A high methylation frequency of VIM (55.6 %) was observed in normal colon tissue, whereas ITGA4 and TFPI2 were completely unmethylated in controls. A significant gain of methylation frequency with progression of disease as well as an age-dependent effect was detectable for TFPI2. ITGA4 methylation frequency was high in precancerous and cancerous tissues as well as in inflammatory bowel diseases (IBD). The already established methylation marker VIM does not permit a specific and sensitive discrimination of healthy and neoplastic tissue. The methylation markers ITGA4 and TFPI2 seem to be suitable risk markers for inflammation-associated colon cancer. KW - Epigenetic KW - DNA methylation KW - Colon cancer KW - Colitis KW - Gastrointestinal tract KW - Biomarker Y1 - 2015 U6 - https://doi.org/10.1007/s00432-015-1972-8 SN - 0171-5216 SN - 1432-1335 VL - 141 IS - 12 SP - 2097 EP - 2107 PB - Springer CY - New York ER - TY - JOUR A1 - Schumacher, Fabian A1 - Chakraborty, Sudipta A1 - Kleuser, Burkhard A1 - Gulbins, Erich A1 - Schwerdtle, Tanja A1 - Aschner, Michael A. A1 - Bornhorst, Julia T1 - Highly sensitive isotope-dilution liquid-chromatography-electrospray ionization-tandem-mass spectrometry approach to study the drug-mediated modulation of dopamine and serotonin levels in Caenorhabditis elegans JF - Talanta : the international journal of pure and applied analytical chemistry N2 - Dopamine (DA) and serotonin (SRT) are monoamine neurotransmitters that play a key role in regulating the central and peripheral nervous system. Their impaired metabolism has been implicated in several neurological disorders, such as Parkinson's disease and depression. Consequently, it is imperative to monitor changes in levels of these low-abundant neurotransmitters and their role in mediating disease. For the first time, a rapid, specific and sensitive isotope-dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantification of DA and SRT in the nematode Caenorhabditis elegans (C. elegans). This model organism offers a unique approach for studying the effect of various drugs and environmental conditions on neurotransmitter levels, given by the conserved DA and SRT biology, including synaptic release, trafficking and formation. We introduce a novel sample preparation protocol incorporating the usage of sodium thiosulfate in perchloric acid as extraction medium that assures high recovery of the relatively unstable neurotransmitters monitored. Moreover, the use of both deuterated internal standards and the multiple reaction monitoring (MRM) technique allows for unequivocal quantification. Thereby, to the best of our knowledge, we achieve a detection sensitivity that clearly exceeds those of published DA and SRT quantification methods in various matrices. We are the first to show that exposure of C elegans to the monoamine oxidase B (MAOB) inhibitor selegiline or the catechol-O-methyltransferase (COMT) inhibitor tolcapone, in order to block DA and SRT degradation, resulted in accumulation of the respective neurotransmitter. Assessment of a behavioral output of the dopaminergic system (basal slowing response) corroborated the analytical LC-MS/MS data. Thus, utilization of the C elegans model system in conjunction with our analytical method is well-suited to investigate drug-mediated modulation of the DA and SRT system in order to identify compounds with neuroprotective or regenerative properties. (C) 2015 Elsevier B.V. All rights reserved. KW - Caenorhabditis elegans KW - Dopamine KW - Serotonin KW - Liquid chromatography-tandem mass spectrometry KW - Isotope-dilution analysis Y1 - 2015 U6 - https://doi.org/10.1016/j.talanta.2015.05.057 SN - 0039-9140 SN - 1873-3573 VL - 144 SP - 71 EP - 79 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Nojima, Hiroyuki A1 - Freeman, Christopher M. A1 - Schuster, Rebecca M. A1 - Japtok, Lukasz A1 - Kleuser, Burkhard A1 - Edwards, Michael J. A1 - Gulbins, Erich A1 - Lentsch, Alex B. T1 - Hepatocyte exosomes mediate liver repair and regeneration via sphingosine-1-phosphate JF - Journal of hepatology N2 - Background & Aims: Exosomes are small membrane vesicles involved in intercellular communication. Hepatocytes are known to release exosomes, but little is known about their biological function. We sought to determine if exosomes derived from hepatocytes contribute to liver repair and regeneration after injury. Methods: Exosomes derived from primary murine hepatocytes were isolated and characterized biochemically and biophysically. Using cultures of primary hepatocytes, we tested whether hepatocyte exosomes induced proliferation of hepatocytes in vitro. Using models of ischemia/reperfusion injury and partial hepatectomy, we evaluated whether hepatocyte exosomes promote hepatocyte proliferation and liver regeneration in vivo. Results: Hepatocyte exosomes, but not exosomes from other liver cell types, induce dose-dependent hepatocyte proliferation in vitro and in vivo. Mechanistically, hepatocyte exosomes directly fuse with target hepatocytes and transfer neutral ceramidase and sphingosine kinase 2 (SK2) causing increased synthesis of sphingosine-1-phosphate (S1P) within target hepatocytes. Ablation of exosomal SK prevents the proliferative effect of exosomes. After ischemia/reperfusion injury, the number of circulating exosomes with proliferative effects increases. Conclusions: Our data shows that hepatocyte-derived exosomes deliver the synthetic machinery to form S1P in target hepatocytes resulting in cell proliferation and liver regeneration after ischemia/reperfusion injury or partial hepatectomy. These findings represent a potentially novel new contributing mechanism of liver regeneration and have important implications for new therapeutic approaches to acute and chronic liver disease. (C) 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved. KW - Liver injury KW - Sphingolipids KW - Sphingosine kinase KW - Ischemia/reperfusion KW - Transplantation Y1 - 2016 U6 - https://doi.org/10.1016/j.jhep.2015.07.030 SN - 0168-8278 SN - 1600-0641 VL - 64 SP - 60 EP - 68 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Sahle, Fitsum Feleke A1 - Balzus, Benjamin A1 - Gerecke, Christian A1 - Kleuser, Burkhard A1 - Bodmeier, Roland T1 - Formulation and in vitro evaluation of polymeric enteric nanoparticles as dermal carriers with pH-dependent targeting potential JF - European journal of pharmaceutical sciences : official journal of the European Federation for Pharmaceutical Sciences, EUFEPS N2 - pH-sensitive nanoparticles which release in a controlled fashion on the skin or dissolve in the hair follicle could significantly improve treatment effectiveness and make transfollicular drug delivery a success. Dexamethasone-loaded Eudragit L 100 nanoparticles were prepared by nanoprecipitation from an organic drug-polymer solution. Their toxicity potential was assessed using isolated human fibroblasts. pH-dependent swelling and erosion kinetics of the nanoparticles were investigated by dynamic light scattering and viscosity measurements and its effect on drug release was assessed in vitro with Franz diffusion cells. Stable, 100-550 nm-sized dexamethasone-loaded Eudragit L 100 nanoparticles with drug loading capacity and entrapment efficiency as high as 83% and 85%, respectively, were obtained by using polyvinyl alcohol as a stabilizer and ethanol as organic solvent The nanoparticles showed little or no toxicity on isolated normal human fibroblasts. Dexamethasone existed in the nanoparticles as solid solution or in amorphous form. The nanoparticles underwent extensive swelling and slow drug release in media with a low buffer capacity (as low as 10 mM) and a higher pH or at a pH close to the dissolution pH of the polymer (pH 6) and a higher buffer capacity. In 40 mM buffer and above pH 6.8, the nanoparticles eroded fast or dissolved completely and thus released the drug rapidly. pH-sensitive nanoparticles which potentially release in a controlled manner on the stratum corneum but dissolve in the hair follicle could be prepared. (C) 2016 Elsevier B.V. All rights reserved. KW - Dexamethasone KW - Enteric polymer KW - Eudragit L 100 KW - pH-sensitive nanoparticles KW - Skin nanocarrier KW - Erosion kinetics Y1 - 2016 U6 - https://doi.org/10.1016/j.ejps.2016.07.004 SN - 0928-0987 SN - 1879-0720 VL - 92 SP - 98 EP - 109 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Balzus, Benjamin A1 - Sahle, Fitsum Feleke A1 - Hönzke, Stefan A1 - Gerecke, Christian A1 - Schumacher, Fabian A1 - Hedtrich, Sarah A1 - Kleuser, Burkhard A1 - Bodmeier, Roland T1 - Formulation and ex vivo evaluation of polymeric nanoparticles for controlled delivery of corticosteroids to the skin and the corneal epithelium JF - European journal of pharmaceutics and biopharmaceutics : EJPB ; official journal of the International Association for Pharmaceutical Technology N2 - Controlled delivery of corticosteroids using nanoparticles to the skin and corneal epithelium may reduce their side effects and maximize treatment effectiveness. Dexamethasone-loaded ethyl cellulose, Eudragit® RS and ethyl cellulose/Eudragit® RS nanoparticles were prepared by the solvent evaporation method. Dexamethasone release from the polymeric nanoparticles was investigated in vitro using Franz diffusion cells. Drug penetration was also assessed ex vivo using excised human skin. Nanoparticle toxicity was determined by MTT and H2DCFDA assays. Eudragit® RS nanoparticles were smaller and positively charged but had a lower dexamethasone loading capacity (0.3–0.7%) than ethyl cellulose nanoparticles (1.4–2.2%). By blending the two polymers (1:1), small (105 nm), positively charged (+37 mV) nanoparticles with sufficient dexamethasone loading (1.3%) were obtained. Dexamethasone release and penetration significantly decreased with decreasing drug to polymer ratio and increased when Eudragit® RS was blended with ethyl cellulose. Ex vivo, drug release and penetration from the nanoparticles was slower than a conventional cream. The nanoparticles bear no toxicity potentials except ethyl cellulose nanoparticles had ROS generation potential at high concentration. In conclusion, the nanoparticles showed great potential to control the release and penetration of corticosteroids on the skin and mucus membrane and maximize treatment effectiveness. KW - Dermal delivery KW - Dexamethasone KW - Ethyl cellulose KW - Eudragit (R) RS KW - Ocular delivery KW - Polymeric nanoparticle Y1 - 2017 U6 - https://doi.org/10.1016/j.ejpb.2017.02.001 SN - 0939-6411 SN - 1873-3441 VL - 115 SP - 122 EP - 130 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Sahle, Fitsum Feleke A1 - Gerecke, Christian A1 - Kleuser, Burkhard A1 - Bodmeier, Roland T1 - Formulation and comparative in vitro evaluation of various dexamethasone-loaded pH-sensitive polymeric nanoparticles intended for dermal applications JF - International Journal of Pharmaceutics N2 - pH-sensitive nanoparticles have a great potential for dermal and transfollicular drug delivery. In this study, pH-sensitive, dexamethasone-loaded Eudragit (R) L 100, Eudragit (R) L 100-55, Eudragit (R) S 100, HPMCP-50, HPMCP-55 and cellulose acetate phthalate nanoparticles were prepared by nanoprecipitation and characterized. The pH-dependent swelling, erosion, dissolution and drug release kinetics were investigated in vitro using dynamic light scattering and Franz diffusion cells, respectively. Their toxicity potential was assessed by the ROS and MTT assays. 100-700 nm nanoparticles with high drug loading and entrapment efficiency were obtained. The nanoparticles bear no toxicity potential. Cellulose phthalates nanoparticles were more sensitive to pH than acrylates nanoparticles. They dissolved in 10 mM pH 7.5 buffer and released > 80% of the drug within 7 h. The acrylate nanoparticles dissolved in 40 mM pH 7.5 buffer and released 65-70% of the drug within 7 h. The nanoparticles remained intact in 10 and 40 mM pH 6.0 buffers (HPMCP nanoparticles dissolved in 40 mM pH 6.0 buffer) and released slowly. The nanoparticles properties could be modulated by blending the different polymers. In conclusion, various pH-sensitive nanoparticles that could release differently on the skin surface and dissolve and release in the hair follicles were obtained. KW - Cellulose acetate phthalate KW - Dexamethasone KW - Eudragit (R) KW - HPMCP KW - pH-sensitive nanoparticle KW - Skin nanocarrier Y1 - 2016 U6 - https://doi.org/10.1016/j.ijpharm.2016.11.029 SN - 0378-5173 SN - 1873-3476 VL - 516 IS - 1-2 SP - 21 EP - 31 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Hausmann, Christian A1 - Zoschke, Christian A1 - Wolff, Christopher A1 - Darvin, Maxim E. A1 - Sochorova, Michaela A1 - Kovacik, Andrej A1 - Wanjiku, Barbara A1 - Schumacher, Fabian A1 - Tigges, Julia A1 - Kleuser, Burkhard A1 - Lademann, Juergen A1 - Fritsche, Ellen A1 - Vavrova, Katerina A1 - Ma, Nan A1 - Schaefer-Korting, Monika T1 - Fibroblast origin shapes tissue homeostasis, epidermal differentiation, and drug uptake JF - Scientific reports N2 - Preclinical studies frequently lack predictive value for human conditions. Human cell-based disease models that reflect patient heterogeneity may reduce the high failure rates of preclinical research. Herein, we investigated the impact of primary cell age and body region on skin homeostasis, epidermal differentiation, and drug uptake. Fibroblasts derived from the breast skin of female 20- to 30-yearolds or 60- to 70-year-olds and fibroblasts from juvenile foreskin (<10 years old) were compared in cell monolayers and in reconstructed human skin (RHS). RHS containing aged fibroblasts differed from its juvenile and adult counterparts, especially in terms of the dermal extracellular matrix composition and interleukin-6 levels. The site from which the fibroblasts were derived appeared to alter fibroblast-keratinocyte crosstalk by affecting, among other things, the levels of granulocyte-macrophage colony-stimulating factor. Consequently, the epidermal expression of filaggrin and e-cadherin was increased in RHS containing breast skin fibroblasts, as were lipid levels in the stratum corneum. In conclusion, the region of the body from which fibroblasts are derived appears to affect the epidermal differentiation of RHS, while the age of the fibroblast donors determines the expression of proteins involved in wound healing. Emulating patient heterogeneity in preclinical studies might improve the treatment of age-related skin conditions. Y1 - 2019 U6 - https://doi.org/10.1038/s41598-019-39770-6 SN - 2045-2322 VL - 9 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Lu, Yong-Ping A1 - Reichetzeder, Christoph A1 - Prehn, Cornelia A1 - von Websky, Karoline A1 - Slowinski, Torsten A1 - Chen, You-Peng A1 - Yin, Liang-Hong A1 - Kleuser, Burkhard A1 - Yang, Xue-Song A1 - Adamski, Jerzy A1 - Hocher, Berthold T1 - Fetal serum metabolites are independently associated with Gestational diabetes mellitus JF - Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry and pharmacology N2 - Background/Aims: Gestational diabetes (GDM) might be associated with alterations in the metabolomic profile of affected mothers and their offspring. Until now, there is a paucity of studies that investigated both, the maternal and the fetal serum metabolome in the setting of GDM. Mounting evidence suggests that the fetus is not just passively affected by gestational disease but might play an active role in it. Metabolomic studies performed in maternal blood and fetal cord blood could help to better discern distinct fetal from maternal disease interactions. Methods: At the time of birth, serum samples from mothers and newborns (cord blood samples) were collected and screened for 163 metabolites utilizing tandem mass spectrometry. The cohort consisted of 412 mother/child pairs, including 31 cases of maternal GDM. Results: An initial non-adjusted analysis showed that eight metabolites in the maternal blood and 54 metabolites in the cord blood were associated with GDM. After Benjamini-Hochberg (BH) procedure and adjustment for confounding factors for GDM, fetal phosphatidylcholine acyl-alkyl C 32:1 and proline still showed an independent association with GDM. Conclusions: This study found metabolites in cord blood which were associated with GDM, even after adjustment for established risk factors of GDM. To the best of our knowledge, this is the first study demonstrating an independent association between fetal serum metabolites and maternal GDM. Our findings might suggest a potential effect of the fetal metabolome on maternal GDM. (c) 2018 The Author(s) Published by S. Karger AG, Basel KW - Gestational diabetes KW - Metabolomics KW - Phosphatidylcholine acyl-alkyl C 32:1 KW - Proline Y1 - 2018 U6 - https://doi.org/10.1159/000487119 SN - 1015-8987 SN - 1421-9778 VL - 45 IS - 2 SP - 625 EP - 638 PB - Karger CY - Basel ER - TY - JOUR A1 - Nitezki, Tina A1 - Kleuser, Burkhard A1 - Krämer, Stephanie T1 - Fatal gastric distension in a gold thioglucose mouse model of obesity JF - Laboratory Animals N2 - This case report addresses the problem of underreporting negative results and adverse side effects in animal testing. We present our findings regarding a hyperphagic mouse model associated with unforeseen high mortality. The results outline the necessity of reporting detailed information in the literature to avoid duplication. Obese mouse models are essential in the study of obesity, metabolic syndrome and diabetes mellitus. An experimental model of obesity can be induced by the administration of gold thioglucose (GTG). After transcending the blood-brain barrier, the GTG molecule interacts with regions of the ventromedial hypothalamus, thereby primarily targeting glucose-sensitive neurons. When these neurons are impaired, mice become insensitive to the satiety effects of glucose and develop hyperphagia. In a pilot study for optimising dosage and body weight development, C57BL/6 mice were treated with GTG (0.5 mg/g body weight) or saline, respectively. Animals were provided a physiological amount of standard diet (5 g per animal) for the first 24 hours after treatment to prevent gastric dilatation. Within 24 hours after GTG injection, all GTG-treated animals died of gastric overload and subsequent circulatory shock. Animals developed severe attacks of hyperphagia, and as the amount of provided chow was restricted, mice exhibited unforeseen pica and ingested bedding material. These observations strongly suggest that restricted feeding is contraindicated concerning GTG application. Presumably, the impulse of excessive food intake was a strong driving force. Therefore, the actual degree of suffering in the GTG-induced model of hyperphagia should be revised from moderate to severe. KW - appetite KW - distress KW - refinement KW - mortality Y1 - 2018 U6 - https://doi.org/10.1177/0023677218803384 SN - 0023-6772 SN - 1758-1117 VL - 53 IS - 1 SP - 89 EP - 94 PB - Sage Publ. CY - Thousand Oaks ER - TY - JOUR A1 - Böhm, Andreas A1 - Flößer, Anja A1 - Ermler, Swen A1 - Fender, Anke C. A1 - Lüth, Anja A1 - Kleuser, Burkhard A1 - Schrör, Karsten A1 - Rauch, Bernhard H. T1 - Factor-Xa-induced mitogenesis and migration require sphingosine kinase activity and S1P formation in human vascular smooth muscle cells JF - Cardiovascular research N2 - Sphingosine-1-phosphate (S1P) is a cellular signalling lipid generated by sphingosine kinase-1 (SPHK1). The aim of the study was to investigate whether the activated coagulation factor-X (FXa) regulates SPHK1 transcription and the formation of S1P and subsequent mitogenesis and migration of human vascular smooth muscle cells (SMC). FXa induced a time- (36 h) and concentration-dependent (330 nmol/L) increase of SPHK1 mRNA and protein expression in human aortic SMC, resulting in an increased synthesis of S1P. FXa-stimulated transcription of SPHK1 was mediated by the protease-activated receptor-1 (PAR-1) and PAR-2. In human carotid artery plaques, expression of SPHK1 was observed at SMC-rich sites and was co-localized with intraplaque FX/FXa content. FXa-induced SPHK1 transcription was attenuated by inhibitors of Rho kinase (Y27632) and by protein kinase C (PKC) isoforms (GF109203X). In addition, FXa rapidly induced the activation of the small GTPase Rho A. Inhibition of signalling pathways which regulate SPHK1 expression, inhibition of its activity or siRNA-mediated SPHK1 knockdown attenuated the mitogenic and chemotactic response of human SMC to FXa. These data suggest that FXa induces SPHK1 expression and increases S1P formation independent of thrombin and that this involves the activation of Rho A and PKC signalling. In addition to its key function in coagulation, this direct effect of FXa on human SMC may increase cell proliferation and migration at sites of vessel injury and thereby contribute to the progression of vascular lesions. KW - Factor-Xa KW - Atherosclerosis KW - Proliferation KW - Smooth muscle cells KW - Sphingosine kinase-1 Y1 - 2013 U6 - https://doi.org/10.1093/cvr/cvt112 SN - 0008-6363 VL - 99 IS - 3 SP - 505 EP - 513 PB - Oxford Univ. Press CY - Oxford ER - TY - JOUR A1 - Kakkassery, Vinodh A1 - Skosyrski, S. A1 - Lüth, A. A1 - Kleuser, Burkhard A1 - van der Giet, Maria A1 - Tate, R. A1 - Reinhard, J. A1 - Faissner, Andreas A1 - Joachim, Stephanie Christine A1 - Kociok, N. T1 - Etoposide Upregulates Survival Favoring Sphingosine-1-Phosphate in Etoposide-Resistant Retinoblastoma Cells JF - Pathology & Oncology Research N2 - Improved knowledge of retinoblastoma chemotherapy resistance is needed to raise treatment efficiency. The objective of this study was to test whether etoposide alters glucosyl-ceramide, ceramide, sphingosine, and sphingosine-1-phosphate (sphingosine-1-P) levels in parental retinoblastoma cells (WERI Rb1) or their etoposide-resistant subclones (WERI EtoR). WERI Rb1 and WERI EtoR were incubated with 400 ng/ml etoposide for 24 h. Levels of glucosyl-ceramides, ceramides, sphingosine, sphingosine-1-P were detected by Q-TOF mass spectrometry. Statistical analysis was done by ANOVA followed by Tukey post-hoc test (p < 0.05). The mRNA expression of sphingolipid pathways enzymes in WERI Rb1, WERI EtoR and four human retinoblastoma tissue samples was analyzed by quantitative real-time PCR. Pathways enzymes mRNA expression confirmed similarities of human sphingolipid metabolism in both cell lines and tissue samples, but different relative expression. Significant up-regulation of sphingosine was seen in both cell lines (p < 0.001). Only sphingosine-1-P up-regulation was significantly increased in WERI EtoR (p < 0.01), but not in WERI Rb1 (p > 0.2). Both cell lines upregulate pro-apoptotic sphingosine after etoposide incubation, but only WERI EtoR produces additional survival favorable sphingosine-1-P. These data may suggest a role of sphingosine-1-P in retinoblastoma chemotherapy resistance, although this seems not to be the only resistance mechanism. KW - Retinoblastoma KW - Sphingosine-1-phosphate KW - Chemotherapy resistance Y1 - 2017 U6 - https://doi.org/10.1007/s12253-017-0360-x SN - 1219-4956 SN - 1532-2807 VL - 25 IS - 1 SP - 391 EP - 399 PB - Springer CY - Dordrecht ER - TY - JOUR A1 - Döge, Nadine A1 - Hönzke, Stefan A1 - Schumacher, Fabian A1 - Balzus, Benjamin A1 - Colombo, Miriam A1 - Hadam, Sabrina A1 - Rancan, Fiorenza A1 - Blume-Peytavi, Ulrike A1 - Schäfer-Korting, Monika A1 - Schindler, Anke A1 - Rühl, Eckart A1 - Skov, Per Stahl A1 - Church, Martin K. A1 - Hedtrich, Sarah A1 - Kleuser, Burkhard A1 - Bodmeier, Roland A1 - Vogt, Annika T1 - Ethyl cellulose nanocarriers and nanocrystals differentially deliver dexamethasone into intact, tape-stripped or sodium lauryl sulfate-exposed ex vivo human skin - assessment by intradermal microdialysis and extraction from the different skin layers JF - Journal of controlled release N2 - Understanding penetration not only in intact, but also in lesional skin with impaired skin barrier function is important, in order to explore the surplus value of nanoparticle-based drug delivery for anti-inflammatory dermatotherapy. Herein, short-termex vivo cultures of (i) intact human skin, (ii) skin pretreated with tape-strippings and (iii) skin pre-exposed to sodium lauryl sulfate (SLS) were used to assess the penetration of dexamethasone (Dex). Intradermal microdialysis was utilized for up to 24 h after drug application as commercial cream, nanocrystals or ethyl cellulose nanocarriers applied at the therapeutic concentration of 0.05%, respectively. In addition, Dex was assessed in culture media and extracts from stratum corneum, epidermis and dermis after 24 h, and the results were compared to those in heat-separated split skin from studies in Franz diffusion cells. Providing fast drug release, nanocrystals significantly accelerated the penetration of Dex. In contrast to the application of cream and ethyl cellulose nanocarriers, Dex was already detectable in eluates after 6 h when applying nanocrystals on intact skin. Disruption of the skin barrier further accelerated and enhanced the penetration. Encapsulation in ethyl cellulose nanocarriers delayed Dex penetration. Interestingly, for all formulations highly increased concentrations in the dialysate were observed in tape-stripped skin, whereas the extent of enhancement was less in SLS-exposed skin. The results were confirmed in tissue extracts and were in line with the predictions made by in vitro release studies and ex vivo Franz diffusion cell experiments. The use of 45 kDa probes further enabled the collection of inflammatory cytokines. However, the estimation of glucocorticoid efficacy by Interleukin (IL)-6 and IL-8 analysis was limited due to the trauma induced by the probe insertion. Ex vivo intradermal microdialysis combined with culture media analysis provides an effective, skin-sparing method for preclinical assessment of novel drug delivery systems at therapeutic doses in models of diseased skin. (C) 2016 Elsevier B.V. All rights reserved. KW - Drug delivery systems KW - Polymeric nanoparticles KW - Dexamethasone KW - Microdialysis KW - Skin penetration KW - Skin barrier disruption Y1 - 2016 U6 - https://doi.org/10.1016/j.jconrel.2016.07.009 SN - 0168-3659 SN - 1873-4995 VL - 242 SP - 25 EP - 34 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Wetzel, Alexandra Nicole A1 - Scholtka, Bettina A1 - Gerecke, Christian A1 - Kleuser, Burkhard T1 - Epigenetic histone modulation contributes to improvements in inflammatory bowel disease via EBI3 JF - Cellular and molecular life sciences N2 - Ulcerative colitis (UC) is characterized by relapsing-remitting inflammatory episodes paralleled by varying cytokine levels, suggesting that switching epigenetic processes might be involved. However, the epigenetic impact on cytokine levels in colitis is mostly unexplored. The heterodimeric interleukin (IL)-12 cytokine family have various functions in both pro- and anti-inflammatory processes. The family member IL-35 (EBI3/IL-12p35) was recently reported to play an anti-inflammatory role in UC. Therefore, we aimed to investigate a possible epigenetic regulation of the IL-35 subunits in vitro and in vivo, and to examine the epigenetic targeting of EBI3 expression as a therapeutic option for UC. Exposure to either the pro-inflammatory TNF alpha or to histone deacetylase inhibitors (HDACi) significantly increased EBI3 expression in Human Colon Epithelial Cells (HCEC) generated from healthy tissue. When applied in combination, a drastic upregulation of EBI3 expression occurred, suggesting a synergistic mechanism. Consequently, IL-35 was increased as well. In vivo, the intestines of HDACi-treated wild-type mice exhibited reduced pathological signs of colitis compared to non-treated colitic mice. However, the improvement by HDACi treatment was completely lost in Ebi3-deficient mice (Ebi3(-/-)). In fact, HDACi appeared to exacerbate the disease phenotype in Ebi3(-/-). In conclusion, our results reveal that under inflammatory conditions, EBI3 is upregulated by the epigenetic mechanism of histone acetylation. The in vivo data show that the deficiency of EBI3 plays a key role in colitis manifestation. Concordantly, our data suggest that conditions promoting histone acetylation, such as upon HDACi application, improve colitis by a mechanism involving the local formation of the anti-inflammatory cytokine IL-35. KW - Histone deacetylase inhibitor KW - Inhibitory cytokines KW - Interleukin-35 KW - SAHA KW - Ulcerative colitis Y1 - 2020 U6 - https://doi.org/10.1007/s00018-020-03451-9 SN - 1420-682X SN - 1420-9071 VL - 77 IS - 23 SP - 5017 EP - 5030 PB - Springer International Publishing AG CY - Cham (ZG) ER - TY - JOUR A1 - Wetzel, Alexandra Nicole A1 - Scholtka, Bettina A1 - Schumacher, Fabian A1 - Rawel, Harshadrai Manilal A1 - Geisendörfer, Birte A1 - Kleuser, Burkhard T1 - Epigenetic DNA methylation of EBI3 modulates human interleukin-35 formation via NFkB signaling BT - a promising therapeutic option in ulcerative colitis JF - International journal of molecular sciences N2 - Ulcerative colitis (UC), a severe chronic disease with unclear etiology that is associated with increased risk for colorectal cancer, is accompanied by dysregulation of cytokines. Epstein-Barr virus-induced gene 3 (EBI3) encodes a subunit in the unique heterodimeric IL-12 cytokine family of either pro- or anti-inflammatory function. After having recently demonstrated that upregulation of EBI3 by histone acetylation alleviates disease symptoms in a dextran sulfate sodium (DSS)-treated mouse model of chronic colitis, we now aimed to examine a possible further epigenetic regulation of EBI3 by DNA methylation under inflammatory conditions. Treatment with the DNA methyltransferase inhibitor (DNMTi) decitabine (DAC) and TNF alpha led to synergistic upregulation of EBI3 in human colon epithelial cells (HCEC). Use of different signaling pathway inhibitors indicated NF kappa B signaling was necessary and proportional to the synergistic EBI3 induction. MALDI-TOF/MS and HPLC-ESIMS/MS analysis of DAC/TNF alpha-treated HCEC identified IL-12p35 as the most probable binding partner to form a functional protein. EBI3/IL-12p35 heterodimers (IL-35) induce their own gene upregulation, something that was indeed observed in HCEC cultured with media from previously DAC/TNF alpha-treated HCEC. These results suggest that under inflammatory and demethylating conditions the upregulation of EBI3 results in the formation of anti-inflammatory IL-35, which might be considered as a therapeutic target in colitis. KW - decitabine KW - DNMT inhibitor KW - EBI3 KW - inhibitory cytokines KW - interleukin-35 KW - TNF alpha KW - Ulcerative colitis Y1 - 2021 U6 - https://doi.org/10.3390/ijms22105329 SN - 1422-0067 VL - 22 IS - 10 PB - MDPI CY - Basel ER - TY - JOUR A1 - Giulbudagian, Michael A1 - Hönzke, Stefan A1 - Bergueiro, Julián A1 - Işık, Doğuş A1 - Schumacher, Fabian A1 - Saeidpour, Siavash A1 - Lohan, Silke A1 - Meinke, Martina A1 - Teutloff, Christian A1 - Schäfer-Korting, Monika A1 - Yealland, Guy A1 - Kleuser, Burkhard A1 - Hedtrich, Sarah A1 - Calderón, Marcelo T1 - Enhanced topical delivery of dexamethasone by beta-cyclodextrin decorated thermoresponsive nanogels JF - Nanoscale N2 - Highly hydrophilic, responsive nanogels are attractive as potential systems for the topical delivery of bioactives encapsulated in their three-dimensional polymeric scaffold. Yet, these drug carrier systems suffer from drawbacks for efficient delivery of hydrophobic drugs. Addressing this, β-cyclodextrin (βCD) could be successfully introduced into the drug carrier systems by exploiting its unique affinity toward dexamethasone (DXM) as well as its role as topical penetration enhancer. The properties of βCD could be combined with those of thermoresponsive nanogels (tNGs) based on dendritic polyglycerol (dPG) as a crosslinker and linear thermoresponsive polyglycerol (tPG) inducing responsiveness to temperature changes. Electron paramagnetic resonance (EPR) studies localized the drug within the hydrophobic cavity of βCD by differences in its mobility and environmental polarity. In fact, the fabricated carriers combining a particulate delivery system with a conventional penetration enhancer, resulted in an efficient delivery of DXM to the epidermis and the dermis of human skin ex vivo (enhancement compared to commercial DXM cream: ∼2.5 fold in epidermis, ∼30 fold in dermis). Furthermore, DXM encapsulated in βCD tNGs applied to skin equivalents downregulated the expression of proinflammatory thymic stromal lymphopoietin (TSLP) and outperformed a commercially available DXM cream. Y1 - 2017 U6 - https://doi.org/10.1039/c7nr04480a SN - 2040-3364 SN - 2040-3372 VL - 10 IS - 1 SP - 469 EP - 479 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Schraplau, Anne A1 - Schewe, Bettina A1 - Neuschäfer-Rube, Frank A1 - Ringel, Sebastian A1 - Neuber, Corinna A1 - Kleuser, Burkhard A1 - Püschel, Gerhard Paul T1 - Enhanced thyroid hormone breakdown in hepatocytes by mutual induction of the constitutive androstane receptor (CAR, NR1I3) and arylhydrocarbon receptor by benzo[a]pyrene and phenobarbital JF - Toxicology N2 - Xenobiotics may interfere with the hypothalamic-pituitary-thyroid endocrine axis by inducing enzymes that inactivate thyroid hormones and thereby reduce the metabolic rate. This induction results from an activation of xeno-sensing nuclear receptors. The current study shows that benzo[a]pyrene, a frequent contaminant of processed food and activator of the arylhydrocarbon receptor (AhR) activated the promoter and induced the transcription of the nuclear receptor constitutive androstane receptor (CAR, NR1I3) in rat hepatocytes. Likewise, phenobarbital induced the AhR transcription. This mutual induction of the nuclear receptors enhanced the phenobarbital-dependent induction of the prototypic CAR target gene Cyp2b1 as well as the AhR-dependent induction of UDP-glucuronosyltransferases. In both cases, the induction by the combination of both xenobiotics was more than the sum of the induction by either substance alone. By inducing the AhR, phenobarbital enhanced the benzo[a]pyrene-dependent reduction of thyroid hormone half-life and the benzo[a]pyrene-dependent increase in the rate of thyroid hormone glucuronide formation in hepatocyte cultures. CAR ligands might thus augment the endocrine disrupting potential of AhR activators by an induction of the AhR. (C) 2014 Elsevier Ireland Ltd. All rights reserved. KW - Endocrine disruption KW - Xenobesity KW - Aryl-hydrocarbon receptor KW - Cyp2b1 KW - Thyroid hormone KW - UDP-glucuronosyltransferase Y1 - 2015 U6 - https://doi.org/10.1016/j.tox.2014.12.004 SN - 0300-483X VL - 328 SP - 21 EP - 28 PB - Elsevier CY - Clare ER - TY - JOUR A1 - Kachler, Katerina A1 - Bailer, Maximilian A1 - Heim, Lisanne A1 - Schumacher, Fabian A1 - Reichel, Martin A1 - Holzinger, Corinna D. A1 - Trump, Sonja A1 - Mittler, Susanne A1 - Monti, Juliana A1 - Trufa, Denis I. A1 - Rieker, Ralf J. A1 - Hartmann, Arndt A1 - Sirbu, Horia A1 - Kleuser, Burkhard A1 - Kornhuber, Johannes A1 - Finotto, Susetta T1 - Enhanced acid sphingomyelinase activity drives immune evasion and tumor growth in non-small cell lung carcinoma JF - Cancer research N2 - The lipid hydrolase enzyme acid sphingomyelinase (ASM) is required for the conversion of the lipid cell membrane component sphingomyelin into ceramide. In cancer cells, ASM-mediated ceramide production is important for apoptosis, cell proliferation, and immune modulation, highlighting ASM as a potential multimodal therapeutic target. In this study, we demonstrate elevated ASM activity in the lung tumor environment and blood serum of patients with non-small cell lung cancer (NSCLC). RNAi-mediated attenuation of SMPD1 in human NSCLC cells rendered them resistant to serum starvation-induced apoptosis. In a murine model of lung adenocarcinoma, ASM deficiency reduced tumor development in a manner associated with significant enhancement of Th1-mediated and cytotoxic T-cell-mediated antitumor immunity. Our findings indicate that targeting ASM in NSCLC can act by tumor cell-intrinsic and-extrinsic mechanisms to suppress tumor cell growth, most notably by enabling an effective antitumor immune response by the host. (C) 2017 AACR. Y1 - 2017 U6 - https://doi.org/10.1158/0008-5472.CAN-16-3313 SN - 0008-5472 SN - 1538-7445 VL - 77 IS - 21 SP - 5963 EP - 5976 PB - American Association for Cancer Research CY - Philadelphia ER - TY - JOUR A1 - Henry, Brian D. A1 - Neill, Daniel R. A1 - Becker, Katrin Anne A1 - Gore, Suzanna A1 - Bricio-Moreno, Laura A1 - Ziobro, Regan A1 - Edwards, Michael J. A1 - Muehlemann, Kathrin A1 - Steinmann, Joerg A1 - Kleuser, Burkhard A1 - Japtok, Lukasz A1 - Luginbuehl, Miriam A1 - Wolfmeier, Heidi A1 - Scherag, Andre A1 - Gulbins, Erich A1 - Kadioglu, Aras A1 - Draeger, Annette A1 - Babiychuk, Eduard B. T1 - Engineered liposomes sequester bacterial exotoxins and protect from severe invasive infections in mice JF - Nature biotechnology : the science and business of biotechnology N2 - Gram-positive bacterial pathogens that secrete cytotoxic pore-forming toxins, such as Staphylococcus aureus and Streptococcus pneumoniae, cause a substantial burden of disease. Inspired by the principles that govern natural toxin-host interactions, we have engineered artificial liposomes that are tailored to effectively compete with host cells for toxin binding. Liposome-bound toxins are unable to lyse mammalian cells in vitro. We use these artificial liposomes as decoy targets to sequester bacterial toxins that are produced during active infection in vivo. Administration of artificial liposomes within 10 h after infection rescues mice from septicemia caused by S. aureus and S. pneumoniae, whereas untreated mice die within 24-33 h. Furthermore, liposomes protect mice against invasive pneumococcal pneumonia. Composed exclusively of naturally occurring lipids, tailored liposomes are not bactericidal and could be used therapeutically either alone or in conjunction with antibiotics to combat bacterial infections and to minimize toxin-induced tissue damage that occurs during bacterial clearance. Y1 - 2015 U6 - https://doi.org/10.1038/nbt.3037 SN - 1087-0156 SN - 1546-1696 VL - 33 IS - 1 SP - 81 EP - U295 PB - Nature Publ. Group CY - New York ER - TY - JOUR A1 - Bhabak, Krishna P. A1 - Kleuser, Burkhard A1 - Huwiler, Andrea A1 - Arenz, Christoph T1 - Effective inhibition of acid and neutral ceramidases by novel B-13 and LCL-464 analogues JF - Bioorganic & medicinal chemistry : a Tetrahedron publication for the rapid dissemination of full original research papers and critical reviews on biomolecular chemistry, medicinal chemistry and related disciplines N2 - Induction of apoptosis mediated by the inhibition of ceramidases has been shown to enhance the efficacy of conventional chemotherapy in several cancer models. Among the inhibitors of ceramidases reported in the literature, B-13 is considered as a lead compound having good in vitro potency towards acid ceramidase. Furthermore, owing to the poor activity of B-13 on lysosoamal acid ceramidase in living cells, LCL-464 a modified derivative of B-13 containing a basic omega-amino group at the fatty acid was reported to have higher potency towards lysosomal acid ceramidase in living cells. In a search for more potent inhibitors of ceramidases, we have designed a series of compounds with structural modifications of B-13 and LCL-464. In this study, we show that the efficacy of B-13 in vitro as well as in intact cells can be enhanced by suitable modification of functional groups. Furthermore, a detailed SAR investigation on LCL-464 analogues revealed novel promising inhibitors of aCDase and nCDase. In cell culture studies using the breast cancer cell line MDA-MB-231, some of the newly developed compounds elevated endogenous ceramide levels and in parallel, also induced apoptotic cell death. In summary, this study shows that structural modification of the known ceramidase inhibitors B-13 and LCL-464 generates more potent ceramidase inhibitors that are active in intact cells and not only elevates the cellular ceramide levels, but also enhances cell death. KW - Sphingolipids KW - Ceramide KW - Ceramidase inhibitors KW - Structure-activity-relationship Y1 - 2013 U6 - https://doi.org/10.1016/j.bmc.2012.12.014 SN - 0968-0896 VL - 21 IS - 4 SP - 874 EP - 882 PB - Elsevier CY - Oxford ER - TY - JOUR A1 - Schoenauer, Roman A1 - Larpin, Yu A1 - Babiychuk, Eduard B. A1 - Drucker, Patrick A1 - Babiychuk, Viktoriia S. A1 - Avota, Elita A1 - Schneider-Schaulies, Sibylle A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Koffel, Rene A1 - Draeger, Annette T1 - Down-regulation of acid sphingomyelinase and neutral sphingomyelinase-2 inversely determines the cellular resistance to plasmalemmal injury by pore-forming toxins JF - The FASEB journal : the official journal of the Federation of American Societies for Experimental Biology N2 - Bacterial pore-forming toxins compromise plasmalemmal integrity, leading to Ca2+ influx, leakage of the cytoplasm, and cell death. Such lesions can be repaired by microvesicular shedding or by the endocytic uptake of the injured membrane sites. Cells have at their disposal an entire toolbox of repair proteins for the identification and elimination of membrane lesions. Sphingomyelinases catalyze the breakdown of sphingomyelin into ceramide and phosphocholine. Sphingomyelin is predominantly localized in the outer leaflet, where it is hydrolyzed by acid sphingomyelinase (ASM) after lysosomal fusion with the plasma membrane. The magnesium-dependent neutral sphingomyelinase (NSM)-2 is found at the inner leaflet of the plasmalemma. Because either sphingomyelinase has been ascribed a role in the cellular stress response, we investigated their role in plasma membrane repair and cellular survival after treatment with the pore-forming toxins listeriolysin O (LLO) or pneumolysin (PLY). Jurkat T cells, in which ASM or NSM-2 was down-regulated [ASM knockdown (KD) or NSM-2 KD cells], showed inverse reactions to toxin-induced membrane damage: ASM KD cells displayed reduced toxin resistance, decreased viability, and defects in membrane repair. In contrast, the down-regulation of NSM-2 led to an increase in viability and enhanced plasmalemmal repair. Yet, in addition to the increased plasmalemmal repair, the enhanced toxin resistance of NSM-2 KD cells also appeared to be dependent on the activation of p38/MAPK, which was constitutively activated, whereas in ASM KD cells, the p38/MAPK activation was constitutively blunted.Schoenauer, R., Larpin, Y., Babiychuk, E. B., Drucker, P., Babiychuk, V. S., Avota, E., Schneider-Schaulies, S., Schumacher, F., Kleuser, B., Koffel, R., Draeger, A. Down-regulation of acid sphingomyelinase and neutral sphingomyelinase-2 inversely determines the cellular resistance to plasmalemmal injury by pore-forming toxins. KW - membrane repair KW - blebbing KW - calcium KW - bacterial toxins KW - annexins Y1 - 2018 U6 - https://doi.org/10.1096/fj.201800033R SN - 0892-6638 SN - 1530-6860 VL - 33 IS - 1 SP - 275 EP - 285 PB - Federation of American Societies for Experimental Biology CY - Bethesda ER - TY - JOUR A1 - Laeger, Thomas A1 - Castano-Martinez, Teresa A1 - Werno, Martin W. A1 - Japtok, Lukasz A1 - Baumeier, Christian A1 - Jonas, Wenke A1 - Kleuser, Burkhard A1 - Schürmann, Annette T1 - Dietary carbohydrates impair the protective effect of protein restriction against diabetes in NZO mice used as a model of type 2 diabetes JF - Diabetologia : journal of the European Association for the Study of Diabetes (EASD) N2 - Aims/hypothesis Low-protein diets are well known to improve glucose tolerance and increase energy expenditure. Increases in circulating fibroblast growth factor 21 (FGF21) have been implicated as a potential underlying mechanism. Methods We aimed to test whether low-protein diets in the context of a high-carbohydrate or high-fat regimen would also protect against type 2 diabetes in New Zealand Obese (NZO) mice used as a model of polygenetic obesity and type 2 diabetes. Mice were placed on high-fat diets that provided protein at control (16 kJ%; CON) or low (4 kJ%; low-protein/high-carbohydrate [LP/HC] or low-protein/high-fat [LP/HF]) levels. Results Protein restriction prevented the onset of hyperglycaemia and beta cell loss despite increased food intake and fat mass. The effect was seen only under conditions of a lower carbohydrate/fat ratio (LP/HF). When the carbohydrate/fat ratio was high (LP/HC), mice developed type 2 diabetes despite the robustly elevated hepatic FGF21 secretion and increased energy expenditure. Conclusion/interpretation Prevention of type 2 diabetes through protein restriction, without lowering food intake and body fat mass, is compromised by high dietary carbohydrates. Increased FGF21 levels and elevated energy expenditure do not protect against hyperglycaemia and type 2 diabetes per se. KW - Energy expenditure KW - FGF21 KW - Hyperglycaemia KW - Insulin resistance KW - NZO KW - Obesity KW - Protein restriction Y1 - 2018 U6 - https://doi.org/10.1007/s00125-018-4595-1 SN - 0012-186X SN - 1432-0428 VL - 61 IS - 6 SP - 1459 EP - 1469 PB - Springer CY - New York ER - TY - JOUR A1 - Rancan, Fiorenza A1 - Volkmann, Hildburg A1 - Giulbudagian, Michael A1 - Schumacher, Fabian A1 - Stanko, Jessica Isolde A1 - Kleuser, Burkhard A1 - Blume-Peytavi, Ulrike A1 - Calderon, Marcelo A1 - Vogt, Annika T1 - Dermal Delivery of the High-Molecular-Weight Drug Tacrolimus by Means of Polyglycerol-Based Nanogels JF - Pharmaceutics : Molecular Diversity Preservation International N2 - Polyglycerol-based thermoresponsive nanogels (tNGs) have been shown to have excellent skin hydration properties and to be valuable delivery systems for sustained release of drugs into skin. In this study, we compared the skin penetration of tacrolimus formulated in tNGs with a commercial 0.1% tacrolimus ointment. The penetration of the drug was investigated in ex vivo abdominal and breast skin, while different methods for skin barrier disruption were investigated to improve skin permeability or simulate inflammatory conditions with compromised skin barrier. The amount of penetrated tacrolimus was measured in skin extracts by liquid chromatography tandem-mass spectrometry (LC-MS/MS), whereas the inflammatory markers IL-6 and IL-8 were detected by enzyme-linked immunosorbent assay (ELISA). Higher amounts of tacrolimus penetrated in breast as compared to abdominal skin or in barrier-disrupted as compared to intact skin, confirming that the stratum corneum is the main barrier for tacrolimus skin penetration. The anti-proliferative effect of the penetrated drug was measured in skin tissue/Jurkat cells co-cultures. Interestingly, tNGs exhibited similar anti-proliferative effects as the 0.1% tacrolimus ointment. We conclude that polyglycerol-based nanogels represent an interesting alternative to paraffin-based formulations for the treatment of inflammatory skin conditions. KW - tacrolimus formulation KW - nanogels KW - skin penetration KW - drug delivery KW - human excised skin KW - Jurkat cells Y1 - 2019 U6 - https://doi.org/10.3390/pharmaceutics11080394 SN - 1999-4923 VL - 11 IS - 8 PB - MDPI CY - Basel ER - TY - JOUR A1 - Radbruch, Moritz A1 - Pischon, Hannah A1 - Ostrowski, Anja A1 - Volz, Pierre A1 - Brodwolf, Robert A1 - Neumann, Falko A1 - Unbehauen, Michael A1 - Kleuser, Burkhard A1 - Haag, Rainer A1 - Ma, Nan A1 - Alexiev, Ulrike A1 - Mundhenk, Lars A1 - Gruber, Achim D. T1 - Dendritic core-multishell nanocarriers in murine models of healthy and atopic skin JF - Nanoscale Research Letters N2 - Dendritic hPG-amid-C18-mPEG core-multishell nanocarriers (CMS) represent a novel class of unimolecular micelles that hold great potential as drug transporters, e. g., to facilitate topical therapy in skin diseases. Atopic dermatitis is among the most common inflammatory skin disorders with complex barrier alterations which may affect the efficacy of topical treatment. Here, we tested the penetration behavior and identified target structures of unloaded CMS after topical administration in healthy mice and in mice with oxazolone-induced atopic dermatitis. We further examined whole body distribution and possible systemic side effects after simulating high dosage dermal penetration by subcutaneous injection. Following topical administration, CMS accumulated in the stratum corneum without penetration into deeper viable epidermal layers. The same was observed in atopic dermatitis mice, indicating that barrier alterations in atopic dermatitis had no influence on the penetration of CMS. Following subcutaneous injection, CMS were deposited in the regional lymph nodes as well as in liver, spleen, lung, and kidney. However, in vitro toxicity tests, clinical data, and morphometry-assisted histopathological analyses yielded no evidence of any toxic or otherwise adverse local or systemic effects of CMS, nor did they affect the severity or course of atopic dermatitis. Taken together, CMS accumulate in the stratum corneum in both healthy and inflammatory skin and appear to be highly biocompatible in the mouse even under conditions of atopic dermatitis and thus could potentially serve to create a depot for anti-inflammatory drugs in the skin. KW - CMS KW - Skin KW - Topical treatment KW - Dermal delivery KW - Atopic dermatitis KW - Oxazolone KW - Fluorescence lifetime imaging microscopy KW - Nanomaterials KW - Multi-domain nanoparticles KW - Penetration enhancement Y1 - 2017 U6 - https://doi.org/10.1186/s11671-017-1835-0 SN - 1556-276X VL - 12 IS - 64 PB - Springer CY - New York ER - TY - JOUR A1 - Michels, Meta A1 - Japtok, Lukasz A1 - Alisjahbana, Bachti A1 - Wisaksana, Rudi A1 - Sumardi, Uun A1 - Puspita, Mita A1 - Kleuser, Burkhard A1 - de Mast, Quirijn A1 - van der Ven, Andre J. A. M. T1 - Decreased plasma levels of the endothelial protective sphingosine-1-phosphate are associated with dengue-induced plasma leakage JF - Journal of infection N2 - Background: A transient endothelial hyperpermeability is a hallmark of severe dengue infections. Sphingosine-1-phosphate (S1P) maintains vascular integrity and protects against plasma leakage. We related plasma S1P levels to dengue-induced plasma leakage and studied mechanisms that may underlie the decrease in S1P levels in dengue. Methods: We determined circulating levels of S1P in 44 Indonesian adults with acute dengue and related levels to plasma leakage, as determined by daily ultrasonography, and to levels of its chaperone apolipoprotein M, other lipoproteins and platelets. Results: Plasma S1P levels were decreased during dengue and patients with plasma leakage had lower median levels compared to those without (638 vs. 745 nM; p < 0.01). ApoM and other lipoprotein levels were also decreased during dengue, but did not correlate to S1P levels. Platelet counts correlated positively with S1P levels, but S1P levels were not higher in frozen-thawed platelet rich plasma, arguing against platelets as an important cellular source of S1P in dengue. Conclusions: Decreased plasma S1P levels during dengue are associated with plasma leakage. We speculate that decreased levels of ApoM underlies the lower S1P levels. Modulation of S1P levels and its receptors may be a novel therapeutic intervention to prevent plasma leakage in dengue. (C) 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved. KW - Sphingosine 1phosphate KW - Dengue KW - APOM protein KW - Human KW - Blood platelets Y1 - 2015 U6 - https://doi.org/10.1016/j.jinf.2015.06.014 SN - 0163-4453 SN - 1532-2742 VL - 71 IS - 4 SP - 480 EP - 487 PB - Elsevier CY - London ER - TY - JOUR A1 - Edlich, Alexander A1 - Volz, Pierre A1 - Brodwolf, Robert A1 - Unbehauen, Michael A1 - Mundhenk, Lars A1 - Gruber, Achim D. A1 - Hedtrich, Sarah A1 - Haag, Rainer A1 - Alexiev, Ulrike A1 - Kleuser, Burkhard T1 - Crosstalk between core-multishell nanocarriers for cutaneous drug delivery and antigen-presenting cells of the skin JF - Biomaterials : biomaterials reviews online N2 - Owing their unique chemical and physical properties core-multishell (CMS) nanocarriers are thought to underlie their exploitable biomedical use for a topical treatment of skin diseases. This highlights the need to consider not only the efficacy of CMS nanocarriers but also the potentially unpredictable and adverse consequences of their exposure thereto. As CMS nanocarriers are able to penetrate into viable layers of normal and stripped human skin ex vivo as well as in in vitro skin disease models the understanding of nanoparticle crosstalk with components of the immune system requires thorough investigation. Our studies highlight the biocompatible properties of CMS nanocarriers on Langerhans cells of the skin as they did neither induce cytotoxicity and genotoxicity nor cause reactive oxygen species (ROS) or an immunological response. Nevertheless, CMS nanocarriers were efficiently taken up by Langerhans cells via divergent endocytic pathways. Bioimaging of CMS nanocarriers by fluorescence lifetime imaging microscopy (FLIM) and flow cytometry indicated not only a localization within the lysosomes but also an energy-dependent exocytosis of unmodified CMS nanocarriers into the extracellular environment. (C) 2018 Elsevier Ltd. All rights reserved. KW - Core-multishell nanocarriers KW - Fluorescence lifetime imaging microscopy KW - Langerhans cells KW - Nanoparticle uptake KW - Nanotoxicology Y1 - 2018 U6 - https://doi.org/10.1016/j.biomaterials.2018.01.058 SN - 0142-9612 SN - 1878-5905 VL - 162 SP - 60 EP - 70 PB - Elsevier CY - Oxford ER - TY - JOUR A1 - Frombach, Janna A1 - Unbehauen, Michael A1 - Kurniasih, Indah N. A1 - Schumacher, Fabian A1 - Volz, Pierre A1 - Hadam, Sabrina A1 - Rancan, Fiorenza A1 - Blume-Peytavi, Ulrike A1 - Kleuser, Burkhard A1 - Haag, Rainer A1 - Alexiev, Ulrike A1 - Vogt, Annika T1 - Core-multishell nanocarriers enhance drug penetration and reach keratinocytes and antigen-presenting cells in intact human skin JF - Journal of controlled release N2 - In reconstructed skin and diffusion cell studies, core-multishell nanocarriers (CMS-NC) showed great potential for drug delivery across the skin barrier. Herein, we investigated penetration, release of dexamethasone (DXM), in excised full-thickness human skin with special focus on hair follicles (HF). Four hours and 16 h after topical application of clinically relevant dosages of 10 mu g DXM/cm(2) skin encapsulated in CMS-NC (12 nm diameter, 5.8% loading), presence of DXM in the tissue as assessed by fluorescence microscopy of anti-DXM-stained tissue sections as well as ELISA and HPLC-MS/MS in tissue extracts was enhanced compared to standard LAW-creme but lower compared to DXM aqueous/alcoholic solution. Such enhanced penetration compared to conventional cremes offers high potential for topical therapies, as recurrent applications of corticosteroid solutions face limitations with regard to tolerability and fast drainage. The findings encourage more detailed investigations on where and how the nanocarrier and drug dissociate within the skin and what other factors, e.g. thermodynamic activity, influence the penetration of this formulations. Microscopic studies on the spatial distribution within the skin revealed accumulation in HF and furrows accompanied by limited cellular uptake assessed by flow cytometry (up to 9% of total epidermal cells). FLIM clearly visualized the presence of CMS-NC in the viable epidermis and dermis. When exposed in situ a fraction of up to 25% CD1a(+) cells were found within the epidermal CMS-NC+ population compared to approximately 3% CD1a(+)/CMS-NC+ cells after in vitro exposure in short-term cultures of epidermal cell suspensions. The latter reflects the natural percentage of Langerhans cells (LC) in epidermis suspensions and indicated that CMS-NC were not preferentially internalized by one cell type. The increased CMS-NC+ LC proportion after exposure within the tissue is in accordance with the strategic suprabasal LC-localization. More specifically we postulate that the extensive dendrite meshwork, their position around HF orifices and their capacity to modulate tight junctions facilitated a preferential uptake of CMS-NC by LC within the skin. This newly identified aspect of CMS-NC penetration underlines the potential of CMS-NC for dermatotherapy and encourages further investigations of CMS-NC for the delivery of other molecule classes for which intracellular delivery is even more crucial. KW - Drug delivery KW - Skin penetration KW - Cellular uptake KW - Nanoparticles KW - Dendritic cells KW - High resolution microscopy Y1 - 2019 U6 - https://doi.org/10.1016/j.jconrel.2019.02.028 SN - 0168-3659 SN - 1873-4995 VL - 299 SP - 138 EP - 148 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Ahlberg, Sebastian A1 - Rancan, Fiorenza A1 - Epple, Matthias A1 - Loza, Kateryna A1 - Höppe, David A1 - Lademann, Jürgen A1 - Vogt, Annika A1 - Kleuser, Burkhard A1 - Gerecke, Christian A1 - Meinke, Martina C. T1 - Comparison of different methods to study effects of silver nanoparticles on the pro- and antioxidant status of human keratinocytes and fibroblasts JF - Methods : focusing on rapidly developing techniques KW - Oxidative stress KW - Dichlorofluorescein assay KW - Electron paramagnetic resonance spectroscopy KW - HaCaT cells KW - Glutathione KW - Free radicals Y1 - 2016 U6 - https://doi.org/10.1016/j.ymeth.2016.05.015 SN - 1046-2023 SN - 1095-9130 VL - 109 SP - 55 EP - 63 PB - Elsevier CY - San Diego ER - TY - JOUR A1 - Reichel, Martin A1 - Rhein, Cosima A1 - Hofmann, Lena M. A1 - Monti, Juliana A1 - Japtok, Lukasz A1 - Langgartner, Dominik A1 - Füchsl, Andrea M. A1 - Kleuser, Burkhard A1 - Gulbins, Erich A1 - Hellerbrand, Claus A1 - Reber, Stefan O. A1 - Kornhuber, Johannes T1 - Chronic Psychosocial Stress in Mice Is Associated With Increased Acid Sphingomyelinase Activity in Liver and Serum and With Hepatic C16:0-Ceramide Accumulation JF - Frontiers in Psychiatry N2 - Chronic psychosocial stress adversely affects human morbidity and is a risk factor for inflammatory disorders, liver diseases, obesity, metabolic syndrome, and major depressive disorder (MDD). In recent studies, we found an association of MDD with an increase of acid sphingomyelinase (ASM) activity. Thus, we asked whether chronic psychosocial stress as a detrimental factor contributing to the emergence of MDD would also affect ASM activity and sphingolipid (SL) metabolism. To induce chronic psychosocial stress in male mice we employed the chronic subordinate colony housing (CSC) paradigm and compared them to non-stressed single housed control (SHC) mice. We determined Asm activity in liver and serum, hepatic SL concentrations as well as hepatic mRNA expression of genes involved in SL metabolism. We found that hepatic Asm activity was increased by 28% (P = 0.006) and secretory Asm activity by 47% (P = 0.002) in stressed mice. C16:0-Cer was increased by 40% (P = 0.008). Gene expression analysis further revealed an increased expression of tumor necrosis factor (TNF)-alpha (P = 0.009) and of several genes involved in SL metabolism (Cers5, P = 0.028; Cers6, P = 0.045; Gba, P = 0.049; Gba2, P = 0.030; Ormdl2, P = 0.034; Smpdl3B; P = 0.013). Our data thus provides first evidence that chronic psychosocial stress, at least in mice, induces alterations in SL metabolism, which in turn might be involved in mediating the adverse health effects of chronic psychosocial stress and peripheral changes occurring in mood disorders. KW - chronic psychosocial stress KW - acid sphingomyelinase KW - ceramide KW - sphingolipid metabolism KW - chronic subordinate colony housing (CSC) KW - liver metabolism Y1 - 2018 U6 - https://doi.org/10.3389/fpsyt.2018.00496 SN - 1664-0640 VL - 9 PB - Frontiers Research Foundation CY - Lausanne ER - TY - JOUR A1 - Nojima, Hiroyuki A1 - Konishi, Takanori A1 - Freeman, Christopher M. A1 - Schuster, Rebecca M. A1 - Japtok, Lukasz A1 - Kleuser, Burkhard A1 - Edwards, Michael J. A1 - Gulbins, Erich A1 - Lentsch, Alex B. T1 - Chemokine Receptors, CXCR1 and CXCR2, Differentially Regulate Exosome Release in Hepatocytes JF - PLoS one N2 - Exosomes are small membrane vesicles released by different cell types, including hepatocytes, that play important roles in intercellular communication. We have previously demonstrated that hepatocyte-derived exosomes contain the synthetic machinery to form sphingosine-1-phosphate (S1P) in target hepatocytes resulting in proliferation and liver regeneration after ischemia/reperfusion (I/R) injury. We also demonstrated that the chemokine receptors, CXCR1 and CXCR2, regulate liver recovery and regeneration after I/R injury. In the current study, we sought to determine if the regulatory effects of CXCR1 and CXCR2 on liver recovery and regeneration might occur via altered release of hepatocyte exosomes. We found that hepatocyte release of exosomes was dependent upon CXCR1 and CXCR2. CXCR1-deficient hepatocytes produced fewer exosomes, whereas CXCR2-deficient hepatocytes produced more exosomes compared to their wild-type controls. In CXCR2-deficient hepatocytes, there was increased activity of neutral sphingomyelinase (Nsm) and intracellular ceramide. CXCR1-deficient hepatocytes had no alterations in Nsm activity or ceramide production. Interestingly, exosomes from CXCR1-deficient hepatocytes had no effect on hepatocyte proliferation, due to a lack of neutral ceramidase and sphingosine kinase. The data demonstrate that CXCR1 and CXCR2 regulate hepatocyte exosome release. The mechanism utilized by CXCR1 remains elusive, but CXCR2 appears to modulate Nsm activity and resultant production of ceramide to control exosome release. CXCR1 is required for packaging of enzymes into exosomes that mediate their hepatocyte proliferative effect. Y1 - 2016 U6 - https://doi.org/10.1371/journal.pone.0161443 SN - 1932-6203 VL - 11 SP - 6900 EP - + PB - PLoS CY - San Fransisco 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 - Wardelmann, Kristina A1 - Rath, Michaela A1 - Castro, José Pedro A1 - Blümel, Sabine A1 - Schell, Mareike A1 - Hauffe, Robert A1 - Schumacher, Fabian A1 - Flore, Tanina A1 - Ritter, Katrin A1 - Wernitz, Andreas A1 - Hosoi, Toru A1 - Ozawa, Koichiro A1 - Kleuser, Burkhard A1 - Weiß, Jürgen A1 - Schürmann, Annette A1 - Kleinridders, André T1 - Central acting Hsp10 regulates mitochondrial function, fatty acid metabolism and insulin sensitivity in the hypothalamus JF - Antioxidants N2 - Mitochondria are critical for hypothalamic function and regulators of metabolism. Hypothalamic mitochondrial dysfunction with decreased mitochondrial chaperone expression is present in type 2 diabetes (T2D). Recently, we demonstrated that a dysregulated mitochondrial stress response (MSR) with reduced chaperone expression in the hypothalamus is an early event in obesity development due to insufficient insulin signaling. Although insulin activates this response and improves metabolism, the metabolic impact of one of its members, the mitochondrial chaperone heat shock protein 10 (Hsp10), is unknown. Thus, we hypothesized that a reduction of Hsp10 in hypothalamic neurons will impair mitochondrial function and impact brain insulin action. Therefore, we investigated the role of chaperone Hsp10 by introducing a lentiviral-mediated Hsp10 knockdown (KD) in the hypothalamic cell line CLU-183 and in the arcuate nucleus (ARC) of C57BL/6N male mice. We analyzed mitochondrial function and insulin signaling utilizing qPCR, Western blot, XF96 Analyzer, immunohistochemistry, and microscopy techniques. We show that Hsp10 expression is reduced in T2D mice brains and regulated by leptin in vitro. Hsp10 KD in hypothalamic cells induced mitochondrial dysfunction with altered fatty acid metabolism and increased mitochondria-specific oxidative stress resulting in neuronal insulin resistance. Consequently, the reduction of Hsp10 in the ARC of C57BL/6N mice caused hypothalamic insulin resistance with acute liver insulin resistance. KW - brain insulin signaling KW - mitochondria KW - oxidative stress KW - fatty acid metabolism Y1 - 2021 U6 - https://doi.org/10.3390/antiox10050711 SN - 2076-3921 VL - 10 IS - 5 PB - MDPI CY - Basel ER - TY - JOUR A1 - Keller, Johannes A1 - Catala-Lehnen, Philip A1 - Huebner, Antje K. A1 - Jeschke, Anke A1 - Heckt, Timo A1 - Lueth, Anja A1 - Krause, Matthias A1 - Koehne, Till A1 - Albers, Joachim A1 - Schulze, Jochen A1 - Schilling, Sarah A1 - Haberland, Michael A1 - Denninger, Hannah A1 - Neven, Mona A1 - Hermans-Borgmeyer, Irm A1 - Streichert, Thomas A1 - Breer, Stefan A1 - Barvencik, Florian A1 - Levkau, Bodo A1 - Rathkolb, Birgit A1 - Wolf, Eckhard A1 - Calzada-Wack, Julia A1 - Neff, Frauke A1 - Gailus-Durner, Valerie A1 - Fuchs, Helmut A1 - de Angelis, Martin Hrabe A1 - Klutmann, Susanne A1 - Tsourdi, Elena A1 - Hofbauer, Lorenz C. A1 - Kleuser, Burkhard A1 - Chun, Jerold A1 - Schinke, Thorsten A1 - Amling, Michael T1 - Calcitonin controls bone formation by inhibiting the release of sphingosine 1-phosphate from osteoclasts JF - Nature Communications N2 - The hormone calcitonin (CT) is primarily known for its pharmacologic action as an inhibitor of bone resorption, yet CT-deficient mice display increased bone formation. These findings raised the question about the underlying cellular and molecular mechanism of CT action. Here we show that either ubiquitous or osteoclast-specific inactivation of the murine CT receptor (CTR) causes increased bone formation. CT negatively regulates the osteoclast expression of Spns2 gene, which encodes a transporter for the signalling lipid sphingosine 1-phosphate (S1P). CTR-deficient mice show increased S1P levels, and their skeletal phenotype is normalized by deletion of the S1P receptor S1P(3). Finally, pharmacologic treatment with the nonselective S1P receptor agonist FTY720 causes increased bone formation in wild-type, but not in S1P(3)-deficient mice. This study redefines the role of CT in skeletal biology, confirms that S1P acts as an osteoanabolic molecule in vivo and provides evidence for a pharmacologically exploitable crosstalk between osteoclasts and osteoblasts. Y1 - 2014 U6 - https://doi.org/10.1038/ncomms6215 SN - 2041-1723 VL - 5 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Henze, Andrea A1 - Homann, Thomas A1 - Rohn, Isabelle A1 - Aschner, Michael A. A1 - Link, Christopher D. A1 - Kleuser, Burkhard A1 - Schweigert, Florian J. A1 - Schwerdtle, Tanja A1 - Bornhorst, Julia T1 - Caenorhabditis elegans as a model system to study post-translational modifications of human transthyretin JF - Scientific reports N2 - The visceral protein transthyretin (TTR) is frequently affected by oxidative post-translational protein modifications (PTPMs) in various diseases. Thus, better insight into structure-function relationships due to oxidative PTPMs of TTR should contribute to the understanding of pathophysiologic mechanisms. While the in vivo analysis of TTR in mammalian models is complex, time-and resource-consuming, transgenic Caenorhabditis elegans expressing hTTR provide an optimal model for the in vivo identification and characterization of drug-mediated oxidative PTPMs of hTTR by means of matrix assisted laser desorption/ionization - time of flight - mass spectrometry (MALDI-TOF-MS). Herein, we demonstrated that hTTR is expressed in all developmental stages of Caenorhabditis elegans, enabling the analysis of hTTR metabolism during the whole life-cycle. The suitability of the applied model was verified by exposing worms to D-penicillamine and menadione. Both drugs induced substantial changes in the oxidative PTPM pattern of hTTR. Additionally, for the first time a covalent binding of both drugs with hTTR was identified and verified by molecular modelling. Y1 - 2016 U6 - https://doi.org/10.1038/srep37346 SN - 2045-2322 VL - 6 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Henze, Andrea A1 - Homann, Thomas A1 - Rohn, Isabelle A1 - Aschner, Michael A. A1 - Link, Christopher D. A1 - Kleuser, Burkhard A1 - Schweigert, Florian J. A1 - Schwerdtle, Tanja A1 - Bornhorst, Julia T1 - Caenorhabditis elegans as a model system to study post-translational modifications of human transthyretin JF - Scientific reports N2 - The visceral protein transthyretin (TTR) is frequently affected by oxidative post-translational protein modifications (PTPMs) in various diseases. Thus, better insight into structure-function relationships due to oxidative PTPMs of TTR should contribute to the understanding of pathophysiologic mechanisms. While the in vivo analysis of TTR in mammalian models is complex, time- and resource-consuming, transgenic Caenorhabditis elegans expressing hTTR provide an optimal model for the in vivo identification and characterization of drug-mediated oxidative PTPMs of hTTR by means of matrix assisted laser desorption/ionization – time of flight – mass spectrometry (MALDI-TOF-MS). Herein, we demonstrated that hTTR is expressed in all developmental stages of Caenorhabditis elegans, enabling the analysis of hTTR metabolism during the whole life-cycle. The suitability of the applied model was verified by exposing worms to D-penicillamine and menadione. Both drugs induced substantial changes in the oxidative PTPM pattern of hTTR. Additionally, for the first time a covalent binding of both drugs with hTTR was identified and verified by molecular modelling. KW - n-acetyl-cysteine KW - s-glutathionylation KW - force-field KW - c. elegans KW - life-span KW - protein KW - cells KW - menadione KW - disease KW - binding Y1 - 2016 U6 - https://doi.org/10.1038/srep37346 SN - 2045-2322 VL - 6 PB - Nature Publishing Group CY - London ER - TY - JOUR A1 - Beckmann, Nadine A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Gulbins, Erich A1 - Nomellini, Vanessa A1 - Caldwell, Charles C. T1 - Burn injury impairs neutrophil chemotaxis through increased ceramide JF - Shock : injury, inflammation, and sepsis, laboratory and clinical approaches N2 - Infection is a common and often deadly complication after burn injury. A major underlying factor is burn-induced immune dysfunction, particularly with respect to neutrophils as the primary responders to infection. Temporally after murine scald injury, we demonstrate impaired bone marrow neutrophil chemotaxis toward CXCL1 ex vivo. Additionally, we observed a reduced recruitment of neutrophils to the peritoneal after elicitation 7 days after injury. We demonstrate that neutrophil ceramide levels increase after burn injury, and this is associated with decreased expression of CXCR2 and blunted chemotaxis. A major signaling event upon CXCR2 activation is Akt phosphorylation and this was reduced when ceramide was elevated. In contrast, PTEN levels were elevated and PTEN-inhibition elevated phospho-Akt levels and mitigated the burn-induced neutrophil chemotaxis defect. Altogether, this study identifies a newly described pathway of ceramide-mediated suppression of neutrophil chemotaxis after burn injury and introduces potential targets to mitigate this defect and reduce infection-related morbidity and mortality after burn. KW - Acid sphingomyelinase KW - Akt KW - burn injury KW - ceramide KW - CXCR2 KW - immune KW - dysfunction KW - neutrophil chemotaxis KW - PTEN Y1 - 2021 U6 - https://doi.org/10.1097/SHK.0000000000001693 SN - 1073-2322 SN - 1540-0514 VL - 56 IS - 1 SP - 125 EP - 132 PB - Lippincott Williams & Wilkins CY - Hagerstown, Md. ER - TY - JOUR A1 - Giulbudagian, Michael A1 - Yealland, Guy A1 - Hönzke, S. A1 - Edlich, A. A1 - Geisendörfer, Birte A1 - Kleuser, Burkhard A1 - Hedtrich, Sarah A1 - Calderon, Marcelo T1 - Breaking the Barrier BT - potent anti-inflammatory activity following efficient topical delivery of etanercept using thermoresponsive nanogels JF - Theranostics N2 - Topical administration permits targeted, sustained delivery of therapeutics to human skin. Delivery to the skin, however, is typically limited to lipophilic molecules with molecular weight of < 500 Da, capable of crossing the stratum corneum. Nevertheless, there are indications protein delivery may be possible in barrier deficient skin, a condition found in several inflammatory skin diseases such as psoriasis, using novel nanocarrier systems. Methods: Water in water thermo-nanoprecipitation; dynamic light scattering; zeta potential measurement; nanoparticle tracking analysis; atomic force microscopy; cryogenic transmission electron microscopy; UV absorption; centrifugal separation membranes; bicinchoninic acid assay; circular dichroism; TNF alpha binding ELISA; inflammatory skin equivalent construction; human skin biopsies; immunohistochemistry; fluorescence microscopy; western blot; monocyte derived Langerhans cells; ELISA Results: Here, we report the novel synthesis of thermoresponsive nanogels (tNG) and the stable encapsulation of the anti-TNFa fusion protein etanercept (ETR) (similar to 150 kDa) without alteration to its structure, as well as temperature triggered release from the tNGs. Novel tNG synthesis without the use of organic solvents was conducted, permitting in situ encapsulation of protein during assembly, something that holds great promise for easy manufacture and storage. Topical application of ETR loaded tNGs to inflammatory skin equivalents or tape striped human skin resulted in efficient ETR delivery throughout the SC and into the viable epidermis that correlated with clear anti-inflammatory effects. Notably, effective ETR delivery depended on temperature triggered release following topical application. Conclusion: Together these results indicate tNGs hold promise as a biocompatible and easy to manufacture vehicle for stable protein encapsulation and topical delivery into barrier-deficient skin. KW - thermoresponsive-nanogel KW - topical KW - anti-inflammatory therapy KW - etanercept KW - skin equivalents Y1 - 2018 U6 - https://doi.org/10.7150/thno.21668 SN - 1838-7640 VL - 8 IS - 2 SP - 450 EP - 463 PB - Ivyspring International Publisher CY - Lake haven ER - TY - JOUR A1 - Gerecke, Christian A1 - Edlich, Alexander A1 - Giulbudagian, Michael A1 - Schumacher, Fabian A1 - Zhang, Nan A1 - Said, Andre A1 - Yealland, Guy A1 - Lohan, Silke B. A1 - Neumann, Falko A1 - Meinke, Martina C. A1 - Ma, Nan A1 - Calderon, Marcelo A1 - Hedtrich, Sarah A1 - Schaefer-Korting, Monika A1 - Kleuser, Burkhard T1 - Biocompatibility and characterization of polyglycerol-based thermoresponsive nanogels designed as novel drug-delivery systems and their intracellular localization in keratinocytes JF - Nanotoxicology N2 - Novel nanogels that possess the capacity to change their physico-chemical properties in response to external stimuli are promising drug-delivery candidates for the treatment of severe skin diseases. As thermoresponsive nanogels (tNGs) are capable of enhancing penetration through biological barriers such as the stratum corneum and are taken up by keratinocytes of human skin, potential adverse consequences of their exposure must be elucidated. In this study, tNGs were synthesized from dendritic polyglycerol (dPG) and two thermoresponsive polymers. tNG_dPG_tPG are the combination of dPG with poly(glycidyl methyl ether-co-ethyl glycidyl ether) (p(GME-co-EGE)) and tNG_dPG_pNIPAM the one with poly(N-isopropylacrylamide) (pNIPAM). Both thermoresponsive nanogels are able to incorporate high amounts of dexamethasone and tacrolimus, drugs used in the treatment of severe skin diseases. Cellular uptake, intracellular localization and the toxicological properties of the tNGs were comprehensively characterized in primary normal human keratinocytes (NHK) and in spontaneously transformed aneuploid immortal keratinocyte cell line from adult human skin (HaCaT). Laser scanning confocal microscopy revealed fluorescently labeled tNGs entered into the cells and localized predominantly within lysosomal compartments. MTT assay, comet assay and carboxy-H2DCFDA assay, demonstrated neither cytotoxic or genotoxic effects, nor any induction of reactive oxygen species of the tNGs in keratinocytes. In addition, both tNGs were devoid of eye irritation potential as shown by bovine corneal opacity and permeability (BCOP) test and red blood cell (RBC) hemolysis assay. Therefore, our study provides evidence that tNGs are locally well tolerated and underlines their potential for cutaneous drug delivery. KW - Drug delivery KW - nanoparticles KW - particle characterization KW - keratinocytes KW - nanotoxicology Y1 - 2017 U6 - https://doi.org/10.1080/17435390.2017.1292371 SN - 1743-5390 SN - 1743-5404 VL - 11 SP - 267 EP - 277 PB - Routledge, Taylor & Francis Group CY - Abingdon ER - TY - JOUR A1 - Dwi Putra, Sulistyo Emantoko A1 - Reichetzeder, Christoph A1 - Hasan, Ahmed Abdallah Abdalrahman Mohamed A1 - Slowinski, Torsten A1 - Chu, Chang A1 - Krämer, Bernhard K. A1 - Kleuser, Burkhard A1 - Hocher, Berthold T1 - Being born large for gestational age is associated with increased global placental DNA methylation JF - Scientific Reports N2 - Being born small (SGA) or large for gestational age (LGA) is associated with adverse birth outcomes and metabolic diseases in later life of the offspring. It is known that aberrations in growth during gestation are related to altered placental function. Placental function is regulated by epigenetic mechanisms such as DNA methylation. Several studies in recent years have demonstrated associations between altered patterns of DNA methylation and adverse birth outcomes. However, larger studies that reliably investigated global DNA methylation are lacking. The aim of this study was to characterize global placental DNA methylation in relationship to size for gestational age. Global DNA methylation was assessed in 1023 placental samples by LC-MS/MS. LGA offspring displayed significantly higher global placental DNA methylation compared to appropriate for gestational age (AGA; p<0.001). ANCOVA analyses adjusted for known factors impacting on DNA methylation demonstrated an independent association between placental global DNA methylation and LGA births (p<0.001). Tertile stratification according to global placental DNA methylation levels revealed a significantly higher frequency of LGA births in the third tertile. Furthermore, a multiple logistic regression analysis corrected for known factors influencing birth weight highlighted an independent positive association between global placental DNA methylation and the frequency of LGA births (p=0.001). KW - fetal origins hypothesis KW - birth weight KW - repetitive elements KW - glucocorticoid receptor KW - nutrient transport KW - growth restriction KW - later health KW - pregnancy KW - genes KW - patterns Y1 - 2020 U6 - https://doi.org/10.1038/s41598-020-57725-0 SN - 2045-2322 VL - 10 IS - 1 SP - 1 EP - 10 PB - Springer Nature CY - London ER - TY - JOUR A1 - Fink, Julian A1 - Schumacher, Fabian A1 - Schlegel, Jan A1 - Stenzel, Philipp A1 - Wigger, Dominik A1 - Sauer, Markus A1 - Kleuser, Burkhard A1 - Seibel, Jürgen T1 - Azidosphinganine enables metabolic labeling and detection of sphingolipid de novo synthesis JF - Organic & biomolecular chemistry : an international journal of synthetic, physical and biomolecular organic chemistry N2 - Here were report the combination of biocompatible click chemistry of omega-azidosphinganine with fluorescence microscopy and mass spectrometry as a powerful tool to elaborate the sphingolipid metabolism. The azide probe was efficiently synthesized over 13 steps starting from l-serine in an overall yield of 20% and was used for live-cell fluorescence imaging of the endoplasmic reticulum in living cells by bioorthogonal click reaction with a DBCO-labeled fluorophore revealing that the incorporated analogue is mainly localized in the endoplasmic membrane like the endogenous species. A LC-MS(/MS)-based microsomal in vitro assay confirmed that omega-azidosphinganine mimics the natural species enabling the identification and analysis of metabolic breakdown products of sphinganine as a key starting intermediate in the complex sphingolipid biosynthetic pathways. Furthermore, the sphinganine-fluorophore conjugate after click reaction was enzymatically tolerated to form its dihydroceramide and ceramide metabolites. Thus, omega-azidosphinganine represents a useful biofunctional tool for metabolic investigations both by in vivo fluorescence imaging of the sphingolipid subcellular localization in the ER and by in vitro high-resolution mass spectrometry analysis. This should reveal novel insights of the molecular mechanisms sphingolipids and their processing enzymes have e.g. in infection. Y1 - 2021 U6 - https://doi.org/10.1039/d0ob02592e SN - 1477-0520 SN - 1477-0539 VL - 19 IS - 10 SP - 2203 EP - 2212 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Müller, S. M. A1 - Finke, Hannah A1 - Ebert, Franziska A1 - Kopp, Johannes Florian A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Francesconi, Kevin A. A1 - Raber, G. A1 - Schwerdtle, Tanja T1 - Arsenic-containing hydrocarbons BT - effects on gene expression, epigenetics, and biotransformation in HepG2 cells JF - Archives of toxicology : official journal of EUROTOX N2 - Arsenic-containing hydrocarbons (AsHCs), a subgroup of arsenolipids found in fish and algae, elicit substantial toxic effects in various human cell lines and have a considerable impact on cellular energy levels. The underlying mode of action, however, is still unknown. The present study analyzes the effects of two AsHCs (AsHC 332 and AsHC 360) on the expression of 44 genes covering DNA repair, stress response, cell death, autophagy, and epigenetics via RT-qPCR in human liver (HepG2) cells. Both AsHCs affected the gene expression, but to different extents. After treatment with AsHC 360, flap structure-specific endonuclease 1 (FEN1) as well as xeroderma pigmentosum group A complementing protein (XPA) and (cytosine-5)-methyltransferase 3A (DNMT3A) showed time- and concentration-dependent alterations in gene expression, thereby indicating an impact on genomic stability. In the subsequent analysis of epigenetic markers, within 72 h, neither AsHC 332 nor AsHC 360 showed an impact on the global DNA methylation level, whereas incubation with AsHC 360 increased the global DNA hydroxymethylation level. Analysis of cell extracts and cell media by HPLC-mass spectrometry revealed that both AsHCs were considerably biotransformed. The identified metabolites include not only the respective thioxo-analogs of the two AsHCs, but also several arsenic-containing fatty acids and fatty alcohols, contributing to our knowledge of biotransformation mechanisms of arsenolipids. KW - Arsenolipids KW - Gene expression KW - Arsenic-containing hydrocarbons KW - Global DNA methylation KW - Arsenic speciation KW - Metabolism Y1 - 2018 U6 - https://doi.org/10.1007/s00204-018-2194-z SN - 0340-5761 SN - 1432-0738 VL - 92 IS - 5 SP - 1751 EP - 1765 PB - Springer CY - Heidelberg ER - TY - JOUR A1 - Gulbins, Anne A1 - Schumacher, Fabian A1 - Becker, Katrin Anne A1 - Wilker, Barbara A1 - Soddemann, Matthias A1 - Boldrin, Francesco A1 - Müller, Christian P. A1 - Edwards, Michael J. A1 - Goodman, Michael A1 - Caldwell, Charles C. A1 - Kleuser, Burkhard A1 - Kornhuber, Johannes A1 - Szabo, Ildiko A1 - Gulbins, Erich T1 - Antidepressants act by inducing autophagy controlled by sphingomyelin-ceramide JF - Molecular psychiatry N2 - Major depressive disorder (MDD) is a common and severe disease characterized by mood changes, somatic alterations, and often suicide. MDD is treated with antidepressants, but the molecular mechanism of their action is unknown. We found that widely used antidepressants such as amitriptyline and fluoxetine induce autophagy in hippocampal neurons via the slow accumulation of sphingomyelin in lysosomes and Golgi membranes and of ceramide in the endoplasmic reticulum (ER). ER ceramide stimulates phosphatase 2A and thereby the autophagy proteins Ulk, Beclin, Vps34/Phosphatidylinositol 3-kinase, p62, and Lc3B. Although treatment with amitriptyline or fluoxetine requires at least 12 days to achieve sphingomyelin accumulation and the subsequent biochemical and cellular changes, direct inhibition of sphingomyelin synthases with tricyclodecan-9-yl-xanthogenate (D609) results in rapid (within 3 days) accumulation of ceramide in the ER, activation of autophagy, and reversal of biochemical and behavioral signs of stress-induced MDD. Inhibition of Beclin blocks the antidepressive effects of amitriptyline and D609 and induces cellular and behavioral changes typical of MDD. These findings identify sphingolipid-controlled autophagy as an important target for antidepressive treatment methods and provide a rationale for the development of novel antidepressants that act within a few days. Y1 - 2018 U6 - https://doi.org/10.1038/s41380-018-0090-9 SN - 1359-4184 SN - 1476-5578 VL - 23 IS - 12 SP - 2324 EP - 2346 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Putra, Sulistyo Emantoko Dwi A1 - Neuber, Corinna A1 - Reichetzeder, Christoph A1 - Hocher, Berthold A1 - Kleuser, Burkhard T1 - Analysis of genomic DNA methylation levels in human placenta using liquid Chromatography-Electrospray ionization tandem mass spectrometry JF - Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry and pharmacology N2 - Background: DNA-methylation is a common epigenetic tool which plays a crucial role in gene regulation and is essential for cell differentiation and embryonic development. The placenta is an important organ where gene activity can be regulated by epigenetic DNA modifications, including DNA methylation. This is of interest as, the placenta is the interface between the fetus and its environment, the mother. Exposure to environmental toxins and nutrition during pregnancy may alter DNA methylation of the placenta and subsequently placental function and as a result the phenotype of the offspring. The aim of this study was to develop a reliable method to quantify DNA methylation in large clinical studies. This will be a tool to analyze the degree of DNA methylation in the human placenta in relationship to clinical readouts. Methods: Liquid chromatography-electrospray ionization/multi-stage mass spectrometry (LC-ESI/MS/MS) technique was used for the quantification of the 5dmC/dG ratio in placentas from 248 healthy pregnancies. We were able to demonstrate that this method is a reliable and stable way to determine global placental DNA methylation in large clinical trials. Results/Conclusion: The degree of placental DNA methylation seen in our pilot study varies substantially from 2% to 5%. The clinical implications of this variation need to be demonstrated in adequately powered large studies. KW - Pregnancy KW - Placenta KW - Methylation KW - Global KW - LC-MS/MS KW - Fetal programming KW - Clinical Y1 - 2014 U6 - https://doi.org/10.1159/000358666 SN - 1015-8987 SN - 1421-9778 VL - 33 IS - 4 SP - 945 EP - 952 PB - Karger CY - Basel ER - TY - JOUR A1 - Reichel, Martin A1 - Hoenig, Stefanie A1 - Liebisch, Gerhard A1 - Lüth, Anja A1 - Kleuser, Burkhard A1 - Gulbins, Erich A1 - Schmitz, Gerd A1 - Kornhuber, Johannes T1 - Alterations of plasma glycerophospholipid and sphingolipid species in male alcohol-dependent patients JF - Biochimica et biophysica acta : Molecular and cell biology of lipids N2 - Background: Alcohol abuse is a major risk factor for somatic and neuropsychiatric diseases. Despite their potential clinical importance, little is known about the alterations of plasma glycerophospholipid (GPL) and sphingolipid (SPL) species associated with alcohol abuse. Methods: Plasma GPL and SPL species were quantified using electrospray ionization tandem mass spectrometry in samples from 23 male alcohol-dependent patients before and after detoxification, as well as from 20 healthy male controls. Results: A comparison of alcohol-dependent patients with controls revealed higher phosphatidylcholine (PC; P-value = 0.008) and phosphatidylinositol (PI; P-value = 0.001) concentrations in patients before detoxification, and higher PI (P-value = 0.001) and phosphatidylethanolamine (PE)-based plasmalogen (PEP; P-value = 0.003) concentrations after detoxification. Lysophosphatidylcholines (LPC) were increased by acute intoxication (P-value = 0.002). Sphingomyelin (SM) concentration increased during detoxification (P-value = 0.011). The concentration of SM 23:0 was lower in patients (P-value = 2.79 x 10(-5)), and the concentrations of ceramide Cer d18:1/16:0 and Cer d18:1/18:0 were higher in patients (P-value = 2.45 x 10(-5) and 3.73 x 10(-5)). Activity of lysosomal acid sphingomyelinase (ASM) in patients correlated positively with the concentrations of eight LPC species, while activity of secreted ASM was inversely correlated with several PE, PI and PC species, and positively correlated with the molar ratio of PC to SM (Pearson's r = 0.432; P-value = 0.039). Conclusion: Plasma concentrations of numerous GPL and SPL species were altered in alcohol-dependent patients. These molecules might serve as potential biomarkers to improve the diagnosis of patients and to indicate health risks associated with alcohol abuse. Our study further indicates that there are strong interactions between plasma GPL concentrations and SPL metabolism. (C) 2015 Elsevier B.V. All rights reserved. KW - Acid sphingomyelinase KW - Alcohol dependence KW - Anxiety KW - Cardiovascular KW - Case-control study KW - Ceramide KW - Clinical KW - Depression KW - Diagnostic KW - Disease KW - Glycerophospholipids KW - Lysophosphatidylcholines KW - Mass spectrometry KW - Phosphatidylcholines KW - Phosphatidylinositols KW - Plasma KW - Plasmalogens KW - Sphingolipids KW - Sphingomyelin KW - Tandem mass spectrometry Y1 - 2015 U6 - https://doi.org/10.1016/j.bbalip.2015.08.005 SN - 1388-1981 SN - 0006-3002 VL - 1851 IS - 11 SP - 1501 EP - 1510 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Gulbins, Erich A1 - Palmada, Monica A1 - Reichel, Martin A1 - Lueth, Anja A1 - Boehmer, Christoph A1 - Amato, Davide A1 - Mueller, Christian P. A1 - Tischbirek, Carsten H. A1 - Groemer, Teja W. A1 - Tabatabai, Ghazaleh A1 - Becker, Katrin Anne A1 - Tripal, Philipp A1 - Staedtler, Sven A1 - Ackermann, Teresa F. A1 - van Brederode, Johannes A1 - Alzheimer, Christian A1 - Weller, Michael A1 - Lang, Undine E. A1 - Kleuser, Burkhard A1 - Grassme, Heike A1 - Kornhuber, Johannes T1 - Acid sphingomyelinase-ceramide system mediates effects of antidepressant drugs JF - Nature medicine N2 - Major depression is a highly prevalent severe mood disorder that is treated with antidepressants. The molecular targets of antidepressants require definition. We investigated the role of the acid sphingomyelinase (Asm)-ceramide system as a target for antidepressants. Therapeutic concentrations of the antidepressants amitriptyline and fluoxetine reduced Asm activity and ceramide concentrations in the hippocampus, increased neuronal proliferation, maturation and survival and improved behavior in mouse models of stress-induced depression. Genetic Asm deficiency abrogated these effects. Mice overexpressing Asm, heterozygous for acid ceramidase, treated with blockers of ceramide metabolism or directly injected with C16 ceramide in the hippocampus had higher ceramide concentrations and lower rates of neuronal proliferation, maturation and survival compared with controls and showed depression-like behavior even in the absence of stress. The decrease of ceramide abundance achieved by antidepressant-mediated inhibition of Asm normalized these effects. Lowering ceramide abundance may thus be a central goal for the future development of antidepressants. Y1 - 2013 U6 - https://doi.org/10.1038/nm.3214 SN - 1078-8956 VL - 19 IS - 7 SP - 934 EP - + PB - Nature Publ. Group CY - New York ER - TY - JOUR A1 - McVey, Mark J. A1 - Kim, Michael A1 - Tabuchi, Arata A1 - Srbely, Victoria A1 - Japtok, Lukasz A1 - Arenz, Christoph A1 - Rotstein, Ori A1 - Kleuser, Burkhard A1 - Semple, John W. A1 - Kuebler, Wolfgang M. T1 - Acid sphingomyelinase mediates murine acute lung injury following transfusion of aged platelets JF - American journal of physiology : Lung cellular and molecular physiology N2 - Pulmonary complications from stored blood products are the leading cause of mortality related to transfusion. Transfusion-related acute lung injury is mediated by antibodies or bioactive mediators, yet underlying mechanisms are incompletely understood. Sphingolipids such as ceramide regulate lung injury, and their composition changes as a function of time in stored blood. Here, we tested the hypothesis that aged platelets may induce lung injury via a sphingolipid-mediated mechanism. To assess this hypothesis, a two-hit mouse model was devised. Recipient mice were treated with 2 mg/kg intraperitoneal lipopolysaccharide (priming) 2 h before transfusion of 10 ml/kg stored (1-5 days) platelets treated with or without addition of acid sphingomyelinase inhibitor ARC39 or platelets from acid sphingomyelinase-deficient mice, which both reduce ceramide formation. Transfused mice were examined for signs of pulmonary neutrophil accumulation, endothelial barrier dysfunction, and histological evidence of lung injury. Sphingolipid profiles in stored platelets were analyzed by mass spectrophotometry. Transfusion of aged platelets into primed mice induced characteristic features of lung injury, which increased in severity as a function of storage time. Ceramide accumulated in platelets during storage, but this was attenuated by ARC39 or in acid sphingomyelinase-deficient platelets. Compared with wild-type platelets, transfusion of ARC39-treated or acid sphingomyelinase-deficient aged platelets alleviated lung injury. Aged platelets elicit lung injury in primed recipient mice, which can be alleviated by pharmacological inhibition or genetic deletion of acid sphingomyelinase. Interventions targeting sphingolipid formation represent a promising strategy to increase the safety and longevity of stored blood products. KW - transfusion-related acute lung injury KW - ceramide KW - acid sphingomyelinase KW - platelets KW - storage Y1 - 2017 U6 - https://doi.org/10.1152/ajplung.00317.2016 SN - 1040-0605 SN - 1522-1504 VL - 312 IS - 5 SP - 625 EP - 637 PB - American Physiological Society CY - Bethesda 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 - Beckmann, Nadine A1 - Becker, Katrin Anne A1 - Kadow, Stephanie A1 - Schumacher, Fabian A1 - Kramer, Melanie A1 - Kuehn, Claudine A1 - Schulz-Schaeffer, Walter J. A1 - Edwards, Michael J. A1 - Kleuser, Burkhard A1 - Gulbins, Erich A1 - Carpinteiro, Alexander T1 - Acid Sphingomyelinase Deficiency Ameliorates Farber Disease JF - International journal of molecular sciences N2 - Farber disease is a rare lysosomal storage disorder resulting from acid ceramidase deficiency and subsequent ceramide accumulation. No treatments for Farber disease are clinically available, and affected patients have a severely shortened lifespan. We have recently reported a novel acid ceramidase deficiency model that mirrors the human disease closely. Acid sphingomyelinase is the enzyme that generates ceramide upstream of acid ceramidase in the lysosomes. Using our acid ceramidase deficiency model, we tested if acid sphingomyelinase could be a potential novel therapeutic target for the treatment of Farber disease. A number of functional acid sphingomyelinase inhibitors are clinically available and have been used for decades to treat major depression. Using these as a therapeutic for Farber disease, thus, has the potential to improve central nervous symptoms of the disease as well, something all other treatment options for Farber disease can’t achieve so far. As a proof-of-concept study, we first cross-bred acid ceramidase deficient mice with acid sphingomyelinase deficient mice in order to prevent ceramide accumulation. Double-deficient mice had reduced ceramide accumulation, fewer disease manifestations, and prolonged survival. We next targeted acid sphingomyelinase pharmacologically, to test if these findings would translate to a setting with clinical applicability. Surprisingly, the treatment of acid ceramidase deficient mice with the acid sphingomyelinase inhibitor amitriptyline was toxic to acid ceramidase deficient mice and killed them within a few days of treatment. In conclusion, our study provides the first proof-of-concept that acid sphingomyelinase could be a potential new therapeutic target for Farber disease to reduce disease manifestations and prolong survival. However, we also identified previously unknown toxicity of the functional acid sphingomyelinase inhibitor amitriptyline in the context of Farber disease, strongly cautioning against the use of this substance class for Farber disease patients. KW - Farber disease KW - lysosomal storage disorders KW - acid ceramidase KW - acid sphingomyelinase KW - amitriptyline Y1 - 2019 U6 - https://doi.org/10.3390/ijms20246253 SN - 1422-0067 VL - 20 IS - 24 PB - MDPI CY - Basel ER - TY - JOUR A1 - Zeitler, Stefanie A1 - Ye, Lian A1 - Andreyeva, Aksana A1 - Schumacher, Fabian A1 - Monti, Juliana A1 - Nürnberg, Bernd A1 - Nowak, Gabriel A1 - Kleuser, Burkhard A1 - Reichel, Martin A1 - Fejtova, Anna A1 - Kornhuber, Johannes A1 - Rhein, Cosima A1 - Friedland, Kristina T1 - Acid sphingomyelinase - a regulator of canonical transient receptor potential channel 6 (TRPC6) activity JF - Journal of neurochemistry N2 - Recent investigations propose the acid sphingomyelinase (ASM)/ceramide system as a novel target for antidepressant action. ASM catalyzes the breakdown of the abundant membrane lipid sphingomyelin to the lipid messenger ceramide. This ASM‐induced lipid modification induces a local shift in membrane properties, which influences receptor clustering and downstream signaling. Canonical transient receptor potential channels 6 (TRPC6) are non‐selective cation channels located in the cell membrane that play an important role in dendritic growth, synaptic plasticity and cognition in the brain. They can be activated by hyperforin, an ingredient of the herbal remedy St. John’s wort for treatment of depression disorders. Because of their role in the context of major depression, we investigated the crosstalk between the ASM/ceramide system and TRPC6 ion channels in a pheochromocytoma cell line 12 neuronal cell model (PC12 rat pheochromocytoma cell line). Ca2+ imaging experiments indicated that hyperforin‐induced Ca2+ influx through TRPC6 channels is modulated by ASM activity. While antidepressants, known as functional inhibitors of ASM activity, reduced TRPC6‐mediated Ca2+ influx, extracellular application of bacterial sphingomyelinase rebalanced TRPC6 activity in a concentration‐related way. This effect was confirmed in whole‐cell patch clamp electrophysiology recordings. Lipidomic analyses revealed a decrease in very long chain ceramide/sphingomyelin molar ratio after ASM inhibition, which was connected with changes in the abundance of TRPC6 channels in flotillin‐1–positive lipid rafts as visualized by western blotting. Our data provide evidence that the ASM/ceramide system regulates TRPC6 channels likely by controlling their recruitment to specific lipid subdomains and thereby fine‐tuning their physical properties. KW - acid sphingomyelinase KW - antidepressants KW - ceramide KW - hyperforin KW - lipid rafts KW - TRPC6 Y1 - 2019 U6 - https://doi.org/10.1111/jnc.14823 SN - 0022-3042 SN - 1471-4159 VL - 150 IS - 6 SP - 678 EP - 690 PB - Wiley CY - Hoboken ER - TY - JOUR A1 - Lang, Judith A1 - Bohn, Patrick A1 - Bhat, Hilal A1 - Jastrow, Holger A1 - Walkenfort, Bernd A1 - Cansiz, Feyza A1 - Fink, Julian A1 - Bauer, Michael A1 - Schumacher, Fabian A1 - Kleuser, Burkhard A1 - Lang, Karl S. T1 - Acid ceramidase of macrophages traps herpes simplex virus in multivesicular bodies and protects from severe disease JF - Nature Communications N2 - Macrophages have important protective functions during infection with herpes simplex virus type 1 (HSV-1). However, molecular mechanisms that restrict viral propagation and protect from severe disease are unclear. Here we show that macrophages take up HSV-1 via endocytosis and transport the virions into multivesicular bodies (MVBs). In MVBs, acid ceramidase (aCDase) converts ceramide into sphingosine and increases the formation of sphingosine-rich intraluminal vesicles (ILVs). Once HSV-1 particles reach MVBs, sphingosine-rich ILVs bind to HSV-1 particles, which restricts fusion with the limiting endosomal membrane and prevents cellular infection. Lack of aCDase in macrophage cultures or in Asah1(-/-) mice results in replication of HSV-1 and Asah1(-/-) mice die soon after systemic or intravaginal inoculation. The treatment of macrophages with sphingosine enhancing compounds blocks HSV-1 propagation, suggesting a therapeutic potential of this pathway. In conclusion, aCDase loads ILVs with sphingosine, which prevents HSV-1 capsids from penetrating into the cytosol. KW - immunology KW - infection KW - membrane fusion KW - phagocytosis KW - sphingolipids Y1 - 2020 U6 - https://doi.org/10.1038/s41467-020-15072-8 SN - 2041-1723 VL - 11 IS - 1 SP - 1 EP - 15 PB - Nature Publishing Group UK CY - London ER - TY - JOUR A1 - Huston, Joseph P. A1 - Kornhuber, Johannes A1 - Muehle, Christiane A1 - Japtok, Lukasz A1 - Komorowski, Mara A1 - Mattern, Claudia A1 - Reichel, Martin A1 - Gulbins, Erich A1 - Kleuser, Burkhard A1 - Topic, Bianca A1 - Silva, Maria A. De Souza A1 - Mueller, Christian P. T1 - A sphingolipid mechanism for behavioral extinction JF - Journal of neurochemistry N2 - Reward-dependent instrumental behavior must continuously be re-adjusted according to environmental conditions. Failure to adapt to changes in reward contingencies may incur psychiatric disorders like anxiety and depression. When an expected reward is omitted, behavior undergoes extinction. While extinction involves active re-learning, it is also accompanied by emotional behaviors indicative of frustration, anxiety, and despair (extinction-induced depression). Here, we report evidence for a sphingolipid mechanism in the extinction of behavior. Rapid extinction, indicating efficient re-learning, coincided with a decrease in the activity of the enzyme acid sphingomyelinase (ASM), which catalyzes turnover of sphingomyelin to ceramide, in the dorsal hippocampus of rats. The stronger the decline in ASM activity, the more rapid was the extinction. Sphingolipid-focused lipidomic analysis showed that this results in a decline of local ceramide species in the dorsal hippocampus. Ceramides shape the fluidity of lipid rafts in synaptic membranes and by that way can control neural plasticity. We also found that aging modifies activity of enzymes and ceramide levels in selective brain regions. Aging also changed how the chronic treatment with corticosterone (stress) or intranasal dopamine modified regional enzyme activity and ceramide levels, coinciding with rate of extinction. These data provide first evidence for a functional ASM-ceramide pathway in the brain involved in the extinction of learned behavior. This finding extends the known cellular mechanisms underlying behavioral plasticity to a new class of membrane-located molecules, the sphingolipids, and their regulatory enzymes, and may offer new treatment targets for extinction- and learning-related psychopathological conditions. KW - acid sphingomyelinase KW - ceramide KW - extinction KW - hippocampus KW - operant behavior KW - sphingomyelin Y1 - 2016 U6 - https://doi.org/10.1111/jnc.13537 SN - 0022-3042 SN - 1471-4159 VL - 137 SP - 589 EP - 603 PB - Wiley-Blackwell CY - Hoboken ER -