Katrin Anne Becker

Ceramide-enriched membrane domains--structure and function.

Membrane lipids seem to be organized and not randomly distributed in the cell membrane. In particular, sphingolipids seem to interact with cholesterol in the outer leaflet of the cell membrane resulting in the formation of distinct membrane domains, i.e. rafts. The generation of ceramide within rafts alters their biophysical properties and results in the formation of large ceramide-enriched membrane platforms. These platforms serve to cluster receptor molecules and to organize intracellular signalling molecules to facilitate signal transduction via a receptor upon stimulation. Thus, ceramide-enriched membrane domains amplify not only receptor-, but also stress-mediated signalling events. Although many receptors cluster, the molecular mechanisms mediating this important and general event in signal transduction need to be identified.

Acid sphingomyelinase–ceramide system mediates effects of antidepressant drugs

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.

Acid Sphingomyelinase Inhibitors Normalize Pulmonary Ceramide and Inflammation in Cystic Fibrosis

Employing genetic mouse models we have recently shown that ceramide accumulation is critically involved in the pathogenesis of cystic fibrosis (CF) lung disease. Genetic or systemic inhibition of the acid sphingomyelinase (Asm) is not feasible for treatment of patients or might cause adverse effects. Thus, a manipulation of ceramide specifically in lungs of CF mice must be developed. We tested whether inhalation of different acid sphingomyelinase inhibitors does reduce Asm activity and ceramide accumulation in lungs of CF mice. The efficacy and specificity of the drugs was determined. Ceramide was determined by mass spectrometry, DAG-kinase assays, and fluorescence microscopy. We determined pulmonary and systemic Asm activity, neutral sphingomyelinase (Nsm), ceramide, cytokines, and infection susceptibility. Mass spectroscopy, DAG-kinase assays, and semiquantitative immune fluorescence microscopy revealed that a standard diet did not influence ceramide in bronchial respiratory epithelial cells, while a diet with Peptamen severely affected the concentration of sphingolipids in CF lungs. Inhalation of the Asm inhibitors amitriptyline, trimipramine, desipramine, chlorprothixene, fluoxetine, amlodipine, or sertraline restored normal ceramide concentrations in murine bronchial epithelial cells, reduced inflammation in the lung of CF mice and prevented infection with Pseudomonas aeruginosa. All drugs showed very similar efficacy. Inhalation of the drugs was without systemic effects and did not inhibit Nsm. These findings employing several structurally different Asm inhibitors identify Asm as primary target in the lung to reduce ceramide concentrations. Inhaling an Asm inhibitor may be a beneficial treatment for CF, with minimal adverse systemic effects.

Ceramide in Redox Signaling and Cardiovascular Diseases

Lipid rafts are distinct cell membrane microdomains that consist of cholesterol, sphingolipids, and some associated proteins. Accumulating evidence suggests that activation of sphingomyelinase and generation of ceramide mediates clustering of lipid rafts to form large ceramide-enriched platforms, in which transmembrane signals are transmitted or amplified. Ceramide and reactive oxygen species (ROS) are involved in the modulation of the cell membrane and intracellular ion channels, cell proliferation and apoptotic cell death, neutrophil adhesion to the vessel wall, and vascular tone and in the development of cardiovascular diseases to name some important examples. Ceramide triggers the generation of ROS and increases oxidative stress in many mammalian cells and animal models. Moreover, inhibition of ROS generating enzymes or treatment of antioxidants impairs sphingomyelinase activation and ceramide production. Thus, a new concept has been proposed that ceramide-enriched raft platforms are important redox signaling platforms that amplify activation of ROS generating enzymes (e.g. NADPH oxidase family enzymes) and sphingomyelinases. The general function of ceramide to form redox signaling platforms amplifying oxdative stress might be critically involved in the dysfunction of vascular cells induced by death receptor ligands and stress stimuli contributing to the development of cardiovascular diseases.

Engineered liposomes sequester bacterial exotoxins and protect from severe invasive infections in mice

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.

Targeting the ceramide system in cancer

Sphingolipids, in particular ceramide, have been described as important components of cellular signalling pathways. Ceramide can be produced via multiple mechanisms including through the hydrolysis of sphingomyelin by acid and neutral sphingomyelinase or by a de novo synthesis pathway. Recent studies have identified sphingomyelinases and ceramide synthases as important targets for γ-irradiation and chemotherapeutic drugs. Likewise, common cancer treatment modalities, such as γ-irradiation and many chemotherapeutic agents, induce cell death via the generation of ceramide. This suggests that the manipulation of ceramide production and metabolism could offer promising means for the enhancement of anti-tumor therapies. The focus of this mini-review will be to discuss contemporary evidence suggesting that ceramide forming pathways and ceramide itself are important targets for the treatment of tumors and the development of novel tumor treatment strategies.

Sphingoid long chain bases prevent lung infection by Pseudomonas aeruginosa

Cystic fibrosis patients and patients with chronic obstructive pulmonary disease, trauma, burn wound, or patients requiring ventilation are susceptible to severe pulmonary infection by Pseudomonas aeruginosa. Physiological innate defense mechanisms against this pathogen, and their alterations in lung diseases, are for the most part unknown. We now demonstrate a role for the sphingoid long chain base, sphingosine, in determining susceptibility to lung infection by P. aeruginosa. Tracheal and bronchial sphingosine levels were significantly reduced in tissues from cystic fibrosis patients and from cystic fibrosis mouse models due to reduced activity of acid ceramidase, which generates sphingosine from ceramide. Inhalation of mice with sphingosine, with a sphingosine analog, FTY720, or with acid ceramidase rescued susceptible mice from infection. Our data suggest that luminal sphingosine in tracheal and bronchial epithelial cells prevents pulmonary P. aeruginosa infection in normal individuals, paving the way for novel therapeutic paradigms based on inhalation of acid ceramidase or of sphingoid long chain bases in lung infection.

Accumulation of ceramide in the trachea and intestine of cystic fibrosis mice causes inflammation and cell death.

Cystic fibrosis is a hereditary metabolic disorder caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene and characterized by severe intestinal and pulmonary symptoms, in particular intestinal obstruction, pancreatic insufficiency, chronic pulmonary inflammation, and microbial lung infections. Recent studies have demonstrated an accumulation of ceramide in the lungs of cystic fibrosis patients and in several mouse models. These findings showed that pulmonary ceramide concentrations play an important role in pulmonary inflammation and infection. In this study we investigated whether ceramide concentrations are also altered in the trachea and the intestine of cystic fibrosis mice and whether an accumulation of ceramide in these organs has functional consequences that are typical of cystic fibrosis. Our findings demonstrate a marked accumulation of ceramide in tracheal and intestinal epithelial cells of cystic fibrosis mice. When acid sphingomyelinase activity is inhibited by treating cystic fibrosis mice with amitriptyline or by genetic heterozygosity of acid sphingomyelinase in cystic fibrosis mice, ceramide concentrations in the trachea and the intestine are normalized. Moreover, increased rates of cell death and increased cytokine concentrations in the trachea, the intestine, or both were normalized by the inhibition of acid sphingomyelinase activity and the concomitant normalization of ceramide concentrations. These findings suggest that ceramide plays a crucial role in inflammation and increased rates of cell death in several organs of cystic fibrosis mice.

The ceramide system as a novel antidepressant target

Major depression is a systems disorder which impairs not only central nervous system aspects of mood and behavior but also peripheral organ systems. Current views on the pathogenesis and treatment of depression are predominantly based on proteins and transmitters and thus are difficult to reconcile central with peripheral pathomechanisms. Recent research showed that there is also a lipid-based pathway involved in the pathology of depression, which is activated by psychosocial stress, oxidative stress, or inflammation. Inducible dysfunction of the ceramide pathway, which is abundant in the brain as well as in peripheral organs, may account for mood disorder, behavioral symptoms, and further promote inflammation and oxidative stress in peripheral systems. As such, the lipid ceramide pathway may provide the missing link between brain dysfunction and somatic symptoms of depression. Pharmacological interventions that reduce ceramide abundance also show antidepressant action and may promise a better treatment of major depression.

Clofazimine, Psora-4 and PAP-1, inhibitors of the potassium channel Kv1.3, as a new and selective therapeutic strategy in chronic lymphocytic leukemia

Clofazimine, Psora-4 and PAP-1, inhibitors of the potassium channel Kv1.3, as a new and selective therapeutic strategy in chronic lymphocytic leukemia