Acid Sphingomyelinase Promotes Endothelial Stress Response in Systemic Inflammation and Sepsis.

The pathophysiology of sepsis involves activation of acid sphingomyelinase (SMPD1) with subsequent generation of the bioactive mediator ceramide. We herein evaluated the hypothesis that the enzyme exerts biological effects in endothelial stress response. Plasma-secreted sphingomyelinase activity, ceramide generation and lipid raft formation were measured in human microcirculatory endothelial cells (HMEC-1) stimulated with serum obtained from sepsis patients. Clustering of receptors relevant for signal transduction was studied by immuno staining. The role of SMPD1 for macrodomain formation was tested by pharmacological inhibition. To confirm the involvement of the stress enzyme, direct inhibitors (amino bisphosphonates) and specific downregulation of the gene was tested with respect to ADAMTS13 expression and cytotoxicity. Plasma activity and amount of SMPD1 were increased in septic patients dependent on clinical severity. Increased breakdown of sphingomyelin to ceramide in HMECs was observed following stimulation with serum from sepsis patients in vitro. Hydrolysis of sphingomyelin, clustering of receptor complexes, such as the CD95L/Fas-receptor, as well as formation of ceramide enriched macrodomains was abrogated using functional inhibitors (desipramine and NB6). Strikingly, the stimulation of HMECs with serum obtained from sepsis patients or mixture of proinflammatory cytokines resulted in cytotoxicity and ADAMTS13 downregulation which was abrogated using desipramine, amino bisphosphonates and genetic inhibitors. SMPD1 is involved in the dysregulation of ceramide metabolism in endothelial cells leading to macrodomain formation, cytotoxicity and downregulation of ADAMTS13 expression. Functional inhibitors, such as desipramine, are capable to improve endothelial stress response during sepsis and might be considered as a pharmacological treatment strategy to favor the outcome.

Inhibition of sphingomyelin hydrolysis: targeting the lipid mediator ceramide as a key regulator of cellular fate.

Ceramide, an intracellular lipid mediator, is generated by transient hydrolysis of sphingomyelin in response to agonists inducing inflammation and apoptosis, ionizing radiation, chemotherapeutics or ischaemia/reperfusion. An elevated intracellular ceramide production is predominantly induced by an elevated hydrolytic activity of sphingomyelinases or by the activity of enzymes controlling de novo synthesis, such as ceramide synthase. Ceramide is implicated in various cellular responses, acting as an autonomous intracellular effector in cell cycle regulation, differentiation, senescence, and apoptosis. Furthermore, by changing membrane properties it contributes to the assembly and interactions of various signal transduction molecules. The lipid mediator is subsequently metabolised by ceramidase and sphingosine kinase, and other key players, which determine the dynamic balance between the intracellular levels of ceramide and its breakdown products (the ceramide/S1P rheostat). Together with sphingomyelinases, they are crucial for cell signalling, cellular survival or initiation of apoptosis. Sphingomyelinases are regarded as key enzymes in the regulated activation of the sphingomyelin cycle. Up to now, five isoforms of sphingomyelinases are known, differentiated by agonists, intracellular localization, pH optimum and essential co-factors. Herein, we focus on the biochemical background and the action of pharmacologically interesting compounds capable of interfering with sphingomyelinases and outline their potential implications for medicinal chemistry.