RESUMO
INTRODUCTION: Ceramide is the central substrate of sphingolipid metabolism and plays a key role in cellular signal transduction pathways, regulating apoptosis, differentiation, and chemotaxis. Alterations in airway ceramide levels are observed in multiple pulmonary diseases and recent human genetic association studies have linked dysregulation of sphingolipid regulatory genes with asthma pathogenesis. METHODS: Utilizing myriocin, a potent inhibitor of sphingolipid synthesis, we evaluated the immune regulatory role of de novo ceramide generation in vitro and in vivo. Intratracheal myriocin was administered alone or during house dust mite sensitization (HDM) of BALB/C mice and airway hyper-responsiveness (AHR) was evaluated by invasive plethysmography followed by bronchial lavage (BAL) cytology and cytokine quantification. RESULTS: Myriocin inhibits and HDM exposure activates de novo ceramide synthesis in bone marrow-derived dendritic cells. Mice receiving intratracheal myriocin developed a mild airway neutrophilic infiltrate without inducing a significant increase in AHR. CXCL1 was elevated in the BAL fluid of myriocin-treated mice while the neutrophilic chemotactic factors anaphylatoxin C5a, leukotriene B4, and IL-17 were unaffected. HDM treatment combined with myriocin led to a dramatic enhancement of AHR (63% increase over HDM alone, p < 0.001) and increased granulocyte pulmonary infiltrates versus HDM or myriocin alone. Elevated Th2 T cell counts and Th2 cytokines/chemokines (IL5, IL13, CCL17) were observed in mice treated with combined HDM/myriocin compared to HDM alone. Myriocin-treated pulmonary CD11c+ cells stimulated with HDM secreted significantly more CXCL1 than cells stimulated with HDM alone while HDM stimulated airway epithelial cells showed no change in CXCL1 secretion following myriocin treatment. CONCLUSIONS: Intratracheal myriocin, likely acting via ceramide synthesis inhibition, enhances allergen-induced airway inflammation, granulocyte and Th2 lymphocyte recruitment, and allergen-induced AHR. Sphingolipid pathways may represent novel targets for possible future anti-inflammatory asthma medications.
