Two mammalian sphingosine kinase (SphK) isoforms, SphK1 and SphK2, possess identical kinase domains but have distinct kinetic properties and subcellular localizations, suggesting each has one or more specific roles in sphingosine-1-phosphate (S1P) generation. Although both kinases use sphingosine as a substrate to generate S1P, the mechanisms controlling SphK activation and subsequent S1P generation during lung injury are not fully understood. In this study, we established a murine lung injury model to investigate LPS-induced lung injury in SphK1 knockout (SphK1(-/-)) and wild-type (WT) mice. We found that SphK1(-/-) mice were much more susceptible to LPS-induced lung injury compared with their WT counterparts, quantified by multiple parameters including cytokine induction. Intriguingly, overexpression of WT SphK1 delivered by adenoviral vector to the lungs protected SphK1(-/-) mice from lung injury and attenuated the severity of the response to LPS. However, adenoviral overexpression of a SphK1 kinase-dead mutant (SphKKD) in SphK1(-/-) mouse lungs further exacerbated the response to LPS as well as the extent of lung injury. WT SphK2 adenoviral overexpression also failed to provide protection and, in fact, augmented the degree of LPS-induced lung injury. This suggested that, in vascular injury, S1P generated by SphK2 activation plays a distinctly separate role compared with SphK1-dependent S1P generation and survival signaling. Microarray and real-time RT-PCR analysis of SphK1 and SphK2 expression levels during lung injury revealed that, in WT mice, LPS treatment caused significantly enhanced SphK1 expression ( approximately 5x) levels within 6 h, which declined back to baseline levels by 24 h posttreatment. In contrast, expression of SphK2 was gradually induced following LPS treatment and was elevated within 24 h. Collectively, our results for the first time demonstrate distinct functional roles of the two SphK isoforms in the regulation of LPS-induced lung injury.

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Mild traumatic brain injury afflicts over 2 million people annually and little can be done for the underlying injury. The Food and Drug Administration-approved drugs Minocycline plus N-acetylcysteine (MINO plus NAC) synergistically improved cognition and memory in a rat mild controlled cortical impact (mCCI) model of traumatic brain injury. The underlying cellular and molecular mechanisms of the drug combination are unknown. This study addressed the effect of the drug combination on white matter damage and neuroinflammation after mCCI. Brain tissue from mCCI rats given either sham-injury, saline, MINO alone, NAC alone, or MINO plus NAC was investigated via histology and qPCR at four time points (2, 4, 7, and 14 days post-injury) for markers of white matter damage and neuroinflammation. MINO plus NAC synergistically protected resident oligodendrocytes and decreased the number of oligodendrocyte precursor cells. Activation of microglia/macrophages (MP/MG) was synergistically increased in white matter two days post-injury after MINO plus NAC treatment. Patterns of M1 and M2 MP/MG were also altered after treatment. The modulation of neuroinflammation is a potential mechanism to promote remyelination and improve cognition and memory. These data also provide new and important insights into how drug treatments can induce repair after traumatic brain injury.

synthase (SMS) catalyzes the synthesis of sphingomyelin (SM) and is required for maintenance of plasma membrane microdomain fluidity. Of the two isoforms of mammalian SMS, SMS1 is mostly present in the trans-Golgi apparatus, whereas SMS2 is predominantly found at the plasma membrane. SMS2 has a role in receptor mediated response to inflammation in macrophages, however, the role of SMS2 in vascular permeability, pulmonary edema, and lung injury have not been investigated. To define the role of SMS activation in lung injury, we utilized a lipopolysaccharide (LPS)-induced lung edema model. SMS activity was measured and correlated with the severity of lung injury. Within 4 h of LPS treatment, SMS activity was increased significantly and remained upregulated up to 24 h. Comparison of LPS-induced lung injury in SMS2 knockout (SMS2 Ϫ/Ϫ ) and wildtype littermate control mice showed that inflammation, cytokine induction, and lung injury were significantly inhibited in SMS2 Ϫ/Ϫ mice. Our results suggest that a deficiency of SMS2 can diminish the extent of pulmonary edema and lung injury. Furthermore, we show that depletion of SMS2 was sufficient to decrease MAP kinase-JNK activation, severity of LPS-induced pulmonary neutrophil influx, and inflammation, suggesting a novel role of SMS2 activation in lung injury.ACUTE LUNG INJURY (ALI) is a condition of acute and persistent lung inflammation characterized by increased vascular permeability (25,40). This vascular leak causes pulmonary edema and hypoxia (5,17). Every year in the United States, ϳ74,500 persons die from ALI due to various causes, a figure comparable to the number of adult deaths attributed to breast cancer or human immunodeficiency virus disease in 1999 (26a). The mortality rate for the most severe form of ALI, acute respiratory distress syndrome, has been ϳ30% in the most recent large clinical trials (e.g., ARDSNet studies), but milder forms of ALI are likely to have significantly less mortality.It is well known that the vascular endothelium plays an important role in maintaining the integrity of the pulmonary vascular and parenchymal interstitium barrier. Any breakdown in the integrity of this barrier leads to vascular leak (5,22,27,40). Sphingosine-1-phosphate (S1P) increases endothelial resistance via lipid rafts by increasing the cortical actin content and decreasing actin stress fiber formation (6, 32). Studies have also shown that exogenous administration of S1P significantly decreased pulmonary and renal vascular leakage and inflammation in murine models of LPS-induced lung injury (24). Moreover, pretreatment with S1P dramatically reduced edema formation in a canine model of lung injury (19). In both the murine and canine models, S1P administration also drastically reduced neutrophil accumulation in the lungs compared with vehicle-treated animals (19,24,29). Furthermore, in sphingosine kinase knockout mice (SphK Ϫ/Ϫ ), endotoxin-induced injury was markedly increased, suggesting a functional role for SphK1 in the regulation of lung injury (4, 39).Althoug...