Attenuation of shock-induced acute lung injury by sphingosine kinase inhibition

Background: Prolonged elevations of cytosolic calcium concentrations ([Ca²⁺J_i) are required for optimal neutrophil (PMN) activation responses to G-Protein coupled chemoattractants. We recently showed that the coupling of endosomal Ca²⁺ store depletion to more prolonged entry of external Ca²⁺ depends on cellular conversion of sphingosine to sphingosine 1-phosphate (S1P) by sphingosine kinase (SK). We therefore hypothesized that inhibition of SK might inhibit PMN activation and thus ameliorate lung injury after trauma and hemorrhagic shock (T/HS). Methods: Chemotaxis (CTX) of human PMN was studied using modified Boyden chambers in the presence or absence of the selective SK inhibitor, SKI-2. After determining the concentration of SKI-2 that inhibited human PMN CTX by 50% (IC₅₀) we subjected rats to T/HS (laparotomy, hemorrhage to 30-40 mm Hg × 90 minutes, 3 hours resuscitation). We then studied rat PMN CD11b expression using flow cytometry and lung injury using the Evans Blue dye technique in the presence of IC₅₀ doses of SKI-2 or vehicle given in pretreatment at laparotomy. Results: Human PMN CTX was suppressed slightly more than 50% by 40 μmol/L SKI-2 (233 ±20 vs103 ± 12 × 10³ cells/well, p < 0.001). Rat PMN expression of CD11b after T/HS was decreased from 352 ± 30 to 232 ± 7 MFU (p < 0.001) in the presence 30 μmol/L SKI-2. Lung permeability to Evans Blue was decreased from 9.5 ± 2 to 4.1 ± 0.7% (p = 0.036.). SKI-2 did not cause hemodynamic instability or alter resuscitation requirements. Conclusions: Modulation of PMN Ca²⁺ entry via SK inhibition inhibits PMN CTX in vitro, and inhibits CD11b expression in vivo without major effects on hemodynamics. These cellular changes were associated with amelioration of lung injury in vivo in a rat model of T/HS. These findings suggest that SK inhibition allows modulation of inflammation via control of [Ca²⁺]; without the cardiovascular compromise expected with Ca²⁺ channel blockade. SK inhibition therefore appears to be an important novel candidate therapy for inflammatory organ injury after shock.