Inhibition of neutral sphingomyelinase decreases elevated levels of nitrative and oxidative stress markers in liver ischemia–reperfusion injury

Abstract

Oxidative stress and excessive nitric oxide production via induction of inducible nitric oxide synthase (NOS)-2 have been shown in the pathogenesis of liver ischemia-reperfusion (IR) injury. Neutral sphingomyelinase (N-SMase)/ceramide pathway can regulate NOS2 expression therefore this study determined the role of selective N-SMase inhibition on nitrative and oxidative stress markers following liver IR injury. Selective N-SMase inhibitor was administered via intraperitoneal injections. Liver IR injury was created by clamping blood vessels supplying the median and left lateral hepatic lobes for 60 min, followed by 60 min reperfusion. Nitrative and oxidative stress markers were determined by evaluating NOS2 expression, protein nitration, nitrite/nitrate levels, 4-hydroxynonenal (HNE) formation, protein carbonyl levels and xanthine oxidase/xanthine dehydrogenase (XO/XDH) activity. Levels of sphingmyelin and ceramide in liver tissue were determined by an optimized multiple reaction monitoring method using ultra-fast liquid chromatography coupled with tandem mass spectrometry (MS/MS). Spingomyelin levels were significantly increased in all IR groups compared to controls. Treatment with a specific N-SMase inhibitor significantly decreased all measured ceramides in IR injury. NOS2 expression, nitrite/nitrate levels and protein nitration were significantly greater in IR injury and decreased with N-SMase inhibition. Treatment with a selective N-SMase inhibitor significantly decreased HNE formation, protein carbonyl levels and the hepatic conversion of XO. Data confirm the role of nitrative and oxidative injury in IR and highlight the protective effect of selective N-SMase inhibition. Future studies evaluating agents blocking N-SMase activity can facilitate the development of treatment strategies to alleviate oxidative injury in liver I/R injury.

European contribution to the study of ROS: A summary of the findings and prospects for the future from the COST action BM1203 (EU-ROS)

“…Other studies showed that exogenous ceramide induces NOS2 expression in rat primary astrocytes [113] and reported positive modulation of NOS2 gene expression by SMase and/or ceramide in vascular smooth muscle cells and microglia [114] , [115] . Neutral SMase inhibition decreased both NOS2 and nitrotyrosine levels in an experimental model of glaucoma [116] and liver IR injury [109] . In summary, studies reveal that cellular stress responses significantly increase N-SMase activity and sphingomyelin/ceramide levels, leading to increased nitrative and oxidative modifications.…”

“…Indeed, activation of neutral sphingomyelinase (N-SMase) is reported to be involved in various disease pathologies connected to “oxidative stress”. Studies have shown that ischemia reperfusion (IR) injury leads to activation of N-SMase in rat cardiac myocytes [108] and rat liver [109] . Repletion of glutathione (GSH) via N -acetylcysteine (NAC) treatment in post-myocardial infarction rat hearts resulted in inhibition of N-SMase and decreased oxidative stress resulting in improved left ventricular function [110] .…”

Decreased Serum Levels of Sphingomyelins and Ceramides in Sickle Cell Disease Patients

“…An optimized multiple reaction monitoring (MRM) method was developed using UFLC coupled with tandem mass spectrometry (MS/MS), as previously described (Unal et al, ). Standards for N ‐palmitoyl‐ d ‐erythro‐sphingosylphosphorylcholine (C16 CerPCho), N ‐stearoyl‐ d ‐erythro‐sphingosylphosphorylcholine (C18 CerPCho), N ‐lignoceroyl‐ d ‐erythro sphingosylphosphorylcholine (C24 CerPCho), N ‐palmitoyl‐ d ‐erythro‐sphingosine (C16 CER), N ‐stearoyl‐ d ‐erythro‐sphingosine (C18 CER), N ‐arachidoyl‐ d ‐erythro‐sphingosine (C20 CER), N ‐behenoyl‐ d ‐erythro‐sphingosine (C22 CER), and N ‐lignoceroyl‐ d ‐erythro‐sphingosine (C24 CER) were purchased from Avanti Polar Lipids (Alabaster, AL, USA).…”

Epigallocatechin-3-gallate protects against hepatic ischaemia–reperfusion injury by reducing oxidative stress and apoptotic cell death

“…7 Damage to DNA bases is usually caused by reactive oxygen species (ROS), which include hydroxyl, the superoxide anion, and hydrogen peroxide. 8 , 9 All ROS respond to varying degrees at the initiation of reperfusion as reduced electron carrier molecules of hypoxic cells donate electrons to oxygen during reperfusion. 10 Therefore, effective therapeutic strategies for hepatic ischaemia–reperfusion injury are urgently needed.…”