gw-4869 and Reperfusion-Injury

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.

Topics: Alanine Transaminase; Aniline Compounds; Animals; Benzylidene Compounds; Ceramides; Dimethyl Sulfoxide; Immunohistochemistry; Liver; Male; Nitric Oxide; Oxidative Stress; Rats, Wistar; Reperfusion Injury; Sphingomyelin Phosphodiesterase; Sphingomyelins; Tandem Mass Spectrometry

This study aimed to determine the role of selective neutral sphingomyelinase (N-SMase) inhibition on arachidonic acid (AA) mediated inflammation following liver ischemia-reperfusion (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. Levels of AA in liver tissue were determined by multiple reaction monitoring (MRM) using ultra fast-liquid chromatography (UFLC) coupled with tandem mass spectrometry (MS/MS). Phospholipase A₂ (PLA₂), cyclooxygenase (COX) and prostaglandin E₂ (PGE₂) were measured in liver tissue. Arachidonic acid levels, activity of PLA₂, COX and PGE₂ levels were significantly increased in postischemic liver tissue compared to nonischemic controls. N-SMase inhibition significantly decreased COX activity and PGE₂ levels in postischemic liver. Future studies evaluating agents blocking N-SMase activity can facilitate the development of treatment strategies to alleviate inflammation in liver I/R injury.

Topics: Aniline Compounds; Animals; Arachidonic Acid; Benzylidene Compounds; Cyclooxygenase 2; Dinoprostone; Inflammation; Liver; Male; Phospholipases A2; Rats; Rats, Wistar; Reperfusion Injury; Sphingomyelin Phosphodiesterase