Mechanism of superoxide anion production by hepatic sinusoidal endothelial cells and Kupffer cells during short-term ethanol perfusion in the rat

Liver. 2002 Aug;22(4):321-9. doi: 10.1034/j.1600-0676.2002.01493.x.

Abstract

Background/aims: The aim of this study was to clarify the candidate cells for and the mechanism of superoxide anion (O2*-) release into the hepatic sinusoids during short-term exposure to ethanol.

Methods: The rat liver was perfused continuously with ethanol (a substrate for alcohol dehydrogenase) or tert-buthanol (not a substrate for alcohol dehydrogenase) for 20 min at a final concentration of 40 mM. In order to detect O2*- production, MCLA (2-methyl-6-[p-methoxyphenyl]-3,7-dihydroimidazo[1,2-a]pyrazin-3-one), a Cypridina luciferin analogue, was simultaneously infused and MCLA-enhanced chemiluminescence was measured. The effects of gadolinium chloride (GdCL3) (a suppressor of Kupffer cells (KCs)), staurosporine (ST) (an inhibitor of serine-threonine kinases, including protein kinase C), diphenyleneiodonium chloride (DPI) (an inhibitor of NADPH oxidase), ibuprofen (IB) (an inhibitor of cyclooxygenase) and 4-methylpyrazole (4MP) (an inhibitor of ethanol metabolism) on the ethanol-induced chemiluminescence were also evaluated. Sites where O2*- could be released were determined by histochemical detection of nitro blue tetrazolium reduction.

Results: Both ethanol and tert-buthanol rapidly caused O2*- release. GdCL3 suppressed the ethanol-induced O2*- release by 61%. Staurosporine and DPI, but neither IB nor 4-MP, also significantly inhibited the ethanol-induced O2*- release. In the histochemical examination, ethanol-stimulated liver showed blue formazan precipitate on both sinusoidal endothelial cells (SECs) and Kupffer cells (KCs), whereas the GdCl3-pretreated liver had the precipitate only on SECs.

Conclusions: This study shows that ethanol itself stimulates both SECs and KCs to release O2*- via activation of NADPH oxidase probably involving protein kinase C (PKC).

MeSH terms

  • Animals
  • Endothelium, Vascular / drug effects
  • Endothelium, Vascular / metabolism*
  • Enzyme Inhibitors / pharmacology
  • Ethanol / administration & dosage
  • Ethanol / toxicity*
  • Fomepizole
  • Gadolinium / pharmacology
  • Ibuprofen / pharmacology
  • Imidazoles / metabolism
  • Kupffer Cells / drug effects
  • Kupffer Cells / metabolism*
  • Luminescent Measurements
  • Male
  • Nitroblue Tetrazolium / metabolism
  • Onium Compounds / pharmacology
  • Perfusion
  • Pyrazines / metabolism
  • Pyrazoles / pharmacology
  • Rats
  • Rats, Wistar
  • Staurosporine / pharmacology
  • Superoxides / metabolism*
  • tert-Butyl Alcohol / administration & dosage
  • tert-Butyl Alcohol / pharmacology

Substances

  • Enzyme Inhibitors
  • Imidazoles
  • Onium Compounds
  • Pyrazines
  • Pyrazoles
  • Superoxides
  • 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo(1,2-alpha)pyrazin-3-one
  • Nitroblue Tetrazolium
  • Ethanol
  • diphenyleneiodonium
  • Fomepizole
  • Gadolinium
  • Staurosporine
  • tert-Butyl Alcohol
  • gadolinium chloride
  • Ibuprofen