Cytoprotection by iloprost against paracetamol-induced toxicity in hamster isolated hepatocytes.
1 The ability of iloprost (ZK36374) to protect hamster isolated hepatocytes from the toxic effects of paracetamol and its reactive metabolite N-acetyl-p-benzoquinoneimine (NABQI) was investigated. The cytoprotection provided by iloprost was compared with that of N-acetyl-L-cysteine. 2 Treatment of hepatocytes with either NABQI (0.4 mM) or paracetamol (2 mM) alone resulted in a considerable loss of cell viability, as assessed by trypan blue exclusion or leakage of lactate dehydrogenase, accompanied by an increase in the percentage of viable cells that were blebbed. N-acetyl-L-cysteine (1.25 mM) pretreatment diminished the loss of cell viability and the percentage of blebbed cells resulting from exposure to NABQI or paracetamol, whereas iloprost (10(-16) M to 10(-10) M) pretreatment reduced only the loss of cell viability, not the percentage of viable cells exhibiting blebbing. Pretreatment with N-acetyl-L-cysteine significantly attenuated the depletion by paracetamol of glutathione and decreased the covalent binding of [14C]-paracetamol to cellular proteins, whereas iloprost was without any such effects. 3 The effects of iloprost and N-acetyl-L-cysteine were also investigated by use of a model of paracetamol toxicity in which it is possible to study the biochemical events leading to cell injury separate from the generation of toxic metabolites. Hamster hepatocytes were incubated with paracetamol (4 mM) for 90 min at 37 degrees C during which metabolism of paracetamol occurs with minimal loss of cell viability. Following washing of cells, to remove paracetamol and its metabolites, there was a progressive loss of viability and increase in the percentage of cells exhibiting blebbing when incubated in buffer alone. Addition of either N-acetyl-L-cysteine (1.25 mM) or iloprost (10 14M to 10 -M), following washing, significantly reduced the expected loss of cell viability. Iloprost at concentrations outside this range was without effect.4. Paracetamol toxicity to isolated hepatocytes could be prevented or delayed by treatment with either N-acetyl-L-cysteine or iloprost, but whereas the former prevented or even reversed plasma membrane blebbing with a resultant reduction in the percentage of viable cells that were blebbed, the prostanoid appeared only to delay the progression from plasma membrane blebbing to loss of viability. Hence, the percentage of viable cells that were ultimately blebbed following exposure to paracetamol was not significantly reduced by addition of iloprost.5. Aspirin or ibuprofen exacerbated the loss of viability induced by prior incubation with paracetamol. Thus, there may be a role for endogenous prostaglandins in protecting hepatocytes from paracetamol toxicity.6. Iloprost is cytoprotective without any effect upon toxin metabolism or detoxication. The mechanism of action of iloprost probably does not involve induction of prostaglandin synthesis or activation of the previously-characterized prostacyclin receptor.
Department of Clinical Pharmacology, Royal Postgraduate Medical School, Hammersmith Hospital, London.
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P
Nasseri-Sina
D J
DJ
Fawthrop
J
J
Wilson
A R
AR
Boobis
D S
DS
Davies
D000082:Acetaminophen / Q000037:antagonists & inhibitors* / Q000633:toxicity; D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D016227:Benzoquinones / Q000633:toxicity; D002470:Cell Survival / Q000187:drug effects*; D006224:Cricetinae; D005978:Glutathione / Q000378:metabolism; D016285:Iloprost / Q000494:pharmacology*; D007097:Imines / Q000633:toxicity; D066298:In Vitro Techniques; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000187:drug effects*; D008297:Male; D008647:Mesocricetus; D011448:Prostaglandin Antagonists / Q000494:pharmacology; D011453:Prostaglandins / Q000096:biosynthesis
D016227:Benzoquinones; D007097:Imines; D011448:Prostaglandin Antagonists; D011453:Prostaglandins; D000082:Acetaminophen; D007770:L-Lactate Dehydrogenase; C028473:N-acetyl-4-benzoquinoneimine; D005978:Glutathione; D016285:Iloprost; D000111:Acetylcysteine