Exogenous phosphatidic acid reduces acetaminophen-induced liver injury in mice by activating hepatic interleukin-6 signaling through inter-organ crosstalk

We previously demonstrated that endogenous phosphatidic acid(PA)promotes liver regeneration after acetaminophen(APAP)hepatotoxicity.Here,we hypothesized that exogenous PA is also beneficial.To test that,we treated mice with a toxic APAP dose at 0 h,followed by PA or vehicle(Veh)posttreatment.We then collected blood and liver at 6,24,and 52 h.Post-treatment with PA 2 h after APAP protected against liver injury at 6 h,and the combination of PA and N-acetyl-L-cysteine(NAC)reduced injury more than NAC alone.Interestingly,PA did not affect canonical mechanisms of APAP toxicity.Instead,transcriptomics revealed that PA activated interleukin-6(IL-6)signaling in the liver.Consistent with that,serum IL-6 and hepatic signal transducer and activator of transcription 3(Stat3)phosphorylation increased in PA-treated mice.Furthermore,PA failed to protect against APAP in IL-6-deficient animals.Interestingly,IL-6 expression increased 18-fold in adipose tissue after PA,indicating that adipose is a source of PA-induced circulating IL-6.Surprisingly,however,exogenous PA did not alter regeneration,despite the importance of endogenous PA in liver repair,possibly due to its short half-life.These data demonstrate that exogenous PA is also beneficial in APAP toxicity and reinforce the protective effects of IL-6 in this model.

Pre-treatment twice with liposomal clodronate protects against acetaminophen hepatotoxicity through a pre-conditioning effect

Background and aim:Acetaminophen(APAP)overdose is a major cause of acute liver injury,but the role of macrophages in the propagation of the hepatotoxicity is controversial.Early research revealed that macrophage inhibitors protect against APAP injury.However,later work demonstrated that macrophage ablation by acute pre-treatment with liposomal clodronate(LC)exacerbates the toxicity.To our surprise,during other studies,we observed that pre-treatment twice with LC seemed to protect against APAP hepatotoxicity,in contrast to acute pre-treatment.The aim of this study was to confirm that observation and to explore the mechanisms.Methods:We treated mice with empty liposomes(LE)or LC twice per week for 1 week before APAP overdose and collected blood and liver tissue at 0,2,and 6 h post-APAP.We then measured liver injury(serum alanine aminotransferase activity,histology),APAP bioactivation(total glutathione,APAP-protein adducts),oxidative stress(oxidized glutathione(GSSG)),glutamate-cysteine ligase subunit c(Gclc)mRNA,and nuclear factor erythroid 2-related factor(Nrf2)immunofluorescence.We also confirmed the ablation of macrophages by F4/80 immunohistochemistry.Results:Pre-treatment twice with LC dramatically reduced F4/80 staining,protected against liver injury,and reduced oxidative stress at 6 h post-APAP,without affecting APAP bioactivation.Importantly,Gclc mRNA was higher in the LC group at 0 h and total glutathione was higher at 2 h,indicating accelerated glutathione re-synthesis after APAP overdose due to greater basal glutamate-cysteine ligase.Oxidative stress was lower in the LC groups at both time points.Finally,total Nrf2 immunofluorescence was higher in the LC group.Conclusions:We conclude that multiple pre-treatments with LC protect against APAP by accelerating glutathione re-synthesis through glutamate-cysteine ligase.Investigators using twice or possibly more LC pre-treatments to deplete macrophages,including peritoneal macrophages,should be aware of this possible confounder.