Neutrophil activation during acetaminophen hepatotoxicity and repair in mice and humans

Abstract

Following acetaminophen (APAP) overdose there is an inflammatory response triggered by release of cellular contents from necrotic hepatocytes into systemic circulation which initiates the recruitment of neutrophils into the liver. It has been demonstrated that neutrophils do not contribute to APAP-induced liver injury, but their role and the role of NADPH oxidase in injury resolution is controversial. C57BL/6 mice were subjected to APAP overdose and neutrophil activation status was determined during liver injury and liver regeneration. Additionally, human APAP overdose patients (ALT: >800U/L) had serial blood draws during the injury and recovery phases for determination of neutrophil activation. Neutrophils in the peripheral blood of mice showed increasing activation status (CD11b expression and ROS priming) during and after the peak of injury but returned to baseline levels prior to complete injury resolution. Hepatic sequestered neutrophils showed an increased and sustained CD11b expression, but no ROS priming was observed. Confirming that NADPH oxidase is not critical to injury resolution, gp91phox-/- mice following APAP overdose displayed no alteration in injury resolution. Peripheral blood from APAP overdose patients also showed increased neutrophil activation status after the peak of liver injury and remained elevated until discharge from the hospital. In mice and humans, markers of activation, like ROS priming, were increased and sustained well after active liver injury had subsided. The similar findings between surviving patients and mice indicate neutrophil activation may be a critical event for host defense or injury resolution following APAP overdose, but not a contributing factor to APAP-induced injury.

Commentary to Choi et al. (2015): CCR5 knockout mice with C57BL6 background are resistant to acetaminophen-mediated hepatotoxicity due to decreased macrophages migration into the liver

“…Importantly, these results are consistent with human data where monocyte and neutrophil activation and recruitment occur after the APAP-induced liver injury during the regeneration phase (Antoniades et al 2012;Williams et al 2014). In addition, APAP hepatotoxicity in humans and human hepatocytes is caused by mitochondrial dysfunction (McGill et al 2012;Xie et al 2014).…”

“…Thus, these interventions affect different cell types. Second, although GdCl 3 has been shown to reduce the capacity of Kupffer cells to generate reactive oxygen (Liu et al 1995), experiments with gp91phox KO mice, which have no functional NADPH oxidase in any phagocytes, demonstrated that these mice have the same oxidant stress, peroxynitrite formation and liver injury after APAP treatment as wild-type animals (James et al 2003;Williams et al 2014). These observations virtually eliminate any involvement of phagocytes including resident Kupffer cells or newly recruited neutrophils and monocytes, as source of oxidant stress during APAP hepatotoxicity.…”

Acetaminophen Toxicity: Novel Insights Into Mechanisms and Future Perspectives

“…Furthermore, there was neither neutrophil activation nor a neutrophil-mediated oxidant stress observed in the liver during the injury phase 94,117 . Most importantly, similar results regarding neutrophil activation were obtained in human patients 119 . Together the findings strongly argue against a direct involvement of neutrophils in the injury phase.…”

“…Although it is not always obvious why so many contradictory results are being reported, it might be useful to focus on the clinically relevant aspects. In APAP overdose patients, pro-inflammatory cytokine formation is limited 128 and neutrophil activation does not occur during the injury phase but more during regeneration 119 . Consistent with these neutrophil findings, monocyte-derived macrophages that are recruited during APAP hepatotoxicity display a pro-regenerative phenotype in mice and in humans suggesting that the inflammatory response is mainly geared towards recovery from the tissue injury 129–131 .…”

Platelets and protease-activated receptor-4 contribute to acetaminophen-induced liver injury in mice

“…It is worth noting, however, that the role of neutrophils in APAP hepatotoxicity continues to be debated. 8,[49][50][51] Although we cannot exclude the possibility that activated platelets promote neutrophildriven exacerbation of injury, it is also possible that reduced neutrophil accumulation in platelet-depleted mice reflects reduced necrosis (ie, a diminished recruitment of neutrophils secondary to necrosis). Additional studies are required to elucidate the exact mechanism, whereby platelets drive early APAP-induced liver damage.…”