Hepatic mitochondrial DNA/Toll‐like receptor 9/MicroRNA‐223 forms a negative feedback loop to limit neutrophil overactivation and acetaminophen hepatotoxicity in mice

Abstract

Acetaminophen (APAP) overdose is a leading cause of acute liver failure worldwide, in which mitochondrial DNA (mtDNA) released by damaged hepatocytes activates neutrophils via the binding of TLR9, further aggravating liver injury. Here, we demonstrated that mtDNA/TLR9 also activates a negative feedback pathway via the induction of microRNA-223 (miR-223) to limit neutrophil over-activation and liver injury. After injection of APAP in mice, levels of miR-223, the most abundant miRNAs in neutrophils, were highly elevated in neutrophils. Disruption of the miR-223 gene exacerbated APAP-induced hepatic neutrophil infiltration, oxidative stress, and injury, and enhanced TLR9 ligand-mediated activation of pro-inflammatory mediators in neutrophils. An additional deletion of the intercellular adhesion molecule 1 (ICAM-1) gene ameliorated APAP-induced neutrophil infiltration and liver injury in miR-223 knockout mice. In vitro experiments revealed that miR-223-deficient neutrophils were more susceptible to TLR9 agonist-mediated induction of pro-inflammatory mediators and NF-κB signaling; whereas overexpression of miR-223 attenuated these effects in neutrophils. Moreover, inhibition of TLR9 signaling via either treatment with a TLR9 inhibitor or via the disruption of TLR9 gene partially but significantly suppressed miR-223 expression in neutrophils post APAP injection. In contrast, activation of TLR9 upregulated miR-223 expression in neutrophils in vivo and in vitro. Mechanistically, activation of TLR9 upregulated miR-223 by enhancing NF-κB binding on miR-223 promoter; while miR-223 attenuated TLR9/NF-κB-mediated inflammation by targeting IKKα expression. Collectively, upregulation of miR-223 plays a key role in terminating the acute neutrophilic response and is a therapeutic target for the treatment of APAP-induced liver failure.

MicroRNA miR-223 as regulator of innate immunity

“…The regulatory role of miR‐223 in neutrophil activation and function was further explored in hepatic injury models. miR‐223 acts as a negative regulator to neutrophil activation during acetaminophen (APAP) overdose . APAP overdose elicits the level of miR‐223 in neutrophils to limit neutrophil activation and prevent further liver injury.…”

MicroRNAs in alcoholic liver disease: Recent advances and future applications

“…MiR‐223, encoded on chromosome 12, one of the most abundant miRNAs in neutrophils, was initially reported to regulate the early and late stages of granulopoiesis as well as in the homeostasis of mature neutrophils (He et al, ; Li et al, ). He et al () previous studies demonstrated that upregulation of miR‐223 plays a key role in terminating the acute neutrophilic response and is a therapeutic target for the treatment of acetaminophen (APAP) induced liver failure. Later studies suggested that there may be a correlation between the level of miR‐223 and ALD.…”

Utilizing Mechanistic Biomarkers in Treating Paracetamol Hepatotoxicity

“…The inflammatory pathways activated by these DAMPs have been elaborately studied. Accordingly, blocking leukocyte DAMP receptors such as toll‐like receptor 9 has shown promise in preclinical models of PCM toxicity . The merits of neutralizing HMGB1 directly have also been demonstrated in animal studies .…”