Toll-like Receptor 4 (TLR4) Antagonist Eritoran Tetrasodium Attenuates Liver Ischemia and Reperfusion Injury through Inhibition of High-Mobility Group Box Protein B1 (HMGB1) Signaling

Abstract

Toll-like receptor 4 (TLR4) is ubiquitously expressed on parenchymal and immune cells of the liver and is the most studied TLR responsible for the activation of proinflammatory signaling cascades in liver ischemia and reperfusion (I/R). Since pharmacological inhibition of TLR4 during the sterile inflammatory response of I/R has not been studied, we sought to determine whether eritoran, a TLR4 antagonist trialed in sepsis, could block hepatic TLR4-mediated inflammation and end organ damage. When C57BL/6 mice were pretreated with eritoran and subjected to warm liver I/R, there was significantly less hepatocellular injury compared to control counterparts. Additionally, we found that eritoran is protective in liver I/R through inhibition of high-mobility group box protein B1 (HMGB1)-mediated inflammatory signaling. When eritoran was administered in conjunction with recombinant HMGB1 during liver I/R, there was significantly less injury, suggesting that eritoran blocks the HMGB1-TLR4 interaction. Not only does eritoran attenuate TLR4-dependent HMGB1 release in vivo, but this TLR4 antagonist also dampened HMGB1's release from hypoxic hepatocytes in vitro and thereby weakened HMGB1's activation of innate immune cells. HMGB1 signaling through TLR4 makes an important contribution to the inflammatory response seen after liver I/R. This study demonstrates that novel blockade of HMGB1 by the TLR4 antagonist eritoran leads to the amelioration of liver injury. Figure 5. Eritoran treatment suppresses HMGB1 release in vivo. (A) Nuclear and cytoplasmic HMGB1 levels were determined by Western blot and quantitative densitometry analysis of the protein expressions in eritoran-treated mice and control mice that underwent ischemia and 6 h of reperfusion determined. Hepatic protein lysates from ischemic lobes were obtained; each lane represents a separate animal. The blots shown are representative of two experiments with similar results. (B) Serum HMGB1 ELISA after 6 h of reperfusion. Data are representative of two experiments with similar results. Gray, eritoran; white, control. *P < 0.05 when compared against control by one-way ANOVA. (C) Immunofluorescent stain of HMGB1 from sections of sham liver and liver 6 h after I/R in placebo control, and eritoran-treated mice (magnification 40×). Images are representative of liver sections from two mice per group. Red, HMGB1; blue, nuclei; green, F-actin.(D) TLR4 wild-type and knockout mice were given eritoran (5 mg/kg body weight) or vehicle control intraperitoneally 30 min before ischemia. HMGB1 levels in serum were collected 6 h after reperfusion and quantified using ELISA. Data represent the mean ± SEM (n ≥ 6 mice per group). Gray, eritoran; white, control. *P < 0.05, by one-way ANOVA.

Acetyl‐3‐Aminoethyl Salicylate Ameliorates Hepatic Ischemia/Reperfusion Injury and Liver Graft Survival Through a High‐Mobility Group Box 1/Toll‐Like Receptor 4–Dependent Mechanism

“…Salicylates directly bind to and inhibit HMGB1 (the key ligand of TLR4 in promoting HIRI), and HMGB1/TLR4 binding antagonism has been shown to attenuate HIRI . Therefore, we next hypothesized that ac3AESA's effects in HIRI are dependent on TLR4.…”

The spleen contributes importantly to myocardial infarct exacerbation during post-ischemic reperfusion in mice via signaling between cardiac HMGB1 and splenic RAGE

“…DAMPs include mediators such as heat shock proteins[23], HMGB1[48], hyaluronic acid[22], S100[35], DNA fragments[51] and fibronectin fragments[36]. As an important DAMP, HMGB1 has been shown to play critical roles in sterile tissue injury such as stroke[25] and I/R injury of the liver[30], kidney[45] and heart[2, 18]. HMGB1 can mediate the chemotaxis of leukocytes[19, 50].…”

High‐Mobility Group Box‐1 and Liver Disease

“…Additional support for the role of HMGB1 in warm I/R comes from the plethora of compounds targeting HMGB1 signaling that have been found to successfully ameliorate injury . Thus, the role of hepatic HMGB1 in warm I/R is still contradictory.…”