The effects of etomidate on expression of high mobility group box 1 via the nuclear factor kappa B pathway in rat model of sepsis

Abstract

Etomidate is an anesthetic agent used in hemodynamically unstable patients, but its use has been controversial in septic patients. The response of high-mobility group box 1 (HMGB1), a late-phase lethal cytokine in sepsis, to etomidate has not been reported. This study investigated the effects of etomidate on the expression and release of HMGB1 and the underlying mechanism using a cecal ligation and puncture (CLP) model. Thirty-six male Sprague-Dawley rats were divided into sham, CLP, and Etomi groups. Sepsis was induced in the CLP and Etomi groups, and intravenous etomidate (4 mg/kg) was infused for 40 min immediately after operation in the Etomi group. Serum creatinine, alanine aminotransferase (ALT), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and HMGB1 levels were measured 6 and 24 hours after surgery. Activation of nuclear factor (NF)-ĸB and HMGB1 mRNA expression in the liver, lung, kidney, and ileum tissues were measured, and immunohistochemical staining of HMGB1 was implemented. Increases of the TNF-α level 6 h after CLP and ALT and IL-6 levels 24 h after CLP were significantly inhibited by etomidate treatment. Etomidate treatment also significantly attenuated the increase in serum HMGB1 level at 6 and 24 h after CLP and suppressed the NF-ĸB and HMGB1 mRNA in multiple organs 24 h after CLP. Immunohistochemical staining also revealed that etomidate treatment inhibited HMGB1 expression. Etomidate inhibited the systemic release of HMGB1 and its expression in various organs. The mechanism may be associated with the inhibitory effects of etomidate on pro-inflammatory cytokine release and NF-ĸB activity.

Keywords: Cecal ligation and puncture; etomidate; high mobility group box1; nuclear factor kappa B; sepsis.

Figure 1.

(a) Histopathologic changes in organs of CLP-induced septic rats. Hematoxylin and eosin-stained sections of liver, lungs and kidneys from rats subjected to sham/CLP operation 24 h after surgery (magnification $\times$400). Etomidate treatment attenuates acute organ injury by CLP-induced sepsis. (b) Quantification of the severity of inflammation in each organ (ranges from 0 to 4). *P<0.05 vs Sham, †P<0.05 vs CLP. Sham: sham operation group, CLP: Cecal ligation and puncture with normal saline group, Etomi: Cecal ligation and puncture with etomidate treatment group.

Figure 2.

Effects of etomidate treatment in systemic inflammatory cytokines concentration 6 and 24 h after sham/CLP operations. ELISA analysis of serum TNF-α, IL-6 and HMGB1 concentration. Data were presented as mean ± SD (n = 6). *P<0.05 vs Sham, †P<0.05 vs CLP. Sham: sham operation group, CLP: Cecal ligation and puncture with normal saline group, Etomi: Cecal ligation and puncture with etomidate treatment group.

Figure 3.

Effect of etomidate on activity of NF-κB in organs of CLP-induced septic rats. Liver, Lungs, kidneys and ileum were collected 24 h after surgery and NF-κB p65 activation was analyzed by Western blot. Etomidate treatment attenuated the increase in NF-κB activity after CLP surgery. *P<0.05 vs Sham, †P<0.05 vs CLP.

Figure 4.

(a) the expressions of HMGB1 mRNA in tissues from rats in each group 24 h after sham/CLP surgeries. Liver, lungs, kidneys and ileum were taken 24 h after surgery. HMGB1 mRNA expression in multiple organs were measured by Real-Time PCR. Etomidate treatment inhibits excessive HMGB1 mRNA expression in various organs. *P<0.05 vs Sham, †P<0.05 vs CLP. (b) Immunohistochemical staining of HMGB1 in liver, lungs, and kidneys from rats 24 h after sham/CLP surgeries (magnification $\times$400). The expression and translocation of HMGB1 were increased in the septic rats that underwent CLP operation compared with rats that underwent the sham operation (indicated by black arrows), but etomidate treatment attenuated the expression of HMGB1. Sham: sham operation group, CLP: Cecal ligation and puncture with normal saline group, Etomi: Cecal ligation and puncture with etomidate treatment group.