Tetramethylpyrazine Showed Therapeutic Effects on Sepsis-Induced Acute Lung Injury in Rats by Inhibiting Endoplasmic Reticulum Stress Protein Kinase RNA-Like Endoplasmic Reticulum Kinase (PERK) Signaling-Induced Apoptosis of Pulmonary Microvascular Endothelial Cells

Abstract

BACKGROUND Acute lung injury (ALI) is a life-threatening complication of sepsis. Tetramethylpyrazine (TMP) has been used in the clinical treatment of vascular diseases. The aim of this study was to investigate the therapeutic effects and possible involved mechanisms on ALI. MATERIAL AND METHODS Cecal ligation and puncture (CLP) was used to establish a sepsis model in rats. TMP at various dosages were administrated to rats using a intragastric method. Animal survival rate was calculated. The lung functions were evaluated by lung weight/dry weight ratio (W/D), PaO2, dynamic compliance (DC), and airway resistance index (ARI). Pulmonary microvascular endothelial cells (PMVECs) were isolated from lungs harvested from rats with sepsis. TUNEL assay was used to detect apoptosis. Protein expression and phosphorylation levels were assessed by western blotting. RESULTS TMP administration increased the survival rate of septic rats. TMP also decreased W/D and DC, but increased PaO2 and ARI in septic rats. Moreover, PMVECs apoptosis was inhibited in septic rats that received TMP treatment. The expression levels of GRP78, ATF4, caspase-12, active caspase-3, as well as the phosphorylation levels of PERK and eIF2α were suppressed in PMVECs isolated from TMP-treated septic rats. CONCLUSIONS TMP alleviated sepsis-induced ALI by suppressing PMVECs apoptosis via PERK/eIF2α/ATF4/CHOP apoptotic signaling in endoplasmic reticulum stress.

Figure 1

The chart shows the Kaplan-Meier survival curves of control and sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. Log-rank test was carried out to analyze the differences between groups. * P<0.05 indicates differences significant between groups.

Figure 2

(A) Columns show the PaO₂ of blood sample collected from aorta in sepsis rats that received TMP at concentrations of 0, 20, 40 and 60 mg/kg. (B) Columns show the wet/dry weight ratio (W/D) of lung in sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (C) Columns show the change of airway resistance index (RI) in sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (D) Columns show the change of dynamic compliance (DC) in sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. * P<0.05 indicates differences significant between groups.

Figure 3

Upper portion shows the captured images of TUNEL, DAPI, and their merged images of PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. * P<0.05 indicates differences significant between groups.

Figure 4

A) Immunoblots of GRP78, p-PERK, PERK, p-eIF2α, eIF2α, ATF4, CHOP, caspase-12, active caspase-3, and GAPDH in PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (B) Columns indicated the relative expression levels of GRP78 (white), ATF4 (blue), and CHOP (deep blue) in PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (C) Columns show the phosphorylation level of PERK in PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (D) Columns show the relative expression levels of caspase-12 (white) and active caspase-3 (deep blue) in PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. (E) Columns show the phosphorylation level of eIF2α in PMVECs isolated from sepsis rats that received TMP at concentrations of 0, 20, 40, and 60 mg/kg. * P<0.05 indicates differences significant between groups.