Emodin alleviates sepsis-mediated lung injury via inhibition and reduction of NF-kB and HMGB1 pathways mediated by SIRT1

Abstract

Inflammation plays an important role during sepsis, and excessive inflammation can result in organ damage, chronic inflammation, fibrosis, and scarring. The study aimed to investigate the specific mechanism of emodin by constructing in vivo and in vitro septic lung injury models via inhibition and reduction of NF-kB and high mobility group box 1 (HMGB1) pathways. A cecal ligation and puncture (CLP) model was built for adult male Sprague-Dawley rats. Concentrations of TNF-α, IL-1β, and IL-6 in bronchoalveolar lavage fluid were determined using commercially available ELISA kits. Hematoxylin and eosin staining was used for the right lung inferior lobes. Myeloperoxidase (MPO) activity of the lung tissue was detected by using the MPO kit. Murine alveolar epithelial cell line (MLE-12) cells were used for flow cytometry and Western blot to analyze the apoptosis rate and protein expression. Emodin significantly decreased CLP-induced cell apoptosis, upregulated expression of sirtuin 1 (SIRT1), and inhibited p-p65/p65 and HMGB1. In lipopolysaccharide (LPS) treated cell model, emodin treatment markedly decreased LPS-induced release of IL-1, IL-6, and tumor necrosis factor (TNF)-α, inhibited LPS-induced cell apoptosis and suppressed protein levels of P-P65/P65 and HMGB1. However, science of SIRT1 reversed the above effects by treatment of emodin. In summarize, this study found that emodin can alleviate sepsis-induced lung injury in vivo and in vitro through regulation of SIRT1.

Keywords: HMGB1; NF-kB; SIRT1; emodin; sepsis.

FIGURE 1

Emodin weakens CLP‐induced injury in the sepsis model. (A) HE staining was used to detect lung tissue in the SHAM, CLP, and CLP + emodin groups of mouse models. (B–D) The levels of (B) IL‐1, (C) IL‐6, and (D) TNF‐α in the lung tissues of eight mice in each group (SHAM, CLP, and CLP + emodin) were detected by ELISA. (E) The MPO activity level of lung tissues in different groups of mouse models was detected by ELISA. (F) Flow cytometry was used to detect the apoptosis level of lung tissue in different groups of mouse models

FIGURE 2

Emodin upregulated SIRT1 protein and inhibited p‐p65/p65 and HMGB1 protein levels. (A) SIRT1, (B) p‐P65/P65, and (C) HMGB1 protein levels in the lung tissues in different groups of mouse models were detected by Western blot. The results show that the SIRT1 protein level was decreased, while the p‐P65/P65 and HMGB1 protein levels were increased. After emodin treatment, the SIRT1 protein level was partially increased, while the p‐P65/P65 and HMGB1 protein levels were partially decreased

FIGURE 3

Emodin attenuates LPS‐induced injury in MLE‐12 cells. (A–C) The levels of (A) IL‐1, (B) IL‐6, and (C) TNF‐α in different groups of MLE‐12 cells were detected by ELISA. (D) Flow cytometry was used to detect the apoptosis level of lung cells in different groups of mouse models. (E–G) (E) SIRT1, (F) p‐P65/P65, and (G) HMGB1 protein levels in different groups of MLE‐12 cells were detected by Western blot. After LPS induction, the SIRT1 protein level was decreased, while the P‐P65/P65 and HMGB1 protein levels were increased. After emodin treatment, the protein level of SIRT1 was partly increased, while the protein levels of P‐P65/P65 and HMGB1 were partly decreased

FIGURE 4

Emodin alleviates sepsis‐induced lung injury via inhibition and reduction of NF‐kB and HMGB1 protein levels mediated by SIRT1. (A) The protein level of SIRT1 in MLE‐12 cells (si‐NC and si‐SIRT1) was detected by Western blot. (B–D) ELISA was used to detect the levels of (B) IL‐1, (C) IL‐6, and (D) TNF‐α in different groups of MLE‐12 cells. IL‐1, IL‐6, and TNF‐α were increased after LPS induction, while IL‐1, IL‐6, and TNF‐α were partially decreased after emodin treatment. IL‐1, IL‐6, and TNF‐α were partially elevated after transfection with si‐SIRT1. (E) Flow cytometry was used to detect the apoptosis level of lung cells in different groups of mouse models. The apoptosis level was increased after LPS induction, partially decreased after emodin treatment, and partially increased after si‐SIRT1 transfection. (F,G) The levels of (F) p‐P65/P65 and (G) HMGB1 were measured in different MLE‐12 groups. The levels of P‐P65/P65 and HMGB1 were increased after LPS induction. After emodin treatment, the protein levels of P‐P65/P65 and HMGB1 were partially reduced. The protein levels of P‐P65/P65 and HMGB1 were partially increased after si‐SIRT1 transfection