Abstract
Chrysophanol (Chr) is the main monomer isolated from Rheum rhabarbarum. This study aimed to identify the potential in vitro and in vivo cytoprotective effects of Chr on lipopolysaccharide (LPS)-triggered acute lung injury (ALI). We used an ALI-murine model and constructed an inflammatory macrophage in vitro cell model to determine the cellular mechanisms involved in Chr-mediated activity. To observe the vital role of histone deacetylase 3 (HDAC3) in abolishing inflammation action, HDAC3 was downregulated using small interfering RNA. Analysis of the expression of nuclear transcription factor-kappa B p65 (NF-κB p65) and molecules of its downstream signaling pathway were assessed in vitro and in lung tissue samples using the mouse model. Concentrations of tumor necrosis factor-α, interleukin-1β, high mobility group protein 1 (HMGB1), and interleukin-16 in supernatants and the bronchoalveolar lavage fluid were measured using enzyme-linked immunosorbent assay. A reporter gene assay measured HMGB1 activity, and NF-κB p65 and HMGB1 intracellular localization was determined by immunofluorescence detection on histological lung samples from Chr-treated mice. The protein interactions between HMGB1, HDAC3, and NF-κB p65 were tested by co-immunoprecipitation. Chr treatment relieved LPS-induced lung lesions. Chr also enhanced superoxide dismutase levels in ALI mice. Chr reduced the LPS-induced protein expression of NF-κB and its related pathway molecules in both in vivo and in vitro models. Moreover, Chr downregulated LPS-enhanced HMGB1 expression, acetylation, and nuclear nucleocytoplasmic translocation. However, HDAC3 knockdown substantially reduced Chr-mediated HDAC3/NF-κB expression.Furthermore, Chr enhanced HMGB1/HDAC3/NF-κB p65 complex interaction, whereas HDAC3 knockdown reduced Chr-mediated HMGB1/HDAC3/NF-κB p65 formation. This study showed that the protective effects induced by Chr were associated with the regulation of the HMGB1/NF-κB pathway via HDAC3.
Highlights
Sepsis, a generalized inflammatory state induced by lipopolysaccharide (LPS) infection, usually leads to acute lung injury (ALI)
We found that under the negative control (NC) or histone deacetylase 3 (HDAC3)-small interfering RNA (siRNA) conditions, high mobility group box 1 (HMGB1) protein was detected in the IP product using the p65 antibody, and p65 protein was detected in the IP product using the HMGB1 antibody (Figures 9A,B)
Sepsis is a life-threatening pathological condition characterized by a dysregulated inflammatory response, a disordered blood coagulation cascade, and multiple organ dysfunctions (Falagas and Kopterides, 2006; Jean-Baptiste, 2007; Prescott et al, 2016)
Summary
A generalized inflammatory state induced by lipopolysaccharide (LPS) infection, usually leads to acute lung injury (ALI). Septic ALI is induced by a lung inflammatory response syndrome, which leads to high mortality rates, high patient management costs, and accelerated morbidity (Ishii, 2011). As a late pro-inflammatory factor, the extracellular high mobility group box 1 (HMGB1) protein triggers responses that cause damage to tissues leading to the activation of the inflammatory cascade in several conditions, including lung injury and septic shock (Czura et al, 2004; El Gazzar, 2007). LPS diffuses into the lung via blood circulation, activates macrophages, and promotes HMGB1 release.
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