6-Gingerol ameliorates alveolar hypercoagulation and fibrinolytic inhibition in LPS-provoked ARDS via RUNX1/NF-κB signaling pathway.

BACKGROUND: Alveolar hypercoagulation and fibrinolytic inhibition play a central role in refractory hypoxemia in acute respiratory distress syndrome (ARDS), but it lacks effective drugs for prevention and treatment of this pathophysiology. Our previous experiment confirmed that RUNX1 promoted alveolar hypercoagulation and fibrinolytic inhibition through NF-κB pathway. Other studies demonstrated that 6-gingerol regulated inflammation and metabolism by inhibiting the NF-κB signaling pathway. We assume that 6-gingerol would ameliorate alveolar hypercoagulation and fibrinolytic inhibition via RUNX1/ NF-κB pathway in LPS-induced ARDS. METHODS: Rat ARDS model was replicated through LPS inhalation. Before LPS inhalation, the rats were intraperitoneally treated with different doses of 6-gingerol or the same volume of normal saline (NS) for 12 h, and then intratracheal inhalation of LPS for 24 h. In cell experiment, alveolar epithelial cell type II (AECII) was treated with 6-gingerol for 6 h and then with LPS for another 24 h. RUNX1 gene was down-regulated both in pulmonary tissue and in cells. Tissue factor (TF), plasminogen Activator Inhibitor 1(PAI-1) and thrombin were determined by Wester-blot (WB), qPCR or by enzyme-linked immunosorbent (ELISA). Lung injury score, pulmonary edema and pulmonary collagen III in rat were assessed. NF-κB pathway were also observed in vivo and in vitro. The direct binding capability of 6-gingerol to RUNX1 was confirmed by using Drug Affinity Responsive Target Stability test (DARTS). RESULTS: 6-gingerol dose-dependently attenuated LPS-induced lung injury and pulmonary edema. LPS administration caused excessive TF and PAI-1 expression both in pulmonary tissue and in AECII cell and a large amount of TF, PAI-1 and thrombin in bronchial alveolar lavage fluid (BALF), which all were effectively decreased by 6-gingerol treatment in a dose-dependent manner. The high collagen Ⅲ level in lung tissue provoked by LPS was significantly abated by 6-gingerol. 6-gingerol was seen to dramatically inhibit the LPS-stimulated activation of NF-κB pathway, indicated by decreases of p-p65/total p65, p-IKKß/total IKKß, and also to suppress the RUNX1 expression. RUNX1 gene knock down or RUNX1 inhibitor Ro5-3335 significantly enhanced the efficacies of 6-gingerol in vivo and in vitro, but RUNX1 over expression remarkably impaired the effects of 6-gingerol on TF, PAI-1 and on NF-κB pathway. DARTS result showed that 6-gingerol directly bond to RUNX1 molecules. CONCLUSIONS: Our experimental data demonstrated that 6-gingerol ameliorates alveolar hypercoagulation and fibrinolytic inhibition via RUNX1/NF-κB pathway in LPS-induced ARDS. 6-gingerol is expected to be an effective drug in ARDS.

Glycyrrhizin alleviates sepsis-induced acute respiratory distress syndrome via suppressing of HMGB1/TLR9 pathways and neutrophils extracellular traps formation.

BACKGROUND: Neutrophil extracellular traps (NETs) are involved in the development of sepsis-induced acute respiratory distress syndrome (ARDS). Glycyrrhizin (GL), the main active ingredient of the traditional Chinese medicine Glycyrrhiza glabra, has anti-inflammatory, anti-viral, and immunomodulatory effects. OBJECTIVE: The study aims to explore the efficacy and potential mechanism of GL on sepsis-induced ARDS in mice. MATERIALS AND METHODS: Mice were randomly divided into 3 groups: Control, CLP, and GL + CLP. Mice sepsis ARDS model was induced by cecal ligation and puncture (CLP) followed by intraperitoneal GL treatment. Then, the 7-day survival rate of mice was recorded. The lung function of mice was determined by whole-body plethysmography. Lung pathology and scores were observed by hematoxylin-eosin staining. The wet/dry ratio (W/D) of the lung was measured by weighing method. The protein concentration in bronchoalveolar lavage fluid (BALF) was measured by the BCA method. NETs formation in lung tissue was detected by immunofluorescence. Furthermore, HMGB1、TLR9、MyD88 and IL6 expression in lung tissue were detected by western blot and by quantitative real-time PCR, respectively. RESULTS: The results showed that GL improved the survival rate, attenuated lung tissue injury and reduced the expression of inflammatory factors in mice with CLP-induced sepsis. Meanwhile, we confirmed that GL could inhibit TLR9 / MyD88 activation from reducing NETs formation by decreasing HMGB1 expression. The formation of NETs is regulated by HMGB1 / TLR9 / MyD88. In addition, GL improved lung function in mice with sepsis-induced ARDS. Lung function suggested that GL increased alveolar ventilation, alleviated ventilator fatigue and reduced airway resistance in mice with ARDS induced by sepsis. CONCLUSIONS: GL ameliorated sepsis-induced ARDS and reduced the NETs formation in lung tissues, which may be associated with the inhibition of the HMGB1 / TLR9 pathway.