Syringic acid attenuates acute lung injury by modulating macrophage polarization in LPS-induced mice

Background: Acute lung injury (ALI) is a continuum of lung changes caused by multiple lung injuries, characterized by a syndrome of uncontrolled systemic inflammation that often leads to significant morbidity and death. Anti-inflammatory is one of its treatment methods, but there is no safe and available drug therapy. Syringic acid (SA) is a natural organic compound commonly found in a variety of plants, especially in certain woody plants and fruits. In modern pharmacological studies, SA has anti-inflammatory effects and therefore may be a potentially safe and available compound for the treatment of acute lung injury. Purpose: This study attempts to reveal the protective mechanism of SA against ALI by affecting the polarization of macrophages and the activation of NF-kappa B signaling pathway. Trying to find a safer and more effective drug therapy for clinical use. Methods: We constructed the ALI model using C57BL/6 mice by intratracheal instillation of LPS (10 mg/kg). Histological analysis was performed with hematoxylin and eosin (H&E). The wet-dry ratio of the whole lung was measured to evaluate pulmonary edema. The effect of SA on macrophage M1-type was detected by flow cytometry. BCA protein quantification method was used to determine the total protein concentration in bronchoalveolar lavage fluid (BALF). The levels of Interleukin (IL)-6, IL-1 beta, and tumor necrosis factor (TNF)-alpha in BALF were determined by the ELISA kits, and RT-qPCR was used to detect the expression levels of IL-6, IL-1 beta and TNF-alpha mRNA of lung tissue. Western blot was used to detect the expression levels of iNOS and COX-2 and the phosphorylation of p65 and I kappa B alpha in the NF-kappa B pathway in lung tissue. In vitro experiments were conducted with RAW267.4 cell inflammation model induced by 100 ng/ml LPS and A549 cell inflammation model induced by 10 mu g/ml LPS. The effects of SA on M1-type and M2-type macrophages of RAW267.4 macrophages induced by LPS were detected by flow cytometry. The toxicity of compound SA to A549 cells was detected by MTT method which to determine the safe dose of SA. The expressions of COX-2 and the phosphorylation of p65 and I kappa B alpha protein in NF-kappa B pathway were detected by Western blot. Results: We found that the pre-treatment of SA significantly reduced the degree of lung injury, and the infiltration of neutrophils in the lung interstitium and alveolar space of the lung. The formation of transparent membrane in lung tissue and thickening of alveolar septum were significantly reduced compared with the model group, and the wet-dry ratio of the lung was also reduced. ELISA and RT-qPCR results showed that SA could significantly inhibit the production of IL -6, IL -1 8, TNF-alpha. At the same time, SA could significantly inhibit the expression of iNOS and COX -2 proteins, and could inhibit the phosphorylation of p65 and I kappa B alpha proteins. in a dose -dependent manner. In vitro experiments, we found that flow cytometry showed that SA could significantly inhibit the polarization of macrophages from M0 type macrophages to M1 -type macrophages, while SA could promote the polarization of M1 -type macrophages to M2 -type macrophages. The results of MTT assay showed that SA had no obvious cytotoxicity to A549 cells when the concentration was not higher than 80 mu M, while LPS could promote the proliferation of A549 cells. In the study of anti-inflammatory effect, SA can significantly inhibit the expression of COX -2 and the phosphorylation of p65 and I kappa B alpha proteins in LPS-induced A549 cells. Conclusion: SA has possessed a crucial anti-ALI role in LPS-induced mice. The mechanism was elucidated, suggesting that the inhibition of macrophage polarization to M1 -type and the promotion of macrophage polarization to M2 -type, as well as the inhibition of NF- kappa B pathway by SA may be the reasons for its anti-ALI. This finding provides important molecular evidence for the further application of SA in the clinical treatment of ALI.