Syringic acid attenuates acute lung injury by modulating macrophage polarization in LPS-induced mice.
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.
This study attempts to reveal the protective mechanism of SA against ALI by affecting the polarization of macrophages and the activation of NF-κB signaling pathway. Trying to find a safer and more effective drug therapy for clinical use.
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-1β, and tumor necrosis factor (TNF)-α in BALF were determined by the ELISA kits, and RT-qPCR was used to detect the expression levels of IL-6, IL-1β and TNF-α mRNA of lung tissue. Western blot was used to detect the expression levels of iNOS and COX-2 and the phosphorylation of p65 and IκBα in the NF-κB 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 μg/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 IκBα protein in NF-κB pathway were detected by Western blot.
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-1β, TNF-α. At the same time, SA could significantly inhibit the expression of iNOS and COX-2 proteins, and could inhibit the phosphorylation of p65 and IκBα 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 μM, 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 IκBα proteins in LPS-induced A549 cells.
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-κB 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.
Wang WT
，Zhang YY
，Li ZR
，Li JM
，Deng HS
，Li YY
，Yang HY
，Lau CC
，Yao YJ
，Pan HD
，Liu L
，Xie Y
，Zhou H
... -
《-》
Knockdown of angiopoietin-like 4 suppresses sepsis-induced acute lung injury by blocking the NF-κB pathway activation and hindering macrophage M1 polarization and pyroptosis.
Sepsis-induced acute lung injury (ALI) is a life-threatening disease. Macrophage pyroptosis has been reported to exert function in ALI. We aimed to investigate the mechanisms of ANGPTL4-mediated cell pyroptosis in sepsis-induced ALI, thus providing new insights into the pathogenesis and prevention and treatment measures of sepsis-induced ALI.
In vivo animal models and in vitro cell models were established by cecal ligation and puncture (CLP) method and lipopolysaccharide-induced macrophages RAW264.7. ANGPTL4 was silenced in CLP mice or macrophages, followed by the determination of ANGPTL4 expression in bronchoalveolar lavage fluid (BALF) or macrophages. Lung histopathology was observed by H&E staining, with pathological injury scores evaluated and lung wet and dry weight ratio recorded. M1/M2 macrophage marker levels (iNOS/CD86/Arg1), inflammatory factor (TNF-α/IL-6/IL-1β/iNOS) expression in BALF, cell death and pyroptosis, NLRP3 inflammasome, cell pyroptosis-related protein (NLRP3/Cleaved-caspase-1/caspase-1/GSDMD-N) levels, NF-κB pathway activation were assessed by RT-qPCR/ELISA/flow cytometry/Western blot, respectively.
ANGPTL4 was highly expressed in mice with sepsis-induced ALI, and ANGPTL4 silencing ameliorated sepsis-induced ALI in mice. In vivo, ANGPTL4 silencing repressed M1 macrophage polarization and macrophage pyroptosis in mice with sepsis-induced ALI. In vitro, ANGPTL4 knockout impeded LPS-induced activation and pyroptosis of M1 macrophages and hindered LPS-induced activation of the NF-κB pathway in macrophages.
Knockdown of ANGPTL4 blocks the NF-κB pathway activation, hinders macrophage M1 polarization and pyroptosis, thereby suppressing sepsis-induced ALI.
Sun B
，Bai L
，Li Q
，Sun Y
，Li M
，Wang J
，Shi X
，Zhao M
... -
《-》
Polygonatum Polysaccharide Regulates Macrophage Polarization and Improves LPS-Induced Acute Lung Injury through TLR4-MAPK/NF-κB Pathway.
To investigate the effects of polygonatum sibiricum polysaccharides (PSPs) on the polarization of macrophages to M1 and M2 phenotypes and their potential mechanism.
PSPs samples were prepared through water extraction and alcohol precipitation assay. The properties of PSPs were identified and analyzed by high-performance liquid chromatography, FT-IR, and NMR assay. Then, the effects of PSPs on mouse macrophage RAW264.7 viability were measured by CCK-8 assay. The cells were randomly divided into the control group, PSPs group, LPS group, and LPS + PSPs group. M1 phenotype polarization of RAW264.7 cells was induced by LPS treatment. The effects of various treatments on expression of M2 phenotype CD206, activation of TLR4-MAPK/NF-κB signal pathway, and translocation of NF-κB into the nucleus were determined by ELISA, western blot, and immunofluorescence assay, respectively. TLR4 inhibitor, TAK-242, and MAPK inhibitor, BIRB 796, were used to verify the effects of PSPs on the TLR4-MAPK/NF-κB pathway. The mice model of acute lung injury (ALI) was established and randomly divided into control group, PSPs group, LPS group, and LPS + PSPs group. Bronchoalveolar lavage fluid (BALF) and lung tissue were collected to measure protein, inflammatory cells, neutrophil and macrophage cells number, and the levels of IL-6 and TNF-α in BALF. Flow cytometry and western blot assay measured the phenotypic changes of macrophages and the activation of the TLR4-MAPK/NF-κB signaling pathway.
The concentrations of PSPs lower than 100 μg/mL showed no toxicity to RAW264.7 cells. PSPs treatment could significantly reverse the reduction of CD206 protein expression (P < 0.05) and the increase of the expression of inflammatory factor TNF-α, IL-1β, and IL-6 (all P < 0.05), TLR4-MAPK/NF-κB signaling pathway activation (all P < 0.05), and NF-κB translocation into the nucleus induced by LPS. The effect of inhibitors TAK-242 and BIRB 796 was consistent with that of PSPs. In the mice model of ALI, PSPs treatment could reduce the total protein levels of BALF and the number of inflammatory cells level, reverse the number changes of neutrophils and macrophages, and downregulate the proinflammatory factors IL-6 and TNF-α caused by LPS (all P < 0.05). In addition, PSPs treatment could also significantly reverse the increase in the number of iNOS expressing macrophages in alveolar lavage fluid induced by LPS (P < 0.05). In contrast, CD206-expressed cells decreased (P < 0.05). PSPs could also reverse LPS-induced TLR4-MAPK/NF-κB signal pathway protein activation (all P < 0.05).
PSPs could suppress TLR4-MAPK/NF-κB activation induced by LPS, inhibit M1 phenotypic polarization of macrophages, and promote M2 phenotypic polarization, thus playing an anti-inflammatory role.
Zhou W
，Hong J
，Liu T
，Li M
，Jin H
，Wang X
... -
《-》
ResearchGate
(全网免费下载)
钛学术
(全网免费下载)
