The Effect And Mechanism Of Resveratrol On TREM-1 Expression In Acute Lung Injury

Acute lung injury(ALI)and its further development cause acute respiratory distress syndrome(ARDS)are important pathological processes of many respiratory diseases.The outbreak of inflammatory response in the lung is one of the important factors leading to the occurrence and development of ALI/ARDS.Therefore,inhibiting the excessive inflammatory response in the early stage of ALI/ARDS can reduce the lung damage and improve the prognosis of ALI/ARDS.Myeloid cells express trigger receptor-1(TREM-1),which is widely expressed on the cell membrane of neutrophils and monocytes/macrophages.Studies have shown that the activation of TREM-1 can directly amplify the inflammatory response,and blocking the activation of TREM-1 can reduce ALI.Resveratrol(RES)is a kind of natural plant extract,which is widely available.Studies have shown that resveratrol has antiinflammatory,anti-oxidation and anti-tumor effects.Whether resveratrol can inhibit the amplification effect of inflammatory by regulating the expression and activation of TREM-1,thus alleviating ALI remains unclear.Objective:To observe the effect of resveratrol on lipopolysaccharide-induced ALI and to explore the effect and mechanism of resveratrol on the expression of inflammatory amplification receptor TREM-1 in ALI.Methods: 1.Kunming mice were studied,and randomly divided into normal group,ALI group and resveratrol +ALI group.Mice in the resveratrol +ALI group were intraperitoneal injected with resveratrol at a dose of 30 mg/kg and were given 10 mg/kg LPS after two hours to establish the animal model of ALI.The respiratory frequency,lung compliance,tidal volume and airway resistance of mice in each group were measured with a small animal respiratory function detector.The pathological changes of lung tissue were evaluated by HE staining and pathological injury scoring system,and the wet/dry weight ratio of lung tissue was determined.Myeloperoxidase(MPO)activity in lung tissue and lactate dehydrogenase(LDH)activity in bronchia alveolar lavage fluid(BALF)of mice were measured.The cells in BALF were used to classify and count,and the protein content of inflammatory factors in BALF was detected by ELISA.The mRNA expression of IL-1?,IL-18 and TREM-1 in lung tissue were detected by Real-time PCR.Immunohistochemistry was used to detect the expression of TREM-1 protein in lung tissue.2.The logarithmic growth phase of macrophages(RAW264.7 cell line)was studied.They were divided into control group,LPS group,resveratrol +LPS group,resveratrol +LPS +PD98059(ERK pathway inhibitor)group,resveratrol +LPS +BAY117082(NF-?B pathway inhibitor)group.The expression of TREM-1 mRNA in macrophages of each group was determined by Real-time PCR.Results: 1.Resveratrol alleviates ALI induced by LPS After the ALI mouse model was established,the respiratory function of the mice was detected by the instrument of respiratory function of small animals.The results showed that: after intraperitoneal injection of LPS for 6 hours,the respiratory rate,lung compliance,tidal volume decreased and airway resistance increased in mouse.The respiratory function of ALI mice induced by LPS was significantly improved by resveratrol intervention at 2 hours before LPS stimulation.HE staining and pathological injury score of lung tissue showed that alveolar hyperemia and hemorrhage,neutrophil infiltration and leukocyte aggregation,alveolar wall thickening in ALI group.Resveratrol could alleviated the pathological injury of lung.Lung wet/dry weight ratio and MPO activity increased in the ALI group compared to the normal group,while the wet/dry weight ratio and MPO activity decreased in the resveratrol +ALI group compared to the ALI group.The content of total protein and the activity of LDH in ALI group were higher than those in normal group,while resveratrol +ALI group decreased in varying degrees compared with ALI group.Total cell number and neutrophil number were significantly increased in the ALI group,while were significantly decreased in the resveratrol +ALI group.Compared with the normal group,the expression of IL-1? and IL-18 protein in the ALI group was increased,and the expression of IL-1? and IL-18 protein in the resveratrol +ALI group was lower than that in the ALI group.2.Resveratrol alleviates lung inflammation in ALI by regulating TREM-12.1 Resveratrol reduces the expression of TREM-1 mRNA and protein in the lungs of mice with ALI The representation of TREM-1 mRNA in the lungs of mice were tested by the Realtime PCR after intraperitoneal LPS injection in 6 hours.We found that TREM-1 mRNA expression of the ALI group has higher level degree than the normal group.On the hand,it was distinctly lower than the ALI group contrasted to the TREM-1 mRNA representation of resveratrol +ALI group.The representation of TREM-1 protein in the lungs of mice was tested by the approach of immunohistochemistry,we found that TREM-1 protein expression of the ALI group has the higher level degree than the normal group.On the hand,it was lower than the ALI group contrasted to the TREM-1 protein representation of resveratrol +ALI group in lung.2.2 Resveratrol reduces the expression of TREM-1 mRNA in macrophages of LPSstressed mice It was revealed that the expression of TREM-1 mRNA in macrophages of LPS group was higher than the control group from the effect of Real-time PCR.Nevertheless,TREM-1 mRNA representation of resveratrol +LPS group appeared lower than the LPS group.The macrophages of LPS-stressed mice were pretreated with different concentrations of resveratrol(0.001,0.01,0.1,1 ?M/L),we found that the representation of TREM-1 mRNA in mice macrophages decreased steadily by increasing of resveratrol concentration.The TREM-1 mRNA representation of mice macrophages reached the lowest degree as the concentration approached 1 ?M/L.After pretreated with RES for 2 hours and carried LPS stimulation,we tested the TREM-1 mRNA representation at different time(0,3,6,12,24 h).The results showed that the expression of TREM-1 mRNA was significantly decreased at 6 h and reached the lowest degree at 24 h.3.Resveratrol inhibits the expression and activation of TREM-1 via ERK and NF-?B pathway The mechanism of TREM-1 regulated by resveratrol was detected by Real-time PCR.The results showed that compared with resveratrol +LPS group,the mRNA expression of TREM-1 in mice macrophages in resveratrol +LPS+PD98059 group and resveratrol +LPS+BAY11-7082 group was increased.Conclusion: 1.Resveratrol can alleviate ALI induced by LPS2.ALI could up-regulate the expression of TREM-1 in the lung of mice,while resveratrol could decrease the expression of TREM-1.3.LPS up-regulated the expression of TREM-1 in mouse macrophages,while resveratrol down-regulated the expression of TREM-1 by inhibiting the activation of ERK and NF-?B.