Mechanism of chlorogenic acid reducing lipopolysaccharide-induced acute lung injury in mice by regulating miR-223/NLRP3 axis / 中南大学学报(医学版)

OBJECTIVES@#Chlorogenic acid has various physiological activities such as antibacterial, anti-inflammatory, and antiviral activities. Studies have shown that chlorogenic acid can alleviate the inflammatory response of mice with acute lung injury (ALI), but the specific mechanism is still unclear. This study aims to investigate whether chlorogenic acid attenuates lipopolysaccharide (LPS)-induced ALI in mice by regulating the microRNA-223 (miR-223)/nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) axis.@*METHODS@#SPF grade BALBc male mice were randomly divided into a control group, a model group, a chlorogenic acid group, a chlorogenic acid+miR-223 negative control (miR-223 NC) group, and a chlorogenic acid+miR-223 inhibitor (miR-223 antagomir) group, 10 mice in each group. Except the control group, the other groups were instilled with 4 mg/kg LPS through the airway to establish the ALI mouse model. After the modeling, the mice in the chlorogenic acid group were continuously given chlorogenic acid (100 mg/kg) by gavage for 7 d. The chlorogenic acid+miR-223 NC group and the chlorogenic acid+miR-223 antagomir group were given 100 mg/kg chlorogenic acid by gavage every day, and then were injected with 10 μL of miR-223 NC (0.5 nmol/μL) and miR-223 antagomir (0.5 nmol/μL) respectively for 7 consecutive days.The control group and the model group were replaced with normal saline. The lung tissues of mice were taken to measure the ratios of lung wet to dry weight (W/D). The bronchoalveolar lavage fluid of mice was collected to measure the levels of TNF-α, IL-6, and IL-1β by ELISA kit and to count the number of eosinophils (EOS), lymphocytes, neutrophils under light microscope. After HE staining, the pathological changes of lung tissues were observed and lung injury was scored. qRT-PCR method were used to determine the expression levels of miR-223 in lung tissues. Western blotting was used to determine the expression levels of NLRP3 protein in mouse lung tissues. Luciferase reporter assay was used to analyze the targeting relationship of miR-223 to NLRP3.@*RESULTS@#Compared with the control group, the lung W/D value, the lung injury score and the level of inflammatory factors in the bronchoalveolar lavage fluid were significantly increased in the model group (all P<0.05); the infiltration of inflammatory cells in the lung tissue was severe; the alveolar space was significantly increased; the alveolar wall was significantly thickened; the number of EOS, lymphocytes, and neutrophils in the bronchoalveolar lavage fluid was significantly increased (all P<0.05); the expression levels of miR-223 in lung tissue were significantly decreased (P<0.05); and the protein expression levels of NLRP3 were significantly increased (P<0.05). Compared with the model group, the W/D value of lungs, lung injury score, and levels of inflammatory factors in bronchoalveolar lavage fluid were significantly decreased in the chlorogenic acid group, the chlorogenic acid+miR-223 NC group, and the chlorogenic acid+miR-223 antagomir group (all P<0.05); lung tissues damage was alleviated; the numbers of EOS, lymphocytes, and neutrophils in bronchoalveolar lavage fluid were significantly decreased (all P<0.05); the expression levels of miR-223 in lung tissues were significantly increased (P<0.05); and the expression levels of NLRP3 protein were significantly decreased (P<0.05). Compared with the chlorogenic acid group, the lung W/D value, lung injury score, and inflammatory factor levels in the bronchoalveolar lavage fluid were significantly increased in the chlorogenic acid+miR-223 antagomir group (all P<0.05); lung tissue damage was aggravated; the number of EOS, lymphocytes and neutrophils in bronchoalveolar lavage fluid significantly increased (all P<0.05); the expression levels of miR-223 in lung tissues were significantly decreased (P<0.05); and the expression levels of NLRP3 protein were significantly increased (P<0.05). The results of luciferase reporter assay showed that miR-223 had a targeting relationship with NLRP3.@*CONCLUSIONS@#Chlorogenic acid may increase the level of miR-223, target the inhibition of NLRP3 expression, reduce LPS-induced inflammatory response in ALI mice, and alleviate pathological damage of lung tissues.

Effects of different temperature stress on cell membrane permeability,active oxygen metabolism and accumulation of effective substances in Lonicera japonicea / 中国中药杂志

The study is aimed to explore the effects of stress at different temperatures( 35,45,55 ℃) on membrane permeability,active oxygen metabolism and accumulation of effective substances in Lonicera japonica,and provide theoretical basis for reducing deterioration and revealing browning mechanism during postharvest processing of L. japonica. The cell membrane permeability( relative conductivity,MDA content),active oxygen metabolism( SOD,POD,PPO,CAT activity) and the accumulation of effective substances( chlorogenic acid,luteolin,neochlorogenic acid,cryptochlorogenic acid,3,5-dicaffeoylquinic acid,3,4-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid) of L. japonica were all studied by constant temperature drying method,and the results were analyzed by the SPSS 17. 0 statistical software. The results showed that MDA content in L. japonica was increased by 151. 14% at 35 ℃,SOD,POD,PPO and CAT activity were 29. 73%,42. 86%,105. 02% and 10. 74% higher than at 45 ℃,respectively. The order of effective substance content in L. japonica was 35 ℃ >45 ℃ >55 ℃. The changes of membrane permeability,activity of active oxygen metabolizing enzyme and accumulation of active components were significantly affected by different temperature stress. The indexes showed that physiological and active oxygen metabolizing enzyme activity of L. japonica was the highest under 35 ℃ stress,chlorogenic acid and luteolin were effectively accumulated,which provides basic data for solving browning problem in the postharvest processing of L. japonica.

Chlorogenic Acid Decreases Retinal Vascular Hyperpermeability in Diabetic Rat Model

To evaluate the effect of chlorogenic acid (CGA), a polyphenol abundant in coffee, on retinal vascular leakage in the rat model of diabetic retinopathy, Sprague-Dawley rats were divided into four groups: controls, streptozotocin-induced diabetic rats, and diabetic rats treated with 10 and 20 mg/kg chlorogenic acid intraperitoneally daily for 14 days, respectively. Blood-retinal barrier (BRB) breakdown was evaluated using FITC-dextran. Vascular endothelial growth factor (VEGF) distribution and expression level was evaluated with immunohistochemistry and Western blot analysis. Expression of tight junction proteins, occludin and claudin-5, and zonula occludens protein, ZO-1 was also evaluated with immunohistochemistry and Western blot analysis. BRB breakdown and increased vascular leakage was found in diabetic rats, with increased VEGF expression and down-regulation of occludin, claudin-5, and ZO-1. CGA treatment effectively preserved the expression of occludin, and decreased VEGF levels, leading to less BRB breakdown and less vascular leakage. CGA may have a preventive role in BRB breakdown in diabetic retinopathy by preserving tight junction protein levels and low VEGF levels.