| 简宇,范婷,代凌云,等.姜黄素通过上调 miR-124减轻脂多糖诱导的大鼠急性肺损伤[J].安徽医药,2022,26(8):1515-1519. |
| 姜黄素通过上调 miR-124减轻脂多糖诱导的大鼠急性肺损伤 |
| Curcumin alleviates lipopolysaccharide-induced acute lung injury in rats by up-regulating miR-124 |
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| DOI：10.3969/j.issn.1009-6469.2022.08.007 |
| 中文关键词: 急性肺损伤 姜黄素 微小 RNA-124 肿瘤坏死因子受体相关因子 6 大鼠 |
| 英文关键词: Acute lung injury Curcumin MicroRNA-124 Tumor necrosis factor receptor associated factor 6 Rats |
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| 中文摘要: |
| 目的探究姜黄素( Cur)对脂多糖( LPS)诱导的大鼠急性肺损伤( ALI)的保护作用及可能机制。方法体外培养大鼠肺泡巨噬细胞 NR8383，采用双荧光素酶报告基因实验验证微小 RNA-124(miR-124)与肿瘤坏死因子受体相关因子 6(TRAF6)的靶向关系。 SD大鼠采用随机数字表法分为对照组( NC组)、 LPS组、 LPS+Cur组、 LPS+Cur+inhibitor-NC组、 LPS+Cur+inhibitor组，每组 10只。 LPS组腹腔注射 LPS 10 mg/kg制备 ALI大鼠模型， LPS+Cur组在 ALI模型制备 30 min后尾静脉注射 Cur 200 mg/ kg，LPS+Cur+inhibitor-NC组、 LPS+Cur+inhibitor组大鼠尾静脉分别注射 miR-124 inhibitor-NC、miR-124 inhibitor 50 mg/kg 1周后腹腔注射 LPS 10 mg/kg，30 min后尾静脉注射 Cur 200 mg/kg，NC组注射等量生理盐水 30 min后尾静脉注射等量生理盐水。干预 24 h后， HE染色观察肺组织病理变化，酶联免疫吸附试验( ELISA)检测血清肿瘤坏死因子 -α(TNF-α)、白细胞介素 -6(IL-6)水平，分别采用实时荧光定量 PCR(RT-qPCR)、蛋白质印迹法( WB)检测肺组织 miR-124、TRAF6 mRNA及蛋白表达。结果双荧光素酶报告基因实验结果显示， miR-124可直接靶向 TRAF6 3 ' UTR区负调控其表达。动物实验结果表明， NC组大鼠肺组织结构清晰，肺泡结构完整，无明显异常改变； LPS组、 LPS+Cur+inhibitor组大鼠肺组织结构破坏严重，肺间质增厚，大量炎性细胞浸润，弥漫性充血、渗出，肺泡萎缩； LPS+Cur组、 LPS+Cur+inhibitor-NC组大鼠肺组织损伤减轻。与 NC组比较， LPS组大鼠血清 TNF-α、IL-6、肺组织 TRAF6 mRNA及蛋白表达水平显著增加， miR-124表达水平显著降低( P<0.05)；与 LPS组比较， LPS+Cur组大鼠血清 TNF-α、IL-6、、肺组织 TRAF6 mRNA及蛋白表达水平显著降低， miR-124表达水平显著增加( P<0.05)；与 LPS+ Cur+ inhibitor-NC组比较， LPS+ Cur+inhibitor组大鼠血清 TNF-α、IL-6、肺组织 TRAF6 mRNA及蛋白表达水平显著增加， miR-124表达水平显著降低( P<0.05)。结论 Cur可通过上调 miR-124靶向抑制 TRAF6减轻 LPS诱导的大鼠 ALI及炎症反应，发挥治疗作用。 |
| 英文摘要: |
| Objective To investigate the protective effect and its possible mechanism of Curcumin (Cur) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rats.Methods Alveolar macrophages NR8383 of rats was cultured in vitro, double luciferase reporter gene assay was used to verify the targeting relationship between microRNA-124 (miR-124) and tumor necrosis factor receptorassociated factor 6 (TRAF6). SD rats were randomly divided into control group (NC group), LPS group, LPS+Cur group, LPS+Cur+inhib-itor NC group, LPS+Cur+inhibitor group, with 10 in each group. In LPS group, 10 mg/kg LPS was injected intraperitoneally to prepareALI rat model. In LPS+Cur group, after 30 minutes of ALI model preparation, Cur 200 mg/kg was injected into tail vein, while the ratsin LPS+Cur+inhibitor NC group and LPS+Cur+inhibitor NC group were injected with miR-124 inhibitor NC and miR-124 inhibitor 50 mg/kg via caudal vein, respectively, and LPS 10 mg/kg was injected intraperitoneally 1 week later, and then Cur 200 mg/kg was inject-ed into tail vein 30 minutes later. The rats in NC group were injected with equal amount of normal saline into tail vein 30 minutes afterinjection of the same amount of normal saline. After 24 hours of intervention, HE staining was used to observe the pathological changesof lung tissue, the levels of serum tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were detected by enzyme-linked immunosor- bent assay (ELISA), and the mRNA and protein expressions of miR-124 and TRAF6 were detected by real-time fluorescence quantita- tive PCR (RT-qPCR) and Western blot (WB) respectively.Results Double luciferase reporter gene showed that miR-124 could direct- ly target TRAF63 'UTR region and negatively regulate its expression. The results of animal experiments showed that the lung tissuestructure of rats in NC group was clear, the alveolar structure was complete, and there was no obvious abnormal change; the lung tissuestructure of rats in LPS group and LPS+Cur+inhibitor group was damaged seriously, the lung interstitium was thickened, a large num-ber of inflammatory cells were infiltrated, diffuse hyperemia, exudation and alveolar atrophy were found; in addition, the lung injury inLPS+Cur group and LPS+Cur+inhibitor NC group was reduced. Compared with those in NC group, the expression levels of TNF-α, IL-6 in serum, TRAF6 mRNA and protein in lung tissue in LPS group were significantly higher, and the expression level of miR-124 was sig- nificantly lower (P<0.05); compared with those in LPS group, the expression levels of TNF-α, IL-6 in serum, TRAF6 mRNA and protein in lung tissue in LPS+Cur group were significantly lower, and the expression level of miR-124 was significantly higher (P<0.05); in ad- dition, compared with those in LPS+Cur+inhibitor NC group, the expression levels of TNF-α, IL-6 in serum, TRAF6 mRNA and proteinin lung tissue in LPS+Cur+inhibitor group were significantly higher, and the expression level of miR-124 was significantly lower (P< 0.05).Conclusion Cur can reduce the ALI and inflammatory response induced by LPS by target inhibition of TRAF6 by up-regulating miR-124, and play a therapeutic role. |
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