异甘草酸镁对成纤维细胞NF-κB信号通路活性的影响

·论著·

异甘草酸镁对成纤维细胞NF-κB信号通路活性的影响

徐建国1，刘继勇2，胡晋红2*

(1.第二军医大学基础医学部病理生理学教研室，上海 200433；2. 第二军医大学长海医院药学部，上海 200433)

[摘要]目的：研究异甘草酸镁(magnesium isoglycyrrhizinate，MgIG)对成纤维细胞核因子-kappa B(nuclear factor kappa B，NF-κB)信号通路活性的影响。方法：实验分6组，为空白对照组、TNF-α模型组、地塞米松(dexamethasone，DEX)阳性对照组和0.1、1、10 mg/ml的MgIG组。DEX组和MgIG组分别给予DEX(1 μg/ml)和MgIG(0.1、1、10 mg/ml)预处理4 h后，除对照组外，其余5组加入TNF-α(20 ng/ml)作用1.5 h。提取细胞核蛋白，以免疫印迹法检测NF-κB p65蛋白质表达。提取总RNA，用实时PCR技术检测细胞内IκBα mRNA表达。成纤维细胞经DEX和MgIG预处理4 h后加入TNF-α作用24 h(浓度同上)，用报告基因技术检测NF-κB基因表达。结果：成纤维细胞经TNF-α作用1.5 h后，细胞核内NF-κB p65蛋白质表达量明显增加，IκBα mRNA表达上调；TNF-α作用24 h后，NF-κB基因上调。DEX预处理4 h，可以明显减少TNF-α引起的核内NF-κB p65蛋白质表达和细胞内IκBα mRNA表达，明显降低与TNF-α共作用24 h后NF-κB基因表达。MgIG处理成纤维细胞4 h后，能够明显降低TNF-α作用1.5 h后核内NF-κB p65蛋白质表达量，但对IκBα mRNA表达水平无明显影响。1、10 mg/ml的MgIG能够明显降低与TNF-α共作用24 h后NF-κB基因表达水平。结论：MgIG的抗炎作用与抑制NF-κB p65转位入核和NF-κB基因表达，从而抑制NF-κB信号通路活性相关，但与DEX不同的是MgIG不能改变IκBαmRNA表达水平。

[关键词]异甘草酸镁；NF-κB信号通路；抗炎作用

[中图分类号]R285.5，R966，R967[文献标志码]A

DOI：10.5428/pcar20150107

基金项目国家自然科学基金(81373896)

作者简介徐建国(男)，博士.E-mail：chxdejia@hotmail.com

[收稿日期]2014-01-10

Effect of magnesium isoglycyrrhizinate on NF-κB signaling pathway activity in fibroblasts

XU JianGuo1，LIU JiYong2，HU JinHong2*(1. Department of Pathophysiology，Faculty of Basic Medical Sciences，Second Military Medical University，Shanghai 200433，China；2. Department of Pharmacy，Changhai Hospital，Second Military Medical University，Shanghai 200433，China)

ABSTRACT[]Objective: To investigate the effect of magnesium isoglycyrrhizinate (MgIG) on the activity of nuclear factor-kappa B (NF-κB) signaling pathway in fibroblast cells (FCs). Methods: The experiment consisted of 6 groups: the blank control group, the TNF-α model group, the dexamethasone (DEX) positive control group, and the 3 different concentrations of MgIG groups (i.e. 0.1,1 and 10 mg/ml subgroups). The DEX group and the MgIG groups were pretreated respectively with DEX (1 μg/ml ) and MgIG (0.1, 1 and 10 mg/ml) for 4 h. Then, with the exception of the control group, all the other 5 groups were treated with TNF-α(20 ng/ml) for 1.5 h. Nuclear proteins and total RNA were collected for detecting the expression levels of NF-κB p65 protein by Western blot. Total RNA was collected and used for detecting the expression levels of IκBα mRNA by real-time PCR. Following pretreatment with DEX and MgIG for 4 h, the FCs were treated with TNF-α(with the same dosage as the above) for 24 h, and then the expression levels of NF-κB were detected by reporter gene technology. Results: Following treatment with TNF-α for 1.5 h, the expression levels of intranuclear NF-κB p65 and intracellular IκBα mRNA in FCs were obviously increased, and 24-h treatment with TNF-α also increased the expression levels of NF-κB. Pretreatment with DEX for 4 h could decrease the expression levels of intranuclear NF-κB p65 and intracellular IκBα mRNA gene induced by TNF-α, and could also significantly reduce the expression levels of NF-κB following combined treatment with TNF-α for 24 h. Treatment of FCs with MgIG for 4 h could significantly lower the expression levels of intranuclear NF-κB p65 after pretreatment with TNF-α for 1.5 h. However, no significant effects could be noted on the expression levels of IκBα mRNA. MgIG with dosages of 1.0 and 10 mg/ml could significantly decrease expression levels of NF-κB following combined treatment with TNF-α for 24 h.Conclusion: The anti-inflammatory effect of MgIG was related with the inhibited NF-κB p65 nuclear transposition and NF-κB gene expression, and was also associated with the inhibited activity of NF-κB signaling pathway. When compared with DEX, MgIG failed to exert significant effect on the expression of IκBα mRNA.

[KEY WORDS]magnesium isoglycyrrhizinate; NF-κB signaling pathway; anti-inflammatory effect

[Pharm Care Res，2015，15(1): 22-26]

*通信作者(Corresponding author)：胡晋红，E-mail: hjhong2016@126.com

异甘草酸镁(magnesium isoglycyrrhizinate，MgIG)属于18α-甘草酸中药单体制剂，与具有抗氧化、抗炎、抗肿瘤和保肝活性的异甘草素[1]同属于18α-甘草酸，于2005年开始应用于慢性肝炎的治疗，具有明显的护肝和抑制肝炎发展的作用，对接触性皮炎小鼠MgIG可明显减轻炎症反应[2]，但其具体抗炎机制不详。……