| 异功散调节巨噬细胞铁代谢的机制研究 |
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| 引用本文： | 姜一陵,郑秦,季玉婷,薛城,张爱萍,石岭,吴志豪,罗梅宏. 异功散调节巨噬细胞铁代谢的机制研究[J]. 上海中医药大学学报, 2019, 33(3): 53-60, 67 |
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| 作者姓名： | 姜一陵 郑秦 季玉婷 薛城 张爱萍 石岭 吴志豪 罗梅宏 |
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| 作者单位： | 上海市宝山区中西医结合医院上海中医药大学附属曙光医院宝山分院 |
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| 基金项目： | 国家自然科学基金面上项目(81774271)；上海市科委引导类项目(17407931500)；上海市自然科学基金项目(16ZR1425400)；上海市卫计委科研项目(20174Y0226)；上海市宝山区中西医结合医院国自然培育项目(GZRPYJJ-201802) |
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| 摘 要： | 目的:研究异功散调节慢性病贫血(ACD)铁代谢的作用机制。方法:①体外培养小鼠巨噬细胞系RAW 264.7细胞,分为空白组和异功散干预组(0.01、0.1、1、1.25、2、2.5、3.75 mg/ml),各组给予相应浓度干预,孵育24 h、48 h和72 h后CCK8检测细胞活性,筛选有效浓度。②取RAW 264.7细胞分为空白组、脂多糖(LPS)组、异功散低剂量(0.1 mg/ml)组、异功散高剂量(1.0 mg/ml)组、LPS+异功散低剂量(0.1 mg/ml)组和LPS+异功散高剂量(1.0 mg/ml)组。预先配置含不同浓度异功散的DMEM培养基,对应加入各药物干预组。预处理1 h后再加入0.1μg/ml LPS刺激。细胞培养24 h、48 h后,比色法检测细胞内外铁含量,PCR检测细胞HAMP和膜铁转运蛋白(FPN)的基因表达水平,Western blot检测细胞信号转导转录激活因子3(STAT3)和磷酸化STAT3(p-STAT3)的蛋白表达水平。③取人肝癌HepG2细胞,在上述分组基础上增加抑制剂+LPS组、LPS+抑制剂+异功散低剂量(0.1 mg/ml)组和LPS+抑制剂+异功散高剂量(1.0 mg/ml)组。预先配置含不同浓度异功散的DMEM培养基,对应加入各药物干预组。预处理1 h后,各抑制剂干预组加入STAT3阻断剂(10μmol)处理10 min,再加入0.1μg/ml LPS刺激。细胞培养48 h后,PCR检测细胞HAMP基因表达水平。结果:①0.01～3.75 mg/ml异功散干预RAW 264.7细胞48 h内无细胞毒性作用,且能促进细胞增殖。选取0.1 mg/ml和1.0 mg/ml为有效浓度。②无LPS刺激状态下,细胞培养24 h、48 h后,异功散低、高剂量组的细胞外铁含量较空白组显著降低(P0.05),但细胞内铁含量无明显变化。LPS刺激状态下,细胞培养24 h、48 h后,LPS组细胞外铁含量较空白组显著降低(P0.05),而细胞内铁含量明显升高(P0.05);与LPS组比较,LPS+异功散低剂量组细胞外铁含量无明显变化,细胞内铁含量降低(P0.05),LPS+异功散高剂量组细胞外铁含量升高(P0.05),细胞内含量铁降低(P0.05)。③无LPS刺激状态下,与空白组相比,异功散低、高剂量组细胞培养24 h、48 h后HAMP mRNA表达无明显变化,异功散高剂量组细胞培养48 h后FPN mRNA表达量显著升高(P0.01),且高剂量组细胞FPN mRNA表达水平较低剂量组显著升高(P0.01)。LPS刺激状态下,细胞培养24 h、48 h后,与空白组相比,LPS组细胞HAMP mRNA表达显著增多(P0.05),FPN mRNA表达明显降低(P0.05,P0.01);与LPS组相比,LPS+异功散高剂量组细胞HAMP mRNA表达显著降低(P0.05),FPN mRNA表达显著升高(P0.05,P0.01);且LPS+异功散高剂量组细胞FPN mRNA表达水平较LPS+异功散低剂量组显著升高(P0.05,P0.01)。④无LPS刺激状态下,细胞培养24 h、48 h后,与空白组相比,异功散低、高剂量组细胞STAT3和p-STAT3蛋白表达无明显变化。LPS刺激状态下,细胞培养24 h、48 h后,与空白组相比,LPS组细胞STAT3蛋白表达无明显变化,p-STAT3蛋白表达有上调趋势;与LPS组相比,LPS+异功散低剂量组和LPS+异功散高剂量组细胞p-STAT3蛋白表达有降低趋势,STAT3蛋白表达无明显变化。⑤LPS刺激状态下,细胞培养48 h后,与LPS组相比,LPS+抑制剂组细胞HAMP mRNA表达显著降低(P0.01);与LPS+抑制剂组比较,LPS+抑制剂+异功散高剂量组细胞HAMP mRNA表达显著下调(P0.05)。结论:异功散可通过降低STAT3磷酸化水平,抑制HAMP mRNA过表达,上调FPN mRNA水平,从而促进巨噬细胞内铁的外流,改善LPS诱导的RAW 264.7细胞铁代谢异常,且异功散与STAT3阻断剂在调控HAMP表达方面具有协同作用。
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| 关 键 词： | 异功散 铁代谢 巨噬细胞 HAMP FPN STAT3 |
| 收稿时间： | 2019-02-22 |
| 修稿时间： | 2019-03-12 |
Mechanism of Yigong Powder on regulation of iron metabolism in macrophages |
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| Affiliation: | Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China,Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China and Shanghai Baoshan District Hospital of Integrated Traditional Chinese and Western Medicine, Baoshan Branch of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 201999, China |
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| Abstract: | |
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| Keywords: | Yigong Powder iron metabolism macrophages HAMP FPN STAT3 |
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