Effect of alcohol exposure on hepatic superoxide generation and hepcidin expression.
Effect of alcohol exposure on hepatic superoxide generation and hepcidin expression.
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DOI：
10.4331/wjbc.v4.i4.119
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发表时间：
2013-11-26
期刊：
影响因子：
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通讯作者：
Zimmerman, Matthew C
中科院分区：
文献类型：
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作者：
Harrison-Findik, Duygu Dee;Lu, Sizhao;Zimmerman, Matthew C
目的：了解线粒体产生的超氧化物（O2（-））在酒精调节肝脏铁调节激素铁调素中的作用。方法：对于酒精实验，锰超氧化物歧化酶敲除小鼠Sod 2基因表达杂合子（Sod 2（+/-））和年龄匹配的同窝对照小鼠（LMC），在两个等位基因上表达Sod 2基因，分别暴露于10%（w/v）的饮用水和普通水（对照）中7 d。用电子顺磁共振波谱法测定从小鼠肝脏分离的肝细胞中总细胞O2（-）水平。采用线粒体靶向、O2（-）敏感的荧光探针MitoSOX Red和流式细胞术检测线粒体内O2（-）。结果：与LMC小鼠相比，Sod 2（+/-）小鼠肝细胞中MnSOD蛋白（SOD 2）的表达量减少了40%。Sod 2等位基因的缺失没有改变肝中hepcidin的基础表达水平。在Sod 2（+/-）和LMC小鼠中，10%乙醇暴露1周抑制肝hepcidin mRNA表达3倍。通过电子顺磁共振光谱法观察到，未处理的Sod 2（+/-）小鼠肝细胞中的O2（-）水平比未处理的LMC小鼠高3倍。通过MitoSOX Red荧光和流式细胞术测量，Sod 2（+/）小鼠线粒体中的O2（-）水平比未处理的LMC小鼠线粒体中的O2（-）水平高4倍。与相应的未处理小鼠相比，酒精诱导LMC小鼠肝细胞中O2（-）水平比Sod 2（+/-）小鼠高两倍。与此相反，1周的酒精暴露并没有改变线粒体O2（-）水平在Sod 2（+/-）和controlmice.CONCLUSION：线粒体O2（-）是不参与抑制肝hepcidin转录，从而调节铁代谢的酒精。这些发现还表明，短期饮酒会显著升高肝细胞中的O2（-）水平，而O2（-）水平似乎并非源自线粒体。
AIM: To understand the role of mitochondrial-produced superoxide (O2 (-)) in the regulation of iron-regulatory hormone, hepcidin by alcohol in the liver.METHODS: For alcohol experiments, manganese superoxide dismutase knockout mice heterozygous for Sod2 gene expression (Sod2 (+/-)) and age-matched littermate control mice (LMC), expressing Sod2 gene on both alleles, were exposed to either 10% (w/v) ethanol in the drinking water or plain water (control) for 7 d. Total cellular O2 (-) levels in hepatocytes isolated from the livers of mice were measured by electron paramagnetic resonance spectroscopy. The mitochondrial-targeted, O2 (-)-sensitive fluorogenic probe, MitoSOX Red and flow cytometry were utilized to measure O2 (-) in mitochondria. Gene and protein expression were determined by Taqman Real-time quantitative PCR and Western blotting, respectively.RESULTS: Sod2 (+/-) mice expressed 40% less MnSOD protein (SOD2) in hepatocytes compared to LMC mice. The deletion of Sod2 allele did not alter the basal expression level of hepcidin in the liver. 10% ethanol exposure for 1 wk inhibited hepatic hepcidin mRNA expression three-fold both in Sod2 (+/-) and LMC mice. O2 (-) levels in hepatocytes of untreated Sod2 (+/-) mice were three-fold higher than in untreated LMC mice, as observed by electron paramagnetic resonance spectroscopy. O2 (-) levels in mitochondria of Sod2 (+/) mice were four-fold higher than in mitochondria of untreated LMC mice, as measured by MitoSOX Red fluorescence and flow cytometry. Alcohol induced a two-fold higher increase in O2 (-) levels in hepatocytes of LMC mice than in Sod2 (+/-) mice compared to respective untreated counterparts. In contrast, 1 wk alcohol exposure did not alter mitochondrial O2 (-) levels in both Sod2 (+/-) and control mice.CONCLUSION: Mitochondrial O2 (-) is not involved in the inhibition of liver hepcidin transcription and thereby regulation of iron metabolism by alcohol. These findings also suggest that short-term alcohol consumption significantly elevates O2 (-) levels in hepatocytes, which appears not to originate from mitochondria.