Validation of control genes and a standardised protocol for quantifying gene expression in the livers of C57BL/6 and ApoE-/- mice.
Validation of control genes and a standardised protocol for quantifying gene expression in the livers of C57BL/6 and ApoE-/- mice.
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DOI：
10.1038/s41598-018-26431-3
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发表时间：
2018-05-24
影响因子：
4.6
通讯作者：
Kroon PA
中科院分区：
文献类型：
--
作者：
Day PEL;Chambers KF;Winterbone MS;García-Blanco T;Vauzour D;Kroon PA
肝脏在食物和药物代谢和解毒中起着关键作用，因此影响健康和疾病中的全身体内平衡。虽然C57 BL/6和ApoE−/−小鼠模型被广泛用于研究肝脏疾病和代谢中的基因表达变化，但目前在这些模型中没有经过验证的稳定表达的内源性基因，也不知道基因表达在肝叶内和跨肝叶如何变化。在这里，我们显示了跨肝叶的Ywhaz、Gak、Gapdh、Hmbs和Act-β内源性基因表达的区域差异;使用来自6只C57 BL/6小鼠的四个肝叶的同质样品，我们测试了12个内源基因的稳定性，并显示Act-β和Eif 2-β在小鼠中的表达。α是所有四个叶中表达最稳定的内源性基因，并证明Abca 1胆固醇流出基因表达的小叶差异。这些结果表明，从一个指定的均匀的粉末状肝叶采样是至关重要的，在提高数据的可靠性和再现性。使用来自20只ApoE−/−小鼠的左肝叶的同质样品进一步测试了12种内源性基因的稳定性，这些小鼠接受标准或高多酚饮食。Act-β和Ywhaz是用于该小鼠模型中基因表达正常化的合适内源基因。
The liver plays a critical role in food and drug metabolism and detoxification and accordingly influences systemic body homeostasis in health and disease. While the C57BL/6 and ApoE−/− mouse models are widely used to study gene expression changes in liver disease and metabolism, currently there are no validated stably expressed endogenous genes in these models, neither is it known how gene expression varies within and across liver lobes. Here we show regional variations in the expression of Ywhaz, Gak, Gapdh, Hmbs and Act-β endogenous genes across a liver lobe; Using homogeneous samples from the four liver lobes of 6 C57BL/6 mice we tested the stability of 12 endogenous genes and show that Act-β and Eif2-α are the most stably expressed endogenous genes in all four lobes and demonstrate lobular differences in the expression of Abca1 cholesterol efflux gene. These results suggest that sampling from a specified homogeneous powdered liver lobe is paramount in enhancing data reliability and reproducibility. The stability of the 12 endogenous genes was further tested using homogeneous samples of left liver lobes from 20 ApoE−/− mice on standard or high polyphenol diets. Act-β and Ywhaz are suitable endogenous genes for gene expression normalisation in this mouse model.
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