Killingsworth Zaria K, Misare Kelly R, Ryan Abigail S, Ampolini Elizabeth A, Mendenhall Tsultrim T, Engevik Melinda A, Hartman Jessica H
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC, United States.
Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC, United States.
Curr Res Toxicol. 2024 Sep 28;7:100195. doi: 10.1016/j.crtox.2024.100195. eCollection 2024.
Cytochrome P450 2E1 (CYP2E1) is a mammalian monooxygenase expressed at high levels in the liver that metabolizes low molecular weight pollutants and drugs, as well as endogenous fatty acids and ketones. Although CYP2E1 has been mainly studied in the endoplasmic reticulum (ER, microsomal fraction), it also localizes in significant amounts to the mitochondria, where it has been far less studied. We investigated the effects of CYP2E1 expression in mitochondria, endoplasmic reticulum, or both organelles in transgenic HepG2 cells exposed to free oleic and palmitic acid, including effects on cytotoxicity, lipid storage, respiration, and gene expression.
We found that HepG2 cells expressing CYP2E1 in both the ER and mitochondria have exacerbated levels of palmitic acid cytotoxicity and inhibited respiration. CYP2E1 expression did not impact lipid accumulation from fatty acid exposures, but mitochondrial CYP2E1 expression promoted lipid droplet depletion during serum starvation. In contrast to HepG2 cells, differentiated HepaRG cells express abundant CYP2E1, but they are not sensitive to palmitic acid cytotoxicity. Oleic acid exposure prompted less cytotoxicity, and CYP2E1 expression in the ER prevented an oleic-acid-induced increase in respiration. HepG2 cells exposed to mixtures of palmitic and oleic acid are protected from palmitic acid cytotoxicity. Additionally, we identified that CYP2E1 was decreased at the gene and protein level in hepatocellular carcinoma. Moreover, patients with tumors that had higher CYP2E1 expression had a better prognosis compared to patients with lower CYP2E1 expression.
This study has demonstrated that transgenic CYP2E1 subcellular localization plays an important role in sensitivity to cytotoxicity, lipid storage, and respiration in the hepatoma cell line HepG2 exposed to palmitic and oleic acid. HepaRG cells, in contrast, were insensitive to palmitic acid. This work demonstrates the clear importance of CYP2E1 in dictating lipotoxicity and differential roles for the mitochondrial and ER forms of the enzyme. Additionally, our data supports a potentially unique role for CYP2E1 in cancer cells.
There lies a role for CYP2E1 in altering lipotoxicity, and since CYP2E1 is known to be upregulated in both liver disease and hepatocellular carcinoma, it is important to better define how the role of CYP2E1 changes during disease progression.
细胞色素P450 2E1（CYP2E1）是一种在肝脏中高表达的哺乳动物单加氧酶，可代谢低分子量污染物、药物以及内源性脂肪酸和酮类。尽管CYP2E1主要在内质网（ER，微粒体部分）中进行研究，但它也大量定位于线粒体，而在线粒体中的研究相对较少。我们研究了在暴露于游离油酸和棕榈酸的转基因HepG2细胞中，CYP2E1在线粒体、内质网或这两种细胞器中的表达所产生的影响，包括对细胞毒性、脂质储存、呼吸作用和基因表达的影响。
我们发现，在内质网和线粒体中均表达CYP2E1的HepG2细胞，其棕榈酸细胞毒性水平加剧，呼吸作用受到抑制。CYP2E1的表达并未影响脂肪酸暴露引起的脂质积累，但线粒体CYP2E1的表达促进了血清饥饿期间脂滴的消耗。与HepG2细胞不同，分化的HepaRG细胞表达丰富的CYP2E1，但它们对棕榈酸细胞毒性不敏感。油酸暴露引起的细胞毒性较小，内质网中CYP2E1的表达可防止油酸诱导的呼吸作用增加。暴露于棕榈酸和油酸混合物的HepG2细胞可免受棕榈酸细胞毒性的影响。此外，我们发现CYP2E1在肝细胞癌中的基因和蛋白水平均降低。此外，与CYP2E1表达较低的患者相比，CYP2E1表达较高的肿瘤患者预后更好。
本研究表明，转基因CYP2E1的亚细胞定位在暴露于棕榈酸和油酸的肝癌细胞系HepG2中，对细胞毒性、脂质储存和呼吸作用的敏感性方面发挥着重要作用。相比之下，HepaRG细胞对棕榈酸不敏感。这项工作证明了CYP2E1在决定脂毒性方面的明确重要性，以及该酶在线粒体和内质网形式中的不同作用。此外，我们的数据支持CYP2E1在癌细胞中可能具有独特的作用。
CYP2E1在改变脂毒性方面发挥作用，并且由于已知CYP2E1在肝病和肝细胞癌中均上调，因此更好地定义CYP2E1在疾病进展过程中的作用变化非常重要。
