Caro Andres A, Barrett Daniel, Garcia Cristobal, Northington Weston, Pinkney Jamya, Shuja Rayan, Stovall Hannah
Hendrix College.
Res Sq. 2023 Mar 22:rs.3.rs-2702878. doi: 10.21203/rs.3.rs-2702878/v1.
Ferroptosis is a recently described form of regulated cell death initiated by the iron-mediated one-electron reduction of lipid hydroperoxides (LOOH). Cytochrome P450 2E1 (CYP2E1) induction, a consequence of genetic polymorphisms or/and gene induction by xenobiotics, may promote ferroptosis by contributing to the cellular pool of LOOH. However, CYP2E1 induction also increases the transcription of anti-ferroptotic genes that regulate the activity of glutathione peroxidase 4 (GPX4), the main ferroptosis inhibitor. Based on the above, we hypothesize that the impact of CYP2E1 induction on ferroptosis depends on the balance between pro- and anti-ferroptotic pathways triggered by CYP2E1. To test our hypothesis, ferroptosis was induced with class 2 inducers (RSL-3 or ML-162) in mammalian COS-7 cancer cells that don't express CYP2E1 (Mock cells), and in cells engineered to express human CYP2E1 (WT cells), and the impact on viability, lipid peroxidation and GPX4 was assessed. CYP2E1 overexpression protected COS-7 cancer cells against ferroptosis, evidenced by an increase in the IC and a decrease in lipid ROS in WT versus Mock cells after exposure to class 2 inducers. CYP2E1 overexpression produced an 80% increase in the levels of the GPX4 substrate glutathione (GSH). Increasing GSH in Mock cells protected cells against ferroptosis by ML-162. Depleting GSH, or inhibiting Nrf2 in WT cells reverted the protective effect mediated by CYP2E1, causing a decrease in the IC and an increase in lipid ROS after exposure to ML-162. These results show that CYP2E1 overexpression protects COS-7 cancer cells against ferroptosis, an effect probably mediated by Nrf2-dependent GSH induction.
铁死亡是一种最近被描述的受调控的细胞死亡形式，由铁介导的脂质氢过氧化物（LOOH）单电子还原引发。细胞色素P450 2E1（CYP2E1）的诱导是遗传多态性或/和外源性物质诱导基因的结果，它可能通过增加细胞内LOOH池来促进铁死亡。然而，CYP2E1的诱导也会增加抗铁死亡基因的转录，这些基因调节主要的铁死亡抑制剂谷胱甘肽过氧化物酶4（GPX4）的活性。基于上述情况，我们假设CYP2E1诱导对铁死亡的影响取决于CYP2E1触发的促铁死亡和抗铁死亡途径之间的平衡。为了验证我们的假设，在不表达CYP2E1的哺乳动物COS-7癌细胞（Mock细胞）和经过基因工程改造以表达人CYP2E1的细胞（WT细胞）中，用2类诱导剂（RSL-3或ML-162）诱导铁死亡，并评估其对细胞活力、脂质过氧化和GPX4的影响。CYP2E1的过表达保护COS-7癌细胞免受铁死亡，这在暴露于2类诱导剂后，WT细胞与Mock细胞相比，IC增加和脂质活性氧减少中得到证明。CYP2E1的过表达使GPX4底物谷胱甘肽（GSH）水平增加了80%。在Mock细胞中增加GSH可保护细胞免受ML-162诱导的铁死亡。耗尽WT细胞中的GSH或抑制Nrf2可逆转CYP2E1介导的保护作用，导致暴露于ML-162后IC降低和脂质活性氧增加。这些结果表明，CYP2E1的过表达保护COS-7癌细胞免受铁死亡，这种作用可能是由Nrf2依赖的GSH诱导介导的。
