Human HMGB1-Lucia Reporter HT-1080 Cells

Cell line description

HT1080-HMGB1-Lucia™ cells are derived from the human fibrosarcoma cell line HT-1080, a standard cell model to study ferroptotic cell death [1-2]. HT1080-HMGB1-Lucia™ cells stably express an HMGB1::Lucia luciferase fusion protein in the cytoplasm, in which the C-terminus of the High-Mobility-Group-Protein B1 (HMGB1) is fused to the Lucia® luciferase.

HMGB1 plays a critical role in the stress response not only inside the cell as a DNA chaperone and cell death regulator but also outside the cell as a prototypical alarmin. Indeed, during non-apoptotic programmed or regulated cell death, such as pyroptosis and ferroptosis, HMGB1 is released along with other pro-inflammatory molecules.

Following ferroptosis activation, the cell membrane ruptures and HMGB1::Lucia is released in the extracellular milieu. Levels of HMGB1::Lucia in the supernatant can be readily monitored by using QUANTI-Luc™ 4 Lucia/Gaussia, a Lucia® luciferase detection reagent (see figures).

Ferroptosis overview

In 2012, Dixon et al. [1] discovered a novel form of non-apoptotic cell death characterized by iron overload and accumulation of lethal lipid peroxidation [3]. Because iron chelators were shown to block this type of regulated cell death, it was originally defined as iron-dependent and was called "Ferroptosis" [1]. In contrast to apoptosis, necrosis, and autophagy, ferroptosis is characterized by the build-up of lipid reactive oxygen species (ROS) triggering membrane damage. Cells undergoing ferroptosis do not exhibit classic apoptosis-like features, such as chromatin condensation or membrane blebbing. Instead, mitochondrial shrinkage and increased membrane density are often observed [1-3]. 

Ferroptosis is regulated by a complex network involving iron homeostasis, lipid metabolism, and glutathione-dependent oxidative-reductive balance [4-5]. Iron metabolism plays a central role, as excessive intracellular iron promotes the Fenton reaction, generating lethal amounts of ROS that drive lipid peroxidation [5]. Moreover, the enzyme glutathione peroxidase 4 (GPX4) is a key regulator that protects cells from ferroptosis by reducing lipid peroxides. GPX4 activity is dependent on Glutathione (GSH) which is synthesized using cystine imported via the cystine-glutamate antiporter, System Xc−. When System Xc− is inhibited (e.g. by Erastin), cystine uptake decreases, leading to GSH depletion and GPX4 inactivation. As a result, lipid peroxides accumulate in the cells ultimately leading to ferroptotic cell death [3-5]. 

Ferroptosis in disease

Ferroptosis is involved in various diseases, including neurodegenerative disorders, organ injury-related conditions, and cancer [3-4]. In neurodegenerative diseases, such as Parkinson's and Alzheimer's, ferroptosis contributes to neuronal loss through iron deposition in the brain and oxidative stress [3]. It has also been identified as a major pathogenic driver in other organ injury-related disorders including acute kidney injury and COVID-19-induced myocarditis [4]. Studies have shown that using ferroptosis-specific inhibitors can protect against severe tissue damage in these cases [5]. On the other hand, triggering ferroptosis is being explored as a potential therapeutic approach for treating therapy-resistant cancers [1, 3-5]. Certain types of cancer, especially those with high iron levels or deficiencies in GPX4, are particularly sensitive to ferroptosis-inducing substances. Compounds like RSL3 or Erastin hold promise to overcome drug resistance in chemotherapy and immunotherapy [3]. Advancing our understanding of ferroptosis will continue to drive progress in cell death research and therapeutic development.

References

1. Dixon SJ, et al., 2012. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell. 2012 May 25;149(5):1060-72.
2. Zheng J, et al. 2021. Sorafenib fails to trigger ferroptosis across a wide range of cancer cell lines. Cell Death Dis. 2021 Jul 13;12(7):698.
3. Du Y, Guo Z. 2022. Recent progress in ferroptosis: inducers and inhibitors. Cell Death Discov. 8(1):501.
4. Sun S, et al., 2023. Targeting ferroptosis opens new avenues for the development of novel therapeutics. Signal Transduct Target Ther. 8(1):372.
5. Li J, et al., 2020. Ferroptosis: past, present and future. Cell Death Dis. 11(2):88.