Ferroptosis: Mechanisms in Disease and Kit Selection

Science Note

[Nov. 26, 2024] 

The Impact of Fatty Acid Types on Ferroptosis Sensitivity

Recent research shows that monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs) play distinct roles in lipid metabolism and ferroptosis, a form of regulated cell death driven by lipid peroxidation. Here are some of the papers that highlight the mechanisms and factors by which MUFAs and PUFAs modulate ferroptosis for cellular homeostasis and disease implications.

Monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs) play distinct roles in lipid metabolism and ferroptosis, a form of regulated cell death driven by lipid peroxidation. PUFAs, with their multiple double bonds, are highly susceptible to peroxidation, making them key drivers of ferroptosis. In contrast, MUFAs, with their single double bond, are less susceptible to peroxidation and may counteract ferroptosis by reducing the availability of oxidizable lipids. This dynamic highlights the opposing roles of MUFAs and PUFAs in modulating ferroptosis, which has implications for cancer therapy and other diseases involving oxidative stress.

Breast cancer secretes anti-ferroptotic MUFAs and depends on selenoprotein synthesis for metastasis
Click here for the original article: Tobias Ackermann, et. al., EMBO Mol. Med., 2024.

Phospholipids with two polyunsaturated fatty acyl tails promote ferroptosis
Click here for the original article: Baiyu Qiu et. al., Cell, 2024.

Polyunsaturated fatty acids-induced ferroptosis suppresses pancreatic cancer growth
Click here for the original article: Akane Suda et. al., Cancer Discovery, 2024.

Point of Interest
- Ferroptosis, driven by iron-dependent lipid peroxidation, is counteracted by GPX4 and inhibited by monounsaturated fatty acid (MUFA) secretion and stearoyl-CoA desaturase (SCD) expression.

- Low SCD expression at low cell density increases the susceptibility of triple-negative breast cancer (TNBC) cells to ferroptosis, which is associated with reduced anti-ferroptotic MUFA secretion.

- Inhibition of Sec-tRNAsec biosynthesis, an essential step for selenoprotein production, induces ferroptosis, which impairs TNBC cell metastasis by disrupting selenoprotein production and MUFA protection.

Point of Interest

- Diacyl polyunsaturated fatty acid phosphatidylcholines (PC-PUFA2s) drive ferroptosis by generating mitochondrial ROS, initiating lipid peroxidation and influencing cancer cell sensitivity.

- Depletion of PC-PUFA2s in aging and disease is associated with ferroptosis, with mitochondrial antioxidants providing protection.

- PC-PUFA2s regulate mitochondrial homeostasis and ferroptosis, providing potential diagnostic and therapeutic targets for ferroptosis modulation.

Point of Interest

- Linoleic acid (LA) and α-linolenic acid (αLA), one of the PUFA, induce ferroptosis in pancreatic cancer cells, reducing tumour growth and improving survival in mice.

- PUFA-induced ferroptosis varies by type; Arachidonic acid and Eicosapentaenoic acid (EPA) show strong effects on pancreatic cancer, while Docosahexaenoic acid mildly suppresses proliferation.

- EPA induced ferroptosis in colorectal adenocarcinoma, while LA and αLA did not show similar effects in this context.

- PUFAs show potential as anti-cancer agents via ferroptosis, effective even in gemcitabine-resistant pancreatic cancer cells.
Related Techniques
Intracellular / mitochondrial ferrous ion (Fe2+) detection FerroOrange(intracellular), Mito-FerroGreen(mitochondrial)
Lipid peroxidation detection Liperfluo(intracellular), MitoPeDPP(mitochondrial)
Lipid Peroxidation Assay Lipid Peroxidation Probe -BDP 581/591 C11-
Total ROS detection Highly sensitive DCFH-DA or Photo-oxidation Resistant DCFH-DA
Mitochondrial superoxide detection MitoBright ROS Deep Red - Mitochondrial Superoxide Detection
Glutathione Quantification GSSG/GSH Quantification Kit
Cystine Uptake detection Cystine Uptake Assay Kit
MDA detection MDA Assay Kit
Cellular senescence detection SPiDER-βGal for live-cell imaging or flow cytometry / microplate reader / tissue samples
NEW SPiDER-βGal Blue for fixed cell and for multiple staining with immunostaining and other methods
Apoptosis detection in multiple samples Annexin V Apoptosis Plate Assay Kit
Glycolysis/Oxidative phosphorylation Assay Glycolysis/OXPHOS Assay Kit 
Cell proliferation/ cytotoxicity assay Cell Counting Kit-8 and Cytotoxicity LDH Assay Kit-WST
Related Applications

Erastin-Induced Ferroptosis: Evaluating Intracellular Uptake and Redox Balance

We investigated the transition of cellular metabolisms in A549 cells treated with erastin, a known ferroptosis inducer. Our results revealed the following.

- The inhibition of cystine uptake by erastin led to a depletion of cysteine, which in turn increased the compensatory uptake of other amino acids.
- Glucose uptake, which typically promotes ferroptosis*, was found to decrease upon erastin treatment, suggesting a potential cellular self-defense mechanism.
- The depletion of cysteine resulted in a decrease in glutathione levels and an increase in Fe2+, ROS, and lipid peroxides, all of which are recognized markers of ferroptosis.


  Cell Line: A549
  Incubation Conditions: 100 μmol/l Erastin/MEM, 37℃, 3h
  *Reference: Xinxin Song, et al., Cell Reports, (2021)

Products in Use

① Amino Acid Uptake : Amino Acid Uptake Assay Kit
② Glucose Uptake : Glucose Uptake Assay Kit-Green
③ Cystine Uptake : Cystine Uptake Assay Kit
④ Intracellular glutathione : GSSG/GSH Quantification Kit
⑤ Intracellular labile Fe : FerroOrange
⑥ Intracellular total ROS : ROS Assay Kit -Highly Sensitive DCFH-DA-
⑦ Lipid Peroxides : Liperfluo

Previous Science Note

- Exploring Ferroptosis Resistance Mechanisms for Cancer Treatment  [Oct. 29, 2024] 
- Diseases Associated with Ferroptosis: Cancer, Neurodegeneration, and Age-Related Diseases  [Jul. 9, 2024] 
- Protein Upregulation to Protect Against Ferroptosis [Jun. 4, 2024]
- Cancer Ferroptosis: Relationship on Metabolism, Lysosome, and Mitochondria [Mar. 12, 2024]
- Ferroptosis Activation, Inhibition, and Sensitization [Feb. 13, 2024]
- Ferroptosis Induced by FSP1-dependent Phase Separation [July 25, 2023]
- Age-related Microglial Phenotype Characterized by Lipid and Iron Contents [July 5, 2023]
- Link Lysosomal Failure to Ferroptosis in Human Neurons [May 9, 2023]
- Mechanisms and role of ferroptosis in disease [Jan. 17, 2023]

 

 

 

What is Ferroptosis?

“Ferroptosis” was coined by Stockwell et al. at Columbia University in 2012 and described as a form of iron-dependent cell death. * It was reported to be a form of programmed cell death by the Nomenclature Committee on Cell Death (NCCD) in 2018.
Ferroptosis is a form of programmed cell death caused by iron ion-dependent accumulation of lipid peroxides. Ferroptosis has been shown to follow a different cell death pathway from apoptosis and thus is attracting attention as a new target for cancer therapy. It has also been found to be associated with various diseases, such as neurodegenerative diseases, cerebral apoplexy, and hepatitis (NASH).

*S. J. Dixon, B. R. Stockwell, et al., Ferroptosis: an iron-dependent form of nonapoptotic cell death., Cell, 2012, 149(5), 1060.
 

How Does Ferroptosis Cause Cell Death?

Open / Close the Article

Ferroptosis is characterized by the accumulation of lipid peroxides. Lipid peroxides are formed from oxidation of polyunsaturated fatty acids (PUFA) in membrane phospholipids, with iron suggested to be involved. Intracellular glutathione peroxidase 4 (GPX4) uses reduced glutathione (GSH), an antioxidant, to reduce lipid peroxides generated by reactive oxygen species (ROS).*
However, when lipid peroxides accumulate due to GPX4 disruption or GSH depletion, ferroptosis is triggered.

*Stockwell et al, a leading researcher in the field of ferroptosis, summarized inhibitors, inducers, and detection indicators of ferroptosis in the following review, in which Dojindo’s Liperfluo is introduced for detection of lipid peroxides.

B. R. Stockwell, et al., "Ferroptosis: A Regulated Cell Death Nexus Linking Metabolism, Redox Biology, and Disease.", Cell, 2017, 171, 273.

 

Research on Related Diseases

Open / Close the Article
Nonalcoholic steatohepatitis (NASH)

Suppression of hepatitis via ferroptosis

In a study involving the livers of NASH model mice, it was confirmed that necrosis precedes apoptosis in the development of fatty liver. Further experiments showed that ferroptosis is involved within necrosis as a trigger for steatohepatitis and that inhibition of ferroptosis almost completely suppressed the onset of hepatitis.

Minoru Tanaka, et al., "Minoru Tanaka, et al., “Hepatic ferroptosis plays an important role as the trigger for initiating inflammation in nonalcoholic steatohepatitis”, Cell Death & Disease, 2019, 10, 449.

Related article: changes in intracellular markers associated with NASH


The article summarizes reports on changes in each indicator of metabolic states and cellular senescence using the NASH model.

(Click on the “NASH” tab in the link)

 

Experimental example: measurement of intracellular metabolism in NASH model tissue

Measurement of ATP, a-KG, and NAD levels in liver tissue of high-fat diet-treated type 1 diabetic model mice. (Please refer to each product’s website for more information, “Experimental Example: Change in Metabolism in Liver Tissue of NASH-Induced Mouse”)
Neurodegenerative disease

Confirmation of the link between lysosomal disorders and ferroptosis

In experiments using human neurons, it is reported that knockdown of the lysosomal protein prosaposin induces formation of lipofuscin, a hallmark of aging. This process involves the iron-catalyzed generation of reactive oxygen species, leading to induction of ferroptosis.

Martin Kampmann, et al., "Genome-wide CRISPRi/a screens in human neurons link lysosomal failure to ferroptosis", Nature Neuroscience, 2021, 24, 1020
Cancer

Regulation of cancer immunity via ferroptosis

CD8+ T cells activated by immunotherapy were found to confer an anti-tumor effect by promoting lipid peroxidation and inducing ferroptosis. The mechanism of immunotherapy-induced inhibition of cystine uptake and promotion of lipid peroxidation in tumor cells is discussed.

Weiping Zou, et al, "CD8+ T cells regulate tumour ferroptosis during cancer immunotherapy", Nature, 2019, 569, 270
Ferroptosis

Ferroptosis – a newly identified, iron-dependent form of programmed cell death

A summary of the current progress in studying ferroptosis, as well as its potential applications in the fields of biology and medicine.

Fudi Wang, et al., “Ferroptosis: Beauty or the Beast“, Dojin News, 2021, 178, 1

 


Ferroptosis-Related Reagent Selection Guide

Lipid Peroxide and Iron (Fe2+) Detection Reagents

Name Liperfluo MitoPeDPP Mito-FerroGreen FerroOrange
Target Lipid Peroxidation Lipid Peroxidation Ferrous Ion(Fe2+) Ferrous Ion(Fe2+)
Localization Intracellular Mitochondria Mitochondria Intracellular
Detection
(Fluorescence:Ex/Em)		 Fluorescence
(524 nm/535 nm)		 Fluorescence
(452 nm/470 nm)		 Fluorescence
(505 nm/580 nm)		 Fluorescence
(543 nm/580 nm)
Instrument Fluorescence Microscope,
FCM		 Fluorescence Microscope,
FCM		 Fluorescence Microscope,
Microplate Reader		 Fluorescence Microscope
Sample Live Cell Live Cell Live Cell Live Cell

Oxidative Stress- and Metabolism-Related Reagents and Kits

Name ROS Assay Kit
-Highly Sensitive DCFH-DA-		 GSSG/GSH Quantification Kit Glutamine Assay Kit-WST Glutamate Assay Kit-WST
Target ROS (Reactive oxygen species) Glutathione (oxidized/reduced) Glutamine Glutamine
Localization Intracellular Intracellular Intracellular/Extracellular Intracellular/Extracellular
Detection
(Fluorescence:Ex/Em)		 Fluorescence
(505 nm/525 nm)		 Colorimetric:412 nm Colorimetric:450 nm Colorimetric:450 nm
Instrument Fluorescence Microscope,
FCM,
Microplate Reader		 Microplate Reader Microplate Reader Microplate Reader
Sample Live Cell Cell, Tissue, Blood Plasma, Red Blood Cell Cell, Cell Culture Cell, Cell Culture

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Product Classification

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