I know that there exists a direct relationship between lipid peroxidation and SOD, Cat, GPX etc., But there is lot of ambiguity when it comes to working with a particular cell line. Please post your inputs here. Thank you.
You are aware that toxic oxygen species [i.e. reactive oxygen species] are invariably generated (especially under stress or due to unusual flow of electrons) in all living cells. Generally, superoxide anion radicals are first to be formed through unusual flow of electron to oxygen. Subsequently, these superoxide anion radicals generate hydrogen peroxide either enzymatically (i.e. through SOD) or non-enzymatically. Hydrogen peroxide (especially through Fenton reaction) generates hydroxyl radicals.
It is hydroxyl radicals that initiate lipid peroxidation. In fact the extent of lipid peroxidation (which is often quantified by measuring malondialdehyde) gives an indication of level of TOS/ROS. As the level of ROS increases under stress, lipid peroxidation (or MDA level) is taken as an indicator/marker of degree of stress.
As far as SOD, catalase, ascorbate peroxidase, glutathione peroxidase, guaicol peroxidase etc. are concerned, their activity increases depending on the level of concerned ROS and place of production. In fact, many of these ROS regulate expression of concerned genes.
Only correlation, I can expect between lipid peroxidation and the concerned enzymes is positive (?). Higher the ROS higher would be lipid peroxidation as well as activities of ROS scavenging/detoxifying enzymes.
Many a times we have problems with expected results (things may not work as per our expectation). One needs to ensure that protocols are followed carefully. If you let me know the actual problem you are facing, I might be in a position to further address your query.
I agree that the hydroxyl radical can initiate unspecific lipid peroxidation. In general, however, lipid peroxidation is achieved by receptor-mediated activation of lipoxygenases. The most important enzyme reducing hydroperoxides of complex lipids is GPx4. Peroxiredoxins are also reported to act on complex lipid peroxides. All the other 'antioxidant enzymes' only affect lipid peroxidation indirectly by removing superoxide, H2O2 or peroxynitrite, which together may initiate unspecific lipid peroxidation via hydroxyl radical formation. Yet as mentioned, this is not an important phenomenon except in severe intoxication by redox cyclers. Read the articles attached and go to PubMed to screen for recent publications by Marcus Conrad on apoptosis and ferroptosis.
The biological effects of free radicals are controlled in human body by a broad range of antioxidants, such as vitamins E, C, carotenoids, GSH, minerals such as zinc, selenium, copper, manganese, and iron, as well as proteins that either catalytically neutralized oxidants, such as SOD, CAT, and GPx, or scavenge oxidants in a sacrificial manner (i.e. replace a highly reactive free radical by a less reactive one), such as albumin, or they sequester transition metals in a way that prevents the metal ion from participating in free radical reactions, such as transferrin, ceruloplasmin, and albumin.
The change in the levels of oxidants and antioxidants may be respected as a biochemical marker of free radical production, which in turn, beneficial to measure the oxidative stress concomitant with diseases.
The most frequently marker used for lipids damage or lipids peroxidation, result from reaction with free radicals is the thiobarbituric acid reactive species in the mean of malondialdehyde (MDA). Protein carbonylation can be used as a marker of radical damage to protein. The most commonly used methods for assessing radical-modified DNA is to measure 8-hydroxyguanine (8-OHG)
Thank you all for your responses. I am working with HL60 cell lines. I treat them with cisplatin (0,1,2 and 3 UM) for 24-96 hours. I wanted to know how the antioxidant enzyme levels would change when compared to control. I'm using lipid peroxidation assay from abcam.
Thank you. This is going to be very useful. I appreciate your valuable suggestion.
It has been implied that mutant RAS-driven tumor cells tend to be specific for the susceptibility to erastin and/ or artesunate, leading to ferroptosis. However, there is not positive correlation between ferroptotic cell death and expression amount of GPX4 and/or xCT transporter.
