Table 1.

Main experimental approaches in the study of the OS theory of disease

Studies based on testing the effect of ROS or antioxidants on a disease‐related process
1. Studies investigating the effect of ROS on disease‐related pathway. For instance, H2O2 induces aggregation of α‐synuclein, a protein important in Alzheimer's disease (Hashimoto et al., 1999); adding H2O2 to immune cells causes the production of inflammatory mediators (DeForge et al., 1993); and injecting a H2O2‐generating enzyme in the mouse knee joint causes inflammation (Schalkwijk et al., 1986).
2. Studies investigating the effect of an antioxidant on a disease‐related pathway. For instance, polyphenols present in red wine inhibit formation of amyloid fibrils that are important in Alzheimer's disease (Ono et al., 2003); antioxidants inhibit TNF production in vitro and in vivo in mice (Chaudhri and Clark, 1989); antioxidants from red wine are protective in animal models of stroke (Yu et al., 2016); and clinical trials with antioxidants.
3. Study of the effect of genetic modification (knock out and overexpression) or modulation of antioxidant enzymes or ROS‐generating enzymes, on a disease‐related pathway or an animal model of disease (Sorce et al., 2014).

Studies based on measuring ROS or antioxidants in a disease or its model
1. Studies showing that ROS are produced in a disease or a disease model. Thus, ROS are generated during the aggregation of amyloid peptides implicated in Alzheimer's disease (Tabner et al., 2005); ischaemia causes activation of a ROS‐producing enzyme, xanthine oxidase (Granger, 1988); and an indicator of ROS production, MDA, is elevated in blood from patients with systemic lupus erythematosus (Wang et al., 2010).
2. Studies showing that endogenous antioxidants are decreased in a disease or disease models in vitro or in vivo, or in patients. For instance, levels of plasma antioxidants are lower in osteoporotic patients (Maggio et al., 2003).
3. Genome‐wide association studies where a mutation in a gene for a ROS‐generating enzyme or an antioxidant protein is associated with a disease.