*3.1. Mechanism of Oxidative Stress*

Reactive oxygen species are mainly produced by a dysfunction in the mitochondria. The range of dysfunction through mitochondria is very wide and comprises a decrease in ATP production, altered regulation of calcium, brain iron accumulation, respiratory chain dysfunction, perturbation in the dynamics of mitochondria, and deregulation of mitochondria clearance [73].

In patients with Parkinson's disease, several genes have been identified that strongly confirm the association of mitochondrial function with oxidative stress in the pathogenesis of the disease. The loss in functioning of the DJ-1 gene causes oxidative stress and exerts a neuroprotective role in response to its antioxidant characteristic in mitochondria [74]. Mutations in the recessive form of the PINK1 gene leads to the onset of Parkinson's disease. PINK1 is a mitochondria kinase that regulates cytosolic calcium, and deficiency in this gene leads to impairment of calcium influx in mitochondria and causes mitochondrial calcium overload, which leads to the opening of the mitochondrial permeability transition pore (MPTP) and permits the translocation of proapoptotic molecules to cytosol from mitochondria [75].

Apart from Parkinson's disease, the dysfunction of mitochondria has also been identified in the pathology of Alzheimer's disease and other common neurodegenerative disorders. The ROS are produced via the oxidation of nicotinamide adenine dinucleotide phosphate (NADPH) and selective neural degeneration. The crucial role of NADPH oxidase in neurodegenerative disease has been confirmed in several animal models. The active form of NADPH oxidase relocates proteins to membranes to enhance healthy neuronal activity and requires the breaching of ion/anion channels to compensate the charge. In microglial cells activated by beta-amyloid proteins, the superoxide production is inhibited [76]. The stimulation of NADPH oxidase by beta-amyloid proteins in combination with the abundant synthesis of nitric oxide can cause the destruction of the surrounding cells. The beta-amyloid proteins also activate NADPH oxidase by causing the entry of calcium into the astrocytes, thereby preventing their entry into the neuronal cells. After NADPH oxidase actuation [77], the mitochondrial membrane is depolarized, which, combined with calcium, leads to the opening of MPTP and changes the membrane structure by phospholipase C stimulation. This signal of oxidative stress is further carried to neighboring neuronal cells that are highly exposed compared to astrocytes. [78]. When the neurons have multiple

synapses and long axons, they have high energy requirements for transportation through axons and long-term plasticity.

To fulfil such big energy requirements, a high demand for ATP is rendered, thereby increasing levels of reactive oxygen species in conditions of stress, which imparts a higher degree of neurodegeneration. For example, the neurons in the hippocampus region of the brain generate greater levels of superoxide anions and comprise increased expression of antioxidant and reactive oxygen species that produce genes [79]. The amplified oxidative stress in intrinsic surroundings leads to the dysfunction of mitochondria, which further generates more free radicals, thereby exaggerating the oxidative stress cycle.

Figure 4 represents the process of oxidative stress-mediated neurodegeneration. Based on external and internal factors, the production of reactive oxygen species increases in the body, which causes an overall increase in the oxidative stress. This oxidative stress causes aberrant phosphorylation of proteins, leading to synthesis of misfolded proteins, which alters the normal functioning of neuronal transmission. Due to altered protein structures, they get aggregated and lose the ability to transport, thereby hampering the synaptic activity and causing the initiation and progression of neurodegeneration.

**Figure 4.** Synthesizing the process of oxidative stress-mediated neurodegeneration. Legend: ROS—reactive oxygen species; RNS—reactive nitrogen species; UV—ultraviolet; NADP—nicotinamide adenine dinucleotide.
