A physical model of axonal damage due to oxidative stress

Abstract

Oxidative damage is implicated in the pathogenesis of neurodegenerative disorders, including Alzheimer's, Parkinson's, and Huntington's diseases, and in normal aging. Here, we model oxidative stress in neurons using photogenerated radicals in a simplified membrane-encapsulated microtubule system. Using fluorescence and differential interference contrast microscopies, we monitor photochemically induced microtubule breakdown on the supported region of membrane in encapsulating synthetic liposomes as a function of lipid composition and environment. Degradation of vesicle-encapsulated microtubules is caused by attack from free radicals formed upon UV excitation of the lipid-soluble fluorescent probe, 6-(9-anthroyloxy)stearic acid. Probe concentration was typically limited to a regime in which microtubule degradation was slow, and microtubule degradation was monitored by changes in the observed protrusion of the membrane surface. The kinetics of microtubule degradation are influenced by lipid saturation level, fluorescent probe concentration, and the presence of free-radical scavengers. This system is sufficient to reproduce some degenerative morphologies found in vivo.O xidative modifications to proteins in neurons have been implicated in a number of degenerative disorders and have become a controversial topic in elucidating the progression of Parkinson's, Huntington's, and Alzheimer's diseases (1-7). Microtubules (MTs) are key cytoskeletal proteins in nerve cell axons. These protein polymers are responsible for many disparate functions of cells, including structural support and imparting polarity, and they are involved in motility and transport of intracellular cargo (8). Fundamental studies of MTs in vivo are complicated by the presence of the full complement of biomolecules that are required for cell survival. Thus, a substantial effort has been devoted to characterizing MT dynamics in vitro, and these studies have led to increased understanding of issues such as dynamic instability (9) and the involvement of molecular motors in producing motility (10). To mimic the confinement of a bilayer, MT assemblies (asters) have previously been examined in rigid, microfabricated wells (11). The drawback of such studies is that interactions between MTs and components of the elastic lipid membrane are neglected. It is important to be able to isolate and to quantify specific factors that impact the structural integrity of the cytoskeleton, while maintaining a simplified cell-like environment.Toward this aim, we have developed a minimal physical system with which to investigate the effects of oxidative stress on the cytoskeleton. Previous studies have demonstrated that incorporating actin and tubulin into liposomes and inducing these proteins to polymerize results in a morphological change in the liposome; a protrusion of the membrane is observed at one or both ends of the protein polymer (12-15). The asymmetry occurs frequently, when the MT wraps around within the encapsulating liposome (16). Here, we employ the membrane as a...

Oxidative Stress and Proinflammatory Cytokines Contribute to Demyelination and Axonal Damage in a Cerebellar Culture Model of Neuroinflammation

“…In turn, this leads to energetic failure, protein and lipid oxidation, and microtubule degradation, thus impairing functions such as axonal transport and structural support [1], [3], [34]. The axonal swelling and mitochondria accumulation were pertinently present in the model and were consistent with a disruption of microtubules by oxidative stress and the subsequent blockade of axonal transport [6].…”

Liraglutide activates autophagyviaGLP-1R to improve functional recovery after spinal cord injury

“…Studies including our work show that pathogenic factors such as reactive oxidative stress (ROS) can trigger apoptosis and delayed death of cells in the ischemic penumbral region due to a buildup of oxidative damage to macromolecules, such as protein and DNA oxidation [ 34 – 36 ]. Moreover, excess oxidative stress compromises the function of axon via accelerating the degradation of microtubules [ 37 ]. Microtubules and microfilaments are susceptible to oxidation, which inhibit the polarization of microtubules and cause the destruction of microfilaments in neurons [ 38 ].…”

Axonal Degeneration in Motor Neuron Disease

“…Lysine-and proline-rich side arms on NF-H and NF-M are one of several theoretical targets for free radical modification that could lead to carbonyl formation, cross-linking, and alterations in secondary structure [95] . In a physical model of the axonal cytoskeleton, free radicals were sufficient to induce a 'catastrophic' level of cytoskeletal collapse, preventable by adding vitamin C or vitamin E [96] .…”