Redox homeostasis and age-related deficits in neuromuscular integrity and function

Abstract

Skeletal muscle is a major site of metabolic activity and is the most abundant tissue in the human body. Age-related muscle atrophy (sarcopenia) and weakness, characterized by progressive loss of lean muscle mass and function, is a major contributor to morbidity and has a profound effect on the quality of life of older people. With a continuously growing older population (estimated 2 billion of people aged >60 by 2050), demand for medical and social care due to functional deficits, associated with neuromuscular ageing, will inevitably increase. Despite the importance of this 'epidemic' problem, the primary biochemical and molecular mechanisms underlying age-related deficits in neuromuscular integrity and function have not been fully determined. Skeletal muscle generates reactive oxygen and nitrogen species (RONS) from a variety of subcellular sources, and age-associated oxidative damage has been suggested to be a major factor contributing to the initiation and progression of muscle atrophy inherent with ageing. RONS can modulate a variety of intracellular signal transduction processes, and disruption of these events over time due to altered redox control has been proposed as an underlying mechanism of ageing. The role of oxidants in ageing has been extensively examined in different model organisms that have undergone genetic manipulations with inconsistent findings. Transgenic and knockout rodent studies have provided insight into the function of RONS regulatory systems in neuromuscular ageing. This review summarizes almost 30 years of research in the field of redox homeostasis and muscle ageing, providing a detailed discussion of the experimental approaches that have been undertaken in murine models to examine the role of redox regulation in age-related muscle atrophy and weakness.

Keywords: Frailty; Mitochondria; Motor neurons; Neuromuscular junction; Redox signalling; Superoxide dismutase.

Figure 1

Schematic representation of the morphological neuromuscular alterations/impairments that occur with the advance of age. Ageing skeletal muscle is associated with increased fibre‐type grouping due to continual cycles of denervation and reinnervation. Axonal degeneration and motor neuron death, inherent with aging, leads to reduced number of motor axons innervating myofibres. These events inevitably result in loss of motor units and atrophy of the remaining muscle cells.

Figure 2

Reactive oxygen derivatives produced by the sequential reduction of O₂ to H₂O. Superoxide (O₂∸), hydrogen peroxide (H₂O₂) and hydroxyl radical (^●OH).

Figure 3

Schematic representation of the non‐mitochondrial sites for nitric oxide and superoxide production in skeletal muscle. Superoxide (O₂∸) is produced by multicomponent nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2), xanthine oxidase and the lipoxygenases (LOX), which activity is regulated by the phospholipase A₂ enzymes (PLA₂). Arachidonic acid (AA) release by the membrane bound calcium‐dependent PLA₂ (sPLA₂) facilitates extracellular O₂∸ release by the membrane bound LOX. It is uncertain whether the cytosolic LOX enzymes contribute to intracellular O₂∸ changes, which substrate availability might be regulated by the cytosolic calcium‐independent PLA₂ (iPLA₂). NAD(P)H oxidase 4 (NOX4) also contributes to ROS changes, although the primary ROS product, O₂∸, or hydrogen peroxide (H₂O₂) of NOX4 is uncertain. Cytosolic and extracellular O₂∸ is dismuted into H₂O₂ by superoxide dismutase (SOD), SOD1 and SOD3, respectively, or reacts rapidly with membrane permeant nitric oxide (NO) produced by the endothelial and neuronal nitric oxide synthase (eNOS and nNOS) to form peroxynitrite (ONOO⁻). H₂O₂ formed within the extracellular space is reduced into H₂O by the action of glutathione peroxidase 3 (GPX3) or peroxiredoxin IV (PRX4), while cytosolic H₂O₂ is reduced into H₂O by glutathione peroxidase 1 (GPX1), catalase (CAT) or peroxiredoxins (PRXs). Reduced glutathione (GSH) provides the electrons to GPX to catalyse the reduction of H₂O₂; GSH is oxidized to glutathione disulfide (GSSG). Reduction of GSSG is catalysed by glutathione reductase (GR), where NADPH is used as the reducing agent. Cytosolic PRXs utilize thioredoxin 1 (Trx1^Red) for their reducing action. Oxidized form of Trx1 (Trx1^Ox) is reduced by thioredoxin reductase 1 (TR1) by utilizing electrons from NAD(P)H. ONOO⁻ can be reduced predominantly into nitrite (NO2⁻) by peroxiredoxin V (PRX5). Sarcoplasmic reticulum (SR).

Figure 4

Schematic representation of the mitochondrial sites for nitric oxide and superoxide production and the channels that mediate the release of superoxide to the cytosolic compartment in skeletal muscle. Superoxide (O₂∸) is produced by complex I, complex II and complex III of the mitochondrial electron transport chain of the inner mitochondrial membrane (IMM) and released into the matrix and the mitochondrial intermembrane space (MIS). Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) also contributes to ROS changes, although the primary ROS product, O₂∸, or hydrogen peroxide (H₂O₂) of NOX4 is uncertain. Arachidonic acid (AA) release by the calcium‐dependent phospholipase A₂ enzymes (sPLA₂) interacts with complex I and enhances superoxide generation by this complex. O₂∸ released into the matrix, and MIS is dismuted into H₂O₂ by superoxide dismutase (SOD), SOD2 and SOD1, respectively, or reacts rapidly with nitric oxide (NO) produced by the endothelial nitric oxide synthase (eNOS) to form peroxynitrite (ONOO⁻). H₂O₂ is reduced into H₂O by the action of glutathione peroxidase 4 (GPX4) or peroxiredoxins (PRXs). Reduced glutathione (GSH) provides the electrons to GPX4 to catalyse the reduction of H₂O₂; GSH is oxidized to glutathione disulfide (GSSG). Reduction of GSSG is catalysed by glutathione reductase (GR), where NADPH is used as the reducing agent. Mitochondrial PRXs utilize thioredoxin 2 (Trx2^Red) for their reducing action. Oxidized form of Trx2 (Trx2^Ox) is reduced by thioredoxin reductase 2 (TR2) by utilizing electrons from NADPH. ONOO⁻ can be reduced predominantly into nitrite (NO2⁻) by peroxiredoxin V (PRX5). O₂∸ is essentially membrane impermeant, while H₂O₂ is readily diffusible. Matrix O₂∸ can diffuse to the cytosol through the inner membrane anion channel (iMAC) that spans the IMM and the outer mitochondrial membrane (OMM) or via the mitochondrial permeability transition pore (mPTP) composed of the voltage‐dependent anion channels (VDAC) on the OMM, the adenine‐nucleotide translocator (ANT) located on the IMM and cyclophilin D (Cyclo D) located in the matrix. Channels of the OMM including VDAC, BAX and possibly the translocase of outer membrane 40 (TOM40) can also mediate the release of O₂∸ from the MIS to the cytosol.

Figure 5

Gross morphology of skinned hindlimb muscles of SOD1^−/− and WT mice at 20 months of age. Redrawn from Jang et al. 2010.276

Figure 6

NMJ immunofluorescence images from AT muscle of SOD1^−/− and WT mice at 10 months of age. Left panels: morphology of presynaptic motor neurons stained with antibodies to synaptotagmin‐2 and neurofilaments (green staining). Middle panels: morphology of postsynaptic AChRs labelled with bungarotoxin (red staining). Right panels: merged images of presynaptic motor neurons and AChRs. Redrawn from Sakellariou et al.29 Original magnification: 60X (scale bar = 10 μm).