Stefania Fulle

AGE-DEPENDENT INCREASES IN OXIDATIVE DAMAGE TO DNA, LIPIDS, AND PROTEINS IN HUMAN SKELETAL MUSCLE

A role for oxidative damage in normal aging is supported by studies in experimental animals, but there is limited evidence in man. We examined markers of oxidative damage to DNA, lipids, and proteins in 66 muscle biopsy specimens from humans aged 25 to 93 years. There were age-dependent increases in 8-hydroxy-2-deoxyguanosine (OH8dG), a marker of oxidative damage to DNA, in malondialdehyde (MDA), a marker of lipid peroxidation, and to a lesser extent in protein carbonyl groups, a marker of protein oxidation. The increases in OH8dG were significantly correlated with increases in MDA. These results provide evidence for a role of oxidative damage in human aging which may contribute to age-dependent losses of muscle strength and stamina.

The contribution of reactive oxygen species to sarcopenia and muscle ageing

In recent years, age-related diseases and disabilities have become of major interest and importance for health. This holds particularly for the Western community, where the remarkable improvement of medical health, standard of living, and hygiene have reduced the main causes of death. Despite numerous theories and intensive research, the principal molecular mechanisms underlying the process of aging are still unknown. Most, if not all, attempts to prevent or stop the onset of typical degenerative diseases associated with aging have so far been futile. Solutions to the major problems of dealing with age-related diseases can only come from a systematic and thorough molecular analysis of the aging process and a detailed understanding of its causes.

Specific oxidative alterations in vastus lateralis muscle of patients with the diagnosis of chronic fatigue syndrome

Chronic fatigue syndrome (CFS) is a poorly understood disease characterized by mental and physical fatigue, most often observed in young white females. Muscle pain at rest, exacerbated by exercise, is a common symptom. Although a specific defect in muscle metabolism has not been clearly defined, yet several studies report altered oxidative metabolism. In this study, we detected oxidative damage to DNA and lipids in muscle specimens of CFS patients as compared to age-matched controls, as well as increased activity of the antioxidant enzymes catalase, glutathione peroxidase, and transferase, and increases in total glutathione plasma levels. From these results we hypothesize that in CFS there is oxidative stress in muscle, which results in an increase in antioxidant defenses. Furthermore, in muscle membranes, fluidity and fatty acid composition are significantly different in specimens from CFS patients as compared to controls and to patients suffering from fibromyalgia. These data support an organic origin of CFS, in which muscle suffers oxidative damage.

Age and sex influence on oxidative damage and functional status in human skeletal muscle

A reduction in muscle mass, with consequent decrease in strength and resistance, is commonly observed with advancing age. In this study we measured markers of oxidative damage to DNA, lipids and proteins, some antioxidant enzyme activities as well Ca2+ transport in sarcoplasmic reticulum membranes in muscle biopsies from vastus lateralis of young and elderly healthy subjects of both sexes in order to evaluate the presence of age- and sex- related differences. We found a significant increase in oxidation of DNA and lipids in the elderly group, more evident in males, and a reduction in catalase and glutathione transferase activities. The experiments on Ca2+ transport showed an abnormal functional response of aged muscle after exposure to caffeine, which increases the opening of Ca2+ channels, as well a reduced activity of the Ca2+ pump in elderly males. From these results we conclude that oxidative stress play an important role in muscle aging and that oxidative damage is much more evident in elderly males, suggesting a gender difference maybe related to hormonal factors.

Age-dependent imbalance of the antioxidative system in human satellite cells

The mature myofibres of human skeletal muscle are surrounded by a type of adult stem cell, known as the satellite cell, which lies outside the sarcolemma but within the basal lamina. These cells remain quiescent until external stimuli trigger their re-entry into the cell cycle. In humans, ageing is characterised by a progressive loss of muscle mass and strength (sarcopenia) associated with a decline in functional ability. One of the possible causes of this decline in muscle performance is a decrease in the antioxidative capacity of skeletal muscle, resulting in an abnormal accumulation of the reactive oxygen species (ROS) critical for cell life. The present study shows that: (i) the antioxidant activity of Catalase and Gluthatione transferase in satellite cells derived from the elderly is drastically reduced compared to that in cells isolated from young individuals; (ii) cell membrane fluidity is considerably different between the two age groups; and (iii) basal [Ca(2+)](i) levels in satellite cells increase significantly in an age-dependent manner. In view of the data obtained, we hypothesise that the destabilising oxidative damage that occurs during ageing in skeletal muscle also affects quiescent satellite cells, which spend their life in close anatomic and functional contact with adult fibres. This status is derived from a decrease in the antioxidative capacity, and may negatively affect the ageing satellite cells ability to repair muscle.

The brain protein S-100ab induces apoptosis in PC12 cells

Incubation of PC12 cells with S-100 protein induces a rapid (0.5-1.0 min) rise of intracellular Ca2+ which lasts for the whole period of incubation. This effect is abolished in a Ca(2+)-free medium or in the presence of 1.0 microM Ni2+, an inhibitor of calcium channels. The rise in intracellular Ca2+ is followed by a progressive increase of cells undergoing degeneration and death. This event is accompanied by the appearance of apoptotic bodies and DNA fragmentation typical of the process known as apoptosis. S-100-induced cell death is prevented by 1 microM Ni2+ or by 0.1 nM cycloheximide, suggesting the involvement of new protein synthesis. It is postulated that the binding of S-100ab to specific sites present in PC12 cells is followed by the formation of Ca2+ channels and/or the stimulation of pre-existing ones with consequent increase of Ca2+ influx and activation of a process of cell death.

The S-100 protein causes an increase of intracellular calcium and death of PC12 cells

The S-100 protein-PC12 cell interaction has been studied as a model system of the possible physiological role played by S-100 protein in the nervous system. The data reported demonstrate that S-100 exerts a cytotoxic action which eventually leads to PC12 cell death, regardless of the cell cycle phase. The effect is specific for the S-100 isoforms, which are made up of two identical subunits and is abolished by a monoclonal antibody directed against the same isoforms. Other isoforms and/or calcium-binding proteins, such as troponin or calmodulin, do not induce the same effects. The action of S-100 on cell viability is not detectable in other cell lines of different embryological origin, such as 3T3, L1210, GH3. S-100 causes a rapid and considerable increase (two- to three-fold) of intracellular Ca2+ concentration in PC12 cells accompanied by cytostatic and cytotoxic action. It is postulated that this action also occurs in vivo, as part of the physiological action of this protein.

Molecular basis of the myogenic profile of aged human skeletal muscle satellite cells during differentiation

Sarcopenia is the age-related loss of muscle mass, strength and function. Human muscle proteins are synthesized at a slower rate in the elderly than in young adults, leading to atrophy and muscle mass loss with a decline in the functional capability. Additionally, aging is accompanied by a decrease in the ability of muscle tissue to regenerate following injury or overuse due to the impairment of intervening satellite cells, in which we previously reported oxidative damage evidences. The aim of the present study was to determine the effects of aging on myoblasts and myotubes obtained from human skeletal muscle, and characterize the transcriptional profile as molecular expression patterns in relation to age-dependent modifications in their regenerative capacity. Our data show that the failure to differentiate does not depend on reduced myogenic cell number, but difficulty to complete the differentiation program. Data reported here suggested the following findings: (i) oxidative damage accumulation in molecular substrates, probably due to impaired antioxidant activity and insufficient repair capability, (ii) limited capability of elderly myoblasts to execute a complete differentiation program; restricted fusion, possibly due to altered cytoskeleton turnover and extracellular matrix degradation and (iii) activation of atrophy mechanism by activation of a specific FOXO-dependent program.

Oxidative stress and muscle homeostasis

Purpose of reviewThe term oxidative stress is often used to indicate a condition in which the accumulation of reactive oxygen species is considered just damaging. We will discuss both the physiological and pathological role of oxidative stress on skeletal muscle homeostasis and function, and how oxidative stress can activates opposite signaling molecule to regulate gene and protein expression to guarantee muscle adaptation and to trigger a pathological condition. Recent findingsEmerging evidences have assigned a critical role to oxidative stress in muscle homeostasis and in the physiopathology of skeletal muscle, suggesting that reactive oxygen species are not merely damaging agent inflicting random destruction to the cell structure and function, but useful signaling molecules to regulate growth, proliferation, differentiation, and adaptation, at least within physiological concentration. SummaryThe role of oxidative stress on muscle homeostasis is quite complex. It is clear that transiently increased levels of oxidative stress might reflect a potentially health promoting process, whereas an uncontrolled accumulation of oxidative stress might have pathological implication. Additional work is, therefore, necessary to understand and define precisely whether the manipulation of the redox balance represents a useful approach in the design of therapeutic strategies for muscle diseases.

Calcium and Fos Involvement in Brain‐Derived Ca2+‐Binding Protein (S100)‐Dependent Apoptosis in Rat Phaeochromocytoma Cells

Brain‐derived calcium‐binding protein S100 induces apoptosis in a significant fraction of rat phaeochromocytoma (PC12) cells. We used single cell techniques (patch clamp, videomicroscopy and immunocytochemistry) to clarify some of the specific aspects of S100‐induced apoptosis, the modality(ies) of early intracellular Ca2+ concentration increase and the expression of some classes of genes (c‐fos, c‐jun, bax, bcl‐x, p‐15, p‐21) known to be implicated in apoptosis of different cells. The results show that S100: (1) causes an increase of [Ca2+]i due to an increased conductance of L‐type Ca2+ channels; (2) induces a sustained increase of the Fos levels which is evident since the first time point tested (3 h) and remains elevated until to the last time point (72 h). All these data suggest that S100‐derived apoptosis in PC12 cells may be the consequence of a system involving an increase in L‐type Ca2+ channel conductance with consequent [Ca2+]i increase which up‐regulates, directly or indirectly, the expression of Fos.