Mitochondrial oxidative stress and increased seizure susceptibility in Sod2(-/+) mice

Epileptic seizures can occur as a result of mitochondrial dysfunction. Mitochondria have vital functions such as energy generation, control of cell death, neurotransmitter synthesis, and free radical production. Which of these critical mitochondrial functions contributes to epileptic seizures is unknown. We demonstrate here that a subset of mice with partial deficiency of the mitochondrial superoxide dismutase (Sod2(-/+)) show increased incidence of spontaneous and handling-induced seizures that correlates with chronic mitochondrial oxidative stress (increased aconitase inactivation and 8-hydroxy-2'-deoxyguanosine formation in mitochondria) and diminished mitochondrial oxygen utilization. Before the age at which spontaneous seizures appear in a subset of the mice, Sod2(-/+) mice demonstrated increased susceptibility to behavioral seizures, mitochondrial aconitase inactivation, and neurodegeneration induced by the administration of kainate. These data suggest that chronic mitochondrial oxidative stress initiated by superoxide (O(2)(.-)) radicals is sufficient to increase seizure susceptibility due to aging, environmental stimulation, or excitotoxin administration. Sod2(-/+) mice showed an age-related decrease in the expression of glial glutamate transporters (GLT-1 and GLAST), suggesting that oxidant-induced inhibition of glutamate transport may play a mechanistic role in rendering some Sod2(-/+) mice susceptible to seizures. In summary, mitochondrial oxidative stress and resultant dysfunction may be an important mechanism underlying certain seizure disorders.     

Early induction of oxidative stress in mouse model of Alzheimer disease with reduced mitochondrial superoxide dismutase activity

While oxidative stress has been linked to Alzheimer's disease, the underlying pathophysiological relationship is unclear. To examine this relationship, we induced oxidative stress through the genetic ablation of one copy of mitochondrial antioxidant superoxide dismutase 2 (Sod2) allele in mutant human amyloid precursor protein (hAPP) transgenic mice. The brains of young (5-7 months of age) and old (25-30 months of age) mice with the four genotypes, wild-type (Sod2(+/+)), hemizygous Sod2 (Sod2(+/-)), hAPP/wild-type (Sod2(+/+)), and hAPP/hemizygous (Sod2(+/-)) were examined to assess levels of oxidative stress markers 4-hydroxy-2-nonenal and heme oxygenase-1. Sod2 reduction in young hAPP mice resulted in significantly increased oxidative stress in the pyramidal neurons of the hippocampus. Interestingly, while differences resulting from hAPP expression or Sod2 reduction were not apparent in the neurons in old mice, oxidative stress was increased in astrocytes in old, but not young hAPP mice with either Sod2(+/+) or Sod2(+/-). Our study shows the specific changes in oxidative stress and the causal relationship with the pathological progression of these mice. These results suggest that the early neuronal susceptibility to oxidative stress in the hAPP/Sod2(+/-) mice may contribute to the pathological and behavioral changes seen in this animal model.     

Skeletal muscle weakness due to deficiency of CuZn-superoxide dismutase is associated with loss of functional innervation

An association between oxidative stress and muscle atrophy and weakness in vivo is supported by elevated oxidative damage and accelerated loss of muscle mass and force with aging in CuZn-superoxide dismutase-deficient (Sod1(-/-)) mice. The purpose was to determine the basis for low specific force (N/cm(2)) of gastrocnemius muscles in Sod1(-/-) mice and establish the extent to which structural and functional changes in muscles of Sod1(-/-) mice resemble those associated with normal aging. We tested the hypothesis that muscle weakness in Sod1(-/-) mice is due to functionally denervated fibers by comparing forces during nerve and direct muscle stimulation. No differences were observed for wild-type mice at any age in the forces generated in response to nerve and muscle stimulation. Nerve- and muscle-stimulated forces were also not different for 4-wk-old Sod1(-/-) mice, whereas, for 8- and 20-mo-old mice, forces during muscle stimulation were 16 and 30% greater, respectively, than those obtained using nerve stimulation. In addition to functional evidence of denervation with aging, fiber number was not different for Sod1(-/-) and wild-type mice at 4 wk, but 50% lower for Sod1(-/-) mice by 20 mo, and denervated motor end plates were prevalent in Sod1(-/-) mice at both 8 and 20 mo and in WT mice by 28 mo. The data suggest ongoing denervation in muscles of Sod1(-/-) mice that results in fiber loss and muscle atrophy. Moreover, the findings support using Sod1(-/-) mice to explore mechanistic links between oxidative stress and the progression of deficits in muscle structure and function.     

Loss of adaptation to oxidative stress as a mechanism for aortic damage in aging rats

Cells are armed with a vast repertoire of antioxidant defence mechanisms to prevent the accumulation of oxidative damage. The cellular adaptive response is an important antioxidant mechanism against physiological and pathophysiological oxidative alterations in a cell's microenvironment. The aim of this paper was to study, in the rat aorta, whether this adaptive response and the inflammation associated with oxidative stress were expressed throughout the aging process. We examined the rat aorta, as it is a very sensitive tissue to oxidative stress. Male Wistar rats of 1.5, 3, 12, 18 and 24 months of age were used. Superoxide anion (O2(-)) generation; levels of two antioxidant enzymes, superoxide dismutase (SOD) and catalase; and the levels of prostaglandin E2 (PGE2), an inflammatory marker, were measured. The results for rats at different ages were compared with those for 3 months of age. A balance between production of O2(-) and SOD activity was found in the aorta of rats from 1.5 to 12 months old. Oxidative stress was present in the aorta of old animals (18-24 months), due to a failure in the mechanisms of adaptation to oxidative stress. The observed increase in PGE2 levels in these rats reflected an inflammatory response. All together suggest that vascular oxidative stress and the inflammatory process observed in the old groups of rats could be closely related to vascular aging. Our results also remark the importance of the adaptative response to oxidative stress.     

Dietary restriction attenuates age-associated muscle atrophy by lowering oxidative stress in mice even in complete absence of CuZnSOD

Age-related loss of muscle mass and function, sarcopenia, has a major impact on the quality of life in the elderly. Among the proposed causes of sarcopenia are mitochondrial dysfunction and accumulated oxidative damage during aging. Dietary restriction (DR), a robust dietary intervention that extends lifespan and modulates age-related pathology in a variety of species, has been shown to protect from sarcopenia in rodents. Although the mechanism(s) by which DR modulates aging are still not defined, one potential mechanism is through modulation of oxidative stress and mitochondrial dysfunction. To directly test the protective effect of DR against oxidative stress-induced muscle atrophy in vivo, we subjected mice lacking a key antioxidant enzyme, CuZnSOD (Sod1) to DR (60% of ad libitum fed diet). We have previously shown that the Sod1(-/-) mice exhibit an acceleration of sarcopenia associated with high oxidative stress, mitochondrial dysfunction, and severe neuromuscular innervation defects. Despite the dramatic atrophy phenotype in the Sod1(-/-) mice, DR led to a reversal or attenuation of reduced muscle function, loss of innervation, and muscle atrophy in these mice. DR improves mitochondrial function as evidenced by enhanced Ca(2+) regulation and reduction of mitochondrial reactive oxygen species (ROS). Furthermore, we show upregulation of SIRT3 and MnSOD in DR animals, consistent with reduced mitochondrial oxidative stress and reduced oxidative damage in muscle tissue measured as F(2) -isoprostanes. Collectively, our results demonstrate that DR is a powerful mediator of mitochondrial function, mitochondrial ROS production, and oxidative damage, providing a solid protection against oxidative stress-induced neuromuscular defects and muscle atrophy in vivo even under conditions of high oxidative stress.     

Absence of CuZn superoxide dismutase leads to elevated oxidative stress and acceleration of age-dependent skeletal muscle atrophy

We describe a novel phenotype in mice lacking the major antioxidant enzyme, CuZn-superoxide dismutase (Sod1(-/-) mice), namely a dramatic acceleration of age-related loss of skeletal muscle mass. Sod1(-/-) mice are 17 to 20% smaller and have a significantly lower muscle mass than wild-type mice as early as 3 to 4 months of age. Muscle mass in the Sod1(-/-) mice is further reduced with age and by 20 months, the hind-limb muscle mass in Sod1(-/-) mice is nearly 50% lower than in age-matched wild-type mice. Skeletal muscle tissue from young Sod1(-/-) mice has elevated oxidative damage to proteins, lipids, and DNA compared to muscle from young wild-type mice. The reduction in muscle mass and elevated oxidative damage are accompanied by a 40% decrease in voluntary wheel running by 6 months of age and decreased performance on the Rota-rod test at 13 months of age, but are not associated with a decline in overall spontaneous activity. In some of the old Sod1(-/-) mice, the loss in muscle mass is also associated with the presence of tremors and gait disturbances. Thus, the absence of CuZnSOD imposes elevated oxidative stress, loss of muscle mass, and physiological consequences that resemble an acceleration of normal age-related sarcopenia.     

Stimulation of the amyloidogenic pathway by cytoplasmic superoxide radicals in an Alzheimer's disease mouse model

Oxidative stress is involved in the pathogenesis of neurodegeneration. Amyloid β (Aβ) oligomer as an intermediate of aggregates causes memory loss in Alzheimer's disease (AD). We have suggested that oxidative stress plays an important role in Aβ oligomerization and cognitive impairment using a human amyloid precursor protein (hAPP) transgenic AD mice lacking cytoplasmic superoxide dismutase (hAPP/Sod1-/-). Recently, clinical trials revealed inhibitors of Aβ production from hAPP as promising therapeutics, but the relationship between oxidative stress and Aβ metabolism remains unclear. Here we found that Sod1 deficiency enhanced β-cleavage of hAPP, suggesting that it increased Aβ production in hAPP/Sod1-/- mice. In contrast, Aβ degradation did not decrease in hAPP/Sod1-/- as compared with hAPP/Sod1+/+ mice. Furthermore, we successfully detected in situ superoxide radicals associated with increased protein carbonylation in hAPP/Sod1-/-. These results suggest that cytoplasmic oxidative stress is involved in Aβ production as well as aggregation during AD progression.     

Superoxide dismutase 1 knockdown induces oxidative stress and DNA methylation loss in the prostate

Increased oxidative stress and concordant DNA methylation changes are found during aging and in many malignant processes including prostate cancer. Increased oxidative stress has been shown to inhibit DNA methyltransferase in in vitro assays, but whether this occurs in vivo is unknown. To generate increased oxidative stress we utilized mice containing mutations in the CuZnSOD (Sod1) gene, a major superoxide dismutase in mammals. Increased 8-hydroxy-2'-deoxyguanosine, an adduct indicating oxidative damage, was found in liver and prostate tissues at 2 and 12 mo Sod1^+/- mice compared to controls. Prostate tissues from Sod1^+/- mice demonstrated decreased weight at 2 mo compared to controls, but this difference was not significant at 12 mo. Histologic changes were not seen. Global DNA methylation was significantly decreased at 2 mo in the prostate in Sod1^+/- mice. 11p15 containing the epigenetically modulated insulin-like growth factor 2 (Igf2) and H19 genes, both which display oncogenic functions, may be particularly sensitive to oxidative stress. CpG island methylation at an intergenic CTCF binding site and the Igf2 P3 promoter was decreased in Sod1 mutants compared to controls. This is the first in vivo study to show that a deficiency of Sod1 leads to a decrease in DNA methylation. These studies indicate that increased oxidative stress, a factor implicated in neoplasia, can induce DNA hypomethylation in prostate tissues.     

Effect of oxidative stress on telomere maintenance in aortic smooth muscle cells

Reactive oxygen species (ROS) and telomere dysfunction are both associated with aging and the development of age-related diseases. Although there is evidence for a direct relationship between ROS and telomere dysfunction as well as an independent association of oxidative stress and telomere attrition with age-related disorders, there has not been sufficient exploration of how the interaction between oxidative stress and telomere function may contribute to the pathophysiology of cardiovascular diseases (CVD). To better understand the complex relationships between oxidative stress, telomerase biology and pathophysiology, we examined the telomere biology of aortic smooth muscle cells (ASMCs) isolated from mutant mouse models of oxidative stress. We discovered that telomere lengths were significantly shorter in ASMCs isolated from superoxide dismutase 2 heterozygous (Sod2^+/?) mice, which exhibit increased arterial stiffness with aging, and the observed telomere attrition occurred over time. Furthermore, the telomere erosion occurred even though telomerase activity increased. In contrast, telomeres remained stable in wild-type and superoxide dismutase 1 heterozygous (Sod1^+/?) mice, which do not exhibit CVD phenotypes. The data indicate that mitochondrial oxidative stress, in particular elevated superoxide levels and decreased hydrogen peroxide levels, induces telomere erosion in the ASMCs of the Sod2^+/? mice. This reduction in telomere length occurs despite an increase in telomerase activity and correlates with the onset of disease phenotype. Our results suggest that the oxidative stress caused by imbalance in mitochondrial ROS, from deficient SOD2 activity as a model for mitochondrial dysfunction results in telomere dysfunction, which may contribute to pathogenesis of CVD.     

The Role of Oxidative Stress in Nervous System Aging

While oxidative stress is implicated in aging, the impact of oxidative stress on aging in the peripheral nervous system is not well understood. To determine a potential mechanism for age-related deficits in the peripheral nervous system, we examined both functional and morphological changes and utilized microarray technology to compare normal aging in wild-type mice to effects in copper/zinc superoxide dismutase-deficient (Sod1^−/−) mice, a mouse model of increased oxidative stress. Sod1^−/− mice exhibit a peripheral neuropathy phenotype with normal sensory nerve function and deficits in motor nerve function. Our data indicate that a decrease in the synthesis of cholesterol, which is vital to myelin formation, correlates with the structural deficits in axons, myelin, and the cell body of motor neurons in the Sod1^+/+ mice at 30 months and the Sod1^−/− mice at 20 months compared with mice at 2 months. Collectively, we have demonstrated that the functional and morphological changes within the peripheral nervous system in our model of increased oxidative stress are manifested earlier and resemble the deficits observed during normal aging.     

Effect of the reduction of superoxide dismutase 1 and 2 or treatment with alpha-tocopherol on tumorigenesis in Atm-deficient mice

Atm-deficient mice, a cancer-prone model of the human disease ataxia-telangiectasia, display increased levels of oxidative stress and damage. Chronic treatment of these mice with the nitroxide antioxidant and superoxide dismutase (SOD) mimetic Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) resulted in an increased latency to tumorigenesis. We initially hypothesized that the chemopreventative effect of Tempol was due to its SOD mimetic activity reducing cellular oxidative stress and damage. However, it is also possible that the chemopreventative effect of Tempol results from mechanisms other than directly reducing superoxide radical-induced oxidative stress and damage. To help distinguish between these possibilities, we attempted to genetically increase oxidative stress in Atm-deficient mice by either removing cytosolic Sod1 or reducing mitochondrial Sod2, or we attempted to decrease oxidative stress by treatment of Atm-deficient mice with alpha-tocopherol. Surprisingly, we found that reducing both Atm and Sod1 or Atm and Sod2 did not shorten latency to tumorigenesis or significantly affect life span. Furthermore, continuous administration of alpha-tocopherol did not affect latency to thymic lymphomas. Thus, genetically reducing Sod in Atm-deficient mice or treatment with alpha-tocopherol had no effect on survival or tumorigenesis, suggesting that the chemopreventative effect of Tempol may be at least partially independent of its effects on reducing oxidative damage and stress.     

Mitochondrial H2O2 regulates the angiogenic phenotype via PTEN oxidation

Recent studies have demonstrated that the tumor suppressor PTEN (phosphatase and tensin homolog deleted from chromosome 10), the antagonist of the phosphosphoinositol-3-kinase (PI3K) signaling cascade, is susceptible to H2O2-dependent oxidative inactivation. This study describes the use of redox-engineered cell lines to identify PTEN as sensitive to oxidative inactivation by mitochondrial H2O2. Increases in the steady state production of mitochondrial derived H2O2, as a result of manganese superoxide dismutase (Sod2) overexpression, led to PTEN oxidation that was reversed by the coexpression of the H2O2-detoxifying enzyme catalase. The accumulation of an oxidized inactive fraction of PTEN favored the formation of phosphatidylinositol 3,4,5-triphosphate at the plasma membrane, resulting in increased activation of Akt and modulation of its downstream targets. PTEN oxidation in response to mitochondrial H2O2 enhanced PI3K signaling, leading to increased expression of the key regulator of angiogenesis, vascular endothelial growth factor. Overexpression of PTEN prevented the H2O2-dependent increase in vascular endothelial growth factor promoter activity and immunoreactive protein, whereas a mutant PTEN (G129R), lacking phosphatase activity, did not. Furthermore, mitochondrial generation of H2O2 by Sod2 promoted endothelial cell sprouting in a three-dimensional in vitro angiogenesis assay that was attenuated by catalase coexpression or the PI3K inhibitor LY2949002. Moreover, Sod2 overexpression resulted in increased in vivo blood vessel formation that was H2O2-dependent as assessed by the chicken chorioallantoic membrane assay. Our findings provide the first evidence for the involvement of mitochondrial H2O2 in regulating PTEN function and the angiogenic switch, indicating that Sod2 can serve as an alternative physiological source of the potent signaling molecule, H2O2.     

Dietary restriction attenuates the accelerated aging phenotype of Sod1−/− mice

Dietary restriction is a powerful aging intervention that extends the life span of diverse biological species ranging from yeast to invertebrates to mammals, and it has been argued that the antiaging action of dietary restriction occurs through reduced oxidative stress/damage. Using Sod1^−/− mice, which have previously been shown to have increased levels of oxidative stress associated with a shorter life span and a high incidence of neoplasia, we were able to test directly the ability of dietary restriction to reverse an aging phenotype due to increased oxidative stress/damage. We found that dietary restriction increased the life span of Sod1^−/− mice 30%, returning it to that of wild-type, control mice fed ad libitum. Oxidative damage in Sod1^−/− mice was markedly reduced by dietary restriction, as indicated by a reduction in liver and brain F₂-isoprostanes, a marker of lipid peroxidation. Analysis of end of life pathology showed that dietary restriction significantly reduced the overall incidence of pathological lesions in the Sod1^−/− mice fed the dietary-restricted diet compared to Sod1^−/− mice fed ad libitum, including the incidence of lymphoma (27 vs 5%) and overall liver pathology. In addition to reduced incidence of overall and liver-specific pathology, the burden and severity of both neoplastic and nonneoplastic lesions was also significantly reduced in the Sod1^−/− mice fed the dietary-restricted diet. These data demonstrate that dietary restriction can significantly attenuate the accelerated aging phenotype observed in Sod1^−/− mice that arises from increased oxidative stress/damage.     

Ionizing radiation induces a Yap1-dependent peroxide stress response in yeast

Repair of DNA damage is fundamental for cellular tolerance to ionizing radiation (IR) and many IR-induced DNA lesions are thought to occur as a result of oxidative stress. We investigated the physiological effects of IR in Saccharomyces cerevisiae by performing protein expression profiles in cells exposed to electron pulse irradiation. Transient induction of several antioxidant enzymes in wild-type cells, but not in cells lacking the oxidative stress regulator Yap1, indicated that IR exposure causes cellular oxidative stress. Yap1 activation involved oxidation to the intramolecular disulfide bond, a signature of activation by peroxide, and was dependent on the Yap1 peroxide sensor Orp1/Gpx3. H(2)O(2) was produced in the culture medium of irradiated cells and was both necessary and sufficient for IR-induced Yap1 activation. When IR was performed in the presence of N(2)O, obviating H(2)O(2) production and increasing hydroxyl radical ((*)OH) production, the Yap1 response was lost, indicating that Yap1 was unable to respond to (*)OH or (*)OH-induced damage. However, the Yap1 response to IR did not seem to be a primary determinant of cellular IR tolerance. Altogether, these data provide a molecular demonstration that cells experience in vivo peroxide stress during IR and indicate that the H(2)O(2) produced cannot account for IR toxicity.     

Fibroblasts derived from Gpx1 knockout mice display senescent-like features and are susceptible to H2O2-mediated cell death

The Free Radical Theory of Aging proposes that reactive oxygen species (ROS) contribute to the pathophysiology of aging. Our previous data highlight the importance of antioxidant enzymes, superoxide dismutase 1 (Sod1) and glutathione peroxidase 1 (Gpx1), in regulating this process. Previously, we demonstrated that a perturbation in the Sod1-to-Gpx1 ratio, as a consequence of Sod1 overexpression, leads to senescence-like changes. We proposed that this was mediated via the Sod1 dismutation product H2O2, because H2O2 induced similar changes in control cells. However, it has been suggested that H2O2 production, via Sod1 dismutation, is rate-limited by the availability of the substrate O2*-, and therefore age-related changes may occur as a result of other functions of Sod1. In this study, we test this notion in fibroblasts derived from Gpx1 null mutant mice (Gpx1-/-) that have elevated H2O2 as a consequence of the lack of its removal by Gpx1. We demonstrate senescence-like changes in Gpx1-/- fibroblasts that include (1) reduced proliferative capacity, DNA synthesis, and responsiveness to EGF and serum; (2) elevated levels of Cip1; (3) increased NF-kappaB activation; and (4) morphological features of senescent cells. Gpx1-/- fibroblasts also demonstrate a dose-dependent susceptibility to H2O2-induced apoptosis. Our findings suggest that Gpx1 is protective against both ROS-mediated senescence-like changes and oxidant-mediated cell death.     

Deletions encompassing the manganese superoxide dismutase gene in the Drosophila melanogaster genome.

Two deletions, Df(2R)Sod2-11 and Df(2R)Sod2-332, are recovered that encompass the manganese superoxide dismutase (MnSOD) gene or a null mutant referred to as SOD2n283 in Drosophila. Molecular analysis has revealed that the Df(2R)Sod2-332 deletion completely uncovered both MnSOD and its adjacent gene, Arp53D, whereas Df(2R)Sod2-11 was missing the promoter region of MnSOD gene. As a consequence of reduced MnSOD expression, these deletion heterozygotes are now sensitive to oxidative stress. Complementation analysis with some recently recovered deletions in the 53C/D region has established that other essential loci exist in this interval, and second, that Arp53D function is not essential for the survival of the organism. These deletions will be instrumental in the recovery of missense substitutions in the MnSOD peptide and their influence on oxidative stress resistance.     

Endogenous mitochondrial oxidative stress in MnSOD-deficient mouse embryonic fibroblasts promotes mitochondrial DNA glycation

The accumulation of somatic mutations in mitochondrial DNA (mtDNA) induced by reactive oxygen species (ROS) is regarded as a major contributor to aging and age-related degenerative diseases. ROS have also been shown to facilitate the formation of certain advanced glycation end-products (AGEs) in proteins and DNA and N(2)-carboxyethyl-2'-deoxyguanosine (CEdG) has been identified as a major DNA-bound AGE. Therefore, the influence of mitochondrial ROS on the glycation of mtDNA was investigated in primary embryonic fibroblasts derived from mutant mice (Sod2(-/+)) deficient in the mitochondrial antioxidant enzyme manganese superoxide dismutase. In Sod2(-/+) fibroblasts vs wild-type fibroblasts, the CEdG content of mtDNA was increased from 1.90 ± 1.39 to 17.14 ± 6.60 pg/μg DNA (p<0.001). On the other hand, the CEdG content of nuclear DNA did not differ between Sod2(+/+) and Sod2(-/+) cells. Similarly, cytosolic proteins did not show any difference in advanced glycation end-products or protein carbonyl contents between Sod2(+/+) and Sod2(-/+). Taken together, the data suggest that mitochondrial oxidative stress specifically promotes glycation of mtDNA and does not affect nuclear DNA or cytosolic proteins. Because DNA glycation can change DNA integrity and gene functions, glycation of mtDNA may play an important role in the decline of mitochondrial functions.     

Reactive oxygen species on bone mineral density and mechanics in Cu,Zn superoxide dismutase (Sod1) knockout mice

Reactive oxygen species (ROS) play a role in a number of degenerative conditions including osteoporosis. Mice deficient in Cu,Zn-superoxide dismutase (Sod1) (Sod1^−/− mice) have elevated oxidative stress and decreased muscle mass and strength compared to wild-type mice (WT) and appear to have an accelerated muscular aging phenotype. Thus, Sod1^−/− mice may be a good model for evaluating the effects of free radical generation on diseases associated with aging. In this experiment, we tested the hypothesis that the structural integrity of bone as measured by bending stiffness (EI; N/mm²) and strength (MPa) is diminished in Sod1^−/− compared to WT mice. Femurs were obtained from male and female WT and Sod1^−/− mice at 8 months of age and three-point bending tests were used to determine bending stiffness and strength. Bones were also analyzed for bone mineral density (BMD; mg/cc) using micro-computed tomography. Femurs were approximately equal in length across all groups, and there were no significant differences in BMD or EI with respect to gender in either genotype. Although male and female mice demonstrated similar properties within each genotype, Sod1^−/− mice exhibited lower BMD and EI of femurs from both males and females compared with gender matched WT mice. Strength of femurs was also lower in Sod1^−/− mice compared to WT as well as between genders. These data indicate that increased oxidative stress, due to the deficiency of Sod1 is associated with decreased bone stiffness and strength and Sod1^−/− mice may represent an appropriate model for studying disease processes in aging bone.