SAM Mice Research Articles

Dehydroascorbic acid, the oxidized form of vitamin C, improves renal histology and function in old mice.

Oxidative stress and inflammation are crucial factors that increase with age. In the progression of multiple age-related diseases, antioxidants and bioactive compounds have been recognized as useful antiaging agents. Oxidized or reduced vitamin C exerts different actions on tissues and has different metabolism and uptake. In this study, we analyzed the antiaging effect of vitamin C, both oxidized and reduced forms, in renal aging using laser microdissection, quantitative reverse-transcription polymerase chain reaction, and immunohistochemical analyses. In the kidneys of old SAM mice (10 months of age), a model of accelerated senescence, vitamin C, especially in the oxidized form (dehydroascorbic acid [DHA]) improves renal histology and function. Serum creatinine levels and microalbuminuria also decrease after treatment with a decline in azotemia. In addition, sodium-vitamin C cotransporter isoform 1 levels, which were increased during aging, are normalized. In contrast, the pattern of glucose transporter 1 expression is not affected by aging or vitamin C treatment. We conclude that oxidized and reduced vitamin C are potent antiaging therapies and that DHA reverses the kidney damage observed in senescence-accelerated prone mouse 8 to a greater degree.

Increase in tetrahydrobiopterin concentration with aging in the cerebral cortex of the senescence-accelerated mouse prone 10 strain caused by abnormal regulation of tetrahydrobiopterin biosynthesis

6R-L-Erythro-5,6,7,8-tetrahydrobiopterin (BH4) is an essential cofactor for tyrosine hydroxylase (TH) activity and is a risk factor for cognitive decline and brain atrophy. Previous studies have shown that the decline in TH activity in the cerebral cortex of senescence-accelerated mouse prone 10 (SAMP10) mice is caused, at least in part, by a decrease in Fe, ferritin, and TH phosphorylation. We determined the concentrations of BH4 and the enzymes GTP cyclohydrolase-1,6-pyruvoyltetrahydropterin synthase and sepiapterin reductase (SPR) in the de novo pathway of BH4 biosynthesis. Dihydrofolate reductase (DHFR), which converts BH2 to BH4 in the salvage pathway of BH4 synthesis was also determined in the cerebral cortex of SAM mice at 3 and 12months of age. The BH4 concentration was measured by HPLC, and the protein levels of enzymes involved in BH4 synthesis were measured by western blot analysis. At 12months of age, BH4 concentration in the cerebral cortex of SAMP10 mice showed significantly higher values as compared to that of control mice. Further, the protein level of SPR in SAMP10 mice was significantly higher than that in SAMR1 mice at 3 and 12months of age. In contrast to SPR, the protein level of DHFR in SAMP10 mice was significantly lower than that in SAMR1 mice. These results indicate that abnormal regulation of BH4 metabolism occurs in the cerebral cortex of SAMP10 where the dysfunction of the salvage pathway of BH4 synthesis may cause overproduction of BH4 through the de novo pathway, which is considered characteristic in the cerebral cortex of SAMP10 with aging. Therefore, there is a possibility that the excess amounts of BH4 lead to age-related brain dysfunction in the cerebral cortex of SAMP10.

Strain- and Age-related Alteration of Proteins in the Brain of SAMP8 and SAMR1 Mice

In order to discover and identify the key protein biomarkers in the aging process, we performed a differential proteomic analysis of hippocampus and cortex in 5- and 15-month old senescence-accelerated mouse prone 8 (SAMP8) as well as in control strain SAM/resistant 1 (SAMR1). Using 2-DE combined with MALDI TOF/TOF mass spectrometry, about 1700 protein spots were isolated, and three groups of differentially expressed proteins were identified. The first group contained the strain-specific and non-age-related differential proteins that were differentially expressed in SAMP8 compared with SAMR1 mice. The changes might be implicated in the genetic difference between SAMP8 and SAMR1 mice; specifically, the proteins ubiquitin carboxyl-terminal esterase L3, mitofilin, adenylate kinase 4, and an unnamed protein product (gi|12847201). The proteins in the second group were age-specific, which were differentially expressed between 5- and 15-month old SAM mice. Those proteins are particularly interesting since the changes were aging-related and some of them were previously reported to be expressed in Alzheimer's disease patients. These proteins included N-myc downstream regulated gene 2, enolase 2, Cu/Zn superoxide dismutase, myosin, and two unnamed protein products (gi|74214304 and gi|74178239). The protein in the third group was SAMP8 specific-age-related protein, which was identified as heme binding protein 1. The present study provides new information about SAMP8 specific and aging-related protein changes in brain. Further investigations will be performed to reveal the significance of these proteins in brain aging process and the potential roles as biomarkers for effective diagnosis and therapy.

D-Tyr-tRNA Tyr Deacylase, a New Role in Alzheimer's-associated Disease in SAMP8 Mice

To assess the expression level of D-Tyr-tRNA(Tyr) deacylase (DTD) in SAMP8 mice and speculate the function of DTD in disorders associated with Alzheimer's disease (AD). Altogether 12 SAMP8 mice and 12 SAMR1 mice were used in this study. Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were performed to detect the mRNA and protein levels of DTD in the mice. Purified DTD protein was injected into lateral ventricle to investigate the function of DTD in SAMP mice. The behavior of the mice was tested by using a Step-through Test System. Both mRNA and protein levels of DTD were found to be significantly lower in SAMP8 mice compared with those in SAMR1 mice (P<0.05). In vivo injection of DTD protein did not lead to an obvious change in behavior of SAM mice. DTD might function in the process of AD-associated pathology and could possibly participate in physiology process in a long-term manner to orchestrate with other regulators in order to maintain the balance of organism.

Melatonin alters cell death processes in response to age‐related oxidative stress in the brain of senescence‐accelerated mice

We studied the effect of age and melatonin on cell death processes in brain aging. Senescence-accelerated prone mice 8 (SAMP8) and senescence-accelerated resistant mice (SAMR1) at 5 and 10 months of age were used as models of the study. Melatonin (10 mg/kg) or its vehicle (ethanol at 0.066%) was administered in the drinking water from 1 to 9 months of age. Neurodegeneration, previously shown in the aged brain of SAMP8 and SAMR1 at 10 months of age, may be due to a drop in age-related proteolytic activities (cathepsin D, calpains, and caspase-3). Likewise, lack of apoptotic and macroautophagic processes were found, without apparent modification by melatonin. However, the caspase-independent cell death, owing to high p53 and apoptosis-inducing factor (AIF) levels, might be an alternative pathway of cell death in the aged brain. The main effects of melatonin treatment were observed in the aged SAMR1 mice; in this strain we observed a marked increase in antioxidant activity (catalase and superoxide dismutase). Likewise, a key antioxidant role of apoptosis-related proteins, Bcl-2 and AIF, was suggested in the aged brain of SAM mice, which was clearly influenced by melatonin. Moreover, the age-related increase of lysosomal activity of cathepsin B and a lysosomal membrane-associated protein 2 supports the possibility of the maintenance of lysosomal viability in addition to age-related impairments of the proteolytic or macroautophagic activities. The effectiveness of melatonin against the oxidative stress-related impairments and apoptosis during the aging process is, once more, corroborated in this article.

P3-409: The establishment of astrocyte cell models expressing endogenous MuLV from SAM mice

P3-409: The establishment of astrocyte cell models expressing endogenous MuLV from SAM mice

Improved mitochondrial function and increased life span after chronic melatonin treatment in senescent prone mice

We investigated whether chronic melatonin administration influences mitochondrial oxidative stress and life span in mice. Diaphragmatic mitochondria from female senescent prone (SAMP8) and senescent resistant (SAMR1) mice at 5 and 10 months of age were studied. Mitochondrial oxidative stress was determined by measuring the levels of lipid peroxidation, glutathione and glutathione disulfide, and glutathione peroxidase and reductase activities. Mitochondrial function was assessed by measuring the activity of the respiratory chain complexes and the ATP content. The results suggest that the age-dependent mitochondrial oxidative damage in the diaphragm of SAMP8 mice was accompanied by a reduction in the electron transport chain complex activities and in ATP levels. Furthermore, melatonin administration between 1 and 10 months of age normalized the redox and the bioenergetic status of the mitochondria and increased the ATP levels. Melatonin also increased both half-life and longevity, mainly in SAMP8 group. These results suggest an age-related increase in mitochondria vulnerability to oxidation in SAM mice at 10 months of age that was counteracted by melatonin therapy. The effects of melatonin on mitochondrial physiology probably underline the ability of the indoleamine to increase maximal life span in these animals.

Chronic melatonin treatment prevents age-dependent cardiac mitochondrial dysfunction in senescence-accelerated mice

Heart mitochondria from female senescence-accelerated (SAMP8) and senescence-resistant (SAMR1) mice of 5 or 10 months of age, were studied. Mitochondrial oxidative stress was determined by measuring the levels of lipid peroxidation, glutathione and glutathione disulfide and glutathione peroxidase and reductase activities. Mitochondrial function was assessed by measuring the activity of the respiratory chain complexes and ATP content. The results show that the age-dependent mitochondrial oxidative damage in the heart of SAMP8 mice was accompanied by a reduction in the electron transport chain complex activities and in ATP levels. Chronic melatonin administration between 1 and 10 months of age normalized the redox and the bioenergetic status of the mitochondria and increased ATP levels. The results support the presence of significant mitochondrial oxidative stress in SAM mice at 10 months of age, and they suggest a beneficial effect of chronic pharmacological intervention with melatonin, which reduces the deteriorative and functional oxidative changes in cardiac mitochondria with age.

Scrapie infection activates the replication of ecotropic, xenotropic, and polytropic murine leukemia virus (MuLV) in brains and spinal cords of senescence-accelerated mice: Implication of MuLV in progression of scrapie pathogenesis

Senescence-accelerated mice (SAMP8) have a short life span, whereas SAMR1 mice are resistant to accelerated senescence. Previously it has been reported that the Akv strain of ecotropic murine leukemia virus (E-MuLV) was detected in brains of SAMP8 mice but not in brains of SAMR1 mice. In order to determine the change of MuLV levels following scrapie infection, we analyzed the E-MuLV titer and the RNA expression levels of E-MuLV, xenotropic MuLV, and polytropic MuLV in brains and spinal cords of scrapie-infected SAM mice. The expression levels of the 3 types of MuLV were increased in scrapie-infected mice compared to control mice; E-MuLV expression was detected in infected SAMR1 mice, but only in the terminal stage of scrapie disease. We also examined incubation periods and the levels of PrP Sc in scrapie-infected SAMR1 (sR1) and SAMP8 (sP8) mice. We confirmed that the incubation period was shorter in sP8 (210 ± 5 days) compared to sR1 (235 ± 10 days) after intraperitoneal injection. The levels of PrP Sc in sP8 were significantly greater than sR1 at 210 ± 5 days, but levels of PrP Sc at the terminal stage of scrapie in both SAM strains were virtually identical. These results show the activation of MuLV expression by scrapie infection and suggest acceleration of the progression of scrapie pathogenesis by MuLV.

Changes of apoptosis-related proteins in hippocampus of SAM mouse in development and aging

Expression of Caspase and Bcl-2 proteins was examined in the hippocampus of senescence-accelerated mice (SAM, P8 and R1 strain) from E19 to 16 months of age. Immunoblotting analysis showed no upregulation of pro-apoptotic proteins (caspase-2L, -3, -6, -8, -9, and Bax) with age while all the anti-apoptotic proteins (caspase-2S, Bcl-2, and Bcl-XL) remained unchanged during aging. Terminal dUTP nick end labeling (TUNEL) and electron microscopy on the hippocampus of 3- and 16-month-old SAM revealed very few TUNEL positive cells in both groups. Morphometric study further showed neuronal loss in the hippocampus was not age-related. Our results suggest apoptosis and cell loss are minor events in the hippocampus of SAM mice and are unlikely to be the cause of functional decline during aging in SAM.

Carnosine-Induced Changes in the Development of Spermatogenic Epithelial Cells in Senescence Accelerated SAM Mice

Carnosine significantly increased the number of spermatogonia and Sertoli cells in mice prone (SAMP1) and resistant (SAMR1) to accelerated aging and appreciably reduced cell yield in meiosis and spermiogenesis in SAMP1 mice. In experimental SAMP1 mice catastrophic changes in the number of gametes were paralleled by intensive degradation of the spermatogenic epithelium. In SAMR1 mice treated with carnosine highly ordered spermatogenic structure was preserved.

Effect of aging on species-typical behaviors in senescence-accelerated mouse

The species-typical behaviors have been extensively studied, especially in the rodents. But little is known about whether the aging impacts on these species-typical behaviors. In the present study, the species-typical behaviors, including burrowing, hoarding and nesting, were assessed in the accelerated senescence-prone mouse 8 (SAMP8, P8) and the control strain senescence-resistant mouse 1 (SAMR1, R1). Total 147 SAM mice including 74 P8 mice and 73 R1 mice were grouped according to the age, 3, 7 and 11 months, respectively. In the hoarding test, an age-related increase was observed in the both P8 and R1 mice, whereas in the burrowing task, the age-related increment only took place in the P8 mice. The nesting ability in the P8 mice at different ages was inferior to that in the age-matched R1 mice, and the 3-month P8 mice showed the poorest nesting ability. The principal component analysis revealed that the burrowing, hoarding and nesting tests detected the different aspects of species-typical behaviors respectively for all mice combined. Our findings indicated that all tasks of hoarding, burrowing and nesting could detect the aging effect in the P8 mice, whereas, only the hoarding test could detect the aging effect in the R1 mice. These different species-typical behaviors were dissociable.

Rose (Rosa rugosa)-flower extract increases the activities of antioxidant enzymes and their gene expression and reduces lipid peroxidation

The effects of rose-flower extract on antioxidant enzymes were studied. The activities of catalase (CAT) and glutathione peroxidase (GPx) in 9-month-old senescence-accelerated mice (SAM mice) were lower than those in 6-month-old SAM mice. Therefore, 9-month-old SAM mice were the most appropriate targets for treatment with the rose-flower extract. The activities of CAT and GPx in SAM mice treated with rose-flower extract showed a marked increase in whole blood and liver. At the same time, the gene-expression level of CAT and GPx was upregulated in the liver, while malondialdehyde content in liver and brain decreased. Male SAM mice were more sensitive than female SAM mice. The mean and the longest lifespan of SAM mice were longer after treatment with rose-flower extract.

Comparison of thiol redox state of mitochondria and homogenates of various tissues between two strains of mice with different longevities

The main purpose of this study was to determine if differences in life spans of two different strains of mice are associated with the thiol redox state of their tissues and mitochondria. A comparison, based on amounts of reduced and oxidized glutathione (GSH, GSSG) and reactive protein thiols, was made between short-lived SAM (P8) mice and the longer-lived C57BL/6 mice at 13 months of age. The average life span of the latter mouse strain is approximately 48% longer than the former strain. Analyses of plasma, tissue homogenates and mitochondria of liver, kidney, heart, brain and skeletal muscle indicated that, in general, amounts of GSH and reactive protein sulfhydryls and GSH:GSSG ratios were lower and concentrations of GSSG were higher in the SAM than the C57BL/6 mice. Differences in the redox state between the two strains were more consistent and pronounced in skeletal muscle than in other tissues, and in mitochondria than in their respective tissue homogenates. Overall, the results support the view that the shorter-lived SAM mice exhibit a relatively higher level of oxidative stress than the longer-lived C57BL/6 mice, which is consistent with the predictions of the oxidative stress hypothesis of aging. Intra-species comparisons may be useful for the identification of biochemical characteristics associated with the variations in life spans.

Morphological and mechanical properties of caudal vertebrae in the SAMP6 mouse model of senile osteoporosis

The senescence-accelerated mouse, strain P6 (SAMP6), is a model of senile osteoporosis with relatively low bone mineral density (BMD), low rates of bone formation and reduced long-bone bending strength. Seeking to extend previous descriptions of the SAMP6 skeletal phenotype, we assessed the morphological and mechanical properties of vertebrae from SAM mice at 4 and 12 months of age. We hypothesized that, relative to SAMR1 controls, vertebrae from SAMP6 mice have: (1) less trabecular bone, (2) increased endosteal and periosteal bone size and (3) decreased whole-bone strength. Caudal vertebrae from adult female mice (SAMR1 and SAMP6; 4 and 12 months; n = 10–11 per group) were evaluated by micro-computed tomography and mechanical compression testing. SAMP6 vertebrae had 33% less trabecular bone volume in cephalad and caudal end regions than SAMR1 ( P < 0.001) due to significant reductions in both trabecular number ( P = 0.002) and thickness ( P < 0.001). In contrast to previous findings in SAMP6 long bones, SAMP6 vertebrae showed no evidence of increased overall bone dimensions compared to SAMR1, and in fact had smaller total bone area (TA) and endosteal area (EA) at 12 months of age. Whereas SAMR1 vertebrae showed signs of age-related expansion from 4 to 12 months, SAMP6 vertebral dimensions did not change with age. Mechanical properties of SAMP6 vertebrae were not significantly different than SAMR1 vertebrae (stiffness, yield force, ultimate force, displacement at ultimate force, energy to ultimate force; P > 0.05), another finding in contrast to previous results in SAMP6 long bones. In summary, reduced vertebral trabecular bone volume is another feature of the SAMP6 mice with relevance to senile osteoporosis. However, age-related bone expansion and reduced whole-bone strength were not evident in SAMP6 vertebrae, indicating that while the SAMP6 mouse has many features relevant to senile osteoporosis in humans, not all features are observed or detectable at all skeletal sites.

Abstracts from Other Presentations at the International Symposium on Toxins and Natural Products in Honor of Professor Anthony T. Tu

Abstracts from Other Presentations at the International Symposium on Toxins and Natural Products in Honor of Professor Anthony T. Tu

Effects of aging and blood pressure on the structure of the thoracic aorta in SAM mice: a model of age-associated degenerative vascular changes.

The effects of aging and blood pressure on the structural alterations of the thoracic aorta were examined using male, accelerated senescence-prone, short-lived SAMP11 mice or accelerated senescence-resistant, long-lived SAMR1 mice. The aortic wall thickness increased significantly by 34% in SAMR1 and by 62% in SAMP11 with advanced age. We observed branching, breakage and disorganization of the elastic lamellae, an increase in thin collagen fibrils between the medial smooth muscle cells and hypertrophy but a significant decrease in the number of medial smooth muscle cells with aging in both strains. These alterations observed in SAMP11 occurred earlier and were more exaggerated with advanced age than in SAMR1. The aortic lumen dilated gradually in SAMR1, but narrowed significantly in SAMP11 with aging. The systolic blood pressure did not differ significantly among SAMP11s aged 3-9months, or among all ages of SAMR1. However, it was elevated in SAMP11 at the terminal stage of their life. Our results suggest that the aorta in SAMR1 might reflect the physiological process of aging, whereas SAMP11 showed earlier changes due to the senescence acceleration of the vascular cells, which were exaggerated by the elevated blood pressure.

Age-related changes in myelopoietic response to lipopolysaccharide in senescence-accelerated (SAM) mice

The effects of in vivo lipopolysaccharide (LPS) administration on myelopoiesis were examined in senescence-accelerated (SAM) mice. Young mice injected with LPS exhibited: (a) increased femoral proliferative pool size; (b) transient reduction in femoral non-proliferative pool size and number of femoral colony forming unit-granulocyte macrophages (CFU-GMs); (c) marked increase in splenic CFU-GMs; and (d) transient increase in S-phase of femoral CFU-GMs. The responses of old mice after LPS administration differed from those of young mice in the following points: (a) no recovery of the femoral non-proliferative pool or femoral CFU-GMs, (b) less significant augmentation of the femoral proliferative pool and splenic CFU-GMs, and (c) prolonged reduction in S-phase of femoral CFU-GM. Injection of LPS into mice resulted in a hyperproduction of colony-stimulating activity (CSA) in bone followed by production of colony-inhibitory activity (CIA) in young mice and in contrast, an excessive CIA secretion from bone without an increase in CSA levels in old mice. These imbalances in the regulatory factors derived from non-hemopoietic cells in the bones may lead to an inappropriate response of myelopoiesis in aged SAM mice after LPS administration, which may play a key role in infections.

Aging of blood-brain barrier and neuronal cells of eye and ear in SAM mice

The SAMP, Senescence-Accelerated Mouse strains show senescence acceleration and age-associated pathological phenotypes similar to geriatric disorders seen in humans. Among these strains, SAMP8 mice show age-associated deficits in learning and memory. Histopathological studies revealed various neurodegenerative changes in the brain, including age-associated appearance of spongiform degeneration in the brain stem and of PAS-positive granular structures in the hippocampus. The blood-brain barrier (BBB) function of SAMP8 mice was also impaired with advancing age. The compromised BBB function in the olfactory bulb, the hippocampus and the pons of SAMP8 mice coincided with and might have been the cause of some morphological changes. Age-associated degeneration of receptor cells and ganglion neurons in the retina and cochlea also occurred in the SAM mice. Oxidative stress partly caused by mitochondrial dysfunction was detected and may be a cause of the neuronal cell degeneration. The SAM strains are useful tool in the attempt to understand the mechanisms of age-dependent neurodegeneration and to develop clinical interventions.

Structural and functional changes in proteins induced by free radical-mediated oxidative stress and protective action of the antioxidants N-tert-butyl-alpha-phenylnitrone and vitamin E.

The free radical theory of aging proposes that reactive oxygen species (ROS) cause oxidative damage over the lifetime of the subject. It is the cumulative and potentially increasing amount of accumulated damage that accounts for the dysfunctions and pathologies seen in normal aging. We have previously demonstrated that both normal rodent brain aging and normal human brain aging are associated with an increase in oxidative modification of proteins and in changes in plasma membrane lipids. Several lines of investigation indicate that one of the likely sources of ROS is the mitochondria. There is an increase in oxidative damage to the mitochondrial genome in aging and a decreased expression of mitochondrial mRNA in aging. We have used a multidisciplinary approach to the characterization of the changes that occur in aging and in the modeling of brain aging, both in vitro and in vivo. Exposure of rodents to acute normobaric hyperoxia for up to 24 h results in oxidative modifications in cytosolic proteins and loss of activity for the oxidation-sensitive enzymes glutamine synthetase and creatine kinase. Cytoskeletal protein spin labeling also reveals synaptosomal membrane protein oxidation following hyperoxia. These changes are similar to the changes seen in senescent brains, compared to young adult controls. The antioxidant spin-trapping compound N-tert-butyl-alpha-phenylnitrone (PBN) was effective in preventing all of these changes. In a related study, we characterized the changes in brain protein spin labeling and cytosolic enzyme activity in a series of phenotypically selected senescence-accelerated mice (SAMP), compared to a resistant line (SAMR1) that was derived from the same original parents. In general, the SAM mice demonstrated greater oxidative changes in brain proteins. In a sequel study, a group of mice from the SAMP8-sensitive line were compared to the SAMR1-resistant mice following 14 days of daily PBN treatment at a dose of 30 mg/kg. PBN treatment resulted in an improvement in the cytoskeletal protein labeling toward that of the normal control line (SAMR1). The results of these and related studies indicate that the changes in brain function seen in several different studies may be related to the progressive oxidation of critical brain proteins and lipids. These components may be critical targets for the beneficial effects of gerontotherapeutics both in normal aging and in disease of aging.