Liver Of Old Mice Research Articles

CIDE-A is expressed in liver of old mice and in type 2 diabetic mouse liver exhibiting steatosis

BackgroundIncreased levels of circulating fatty acids caused by insulin resistance and increased adipocyte lipolysis can accumulate within the liver resulting in steatosis. This steatosis sensitizes the liver to inflammation and further injury which can lead to liver dysfunction. We performed microarray analysis on normal mouse liver tissue at different ages and type 2 diabetic liver exhibiting steatosis to identify differentially expressed genes involved in lipid accumulation and liver dysfunction.ResultsMicroarray analysis identified CIDE-A as the most differentially expressed gene as a function of age. Mice fed a high fat diet developed hyperinsulinemia, hyperglycemia and liver steatosis, all features of the human metabolic syndrome. Increased CIDE-A expression was observed in type 2 diabetic liver and the elevated CIDE-A expression could be reversed by weight loss and normalization of plasma insulin. Also, CIDE-A expression was found to be correlated with hepatic lipid accumulation.ConclusionThe corresponding increase in CIDE-A expression with hyperinsulinemia and liver steatosis suggests a novel pathway for lipid accumulation in the liver.

Release of superoxide from skeletal muscle of adult and old mice: an experimental test of the reductive hotspot hypothesis

Increased extracellular generation of reactive oxygen species (ROS) as a result of increasing reliance on glycolytic metabolism by old mitochondria-rich tissues has been claimed to contribute to the propagation of oxidative damage during aging (the reductive hotspot hypothesis), but the process has not been examined experimentally in old animals. Superoxide activity in the extracellular fluid of gastrocnemius muscle and markers of oxidation in blood and the liver were examined in adult and old mice at rest and following a period of demanding isometric contractions. The activity of superoxide in muscle microdialysates did not differ between adult and old mice at rest, but during contractile activity, there was a significant increase in the superoxide activity in microdialysates from adult muscle but no increase in microdialysates from old muscle. At rest, the liver of old mice contained an increased malonaldehyde content and a decreased protein thiol content in comparison with adult mice, but following the contraction protocol, only the adult mice showed significant, transient increases in the serum and liver malonaldehyde content and a decrease in liver glutathione and protein thiol content. Further studies revealed that the lack of superoxide release from contracting muscle of old mice was not due to reduced force generation by these muscles. These data provide no evidence for an increased extracellular superoxide in resting or contracting skeletal muscle of old mice, or that release of superoxide from muscle contributes to oxidation of blood components in the liver in old mice as is predicted from the reductive hotspot hypothesis.

Growth Hormone Corrects Proliferation and Transcription of Phosphoenolpyruvate Carboxykinase in Livers of Old Mice via Elimination of CCAAT/Enhancer-binding Protein α-Brm Complex

Growth hormone (GH), which is reduced with age, corrects the impaired proliferative capacity of livers of old animals. In this paper, we present a mechanism by which GH eliminates age-dependent negative control of proliferation and increases transcription of liver-specific genes in livers of old mice. The reduced proliferative capacities of the liver of old animals are associated with the CCAAT/enhancer-binding protein alpha (C/EBPalpha)-Brm complex, which inhibits E2F-dependent promoters. We found that a sequestration of C/EBPalpha into complexes with Brm leads to a weak interaction of C/EBPalpha with promoters of liver-specific genes, expression of which is reduced in old animals. Injection of either GH or the regulator of the amplitude of endogenous GH release, ghrelin, reduces the C/EBPalpha-Brm complex in livers of old mice, leading to a derepression of E2F targets, to increased interactions of C/EBPalpha with promoters of liver-specific genes, and to correction of their expression. GH-dependent elimination of the complex is mediated by the inhibition of cyclin D3-CDK4 activity and by elevation of a phosphatase, protein phosphatase 2A, which dephosphorylates C/EBPalpha and dissociates the complex.

Hepatic pseudocapillarization in aged mice

Age-related changes in the hepatic sinusoid of the rat, human and baboons called pseudocapillarization have been discovered and are important because they are considered to be implicated in the pathogenesis of some age-related diseases. In this study, we investigated whether similar changes occur in the livers of old mice. Livers of young (3–4 months) and old (20–24 months) mice were perfusion-fixed and studied using electron microscopy and immunohistochemistry. The thickness of the sinusoidal endothelium was increased in old mice (154±4 versus 244±8 nm, P<0.001). There was a reduction in fenestrations within the endothelium (porosity decreased from 4.1±0.3 to 2.2±0.2%, P<0.001). There was perisinusoidal staining with Sirius red in old mice, however, expression of laminin and von Willebrands factor was similar in young and old mice. Novel perisinusoidal fat-engorged stellate cells were found extensively in the old mice. This study confirmed that pseudocapillarization is a widespread aging change in the liver, now documented in several species including the mouse. Mice are an appropriate animal model for studying aging and the hepatic sinusoid.

Krebs cycle enzymes from livers of old mice are differentially regulated by caloric restriction

Krebs cycle enzyme activities and levels of five metabolites were determined from livers of old mice (30 months) maintained either on control or on long-term caloric restriction (CR) diets (28 months). In CR mice, the cycle was divided into two major blocks, the first containing citrate synthase, aconitase and NAD-dependent isocitrate dehydrogenase which showed decreased activities, while the second block, containing the remaining enzymes, displayed increased activity (except for fumarase, which was unchanged). CR also resulted in decreased levels of citrate, glutamate and α-ketoglutarate, increased levels of malate, and unchanged levels of aspartate. The α-ketoglutarate/glutamate and malate/α-ketoglutarate ratios were higher in CR, in parallel with previously reported increases with CR in pyruvate carboxylase activity and glucagon levels, respectively. The results indicate that long-term CR induces a differential regulation of Krebs cycle in old mice and this regulation may be the result of changes in gene expression levels, as well as a complex interplay between enzymes, hormones and other effectors. Truncation of Krebs cycle by CR may be an important adaptation to utilize available substrates for the gluconeogenesis necessary to sustain glycolytic tissues, such as brain.

Age-related bias in function of natural killer T cells and granulocytes after stress: reciprocal association of steroid hormones and sympathetic nerves.

Stress-associated immune responses were compared between young (8 weeks of age) and old (56 weeks) mice. Since stress suppresses the conventional immune system (i.e. T and B cells) but inversely activates the primordial immune system (i.e. extrathymic T cells, NKT cells, and granulocytes), these parameters were analysed after restraint stress for 24 h. The thymus became atrophic as a function of age, and an age-related increase in the number of lymphocytes was seen in the liver. Although the number of lymphocytes in both the thymus and liver decreased as the result of stress, the magnitude was much more prominent in the thymus. To determine stress-resistant lymphocyte subsets, two-colour immunofluorescence tests were conducted in the liver and spleen. NKT cells were found to be such cells in the liver of young mice. On the other hand, an infiltration of granulocytes due to stress was more prominent in the liver of old mice than in young mice. Liver injury as a result of stress was prominent in young mice. This age-related bias in the function of NKT cells and granulocytes seemed to be associated with a difference in the responses of catecholamines (high in old mice) and corticosterone (high in young mice) after stress. Indeed, an injection of adrenaline mainly induced the infiltration of granulocytes while that of cortisol activated NKT cells. The present results suggest the existence of age-related bias in the function of NKT cells and granulocytes after stress and that such bias might be produced by different responses of sympathetic nerves and steroid hormones between young and old mice.

Superoxide dismutase, catalase and glutathione peroxidase activities in the liver of young and old mice: linear regression and correlation

Activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GP) were determined in the liver of 15 young (3–5 months) and 15 old (23–26 months) female CBA mice. Although, the activities of all three enzymes decreased in aging, only the SOD decline was statistically significant ( P<0.01). The pair wise correlation analysis exhibited an almost identical positive correlation between SOD and CAT in the liver of both young and old mice ( r=0.57), whereas the correlative links between the enzymatic pairs of SOD–GP and CAT–GP were increased in aging. Close to zero in the young group ( r=−0.08), the coefficient of correlation between SOD and GP became highly significant in the group of old mice ( r=0.66; P<0.01). When the coefficients of pair wise and partial correlation were compared, practically no differences were found for the young mice, whereas all three partial coefficients were decreased in the group of old animals. Estimation of the linear regression between the enzymatic pairs revealed higher coefficients of regression and lower intercepts in the group of old mice. The results imply stronger correlative links between the antioxidant enzymes in the liver of old mice, compared with the same indices in the liver of young animals.

Characterization of Two Age-induced Intracisternal A-particle-related Transcripts in the Mouse Liver: TRANSCRIPTIONAL READ-THROUGH INTO AN OPEN READING FRAME WITH SIMILARITIES TO THE YEAST CCR4 TRANSCRIPTION FACTOR

Intracisternal A-particle (IAP) sequences are endogenous retrovirus-like elements present at 1,000 copies in the mouse genome. We had previously identified IAP-related transcripts of unusual size (6 and 10 kilobases (kb)), which are observed exclusively in the liver of the aging mouse. In this report, using cDNA libraries that we have constructed from the liver mRNAs of an aged DBA/2 mouse, we have cloned and entirely sequenced the corresponding cDNAs. Both are initiated within the 5' long terminal repeat of a type IDelta1 IAP sequence, and correspond to a read-through into a unique flanking cellular sequence containing a 966-nucleotide open reading frame, located 3' to the IAP sequence. The 6-kb IAP-related transcript corresponds to a post-transcriptional modification of the 10-kb mRNA, and is generated by a splicing event with the donor site in the IAP sequence, and the acceptor site 5' to the open reading frame. This open reading frame is located on chromosome 3, is evolutionarily conserved, and discloses significant similarity to the yeast CCR4 transcription factor at the amino acid level. The specific expression of these age-induced transcripts, which account for more than 50% of the IAP-related transcripts in the liver of old mice, is therefore entirely consistent with the induction of a single genomic locus, thus strengthening the importance of position effects for the expression of transposable elements. Characterization of this locus should now allow studies on its chromatin and methylation status, and on the "molecular factors of senescence" possibly involved in its induction.

William Carver, practical horse farrier.

Protection of polyunsaturated fatty acids within the liver of old mice was realized by the ingestion of culinary and medicinal plant volatile oils containing natural antioxidants obtained by hydrodistillation. This protection effectively reversed the normal trend in polyunsaturated fatty acid metabolism during aging wherein a decrease in levels is concomitant with a reduction in tissue function and integrity. Volatile oil from thyme was overall the most effective in this protective capacity.

Age-associated changes in dolichol and dolichyl phosphate metabolism in the kidneys and liver of mice

The total amounts of cholesterol, dolichol and dolichyl phosphate (Dol-P) in kidneys and liver from 1- and 24-month-old mice were measured after saponification of the tissues. The cholesterol content of kidneys decreased by about 4-fold per g of tissue between the two ages, while the amount of cholesterol/g liver remained constant. The dolichol content of both tissues increased by about 2-fold. The amount of Dol-P in kidneys increased by 7-fold over 23 months, while the concentration in liver decreased some 6-fold. Metabolic labelling experiments using tritiated water demonstrated that the rate of synthesis of total dolichol (dolichol + Dol-P) in the kidneys did not change as a function of age. However, the kidneys of young mice incorporated 3-fold more radioactivity into dolichol than Dol-P, whereas this distribution was reversed in old mice. In liver, the rate of dolichol synthesis decreased 2-fold over 23 months. It was not possible to compare the rates of Dol-P biosynthesis between ages as Dol-P was undetectable in liver of old mice. The relative abundance of the 19 isoprene unit homologs of both dolichol and Dol-P decreased approx. 5% while the 17 isoprene unit homologs increased by a similar amount in both tissues of the old mice.

Influence of age upon the metabolism of zinc in livers of C57BL/6J mice

The kinetics of accumulation and loss of zinc from the liver following subcutaneous administration of 10 mg of zinc per kg were examined in young adult (6 months old) and old (24 months old) male C57BL/6J mice. After zinc treatment, total liver zinc concentrations rose equally in both groups and returned to basal levels at 96 h post treatment. However, differences were found in the subcellular distribution of zinc in these two age groups. The concentration of zinc in the cytosolic fraction (104 000 g supernate) prepared from the livers of old mice attained its maximum at 24 h post treatment. In contrast, the concentration of zinc in the cytosolic fraction of liver from young adult mice peaked at 48 h post treatment. This difference in accumulation of zinc in the cytosol was reflected by differences in the binding of zinc to metallothionein, a cytosolic transition metal binding protein. In old mice the highest amounts of zinc bound to metallothionein were found at 24 h post treatment; in young adults the maximal zinc binding to this protein occurred at 48 h post treatment. Examination of the relationship between cytosolic zinc contents and the binding of zinc to metallothionein in young adult and old mice suggested similar regulatory processes in the two age groups. Thus, age-dependent differences in accumulation and loss of zinc from the cytosolic fraction of liver probably reflect factors other than differences in regulationof the synthesis of metallothionein by this essential metal.

Effect of aging on oxidative metabolism and conjugation in perfused livers of an inbred strain of mice: A pilot study

A study was carried out to examine the possibility that altered mitochondrial function with aging might cause changes in hepatic rates of oxygen uptake and β-oxidation of fatty acids. Livers of 2–4 month and 20–25 month old CBA mice were perfused, and rates of respiration and ketogenesis were measured. Livers of old mice had significantly decreased rates of oxygen utilization (190 ± 16 vs 151 ± 9 μmol/gm/hr, p.05) and significantly lower rates of ketogenesis as indicated by rates of acetoacetate and β-hydroxybutyrate production (84 ± 8 vs 63 ± 7, p <.05). To determine whether NADPH supply, which is largely derived from β-oxidation of fatty acids in the fasted state, might be rate limiting for mixed-function oxidation in aged mice, rates of p-nitroanisole O-demethylation were measured in livers of young and old mice. Age had no effect on mixed-function oxidation of p-nitroanisole. Rates of glucuronidation of p-nitrophenol were also not affected by age; however, rates of sulfation of p-nitrophenol were significantly higher in livers of old (7.2 ±.6) than in young (3.5 ± 0.8) mice. This pilot study raises the possibility that the disposition of ingested fatty acids and xenobiotic compounds requiring sulfation may be significantly altered with aging. Moreover, it demonstrates that the perfused mouse livers can be used as a model in aging studies.

Changes with age in cadmium and copper levels in C57BL/6J mice

Cadmium concentrations in C57BL/6J male mice were found to increase with age in kidney from 0.1 ng/mg dry wt. at 45 days of age to 1.7 ng/mg dry wt. at 880 days of age. Cadmium in liver increased exponentially with age with a doubling time of 242 days, from a value of 0.03 ng/mg dry wt. at 45 days of age to 0.29 ng/mg dry wt. at 880 days of age. Kidney copper declined by only 14% between 45 and 400 days of age and remained unchanged between 400 and 880 days of age. Liver copper declined 41% between 45 and 500 days of age and showed no change between 500 and 880 days of age. Most of the decline in kidney and liver copper concentrations occurred before 250 days of age, possibly reflecting developmental changes. Feeding cadmium chloride in drinking water at concentrations of up to 100 μg/ml did not change the copper concentrations in kidney, liver, heart or brain of young mice. Feeding copper gluconate did not change the cadmium concentration in livers of old mice. However, these treatments did change the cadmium/copper ratios in tissues.

The effect of age on protein synthesis in mouse liver.

1. A system of microsomes and 105000g supernatant from livers of old mice is less able to promote the incorporation of [(14)C]phenylalanine into protein than a similar system from livers of young animals. 2. The decrease in [(14)C]phenylalanine incorporation is attributable to changes in microsomes from old animals rather than in the cell-sap fraction. 3. Decreased synthetic ability is found in various classes of microsomes from older animals, namely unfractionated, light and heavy microsomes, but not in detergent-washed ribonucleoprotein particles. 4. Deletions of certain detergent-soluble microsomal proteins accompany the decreased synthetic ability of microsomes from older animals. 5. Microsomes from old mice are less responsive to a synthetic messenger RNA, polyuridylic acid, and this is partly due to a higher rate of hydrolysis in the presence of cell sap from animals of extreme age. 6. Other more direct evidence, from the priming of a cell-free protein-synthesizing system from bacteria and the examination of ribonucleoprotein particles on sucrose density gradients, suggests that senescence is accompanied by a decrease in messenger RNA content.