Hyon Jeen Kim

The Inflammation Hypothesis of Aging

Abstract: Current evidence strongly indicates that reactive oxygen species (ROS) and reactive nitrogen species (RNS) are widely implicated in the inflammatory process. However, mechanistic information is not readily available on the extent to which ROS/RNS contributes to the proinflammatory states of the aging process. The involvement of the underlying inflammation during the aging process and the molecular delineation of anti‐inflammatory action of calorie restriction (CR) is described. Age‐related upregulations of NF‐κB, IL‐β, IL‐6, TNFα, cyclooxygenase‐2, and inducible NO synthase are all attenuated by CR. The suppression of the NF‐κB activation was accomplished by blocking the dissociation of inhibitory IκBα and IκBβ by CR. These findings provide underlying molecular insights into the anti‐inflammatory action of CR in relation to the aging process. Based on these and other available data, it is suggested that the “Inflammation Hypothesis of Aging” supports the molecular basis of the inflammatory process as a plausible cause of the aging process.

Age-associated increases in oxidative stress and nuclear transcription factor κB activation are attenuated in rat liver by regular exercise

The combined effects of aging and regular physical exercise was investigated on the production of reactive oxygen species (ROS), lipid peroxidation, glutathione status, and the activity of nuclear factor‐κB (NF‐κB) in rat liver. A group of 24 male F344 rats was divided into the following categories: adult control (18 months), adult exercised (18 months), and aged control (28 months) and aged exercised (28 months). The ROS formation increased as a function of age and exercise training decreased the rate of ROS formation in the two age groups. Significant positive correlation was found between ROS production and lipid peroxidation (LIPOX). The reduced glutathione (GSH) level was higher and the oxidized glutathione (GSSG) level lower in exercised groups compared with the sedentary controls (P<0.05). An age‐associated increase in NF‐κB activity was attenuated by the regular exercise. The content of p50 and p65 subunits of NF‐κB increased with age and decreased with exercise training. The content of inhibitory factor‐κB was inversely related to NF‐κB activation. Regular exercise‐induced adaptive responses, including attenuation of an increase in ROS production, LIPOX level, NF‐κB activation, and reduced GSH/GSSG ratio, appear to be capable, even in old age, of reducing increases in inflammatory and other detrimental consequences that are often associated with advancing age.

Modulation of glutathione and thioredoxin systems by calorie restriction during the aging process

Accumulating evidence strongly suggests that oxidative stress underlies aging processes and that calorie restriction (CR) retards aging processes, leading to an extended lifespan for various organisms. Recent studies revealed that the anti-aging action of CR depends on its anti-oxidative mechanism. However, at present, the status of glutathione (GSH) and thioredoxin (Trx) system, two major thiol redox systems in animal cells during aging and its modulation by CR has not fully been explored. The purpose of this study is two-fold: one, to determine whether these two systems in rat kidney are altered as a consequence of aging; two, to determine whether these systems can be modulated by anti-oxidative CR. The results of our study showed that GSH and GSH-related enzyme activities decreased with age in ad libitum (AL)-fed rats, while CR rats consistently showed resistance to decreases in these activities. Data from the present data further showed that while Trx and Trx reductase (TrxR) in cytoplasm decrease with age in AL-fed rats, CR prevents these decreases. In contrast, we also found that the nuclear translocation of the redox regulators, Trx and Ref-1, increase with age, which was suppressed in CR rats. Therefore, increases in nuclear Trx and Ref-1 during aging may result in the up-regulation of redox-sensitive transcription factors, such as NF-kappaB or AP-1, via the interaction of Ref-1 and Trx in a redox-dependent manner. Our conclusion is that a redox imbalance occurs during aging and that redox changes are minimized through the anti-oxidative action of CR.

The inflammatory process in aging

Aging processes are time-dependent, deteriorative functional changes. These functional changes lead to a progressive loss of the organism’s ability to withstand both internal and environmental stresses, causing the failure of cellular homeostasis. Among the modern hypotheses, the ‘Oxidative Stress Hypothesis’ offers the best mechanistic elucidation of aging phenomena. Based on the ‘Free Radical Theory of Aging’, this hypothesis has gained popularity among researchers in the field of gerontology as well as other biomedical fields. Its primary premise proposes that aging and its related disease processes are the net result of free radical-induced damage, asserting further that an organism’s inability to produce counterbalancing antioxidative defences, i.e. defences that offset disturbances in the redox state, underlies its cause.

Influence of aging and calorie restriction on MAPKs activity in rat kidney.

Mitogen-activated protein kinases (MAPK), which include the extracellular signal-related kinases (ERK), the c-Jun N-terminal kinases (JNK), and the p38 MAPK, are important regulatory proteins by which a wide variety of extracellular signals are transduced into intracellular sites. Recent studies reported that mitogenic signal transduction in various cell types are exquisitely sensitive to reactive oxygen species (ROS) and the celluar redox status. In the present study, we investigated the activation of MAPK activity by aging and calorie restriction (CR) in rat kidneys isolated from Fischer 344 rats, ages 6, 12, 18, and 24 months fed ad libitum (AL) and CR diets. Results showed that the aging process strongly enhanced all three of the MAPK activities studied, ERK, JNK, and p38 MAPK, in parallel to increased ROS status. In contrast, we observed CR to markedly suppress the age-related activation of MAPKs. Based on these data, we concluded that an age-related increase in MAPK activity is associated with increased ROS, which was effectively suppressed by the anti-oxidative action of CR.

Suppression of apoptosis by calorie restriction in aged kidney.

Programmed cell death by apoptosis is regarded as an organisms protective mechanism against the accumulation of defective cells. Apoptotic activity is shown to be elevated in most aged tissues, and its intracellular regulation is intricately manipulated by mitochondria. In this study, to determine the progression of apoptosis during aging, we investigated the expression of several key apoptosis-related markers in kidney of 12- and 24-month-old rats. Mitochondrial damage was detected by lipid peroxidation and Western blot analysis in several target apoptotic proteins in aged rat kidney. Our results showed that the expression levels of a pro-apoptotic Bax protein, was significantly enhanced at the age of 24 months, while an anti-apoptotic protein, Bcl-2, was reduced in the aged rat kidney. We also found that the cytosolic cytochrome c level was significantly increased in the aged kidney. However, these age-related changes were reversed by calorie restriction (CR), exhibiting its modulatory action on apoptotic activity. Furthermore, caspase-3 activation was markedly increased in kidney of 24-month-old rats fed ad libitum (AL), as indicated by the cleaved, active form of caspase-3 (17-19 kDa), which we found was replaced with the procaspase (32 kDa) in the CR rats of both age groups. We also found that a cleaved active form (85 kDa) of poly (ADP-ribose) polymerase (116 kDa inactivated form), which serves as a nuclear substrate for active caspase-3, was increased in aged AL kidney and was blunted by CR. In addition, to investigate the oxidative status in aged kidney, we measured and compared the malondialdehyde (MDA) and 4-hydroxynonenal (HNE) levels in aged AL and CR rat kidneys. Our results showed increased MDA and HNE levels in aged AL rats, while these levels were markedly lower in CR rats, even at 24 months. These results indicate that the kidneys of rats fed ad libitum are under the influence of high oxidative stress compared to CR rats. Thus, our present data strongly suggest that the apoptotic activity observed in the aged kidney is likely modulated by the age-related oxidative status, and reversed by CR as a result of its anti-oxidative and anti-aging actions.

The effect of age and calorie restriction on HIF-1-responsive genes in aged liver

Hypoxia inducible factor-1 (HIF-1) regulates transactivation of several genes in response to hypoxia condition. We explore hepatic HIF-1 responsive gene regulation during aging and the age-related changes of the HIF-1 related gene activation in young and old rats. Results indicate that the aging process induces the activation of HIF-1α, which is accompanied by increased HIF-1 DNA binding. This increased binding activity is accompanied by the increase of HIF-1-dependent genes, heme oxygenase-1 (HO-1), vascular endothelial growth factor (VEGF), erythropoietin (EPO), and inducible nitric oxide synthase (iNOS), which all showed remarkable up-regulation during aging process. In contrast, the increased HIF-1 related gene expression was effectively blunted by the anti-oxidative action of calorie restriction in aged rat liver. We propose that age-related HIF-1 binding activity may well be influenced by the increased pro-oxidative conditions of aged animals, which up-regulate HIF-1-dependent gene expression.

Calorie restriction modulates redox-sensitive AP-1 during the aging process.

Oxidative stress is claimed to be a major cause of aging. Recent data suggest that calorie restriction (CR) prolongs life span by its ability to retard aging, possibly by regulating the intracellular redox status through its antioxidative actions. Currently, there is little information showing the influences of age and CR on the redox-sensitive transcription factor activator protein-1 (AP-1). In the present study, we investigated how age affects the status of AP-1 and whether CR modulates the age effect. For our study, we used the kidney from male Fischer 344 rats, ages 6, 12, 18, and 24 months fed ad libitum (AL) or a CR diet. Results from our study showed that AP-1 binding activity markedly increases with age, while CR keeps this activity at the level of 6-month-old rats. We found that c-Jun and c-Fos protein levels increase during aging, and that aging induces phosphorylation of c-Jun, which might enhance AP-1 transcriptional activity. For CR’s action, we found that in the nucleus of aged rats, AP-1 activation was blunted by decreasing c-Jun and c-Fos levels and inhibiting c-Jun protein phosphorylation. Results also indicated that matrix metalloproteinase-13 and heme oxygenase-1, which have an AP-1 binding site in their promoter regions, have a similar tendency toward AP-1 binding activity. Based on the data of these findings, we concluded that AP-1 activity increases in rat kidney with age and that CR reduces AP-1 activity.