Osteoporosis is negatively correlated with body mass, whereas both osteoporosis and weight loss occur at higher incidence during the progression of Alzheimer's disease (AD) than the age-matched non-dementia individuals. Given that there is no evidence that being overweight is associated with AD-type cognitive dysfunction, we hypothesized that moderate weight gain might have a protective effect on the bone loss in AD without exacerbating cognitive dysfunction. In this study, feeding a high-fat diet (HFD, 45% calorie from fat) to female APP/PS1 transgenic mice, an AD animal model, induced weight gain. The bone mineral density, microarchitecture, and biomechanical properties of the femurs were then evaluated. The results showed that the middle-aged female APP/PS1 transgenic mice were susceptible to osteoporosis of the femoral bones and that weight gain significantly enhanced bone mass and mechanical properties. Notably, HFD was not detrimental to brain insulin signaling and A?PP processing, as well as to exploration ability and working, learning, and memory performance of the transgenic mice measured by T maze and Morris water maze, compared with the mice fed a normal-fat diet (10% calorie from fat). In addition, the circulating levels of leptin but not estradiol were remarkably elevated in HFD-treated mice. These results suggest that a body weight gain induced by the HFD feeding regimen significantly improved bone mass in female APP/PS1 mice with no detriments to exploration ability and spatial memory, most likely via the action of elevated circulating leptin.
Prenatal stress induces cognitive functional impairment in offspring, an eventuality in which mitochondrial dysfunction and oxidative stress are believed to be closely involved. In this study, the involvement of the AMP-activated protein kinase (AMPK) pathway was investigated. A well-known activator, resveratrol (Res), was used to induce AMPK activation in SH-SY-5Y cells. Significant mitochondrial biogenesis and phase II enzyme activation, accompanied by decreased protein oxidation and GSSG content, were observed after Res treatment, and inhibition of AMPK with Compound c abolished the induction effects of Res. Further study utilizing a prenatal restraint stress (PRS) animal model indicated that maternal supplementation of Res may activate AMPK in the hippocampi of both male and female offspring, and that PRS-induced mitochondrial loss in the offspring hippocampus was inhibited by Res maternal supplementation. In addition, Res activated Nrf2-mediated phase II enzymes and reduced PRS-induced oxidative damage in both male and female offspring. Moreover, PRS markedly decreased mRNA levels of various neuron markers, as well as resultant offspring cognitive function, based on spontaneous alternation performance and Morris water maze tests, the results of which were significantly improved by maternal Res supplementation. Our results provide evidence indicating that AMPK may modulate mitochondrial content and phase II enzymes in neuronal cells, a process which may play an essential role in preventing PRS-induced cognitive impairment. Through the coupling of mitochondrial biogenesis and the Nrf2 pathway, AMPK may modulate oxidative stress and be a promising target against neurological disorders.
4-Methylene-2-octyl-5-oxotetrahydrofuran-3-carboxylic acid (C75) is a synthetic fatty-acid synthase (FASN) inhibitor with potential therapeutic effects in several cancer models. Human mitochondrial ?-ketoacyl-acyl carrier protein synthase (HsmtKAS) is a key enzyme in the newly discovered mitochondrial fatty acid synthesis pathway that can produce the substrate for lipoic acid (LA) synthesis. HsmtKAS shares conserved catalytic domains with FASN, which are responsible for binding to C75. In our study, we explored the possible effect of C75 on HsmtKAS and mitochondrial function. C75 treatment decreased LA content, impaired mitochondrial function, increased reactive oxygen species content, and reduced cell viability. HsmtKAS but not FASN knockdown had an effect that was similar to C75 treatment. In addition, an LA supplement efficiently inhibited C75-induced mitochondrial dysfunction and oxidative stress. Overexpression of HsmtKAS showed cellular protection against low dose C75 addition, whereas there was no protective effect upon high dose C75 addition. In summary, the mitochondrial fatty acid synthesis pathway has a vital role in mitochondrial function. Besides FASN, C75 might also inhibit HsmtKAS, thereby reducing LA production, impairing mitochondrial function, and potentially having toxic effects. LA supplements sufficiently ameliorated the toxicity of C75, showing that a combination of C75 and LA may be a reliable cancer treatment.
This study was to undertaken to investigate the impacts of AhR, CYP1A1, GSTM1 genetic polymorphisms on the R273G mutation in exon 8 of the tumor suppressor p53 gene (TP53) among polycyclic aromatic hydrocarbons (PAHs) exposed to coke-oven workers. One hundred thirteen workers exposed to PAH and 82 control workers were recruited. We genotyped for polymorphisms in the AhR, CYP1A1, GSTM1, and TP53 R273G mutation in blood by PCR methods, and determined the levels of 1-hydroxypyrene as PAH exposure marker in urine using the high pressure liquid chromatography assay. We found that the distribution of alcohol users and the urinary excretion of 1-OHP in the exposed workers were significantly higher than that of the control workers (p=0.004, p<0.001, respectively). Significant differences were observed in the p53 genotype distributions of smoking subjects (p=0.01, 95%CI: 1.23-6.01) and PAH exposure (p=0.008, 95%CI: 1.24-4.48), respectively. Further, significant differences were observed in the p53 exon 8 mutations for the genetic polymorphisms of Lys/Arg for AhR (p=0.02, 95%CI: 0.70-15.86), Val/Val for CYP1A1 (p=0.04, 95%CI: 0.98-19.09) and null for GSTM1 (p=0.02, 95%CI: 1.19-6.26), respectively. Our findings indicated that polymorphisms of PAH metabolic genes, such as AhR, CYP1A1, GSTM1 polymorphisms may interact with p53 genetic variants and may contribute to PAH related cancers.
FoxO1 acts as a pivotal transcription factor in insulin signaling. However, in hyperglycemia induced cardiac complications, whether FoxO1 is involved remains unclear. The goal of this study was to delineate the potential role of FoxO1 under high-glucose condition.
First proposed by D Harman in the 1950s, the Mitochondrial Free Radical Theory of Aging (MFRTA) has become one of the most tested and well-known theories in aging research. Its core statement is that aging results from the accumulation of oxidative damage, which is closely linked with the release of reactive oxygen species (ROS) from mitochondria. Although MFRTA has been well acknowledged for more than half a century, conflicting evidence is piling up in recent years querying the causal effect of ROS in aging. A critical idea thus emerges that contrary to their conventional image only as toxic agents, ROS at a non-toxic level function as signaling molecules that induce protective defense in responses to age-dependent damage. Furthermore, the peroxisome, another organelle in eukaryotic cells, might have a say in longevity modulation. Peroxisomes and mitochondria are two organelles closely related to each other, and their interaction has major implications for the regulation of aging. The present review particularizes the questionable sequiturs of the MFRTA, and recommends peroxisome, similarly as mitochondrion, as a possible candidate for the regulation of aging. Geriatr Gerontol Int 2014; 14: 740-749.
Punicalagin (PU) is one of the major ellagitannins found in the pomegranate (Punica granatum), which is a popular fruit with several health benefits. So far, no studies have evaluated the effects of PU on nonalcoholic fatty liver disease (NAFLD). Our work aims at studying the effect of PU-enriched pomegranate extract (PE) on high fat diet (HFD)-induced NAFLD.
Accumulating research has shown that chronic D-galactose (D-gal) exposure induces symptoms similar to natural aging in animals. Therefore, rodents chronically exposed to D-gal are increasingly used as a model for aging and delay-of-aging pharmacological research. Mitochondrial dysfunction is thought to play a vital role in aging and age-related diseases; however, whether mitochondrial dysfunction plays a significant role in mice exposed to D-gal remains unknown. In the present study, we investigated cognitive dysfunction, locomotor activity, and mitochondrial dysfunction involved in D-gal exposure in mice. We found that D-gal exposure (125?mg/kg/day, 8 weeks) resulted in a serious impairment in grip strength in mice, whereas spatial memory and locomotor coordination remained intact. Interestingly, muscular mitochondrial complex I deficiency occurred in the skeletal muscle of mice exposed to D-gal. Mitochondrial ultrastructure abnormality was implicated as a contributing factor in D-gal-induced muscular impairment. Moreover, three combinations (A, B, and C) of nutrients applied in this study effectively reversed D-gal-induced muscular impairment. Nutrient formulas B and C were especially effective in reversing complex I dysfunction in both skeletal muscle and heart muscle. These findings suggest the following: (1) chronic exposure to D-gal first results in specific muscular impairment in mice, rather than causing general, premature aging; (2) poor skeletal muscle strength induced by D-gal might be due to the mitochondrial dysfunction caused by complex I deficiency; and (3) the nutrient complexes applied in the study attenuated the skeletal muscle impairment, most likely by improving mitochondrial function.
Adipogenesis and lipid accumulation during aging have a great impact on the aging process and the pathogenesis of chronic, age-related diseases. However, little is known about the age-related molecular changes in lipid accumulation and the mechanisms underlying them. Here, using 5-month- and 25-month-old rats (young and old, respectively), we found that epididymal fat is the only tissue to accumulate during aging. By testing tissues rich with mitochondria in old and young animals, we found that the old animals had elevated levels of triglycerides in their muscle, heart and liver tissues but not in their kidneys, while, the mRNA level of fatty acid synthase remained unchanged among the four tissues. Regarding lipid catabolism, we determined that the activities of mitochondrial ETC. complexes changed in aged rats (muscle: decreased complex I and V activities; heart: decreased complex I activity; liver: increased complex I and III activities; kidney: decreased complex I and increased complex II activities), while changes in mitochondrial content were not observed in the muscle, heart nor in the liver tissue except increased complex IV and V subunits in aged kidneys. Furthermore, decreased mitochondrial fusion marker Mfn2 and decreased PGC-1? level were observed in the aged muscle, heart and liver but remained unchanged in the kidneys. Down-regulation of Mfn2 with siRNA in 293T cells induced significant mitochondrial dysfunction including decreased oxygen consumption, decreased ATP production, and increased ROS production, followed by increased triglyceride content suggesting a contributing role of decreased mitochondrial fusion to lipid deposit. Meanwhile, judging from autophagy marker p62/SQSTM1 and LC3-II, autophagy was suppressed in the aged muscle, heart and liver but remained unchanged in the kidneys. Taken together, these data suggest that reduction in PGC-1? expression and disruption of mitochondrial dynamics and autophagy might contribute to lipid accumulation during aging.
Superparamagnetic iron oxide nanoparticles (IONPs) have been widely applied in numerous biomedical fields. The evaluation of the toxicity of IONPs to the environment and human beings is indispensable to guide their applications. IONPs are usually considered to have good biocompatibility; however, some literatures have reported the toxicity of IONPs in vitro and in vivo. The controversy surrounding the biocompatibility of IONPs prompted us to carefully consider the biological effects of IONPs, especially under stress conditions. However, the potential risks of IONPs under stress conditions have not yet been evaluated in depth. Acrolein is widespread in the environment and modulates stress-induced gene activation and cell death in many organs and tissues. In this study, we assessed the sensitivity of H9c2 cardiomyocyte cells embedded with IONPs to acrolein and investigated the possible molecular mechanisms involved in this sensitivity. IONPs, which alone exhibited no toxicity, sensitized the H9c2 cardiomyocytes to acrolein-induced dysfunction. The IONP/acrolein treatment induced a loss of viability, membrane disruption, reactive oxygen species (ROS) generation, Erk activation, mitochondrial and lysosomal dysfunction, and necrosis in H9c2 cells. Treatment with an ROS generation inhibitor (diphenyleneiodonium) or an iron chelator (deferoxamine) prevented the IONP/acrolein-induced loss of viability, suggesting that ROS and IONP degradation facilitated the toxicity of the IONP/acrolein treatment in H9c2 cells. Our data suggest that cells embedded in IONPs are more vulnerable to oxidative stress, which confirms the hypothesis that nanoparticles can sensitize cells to the adverse effects of external stimulation. The present work provides a new perspective from which to evaluate the interactions between nanoparticles and cells.
Hypertension leads to cardiac hypertrophy as an adaptive response to increased workload. While initial development of hypertrophy is compensatory when contractile function is maintained, persistent stress on heart leads to deteriorated cardiac function and onset of heart failure. Mitochondrial dysfunction develops in the failing heart; however, whether it presents in compensatory cardiac hypertrophy is controversial.
We previously found that mitochondrial dysfunction occurs in disuse-induced muscle atrophy. However, the mitochondrial remodeling that occurs during reloading, an effective approach for rescuing unloading-induced atrophy, remains to be investigated. In this study, using a rat model of 3-week hindlimb unloading plus 7-day reloading, we found that reloading protected mitochondria against dysfunction, including mitochondrial loss, abnormal mitochondrial morphology, inhibited biogenesis, and activation of mitochondria-associated apoptotic signaling. Interestingly, a combination of nutrients, including ?-lipoic acid, acetyl-L-carnitine, hydroxytyrosol, and CoQ10, which we designed to target mitochondria, was able to efficiently rescue muscle atrophy via a reloading-like action. It is suggested that reloading ameliorates skeletal muscle atrophy through the activation of mitochondrial biogenesis and the amelioration of oxidative stress. Nutrient administration acted similarly in unloaded rats. Here, the study of mitochondrial remodeling in rats during unloading and reloading provides a more detailed picture of the pathology of muscle atrophy.
Copper oxide nanoparticles (CuONP) have attracted increasing attention due to their unique properties and have been extensively utilized in industrial and commercial applications. For example, their antimicrobial capability endows CuONP with applications in dressings and textiles against bacterial infections. Along with the wide applications, concerns about the possible effects of CuONP on humans are also increasing. It is crucial to evaluate the safety and impact of CuONP on humans, and especially the skin, prior to their practical application. The potential toxicity of CuONP to skin keratinocytes has been reported recently. However, the underlying mechanism of toxicity in skin cells has remained unclear. In the present work, we explored the possible mechanism of the cytotoxicity of CuONP in HaCaT human keratinocytes and mouse embryonic fibroblasts (MEF). CuONP exposure induced viability loss, migration inhibition, and G2/M phase cycle arrest in both cell types. CuONP significantly induced mitogen-activated protein kinase (extracellular signal-regulated kinase [Erk], p38, and c-Jun N-terminal kinase [JNK]) activation in dose- and time-dependent manners. U0126 (an inhibitor of Erk), but not SB 239063 (an inhibitor of p38) or SP600125 (an inhibitor of JNK), enhanced CuONP-induced viability loss. CuONP also induced decreases in p53 and p-p53 levels in both cell types. Cyclic pifithrin-?, an inhibitor of p53 transcriptional activity, enhanced CuONP-induced viability loss. Nutlin-3?, a p53 stabilizer, prevented CuONP-induced viability loss in HaCaT cells, but not in MEF cells, due to the inherent toxicity of nutlin-3? to MEF. Moreover, the experiments on primary keratinocytes are in accordance with the conclusions acquired from HaCaT and MEF cells. These data demonstrate that the activation of Erk and p53 plays an important role in CuONP-induced cytotoxicity, and agents that preserve Erk or p53 activation may prevent CuONP-induced cytotoxicity.
A Mediterranean diet rich in olive oil has profound influence on health outcomes including metabolic syndrome. However, the active compound and detailed mechanisms still remain unclear. Hydroxytyrosol (HT), a major polyphenolic compound in virgin olive oil, has received increased attention for its antioxidative activity and regulation of mitochondrial function. Here, we investigated whether HT is the active compound in olive oil exerting a protective effect against metabolic syndrome. In this study, we show that HT could prevent high-fat-diet (HFD)-induced obesity, hyperglycemia, hyperlipidemia, and insulin resistance in C57BL/6J mice after 17 weeks supplementation. Within liver and skeletal muscle tissues, HT could decrease HFD-induced lipid deposits through inhibition of the SREBP-1c/FAS pathway, ameliorate HFD-induced oxidative stress by enhancing antioxidant enzyme activities, normalize expression of mitochondrial complex subunits and mitochondrial fission marker Drp1, and eventually inhibit apoptosis activation. Moreover, in muscle tissue, the levels of mitochondrial carbonyl protein were decreased and mitochondrial complex activities were significantly improved by HT supplementation. In db/db mice, HT significantly decreased fasting glucose, similar to metformin. Notably, HT decreased serum lipid, at which metformin failed. Also, HT was more effective at decreasing the oxidation levels of lipids and proteins in both liver and muscle tissue. Similar to the results in the HFD model, HT decreased muscle mitochondrial carbonyl protein levels and improved mitochondrial complex activities in db/db mice. Our study links the olive oil component HT to diabetes and metabolic disease through changes that are not limited to decreases in oxidative stress, suggesting a potential pharmaceutical or clinical use of HT in metabolic syndrome treatment.
Total paeony glucosides (TPG) extracted from the roots of Radix Paeoniae Rubrae, have been approved for the therapy of rheumatoid arthritis by the State Food and Drug Administration. We previously demonstrated the myocardial protective effects of TPG in both isoprenaline-induced myocardial ischemia rat and acute myocardial infarction rat. However, the underlying mechanism of TPG effect in cardiomyocytes remains to be investigated. The aims of this study were to elucidate the effect of TPG on the activities of antioxidant defense targets and the bioenergetic system in rat cardiomyocytes. The changes of viability, antioxidant defense system activities, protein contents, and mitochondrial functions in tert-butyl hydroperoxide challenged H9c2 rat cardiomyoblasts were evaluated. The results suggest that TPG ameliorated cardiomyoblast dysfunction by preserving antioxidant defense and bioenergetic system.
Cigarette smoking causes various diseases, including lung cancer and cardiovascular disease, and reduces life span, though the mechanisms are not well understood. We hypothesize that smoking may cause cellular mitochondrial dysfunction and oxidative stress, leading to aging acceleration. In the present study, we tested the effects of acrolein, a major representative smoking toxicant, on human lung fibroblast IMR-90 cells with regard to cellular senescence, oxidative stress, and mitochondrial function. The results showed that subacute treatment with low dose of acrolein induces the following events compared to the control cells: cell senescence demonstrated by increases in the activity of ?-galactosidase, the higher expression of p53 and p21, decreases in DNA synthesis, Sirt1 expression, and telomere length; oxidative stress occurred as the increases in the production of reactive oxygen species, DNA damage, and protein oxidation; and mitochondrial dysfunction shown as decreases in the mitochondrial membrane potential, mitochondrial biogenesis regulator PGC-1 alpha and mitochondria complex I, II, III, and V. These results suggest that acrolein may accelerate aging through the mechanism of increasing oxidative stress and mitochondrial dysfunction.
Mitochondrial dynamics maintains normal mitochondrial function by degrading damaged mitochondria and generating newborn mitochondria. The accumulation of damaged mitochondria influences the intracellular environment by promoting mitochondrial dysfunction, and thus initiating a vicious cycle. Oxidative stress induces mitochondrial malfunction, which is involved in many cardiovascular diseases. However, the mechanism of mitochondrial accumulation in cardiac myoblasts remains unclear. We observed mitochondrial dysfunction and an increase in mitochondrial mass under the oxidative conditions produced by tert-butyl hydroperoxide (tBHP) in cardiac myoblast H9c2 cells. However, in contrast to the increase in mitochondrial mass, mitochondrial DNA (mtDNA) decreased, suggesting that enhanced mitochondrial biogenesis may be not the primary cause of the mitochondrial accumulation. Therefore, we investigated changes in a number of proteins involved in autophagy. Beclin1, Atg12-Atg5 conjugate, Atg7 contents decreased but LC3-II accumulated in tBHP-treated H9c2 cells. Moreover, the capacity for acid hydrolysis decreased in H9c2 cells. We also demonstrated a decrease in DJ-1 protein under the oxidative conditions that deregulate mitochondrial dynamics. These results reveal that autophagy became defective under oxidative stress. We therefore suggest that defects in autophagy mediate mitochondrial accumulation under these conditions.
Hydroxytyrosol, an important polyphenolic compound found in olive oil, has shown anti-tumor activity both in vitro and in vivo. However, effects of hydroxytyrosol on prostate cancer are largely unkown. We found that hydroxytyrosol preferentially reduces the viability of human prostate cancer cells (PC-3, DU145) compared to an immortalized non-malignant prostate epithelial cell line (RWPE-1). Exposure of PC-3 cells to 80 µmol/L hydroxytyrosol resulted in significant increases in both superoxide production and activation of apoptosis. These increases were accompanied by mitochondrial dysfunction, defects in autophagy, and activation of MAP kinases. Moreover, N-acetylcysteine (NAC), an efficient reactive oxygen species (ROS) scavenger, was able to reverse the hydroxytyrosol-induced effects of cell viability loss, defects in autophagy, and activation of apoptosis. This evidence suggests that ROS play a vital role in the loss of PC-3 cell viability. However, MAPK inhibitors including U0126 (for Erk1/2), SB203580 (for p38MAPK) and SP600125 (for JNK) did not decrease hydroxytyrosol-induced growth inhibition, suggesting that these kinases may not be required for the growth inhibitory effect of hydroxytyrosol. Moreover, addition of ROS scavengers (i.e. NAC, catalase, pyruvate, SOD) in the growth media can prevent hydroxytyrosol induced cell viability loss, suggesting that extracellular ROS (superoxide and hydrogen peroxide) facilitate the anti-proliferation effect of hydroxytyrosol in prostate cancer cells. The present work firstly shows that hydroxytyrosol induces apoptotic cell death and mitochondrial dysfunction by generating superoxide in PC-3 cells. This research presents preliminary evidence on the in vitro chemopreventive effect of hydroxytyrosol, and will contribute to further investigation of hydroxytyrosol as an anti-cancer agent.
Benzo[a]pyrene (BaP) is a representative compound of polycyclic aromatic hydrocarbons exerting cytotoxicity and genotoxicity in the human liver, lung, stomach and skin. However, the toxic effect of BaP on cervical tissue remains unclear. This study was carried out to investigate the toxic effects of BaP on the cervix of ICR mice. Female mice were treated with BaP by intraperitoneal injection and oral gavage at a dose of 2.5, 5 and 10 mg/kg body-weight, twice a week for 14 weeks. BaP treatment caused a significant increase in the levels of MDA and IL-6 with significantly increased activity of CYP1A1, creatine kinase and aspartate aminotransferase (AST) and decreased activity of glutathione-S-transferase in the cervix and liver. The relative cervix weight was markedly reduced in the intraperitoneal BaP injection groups, whereas only a slight reduction was observed in the oral gavage groups. The increase in weight decreased with increasing BaP dose. Moreover, BaP treatment induced significant pathomorphological changes in the cervical tissue and increased the mortality of the mice. Taken together, these results suggest that BaP causes a certain toxic effect on cervical tissue.
We developed a complex dietary supplement designed to offset five key mechanisms of aging and tested its effectiveness in ameliorating age-related cognitive decline using a visually cued Morris water maze test. All younger mice (<1 year old) learned the task well. However, older untreated mice (>1 year) were unable to learn the maze even after 5 days, indicative of strong cognitive decline at older ages. In contrast, no cognitive decline was evident in older supplemented mice, even when ?2 years old. Supplemented older mice were nearly 50% better at locating the platform than age-matched controls. Brain weights of supplemented mice were significantly greater than controls, even at younger ages. Reversal of cognitive decline in activity of complexes III and IV by supplementation was significantly associated with cognitive improvement, implicating energy supply as one possible mechanism. These results represent proof of principle that complex dietary supplements can provide powerful benefits for cognitive function and brain aging.
Treatment with a combination of four nutrients, i.e. R-?-lipoic acid, acetyl-l-carnitine, nicotinamide and biotin, just as with pioglitazone, significantly improves glucose tolerance, insulin release, plasma NEFA, skeletal muscle mitochondrial biogenesis and oxidative stress in Goto-Kakizaki (GK) rats. However, it is not known whether treatment with these nutrients can improve mitochondrial function and reduce oxidative stress in GK rats. The effects of a combination of these four nutrients on mitochondrial function, oxidative stress and apoptosis in GK rat liver were investigated. Livers of untreated GK rats showed (1) abnormal changes in the activities of mitochondrial complexes (decreases in I, III and IV and increases in II and V), (2) increases in protein oxidation, (3) decreases in antioxidant enzymes (superoxide dismutase, glutathione S-transferase, NADH-quinone oxidoreductase-1), (4) a decrease in total antioxidant capacity but increases in reduced glutathione level and glyceraldehyde 3-phosphate dehydrogenase expression and (5) significant increases in apoptosis biomarkers, including expression of p21 and p53. A 3-month treatment with the four nutrients significantly improved most of these abnormalities in GK rats, and the effects of the nutrient combination were greater than those of pioglitazone for most of these indices. These results suggest that dietary supplementation with nutrients that are thought to influence mitochondrial function may be an effective strategy for improving liver dysfunction in GK diabetic rats.
Oxidative stress and mitochondrial dysfunction are known to play important roles in type 2 diabetes mellitus (T2DM) and insulin resistance. However, the pathology of T2DM remains complicated; in particular, the mechanisms of mitochondrial dysfunction in skeletal muscle and other insulin-sensitive tissues are as yet unclear. In the present study, we investigated the underlying mechanisms of oxidative stress and mitochondrial dysfunction by focusing on mitochondrial dynamics, including mitochondrial biogenesis and autophagy, in skeletal muscle of a nonobese diabetic animal model--the Goto-Kakizaki (GK) rat. The results showed that GK rats exhibited impaired glucose metabolism, increased oxidative stress and decreased mitochondrial function. These dysfunctions were found to be associated with induction of LC3B, Beclin1 and DRP1 (key molecules mediating the autophagy pathway), while they appeared not to affect the mitochondrial biogenesis pathway. In addition, (-)-epigallocatechin-3-gallate (EGCG) was tested as a potential autophagy-targeting nutrient, and we found that EGCG treatment improved glucose tolerance and glucose homeostasis in GK rats, and reduced oxidative stress and mitochondrial dysfunction in skeletal muscle. Amelioration of excessive muscle autophagy in GK rats through the down-regulation of the ROS-ERK/JNK-p53 pathway leads to improvement of glucose metabolism, reduction of oxidative stress and inhibition of mitochondrial loss and dysfunction. These results suggest (a) that hyperglycemia-associated oxidative stress may induce autophagy through up-regulation of the ROS-ERK/JNK-p53 pathway, which may contribute to mitochondrial loss in soleus muscle of diabetic GK rats, and (b) that EGCG may be a potential autophagy regulator useful in treatment of insulin resistance.
Benzo[a]pyrene (BaP) is cytotoxic and/or genotoxic to lung, stomach and skin tissue in the body. However, the effect of BaP on cervical tissue remains unclear. The present study detected DNA damage and the expression of the p53 gene in BaP-induced cervical tissue in female mice. Animals were intraperitoneally injected and orally gavaged with BaP at the doses of 2.5, 5, and 10mg/kg twice a week for 14 weeks. The single-cell gel electrophoresis (SCGE) assay was used to detect the DNA damage. Immunohistochemistry (IHC) and in situ hybridization (ISH) were used to detect the expression of p53 protein and p53 mRNA, respectively. The results showed that BaP induced a significant and dose-dependent increase of the number of cells with DNA damaged and the tail length as well as Comet tail moment in cervical tissue. The expression level of p53 protein and mRNA was increased. The results demonstrate that BaP may show toxic effect on the cervix by increasing DNA damage and the expression of the p53 gene.
As brain testosterone plays both androgenic and estrogenic actions due to its conversion into estrogen via aromatase naturally, it is unclear that the age-related reduction of testosterone increased risk of Alzheimers disease (AD) in men is mediated through androgen alone or both androgen and estrogen mechanisms. Our previous studies using a gene-based approach in mouse model to block the conversion of testosterone into estrogen (aromatase gene knock-out, ArKO), found a depletion of estrogen and increase in testosterone endogenously in males. Here, we use crossing the ArKO mice with APP23 transgenic mice, a mouse model of AD, to produce APP23/Ar(+/-) mice to study the estrogen-independent effect of testosterone on AD. We found a significant reduction in brain plaque formation, improved cognitive function and increase NEP activity in male APP23/Ar(+/-) mice compared with age-matched male APP23 controls. In addition, we found, for the first time, a reduction of beta-secretase (BACE1) enzyme activity, mRNA level and protein expression in the male APP23/Ar(+/-) mice, suggesting that endogenous testosterone, independent from estrogen, may protect against AD in males via two major mechanisms, downregulation of BACE1 activities at transcriptional level to reduce beta amyloid production and upregulation of NEP activities to enhance bate amyloid degradation.
Benzo[a]pyrene (BaP) is a polycyclic aromatic hydrocarbon and a potent inducer of carcinogenesis. Many studies have reported that the carcinogenic effects of BaP might be due to its intermediate metabolites and to reactive oxygen species (ROS) that cause oxidative damage to the cells. However, the mechanisms of BaP-induced oxidative damage in cervical tissue are still not clear. We studied these mechanisms in female ICR mice treated with BaP either orally or intraperitoneally by measuring (1) several general biomarkers of oxidative stress in serum, (2) mitochondrial function in the cervix, and (3) the morphology of mitochondria in cervical tissue. BaP treatment (1) significantly lowered levels of vitamins A, C, and E and of glutathione; (2) reduced activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione S-transferases; and (3) significantly increased lipid peroxidation levels. In addition, significant increases in the levels of superoxide anion, hydrogen peroxide, and hydroxyl radical were observed. These results were confirmed by morphological changes in mitochondria and by decreases in membrane potential levels and in succinate dehydrogenase and malate dehydrogenase activities. The changes in these biomarkers and mitochondrial damage were BaP-dose-dependent and eventually induced both cell apoptosis and necrosis in cervical tissue. As mitochondria are the major sites of ROS generation, these findings show that mitochondrial decay greatly contributes to BaP-induced cervical damage.
Benzo[a]pyrene [B(a)P] is one of the most prevalent environmental carcinogens and genotoxic agents. However, the mechanisms of B(a)P-induced oxidative damage in cervical tissue are still not clear. The present study was to investigate the oxidative stress and DNA damage in cervix of ICR female mice induced by acute treatment with B(a)P. Oxidative stress was assayed by analysis of malondialdehyde (MDA), superoxide anion and H(2)O(2), and antioxidant enzymes. The alkaline single-cell electrophoresis (SCGE) was used to measure DNA damage. The contents of MDA and glutathione (GSH), and the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione S-transferase (GST) were significantly increased in cervix 24, 48 and 72h after B(a)P treatment of a single dose of 12.5 and 25mg/kg, while GSH, CAT, SOD and GST had no significant difference with the dose of 50mg/kg B(a)P at post-treatment time 48 and 72h except for SOD activity at 48h which was significant. The maximum values of SOD, CAT, GST and GSH were peaked at 24h and then decreased gradually while GPx activities and MDA levels persisted for up to 72h. Superoxide anion, H(2)O(2) and DNA damage changed similarly as the activity of SOD, CAT or GST. Additionally, increases of formamidopyrimidine DNA glycosylase (FPG) specific DNA damage were observed and can be greatly rescued by vitamin C pretreatment. Overall, B(a)P demonstrated a time- and dose- related oxidative stress and DNA damage in cervix.
Aging degrades motivation, cognition, sensory modalities and physical capacities, essentially dimming zestful living. Bradykinesis (declining physical movement) is a highly reliable biomarker of aging and mortality risk. Mice fed a complex dietary supplement (DSP) designed to ameliorate five mechanisms associated with aging showed no loss of total daily locomotion compared with >50% decrement in old untreated mice. This was associated with boosted striatal neuropeptide Y, reversal of age-related declines in mitochondrial complex III activity in brain and amelioration of oxidative stress (brain protein carbonyls). Supplemented mice expressed approximately 50% fewer mitochondrial protein carbonyls per unit of complex III activity. Reduction of free radical production by mitochondria may explain the exceptional longevity of birds and dietary restricted animals and no DSP is known to impact this mechanism. Functional benefits greatly exceeded the modest longevity increases documented for supplemented normal mice. Regardless, for aging humans maintaining zestful health and performance into later years may provide greater social and economic benefits than simply prolonging lifespan. Although identifying the role of specific ingredients and interactions remains outstanding, results provide proof of principle that complex dietary cocktails can powerfully ameliorate biomarkers of aging and modulate mechanisms considered ultimate goals for aging interventions.
A botanical extract (Regrapex-R) prepared from whole grape (Vitis vinifera) and Polygonum cuspidatum, which contains polyphenols, including flavans, anthocyanins, emodin, and resveratrol, exhibited dose-dependent scavenging effects on reactive oxygen species (ROS). The extract inhibited increases of ROS and protein carbonyl in isolated rat liver mitochondria following exposure to 2,2-azobis (2-amidino propane) dihydrocholoride (AAPH), a potent lipid oxidant generator. The antioxidant effects of this extract were further demonstrated by protecting enzyme activities of the mitochondrial respiratory electron transport chain (complexes I and II) and pyruvate dehydrogenase in isolated liver mitochondria with AAPH insult. In human neuroblastoma cells (SKN-MC), pretreatment of extract protected cells against AAPH induced oxidation in maintaining cell viability and inhibiting excessive ROS generation. Extract was fed to transgenic Drosophila expressing human alpha-synuclein. This model for Parkinson disease recapitulates essential features of the disorder, including loss of dopaminergic neurons in the substantia nigra and a locomotor dysfunction that is displayed by a progressive loss of climbing ability measured using a geotaxis assay. Male transgenic flies fed the extract (0.16-0.64 mg/100 g of culture medium) showed a significant improvement in climbing ability compared to controls. Female transgenic flies showed a significant extension in average lifespan. These results suggest that Regrapex-R is a potent free radical scavenger, a mitochondrial protector, and a candidate for further studies to assess its ability to protect against neurodegenerative disease and potentially extend lifespan.
The development of type 2 diabetes is accompanied by decreased immune function and the mechanisms are unclear. We hypothesize that oxidative damage and mitochondrial dysfunction may play an important role in the immune dysfunction in diabetes. In the present study, we investigated this hypothesis in diabetic Goto-Kakizaki rats by treatment with a combination of four mitochondrial-targeting nutrients, namely, R-alpha-lipoic acid, acetyl-L-carnitine, nicotinamide and biotin. We first studied the effects of the combination of these four nutrients on immune function by examining cell proliferation in immune organs (spleen and thymus) and immunomodulating factors in the plasma. We then examined, in the plasma and thymus, oxidative damage biomarkers, including lipid peroxidation, protein oxidation, reactive oxygen species, calcium and antioxidant defence systems, mitochondrial potential and apoptosis-inducing factors (caspase 3, p53 and p21). We found that immune dysfunction in these animals is associated with increased oxidative damage and mitochondrial dysfunction and that the nutrient treatment effectively elevated immune function, decreased oxidative damage, enhanced mitochondrial function and inhibited the elevation of apoptosis factors. These effects are comparable to, or greater than, those of the anti-diabetic drug pioglitazone. These data suggest that a rational combination of mitochondrial-targeting nutrients may be effective in improving immune function in type 2 diabetes through enhancement of mitochondrial function, decreased oxidative damage, and delayed cell death in the immune organs and blood.
Malondialdehyde (MDA) is a product of oxidative damage to lipids, amino acids and DNA, and accumulates with aging and diseases. MDA can possibly react with amines so as to modify proteins and inactivate enzymes; it can also modify nucleosides so as to cause mutagenicity. Brain mitochondrial dysfunction is a major contributor to aging and neurodegenerative diseases. We hypothesize that MDA accumulated during aging targets mitochondrial enzymes so as to cause further mitochondrial dysfunction and additional contributions to aging and neurodegeneration. Herein, we investigated the neuronal mitochondrial toxic effects of MDA on mitochondrial respiration and activities of enzymes (mitochondrial complexes I-V, alpha-ketoglutarate dehydrogenase (KGDH) and pyruvate dehydrogenase (PDH)), in isolated rat brain mitochondria. MDA depressed mitochondrial membrane potential, and also showed a dose-dependent inhibition of mitochondrial complex I- and complex II-linked respiration. Complex I and II, and PDH activities were depressed by MDA at >or=0.2 micromol/mg; KGDH and complex V were inhibited by >or=0.4 and >or=1.6 micromol MDA/mg, respectively. However, MDA did not have any toxic effects on complex III and IV activities over the range 0-2 micromol/mg. MDA significantly elevated mitochondrial reactive oxygen species (ROS) and protein carbonyls at 0.2 and 0.002 micromol/mg, respectively. As for the antioxidant defense system, a high dose of MDA slightly decreased mitochondrial GSH and superoxide dismutase. These results demonstrate that MDA causes neuronal mitochondrial dysfunction by directly promoting generation of ROS and modifying mitochondrial proteins. The results suggest that MDA-induced neuronal mitochondrial toxicity may be an important contributing factor to brain aging and neurodegenerative diseases.
To investigate the mitochondrial decay and oxidative damage resulting from aging, the activities/kinetics of the mitochondrial complexes were examined in the brains of young and old rats as well as in old rats fed R-alpha-lipoic acid plus acetyl-L-carnitine (LA/ALC). The brain mitochondria of old rats, compared with young rats, had significantly decreased endogenous antioxidants and superoxide dismutase activity; more oxidative damage to lipids and proteins; and decreased activities of complex I, IV and V. Complex I showed a decrease in binding affinity (increase in K(m)) for substrates. Feeding LA/ALC to old rats partially restored age-associated mitochondrial dysfunction to the levels of the young rats. These results indicate that oxidative mitochondrial decay plays an important role in brain aging and that a combination of nutrients targeting mitochondria, such as LA/ALC, could ameliorate mitochondrial decay through preventing mitochondrial oxidative damage.
Astragalus mongholicus (AM) is a traditional medicinal herb used as a neuroprotective agent for its anxiolytic, antidepressant, antiamnestic, and antiaggresive effects. However, the mechanisms underlying its anti-convulsant properties are not well studied. In the present study, we examined the anticonvulsant effects on pentylenetetrazol (PTZ)-induced seizures in mice and the possible mechanisms of protection against oxidative damage and mitochondrial dysfunction in vitro. The behavioral studies showed that the root extract of AM had powerful anticonvulsant effects against seizures induced by PTZ and the biochemical studies showed that root extract of AM inhibited PTZ-induced increase in lipid peroxidation, protein oxidation and reactive oxygen species, and enhanced mitochondrial function. Electron spin resonance spectroscopy studies demonstrated that the extracts from the root and aerial parts of AM possess potent effects on scavenging hydroxyl and lipid free radicals. We found that AM extract significantly protected malondialdehyde-induced oxidative damage by ameliorating activities of the mitochondrial complexes I, II, malate dehydrogenase and mitochondrial membrane potential. These data suggest that the anti-convulsant effects of AM extract may be mediated by its protective actions against oxidative damage and amelioration of mitochondrial dysfunction.
Plants are a promising expression system for the production of recombinant proteins. However, low protein productivity remains a major obstacle that limits extensive commercialization of whole plant and plant cell bioproduction platform. Plastid genetic engineering offers several advantages, including high levels of transgenic expression, transgenic containment via maternal inheritance, and multigene expression in a single transformation event. In recent years, the development of optimized expression strategies has given a huge boost to the exploitation of plastids in molecular farming. The driving forces behind the high expression level of plastid bioreactors include codon optimization, promoters and UTRs, genotypic modifications, endogenous enhancer and regulatory elements, posttranslational modification, and proteolysis. Exciting progress of the high expression level has been made with the plastid-based production of two particularly important classes of pharmaceuticals: vaccine antigens, therapeutic proteins, and antibiotics and enzymes. Approaches to overcome and solve the associated challenges of this culture system that include low transformation frequencies, the formation of inclusion bodies, and purification of recombinant proteins will also be discussed.
Mitochondrial dynamics is highly involved in muscle atrophy, the slow twitch muscle as soleus, preferentially affected by hindlimb unloading (HU), was well characterized by mitochondrial dysfunction in biogenesis. However, the fast twitch muscles like tibialis anterior (TA) and gastrocnemius (GAS), which are the most massive parts of the hindlimb muscles, are less elucidated on mitochondrial adaptations responding to HU. To investigate the mitochondrial dynamic responses and the involved molecules mediating atrophy in TA and GAS, we studied a 4-week HU mouse model. We found GAS was preferentially affected to atrophy by unloading compared with TA. Furthermore, the depressed mitochondrial biogenesis occurred, accounting for mitochondrial loss in GAS by unloading. PGC-1?, as well as its transcriptional/post-translational modification regulators, such as p-CREB, SIRT1, and p-AMPK, were consistently reduced in response to unloading in GAS. Molecules relevant to autophagy, mitochondrial fusion, and fission, were compromised following unloading both in TA and GAS. These results suggested that TA exhibited resistance to unloading induced muscle atrophy while GAS presented significant mitochondrial loss, which might be due to the mitochondrial biogenesis suppressed by the inactivation of PGC-1?. However, both in TA and GAS muscles, a similar sedentary mitochondrial dynamics of mitochondrial fusion and fission was induced by unloading though TA exhibited little muscle atrophy.
We examined whether transgenic growth hormone mice (Tg) that exhibit accelerated cognitive aging and exceptional free radical damage also express elevated nitrative stress. We characterized age-related patterns of 3-nitrotyrosine (3-NT) in brain homogenate and mitochondria of Tg and normal (Nr) mice as modulated by a complex anti-aging dietary supplement. Levels of 3-NT rose rapidly with age in Tg brain homogenate whereas normal controls maintained constant lower levels. The age-related slope for 3-NT was 3.6-fold steeper in untreated Tg compared to treated Tg (p<0.009), although treated Tg showed elevation in youth. Opposite to Tg, treated Nr mice had reduced 3-NT in youth (p<0.02). The age-related pattern of mitochondrial 3-NT in Nr mice was parabolic (p<0.005). Remarkably, levels in treated Nr were reduced by ~50% (p<0.0007). Untreated Tg showed strongly increasing mitochondrial 3-NT with higher mitochondrial activity (p<0.01) whereas treated Tg showed lower nitrosylation at higher levels of mitochondrial activity. Tg mice also expressed a postural abnormality that is a biomarker of neurodegeneration and/or nitrative stress. Tg represent a promising new model of nitrative stress associated with brain deterioration and results provide proof of principle that complex dietary supplements may be ameliorating.
Paeoniae radix is a traditional Chinese medicinal herb for treating some diseases; important components are total paeony glycosides (TPGs), an approved drug by the State Food and Drug Administration (SFDA) for the therapy of rheumatoid arthritis (RA). We firstly reported myocardial benefits of TPGs previously, and the present study is to further investigate the underlying mechanisms for preventing oxidative damage in cardiomyopathy. We measured the capacity of TPGs to scavenge free radicals in vitro. Then 60 SD rats were randomly divided into five groups: (1) a normal control group, (2) an isoprenaline (ISO)-induced myocardial ischemic model group, (3) a TPG treatment group (TPGs 269.4 mg/kg delivered by intragastric administration for 3 days before ISO administration and TPGs 449 mg/kg delivered for 3 days after ISO administration), (4) a TPG therapy group (TPGs 449 mg/kg delivered for 3 days after ISO administration), and (5) a positive control group (propranolol 15 mg/kg for 3 days after ISO administration). The ISO-induced myocardial ischemic model was established by subcutaneous injection of 1mg/kg/8h ISO (2 times). The activities of myocardial enzymes, including glutamic oxaloacetic transaminase (GOT), creatine kinase (CK), lactate dehydrogenase (LDH), antioxidant enzyme superoxide dismutase (SOD) as well as the content of lipid peroxidation product malondialdehyde (MDA) were detected. We found that TPGs potently eliminated hydroxyl radicals and superoxide in vitro using ESR assays. Compared with model rats, TPG treatment, TPG therapy and the positive control treatment exhibited significantly reduced activities of GOT, LDH, and CK (p < 0.01), increased activity of SOD (p < 0.01) and lower levels of MDA (p < 0.05). More interestingly, the protective effect of TPG treatment was even better than that of propranolol. These results suggest that TPGs significantly ameliorate ISO-induced myocardial ischemia and their action might be through reducing oxidative stress in ischemic myocardium.
Prostate carcinoma is one of the leading causes of cancer-related morbidity and mortality in males in western countries. Curcumin exhibits growth-suppressive activity against several cancers, including prostate cancer, but it has poor bioavailability. The purpose of this study was to evaluate the anticancer potency and mechanism of a curcumin analogue, 1,5-bis(3-hydroxyphenyl)-1,4-pentadiene-3-one (Ca 37), in human prostate cancer. Studies were performed in established human prostate cancer cell lines (PC-3 and DU145) as well as in a murine xenograft tumor (PC-3) model. Ca 37 presented a preferential suppression capacity against growth and migration toward prostate cancer cells compared with curcumin. Ca 37 impaired the bioenergetics system, promoted cell cycle arrest and apoptosis activation in PC-3 cells. In addition, 0.5 ?mol (6.65 mg/kg body weight) of Ca 37 significantly inhibited the growth of the prostate xenografted tumors, whereas 6 ?mol (110 mg/kg body weight) of curcumin had little effect. Furthermore, a combination of Ca 37 and curcumin resulted in enhanced antitumor activity in prostate cancer cells. N-Acetylcysteine abrogated both reactive oxygen species (ROS) production and viability loss induced by Ca 37 but partially prevented growth inhibition in PC-3 cells treated with curcumin alone, or a combination with Ca 37. The data indicate that induction of ROS plays a vital role in the growth inhibitory effect of Ca 37 in PC-3 cells. This study suggests that Ca 37, alone or in combination with curcumin, may be a promising anticancer agent for prostate cancer therapy.
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