[Show abstract][Hide abstract] ABSTRACT: Background
Methionine, an essential amino acid, is required for protein synthesis and normal cell metabolism. The transmethylation pathway and methionine salvage pathway (MTA cycle) are two major pathways regulating methionine metabolism. Recently, methionine has been reported to play a key role in Drosophila fecundityResultsHere, we revealed that the MTA cycle plays a crucial role in Drosophila fecundity using the mutant of aci-reductone dioxygenase 1 (DADI1), an enzyme in the MTA cycle. In dietary restriction condition, the egg production of adi1 mutant flies was reduced compared to that of control flies. This fecundity defect in mutant flies was rescued by reintroduction of Dadi1 gene. Moreover, a functional homolog of human ADI1 also recovered the reproduction defect, in which the enzymatic activity of human ADI1 is required for normal fecundity. Importantly, methionine supply rescued the fecundity defect in Dadi1 mutant flies. The detailed analysis of Dadi1 mutant ovaries revealed a dramatic change in the levels of methionine metabolism. In addition, we found that three compounds namely, methionine, SAM and Methionine sulfoxide, respectively, may be required for normal fecundityConclusions
In summary, these results suggest that ADI1, an MTA cycle enzyme, affects fly fecundity through the regulation of methionine metabolism.
[Show abstract][Hide abstract] ABSTRACT: Abstract Glucose-6-phosphate dehydrogenase (G6PD) is critical to the maintenance of NADPH pool and redox homeostasis. Coventionally, G6PD deficiency has been associated with hemolytic disorders. Most biochemical variants were identified and characterized at molecular level. Recently, a number of studies have shone light on the roles of G6PD in aspects of physiology other than erythrocytic pathophysiology. G6PD deficiency alters the redox homeostasis, and affects dysfunctional cell growth and signaling, anomalous embryonic development, and altered susceptibility to infection. The present article gives a brief review of basic science and clinical findings about G6PD, and covers the latest development in the field. Moreover, how G6PD status alters the susceptibility of the affected individuals to certain degenerative diseases is also discussed.
Free Radical Research 04/2014; · 3.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Spinocerebellar ataxia (SCAs) types 1, 2, 3, 6, 7, 17, and dentatorubropallidoluy-sianatrophy (DRPLA) as well as Huntington's disease (HD) are a group of neurodegenerative disorders caused by a CAG triplet repeat expansion encoding a long polyglutamine (polyQ) tract in the respective mutant proteins. The cytoplasmic and nuclear aggregate formation, pathological hallmark of polyQ diseases, is likely the initial process to trigger the subsequent pathological events. Compromised oxidative stress defense capacity and mitochondrial dysfunction have emerged as contributing factors to the pathogenesis of polyQ diseases. The roots of licorice (Glycyrrhiza) species have long been used as an herbal medicine. In the present study, we demonstrated the aggregate-inhibitory effect of Glycyrrhiza inflata (G. inflata) herb extract and its constituents licochalcone A and ammonium glycyrrhizinate (AMGZ) in both 293 and SH-SY5Y ATXN3/Q75 cells, SCA3 cell models. The reporter assay showed that G. inflata herb extract, licochalcone A and AMGZ could enhance the promoter activity of peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PPARGC1A), a known regulator of mitochondrial biogenesis and anti-oxidative response genes. G. inflata extract, licochalcone A and AMGZ up-regulated PPARGC1A expression and its downstream target genes, SOD2 and CYCS, in 293 ATXN3/Q75 cell model. The expression of nuclear factor erythroid 2-related factor 2 (NFE2L2), the principal transcription factor that binds to anti-oxidant responsive elements (ARE) to promote ARE-dependent gene expression when the cells respond to oxidative stress, and its downstream genes, HMOX1, NQO1, GCLC and GSTP1 were also increased by G. inflata herb extract, licochalcone A and AMGZ. Knock down of PPARGC1A increased aggregates in ATXN3/Q75 cells and also attenuated the aggregate-inhibiting effect of the tested compounds. G. inflata extract and its constituents significantly elevated GSH/GSSG ratio and reduced reactive oxidative species in ATXN3/Q75 cells. The study results suggest that the tested agents activate PPARGC1A activity and NFE2L2-ARE signaling to increase mitochondrial biogenesis, decrease oxidative stress, and reduce aggregate formation in SCA3 cellular models.
Free Radical Biology & Medicine 03/2014; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glucose-6-phosphate dehydrogenase (G6PD) is pivotal to reduced nicotinamide adenine dinucleotide phosphate (NADPH) production and cellular redox balance. Cells with G6PD deficiency are susceptible to oxidant-induced death at high oxidative stress. However, it remains unclear what precise biological processes are affected by G6PD deficiency due to altered cellular redox homeostasis, particularly at low oxidative stress. To further explore the biological role of G6PD, we generated G6PD-knockdown cell clones using lung cancer line A549. We identified proteins differentially expressed in the knockdown clones without the addition of exogenous oxidant by means of isobaric tags for relative and absolute quantification (iTRAQ) labeling coupled with multidimensional liquid chromatography–mass spectrometry (LC–MS/MS). We validated a panel of proteins that showed altered expression in G6PD-knockdown clones and were involved in metabolism of xenobiotic and glutathione (GSH) as well as energy metabolism. To determine the physiological relevancy of our findings, we investigated the functional consequence of G6PD depletion in cells treated with a prevalent xenobiotic, aflatoxin B1 (AFB1). We found a protective role of G6PD in AFB1-induced cytotoxicity, possibly via providing NADPH for NADPH oxidase to induce epoxide hydrolase 1 (EPHX1), a xenobiotic-metabolizing enzyme. Collectively, our findings reveal for the first time a proteome-wide dysregulation by G6PD depletion under the condition without exogenous oxidant challenge, and we suggest a novel association of G6PD activity with AFB1-related xenobiotic metabolism.
Journal of Proteome Research 06/2013; 12(7):3434–3448. · 5.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Abstract In response to infection, neutrophils employ various strategies to defend against the invading microbes. One of such defense mechanism is the formation of neutrophil extracellular traps (NETs). Recent studies suggest that reactive oxygen species is a signal critical to NET formation. This prompts us to examine whether neutrophils from individuals with glucose-6-phosphate dehydrogenase (G6PD) Taiwan-Hakka variant, which are prone to oxidative stress generation, have altered ability to form NET. We adopted an image-based method to study the NET formation potential in neutrophils from G6PD-deficient patients. Neutrophils from either normal or G6PD-deficient individuals underwent NETosis in response to phorbol 12-myristate 13-acetate (PMA). The extent of NETosis in the former did not significantly differ from that of the latter. Diphenyleneiodonium sulfate (DPI) and 3-methyladenine (MA) inhibited PMA-stimulated NET formation in these cells, suggesting the involvement of NADPH oxidase and autophagy in the process. Glucose oxidase (GO) and xanthine oxidase/xanthine (XO/X) could induce a similar extent of NET formation in normal and G6PD-deficient neutrophils. GO- or XO-induced NETosis was not inhibitable by MA, implying that reactive oxygen species (ROS) can act as an independent signal for activation of NETosis. Mechanistically, enhanced superoxide production in neutrophils was associated with increases in levels of NAD(+) and NADP(+), as well as activation of NAD(+) kinase. Taken together, these findings suggest that G6PD-deficient neutrophils are as equally efficient as normal cells in NET formation, and their deficiency in G6PD-associated NADPH regeneration capacity is largely compensated for by nicotinamide nucleotide biosynthesis.
Free Radical Research 06/2013; · 3.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE
Metabolic syndrome is a multiplex disorder and puts patients on the road to type 2 diabetes and atherosclerotic cardiovascular diseases. However, a surrogate biomarker in plasma or urine in fully reflecting features of metabolic syndrome has not been explored.RESEARCH DESIGN AND METHODS
Urine metabolomics has potential utility in metabolic profiling because urine metabolites analysis reflects global outflux of metabolic change. Accordingly, we collected data on subjects (n = 99) with overweight, dyslipidemia, hypertension or impaired glucose tolerance and took a metabolomics approach to analyze the metabolites of urine revealed in metabolic syndrome by high-performance liquid chromatography-time-of-flight mass spectrometry and elicit potential biomarkers to picture metabolic syndrome.RESULTSOur results revealed that the urine nicotinuric acid value of subjects with diabetes (HbA(1c) ≥6.5% or those receiving diabetes medications) (n = 25) was higher than subjects without diabetes (n = 37) (221 ± 31 vs. 152 ± 13 × 10(3) mAU, P = 0.0268). Moreover, urinary nicotinuric acid level was positively correlated with body mass index, blood pressure, total cholesterol, low-density lipoprotein cholesterol, triacylglycerol and high sensitivity C-reactive protein, but negatively correlated with high-density lipoprotein cholesterol.CONCLUSIONS
This is the first study, to our knowledge, to propose that nicotinuric acid represents an important pathogenic mechanism in process from metabolic syndrome to diabetes and atherosclerotic cardiovascular disease.
[Show abstract][Hide abstract] ABSTRACT: G6PD is crucial to NADPH generation and redox homeostasis. We have recently shown that G6PD deficiency predisposes cells to oxidant-induced cell death, and it is associated with the impairment of GSH regeneration. It remains unclear what other metabolic pathways are affected by G6PD deficiency, and whether the altered metabolism disturbs cellular redox homeostasis and underlies the increased susceptibility to oxidants. In the study, we examined the effect of diamide on global metabolite profiles of SK-Hep1-derived SK-i-Gi and SK-i-Sc cells, which could inducibly express shRNAs against G6PD (Gi) and control shRNA (Sc), respectively. There was no significant difference in their metabolite profiles under uninduced condition. Doxycycline (Dox) addition resulted in over 70% decrease in G6PD activity in SK-i-Gi cells. It was accompanied by relatively minor changes in the metabolome of SK-i-Gi cells. Upon further diamide treatment, the metabolite profiles of both SK-i-Gi and SK-i-Sc cells changed in a time-dependent manner. A number of metabolic pathways, including those involved in energy metabolism and metabolism of amino acid and glutathione, were affected. However, the changes in metabolite profile of Dox-treated SK-i-Gi cells were distinct from those of control cells (i.e. Dox-treated SK-i-Sc, SK-i-Gi, and SK-i-Sc cells). Cellular glutathione was depleted, while its disulfide form increased significantly in diamide, Dox-treated SK-i-Gi cells. Metabolites related to energy metabolism, such as AMP, ADP and acetylcarnitine, increased to a greater extent in these cells than in diamide-treated control cells. In contrast, nicotinamide adenine dinucleotide (NAD) and glutathione dropped to a lower level in SK-i-Gi cells than in control cells. The NAD depletion in SK-i-Gi cells was accompanied by a significant increase in NAD kinase activity. Targeted analyses revealed that NADP and NADPH increased significantly in diamide, Dox-treated SK-i-Gi cells as compared with similarly treated control cells. Our results suggest that diamide induces oxidation and depletion of glutathione in SK-i-Gi cells under the condition of G6PD shRNA induction, and subsequently induces conversion of NAD to NADP through enhanced NAD kinase activity. This may represent a compensatory mechanism to restore cellular NADPH reserve in G6PD-deficient cells. It is accompanied by alteration in pathways of cellular energy metabolism, such as glycolysis and β-oxidation.
Free Radical Biology & Medicine 11/2012; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oxidative stress may play a role in spontaneous intracerebral hemorrhage (ICH), but data on oxidative burden in cerebral hemorrhage are limited, and it is not clear whether oxidative markers add predictive power regarding ICH outcome beyond that of traditional factors. The authors therefore examined redox status and traditional factors in ICH patients within 3 days of hemorrhage onset to delineate redox status in ICH and investigate the predictive value with respect to 30-day functional outcome.
Sixty-four patients with ICH and 114 controls were prospectively enrolled in this study. Blood samples were collected within 3 days of ICH onset and processed for isolation of plasma, erythrocytes, and leukocytes. The authors evaluated levels or activities of leukocyte 8-hydroxy-2'-deoxyguanosine (8-OHdG), erythrocyte glucose-6-phosphate dehydrogenase (G6PD), erythrocyte glutathione peroxidase (GPx), plasma malondialdehyde (MDA), vitamin E, and vitamin A, as well as traditional factors including the presence of hypertension or diabetes mellitus, total cholesterol level, and measures of liver function. A general linear model and multivariable logistic regression were used for analyses where appropriate.
After adjustment for age and sex and traditional risk factors, ICH was significantly associated with an increased level of 8-OHdG (p < 0.0001), decreased GPx activity (p = 0.0002), and a decreased level of vitamin E (p = 0.003). There was no association of ICH risk with G6PD activity or MDA or vitamin A level. Considering all the oxidative markers and traditional risk factors together, logistic regression showed an independent association of ICH with 8-OHdG (OR 2.7, 95% CI 1.7-4.2, p < 0.0001). The association between increased 8-OHdG level and lower 30-day Barthel Index was also independent of the effects of age, sex, hemorrhage location and size, and traditional factors (p = 0.026). Unfavorable outcome (modified Rankin Scale score ≥ 3) at 30 days after ICH onset was not significantly associated with any of the examined oxidative markers.
Increased leukocyte 8-OHdG levels, as well as decreased GPx activity and vitamin E levels, were found during acute ICH. Only 8-OHdG was associated with ICH and the 30-day outcome independently from the other oxidative markers and traditional factors. Leukocyte 8-OHdG may add power beyond the traditional factors in predicting ICH outcome and thus may be used as an independent surrogate for clinical ICH study.
Journal of Neurosurgery 08/2011; 115(6):1184-90. · 3.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: DHEA is known to have chemopreventive and antiproliferative activities, and was initially thought to be mediated by inhibition of G6PD. Our previous study has shown that DHEA may act through interference with energy metabolism. To study the effect of pharmacological dose of DHEA on cellular metabolism, and to further delineate the mechanism underlying its antiproliferative effect, we applied a metabolomic approach to globally profile the changes in metabolites in SK-Hep1 cells underexpressing G6PD (Sk-Gi) and control cells (Sk-Sc) after DHEA treatment. RRLC-TOF-MS was used to identify metabolites, and tandem mass spectrometry was used to confirm their identity. DHEA induced changes in glutathione metabolism, lipid metabolism, s-adenosylmethionine (SAM) metabolism, as well as lysine metabolism. Elevation in level of glutathione disulfide, together with a concomitant decrease in level of reduced glutathione, was indicative of increased oxidative stress. Depletion of carnitine and its acyl derivatives reflected decline in fatty acid catabolism. These changes were associated with mitochondrial malfunction and reduction in cellular ATP content. Cardiolipin (CL) and phosphatidylcholine (PC) levels decreased significantly, suggesting that alterations in lipid composition are causally related to decline in mitochondrial function after DHEA treatment. The decline in cellular SAM content was accompanied by decreased expression of methionine adenosyltransferase genes MAT2A and MAT2B. SAM supplementation partially rescued cells from DHEA-induced growth stagnation. Our findings suggest that DHEA causes perturbation of multiple pathways in cellular metabolism. Decreased SAM production, and cardiolipin depletion and the resulting mitochondrial dysfunction underlie the antiproliferative effect of DHEA.
[Show abstract][Hide abstract] ABSTRACT: Diabetes mellitus (DM) is characterized by dysregulated energy metabolism. Resveratrol (RSV) has been shown to ameliorate hyperglycemia and hyperlipidemia in diabetic animals. However, its overall in vivo effects on energy metabolism and the underlying mechanism require further investigation. In the present study, electrospray ionization-tandem mass spectrometry was employed to characterize the urine and plasma metabolomes of control, streptozotocin-induced DM and RSV-treated DM rats. Using principal component analysis (PCA) and heat map analysis, we discovered significant differences among control and experimental groups. RSV treatment significantly reduced the metabolic abnormalities in DM rats. Compared with the age-matched control rats, the level of carnitine was lower, and the levels of acetylcarnitine and butyrylcarnitine were higher in the urine and plasma of DM rats. RSV treatment ameliorated the deranged carnitine metabolism in DM rats. In addition, RSV treatment attenuated the diabetic ketoacidosis and muscle protein degradation, as evidenced from the attenuation of elevated urinary methyl-histidine and plasma branched-chain amino acids levels in DM rats. The beneficial effects of RSV in DM rats were correlated with activation of hepatic AMP-activated protein kinase and SIRT1 expression, increase of hepatic and muscular mitochondrial biogenesis and inhibition of muscle NF-κB activities. We concluded that RSV possesses multiple beneficial metabolic effects in insulin-deficient DM rats, particularly in improving energy metabolism and reducing protein wasting.
AJP Endocrinology and Metabolism 07/2011; 301(5):E853-63. · 4.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Huntington disease (HD) is a degenerative disorder caused by expanded CAG repeats in exon 1 of the huntingtin gene (HTT). Patients with late-stage HD are known to have abnormal auditory processing, but the peripheral auditory functions of HD patients have yet to be thoroughly assessed. In this study, 19 HD patients (aged 40-59 years) were assessed for hearing impairment using pure-tone audiometry and assessment of auditory brainstem responses (ABRs). PTA thresholds were markedly elevated in HD patients. Consistent with this, elevated ABR thresholds were also detected in two mouse models of HD. Hearing loss thus appears to be an authentic symptom of HD. Immunohistochemical analyses demonstrated the presence of mutant huntingtin that formed intranuclear inclusions in the organ of Corti of HD mice, which might interfere with normal auditory function. Quantitative RT-PCR and Western blot analyses further revealed reduced expression of brain creatine kinase (CKB), a major enzyme responsible for ATP regeneration via the phosphocreatine-creatine kinase (PCr-CK) system, in the cochlea of HD mice. Treatment with creatine supplements ameliorated the hearing impairment of HD mice, suggesting that the impaired PCr-CK system in the cochlea of HD mice may contribute to their hearing impairment. These data also suggest that creatine may be useful for treating the hearing abnormalities of patients with HD.
The Journal of clinical investigation 03/2011; 121(4):1519-23. · 15.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Despite their prominent microbicidal roles, NADPH oxidases (NOXs) have been recently found to regulate a wide variety of physiological activities. Through generation of reactive oxygen species (ROS), NOXs actively participate in cellular activities, including NET formation, inflammasome activation and wound sensing. The microbicidal and cytoregulatory roles of NOXs are contrasted in this review.
Microbes and Infection 10/2010; 13(2):109-20. · 2.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glucose-6-phosphate dehydrogenase (G6PD) plays a key role in the regeneration of NADPH and maintenance of cellular redox balance. In the present study, we investigate the effect of G6PD deficiency on H(2)O(2)-elicited signaling in HepG2 cells. H(2)O(2) was found to inhibit cellular protein tyrosine phosphatase (PTP) activity, resulting in activation of MAPKs. MKP-1 expression increased in the late phase of H(2)O(2) signaling. Using RNAi technology, we found that G6PD knockdown enhanced the inhibitory effect of H(2)O(2) on PTPs and led to sustained MAPK activation. This was accompanied by delayed expression and inhibition of MKP-1. Using a pharmacological inhibitor and siRNA, we demonstrate that MKP-1 acts as a regulator of MAPK activation in H(2)O(2) signaling. The prolonged MAPK activation in G6PD-knockdown cells was associated with an increased susceptibility to H(2)O(2)-induced apoptosis and growth retardation. Treatment with p38 and JNK inhibitors or N-acetylcysteine ameliorated such cellular effect, while triptolide and MKP-1-siRNA did the opposite. Glucose oxidase treatment had similar effects as addition of H(2)O(2). Taken together, these findings suggest that G6PD knockdown enhances the magnitude and duration of H(2)O(2)-induced MAPK signaling through inhibition of cellular PTPs, and the resultant anomalous signaling may lead to cell demise.
[Show abstract][Hide abstract] ABSTRACT: Glucose-6-phosphate dehydrogenase (G6PD) has been recently found to play growth-regulatory roles in nucleated cells. To identify any other physiologic roles of G6PD, we generated G6PD-knockdown Hep G2 cells and investigated their susceptibility to oxidants. Hep G2 cells expressing shRNA against G6PD (Gi) were more susceptible to diamide-induced cytotoxicity than control cells expressing scrambled control shRNA (Sc). The level of reactive oxygen species in the Gi cells substantially exceeded that in Sc cells. This was accompanied by increased membrane peroxidation and the appearance of high-molecular-weight aggregates of membrane-associated cytoskeletal proteins in Gi cells. G6PD knockdown was associated with an impaired ability to regenerate glutathione. Diamide caused a considerable decrease in cellular glutathione level and a concomitant increase in glutathione disulfide in Gi cells. Consistent with this finding, N-acetylcysteine mitigated diamide-induced oxidative stress and cell death. Our findings suggest that G6PD confers protection against oxidant-induced cytotoxicity through effective glutathione regeneration.
[Show abstract][Hide abstract] ABSTRACT: Oxidative stress is known to be a determinant of a host's susceptibility to pathogens. Natural compounds with antioxidant activity may provide a preventive measure against infection. Tea polyphenols were evaluated for their ability to inhibit enterovirus 71 (EV71) replication in Vero cell culture. Among the polyphenolic compounds tested, epigallocatechin gallate (EGCG) and gallocatechin gallate (GCG) potently inhibited replication of EV71. EGCG and GCG reduced the titer of infectious progeny virus by 95%. Quantitative RT-PCR analysis also revealed that EGCG suppressed replication of genomic RNA. It was accompanied by an increased cytoprotective effect. EGCG and GCG caused 5-fold increase in the viability of EV71-infected cells. The viral inhibitory effect correlated well with the antioxidant capacity of polyphenol. Mechanistically, EV71 infection led to increased oxidative stress, as shown by increased dichlorofluorescein and MitoSOX Red fluorescence. Upon EGCG treatment, reactive oxygen species (ROS) generation was significantly reduced. Consistent with this, EV71 replication was enhanced in glucose-6-phosphate dehydrogenase deficient cells, and such enhancement was largely reversed by EGCG. These findings suggest that EGCG may suppress viral replication via modulation of cellular redox milieu.
Journal of Agricultural and Food Chemistry 07/2009; 57(14):6140-7. · 2.91 Impact Factor