Markus Bachschmid

Whitaker Wellness Institute, Newport Beach, California, United States

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Publications (75)393.04 Total impact

  • Colin E Murdoch, Markus M Bachschmid, Reiko Matsui
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    ABSTRACT: S-glutathionylation occurs when reactive oxygen or nitrogen species react with protein-cysteine thiols. Glutaredoxin-1 (Glrx) is a cytosolic enzyme which enzymatically catalyses the reduction in S-glutathionylation, conferring reversible signalling function to proteins with redox-sensitive thiols. Glrx can regulate vascular hypertrophy and inflammation by regulating the activity of nuclear factor κB (NF-κB) and actin polymerization. Vascular endothelial growth factor (VEGF)-induced endothelial cell (EC) migration is inhibited by Glrx overexpression. In mice overexpressing Glrx, blood flow recovery, exercise function and capillary density were significantly attenuated after hindlimb ischaemia (HLI). Wnt5a and soluble Fms-like tyrosine kinase-1 (sFlt-1) were enhanced in the ischaemic-limb muscle and plasma respectively from Glrx transgenic (TG) mice. A Wnt5a/sFlt-1 pathway had been described in myeloid cells controlling retinal blood vessel development. Interestingly, a Wnt5a/sFlt-1 pathway was found also to play a role in EC to inhibit network formation. S-glutathionylation of NF-κB components inhibits its activation. Up-regulated Glrx stimulated the Wnt5a/sFlt-1 pathway through enhancing NF-κB signalling. These studies show a novel role for Glrx in post-ischaemic neovascularization, which could define a potential target for therapy of impaired angiogenesis in pathological conditions including diabetes.
    Biochemical Society Transactions 12/2014; 42(6):1665-70. DOI:10.1042/BST20140213 · 3.24 Impact Factor
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    ABSTRACT: Background: Diet-induced obesity leads to metabolic heart disease (MHD) characterized by increased oxidative stress that may cause oxidative post-translational modifications (OPTM) of cardiac mitochondrial proteins. The functional consequences of OPTM of cardiac mitochondrial proteins in MHD are unknown. Our objective was to determine whether cardiac mitochondrial dysfunction in MHD due to diet-induced obesity is associated with cysteine OPTM. Methods and results: Male C57BL/6J mice were fed either a high-fat, high-sucrose (HFHS) or control diet for 8months. Cardiac mitochondria from HFHS-fed mice (vs. control diet) had an increased rate of H2O2 production, a decreased GSH/GSSG ratio, a decreased rate of complex II substrate-driven ATP synthesis and decreased complex II activity. Complex II substrate-driven ATP synthesis and complex II activity were partially restored ex-vivo by reducing conditions. A biotin switch assay showed that HFHS feeding increased cysteine OPTM in complex II subunits A (SDHA) and B (SDHB). Using iodo-TMT multiplex tags we found that HFHS feeding is associated with reversible oxidation of cysteines 89 and 231 in SDHA, and 100, 103 and 115 in SDHB. Conclusions: MHD due to consumption of a HFHS "Western" diet causes increased H2O2 production and oxidative stress in cardiac mitochondria associated with decreased ATP synthesis and decreased complex II activity. Impaired complex II activity and ATP production are associated with reversible cysteine OPTM of complex II. Possible sites of reversible cysteine OPTM in SDHA and SDHB were identified by iodo-TMT tag labeling. Mitochondrial ROS may contribute to the pathophysiology of MHD by impairing the function of complex II. This article is part of a Special Issue entitled 'Mitochondria'.
    Journal of Molecular and Cellular Cardiology 08/2014; 78. DOI:10.1016/j.yjmcc.2014.07.018 · 5.22 Impact Factor
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    ABSTRACT: Glutaredoxin-1 (Glrx) is a cytosolic enzyme that regulates diverse cellular function by removal of GSH adducts from S-glutathionylated proteins including signaling molecules and transcription factors. Glrx is up-regulated during inflammation and diabetes. Glrx overexpression inhibits VEGF-induced endothelial cell (EC) migration. The aim was to investigate the role of up-regulated Glrx in EC angiogenic capacities and in vivo revascularization in the setting of hind limb ischemia. Glrx overexpressing EC from Glrx transgenic mice (TG) showed impaired migration and network formation and secreted higher level of soluble VEGF receptor 1 (sFlt), an antagonizing factor to VEGF. After hind limb ischemia surgery Glrx TG mice demonstrated impaired blood flow recovery, associated with lower capillary density and poorer limb motor function compared to wild type littermates. There were also higher levels of anti-angiogenic sFlt expression in the muscle and plasma of Glrx TG mice after surgery. Non-canonical Wnt5a is known to induce sFlt. Wnt5a was highly expressed in ischemic muscles and EC from Glrx TG mice, and exogenous Wnt5a induced sFlt expression and inhibited network formation in human microvascular EC. Adenoviral Glrx-induced sFlt in EC was inhibited by a competitive Wnt5a inhibitor. Furthermore, Glrx overexpression removed GSH adducts on p65 in ischemic muscle and EC, and enhanced nuclear factor kappa B (NF-kB) activity which was responsible for Wnt5a-sFlt induction. Taken together, up-regulated Glrx induces sFlt in EC via NF-kB -dependent Wnt5a, resulting in attenuated revascularization in hind limb ischemia. The Glrx-induced sFlt may be a part of mechanism of redox regulated VEGF signaling.
    Journal of Biological Chemistry 01/2014; 289(12). DOI:10.1074/jbc.M113.517219 · 4.60 Impact Factor
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    ABSTRACT: Using a novel cysteine thiol labeling strategy coupled with mass spectrometric analysis, we identified and quantified the changes in global reversible cysteine oxidation of proteins in the left ventricle of hearts from mice with metabolic syndrome-associated diastolic dysfunction. This phenotype was induced by feeding a high-fat, high-sucrose, type-2 diabetogenic diet to C57BL/6J mice for 8 mo. The extent of reversible thiol oxidation in relationship to the total available (free and reducible) level of each cysteine could be confidently determined for 173 proteins, of which 98 contained cysteines differentially modified ≥1.5-fold by the diet. Our findings suggest that the metabolic syndrome leads to potentially deleterious changes in the oxidative modification of metabolically active proteins. These alterations may adversely regulate energy substrate flux through glycolysis, β-oxidation, citric acid (TCA) cycle, and oxidative phosphorylation (oxphos), thereby contributing to maladaptive tissue remodeling that is associated with, and possibly contributing to, diastolic left ventricular dysfunction.-Behring, J. B., Kumar, V., Whelan, S. A., Chauhan, P., Siwik, D. A., Costello, C. E., Colucci, W. S., Cohen, R. A., McComb M. E., Bachschmid, M. M. Does reversible cysteine oxidation link the Western diet to cardiac dysfunction?
    The FASEB Journal 01/2014; 28(5). DOI:10.1096/fj.13-233445 · 5.48 Impact Factor
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    ABSTRACT: Sirtuin-1 (SirT1), a member of the NAD+-dependent class III histone deacetylase family, is inactivated in vitro by oxidation of critical cysteine thiols. In a model of metabolic disease, SirT1 activation attenuated apoptosis of hepatocytes and improved liver function including insulin and lipid metabolism. We show in SirT1 overexpressing HepG2 cells that oxidants (nitrosocysteine or hydrogen peroxide) or metabolic stress (high palmitate and high glucose) inactivate SirT1 by reversible oxidative post-translational modifications (OPTM) on cysteines. Mutating these oxidation-sensitive cysteines to serine preserves SirT1 activity and abolishes reversible OPTMs. Overexpressed mutant SirT1 maintains deacetylase activity and attenuates proapoptotic signaling, while overexpressed wild type SirT1 is less protective in metabolically or oxidant stressed cells. To prove that OPTMs of SirT1 are glutathione (GSH) adducts, glutaredoxin-1 (Glrx) was overexpressed to remove this modification. Glrx overexpression maintains endogenous SirT1 activity and prevents proapoptotic signaling in metabolically stressed HepG2 cells. The in vivo significance of oxidative inactivation of SirT1 was investigated in livers of high fat diet-fed C57/B6J mice. SirT1 deacetylase activity was decreased in the absence of changes in SirT1 expression and associated with a marked increase in OPTMs. These results indicate that glutathione adducts on specific SirT1 thiols may be responsible for dysfunctional SirT1 associated with liver disease in metabolic syndrome.
    Journal of Biological Chemistry 01/2014; 289(11). DOI:10.1074/jbc.M113.520403 · 4.60 Impact Factor
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    ABSTRACT: Recently, we demonstrated that gene ablation of mitochondrial manganese superoxide dismutase and aldehyde dehydrogenase-2 markedly contributed to age-related vascular dysfunction and mitochondrial oxidative stress. The present study has sought to investigate the extent of vascular dysfunction and oxidant formation in glutathione peroxidase-1-deficient (GPx-1(-/-)) mice during the aging process with special emphasis on dysregulation (uncoupling) of the endothelial NO synthase. GPx-1(-/-) mice on a C57 black 6 (C57BL/6) background at 2, 6, and 12 months of age were used. Vascular function was significantly impaired in 12-month-old GPx-1(-/-) -mice as compared with age-matched controls. Oxidant formation, detected by 3-nitrotyrosine staining and dihydroethidine-based fluorescence microtopography, was increased in the aged GPx-1(-/-) mice. Aging per se caused a substantial protein kinase C- and protein tyrosine kinase-dependent phosphorylation as well as S-glutathionylation of endothelial NO synthase associated with uncoupling, a phenomenon that was more pronounced in aged GPx-1(-/-) mice. GPx-1 ablation increased adhesion of leukocytes to cultured endothelial cells and CD68 and F4/80 staining in cardiac tissue. Aged GPx-1(-/-) mice displayed increased oxidant formation as compared with their wild-type littermates, triggering redox-signaling pathways associated with endothelial NO synthase dysfunction and uncoupling. Thus, our data demonstrate that aging leads to decreased NO bioavailability because of endothelial NO synthase dysfunction and uncoupling of the enzyme leading to endothelial dysfunction, vascular remodeling, and promotion of adhesion and infiltration of leukocytes into cardiovascular tissue, all of which was more prominent in aged GPx-1(-/-) mice.
    Hypertension 12/2013; 63(2). DOI:10.1161/HYPERTENSIONAHA.113.01602 · 7.63 Impact Factor
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    ABSTRACT: Direct detection and quantification of protein/peptide palmitoylation by mass spectrometry (MS) is a challenging task because of the tendency of palmitoyl loss during sample preparation and tandem MS analysis. In addition, the large difference in hydrophobicity between the palmitoyl peptides and their unmodified counterparts could prevent their simultaneous analysis in a single liquid chromatography-MS experiment. Here, the stability of palmitoylation in several model palmitoyl peptides under different incubation and fragmentation conditions was investigated. It was found that the usual trypsin digestion protocol using dithiothreitol as the reducing agent in ammonium bicarbonate buffer could result in significant palmitoyl losses. Instead, it is recommended that sample preparation be performed in neutral Tris buffer with tris(2-carboxyethyl)phosphine as the reducing agent, conditions under which palmitoylation was largely preserved. For tandem MS analysis, collision-induced dissociation often led to facile palmitoyl loss, and electron capture dissociation frequently produced secondary side-chain losses remote from the backbone cleavage site, thus discouraging their use for accurate palmitoylation site determination. In contrast, the palmitoyl group was mostly preserved during electron transfer dissociation, which produced extensive inter-residue cleavage coverage, making it the ideal fragmentation method for palmitoyl peptide analysis. Finally, derivatization of the unmodified peptides with a perfluoroalkyl tag, N-[(3-perfluorooctyl)propyl] iodoacetamide, significantly increased their hydrophobicity, allowing them to be simultaneously analyzed with palmitoyl peptides for relative quantification of palmitoylation.
    Analytical Chemistry 11/2013; 85(24). DOI:10.1021/ac402850s · 5.83 Impact Factor
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    ABSTRACT: Global changes in reversible cysteine oxidation of cardiac proteins from mice fed a high fat, high sucrose (HFHS) diet for 8 months were quantified using novel 6-plex Iodo-based isobaric labels in a switch assay and high resolution mass spectrometry. 400 reversibly oxidized proteins were identified, 56 with cysteines exhibiting more than 1.5-fold change in oxidation by HFHS diet. As an improvement on "fold-change", our design also defined the percentage of oxidized cysteine at each site, or "site occupancy". Nearly 50% of the proteins localized to mitochondria, consistent with this organelle being the primary target of metabolic stress resulting in mitochondrial oxidant formation. Multiple proteins in each of the critical metabolic pathways, including: the electron transport chain, tricarboxylic acid cycle, acyl-carnitine shuttle, beta-oxidation, and ketolysis, were reversibly oxidized. This suggests that substrate utilization pathways are dysfunctional. The condition further worsens as arterial stiffness increases energy demand of an overworked heart. All of these factors likely contribute to cardiac diastolic dysfunction and hypertrophy that occurs in the mice. Identifying and quantifying reversible oxidative protein changes will elucidate novel signaling mechanisms for switches in metabolism; these modifications could potentially serve as biomarkers for diet-induced metabolic disease. NHLBI HHSN268201000031C
    Experimental Biology FASEB J April 9, 2013 27:558.3, Boston, MA; 04/2013
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    ABSTRACT: Unfavorable metabolic conditions (metabolic disorders) associated with obesity, diabetes, and hyperlipidemia are major causes for cardiovascular disease (CVD). One major environmental cause of this may be attributed to poor diet, aka the American diet. The early detection and monitoring of the adverse effects of metabolic disease on the heart and vasculature, although well studied, remain elusive. Our hypothesis is that poor diet causes unfavorable metabolic conditions in heart tissue resulting in inflammation and oxidative stress reflected in protein changes. Here we apply label-free proteomics to elucidate potential biomarkers of CVD including both protein changes and changes in post-translational modifications (PTMs). Heart tissue was from control mice and mice fed a high fat high sucrose diet (HFHS). MS/MS data was analyzed with Proteome Discoverer and Mascot software, using both variable-modification and error-tolerant search modes. Label-free quantification was conducted using Scaffold and Progenesis; typically yielding >1,000 features. Using IPA software revealed a number of cardiovascular disease related proteins were observed. This is the first step in biomarker panel development for disease diagnosis and progression. This project was funded by NIH-NCRR grants P41 RR010888/GM104603, S10 RR015942, S10 RR020946, S10 RR025082 and NIH-NHLBI contract N01 HV00239.
    Experimental Biology FASEB J April 9, 2013 27:794.17, Boston, MA; 04/2013
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    ABSTRACT: Unfavorable metabolic conditions (metabolic disorders) associated with obesity and diabetes are major causes for cardiovascular disease. Early detection of the adverse effects of metabolic disease remain elusive. We believe that nonspecific changes which occur in plasma proteins, indicators of inflammation and oxidants, may act as evidence of systemic metabolic disease. Here we elucidate potential biomarkers of CVD including both protein changes and changes in post-translational modifications (PTMs). Label-free mass spectrometry-based proteomics was used to interrogate changes in differential protein and PTM expression in plasma samples from mouse and human models. Label-free LCMS/MS typically yielded >1,000 features ( p<0.05, >2fold). A number of cardiovascular disease related proteins were observed. Up regulated proteins were: haptoglobin, a known biomarker related to inflammation, low mannose binding protein associated with inflammation and CVD in type 2 diabetes, superoxide dismutase and extracellular matrix protein both implicated in type 2 diabetes. We also observed different PTMs associated with oxidative stress including lipid peroxidation products such as hydroxynonenal and multiple forms of oxidation such as cysteine sulfonic acid. Development of a metabolic disorder/CVD-specific protein panel will afford the first step in biomarker panel development such that disease diagnosis and progression may be performed directly at the molecular level. This project was funded by NIH-NCRR grants P41 RR010888/ GM104603, S10 RR015942, S10 RR020946, S10 RR025082 and NIH-NHLBI contract N01 HV00239.
    Experimental Biology FASEB J April 9, 2013 27:663.10, Boston, MA; 04/2013
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    ABSTRACT: We demonstrate for the first time that endomembrane-delimited HRas mediates VEGF-induced activation of eNOS and migratory response of human endothelial cells. Using thiol labeling strategies and immunofluorescent cell staining, we found that only 31% of total HRas is S-palmitoylated, tethering the small GTPase to the plasma membrane, but leaving the function of the large majority of endomembrane-localized HRas unexplained. Knock-down of HRas blocked VEGF-induced PI3K-dependent Akt (S473) and eNOS (S1177) phosphorylation and nitric oxide-dependent cell migration, demonstrating the essential role of HRas. Activation of endogenous HRas led to recruitment and phosphorylation of eNOS at endomembranes. The loss of migratory response in cells lacking endogenous HRas was fully restored by modest over-expression of an endomembrane-delimited HRas palmitoylation mutant. These studies define a newly recognized role for endomembrane-localized HRas in mediating nitric oxide-dependent pro-angiogenic signaling.
    Journal of Biological Chemistry 04/2013; DOI:10.1074/jbc.M112.427765 · 4.60 Impact Factor
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    ABSTRACT: The reaction product of nitric oxide and superoxide, peroxynitrite, is a potent biological oxidant. The most important oxidative protein modifications described for peroxynitrite are cysteine-thiol oxidation and tyrosine nitration. We have previously demonstrated that intrinsic heme-thiolate (P450)-dependent enzymatic catalysis increases the nitration of tyrosine 430 in prostacyclin synthase and results in loss of activity which contributes to endothelial dysfunction. We here report the sensitive peroxynitrite-dependent nitration of an over-expressed and partially purified human prostacyclin synthase (3.3 μM) with an EC50 value of 5 μM. Microsomal thiols in these preparations effectively compete for peroxynitrite and block the nitration of other proteins up to 50 μM peroxynitrite. Purified, recombinant PGIS showed a half-maximal nitration by 10 μM 3-morpholino sydnonimine (Sin-1) which increased in the presence of bicarbonate, and was only marginally induced by freely diffusing NO2-radicals generated by a peroxidase/nitrite/hydrogen peroxide system. Based on these observations, we would like to emphasize that prostacyclin synthase is among the most efficiently and sensitively nitrated proteins investigated by us so far. In the second part of the study, we identified two classes of peroxynitrite scavengers, blocking either peroxynitrite anion-mediated thiol oxidations or phenol/tyrosine nitrations by free radical mechanisms. Dithiopurines and dithiopyrimidines were highly effective in inhibiting both reaction types which could make this class of compounds interesting therapeutic tools. In the present work, we highlighted the impact of experimental conditions on the outcome of peroxynitrite-mediated nitrations. The limitations identified in this work need to be considered in the assessment of experimental data involving peroxynitrite.
    International Journal of Molecular Sciences 04/2013; 14(4):7542-70. DOI:10.3390/ijms14047542 · 2.34 Impact Factor
  • Free Radical Biology and Medicine 11/2012; 53:S138. DOI:10.1016/j.freeradbiomed.2012.10.377 · 5.71 Impact Factor
  • Free Radical Biology and Medicine 11/2012; 53:S127-S128. DOI:10.1016/j.freeradbiomed.2012.10.317 · 5.71 Impact Factor
  • Free Radical Biology and Medicine 11/2012; 53:S31. DOI:10.1016/j.freeradbiomed.2012.10.077 · 5.71 Impact Factor
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    ABSTRACT: Abstract Significance: Oxidative post-translational modifications (OPTMs) have been demonstrated as contributing to cardiovascular physiology and pathophysiology. These modifications have been identified using antibodies as well as advanced proteomic methods, and the functional importance of each is beginning to be understood using transgenic and gene deletion animal models. Given that OPTMs are involved in cardiovascular pathology, the use of these modifications as biomarkers and predictors of disease has significant therapeutic potential. Adequate understanding of the chemistry of the OPTMs is necessary to determine what may occur in vivo and which modifications would best serve as biomarkers. Recent Advances: By using mass spectrometry, advanced labeling techniques, and antibody identification, OPTMs have become accessible to a larger proportion of the scientific community. Advancements in instrumentation, database search algorithms, and processing speed have allowed MS to fully expand on the proteome of OPTMs. In addition, the role of enzymatically reversible OPTMs has been further clarified in preclinical models. Critical Issues: The identification of OPTMs suffers from limitations in analytic detection based on the methodology, instrumentation, sample complexity, and bioinformatics. Currently, each type of OPTM requires a specific strategy for identification, and generalized approaches result in an incomplete assessment. Future Directions: Novel types of highly sensitive MS instrumentation that allow for improved separation and detection of modified proteins and peptides have been crucial in the discovery of OPTMs and biomarkers. To further advance the identification of relevant OPTMs in advanced search algorithms, standardized methods for sample processing and depository of MS data will be required. Antioxid. Redox Signal. 17, 1528-1559.
    Antioxidants & Redox Signaling 05/2012; 17(11):1528-59. DOI:10.1089/ars.2012.4706 · 8.20 Impact Factor
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    ABSTRACT: Abstract Aims: Prostaglandin endoperoxide H(2) synthase (PGHS) is a well-known target for peroxynitrite-mediated nitration. In several experimental macrophage models, however, the relatively late onset of nitration failed to coincide with the early peak of endogenous peroxynitrite formation. In the present work, we aimed to identify an alternative, peroxynitrite-independent mechanism, responsible for the observed nitration and inactivation of PGHS-2 in an inflammatory cell model. Results: In primary rat alveolar macrophages stimulated with lipopolysaccharide (LPS), PGHS-2 activity was suppressed after 12 h, although the prostaglandin endoperoxide H(2) synthase (PGHS-2) protein was still present. This coincided with a nitration of the enzyme. Coincubation with a nitric oxide synthase-2 (NOS-2) inhibitor preserved PGHS-2 nitration and at the same time restored thromboxane A(2) (TxA(2)) synthesis in the cells. Formation of reactive oxygen species (ROS) was maximal at 4 h and then returned to baseline levels. Nitrite (NO(2)(-)) production occurred later than ROS generation. This rendered generation of peroxynitrite and the nitration of PGHS-2 unlikely. We found that the nitrating agent was formed from NO(2)(-), independent from superoxide ((•)O(2)(-)). Purified PGHS-2 treated with NO(2)(-) was selectively nitrated on the active site Tyr(371), as identified by mass spectrometry (MS). Exposure to peroxynitrite resulted in the nitration not only of Tyr(371), but also of other tyrosines (Tyr). Innovation and Conclusion: The data presented here point to an autocatalytic nitration of PGHS-2 by NO(2)(-), catalyzed by the enzyme's endogenous peroxidase activity and indicate a potential involvement of this mechanism in the termination of prostanoid formation under inflammatory conditions. Antioxid. Redox Signal. 17, 1393-1406.
    Antioxidants & Redox Signaling 05/2012; 17(10):1393-406. DOI:10.1089/ars.2011.4485 · 8.20 Impact Factor
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    ABSTRACT: Reactive oxygen and nitrogen species contributing to homeostatic regulation and the pathogenesis of various cardiovascular diseases, including atherosclerosis, hypertension, endothelial dysfunction, and cardiac hypertrophy, is well established. The ability of oxidant species to mediate such effects is in part dependent on their ability to induce specific modifications on particular amino acids, which alter protein function leading to changes in cell signaling and function. The thiol containing amino acids, methionine and cysteine, are the only oxidized amino acids that undergo reduction by cellular enzymes and are, therefore, prime candidates in regulating physiological signaling. Various reports illustrate the significance of reversible oxidative modifications on cysteine thiols and their importance in modulating cardiovascular function and physiology. RECENT ADVANCES: The use of mass spectrometry, novel labeling techniques, and live cell imaging illustrate the emerging importance of reversible thiol modifications in cellular redox signaling and have advanced our analytical abilities. Distinguishing redox signaling from oxidative stress remains unclear. S-nitrosylation as a precursor of S-glutathionylation is controversial and needs further clarification. Subcellular distribution of glutathione (GSH) may play an important role in local regulation, and targeted tools need to be developed. Furthermore, cellular redundancies of thiol metabolism complicate analysis and interpretation. The development of novel pharmacological analogs that specifically target subcellular compartments of GSH to promote or prevent local protein S-glutathionylation as well as the establishment of conditional gene ablation and transgenic animal models are needed.
    Antioxidants & Redox Signaling 03/2012; 16(6):524-42. DOI:10.1089/ars.2011.4336 · 8.20 Impact Factor
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    ABSTRACT: Characteristic morphological and molecular alterations such as vessel wall thickening and reduction of nitric oxide occur in the aging vasculature leading to the gradual loss of vascular homeostasis. Consequently, the risk of developing acute and chronic cardiovascular diseases increases with age. Current research of the underlying molecular mechanisms of endothelial function demonstrates a duality of reactive oxygen and nitrogen species in contributing to vascular homeostasis or leading to detrimental effects when formed in excess. Furthermore, changes in function and redox status of vascular smooth muscle cells contribute to age-related vascular remodeling. The age-dependent increase in free radical formation causes deterioration of the nitric oxide signaling cascade, alters and activates prostaglandin metabolism, and promotes novel oxidative posttranslational protein modifications that interfere with vascular and cell signaling pathways. As a result, vascular dysfunction manifests. Compensatory mechanisms are initially activated to cope with age-induced oxidative stress, but become futile, which results in irreversible oxidative modifications of biological macromolecules. These findings support the 'free radical theory of aging' but also show that reactive oxygen and nitrogen species are essential signaling molecules, regulating vascular homeostasis.
    Annals of Medicine 03/2012; 45(1). DOI:10.3109/07853890.2011.645498 · 4.73 Impact Factor
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    ABSTRACT: Here we demonstrate a new paradigm in redox signaling, whereby oxidants resulting from metabolic stress directly alter protein palmitoylation by oxidizing reactive cysteine thiolates. In mice fed a high-fat, high-sucrose diet and in cultured endothelial cells (ECs) treated with high palmitate and high glucose (HPHG), there was decreased HRas palmitoylation on Cys181/184 (61±24% decrease for cardiac tissue and 38±7.0% in ECs). This was due to oxidation of Cys181/184, detected using matrix-assisted laser desorption/ionization time of flight (MALDI TOF)-TOF. Decrease in HRas palmitoylation affected its compartmentalization and Ras binding domain binding activity, with a shift from plasma membrane tethering to Golgi localization. Loss of plasma membrane-bound HRas decreased growth factor-stimulated ERK phosphorylation (84±8.6% decrease) and increased apoptotic signaling (24±6.5-fold increase) after HPHG treatment that was prevented by overexpressing wild-type but not C181/184S HRas. The essential role of HRas in metabolic stress was made evident by the similar effects of expressing an inactive dominant negative N17-HRas or a MEK inhibitor. Furthermore, the relevance of thiol oxidation was demonstrated by overexpressing manganese superoxide dismutase, which improved HRas palmitoylation and ERK phosphorylation, while lessening apoptosis in HPHG treated ECs.
    The FASEB Journal 11/2011; 26(2):832-41. DOI:10.1096/fj.11-189415 · 5.48 Impact Factor

Publication Stats

3k Citations
393.04 Total Impact Points


  • 2008–2013
    • Whitaker Wellness Institute
      Newport Beach, California, United States
    • Beverly Hospital, Boston MA
      Beverly, Massachusetts, United States
  • 2009–2012
    • Boston University
      • Department of Medicine
      Boston, Massachusetts, United States
  • 1999–2012
    • Universität Konstanz
      • • Department of Biology
      • • Faculty of Sciences
      Konstanz, Baden-Wuerttemberg, Germany
  • 2007–2011
    • Boston Medical Center
      Boston, Massachusetts, United States
    • Johannes Gutenberg-Universität Mainz
      • III. Department of Medicine
      Mainz, Rhineland-Palatinate, Germany
  • 2002
    • University of Zurich
      • Institute of Physiology
      Zürich, Zurich, Switzerland