Biomedical subjects

Matthew J Picklo

Publications and source records attributed to Matthew J Picklo.

16 recordsLinked to original sources

PubMed2007

Carbonylation of adipose proteins in obesity and insulin resistance: identification of adipocyte fatty acid-binding protein as a cellular target of 4-hydroxynonenal.

Obesity is a state of mild inflammation correlated with increased oxidative stress. In general, pro-oxidative conditions lead to production of reactive aldehydes such as trans-4-hydroxy-2-nonenal (4-HNE) and trans-4-oxo-2-nonenal implicated in the development of a variety of metabolic diseases. To investigate protein modification by 4-HNE as a consequence of obesity and its potential relationship to the development of insulin resistance, proteomics technologies were utilized to identify aldehyde-modified proteins in adipose tissue. Adipose proteins from lean insulin-sensitive and obese insulin-resistant C57Bl/6J mice were incubated with biotin hydrazide and detected using horseradish peroxidase-conjugated streptavidin. High carbohydrate, high fat feeding of mice resulted in a approximately 2-3-fold increase in total adipose protein carbonylation. Consistent with an increase in oxidative stress in obesity, the abundance of glutathione S-transferase A4 (GSTA4), a key enzyme responsible for metabolizing 4-HNE, was decreased approximately 3-4-fold in adipose tissue of obese mice. To identify specific carbonylated proteins, biotin hydrazide-modified adipose proteins from obese mice were captured using avidin-Sepharose affinity chromatography, proteolytically digested, and subjected to LC-ESI MS/MS. Interestingly enzymes involved in cellular stress response, lipotoxicity, and insulin signaling such as glutathione S-transferase M1, peroxiredoxin 1, glutathione peroxidase 1, eukaryotic elongation factor 1alpha-1 (eEF1alpha1), and filamin A were identified. The adipocyte fatty acid-binding protein, a protein implicated in the regulation of insulin resistance, was found to be carbonylated in vivo with 4-HNE. In vitro modification of adipocyte fatty acid-binding protein with 4-HNE was mapped to Cys-117, occurred equivalently using either the R or S enantiomer of 4-HNE, and reduced the affinity of the protein for fatty acids approximately 10-fold. These results indicate that obesity is accompanied by an increase in the carbonylation of a number of adipose-regulatory proteins that may serve as a mechanistic link between increased oxidative stress and the development of insulin resistance.

Adipose Tissue↗

PubMed2006

Inhibition of aldehyde detoxification in CNS mitochondria by fungicides.

Among the several converging factors leading to Parkinson's disease, epidemiological studies indicate a correlation between Parkinson's disease (PD) with living in a rural area and/or exposure to agricultural pesticides. In this present study, we examined the potential of multiple agricultural pesticides for their ability to inhibit the function of whole, respiring rat brain mitochondria using the oxidation of the neurotoxic lipid-aldehyde trans-4-hydroxy-2-nonenal (HNE) as a biomarker for mitochondrial aldehyde dehydrogenase (ALDH) activity in situ. We chose an arbitrary cutoff concentration of 10 microM of each pesticide. Our data demonstrate that only four of the eighteen compounds tested inhibited oxidation of HNE to trans-4-hydroxy-2-nonenoic acid (HNEAcid). These compounds included rotenone, maneb, mancozeb, and benomyl. Surprisingly, maneb, mancozeb, and benomyl did not inhibit mitochondrial respiration but inhibited the activity of purified rat ALDH2 and rat ALDH5A, enzymes found in brain mitochondria that oxidize HNE and aldehydes derived from neurotransmitters. Our data demonstrate that mitochondrial ALDHs are sensitive targets of pesticide inactivation and that pesticides such as maneb and benomyl can decrease the detoxification of lipid peroxidation derived aldehydes such as HNE and, likely, aldehydes derived from neurotransmitters.

Aldehyde Dehydrogenase↗

PubMed2006

Elevated oxidation of docosahexaenoic acid, 22:6 (n-3), in brain regions of rats undergoing ethanol withdrawal.

Ethanol withdrawal is a serious clinical problem owing in part to over stimulation of ionotropic glutamate receptors in the brain and is linked to elevated oxidative damage. In this study, we tested the hypothesis that lipid peroxidation is elevated in the brain tissue of rats fed an ethanol-containing diet for 6 weeks followed by 24h of withdrawal. We measured F(2)-isoprostanes (IsoPs), as products of arachidonic acid (20:4, n-6) oxidation and F(4)-neuroprostanes (NeuroPs), as products of docosahexaenoic acid (22:6, n-3; DHA) oxidation. Levels of NeuroPs were significantly elevated in the cerebral cortex (97%) and brainstem (68%) of animals undergoing ethanol withdraw versus control. In contrast, elevations in IsoP content (39%) occurred only in the cerebellum of animals in withdrawal versus control animals. These data demonstrate that DHA, versus arachidonic acid, is particularly vulnerable to oxidative damage in ethanol withdrawal.

Animals↗

PubMed2006

Midpolarity and nonpolar wood smoke particulate matter fractions deplete glutathione in RAW 264.7 macrophages.

Wood smoke particulate matter (PM) is a complex mixture of components falling in a spectrum of highly polar to nonpolar species. Wood smoke PM is a likely factor in pulmonary disease and induces oxidative damage. Most toxicity studies focus upon nonpolar species such as polycyclic aromatic hydrocarbons (PAHs). However, the role of more polar PM constituents as toxicants is not clear. In this work, we evaluated the ability of multiple fractions of varying polarity to deplete glutathione (GSH) in RAW 264.7 macrophages and BEAS-2B bronchial epithelial cells. We utilized hot pressurized (subcritical) water to fractionate wood smoke PM into seven fractions of decreasing polarity. In contrast to polar fractions, midpolarity and nonpolar fractions exhibited greater GSH depletion (ED50 at PM concentrations of approximately 50 microg/mL). GSH depletion caused by nonpolar fractions (extracted at 250-300 degrees C) was associated with the presence of PAHs. In midpolarity fractions (extracted at 100-150 degrees C), oxy-PAHs, syringylguaiacyls, disyringyls, and lower molecular weight PAHs were found. Direct comparison of GSH depletion by individual oxy-PAHs and PAHs suggests that oxy-PAHs are contributors to oxidative stress caused by wood smoke PM. However, other unidentified PM constituents contribute to GSH depletion as well. The results indicate the toxicological importance of oxygenated organics found in midpolarity PM fractions.

Animals↗

PubMed2006

Astrocytic biotransformation of trans-4-hydroxy-2-nonenal is dose-dependent.

Elevated levels of trans-4-hydroxy-2-nonenal (HNE) are observed in brain tissues in patients with neurodegenerative diseases. Although astrocytes are known to play a crucial role in regulating and supporting neuronal processes, their capacity to detoxify HNE is unknown. In this work, we examined the extent to which HNE undergoes phase I and phase II metabolism in astrocytes. Murine astrocytes were exposed to three different concentrations of HNE. The loss of HNE was approximately 90%, 80%, and 70% for 1, 5, and 15 microM HNE, respectively, following a 10 min incubation. The expected metabolites trans-4-hydroxy-2-nonenoic acid (HNEAcid), (4-hydroxynonanal-3-yl)glutathione (GSHNE), and (1,4-dihydroxynonane-3-yl)glutathione (GSDHN) accounted for 90% of HNE lost at 1 microM HNE. However, when astrocytes were exposed to 5 and 15 microM HNE, those metabolites accounted only for 50% and 17%, respectively. Binding to macromolecules accounted for only 5-10% of HNE loss. Furthermore, depletion of GSH content had only a small effect on HNE metabolism without elevating HNE oxidation and suggests that other unidentified metabolic pathways are functioning. We identified two novel metabolites of HNE, gamma-nonalactone and the potent pyrrole forming compound, 4-oxo-nonanal (ONA). Occurrence of 1,4-dihydroxynonene was observed as well. These data suggest that the biotransformation of HNE yields products with differing or enhanced toxicity, as well as nontoxic products.

Aldehydes↗

PubMed2005

Enantioselective metabolism of trans-4-hydroxy-2-nonenal by brain mitochondria.

Trans-4-hydroxy-2-nonenal (HNE) is a product of lipid peroxidation with many cellular effects. HNE possesses a stereogenic center at the C4 carbon that influences the metabolism and alkylation targets of HNE. We tested the hypothesis that rat brain mitochondria metabolize HNE in an enantioselective manner after exposure to racemic HNE. The study of HNE chirality, however, is hindered by the lack of facile methods to chromatographically resolve (R)-HNE and (S)-HNE. We used a chiral hydrazine, (S)-carbidopa, as a derivatization reagent to form diastereomers with (R)-HNE and (S)-HNE that were separated by reverse-phase HPLC. After exposure to racemic HNE, rat brain mitochondria metabolized HNE enantioselectively with a higher rate of (R)-HNE metabolism. By using the purified enantiomers of HNE, we found that this enantioselective metabolism of HNE was the result of higher rates of enzymatic oxidation of (R)-HNE by aldehyde dehydrogenases compared to (S)-HNE. Conjugation of HNE to glutathione was a minor metabolic pathway and was not enantioselective. These studies demonstrate that the chirality of HNE affects its mitochondrial metabolism and potentially other processes in the central nervous system.

Aldehydes↗

PubMed2005

Nitrate-based vasodilators inhibit multiple vascular aldehyde dehydrogenases.

Nitrate-based vasodilators (NBVs) are commonly used to treat multiple sequelae of atherosclerosis. A commonly used NBV, glyceryl trinitrate (GTN) is bioactivated by mitochondrial, class 2 aldehyde dehydrogenase (ALDH2). ALDH2 and other ALDHs are NAD(P)+-dependent enzymes critical to the detoxification of cytotoxic lipid-aldehydes elevated in atherosclerotic lesions, such as trans-4-hydroxy-2-nonenal (HNE). The GTN bioactivation step, however, inac-tivates ALDH2 and may alter the metabolism of these aldehydes. In this study, we tested the hypothesis that multiple ALDH enzymes are inhibited by different NBVs. ALDH2, ALDH3A, and ALDH5A were present in aorta with ALDH2 and ALDH3A localized to the smooth muscle layers. GTN (1 microM) inhibited ALDH2 activity (55 +/- 6% of control) and ablated ALDH3 activity. In contrast, isosorbide-2,5-dinitrate (ISDN, 1 microM) inhibited ALDH3 activity (1.1 +/- 0.4% of control) but did not inhibit ALDH2 activity even up to 50 microM ISDN. In homogenates of rat aorta, GTN (1 microM) inhibited the NAD+-dependent (41 +/- 5% of control) and NADP+-dependent (25 +/- 6% of control) detoxification of HNE. The inhibition of ALDH3A, but not ALDH2, could be prevented by the addition of dithiothreitol. These studies demonstrate that GTN and ISDN possess selectivity for ALDH inactivation with different mechanisms of inactivation.

Aldehyde Dehydrogenase↗

PubMed2004

Metabolism of 4-hydroxy-trans-2-nonenal by central nervous system mitochondria is dependent on age and NAD+ availability.

Lipid peroxidation and mitochondrial dysfunction are associated with multiple neurodegenerative disorders including Alzheimer's disease and Parkinson's disease. 4-Hydroxy-trans-2-nonenal (HNE) is a major, neurotoxic product of lipid peroxidation whose levels are elevated in these diseases. Previous data from this laboratory demonstrate that mitochondria play an important role in the detoxification of HNE particularly through the oxidation of HNE to 4-hydroxy-trans-2-nonenoate (HNEAcid). In this work, we examined the disposition of HNE when incubated with intact, well-coupled, rat brain mitochondria. Our results demonstrated that HNE loss occurred in a time- and concentration-dependent, saturable manner with a K(M) of 28.0 +/- 11.8 microM HNE and a V(Max) of 10.0 +/- 1.7 nmol/min/mg. HNEAcid formation occurred in a saturable manner with a K(M) of 25.3 +/- 6.3 microM HNE and a V(Max) of 4.4 +/- 0.43 nmol/min/mg. The formation of HNE-glutathione adducts and HNE-protein adducts comprised only a small percentage of HNE consumption. HNE metabolism was significantly diminished in rat brain mitochondria isolated from older animals. We then tested the hypothesis that the mitochondrial NADH/NAD(+) ratio regulated matrix aldehyde dehydrogenase activity. Our results demonstrate that HNE oxidation was significantly inhibited to a greater extent with pyruvate and malate as substrates vs succinate. Complex I inhibition with respiratory substrates further blocked HNE detoxification. Rotenone (100 nM) inhibited respiration by 15% whereas HNEAcid formation was decreased to 72% of control levels. These results demonstrate that in situ mitochondrial aldehyde detoxification is affected by decrements in NAD(+) availability and complex I activity.

Acetaldehyde↗

PubMed2004

4-Hydroxy-trans-2-nonenoic acid is a gamma-hydroxybutyrate receptor ligand in the cerebral cortex and hippocampus.

Elevated production of 4-hydroxy-trans-2-nonenal (HNE) occurs in numerous neurological disorders involving oxidative damage. HNE is metabolized to the non-toxic 4-hydroxy-trans-2-nonenoic acid (HNEAcid) by aldehyde dehydrogenases in the rat cerebral cortex. Based upon the structural similarity of HNEAcid to ligands of the gamma-hydroxybutyrate (GHB) receptor, we hypothesized that HNEAcid is an endogenous ligand for the GHB receptor. HNEAcid displaced the specific binding of the GHB receptor ligand (3)H-NCS382 (30 nm) in membrane preparations of human frontal cerebral cortex and whole rat cerebral cortex with IC(50s) of 3.9 +/- 1.1 and 5.6 +/- 1.2 micro m, respectively. Inhibition was attenuated when the carboxyl group of HNEAcid was replaced with an aldehyde or an alcohol. HNEAcid (300 micro m) did not displace the binding of beta-adrenergic receptor and GABA(B) receptor antagonists, demonstrating the selectivity of HNEAcid for the GHB receptor. HNEAcid is formed in homogenates of human frontal cortical gray matter in an NAD(+)-dependent (V(Max), 0.71 nmol/min/mg) and NADP(+)-dependent (V(Max), 0.12 nmol/min/mg) manner. Lastly, (3)H-NCS382 binding is elevated 2.7-fold with age in the cerebral cortex of rats. Our data demonstrate that an HNE metabolite, formed in rat and human brain, is a signaling molecule analogous to other bioactive lipid peroxidation products.

Adrenergic beta-Antagonists↗

PubMed2004

Inhibition of cardiac myocyte contraction by 4-hydroxy-trans-2-nonenal.

Lipid peroxidation, initiated by hydroxyl radicals, results in production of 4-Hydroxy-trans-2-nonenal (HNE) and leads to cardiac injury. However, impact of HNE on ventricular function has not been clearly defined. This study was to examine the direct effect of HNE on cardiac contractile function at cardiac myocyte level. Adult male rat ventricular myocytes were isolated and electrically stimulated to contract at 0.5 Hz. Mechanical and intracellular Ca2+ properties were evaluated using an Ionoptix Myocam system. Contractile properties analyzed included peak shortening (PS), time-PS, time-to-90% relengthening, maximal velocities of shortening and relengthening (+/-dL/dt), change of electrically stimulated intracellular Ca2+ fura-2 fluorescent intensity, and intracellular Ca2+ decay. Our results indicated that short-term incubation of HNE (10(-6) to 10(-4) M) with myocytes depressed PS, +/-dL/dt, and fura-2 fluorescent intensity; shortened time-PS; and elevated resting intracellular Ca2+ levels without affecting time-to-90% relengthening and intracellular Ca2+ decay. Interestingly, the HNE-induced cardiac mechanical effects (with the exception of shortened time-PS) were abolished by either the aldehyde dehydrogenase inhibitor cyanamide or the p38 mitogen-activated protein kinase inhibitor SB203580. These findings reveal a possible role of HNE in the lipid peroxidation-associated cardiac contractile dysfunction that is likely mediated through HNE metabolism and mitogen-activated protein kinase activation.

Aldehyde Dehydrogenase↗

PubMed2004

Toxicity of wide-range polarity fractions from wood smoke and diesel exhaust particulate obtained using hot pressurized water.

Epidemiological and toxicological studies correlate the adverse health effects of particulate matter (PM) with the available information regarding their chemical characterization, which has focused on nonpolar organics (e.g., polycyclic aromatic hydrocarbons [PAHs]), which are limited to 15 to 50% of total organic carbon. To study both polar and nonpolar species, we have employed hot pressurized (liquid) water for the fractionation of diesel exhaust and wood smoke PM. In agreement with the results of previous studies, nonpolar fractions from both PM samples showed strong cytotoxicity (cell viability decreased to 50-60%) corresponding to the presence of PAHs. Surprisingly, similar decreases in cell viability were also found in polar fractions (50 degrees C) from both diesel exhaust and wood smoke PM. The midpolarity fractions (100-150 degrees C) from wood smoke PM also displayed high cytotoxicity corresponding to methoxyphenols and oxy-PAHs. Although the midpolarity fractions from diesel exhaust PM showed no cytotoxicity, genotoxicity was found in the 150 degrees C fraction, possibly corresponding to nitroaromatics. The present study demonstrates the suitability of hot pressurized water (as a single, nontoxic solvent) for the fractionation and toxicological characterization of wide-range polarity constituents of PM and, possibly, other environmental matrices.

Air Pollutants↗

PubMed2003

Oxidation of 4-hydroxynonenal in rat brain slices.

4-Hydroxy-2-nonenal (HNE) is implicated as a neurotoxic 'second messenger' of oxidative damage in Alzheimer's disease (AD). The mechanism of HNE toxicity is due to alkylation of cellular nucleophilic groups. The C1 aldehyde is key to the alkylation ability of HNE. Oxidation of the C1 aldehyde to 4-hydroxy-2-nonenoic acid is catalyzed by aldehyde dehydrogenases. In this work, we tested the hypothesis that HNE oxidation to HNEAcid occurs in rat cerebral cortex utilizing rat cerebral cortical slices exposed extracellularly to HNE. HNEAcid formation occurs in a dose dependent manner with approximately 18-25% of the HNE consumed accounted for by HNEAcid formation. HNEAcid was found exclusively in the incubation media, suggesting that HNEAcid is exported from the cells of the slice. These data demonstrate that HNE detoxification through the oxidation pathway occur in the cerebral cortex.

Aldehydes↗

PubMed2003

Inhibition of succinic semialdehyde dehydrogenase activity by alkenal products of lipid peroxidation.

Lipid peroxidation causes the generation of the neurotoxic aldehydes acrolein and 4-hydroxy-trans-2-nonenal (HNE). These products are elevated in neurodegenerative diseases and acute CNS trauma. Previous studies demonstrate that mitochondrial class 2 aldehyde dehydrogenase (ALDH2) is susceptible to inactivation by these alkenals. In the liver and brain another mitochondrial aldehyde dehydrogenase, succinic semialdehyde dehydrogenase (SSADH/ALDH5A1), is present. In this study, we tested the hypothesis that aldehyde products of lipid peroxidation inhibit SSADH activity using the endogenous substrate, succinic semialdehyde (SSA, 50 microM). Acrolein potently inhibited SSADH activity (IC(50)=15 microM) in rat brain mitochondrial preparations. This inhibition was of an irreversible and noncompetitive nature. HNE inhibited activity with an IC(50) of 110 microM. Trans-2-hexenal (HEX) and crotonaldehyde (100 microM each) did not inhibit activity. These data suggest that acrolein and HNE disrupt SSA metabolism and may have subsequent effects on CNS neurochemistry.

Acrolein↗

PubMed2003

Mitochondrial oxidation of 4-hydroxy-2-nonenal in rat cerebral cortex.

4-hydroxy-trans-2-nonenal (HNE) is a neurotoxic product of lipid peroxidation whose levels are elevated in multiple neurodegenerative diseases and CNS trauma. The detoxification of HNE may take the route of glutathione conjugation to the C3 carbon and the oxidation or reduction of the C1 aldehyde. In this work, we examined whether the oxidation of HNE to its corresponding carboxylic acid, 4-hydroxy-trans-2-nonenoate (HNEAcid) was detoxifying event, if it occurred in rat cerebral cortex, and in which subcellular compartments. Our results show that HNEAcid did not form protein adducts and was non-toxic to Neuro 2A cells. HNEAcid formation occurred in rat cerebral cortex slices following exposure to HNE in a time-dependent and dose-dependent fashion. Homogenate studies indicated that HNEAcid formation was NAD+ dependent. Subcellular fractionation demonstrated that mitochondria had the highest specific activity for HNEAcid formation with a KM of 21 micro m HNE. These data indicate that oxidation of HNE to its corresponding acid is a major detoxification pathway of HNE in the CNS and that mitochondria play a role in this process.

Aldehydes↗

PubMed2003

Oxidation of 4-hydroxy-2-nonenal by succinic semialdehyde dehydrogenase (ALDH5A).

Elevated levels of 4-hydroxy-trans-2-nonenal (HNE) are implicated in the pathogenesis of numerous neurodegenerative disorders. Although well-characterized in the periphery, the mechanisms of detoxification of HNE in the CNS are unclear. HNE is oxidized to a non-toxic metabolite in the rat cerebral cortex by mitochondrial aldehyde dehydrogenases (ALDHs). Two possible ALDH enzymes which might oxidize HNE in CNS mitochondria are ALDH2 and succinic semialdehyde dehydrogenase (SSADH/ALDH5A). It was previously established that hepatic ALDH2 can oxidize HNE. In this work, we tested the hypothesis that SSADH oxidizes HNE. SSADH is critical in the detoxification of the GABA metabolite, succinic semialdehyde (SSA). Recombinant rat SSADH oxidized HNE and other alpha,beta-unsaturated aldehydes. Inhibition and competition studies in rat brain mitochondria showed that SSADH was the predominant oxidizing enzyme for HNE but only contributed a portion of the total oxidizing activity in liver mitochondria. In vivo administration of diethyldithiocarbamate (DEDC) effectively inhibited (86%) ALDH2 activity but not HNE oxidation in liver mitochondria. The data suggest that a relationship between the detoxification of SSA and the neurotoxic aldehyde HNE exists in the CNS. Furthermore, these studies show that multiple hepatic aldehyde dehydrogenases are able to oxidize HNE.

Aldehyde Dehydrogenase↗

PubMed2002

Carbonyl toxicology and Alzheimer's disease.

A large amount of data has implicated reactive carbonyls as neurotoxic mediators of oxidative damage in the progression of Alzheimer's disease (AD) and other neurodegenerative diseases. The oxidation of polyunsaturated fatty acids, reducing sugars, and amino acids leads to the formation of carbonyls and carbonyl adduction products such as 4-hydroxy-2-nonenal (HNE), advanced glycation end products (AGEs), and protein-bound carbonyls. Levels of these products are elevated in AD. In this review, we examine the role that carbonyls may play in the development of this disease. We focus upon the chemistry of these molecules and the evidence for their involvement in AD. The biological effects of these carbonyl species in model systems and their relationship to AD are discussed. Lastly, we examine the potential mechanisms that the brain utilizes to detoxify carbonyl species and possible therapeutic interventions based on carbonyl detoxification.

Aldehydes↗