Aldehyde dehydrogenase 2 inhibited oxidized LDL-induced NLRP3 inflammasome priming and activation via attenuating oxidative stress.

Oxidized low-density lipoprotein (ox-LDL)-mediated NLRP3 inflammasome activation is crucial in atherosclerosis (AS) initiation and progression. Aldehyde dehydrogenase 2 (ALDH2) has been reported to display protective effects during AS development; however, the underlying mechanisms are largely unknown. Here we investigate the role of ALDH2 in ox-LDL-induced NLRP3 inflammasome priming and activation. We treated RAW264.7 murine macrophages with ox-LDL with or without ALDH2 activator Alda-1 and measured NLRP3 inflammasome priming and activation, ALDH2 protein expression and enzyme activity, IL-1ß release, and DNA damage. It was found that ox-LDL impaired ALDH2 activity and induced NLRP3 inflammasome priming and activation. Alda-1 inhibited both of the priming and activation steps of NLRP3 inflammasome as well as subsequent cell pyroptosis and attenuated ROS and 4-HNE levels in ox-LDL-treated macrophages. Taken together, ALDH2 activation inhibits priming and activation of NLRP3 inflammasome via reducing oxidative stress, which suggests that ALDH2 may be a potential target for anti-inflammatory therapies in AS treatment.

New insights into two yeast BDHs from the PDH subfamily as aldehyde reductases in context of detoxification of lignocellulosic aldehyde inhibitors.

At least 24 aldehyde reductases from Saccharomyces cerevisiae have been characterized and most function in in situ detoxification of lignocellulosic aldehyde inhibitors, but none is classified into the polyol dehydrogenase (PDH) subfamily of the medium-chain dehydrogenase/reductase (MDR) superfamily. This study confirmed that two (2R,3R)-2,3-butanediol dehydrogenases (BDHs) from industrial (denoted Y)/laboratory (denoted B) strains of S. cerevisiae, Bdh1p(Y)/Bdh1p(B) and Bdh2p(Y)/Bdh2p(B), were members of the PDH subfamily with an NAD(P)H binding domain and a catalytic zinc binding domain, and exhibited reductive activities towards lignocellulosic aldehyde inhibitors, such as acetaldehyde, glycolaldehyde, and furfural. Especially, the highest enzyme activity towards acetaldehyde by Bdh2p(Y) was 117.95 U/mg with cofactor nicotinamide adenine dinucleotide reduced (NADH). Based on the comparative kinetic property analysis, Bdh2p(Y)/Bdh2p(B) possessed higher specific activity, substrate affinity, and catalytic efficiency towards glycolaldehyde than Bdh1p(Y)/Bdh1p(B). This was speculated to be related to their 49% sequence differences and five nonsynonymous substitutions (Ser41Thr, Glu173Gln, Ile270Leu, Ile316Met, and Gly317Cys) occurred in their conserved NAD(P)H binding domains. Compared with BDHs from a laboratory strain, Bdh1p(Y) and Bdh2p(Y) from an industrial strain displayed five nonsynonymous mutations (Thr12, Asn61, Glu168, Val222, and Ala235) and three nonsynonymous mutations (Ala34, Ile96, and Ala369), respectively. From a first analysis with selected aldehydes, their reductase activities were different from BDHs of laboratory strain, and their catalytic efficiency was higher towards glycolaldehyde and lower towards acetaldehyde. Comparative investigation of kinetic properties of BDHs from S. cerevisiae as aldehyde reductases provides a guideline for their practical applications in in situ detoxification of aldehyde inhibitors during lignocellulose bioconversion.Key Points• Two yeast BDHs have enzyme activities for reduction of aldehydes.• Overexpression of BDHs slightly improves yeast tolerance to acetaldehyde and glycolaldehyde.• Bdh1p and Bdh2p differ in enzyme kinetic properties.• BDHs from strains with different genetic backgrounds differ in enzyme kinetic properties.

Differential cell death decisions in the testis: evidence for an exclusive window of ferroptosis in round spermatids.

Oxidative stress is a major aetiology in many pathologies, including that of male infertility. Recent evidence in somatic cells has linked oxidative stress to the induction of a novel cell death modality termed ferroptosis. However, the induction of this iron-regulated, caspase-independent cell death pathway has never been explored outside of the soma. Ferroptosis is initiated through the inactivation of the lipid repair enzyme glutathione peroxidase 4 (GPX4) and is exacerbated by the activity of arachidonate 15-lipoxygenase (ALOX15), a lipoxygenase enzyme that facilitates lipid degradation. Here, we demonstrate that male germ cells of the mouse exhibit hallmarks of ferroptosis including; a caspase-independent decline in viability following exposure to oxidative stress conditions induced by the electrophile 4-hydroxynonenal or the ferroptosis activators (erastin and RSL3), as well as a reciprocal upregulation of ALOX15 and down regulation of GPX4 protein expression. Moreover, the round spermatid developmental stage may be sensitized to ferroptosis via the action of acyl-CoA synthetase long-chain family member 4 (ACSL4), which modifies membrane lipid composition in a manner favourable to lipid peroxidation. This work provides a clear impetus to explore the contribution of ferroptosis to the demise of germline cells during periods of acute stress in in vivo models.

4-HNE induces proinflammatory cytokines of human retinal pigment epithelial cells by promoting extracellular efflux of HSP70.

Oxidative stress and subsequent chronic inflammation result in dysfunction of the retinal pigment epithelium (RPE) and represent therapeutic targets in the context of age-related macular degeneration (AMD). However, molecular mechanisms that linked oxidative stress and inflammation still unclear. As an important byproduct of oxidative stress, 4-hydroxynonenal (4-HNE) induces apoptosis and lysosome dysregulation of RPE cells. In the present study, we evaluated cytokines production of RPE cells induced by 4-HNE by using cytokine array and confirmed that 4-HNE induced IL-6, IL-1ß and TNF-α production in a concentration dependent manner. Specifically, 4-HNE also induced IL-10 and TGF-ß production in low concentration. Molecular analysis revealed that intracellular HSP70 inhibited 4-HNE-induced production of pro-inflammatory cytokines, and 4-HNE exerted proinflammatory effects in RPE cells by enhancing extracellular release of HSP70, as efflux inhibitor Methyl-ß-cyclodextrin (MBC) treatment significantly blocked the release of HSP70 and decreased IL-6 production of RPE cells induced by 4-HNE. Meanwhile, HSP70 inducer arimoclomol increased intracellular HSP70 production, but showed no influence on its extracellular level, also performed anti-inflammatory effects in 4-HNE-stimulated RPE cells. Whereas the anti-inflammatory effects of paeoniflorin, an HSP70 inducer simultaneously promoted its extracellular efflux, was lower than arimoclomol. In addition, we further confirmed that MBC exhibited synergetic effect with both paeoniflorin and arimoclomol to inhibit the production of proinflammatory cytokines induced by 4-HNE. Taken together, these results indicate that HSP70 plays a vital role in regulating inflammation of RPE cells induced by oxidative stress and might be a potential novel target for clinical treatment of AMD.

Integration of Epigallocatechin Gallate in Gelatin Sponges Attenuates Matrix Metalloproteinase-Dependent Degradation and Increases Bone Formation.

Matrix metalloproteinase (MMP)-2 and MMP-9 are well-known gelatinases that disrupt the extracellular matrix, including gelatin. However, the advantages of modulating MMP expression in gelatin-based materials for applications in bone regenerative medicine have not been fully clarified. In this study, we examined the effects of epigallocatechin gallate (EGCG), a major polyphenol catechin isolated from green tea, on MMP expression in gelatin sponges and its association with bone formation. Four gelatin sponges with or without EGCG were prepared and implanted into bone defects for up to 4 weeks. Histological and immunohistological staining were performed. Micro-computed tomography was used to estimate the bone-forming capacity of each sponge. Our results showed that EGCG integration attenuated MMP-2 (70.6%) and -9 expression (69.1%) in the 1 week group, increased residual gelatin (118.7%), and augmented bone formation (101.8%) in the 4 weeks group in critical-sized bone defects of rat calvaria compared with vacuum-heated gelatin sponges without EGCG. Moreover, vacuum-heated gelatin sponges with EGCG showed superior bone formation compared with other sponges. The results indicated that integration of EGCG in gelatin-based materials modulated the production and activity of MMP-2 and -9 in vivo, thereby enhancing bone-forming capacity.

Protective effects of 6-ureido/thioureido-2,4,5-trimethylpyridin-3-ols against 4-hydroxynonenal-induced cell death in adult retinal pigment epithelial-19 cells.

Dysfunction or progressive degeneration of retinal pigment epithelium (RPE) contributes in the initial pathogenesis of age-related macular degeneration (AMD) causing irreversible vision loss, which makes RPE the prime target of the disease. The present study aimed to identify compounds to protect 4-hydroxynonenal (4-HNE)-induced RPE cell death by inhibiting NADPH oxidase 4 (NOX4) activity, not just as free radical scavengers, using ARPE-19, a human adult retinal pigment epithelial cell line, as a RPE representative. Novel thirty-two 6-ureido/thioureido-2,4,5-trimethylpyridin-3-ol derivatives 17 were synthesized and tested. We found that there was a strong correlation between level of protective effect of compounds 17 against 4-HNE-induced APRE-19 cell death and that of inhibitory activity against 4-HNE-induced superoxide production, and that most of the compounds 17 showed minimal DPPH radical scavenging activity. Compound 17-28 showed the best protective activity against 4-HNE-induced superoxide production (79.5% inhibition) and cell death (85.1% recovery) at 10 µM concentration, which was better than that of VAS2870, a NOX2/4 inhibitor. In addition, compound 17-28 blocked 4-HNE-induced apoptosis of ARPE-19 cells in a concentration-dependent manner. The results indicate that compound 17-28 may be a lead compound to develop AMD therapeutics.

Functions of aldehyde reductases from Saccharomyces cerevisiae in detoxification of aldehyde inhibitors and their biotechnological applications.

Bioconversion of lignocellulosic biomass to high-value bioproducts by fermentative microorganisms has drawn extensive attentions worldwide. Lignocellulosic biomass cannot be efficiently utilized by microorganisms, such as Saccharomyces cerevisiae, but has to be pretreated prior to fermentation. Aldehyde compounds, as the by-products generated in the pretreatment process of lignocellulosic biomass, are considered as the most important toxic inhibitors to S. cerevisiae cells for their growth and fermentation. Aldehyde group in the aldehyde inhibitors, including furan aldehydes, aliphatic aldehydes, and phenolic aldehydes, is identified as the toxic factor. It has been demonstrated that S. cerevisiae has the ability to in situ detoxify aldehydes to their corresponding less or non-toxic alcohols. This reductive reaction is catalyzed by the NAD(P)H-dependent aldehyde reductases. In recent years, detoxification of aldehyde inhibitors by S. cerevisiae has been extensively studied and a huge progress has been made. This mini-review summarizes the classifications and structural features of the characterized aldehyde reductases from S. cerevisiae, their catalytic abilities to exogenous and endogenous aldehydes and effects of metal ions, chemical protective additives, and salts on enzyme activities, subcellular localization of the aldehyde reductases and their possible roles in protection of the subcellular organelles, and transcriptional regulation of the aldehyde reductase genes by the key stress-response transcription factors. Cofactor preference of the aldehyde reductases and their molecular mechanisms and efficient supply pathways of cofactors, as well as biotechnological applications of the aldehyde reductases in the detoxification of aldehyde inhibitors derived from pretreatment of lignocellulosic biomass, are also included or supplemented in this mini-review.

Fancd2 counteracts the toxic effects of naturally produced aldehydes in mice.

Reactive aldehydes are common carcinogens. They are also by-products of several metabolic pathways and, without enzymatic catabolism, may accumulate and cause DNA damage. Ethanol, which is metabolised to acetaldehyde, is both carcinogenic and teratogenic in humans. Here we find that the Fanconi anaemia DNA repair pathway counteracts acetaldehyde-induced genotoxicity in mice. Our results show that the acetaldehyde-catabolising enzyme Aldh2 is essential for the development of Fancd2(-/-) embryos. Nevertheless, acetaldehyde-catabolism-competent mothers (Aldh2(+/-)) can support the development of double-mutant (Aldh2(-/-)Fancd2(-/-)) mice. However, these embryos are unusually sensitive to ethanol exposure in utero, and ethanol consumption by postnatal double-deficient mice rapidly precipitates bone marrow failure. Lastly, Aldh2(-/-)Fancd2(-/-) mice spontaneously develop acute leukaemia. Acetaldehyde-mediated DNA damage may critically contribute to the genesis of fetal alcohol syndrome in fetuses, as well as to abnormal development, haematopoietic failure and cancer predisposition in Fanconi anaemia patients.

Quercetin alleviates 4-hydroxynonenal-induced cytotoxicity and inflammation in ARPE-19 cells.

Retinal pigment epithelium (RPE) plays the principal role in age-related macular degeneration (AMD), a progressive eye disease with no cure and limited therapeutical options. In the pathogenesis of AMD, degeneration of RPE cells by multiple factors including increased oxidative stress and chronic inflammation precedes the irreversible loss of photoreceptors and central vision. Here, we report that the plant-derived polyphenol, quercetin, increases viability and decreases inflammation in stressed human ARPE-19 cells after exposure to the lipid peroxidation end product 4-hydroxynonenal (HNE). Several previous studies have been conducted using the direct oxidant H2O2 but we preferred HNE since natural characteristics predispose RPE cells to the type of oxidative damage evoked by lipid peroxidation. Quercetin improved cell membrane integrity and mitochondrial function as assessed in LDH and MTT tests. Decreased production of proinflammatory mediators IL-6, IL-8, and MCP-1 were indicated at the RNA level by qPCR and at the protein level by the ELISA technique. In addition, we probed the signaling behind the effects and observed that p38 and ERK MAPK pathways, and CREB signaling are regulated by quercetin in ARPE-19 cells. In conclusion, our present data suggests that HNE is highly toxic to serum-starved ARPE-19 cells but quercetin is able to reverse these adverse effects even when administered after an oxidative insult.

Differential metabolism of 4-hydroxynonenal in liver, lung and brain of mice and rats.

The lipid peroxidation end-product 4-hydroxynonenal (4-HNE) is generated in tissues during oxidative stress. As a reactive aldehyde, it forms Michael adducts with nucleophiles, a process that disrupts cellular functioning. Liver, lung and brain are highly sensitive to xenobiotic-induced oxidative stress and readily generate 4-HNE. In the present studies, we compared 4-HNE metabolism in these tissues, a process that protects against tissue injury. 4-HNE was degraded slowly in total homogenates and S9 fractions of mouse liver, lung and brain. In liver, but not lung or brain, NAD(P)+ and NAD(P)H markedly stimulated 4-HNE metabolism. Similar results were observed in rat S9 fractions from these tissues. In liver, lung and brain S9 fractions, 4-HNE formed protein adducts. When NADH was used to stimulate 4-HNE metabolism, the formation of protein adducts was suppressed in liver, but not lung or brain. In both mouse and rat tissues, 4-HNE was also metabolized by glutathione S-transferases. The greatest activity was noted in livers of mice and in lungs of rats; relatively low glutathione S-transferase activity was detected in brain. In mouse hepatocytes, 4-HNE was rapidly taken up and metabolized. Simultaneously, 4-HNE-protein adducts were formed, suggesting that 4-HNE metabolism in intact cells does not prevent protein modifications. These data demonstrate that, in contrast to liver, lung and brain have a limited capacity to metabolize 4-HNE. The persistence of 4-HNE in these tissues may increase the likelihood of tissue injury during oxidative stress.

4-Hydroxy-2-nonenal modified histone-H2A: a possible antigenic stimulus for systemic lupus erythematosus autoantibodies.

Protein modifications by 4-hydroxy-2-nonenals (HNE) are involved in various diseases. Histones are DNA protective nucleoprotein, which adopt different structures under oxidative stress. This study was undertaken to test the role of HNE-modified-histone-H2A (HNE-H2A) in systemic lupus erythematosus (SLE). Our data revealed that HNE-mediated-lipid peroxidation in histone-H2A caused alteration in histidine, lysine and cystein residues. In addition, protein carbonyl contents were also high in HNE-H2A. HNE-specific quencher, L-carnosine further reiterates HNE-modifications. Specificity of autoantibodies from SLE patients (n=48) were analyzed towards HNE-H2A and their results were compared with sex- and age-matched controls (n=36). SLE autoantibodies show preferential binding to HNE-H2A in comparison with histone-H2A (p<0.0001). Furthermore, HNE-H2A was also detected in SLE peripheral blood mononuclear cells. In conclusion, this is the first study to demonstrate the role of HNE-modified-histone in SLE. Preferential binding of HNE-H2A by affinity purified SLE-IgG pointed out the likely role of HNE-H2A in the initiation/progression of SLE.

Citronellal, a monoterpene present in Java citronella oil, attenuates mechanical nociception response in mice.

CONTEXT: Citronellal is a monoterpene present in the oil of many species, including Cymbopogon winterianus Jowitt (Poaceae). OBJECTIVE: The present study investigated the effect of citronellal on inflammatory nociception induced by different stimuli and examined the involvement of the NO-cGMP-ATP-sensitive K⁺ channel pathway. MATERIALS AND METHODS: We used male Swiss mice (n = 6 per group) that were treated intraperitoneally with citronellal (25, 50 or 100 mg/kg) 0.5 h after the subplantar injection of 20 µl of carrageenan (CG; 300 µg/paw), tumor necrosis factor-α (TNF-α; 100 pg/paw), prostaglandin E2 (PGE2; 100 ng/paw) or dopamine (DA; 30 µg/paw). The mechanical nociception was evaluated at 0.5, 1, 2 and 3 h after the injection of the agents, using a digital analgesimeter (von Frey). The effects of citronellal were also evaluated in the presence of L-NAME (30 mg/kg) or glibenclamide (5 mg/kg). RESULTS: At all times, citronellal in all doses inhibited the development of mechanical nociception induced by CG (p < 0.001 and p < 0.01) and TNF-α (p < 0.001, p < 0.01, and p < 0.05). The citronellal was able to increase the pain threshold in the DA test (p < 0.001, p < 0.01, and p < 0.05) and in the PGE2 test at all times (p < 0.001 and p < 0.05). L-NAME and glibenclamide reversed the antinociceptive effects of the citronellal at higher doses in the PGE2 test. DISCUSSION AND CONCLUSION: These data suggest that citronellal attenuated mechanical nociception, mediated in part by the NO-cGMP-ATP-sensitive K⁺ channel pathway.

Genipin inhibits UCP2-mediated proton leak and acutely reverses obesity- and high glucose-induced beta cell dysfunction in isolated pancreatic islets.

Uncoupling protein 2 (UCP2) negatively regulates insulin secretion. UCP2 deficiency (by means of gene knockout) improves obesity- and high glucose-induced beta cell dysfunction and consequently improves type 2 diabetes in mice. In the present study, we have discovered that the small molecule, genipin, rapidly inhibits UCP2-mediated proton leak. In isolated mitochondria, genipin inhibits UCP2-mediated proton leak. In pancreatic islet cells, genipin increases mitochondrial membrane potential, increases ATP levels, closes K(ATP) channels, and stimulates insulin secretion. These actions of genipin occur in a UCP2-dependent manner. Importantly, acute addition of genipin to isolated islets reverses high glucose- and obesity-induced beta cell dysfunction. Thus, genipin and/or chemically modified variants of genipin are useful research tools for studying biological processes thought to be controlled by UCP2. In addition, these agents represent lead compounds that comprise a starting point for the development of therapies aimed at treating beta cell dysfunction.

Hydralazine modifies Aß fibril formation and prevents modification by lipids in vitro.

Lipid oxidative damage and amyloid ß (Aß) misfolding contribute to Alzheimer's disease (AD) pathology. Thus, the prevention of oxidative damage and Aß misfolding are attractive targets for drug discovery. At present, no AD drugs approved by the Food and Drug Administration (FDA) prevent or halt disease progression. Hydralazine, a smooth muscle relaxant, is a potential drug candidate for AD drug therapy as it reduces Aß production and prevents oxidative damage via its antioxidant hydrazide group. We evaluated the efficacy of hydralazine, and related hydrazides, in reducing (1) Aß misfolding and (2) Aß protein modification by the reactive lipid 4-hydroxy-2-nonenal (HNE) using transmission electron microscopy and Western blotting. While hydralazine did not prevent Aß aggregation as measured using the protease protection assay, there were more oligomeric species observed by electron microscopy. Hydralazine prevented lipid modification of Aß, and Aß was used as a proxy for classes of proteins which either misfold or are modified by HNE. All of the other hydrazides prevented lipid modification of Aß and also did not prevent Aß aggregation. Surprisingly, a few of the compounds, carbazochrome and niclosamide, appeared to augment Aß formation. Thus, hydrazides reduced lipid oxidative damage, and hydralazine additionally reduced Aß misfolding. While hydralazine would require specific chemical modifications for use as an AD therapeutic itself (to improve blood brain barrier permeability, reduce vasoactive side effects, and optimization for amyloid inhibition), this study suggests its potential merit for further AD drug development.

Can Medicinal Plants and Bioactive Compounds Combat Lipid Peroxidation Product 4-HNE-Induced Deleterious Effects?

The toxic reactive aldehyde 4-hydroxynonenal (4-HNE) belongs to the advanced lipid peroxidation end products. Accumulation of 4-HNE and formation of 4-HNE adducts induced by redox imbalance participate in several cytotoxic processes, which contribute to the pathogenesis and progression of oxidative stress-related human disorders. Medicinal plants and bioactive natural compounds are suggested to be attractive sources of potential agents to mitigate oxidative stress, but little is known about the therapeutic potentials especially on combating 4-HNE-induced deleterious effects. Of note, some investigations clarify the attenuation of medicinal plants and bioactive compounds on 4-HNE-induced disturbances, but strong evidence is needed that these plants and compounds serve as potent agents in the prevention and treatment of disorders driven by 4-HNE. Therefore, this review highlights the pharmacological basis of these medicinal plants and bioactive compounds to combat 4-HNE-induced deleterious effects in oxidative stress-related disorders, such as neurotoxicity and neurological disorder, eye damage, cardiovascular injury, liver injury, and energy metabolism disorder. In addition, this review briefly discusses with special attention to the strategies for developing potential therapies by future applications of these medicinal plants and bioactive compounds, which will help biological and pharmacological scientists to explore the new vistas of medicinal plants in combating 4-HNE-induced deleterious effects.

Oleacein inhibits STAT3, activates the apoptotic machinery, and exerts anti-metastatic effects in the SH-SY5Y human neuroblastoma cells.

Several studies published in the last decade suggest that the beneficial role of extra-virgin olive oil (EVOO) in human health is mostly attributable to the main secoiridoid derivatives (oleuropein, oleocanthal, and oleacein). Anti-cancer properties have also been demonstrated for certain compounds present in small quantities in EVOO, including oleuropein and hydroxytyrosol, which have been extensively studied, while minor attention has been given to the most abundant secoiridoid oleacein. The aim of our research was to study the molecular mechanisms underlying the anti-proliferative and anti-metastatic capacity of oleacein in the SH-SY5Y human neuroblastoma cell line. Our results demonstrate that oleacein is able to reduce the proliferation of the SH-SY5Y cells by blocking the cell cycle in the S phase and inducing apoptotic cell death through the increase in both Bax and p53 as well as a reduction in the Bcl-2 expression and STAT3 phosphorylation. Moreover, oleacein caused reduction in the SH-SY5Y cell adhesion and migration. Overall, these findings indicate that oleacein exerts anti-cancer effects against neuroblastoma cells, suggesting a promising role as a candidate against this type of cancer.

Prostate-specific antigen is a "chymotrypsin-like" serine protease with unique P1 substrate specificity.

Prostate-specific antigen (PSA), a serine protease belonging to the human kallikrein family, is best known as a prostate cancer biomarker. Emerging evidence suggests that PSA may also play a salient role in prostate cancer development and progression. With large amounts of enzymatically active PSA continuously and selectively produced by all stages of prostate cancer, PSA is an attractive target. PSA inhibitors, therefore, may represent a promising class of therapeutics and/or imaging agents. PSA displays chymotrypsin-like specificity, cleaving after hydrophobic residues, in addition to possessing a unique ability to cleave after glutamine in the P1 position. In this study, we investigated the structural motifs of the PSA S1 pocket that give it a distinct architecture and specificity when compared to the S1 pocket of chymotrypsin. Using the previously described PSA substrate Ser-Ser-Lys-Leu-Gln (SSKLQ) as a template, peptide aldehyde based inhibitors containing novel P1 aldehydes were made and tested against both proteases. Glutamine derivative aldehydes were highly specific for PSA while inhibitors with hydrophobic P1 aldehydes were potent inhibitors of both proteases with K(i) values <500 nM. The crystal structure of PSA was used to generate a model that allowed GOLD docking studies to be performed to further understand the critical interactions required for inhibitor binding to the S1 pockets of PSA and chymotrypsin. In conclusion, these results provide experimental and structural evidence that the S1 specificity pocket of PSA is distinctly different from that of chymotrypsin and that the development of highly specific PSA inhibitors is feasible.

Odor interaction between Bourgeonal and its antagonist undecanal.

The perceived quality of a binary mixture will, as a rule of thumb, be dominated by the quality of the stronger unmixed component. On the other hand, there are mechanisms that, in theory, suggest that this will not always be true; one example being receptor antagonism. Undecanal has been indicated as an antagonist for bourgeonal-sensitive receptors in the human olfactory epithelium. Therefore, we investigated mixtures of isointense concentrations of bourgeonal and undecanal and, as a control, mixtures of isointense concentrations of bourgeonal and n-butanol. Both mixture types were investigated at 2 levels of concentration. The particular aim was to see if the bourgeonal-undecanal mixtures would exhibit asymmetric odor quality favoring the perception of the antagonist and the control mixture would not. For the control mixture, indeed odor quality tended to be dominated by the strongest component before mixing as would be suggested from previous studies. In line with the hypothesis, the bourgeonal-undecanal mixture was dominated by the antagonist's quality, but only when mixed at higher concentrations, altogether suggesting the effects of a low-affinity receptor antagonism. This is, to our knowledge, the first demonstration of how antagonistic interaction at the level of the receptor can affect the perception of odor mixtures in humans.

Modification of endothelial nitric oxide synthase by 4-oxo-2(E)-nonenal(ONE) in preeclamptic placentas.

Preeclampsia (PE) is a leading cause of pregnancy complications, affecting 3-7% of pregnant women worldwide. The pathophysiology of preeclampsia involves a redox imbalance, oxidative stress and a reduced nitric oxide (NO) bioavailability. The molecular and cellular mechanisms leading to the dysfunction of the placental endothelial NO synthase (eNOS) are not clarified. This study was designed to investigate whether aldehydes generated by lipid peroxidation products (LPP), may contribute to placental eNOS dysfunction in PE. The analysis of placentas from PE-affected patients and normal pregnancies, showed a significant increase in protein carbonyl content, indicative of oxidative stress-induced protein modification, as shown by the accumulation of acrolein, 4-hydroxynonenal (HNE), and 4-oxo-2(E)-nonenal (ONE) adducts in PE placentas. In contrast, the levels of these LPP-adducts were low in placentas from normal pregnancies. Immunofluorescence and confocal experiments pointed out a colocalization of eNOS with ONE-Lys adducts, whereas eNOS was not modified in normal placentas. LC-MS/MS analysis of recombinant eNOS preincubated with ONE, allowed to identify several ONE-modified Lys-containing peptides, confirming that eNOS may undergo post-translational modification by LPP. The preincubation of HTR-8/SVneo human trophoblasts (HTR8) with ONE, resulted in ONE-Lys modification of eNOS and a reduced generation of NO. ONE inhibited the migration of HTR8 trophoblasts in the wound closure model, and this was partly restored by the NO donor, NOC-18, which confirmed the important role of NO in the invasive potential of trophoblasts. In conclusion, placental eNOS is modified by ONE in PE placentas, which emphasizes the sensitivity of this protein to oxidative stress in the disturbed redox environment of preeclamptic pregnancies.

Reactive carbonyls and oxidative stress: potential for therapeutic intervention.

Reactive aldehydes and ketones are produced as a result of oxidative stress in several disease processes. Considerable evidence is now accumulating that these reactive carbonyl products are also involved in the progression of diseases, including neurodegenerative disorders, diabetes, atherosclerosis, diabetic complications, reperfusion after ischemic injury, hypertension, and inflammation. To counter carbonyl stress, cells possess enzymes that can decrease aldehyde load. These enzymes include aldehyde dehydrogenases (ALDH), aldo-keto reductases (AKR), carbonyl reductase (CBR), and glutathione S-transferases (GST). Some of these enzymes are inducible by chemoprotective compounds via Nrf2/ARE- or AhR/XRE-dependent mechanisms. This review describes the metabolism of reactive carbonyls and discusses the potential for manipulating levels of carbonyl-metabolizing enzymes through chemical intervention.