Polyubiquitinylation Profile in Down Syndrome Brain Before and After the Development of Alzheimer Neuropathology.

AIMS: Among the putative mechanisms proposed to be common factors in Down syndrome (DS) and Alzheimer's disease (AD) neuropathology, deficits in protein quality control (PQC) have emerged as a unifying mechanism of neurodegeneration. Considering that disturbance of protein degradation systems is present in DS and that oxidized/misfolded proteins require polyubiquitinylation for degradation via the ubiquitin proteasome system, this study investigated if dysregulation of protein polyubiquitinylation is associated with AD neurodegeneration in DS. RESULTS: Postmortem brains from DS cases before and after development of AD neuropathology and age-matched controls were analyzed. By selectively isolating polyubiquitinated proteins, we were able to identify specific proteins with an altered pattern of polyubiquitinylation as a function of age. Interestingly, we found that oxidation is coupled with polyubiquitinylation for most proteins mainly involved in PQC and energy metabolism. INNOVATION: This is the first study showing alteration of the polyubiquitinylation profile as a function of aging in DS brain compared with healthy controls. Understanding the onset of the altered ubiquitome profile in DS brain may contribute to identification of key molecular regulators of age-associated cognitive decline. CONCLUSIONS: Disturbance of the polyubiquitinylation machinery may be a key feature of aging and neurodegeneration. In DS, age-associated deficits of the proteolytic system may further exacerbate the accumulation of oxidized/misfolded/polyubiquitinated proteins, which is not efficiently degraded and may become harmful to neurons and contribute to AD neuropathology. Antioxid. Redox Signal. 26, 280-298.

Redox proteomic analysis of serum from aortic anerurysm patients: insights on oxidation of specific protein target.

OBJECTIVES: oxidative stress is undoubtedly one of the main players in abdominal aortic aneurysm (AAA) pathophysiology. Recent studies in AAA patients reported an increase in the indices of oxidative damage at the tissue level and in biological fluids coupled with the loss of counter-regulatory mechanisms of protection from oxidative stress. We recently reported, in a proteomic analysis of AAA patient sera, changes in the expression of several proteins exerting important modulatory activities on cellular proliferation, differentiation and response to damage. This study aimed to explore the involvement of protein oxidation, at peripheral levels, in AAA. METHODS: a redox proteomic approach was used to investigate total and specific protein carbonylation and protein-bound 4-hydroxy-2-nonenal (HNE) in the serum of AAA patients compared with age-matched controls. RESULTS: our results show increased oxidative damage to protein as indexed by the total carbonyl levels and total protein-bound HNE. By redox proteomics we identified specific carbonylation of three serum proteins: serum retinol-binding protein, vitamin D-binding protein and fibrinogen α-chain HNE. We also identified increased protein-bound HNE levels for hemopexin, IgK chain C region and IgK chain V-III region SIE. In addition we found a high correlation between specific protein carbonylation and protein-bound HNE and the aortic diameter. Moreover the analysis of serum proteins with antioxidant activity demonstrates the oxidation of albumin together with the overexpression of transferrin, haptoglobin and HSPs 90, 70, 60 and 32. CONCLUSIONS: this study support the involvement of oxidative stress in the pathogenesis of AAA and might provide a further degree of knowledge in the cause-effect role of oxidative stress shedding new light on the molecular candidates involved in the disease.

Impairment of biliverdin reductase-A promotes brain insulin resistance in Alzheimer disease: A new paradigm.

Clinical studies suggest a link between peripheral insulin resistance and cognitive dysfunction. Interestingly, post-mortem analyses of Alzheimer disease (AD) subjects demonstrated insulin resistance in the brain proposing a role for cognitive deficits observed in AD. However, the mechanisms responsible for the onset of brain insulin resistance (BIR) need further elucidations. Biliverdin reductase-A (BVR-A) emerged as a unique Ser/Thr/Tyr kinase directly involved in the insulin signaling and represents an up-stream regulator of the insulin signaling cascade. Because we previously demonstrated the oxidative stress (OS)-induced impairment of BVR-A in human AD brain, we hypothesize that BVR-A dysregulation could be associated with the onset of BIR in AD. In the present work, we longitudinally analyze the age-dependent changes of (i) BVR-A protein levels and activation, (ii) total oxidative stress markers levels (PC, HNE, 3-NT) as well as (iii) IR/IRS1 levels and activation in the hippocampus of the triple transgenic model of AD (3xTg-AD) mice. Furthermore, ad hoc experiments have been performed in SH-SY5Y neuroblastoma cells to clarify the molecular mechanism(s) underlying changes observed in mice. Our results show that OS-induced impairment of BVR-A kinase activity is an early event, which starts prior the accumulation of Aß and tau pathology or the elevation of TNF-α, and that greatly contribute to the onset of BIR along the progression of AD pathology in 3xTg-Ad mice. Based on these evidence we, therefore, propose a new paradigm for which: OS-induced impairment of BVR-A is firstly responsible for a sustained activation of IRS1, which then causes the stimulation of negative feedback mechanisms (i.e. mTOR) aimed to turn-off IRS1 hyper-activity and thus BIR. Similar alterations characterize also the normal aging process in mice, positing BVR-A impairment as a possible bridge in the transition from normal aging to AD.

Alteration of mTOR signaling occurs early in the progression of Alzheimer disease (AD): analysis of brain from subjects with pre-clinical AD, amnestic mild cognitive impairment and late-stage AD.

The clinical symptoms of Alzheimer disease (AD) include a gradual memory loss and subsequent dementia, and neuropathological deposition of senile plaques and neurofibrillary tangles. At the molecular level, AD subjects present overt amyloid ß (Aß) production and tau hyperphosphorylation. Aß species have been proposed to overactivate the phosphoinositide3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) axis, which plays a central role in proteostasis. The current study investigated the status of the PI3K/Akt/mTOR pathway in post-mortem tissue from the inferior parietal lobule (IPL) at three different stages of AD: late AD, amnestic mild cognitive impairment (MCI) and pre-clinical AD (PCAD). Our findings suggest that the alteration of mTOR signaling and autophagy occurs at early stages of AD. We found a significant increase in Aß (1-42) levels, associated with reduction in autophagy (Beclin-1 and LC-3) observed in PCAD, MCI, and AD subjects. Related to the autophagy impairment, we found a hyperactivation of PI3K/Akt/mTOR pathway in IPL of MCI and AD subjects, but not in PCAD, along with a significant decrease in phosphatase and tensin homolog. An increase in two mTOR downstream targets, p70S6K and 4EBP1, occurred in AD and MCI subjects. Both AD and MCI subjects showed increased, insulin receptor substrate 1, a candidate biomarker of brain insulin resistance, and GSK-3ß, a kinase targeting tau phosphorylation. Nevertheless, tau phosphorylation was increased in the clinical groups. The results hint at a link between Aß and the PI3K/Akt/mTOR axis and provide further insights into the relationship between AD pathology and insulin resistance. In addition, we speculate that the alteration of mTOR signaling in the IPL of AD and MCI subjects, but not in PCAD, is due to the lack of substantial increase in oxidative stress. The figure represents the three different stages of Alzheimer Disease: Preclinical Alzheimer Disease (PCAD), Mild cognitive impairment (MCI) and late stage of Alzheimer Disease. The progression of the disease is associated with a reduction in autophagy (Beclin-1 and LC-3) observed in Inferior parietal lobe of PCAD, MCI, and AD subjects (light red). Related to the autophagy impairment, the graph shows the impairment of PI3K/Akt/mTOR in MCI and AD subjects (dark red).

Neuropathological role of PI3K/Akt/mTOR axis in Down syndrome brain.

Down syndrome (DS) is the most frequent genetic cause of intellectual disability characterized by the presence of three copies of chromosome 21 (Chr21). Individuals with DS have sufficient neuropathology for a diagnosis of Alzheimer's disease (AD) after the age of 40years. The aim of our study is to gain new insights in the molecular mechanisms impaired in DS subjects that eventually lead to the development of dementia. We evaluate the PI3K/Akt/mTOR axis in the frontal cortex from DS cases (under the age of 40years) and DS with AD neuropathology compared with age-matched controls (Young and Old). The PI3K/Akt/mTOR axis may control several key pathways involved in AD that, if aberrantly regulated, affect amyloid beta (Aß) deposition and tau phosphorylation. Our results show a hyperactivation of PI3K/Akt/mTOR axis in individuals with DS, with and without AD pathology, in comparison with respective controls. The PI3K/Akt/mTOR deregulation results in decreased autophagy, inhibition of IRS1 and GSK3ß activity. Moreover, our data suggest that aberrant activation of the PI3K/Akt/mTOR axis acts in parallel to RCAN1 in phosphorylating tau, in DS and DS/AD. In conclusion, this study provides insights into the neuropathological mechanisms that may be engaged during the development of AD in DS. We suggest that deregulation of this signaling cascade is already evident in young DS cases and persist in the presence of AD pathology. The impairment of the PI3K/Akt/mTOR axis in DS population might represent a key-contributing factor to the neurodegenerative process that culminates in Alzheimer-like dementia.

Redox proteomics analysis of HNE-modified proteins in Down syndrome brain: clues for understanding the development of Alzheimer disease.

Down syndrome (DS) is the most common genetic cause of intellectual disability, due to partial or complete triplication of chromosome 21. DS subjects are characterized by a number of abnormalities including premature aging and development of Alzheimer disease (AD) neuropathology after approximately 40 years of age. Several studies show that oxidative stress plays a crucial role in the development of neurodegeneration in the DS population. Increased lipid peroxidation is one of the main events causing redox imbalance within cells through the formation of toxic aldehydes that easily react with DNA, lipids, and proteins. In this study we used a redox proteomics approach to identify specific targets of 4-hydroxynonenal modifications in the frontal cortex from DS cases with and without AD pathology. We suggest that a group of identified proteins followed a specific pattern of oxidation in DS vs young controls, probably indicating characteristic features of the DS phenotype; a second group of identified proteins showed increased oxidation in DS/AD vs DS, thus possibly playing a role in the development of AD. The third group of comparison, DS/AD vs old controls, identified proteins that may be considered specific markers of AD pathology. All the identified proteins are involved in important biological functions including intracellular quality control systems, cytoskeleton network, energy metabolism, and antioxidant response. Our results demonstrate that oxidative damage is an early event in DS, as well as dysfunctions of protein-degradation systems and cellular protective pathways, suggesting that DS subjects are more vulnerable to oxidative damage accumulation that might contribute to AD development. Further, considering that the majority of proteins have been already demonstrated to be oxidized in AD brain, our results strongly support similarities with AD in DS.

Simvastatin attenuates the endothelial pro-thrombotic shift in saphenous vein grafts induced by Advanced glycation endproducts.

BACKGROUND: Advanced glycation endproducts (AGEs) and its receptors (RAGEs) are heterogeneous signaling proteins associated to diabetes and responsible of endothelial alterations leading to atherosclerosis progression and graft failure. The aim of this study was to investigate the role of statin in reducing AGEs related endothelial damage. METHODS: Endothelial cell(EC) obtained from leftovers of saphenous vein grafts of non-diabetic patients were incubated with AGEs (2 and 20 µM) and subsequently treated with Simvastatin. Neutrophils (PNM) adherence, ROS production and RAGE and peroxisome proliferator-activated receptors-gamma (PPAR-γ) expression were analyzed. As clinical validation of the in vitro findings, ECs of diabetic patients in optimized glycaemic control administered with a 3 weeks Simvastatin regimen were similarly processed. RESULTS: Simvastatin blunted the rise in PMN adhesion and ROS generation following stimulation of saphenous vein EC culture with AGEs in vitro. This effect was time dependent and was associated to an increase in PPAR-γ induction paralleled by a decrease in RAGEs expression. Parallely, data from diabetic patients administered with Simvastatin showed a similar significant reduction in PNM adhesion and ROS generation. Simvastatin treatment significantly decreased RAGEs expression in ECs from diabetic patients and determined a slight increase in PPAR-γ expression but the latter failed to reach statistical significance. Interference in the function of these two crucial pathways might be at the root of the statin antinflammatory and antithrombotic effect in the context of AGEs-associated damage. CONCLUSIONS: Despite the recently raised warning on the use of statins in the diabetic population, this study elucidates their cornerstone position in endothelial homeostasis of saphenous grafts in patients with controlled diabetes.

Involvement of oxidative stress in occurrence of relapses in multiple sclerosis: the spectrum of oxidatively modified serum proteins detected by proteomics and redox proteomics analysis.

Multiple sclerosis (MS) is an autoimmune inflammatory demyelinating disease of the central nervous system. Several evidences suggest that MS can be considered a multi-factorial disease in which both genetics and environmental factors are involved. Among proposed candidates, growing results support the involvement of oxidative stress (OS) in MS pathology. The aim of this study was to investigate the role of OS in event of exacerbations in MS on serum of relapsing-remitting (RR-MS) patients, either in relapsing or remitting phase, with respect to serum from healthy subjects. We applied proteomics and redox proteomics approaches to identify differently expressed and oxidatively modified proteins in the low-abundant serum protein fraction. Among differently expressed proteins ceruloplasmin, antithrombin III, clusterin, apolipoprotein E, and complement C3, were up-regulated in MS patients compared with healthy controls. Further by redox proteomics, vitamin D-binding protein showed a progressive trend of oxidation from remission to relapse, respect with controls. Similarly, the increase of oxidation of apolipoprotein A-IV confirmed that levels of OS are elevated with the progression of the disease. Our findings support the involvement of OS in MS and suggest that dysfunction of target proteins occurs upon oxidative damage and correlates with the pathology.

Impairment of proteostasis network in Down syndrome prior to the development of Alzheimer's disease neuropathology: redox proteomics analysis of human brain.

DS is the most frequent genetic cause of intellectual disability characterized by the anomalous presence of three copies of chromosome 21. One of the peculiar features of DS is the onset of Alzheimer's disease neuropathology after the age of 40years characterized by deposition of senile plaques and neurofibrillary tangles. Growing studies demonstrated that increased oxidative damage, accumulation of unfolded/damaged protein aggregates and dysfunction of intracellular degradative system are key players in neurodegenerative processes. In this study, redox proteomics approach was used to analyze the frontal cortex from DS subjects under the age of 40 compared with age-matched controls, and proteins found to be increasingly carbonylated were identified. Interestingly, our results showed that oxidative damage targets specifically different components of the intracellular quality control system such as GRP78, UCH-L1, V0-ATPase, cathepsin D and GFAP that couples with decreased activity of the proteasome and autophagosome formation observed. We also reported a slight but consistent increase of Aß 1-42 SDS- and PBS-soluble form and tau phosphorylation in DS versus CTR. We suggest that disturbance in the proteostasis network could contribute to the accumulation of protein aggregates, such as amyloid deposits and NFTs, which occur very early in DS. It is likely that a sub-optimal functioning of degradative systems occur in DS neurons, which in turn provide the basis for further accumulation of toxic protein aggregates. The results of this study suggest that oxidation of protein members of the proteostatis network is an early event in DS and might contribute to neurodegenerative phenomena.

Usefulness of preprocedural levels of advanced glycation end products to predict restenosis in patients with controlled diabetes mellitus undergoing drug-eluting stent implantation for stable angina pectoris (from the Prospective ARMYDA-AGEs Study).

Diabetes mellitus (DM) remains the main predictor of restenosis rates and cardiovascular events following successful percutaneous coronary intervention (PCI) despite the use of drug-eluting stents (DES). HbA1c <6.0% is considered an index of optimized metabolic control in patients with DM, but several studies are downsizing its role in the clinical management of these patients. Increasing evidence points at the role of advanced glycation end products (AGEs) in restenosis pathogenesis independently on Hb1AC levels. Thus, we investigated the predictive value of preprocedural AGE levels for in-stent restenosis in a population of euglycaemic diabetic patients undergoing PCI with DES implantation. One hundred twenty-five consecutive patients with DM in optimized glycemic control admitted for stable angina pectoris and treated with elective DES implantation at a tertiary hospital were prospectively included. The primary end point of the ARMYDA-AGEs study was to compare rates of angiographic ISR at 6 months after the intervention according to pre-PCI levels of AGEs. Secondary end points were the correlations of AGE levels with occurrence of periprocedural myocardial damage, major adverse cardiac events, and in-stent late loss at 6-month control coronary angiography. AGE levels >17 µM was found to be an independent predictor of ISR at 6 months and stent lumen loss. AGEs failed to predict occurrence of secondary endpoints. In conclusion, elevated AGE levels predict occurrence of in-stent restenosis after DES implantation in patients with DM on optimized glycemic control and might represent a dosable marker of adverse outcome after PCI.

HO-1/BVR-a system analysis in plasma from probable Alzheimer's disease and mild cognitive impairment subjects: a potential biochemical marker for the prediction of the disease.

Several studies showed increased oxidative and nitrosative stress in plasma from patients with Alzheimer's disease (AD), however, little and controversial knowledge has emerged about the antioxidant functionality of the heme oxygenase-1/biliverdin reductase-A (HO-1/BVR-A) system in blood. The current study reports increased levels of both HO-1 and BVR-A in plasma from probable AD patients, as a result of the increased oxidative environment. However, the increase of oxidative stress in plasma result also in the increase of BVR-A 3-nitrotyrosine levels and the decrease of BVR-A phosphotyrosine levels and reductase activity, suggesting that nitrosative stress play the prominent oxidative role in plasma during AD. Our data on HO-1/BVR-A status in plasma closely correlate with recent reports in hippocampus of subjects with AD and arguably its early form, mild cognitive impairment. Moreover, we show that alterations on HO-1/BVR-A system are tightly connected with cognitive decline indexed by Mini-Mental Status Exam scores. We hypothesize that the HO-1/BVR-A system status in plasma might reflect the ongoing situation in the brain, offering an important biochemical tool for the potential prediction of AD at the earliest stages of the disease.

Heme oxygenase-1 posttranslational modifications in the brain of subjects with Alzheimer disease and mild cognitive impairment.

Alzheimer disease (AD) is a neurodegenerative disorder characterized by progressive cognitive impairment and neuropathology. Oxidative and nitrosative stress plays a principal role in the pathogenesis of AD. The induction of the heme oxygenase-1/biliverdin reductase-A (HO-1/BVR-A) system in the brain represents one of the earliest mechanisms activated by cells to counteract the noxious effects of increased reactive oxygen species and reactive nitrogen species. Although initially proposed as a neuroprotective system in AD brain, the HO-1/BVR-A pathophysiological features are under debate. We previously reported alterations in BVR activity along with decreased phosphorylation and increased oxidative/nitrosative posttranslational modifications in the brain of subjects with AD and those with mild cognitive impairment (MCI). Furthermore, other groups proposed the observed increase in HO-1 in AD brain as a possible neurotoxic mechanism. Here we provide new insights about HO-1 in the brain of subjects with AD and MCI, the latter condition being the transitional phase between normal aging and early AD. HO-1 protein levels were significantly increased in the hippocampus of AD subjects, whereas HO-2 protein levels were significantly decreased in both AD and MCI hippocampi. In addition, significant increases in Ser-residue phosphorylation together with increased oxidative posttranslational modifications were found in the hippocampus of AD subjects. Interestingly, despite the lack of oxidative stress-induced AD neuropathology in cerebellum, HO-1 demonstrated increased Ser-residue phosphorylation and oxidative posttranslational modifications in this brain area, suggesting HO-1 as a target of oxidative damage even in the cerebellum. The significance of these findings is profound and opens new avenues into the comprehension of the role of HO-1 in the pathogenesis of AD.

Oxidative stress in HPV-driven viral carcinogenesis: redox proteomics analysis of HPV-16 dysplastic and neoplastic tissues.

Genital infection by high risk Human Papillomavirus (HR-HPV), although recognized as the main etio-pathogenetic factor of cervical cancer, is not per se sufficient to induce tumour development. Oxidative stress (OS) represents an interesting and under-explored candidate as a promoting factor in HPV-initiated carcinogenesis. To gain insight into the role of OS in cervical cancer, HPV-16 positive tissues were collected from patients with invasive squamous cervical carcinoma, from patients with High Grade dysplastic HPV lesions and from patients with no clinical evidence of HPV lesions. After virological characterization, modulation of proteins involved in the redox status regulation was investigated. ERp57 and GST were sharply elevated in dysplastic and neoplastic tissues. TrxR2 peaked in dysplastic samples while iNOS was progressively reduced in dysplastic and neoplastic samples. By redox proteomic approach, five proteins were found to have increased levels of carbonyls in dysplastic samples respect to controls namely: cytokeratin 6, actin, cornulin, retinal dehydrogenase and GAPDH. In carcinoma samples the peptidyl-prolyl cis-trans isomerase A, ERp57, serpin B3, Annexin 2 and GAPDH were found less oxidized than in dysplastic tissues. HPV16 neoplastic progression seems associated with increased oxidant environment. In dysplastic tissues the oxidative modification of DNA and proteins involved in cell morphogenesis and terminal differentiation may provide the conditions for the neoplastic progression. Conversely cancer tissues seem to attain an improved control on oxidative damage as shown by the selective reduction of carbonyl adducts on key detoxifying/pro-survival proteins.

Redox proteomics analyses of the influence of co-expression of wild-type or mutated LRRK2 and Tau on C. elegans protein expression and oxidative modification: relevance to Parkinson disease.

AIMS: The human LRRK2 gene has been identified as the most common causative gene of autosomal-dominantly inherited and idiopathic Parkinson disease (PD). The G2019S substitution is the most common mutation in LRRK2. The R1441C mutation also occurs in cases of familial PD, but is not as prevalent. Some cases of LRRK2-based PD exhibit Tau pathology, which suggests that alterations on LRRK2 activity affect the pathophysiology of Tau. To investigate how LRRK2 might affect Tau and the pathophysiology of PD, we generated lines of C. elegans expressing human LRRK2 [wild-type (WT) or mutated (G2019S or R1441C)] with and without V337M Tau. Expression and redox proteomics were used to identify the effects of LRRK2 (WT and mutant) on protein expression and oxidative modifications. RESULTS: Co-expression of WT LRRK2 and Tau led to increased expression of numerous proteins, including several 60S ribosomal proteins, mitochondrial proteins, and the V-type proton ATPase, which is associated with autophagy. C. elegans expressing mutant LRRK2 showed similar changes, but also showed increased protein oxidation and lipid peroxidation, the latter indexed as increased protein-bound 4-hydroxy-2-nonenal (HNE). INNOVATION: Our study brings new knowledge about the possible alterations induced by LRRK2 (WT and mutated) and Tau interactions, suggesting the involvement of G2019S and R1441C in Tau-dependent neurodegenerative processes. CONCLUSION: These results suggest that changes in LRRK2 expression or activity lead to corresponding changes in mitochondrial function, autophagy, and protein translation. These findings are discussed with reference to the pathophysiology of PD.

4-Hydroxy-2-nonenal, a reactive product of lipid peroxidation, and neurodegenerative diseases: a toxic combination illuminated by redox proteomics studies.

SIGNIFICANCE: Among different forms of oxidative stress, lipid peroxidation comprises the interaction of free radicals with polyunsaturated fatty acids, which in turn leads to the formation of highly reactive electrophilic aldehydes. Among these, the most abundant aldehydes are 4-hydroxy-2-nonenal (HNE) and malondialdehyde, while acrolein is the most reactive. HNE is considered a robust marker of oxidative stress and a toxic compound for several cell types. Proteins are particularly susceptible to modification caused by HNE, and adduct formation plays a critical role in multiple cellular processes. RECENT ADVANCES: With the outstanding progress of proteomics, the identification of putative biomarkers for neurodegenerative disorders has been the main focus of several studies and will continue to be a difficult task. CRITICAL ISSUES: The present review focuses on the role of lipid peroxidation, particularly of HNE-induced protein modification, in neurodegenerative diseases. By comparing results obtained in different neurodegenerative diseases, it may be possible to identify both similarities and specific differences in addition to better characterize selective neurodegenerative phenomena associated with protein dysfunction. Results obtained in our laboratory and others support the common deregulation of energy metabolism and mitochondrial function in neurodegeneration. FUTURE DIRECTIONS: Research towards a better understanding of the molecular mechanisms involved in neurodegeneration together with identification of specific targets of oxidative damage is urgently required. Redox proteomics will contribute to broaden the knowledge in regard to potential biomarkers for disease diagnosis and may also provide insight into damaged metabolic networks and potential targets for modulation of disease progression.

Serum proteomics in patients with diagnosis of abdominal aortic aneurysm.

BACKGROUND: Molecular mechanisms underlying abdominal aneurysm (AAA) formation and rupture are not well understood. Early detection and repair of AAA may reduce the high mortality rates associated with rupture. Serum proteomics allows the detection of alterations in the expression of proteins, guiding further studies on these target molecules as potential markers. Analysis of proteomic profile of asymptomatic patients with AAA allows the identification of reliable predictors or markers of disease presence or progression. METHODS: A proteomics approach based on two-dimensional electrophoresis and mass spectrometry was used to compare serum proteomic profiles of patients with AAA who are candidates for surgical repair compared with healthy controls. We analyzed in parallel the proteomic profile of subjects with cardiac heart failure to discriminate these two pathologies, which show similar pattern of systemic inflammation process. RESULTS: We identified in AAA subjects four serum proteins that show altered expression profile and that could be specifically linked to AAA pathology. We discuss the role of our identified proteins with their possible implications in disease outcome. CONCLUSIONS: This approach could provide an initial screening tool that may drive the basis for further research in the field of cardiovascular diseases. These results need to be validated in larger studies to find potential markers of AAA presence or progression to use in clinical settings. SUMMARY: A proteomics approach was used to compare serum proteomic profiles of patients with abdominal aortic aneurysm who are candidates for surgical repair compared with healthy controls. Four serum proteins showed altered expression profile that could be correlated with the pathology. This approach could provide an initial screening tool that may drive the basis for further research in the field of cardiovascular diseases.

Circulating biomarkers of protein oxidation for Alzheimer disease: expectations within limits.

Alzheimer disease (AD), the most common dementing disorder, is a multifactorial disease with complex etiology. Among different hypotheses proposed for AD one of the most corroborated is the "oxidative stress hypothesis". Although recent studies extensively demonstrated the specific oxidative modification of selected proteins in the brain of AD patients and how their dysfunction possibly correlates with the pathology, there is still an urgent need to extend these findings to peripheral tissue. So far very few studies showed oxidative damage of proteins in peripheral tissues and current findings need to be replicated. Another limit in AD research is represented by the lack of highly specific diagnostic tools for early diagnosis. For a full screening and early diagnosis, biomarkers easily detectable in biological samples, such as blood, are needed. The search of reliable biomarkers for AD in peripheral blood is a great challenge. A few studies described a set of plasma markers that differentiated AD from controls and were shown to be useful in predicting conversion from mild cognitive impairment, which is considered a prodromal stage, to AD. We review the current state of knowledge on peripheral oxidative biomarkers for AD, including proteomics, which might be useful for early diagnosis and prognosis.

Oxidative and nitrosative modifications of biliverdin reductase-A in the brain of subjects with Alzheimer's disease and amnestic mild cognitive impairment.

Biliverdin reductase-A (BVR-A) is a pleiotropic enzyme and plays pivotal role in the antioxidant defense against free radicals as well as in cell homeostasis. Together with heme oxygenase, BVR-A forms a powerful system involved in the cell stress response during neurodegenerative disorders including Alzheimer's disease (AD), whereas due to the serine/threonine/tyrosine kinase activity the enzyme regulates glucose metabolism and cell proliferation. In this paper, we report results that demonstrate BVR-A undergoes post-translational oxidative and nitrosative modifications in the hippocampus, but not cerebellum, of subjects with AD and amnestic mild cognitive impairment (MCI). A significant increase of nitrated BVR-A was demonstrated only in AD and MCI hippocampi, whereas no significant modifications were found in cerebellar tissue. In addition, a significant reduction in protein carbonyl-derivatives of BVR-A was found in both AD and MCI hippocampi (15% and 18%, respectively). Biliverdin reductase-bound 4-hydroxynonenals were not modified in hippocampi and cerebella from AD and MCI subjects. These results supported the hypothesis of a prevalence of nitrosative stress-induced modifications on BVR-A structure, and this evidence was confirmed by a significant upregulation of inducible nitric oxide synthase in hippocampal tissue of subjects with AD and MCI that was not present in cerebellum. In conclusion, nitrosative stress-induced modifications on hippocampal BVR-A are an early event in the pathogenesis of AD since they appear also in MCI subjects and could contribute to the antioxidant and metabolic derangement characteristic of these neurodegenerative disorders.

Oxidative stress occurs early in Down syndrome pregnancy: A redox proteomics analysis of amniotic fluid.

PURPOSE: The present study aims to evaluate a set of oxidative stress biomarkers in the amniotic fluid (AF) of women carrying Down syndrome (DS) fetuses that could prove in vivo the early occurrence of oxidative damage in DS. EXPERIMENTAL DESIGN: To assess the extent of protein oxidation in DS AF, we measured protein carbonylation and protein-bound HNE by slot-blot analysis, total and oxidized GSH levels by enzymatic assay and heat shock proteins (HSPs) thioredoxin (Trx) induction by Western blot. Further, by a redox proteomics approach specific targets of protein carbonylation were identified. RESULTS: We found increased levels of oxidative stress, as indexed by increased protein oxidation, lipid peroxidation, reduction of GSH and Trx levels and induction of the HSP response. By a redox proteomics approach, we identified selective proteins which showed increased oxidation in DS fetuses compared with healthy controls. The identified proteins are involved in iron homeostasis (ceruloplasmin and transferin), lipid metabolism (zinc-α2-glycoprotein, retinol-binding protein 4 and apolipoprotein A1) and inflammation (complement C9, α-1B-glycoprotein, collagen α-1V chain) with critical relevance in the clinical outcome of DS. CONCLUSIONS AND CLINICAL RELEVANCE: Our results indicate that oxidative damage is an early event in the DS pathogenesis and might contribute to the development of deleterious DS phenotypes, including abnormal development and AD-like neuropathology.