Glycosylated proteins play important roles in cell-to-cell interactions, immunosurveillance, and a variety of receptor-mediated and specific protein functions through a highly complex repertoire of ...glycan structures. Aberrant glycosylation has been implicated in cancer for many years.
We performed specific MALDI mass spectrometry (MS)-based glycomic profile analyses of permethylated glycans in sera from breast cancer patients (12, stage I; 11, stage II; 9, stage III; and 50, stage IV) along with sera from 27 disease-free women. The serum glycoproteins were enzymatically deglycosylated, and the released glycans were purified and quantitatively permethylated before their MALDI-MS analyses. We applied various statistical analysis tools, including ANOVA and principal component analysis, to evaluate the MS profiles.
Two statistical procedures implicated several sialylated and fucosylated N-glycan structures as highly probable biomarkers. Quantitative changes according to a cancer stage resulted when we categorized the glycans according to molecular size, number of oligomer branches, and abundance of sugar residues. Increases in sialylation and fucosylation of glycan structures appeared to be indicative of cancer progression. Different statistical evaluations confirmed independently that changes in the relative intensities of 8 N-glycans are characteristic of breast cancer (P < 0.001), whereas other glycan structures might contribute additionally to distinctions in the statistically recognizable patterns (different stages).
MS-based N-glycomic profiling of serum-derived constituents appears promising as a highly sensitive and informative approach for staging the progression of cancer.
Comparative glycan quantification has thus far been a challenging task due to the lack of sensitive and reproducible analytical techniques. We introduce here a combination of quantitative ...permethylation and isotope labeling of glycans as an approach (C-GlycoMAP) allowing precise comparison between different samples in a single MALDI-MS analysis. Samples are either methylated or deuteriomethylated prior to their mixing and mass spectrometric acquisitions. Comparative analyses are based on the ratio of the two isotopically distinct forms of the same glycan structure, thus allowing a direct absolute evaluation of the intensities of the two forms originating from two different biological samples (e.g., control and diseased). The direct comparison between the two forms eliminates a MALDI-MS low m/z bias commonly associated with this technique. These comparative analyses are highly reliable when the intensity ratios of the two forms lie between 0.125 and 6, an overall reproducibility better than 30% (RSD). The value of C-GlycoMAP is demonstrated here for N-glycans derived from human blood serum collected from a healthy individual and a breast cancer patient as well as for O-glycans derived from normal and cancer cell extracts.
Glycomic profiles derived from human blood sera of 10 healthy males were compared to those from 24 prostate cancer patients. The profiles were acquired using MALDI−MS of permethylated N-glycans ...released from 10-μL sample aliquots. Quantitative permethylation was attained using solid-phase permethylation. Principal component analysis of the glycomic profiles revealed significant differences among the two sets, allowing their distinct clustering. The first principal component distinguished the 24 prostate cancer patients from the healthy individuals. It was determined that fucosylation of glycan structures is generally higher in cancer samples (ANOVA test p-value of 0.0006). Although more than 50 N-glycan structures were determined, 12 glycan structures, of which six were fucosylated, were significantly different between the two sample sets. Significant differences were confirmed through two independent statistical tests (ANOVA and ROC analyses). Ten of these structures had significantly higher relative intensities in the case of the cancer samples, while the other two were less abundant in the cancer samples. All 12 structures were statistically significant, as suggested by their very low ANOVA scores (<0.001) and ROC analysis, with area under the curve values close to 1 or 0. Accordingly, these structures can be considered as cancer−specific glycans and potential prostate cancer biomarkers. Therefore, serum glycomic profiling appears worthy of further investigation to define its role in cancer early detection and prognostication. Keywords: prostate cancer • glycomics • human blood serum • biomarkers • MALDI-TOF • mass spectrometry
Hepatocellular carcinoma (HCC) represents an increasing health problem in the United States. Serum alpha-fetoprotein, the currently used clinical marker, is elevated in only approximately 60% of HCC ...patients; therefore, the identification of additional markers is expected to have significant public health impact. The objective of our study was to quantitatively assess N-glycans originating from serum glycoproteins as alternative markers for the detection of HCC.
We used matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for quantitative comparison of 83 N-glycans in serum samples of 202 participants (73 HCC cases, 77 age- and gender-matched cancer-free controls, and 52 patients with chronic liver disease). N-glycans were enzymatically released from serum glycoproteins and permethylated before mass spectrometric quantification.
The abundance of 57 N-glycans was significantly altered in HCC patients compared with controls. The sensitivity of six individual glycans evaluated for separation of HCC cases from population controls ranged from 73% to 90%, and the specificity ranged from 36% to 91%. A combination of three selected N-glycans was sufficient to classify HCC with 90% sensitivity and 89% specificity in an independent validation set of patients with chronic liver disease. The three N-glycans remained associated with HCC after adjustment for chronic viral infection and other known covariates, whereas the other glycans increased significantly at earlier stages of the progression of chronic viral infection to HCC.
A set of three identified N-glycans is sufficient for the detection of HCC with 90% prediction accuracy in a population with high rates of hepatitis C viral infection. Further evaluation of a wider clinical utility of these candidate markers is warranted.
The consumption of a diet low in fat and enhanced by fruits and vegetables, especially rich in phenolic compounds, may reduce risks of many civilization diseases. The use of traditional medicines, ...mainly derived from plant sources, has become an attractive segment in the management of many lifestyle diseases. Concerning the application of dietary supplements (based on phenolic compounds) in common practice, the ongoing debate over possible adverse effects of certain nutrients and dosage levels is of great importance. Since dietary supplements are not classified as drugs, their potential toxicities and interactions have not been thoroughly evaluated. First, this review will introduce phenolic compounds as natural substances beneficial for human health. Second, the potential dual mode of action of flavonoids will be outlined. Third, potential deleterious impacts of phenolic compounds utilization will be discussed: pro-oxidant and estrogenic activities, cancerogenic potential, cytotoxic effects, apoptosis induction and flavonoid-drug interaction. Finally, future trends within the research field will be indicated.
This paper presents computational methods to analyze MALDI-TOF mass spectrometry data for quantitative comparison of peptides and glycans in serum. The methods are applied to identify candidate ...biomarkers in serum samples of 203 participants from Egypt; 73 hepatocellular carcinoma (HCC) cases, 52 patients with chronic liver disease (CLD) consisting of cirrhosis and fibrosis cases, and 78 population controls. Two complementary sample preparation methods were applied prior to generating mass spectra: (1) low molecular weight (LMW) enrichment of each serum sample was carried out for MALDI-TOF quantification of peptides, and (2) glycans were enzymatically released from proteins in each serum sample and permethylated for MALDI-TOF quantification of glycans. A peak selection algorithm was applied to identify the most useful peptide and glycan peaks for accurate detection of HCC cases from high-risk population of patients with CLD. In addition to global peaks selected by the whole population based approach, where identically labeled patients are treated as a single group, subgroup-specific peaks were identified by searching for peaks that are differentially abundant in a subgroup of patients only. The peak selection process was preceded by peak screening, where we eliminated peaks that have significant association with covariates such as age, gender, and viral infection based on the peptide and glycan spectra from population controls. The performance of the selected peptide and glycan peaks was evaluated in terms of their ability in detecting HCC cases from patients with CLD in a blinded validation set and through the cross-validation method. Finally, we investigated the possibility of using both peptides and glycans in a panel to enhance the diagnostic capability of these candidate markers. Further evaluation is needed to examine the potential clinical utility of the candidate peptide and glycan markers identified in this study.
Cataract, the opacification of eye lens, is the leading cause of blindness worldwide. At present, the only remedy is surgical removal of the cataractous lens and substitution with a lens made of ...synthetic polymers. However, besides significant costs of operation and possible complications, an artificial lens just does not have the overall optical qualities of a normal one. Hence it remains a significant public health problem, and biochemical solutions or pharmacological interventions that will maintain the transparency of the lens are highly required. Naturally, there is a persistent demand for suitable biological models. The ocular lens would appear to be an ideal organ for maintaining culture conditions because of lacking blood vessels and nerves. The lens in vivo obtains its nutrients and eliminates waste products via diffusion with the surrounding fluids. Lens opacification observed in vivo can be mimicked in vitro by addition of the cataractogenic agent sodium selenite (Na2SeO3) to the culture medium. Moreover, since an overdose of sodium selenite induces also cataract in young rats, it became an extremely rapid and convenient model of nuclear cataract in vivo. The main focus of this review will be on selenium (Se) and its salt sodium selenite, their toxicological characteristics and safety data in relevance of modelling cataractogenesis, either under in vivo or in vitro conditions. The studies revealing the mechanisms of lens opacification induced by selenite are highlighted, the representatives from screening for potential anti-cataract agents are listed.
Hyperglycaemia-induced oxidative stress makes an important contribution to the aetiology of diabetic neuropathy. The aim of our study was to evaluate the effect of the antioxidant stobadine (STB) in ...comparison with a treatment by a high-dose of alpha-lipoic acid (ALA), on the neurological consequences of chronic hyperglycaemia in an animal model of diabetes in Wistar rats (16 weeks old), made diabetic by streptozotocin (STZ 3 x 20 mg i.v.). Neuropathy was evaluated electrophysiologically by measuring motor nerve conduction velocity (NCV) in the 4th and 8th week in vivo and motor NCV and resistance to ischaemic conduction failure (RICF) of the sciatic nerve in the 10th week of the experiment in vitro. The therapy with ALA (100 mg/kg i.p., 5 times a week) and STB (25 mg/kg i.p., 5 times a week) had a significant effect on NCV in vivo in the 8th week of the experiment and no effect in the 10th week in vitro. The RICF elevated in diabetic animals was significantly modified by ALA. The effect of the antioxidant STB on NCV was comparable with that of ALA, while RICF was affected only by ALA. We conclude that treatment with appropriate antioxidants might partially prevent nerve dysfunction in diabetic rats.
Abstract Measurements of enzyme kinetics of renal Na,K-ATPase were used for characterization of ATP- and Na+ -binding sites in rats that were subjected to 10 days of moderate inflammation that was ...induced by a single dose of Escherichia coli lipopolysaccharides (LPSs) at a dose of 1 mg kg−1 body weight. We hypothesized that LPSs might initiate a malfunction of renal Na,K-ATPase, which is a key enzyme involved in regulation of sodium homeostasis in the organism. We also investigated the potential effect that fish oil (FO) has in the prevention of Na,K-ATPase alterations by administering FO daily at a dose of 30 mg kg−1 . Alone, LPS elevated the level of C-reactive protein by more than 500% and free radicals by 36% in plasma, as indicated by an increased level of malondialdehyde. The Na,K-ATPase was slightly altered in the vicinity of the ATP-binding site as suggested by the 9% increase of the concentration of ATP necessary for half-maximal activation of the enzyme, thus indicating a deteriorated binding of ATP as a consequence of inflammation. Daily supplementation of FO partly attenuated LPS-induced injury, as observed by a significant decrease in the plasma levels of C-reactive protein and free radicals, hence maintaining the activity of renal Na,K-ATPase to the level of healthy control animals. In conclusion, our findings showed that FO prevented an excessive malondialdehyde production in LPS-treated animals and stabilized renal Na,K-ATPase.
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