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Sample records for plasma protein carbonylation

  1. Plasma protein carbonyl levels and breast cancer risk.

    Science.gov (United States)

    Rossner, Pavel; Terry, Mary Beth; Gammon, Marilie D; Agrawal, Meenakshi; Zhang, Fang Fang; Ferris, Jennifer S; Teitelbaum, Susan L; Eng, Sybil M; Gaudet, Mia M; Neugut, Alfred I; Santella, Regina M

    2007-01-01

    To study the role of oxidative stress in breast cancer risk, we analysed plasma levels of protein carbonyls in 1050 cases and 1107 controls. We found a statistically significant trend in breast cancer risk in relation to increasing quartiles of plasma protein carbonyl levels (OR = 1.2, 95% CI = 0.9-1.5; OR = 1.5, 95% CI = 1.2-2.0; OR = 1.6, 95% CI = 1.2-2.1, for the 2(nd), 3(rd) and 4(th) quartile relative to the lowest quartile, respectively, P for trend = 0.0001). The increase in risk was similar for younger ( or = 15 grams/day for 4(th) quartile versus lowest quartile OR = 2.3, 95% CI = 1.1-4.7), and hormone replacement therapy use (HRT, OR = 2.6, 95% CI = 1.6-4.4 for 4(th) quartile versus lowest quartile). The multiplicative interaction terms were statistically significant only for physical activity and HRT. The positive association between plasma protein carbonyl levels and breast cancer risk was also observed when the analysis was restricted to women who had not received chemotherapy or radiation therapy prior to blood collection. Among controls, oxidized protein levels significantly increased with cigarette smoking and higher fruit and vegetable consumption, and decreased with alcohol consumption >30 grams per day. Women with higher levels of plasma protein carbonyl and urinary 15F(2t)-isoprostane had an 80% increase in breast cancer risk (OR = 1.8, 95% CI = 1.2-2.6) compared to women with levels below the median for both markers of oxidative stress. In summary, our results suggest that increased plasma protein carbonyl levels may be associated with breast cancer risk.

  2. Age-related variations of protein carbonyls in human saliva and plasma: is saliva protein carbonyls an alternative biomarker of aging?

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    Wang, Zhihui; Wang, Yanyi; Liu, Hongchen; Che, Yuwei; Xu, Yingying; E, Lingling

    2015-06-01

    Free radical hypothesis which is one of the most acknowledged aging theories was developed into oxidative stress hypothesis. Protein carbonylation is by far one of the most widely used markers of protein oxidation. We studied the role of age and gender in protein carbonyl content of saliva and plasma among 273 Chinese healthy subjects (137 females and 136 males aged between 20 and 79) and discussed the correlation between protein carbonyl content of saliva and plasma. Protein carbonyl content of saliva and plasma were, respectively, 2.391 ± 0.639 and 0.838 ± 0.274 nmol/mg. Variations of saliva and plasma different age groups all reached significant differences in both male and female (all p saliva and plasma protein carbonyls were found to be significantly correlated with age (r = 0.6582 and r = 0.5176, all p saliva and plasma protein carbonyl levels (all p > 0.05). Saliva and plasma protein carbonyls were positively related (r = 0.4405, p saliva and plasma protein carbonyls/ferric reducing ability of plasma (FRAP) ratios were proved to be significantly correlated with age (r = 0.7796 and r = 0.6938, all p saliva protein carbonyls/FRAP ratio and plasma protein carbonyls/FRAP ratio were also correlated (r = 0.5573, p saliva protein carbonyls seem to be an alternative biomarker of aging while the mechanisms of protein carbonylation and oxidative stress and the relationship between saliva protein carbonyls and diseases need to be further investigated.

  3. A biotin enrichment strategy identifies novel carbonylated amino acids in proteins from human plasma

    DEFF Research Database (Denmark)

    Havelund, Jesper F.; Wojdyla, Katarzyna; Davies, Michael J.

    2017-01-01

    Protein carbonylation is an irreversible protein oxidation correlated with oxidative stress, various diseases and ageing. Here we describe a peptide-centric approach for identification and characterisation of up to 14 different types of carbonylated amino acids in proteins. The modified residues...... in vitro metal ion-catalysed oxidation. Furthermore, we assigned 133 carbonylated sites in 36 proteins in native human plasma protein samples. The optimised workflow enabled detection of 10 hitherto undetected types of carbonylated amino acids in proteins: aldehyde and ketone modifications of leucine...

  4. A biotin enrichment strategy identifies novel carbonylated amino acids in proteins from human plasma.

    Science.gov (United States)

    Havelund, Jesper F; Wojdyla, Katarzyna; Davies, Michael J; Jensen, Ole N; Møller, Ian Max; Rogowska-Wrzesinska, Adelina

    2017-03-06

    Protein carbonylation is an irreversible protein oxidation correlated with oxidative stress, various diseases and ageing. Here we describe a peptide-centric approach for identification and characterisation of up to 14 different types of carbonylated amino acids in proteins. The modified residues are derivatised with biotin-hydrazide, enriched and characterised by tandem mass spectrometry. The strength of the method lies in an improved elution of biotinylated peptides from monomeric avidin resin using hot water (95°C) and increased sensitivity achieved by reduction of analyte losses during sample preparation and chromatography. For the first time MS/MS data analysis utilising diagnostic biotin fragment ions is used to pinpoint sites of biotin labelling and improve the confidence of carbonyl peptide assignments. We identified a total of 125 carbonylated residues in bovine serum albumin after extensive in vitro metal ion-catalysed oxidation. Furthermore, we assigned 133 carbonylated sites in 36 proteins in native human plasma protein samples. The optimised workflow enabled detection of 10 hitherto undetected types of carbonylated amino acids in proteins: aldehyde and ketone modifications of leucine, valine, alanine, isoleucine, glutamine, lysine and glutamic acid (+14Da), an oxidised form of methionine - aspartate semialdehyde (-32Da) - and decarboxylated glutamic acid and aspartic acid (-30Da).

  5. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins: Focus on sample preparation and derivatization conditions.

    Science.gov (United States)

    Weber, Daniela; Davies, Michael J; Grune, Tilman

    2015-08-01

    Protein oxidation is involved in regulatory physiological events as well as in damage to tissues and is thought to play a key role in the pathophysiology of diseases and in the aging process. Protein-bound carbonyls represent a marker of global protein oxidation, as they are generated by multiple different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples.

  6. Validation of protein carbonyl measurement

    DEFF Research Database (Denmark)

    Augustyniak, Edyta; Adam, Aisha; Wojdyla, Katarzyna;

    2015-01-01

    Protein carbonyls are widely analysed as a measure of protein oxidation. Several different methods exist for their determination. A previous study had described orders of magnitude variance that existed when protein carbonyls were analysed in a single laboratory by ELISA using different commercial...

  7. Determination of Carbonyl Group Content in Plasma Proteins as a Useful Marker to Assess Impairment in Antioxidant Defense in Patients with Eales′ Disease

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    Rajesh Mohanraj

    2004-01-01

    Full Text Available Purpose: Formation of protein carbonyl groups is considered an early biomarker for the oxidant/antioxidant barrier impairment in various inflammatory diseases. We evaluated the intensity of free radical reactions in patients with Eales′ disease, an idiopathic inflammatory condition of the retina. Methods: Twenty patients with Eales′ disease in active vasculitis stage, 15 patients with Eales′ disease in healed vasculitis stage and 20 healthy control subjects were recruited for the study. Plasma protein carbonyl groups,plasma glutathione (GSH superoxide dismutase (SOD activity and thiobarbituric acid reactive substances (TBARS were determined in erythrocytes. Results: Plasma protein carbonyl content was elevated by a factor of 3.5 and 1.8 respectively in active and healed vasculitis stages. The increase of carbonyl group content in active and healed stage of patients with Eales′ disease correlated with diminished SOD activity and GSH content. There was also increased accumulation of TBARS in active and healed vasculitis stages of Eales′ disease, and this correlated with diminished SOD activity. Conclusion: Our results showed that protein carbonyl group content increases with severity of Eales′ disease. The increase in carbonyl content correlated with diminished antioxidant status. This confirms an earlier report that free radical mediated tissue damage occurs in Eales′ disease. The determination of protein carbonyl content may be used as a simple biomarker to monitor the efficacy of antioxidant supplementation in controlling retinal vasculitis in patients with Eales′ disease.

  8. Validation of protein carbonyl measurement

    DEFF Research Database (Denmark)

    Augustyniak, Edyta; Adam, Aisha; Wojdyla, Katarzyna

    2015-01-01

    Protein carbonyls are widely analysed as a measure of protein oxidation. Several different methods exist for their determination. A previous study had described orders of magnitude variance that existed when protein carbonyls were analysed in a single laboratory by ELISA using different commercial...... kits. We have further explored the potential causes of variance in carbonyl analysis in a ring study. A soluble protein fraction was prepared from rat liver and exposed to 0, 5 and 15min of UV irradiation. Lyophilised preparations were distributed to six different laboratories that routinely undertook...... protein carbonyl analysis across Europe. ELISA and Western blotting techniques detected an increase in protein carbonyl formation between 0 and 5min of UV irradiation irrespective of method used. After irradiation for 15min, less oxidation was detected by half of the laboratories than after 5min...

  9. Protein carbonylation and adipocyte mitochondrial function.

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    Curtis, Jessica M; Hahn, Wendy S; Stone, Matthew D; Inda, Jacob J; Droullard, David J; Kuzmicic, Jovan P; Donoghue, Margaret A; Long, Eric K; Armien, Anibal G; Lavandero, Sergio; Arriaga, Edgar; Griffin, Timothy J; Bernlohr, David A

    2012-09-21

    Carbonylation is the covalent, non-reversible modification of the side chains of cysteine, histidine, and lysine residues by lipid peroxidation end products such as 4-hydroxy- and 4-oxononenal. In adipose tissue the effects of such modifications are associated with increased oxidative stress and metabolic dysregulation centered on mitochondrial energy metabolism. To address the role of protein carbonylation in the pathogenesis of mitochondrial dysfunction, quantitative proteomics was employed to identify specific targets of carbonylation in GSTA4-silenced or overexpressing 3T3-L1 adipocytes. GSTA4-silenced adipocytes displayed elevated carbonylation of several key mitochondrial proteins including the phosphate carrier protein, NADH dehydrogenase 1α subcomplexes 2 and 3, translocase of inner mitochondrial membrane 50, and valyl-tRNA synthetase. Elevated protein carbonylation is accompanied by diminished complex I activity, impaired respiration, increased superoxide production, and a reduction in membrane potential without changes in mitochondrial number, area, or density. Silencing of the phosphate carrier or NADH dehydrogenase 1α subcomplexes 2 or 3 in 3T3-L1 cells results in decreased basal and maximal respiration. These results suggest that protein carbonylation plays a major instigating role in cytokine-dependent mitochondrial dysfunction and may be linked to the development of insulin resistance in the adipocyte.

  10. Pattern of occurrence and occupancy of carbonylation sites in proteins

    DEFF Research Database (Denmark)

    Rao, R Shyama Prasad; Møller, Ian Max

    2011-01-01

    Proteins are targets for modification by reactive oxygen species, and carbonylation is an important irreversible modification that increases during oxidative stress. While information on protein carbonylation is accumulating, its pattern is not yet understood. We have made a meta-analysis of the ......Proteins are targets for modification by reactive oxygen species, and carbonylation is an important irreversible modification that increases during oxidative stress. While information on protein carbonylation is accumulating, its pattern is not yet understood. We have made a meta...... sites. Comparison of metal-catalyzed oxidation of two closely related proteins indicates that this type of carbonylation might not be very specific in proteins. Interestingly, carbonylated sites show a very strong tendency to cluster together in the protein primary sequence hinting at some sort...

  11. Proteomic identification of carbonylated proteins in F344 rat hippocampus after 1-bromopropane exposure

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    Huang, Zhenlie [Department of Occupational and Environmental Health, Nagoya University Graduate School of Medicine, Nagoya 466‐8550 (Japan); Department of Toxicology, Guangdong Prevention and Treatment Center for Occupational Diseases, Guangzhou 510‐300 (China); Ichihara, Sahoko [Graduate School of Regional Innovation Studies, Mie University, Tsu 514‐8507 (Japan); Oikawa, Shinji [Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Mie 514‐8507 (Japan); Chang, Jie; Zhang, Lingyi; Subramanian, Kaviarasan; Mohideen, Sahabudeen Sheik [Department of Occupational and Environmental Health, Nagoya University Graduate School of Medicine, Nagoya 466‐8550 (Japan); Ichihara, Gaku, E-mail: gak@med.nagoya-u.ac.jp [Department of Occupational and Environmental Health, Nagoya University Graduate School of Medicine, Nagoya 466‐8550 (Japan)

    2012-08-15

    1-Bromopropane (1-BP) is neurotoxic in both experimental animals and humans. Previous proteomic analysis of rat hippocampus implicated alteration of protein expression in oxidative stress, suggesting that oxidative stress plays a role in 1-BP-induced neurotoxicity. To understand this role at the protein level, we exposed male F344 rats to 1-BP at 0, 400, or 1000 ppm for 8 h/day for 1 week or 4 weeks by inhalation and quantitated changes in hippocampal protein carbonyl using a protein carbonyl assay, two-dimensional gel electrophoresis (2-DE), immunoblotting, and matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF-TOF/MS). Hippocampal reactive oxygen species and protein carbonyl were significantly increased, demonstrating 1-BP-associated induction of oxidative stress and protein damage. MALDI-TOF-TOF/MS identified 10 individual proteins with increased carbonyl modification (p < 0.05; fold-change ≥ 1.5). The identified proteins were involved in diverse biological processes including glycolysis, ATP production, tyrosine catabolism, GTP binding, guanine degradation, and neuronal metabolism of dopamine. Hippocampal triosephosphate isomerase (TPI) activity was significantly reduced and negatively correlated with TPI carbonylation (p < 0.001; r = 0.83). Advanced glycation end-product (AGE) levels were significantly elevated both in the hippocampus and plasma, and hippocampal AGEs correlated negatively with TPI activity (p < 0.001; r = 0.71). In conclusion, 1-BP-induced neurotoxicity in the rat hippocampus seems to involve oxidative damage of cellular proteins, decreased TPI activity, and elevated AGEs. -- Highlights: ► 1-BP increases hippocampal ROS levels and hippocampal and plasma protein carbonyls. ► 1-BP increases TPI carbonylation and decreases TPI activity in the hippocampus. ► 1-BP increases hippocampal and plasma AGE levels.

  12. Proteomic identification of carbonylated proteins in F344 rat hippocampus after 1-bromopropane exposure.

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    Huang, Zhenlie; Ichihara, Sahoko; Oikawa, Shinji; Chang, Jie; Zhang, Lingyi; Subramanian, Kaviarasan; Mohideen, Sahabudeen Sheik; Ichihara, Gaku

    2012-08-15

    1-Bromopropane (1-BP) is neurotoxic in both experimental animals and humans. Previous proteomic analysis of rat hippocampus implicated alteration of protein expression in oxidative stress, suggesting that oxidative stress plays a role in 1-BP-induced neurotoxicity. To understand this role at the protein level, we exposed male F344 rats to 1-BP at 0, 400, or 1000 ppm for 8h/day for 1 week or 4 weeks by inhalation and quantitated changes in hippocampal protein carbonyl using a protein carbonyl assay, two-dimensional gel electrophoresis (2-DE), immunoblotting, and matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF-TOF/MS). Hippocampal reactive oxygen species and protein carbonyl were significantly increased, demonstrating 1-BP-associated induction of oxidative stress and protein damage. MALDI-TOF-TOF/MS identified 10 individual proteins with increased carbonyl modification (p < 0.05; fold-change ≥ 1.5). The identified proteins were involved in diverse biological processes including glycolysis, ATP production, tyrosine catabolism, GTP binding, guanine degradation, and neuronal metabolism of dopamine. Hippocampal triosephosphate isomerase (TPI) activity was significantly reduced and negatively correlated with TPI carbonylation (p < 0.001; r = 0.83). Advanced glycation end-product (AGE) levels were significantly elevated both in the hippocampus and plasma, and hippocampal AGEs correlated negatively with TPI activity (p < 0.001; r = 0.71). In conclusion, 1-BP-induced neurotoxicity in the rat hippocampus seems to involve oxidative damage of cellular proteins, decreased TPI activity, and elevated AGEs.

  13. Protein carbonylation and metal-catalyzed protein oxidation in a cellular perspective

    DEFF Research Database (Denmark)

    Møller, Ian Max; Rogowska-Wrzesinska, Adelina; Rao, R S P

    2011-01-01

    Proteins can become oxidatively modified in many different ways, either by direct oxidation of amino acid side chains and protein backbone or indirectly by conjugation with oxidation products of polyunsaturated fatty acids and carbohydrates. While reversible oxidative modifications are thought...... to be relevant in physiological processes, irreversible oxidative modifications are known to contribute to cellular damage and disease. The most well-studied irreversible protein oxidation is carbonylation. In this work we first examine how protein carbonylation occurs via metal-catalyzed oxidation (MCO) in vivo...... and in vitro with an emphasis on cellular metal ion homeostasis and metal binding. We then review proteomic methods currently used for identifying carbonylated proteins and their sites of modification. Finally, we discuss the identified carbonylated proteins and the pattern of carbonylation sites in relation...

  14. Protein carbonylation, protein aggregation and neuronal cell death in a murine model of multiple sclerosis

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    Dasgupta, Anushka

    Many studies have suggested that oxidative stress plays an important role in the pathophysiology of both multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Yet, the mechanism by which oxidative stress leads to tissue damage in these disorders is unclear. Recent work from our laboratory has revealed that protein carbonylation, a major oxidative modification caused by severe and/or chronic oxidative stress conditions, is elevated in MS and EAE. Furthermore, protein carbonylation has been shown to alter protein structure leading to misfolding/aggregation. These findings prompted me to hypothesize that carbonylated proteins, formed as a consequence of oxidative stress and/or decreased proteasomal activity, promote protein aggregation to mediate neuronal apoptosis in vitro and in EAE. To test this novel hypothesis, I first characterized protein carbonylation, protein aggregation and apoptosis along the spinal cord during the course of myelin-oligodendrocyte glycoprotein (MOG)35-55 peptide-induced EAE in C57BL/6 mice [Chapter 2]. The results show that carbonylated proteins accumulate throughout the course of the disease, albeit by different mechanisms: increased oxidative stress in acute EAE and decreased proteasomal activity in chronic EAE. I discovered not only that there is a temporal correlation between protein carbonylation and apoptosis but also that carbonyl levels are significantly higher in apoptotic cells. A high number of juxta-nuclear and cytoplasmic protein aggregates containing the majority of the oxidized proteins are also present during the course of EAE, which seems to be due to reduced autophagy. In chapter 3, I show that when gluthathione levels are reduced to those in EAE spinal cord, both neuron-like PC12 (nPC12) cells and primary neuronal cultures accumulate carbonylated proteins and undergo cell death (both by necrosis and apoptosis). Immunocytochemical and biochemical studies also revealed a temporal

  15. Two-dimensional gel electrophoretic detection of protein carbonyls derivatized with biotin-hydrazide.

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    Wu, Jinzi; Luo, Xiaoting; Jing, Siqun; Yan, Liang-Jun

    2016-04-15

    Protein carbonyls are protein oxidation products that are often used to measure the magnitude of protein oxidative damage induced by reactive oxygen or reactive nitrogen species. Protein carbonyls have been found to be elevated during aging and in age-related diseases such as stroke, diabetes, and neurodegenerative diseases. In the present article, we provide detailed protocols for detection of mitochondrial protein carbonyls labeled with biotin-hydrazide followed by 2-dimensional isoelectric focusing (IEF)/SDS-PAGE and Western blotting probed with horse-radish peroxidase-conjugated streptavidin. The presented procedures can also be modified for detection of carbonylation of non-mitochondrial proteins.

  16. Structural characteristics of green tea catechins for formation of protein carbonyl in human serum albumin.

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    Ishii, Takeshi; Mori, Taiki; Ichikawa, Tatsuya; Kaku, Maiko; Kusaka, Koji; Uekusa, Yoshinori; Akagawa, Mitsugu; Aihara, Yoshiyuki; Furuta, Takumi; Wakimoto, Toshiyuki; Kan, Toshiyuki; Nakayama, Tsutomu

    2010-07-15

    Catechins are polyphenolic antioxidants found in green tea leaves. Recent studies have reported that various polyphenolic compounds, including catechins, cause protein carbonyl formation in proteins via their pro-oxidant actions. In this study, we evaluate the formation of protein carbonyl in human serum albumin (HSA) by tea catechins and investigate the relationship between catechin chemical structure and its pro-oxidant property. To assess the formation of protein carbonyl in HSA, HSA was incubated with four individual catechins under physiological conditions to generate biotin-LC-hydrazide labeled protein carbonyls. Comparison of catechins using Western blotting revealed that the formation of protein carbonyl in HSA was higher for pyrogallol-type catechins than the corresponding catechol-type catechins. In addition, the formation of protein carbonyl was also found to be higher for the catechins having a galloyl group than the corresponding catechins lacking a galloyl group. The importance of the pyrogallol structural motif in the B-ring and the galloyl group was confirmed using methylated catechins and phenolic acids. These results indicate that the most important structural element contributing to the formation of protein carbonyl in HSA by tea catechins is the pyrogallol structural motif in the B-ring, followed by the galloyl group. The oxidation stability and binding affinity of tea catechins with proteins are responsible for the formation of protein carbonyl, and consequently the difference in these properties of each catechin may contribute to the magnitude of their biological activities.

  17. A step-by-step protocol for assaying protein carbonylation in biological samples.

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    Colombo, Graziano; Clerici, Marco; Garavaglia, Maria Elisa; Giustarini, Daniela; Rossi, Ranieri; Milzani, Aldo; Dalle-Donne, Isabella

    2016-04-15

    Protein carbonylation represents the most frequent and usually irreversible oxidative modification affecting proteins. This modification is chemically stable and this feature is particularly important for storage and detection of carbonylated proteins. Many biochemical and analytical methods have been developed during the last thirty years to assay protein carbonylation. The most successful method consists on protein carbonyl (PCO) derivatization with 2,4-dinitrophenylhydrazine (DNPH) and consequent spectrophotometric assay. This assay allows a global quantification of PCO content due to the ability of DNPH to react with carbonyl giving rise to an adduct able to absorb at 366 nm. Similar approaches were also developed employing chromatographic separation, in particular HPLC, and parallel detection of absorbing adducts. Subsequently, immunological techniques, such as Western immunoblot or ELISA, have been developed leading to an increase of sensitivity in protein carbonylation detection. Currently, they are widely employed to evaluate change in total protein carbonylation and eventually to highlight the specific proteins undergoing selective oxidation. In the last decade, many mass spectrometry (MS) approaches have been developed for the identification of the carbonylated proteins and the relative amino acid residues modified to carbonyl derivatives. Although these MS methods are much more focused and detailed due to their ability to identify the amino acid residues undergoing carbonylation, they still require too expensive equipments and, therefore, are limited in distribution. In this protocol paper, we summarise and comment on the most diffuse protocols that a standard laboratory can employ to assess protein carbonylation; in particular, we describe step-by-step the different protocols, adding suggestions coming from our on-bench experience.

  18. Analysis of protein carbonylation - pitfalls and promise in commonly used methods

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, A.; Wojdyla, K.; Nedic, O.

    2014-01-01

    . Several methods for measuring protein carbonylation have been implemented in different laboratories around the world. However, to date no methods prevail as the most accurate, reliable, and robust. The present paper aims at giving an overview of the common methods used to determine protein carbonylation...

  19. Protein carbonylation and heat shock proteins in human skeletal muscle: relationships to age and sarcopenia.

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    Beltran Valls, Maria R; Wilkinson, Daniel J; Narici, Marco V; Smith, Kenneth; Phillips, Bethan E; Caporossi, Daniela; Atherton, Philip J

    2015-02-01

    Aging is associated with a gradual loss of muscle mass termed sarcopenia, which has significant impact on quality-of-life. Because oxidative stress is proposed to negatively impact upon musculoskeletal aging, we investigated links between human aging and markers of oxidative stress, and relationships to muscle mass and strength in young and old nonsarcopenic and sarcopenic adults. Sixteen young and 16 old males (further subdivided into "old" and "old sarcopenic") were studied. The abundance of protein carbonyl adducts within skeletal muscle sarcoplasmic, myofibrillar, and mitochondrial protein subfractions from musculus vastus lateralis biopsies were determined using Oxyblot immunoblotting techniques. In addition, concentrations of recognized cytoprotective proteins (eg, heat shock proteins [HSP], αβ-crystallin) were also assayed. Aging was associated with increased mitochondrial (but not myofibrillar or sarcoplasmic) protein carbonyl adducts, independently of (stage-I) sarcopenia. Correlation analyses of all subjects revealed that mitochondrial protein carbonyl abundance negatively correlated with muscle strength ([1-repetition maximum], p = .02, r (2) = -.16), but not muscle mass (p = .13, r (2) = -.08). Abundance of cytoprotective proteins, including various HSPs (HSP 27 and 70), were unaffected by aging/sarcopenia. To conclude, these data reveal that mitochondrial protein carbonylation increases moderately with age, and that this increase may impact upon skeletal muscle function, but is not a hallmark of (stage-I) sarcopenia, per se. © The Author 2014. Published by Oxford University Press on behalf of The Gerontological Society of America.

  20. Paraquat exposure and Sod2 knockdown have dissimilar impacts on the Drosophila melanogaster carbonylated protein proteome.

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    Narayanasamy, Suresh K; Simpson, David C; Martin, Ian; Grotewiel, Mike; Gronert, Scott

    2014-11-01

    Exposure to Paraquat and RNA interference knockdown of mitochondrial superoxide dismutase (Sod2) are known to result in significant lifespan reduction, locomotor dysfunction, and mitochondrial degeneration in Drosophila melanogaster. Both perturbations increase the flux of the progenitor ROS, superoxide, but the molecular underpinnings of the resulting phenotypes are poorly understood. Improved understanding of such processes could lead to advances in the treatment of numerous age-related disorders. Superoxide toxicity can act through protein carbonylation. Analysis of carbonylated proteins is attractive since carbonyl groups are not present in the 20 canonical amino acids and are amenable to labeling and enrichment strategies. Here, carbonylated proteins were labeled with biotin hydrazide and enriched on streptavidin beads. On-bead digestion was used to release carbonylated protein peptides, with relative abundance ratios versus controls obtained using the iTRAQ MS-based proteomics approach. Western blotting and biotin quantitation assay approaches were also investigated. By both Western blotting and proteomics, Paraquat exposure, but not Sod2 knockdown, resulted in increased carbonylated protein relative abundance. For Paraquat exposure versus control, the median carbonylated protein relative abundance ratio (1.53) determined using MS-based proteomics was in good agreement with that obtained using a commercial biotin quantitation kit (1.36).

  1. Coffee polyphenols protect human plasma from postprandial carbonyl modifications.

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    Sirota, Roman; Gorelik, Shlomit; Harris, Raviv; Kohen, Ron; Kanner, Joseph

    2013-05-01

    The antioxidant capability of coffee polyphenols to inhibit red-meat lipid peroxidation in stomach medium and absorption into blood of malondialdehyde (MDA) in humans was studied. Roasted-ground coffee polyphenols that were found to inhibit lipid peroxidation in stomach medium are 2- to 5-fold more efficient antioxidant than those found in instant coffee. Human plasma from ten volunteers analyzed after a meal of red-meat cutlets (250 g) revealed a rapid accumulation of MDA. The accumulation of MDA in human plasma modified low-density lipoprotein is known to trigger atherogenesis. Consumption of 200 mL roasted coffee by ten volunteers during a meal of red-meat cutlets, resulted after 2 and 4 h in the inhibition by 80 and 50%, respectively, of postprandial plasma MDA absorption. The results obtained in vitro simulated stomach model on MDA accumulation were predictive for the amount of MDA absorbed into circulating human plasma, in vivo. Timing the consumption of coffee during the meals may make it a very active functional food.

  2. Novel DNPH-based method for determination of protein carbonylation in muscle and meat.

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    Soglia, Francesca; Petracci, Massimiliano; Ertbjerg, Per

    2016-04-15

    Protein oxidation is considered an ongoing deteriorative process during storage of fresh and processed meat. Carbonyl compounds have traditionally been detected spectrophotometrically after derivatization with 2,4-dinitrophenylhydrazine (DNPH) to form protein-bound hydrazones with absorbance at 370 nm. Here we describe a novel DNPH-based method to quantify protein carbonylation in muscle and meat. The additional steps of the novel method aimed at increasing the protein solubility and inducing protein unfolding before labeling with DNPH. Compared to the traditional method, the new procedure reflected an increased protein carbonylation level measuring overall two to fourfold more carbonyls in muscles from different species as well as in soluble, salt-soluble and insoluble protein fractions. The study suggested that protein unfolding is a more important phenomenon than solubilization for increased DNPH labeling. The novel method resulted in three to fourfold larger carbonyl content determined in chicken, pork and beef (2.8, 3.6 and 3.1 nmol/mg of protein, respectively). Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. A cleavable biotin tagging reagent that enables the enrichment and identification of carbonylation sites in proteins.

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    Coffey, Chelsea M; Gronert, Scott

    2016-01-01

    The utility of a new, cleavable tag for identifying and enriching protein carbonyls is examined. Using a model system, human serum albumin modified with acrolein, the EZ-Link alkoxyamine-PEG4-SS-PEG4-biotin affinity tag, was tested for its ability to label protein carbonyls in proteomic analyses of protein carbonylation. The efficiency of the labeling was assayed and compared to standard biotin hydrazide reagents. The label was also tested in liquid chromatography-tandem mass spectrometry (LC/MS/MS) experiments. The quality of the fragmentation spectra was assessed and the relative detection efficiency of various modification sites was compared to standard biotin hydrazide reagents. Finally, the viability of using the label with streptavidin bead enrichment protocols in a standard proteomics workflow was probed.

  4. Mitochondrial ATP synthase is a target for TNBS-induced protein carbonylation in XS-106 dendritic cells.

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    Je, Jeong Hwan; Lee, Tae Hyung; Kim, Dong Hyun; Cho, Young Hun; Lee, Ju Hee; Kim, Soo Chan; Lee, Sang-Kyou; Lee, Jaewon; Lee, Min-Geol

    2008-06-01

    ROS are produced in dendritic cells (DCs) during antigen presentation in contact hypersensitivity (CHS). As a result, ROS cause a number of nonenzymatic protein modifications, including carbonylation, which is the most widely used marker of oxidative stress. 2,4,6-Trinitrobenzene sulfonic acid (TNBS) is a well-characterized contact allergen that results in the formation of ROS. However, proteins that are carbonylated in DCs in response to TNBS have not been identified. To study ROS-dependent protein carbonylation in response to TNBS, we used the well-established mouse DC line, XS-106. We focused on the effects of TNBS on oxidation by examining selected oxidative markers. We identified TNBS-induced ROS and myeloperoxidase (MPO) proteins and demonstrated that the increase in ROS resulted in IL-12 production. The increase in oxidation was further confirmed by an oxidation-dependent increase in protein modifications, such as carbonylation. In fact, TNBS strongly induced carbonylation of mitochondrial adenosine triphosphate (ATP) synthase in XS-106 DCs, as determined by MALDI-TOF analysis and 2-D Western blotting. ROS production and protein carbonylation were confirmed in human monocyte-derived DCs (Mo-DCs). Furthermore, glutathione (GSH) decreased ROS and protein carbonylation in Mo-DCs. Carbonylation of ATP synthase in DCs may contribute to the pathophysiology of CHS.

  5. Impact of lipid content and composition on lipid oxidation and protein carbonylation in experimental fermented sausages.

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    Fuentes, Verónica; Estévez, Mario; Ventanas, Jesús; Ventanas, Sonia

    2014-03-15

    This study aims to investigate the effect of lipid content (∼4%, ∼10% and ∼15%) and composition (different lipid sources; animal fat and sunflower oil) on the oxidative stability of proteins and lipids in experimental fermented sausages. Increasing the lipid content of sausages enhanced the susceptibility of lipids to oxidation whereas the effect on the formation of specific carbonyls from protein oxidation was not so evident. Sausages manufactured with different lipid sources affected the susceptibility of lipids and proteins to oxidation as a likely result of the modifications in the fatty acid profile, as well as to the presence of antioxidant compounds. While the fatty acid profile had a major effect on the occurrence and extent of lipid oxidation, the presence of compounds with potential antioxidant activity may be more influential on the extent of protein carbonylation.

  6. Nitrite promotes protein carbonylation and Strecker aldehyde formation in experimental fermented sausages: are both events connected?

    Science.gov (United States)

    Villaverde, A; Ventanas, J; Estévez, M

    2014-12-01

    The role played by curing agents (nitrite, ascorbate) on protein oxidation and Strecker aldehyde formation is studied. To fulfill this objective, increasing concentrations of nitrite (0, 75 and 150ppm) and ascorbate (0, 250 and 500ppm) were added to sausages subjected to a 54day drying process. The concurrence of intense proteolysis, protein carbonylation and formation of Strecker aldehydes during processing of sausages suggests that α-aminoadipic semialdehyde (AAS) and γ-glutamic semialdehyde (GGS) may be implicated in the formation of Strecker aldehydes. The fact that nitrite (150ppm, ingoing amount) significantly promoted the formation of protein carbonyls at early stages of processing and the subsequent formation of Strecker aldehydes provides strength to this hypothesis. Ascorbate (125 and 250ppm) controlled the overall extent of protein carbonylation in sausages without declining the formation of Strecker aldehydes. These results may contribute to understanding the chemistry fundamentals of the positive influence of nitrite on the flavor and overall acceptability of cured muscle foods.

  7. Gel-free proteomic methodologies to study reversible cysteine oxidation and irreversible protein carbonyl formation.

    Science.gov (United States)

    Boronat, S; García-Santamarina, S; Hidalgo, E

    2015-05-01

    Oxidative modifications in proteins have been traditionally considered as hallmarks of damage by oxidative stress and aging. However, oxidants can generate a huge variety of reversible and irreversible modifications in amino acid side chains as well as in the protein backbones, and these post-translational modifications can contribute to the activation of signal transduction pathways, and also mediate the toxicity of oxidants. Among the reversible modifications, the most relevant ones are those arising from cysteine oxidation. Thus, formation of sulfenic acid or disulfide bonds is known to occur in many enzymes as part of their catalytic cycles, and it also participates in the activation of signaling cascades. Furthermore, these reversible modifications have been usually attributed with a protective role, since they may prevent the formation of irreversible damage by scavenging reactive oxygen species. Among irreversible modifications, protein carbonyl formation has been linked to damage and death, since it cannot be repaired and can lead to protein loss-of-function and to the formation of protein aggregates. This review is aimed at researchers interested on the biological consequences of oxidative stress, both at the level of signaling and toxicity. Here we are providing a concise overview on current mass-spectrometry-based methodologies to detect reversible cysteine oxidation and irreversible protein carbonyl formation in proteomes. We do not pretend to impose any of the different methodologies, but rather to provide an objective catwalk on published gel-free approaches to detect those two types of modifications, from a biologist's point of view.

  8. Detection of Protein Carbonyls by Means of Biotin Hydrazide-Streptavidin Affinity Methods.

    Science.gov (United States)

    Hensley, Kenneth

    2015-01-01

    Oxidative posttranslational protein modifications occur as a normal process of cell biology and to a greater extent during pathogenic conditions. The detection and quantitation of protein oxidation has posed a continuing challenge to bioanalytical chemists because of the following reasons: The products of oxidative protein damage are chemically diverse; protein oxidation generally occurs at low background levels; and the complexity of biological samples introduces high background noise when standard techniques such as immunolabeling are applied to "dirty" tissue extracts containing endogenous immunoglobulins or small molecular weight, chemically reactive compounds has been developed which circumvents these difficulties by incorporating a biotin label at sites of protein carbonylation. Biotin hydrazide-labeled proteins are detectable using standard streptavidin-coupled detection techniques such as peroxidase-catalyzed chemiluminescence of immunoblots. Advantages of the biotin hydrazide-labeling technique are its sensitivity and its lack of reliance upon antibodies that inevitably suffer from nonspecific background noise and contaminating endogenous immunoglobulins.

  9. Time and dose effects of cigarette smoke and acrolein on protein carbonyl formation in HaCaT keratinocytes.

    Science.gov (United States)

    Avezov, K; Reznick, A Z; Aizenbud, D

    2015-01-01

    Cigarette smoke (CS) is an important environmental source of human exposure to a highly toxic and chemically active α,β-unsaturated aldehyde: acrolein. It is capable of causing protein carbonylation and dysfunction, especially in oral tissues of smokers, constantly exposed to CS toxic constituents. The foremost damage is considered to be cumulative, but even a short exposure can be potentially harmful. The objectives of the current study were to examine the short time and dose effects of direct CS and acrolein exposure on intracellular protein carbonylation in epithelial cells. HaCaT-keratinocytes were exposed to different doses of acrolein and whole phase CS using a unique smoking simulator apparatus that mimics the exposure in smokers. The rate of intracellular protein carbonyl modification was examined 10-60 min after the exposure by Western blot. In addition, the effect of pre-incubation with a thiol scavenger N-acetylcysteine (NAC) was also assessed. We found that intracellular protein carbonyls increased as fast as 10 min after CS exposure and their concentration doubled after 20 min, with a slight elevation afterwards. Also, carbonyl levels increased gradually as CS and acrolein doses were elevated. Addition of 1 mM NAC neutralized part of the damage. We conclude that CS and acrolein intracellular protein carbonylation is dose- and time- dependent. Even a short time exposure to CS and its aldehydic constituents can be potentially harmful.

  10. Cross-talk between lipid and protein carbonylation in a dynamic cardiomyocyte model of mild nitroxidative stress

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    Eva Griesser

    2017-04-01

    Full Text Available Reactive oxygen and nitrogen species (ROS/RNS play an important role in the regulation of cardiac function. Increase in ROS/RNS concentration results in lipid and protein oxidation and is often associated with onset and/or progression of many cardiovascular disorders. However, interplay between lipid and protein modifications has not been simultaneously studied in detail so far. Biomolecule carbonylation is one of the most common biomarkers of oxidative stress. Using a dynamic model of nitroxidative stress we demonstrated rapid changes in biomolecule carbonylation in rat cardiomyocytes. Levels of carbonylated species increased as early as 15 min upon treatment with the peroxynitrite donor, 3-morpholinosydnonimine (SIN-1, and decreased to values close to control after 16 h. Total (lipids+proteins vs. protein-specific carbonylation showed different dynamics, with a significant increase in protein-bound carbonyls at later time points. Treatment with SIN-1 in combination with inhibitors of proteasomal and autophagy/lysosomal degradation pathways allowed confirmation of a significant role of the proteasome in the degradation of carbonylated proteins, whereas lipid carbonylation increased in the presence of autophagy/lysosomal inhibitors. Electrophilic aldehydes and ketones formed by lipid peroxidation were identified and relatively quantified using LC-MS/MS. Molecular identity of reactive species was used for data-driven analysis of their protein targets. Combination of different enrichment strategies with LC-MS/MS analysis allowed identification of more than 167 unique proteins with 332 sites modified by electrophilic lipid peroxidation products. Gene ontology analysis of modified proteins demonstrated enrichment of several functional categories including proteins involved in cytoskeleton, extracellular matrix, ion channels and their regulation. Using calcium mobilization assays, the effect of nitroxidative stress on the activity of several ion

  11. Comparative Proteomic Analysis of Carbonylated Proteins from the Striatum and Cortex of Pesticide-Treated Mice

    Science.gov (United States)

    Coughlan, Christina; Walker, Douglas I.; Lohr, Kelly M.; Richardson, Jason R.; Saba, Laura M.; Caudle, W. Michael; Fritz, Kristofer S.; Roede, James R.

    2015-01-01

    Epidemiological studies indicate exposures to the herbicide paraquat (PQ) and fungicide maneb (MB) are associated with increased risk of Parkinson's disease (PD). Oxidative stress appears to be a premier mechanism that underlies damage to the nigrostriatal dopamine system in PD and pesticide exposure. Enhanced oxidative stress leads to lipid peroxidation and production of reactive aldehydes; therefore, we conducted proteomic analyses to identify carbonylated proteins in the striatum and cortex of pesticide-treated mice in order to elucidate possible mechanisms of toxicity. Male C57BL/6J mice were treated biweekly for 6 weeks with saline, PQ (10 mg/kg), MB (30 mg/kg), or the combination of PQ and MB (PQMB). Treatments resulted in significant behavioral alterations in all treated mice and depleted striatal dopamine in PQMB mice. Distinct differences in 4-hydroxynonenal-modified proteins were observed in the striatum and cortex. Proteomic analyses identified carbonylated proteins and peptides from the cortex and striatum, and pathway analyses revealed significant enrichment in a variety of KEGG pathways. Further analysis showed enrichment in proteins of the actin cytoskeleton in treated samples, but not in saline controls. These data indicate that treatment-related effects on cytoskeletal proteins could alter proper synaptic function, thereby resulting in impaired neuronal function and even neurodegeneration. PMID:26345149

  12. Comparative Proteomic Analysis of Carbonylated Proteins from the Striatum and Cortex of Pesticide-Treated Mice

    Directory of Open Access Journals (Sweden)

    Christina Coughlan

    2015-01-01

    Full Text Available Epidemiological studies indicate exposures to the herbicide paraquat (PQ and fungicide maneb (MB are associated with increased risk of Parkinson’s disease (PD. Oxidative stress appears to be a premier mechanism that underlies damage to the nigrostriatal dopamine system in PD and pesticide exposure. Enhanced oxidative stress leads to lipid peroxidation and production of reactive aldehydes; therefore, we conducted proteomic analyses to identify carbonylated proteins in the striatum and cortex of pesticide-treated mice in order to elucidate possible mechanisms of toxicity. Male C57BL/6J mice were treated biweekly for 6 weeks with saline, PQ (10 mg/kg, MB (30 mg/kg, or the combination of PQ and MB (PQMB. Treatments resulted in significant behavioral alterations in all treated mice and depleted striatal dopamine in PQMB mice. Distinct differences in 4-hydroxynonenal-modified proteins were observed in the striatum and cortex. Proteomic analyses identified carbonylated proteins and peptides from the cortex and striatum, and pathway analyses revealed significant enrichment in a variety of KEGG pathways. Further analysis showed enrichment in proteins of the actin cytoskeleton in treated samples, but not in saline controls. These data indicate that treatment-related effects on cytoskeletal proteins could alter proper synaptic function, thereby resulting in impaired neuronal function and even neurodegeneration.

  13. Protein conjugated with aldehydes derived from lipid peroxidation as an independent parameter of the carbonyl stress in the kidney damage

    Directory of Open Access Journals (Sweden)

    Medina-Navarro Rafael

    2011-11-01

    Full Text Available Abstract Background One of the well-defined and characterized protein modifications usually produced by oxidation is carbonylation, an irreversible non-enzymatic modification of proteins. However, carbonyl groups can be introduced into proteins by non-oxidative mechanisms. Reactive carbonyl compounds have been observed to have increased in patients with renal failure. In the present work we have described a procedure designed as aldehyde capture to calculate the protein carbonyl stress derived solely from lipid peroxidation. Methods Acrolein-albumin adduct was prepared as standard at alkaline pH. Rat liver microsomal membranes and serum samples from patients with diabetic nephropathy were subjected to the aldehyde capture procedure and aldol-protein formation. Before alkalinization and incubation, samples were precipitated and redisolved in 6M guanidine. The absorbances of the samples were read with a spectrophotometer at 266 nm against a blank of guanidine. Results Evidence showed abundance of unsaturated aldehydes derived from lipid peroxidation in rat liver microsomal membranes and in the serum of diabetic patients with advanced chronic kidney disease. Carbonyl protein and aldol-proteins resulted higher in the diabetic nephropathy patients (p Conclusion The aldehyde-protein adduct represents a non oxidative component of carbonyl stress, independent of the direct amino acid oxidation and could constitute a practical and novelty strategy to measure the carbonyl stress derived solely from lipid peroxidation and particularly in diabetic nephropathy patients. In addition, we are in a position to propose an alternative explanation of why alkalinization of urine attenuates rhabdomyolysis-induced renal dysfunction.

  14. [Association between carbonyl proteins and tumor necrosis factor alpha with muscle strength in young and older women: exploratory study].

    Science.gov (United States)

    Martínez Huenchullán, Sergio Francisco; Mancilla Solorza, Eladio Bernabé

    2015-01-01

    It has recently been proposed that there is a close relationship between oxidative stress and low-grade chronic inflammation. Both processes have been related separately to muscle function in older adults (OA). Nevertheless, it still has not been determined if this relationship is present particularly in OA. The objective of this study was to determine the relationship between the plasma levels of TNF-α and carbonyl proteins (CP) and muscle strength in a group of young and older women. An exploratory study was conducted on 13 older and 8 young women, in whom the plasma levels of CP and TNF-α were measured. Muscle strength was measured by handgrip test, quadriceps voluntary maximal isometric strength, arm curl, and the 30 second sit to stand test. There were no differences in the plasma levels of CP and TNF-α between the groups, but there was relationship between the biomarkers only in the OA group. A non-linear relationship was observed between CP and quadriceps voluntary maximal isometric strength only in the OA group (R(2)=36.2; P=.038). For TNF-α there were no significant association with any of the applied tests. There is an association between CP and quadriceps voluntary maximal isometric strength only in the OA group, which could indicate a deleterious action of oxidative stress on muscle function, particularly in aging. Copyright © 2015 SEGG. Published by Elsevier Espana. All rights reserved.

  15. Protein Mediated Oxidative Stress in Patients with Diabetes and its Associated Neuropathy: Correlation with Protein Carbonylation and Disease Activity Markers

    Science.gov (United States)

    Almogbel, Ebtehal

    2017-01-01

    Introduction Free radicals have been implicated as Diabetes Mellitus (DM) contributors in type 2 DM and its associated Diabetes Mellitus Neuropathy (DMN). However, the potential for protein mediated oxidative stress to contribute disease pathogenesis remains largely unexplored. Aim To investigate the status and contribution of protein mediated oxidative stress in patients with DM or DMN and to explore whether oxidative protein modification has a role in DM progression to DM associated neuropathy. Materials and Methods Sera from 42 DM and 37 DMN patients with varying levels of disease activities biomarkers (HbA1C, patients’ age or disease duration) and 21 age- and sex-matched healthy controls were evaluated for serum levels of protein mediated oxidative stress. Results Serum analysis showed significantly higher levels of protein carbonyl contents in both DM and DMN patients compared with healthy controls. Importantly, not only was there an increased number of subjects positive for protein carbonylation, but also the levels of protein carbonyl contents were significantly higher among DM and DMN patients, whose HbA1C were ≥8.8 as compared with patients with lower HbA1C (HbA1Cdiabetes to diabetes neuropathy. Conclusion These findings support an association between protein oxidation and DM or DMN progression. The stronger response observed in patients with higher HbA1C or patients’ ages or disease durations suggests, that protein mediated oxidative stress may be useful in evaluating the progression of DM and its associated DMN and in elucidating the mechanisms of these disorders pathogenesis.

  16. Protein hydroperoxides and carbonyl groups generated by porphyrin-induced photo-oxidation of bovine serum albumin

    DEFF Research Database (Denmark)

    Silvester, J A; Timmins, G S; Davies, Michael Jonathan

    1998-01-01

    through type I processes (i.e., independent of singlet oxygen), while type II (singlet oxygen) mechanisms may play a significant role in protein carbonyl formation. Reaction of the protein hydroperoxide species with metal ion complexes is shown to produce further protein-derived radicals which...

  17. Engineered Trx2p industrial yeast strain protects glycolysis and fermentation proteins from oxidative carbonylation during biomass propagation

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    Gómez-Pastor Rocío

    2012-01-01

    Full Text Available Abstract Background In the yeast biomass production process, protein carbonylation has severe adverse effects since it diminishes biomass yield and profitability of industrial production plants. However, this significant detriment of yeast performance can be alleviated by increasing thioredoxins levels. Thioredoxins are important antioxidant defenses implicated in many functions in cells, and their primordial functions include scavenging of reactive oxygen species that produce dramatic and irreversible alterations such as protein carbonylation. Results In this work we have found several proteins specifically protected by yeast Thioredoxin 2 (Trx2p. Bidimensional electrophoresis and carbonylated protein identification from TRX-deficient and TRX-overexpressing cells revealed that glycolysis and fermentation-related proteins are specific targets of Trx2p protection. Indeed, the TRX2 overexpressing strain presented increased activity of the central carbon metabolism enzymes. Interestingly, Trx2p specifically preserved alcohol dehydrogenase I (Adh1p from carbonylation, decreased oligomer aggregates and increased its enzymatic activity. Conclusions The identified proteins suggest that the fermentative capacity detriment observed under industrial conditions in T73 wine commercial strain results from the oxidative carbonylation of specific glycolytic and fermentation enzymes. Indeed, increased thioredoxin levels enhance the performance of key fermentation enzymes such as Adh1p, which consequently increases fermentative capacity.

  18. Amelioration of age-dependent increase in protein carbonyls of cerebral hemispheres of mice by melatonin and ascorbic acid.

    Science.gov (United States)

    Dkhar, Preeticia; Sharma, Ramesh

    2011-12-01

    Melatonin secreted by the pineal gland acts as a free radical scavenger besides its role as a hormonal signaling agent. It detoxifies a variety of free radicals and reactive oxygen intermediates including hydroxyl radical, peroxynitrite anion and singlet oxygen. Ascorbic acid (Vitamin C), a water soluble vitamin, is a naturally occurring antioxidant and cofactor in various enzymes. Protein carbonyls are formed as a consequence of the oxidative modification of proteins by reactive oxygen species. Oxidative modification alters the function of protein and is thought to play an important role in the decline of cellular functions during aging. In the present study, the effect of melatonin and ascorbic acid on age-related carbonyl content of cerebral hemispheres in mice was investigated. Protein carbonyls of cerebral hemispheres have been found to be significantly higher in 18-month-old mice as compared to 1-month old mice. Administration of a single dose of melatonin (10 mg/kg body weight) and ascorbic acid (10 mg/kg body weight) intraperitoneally for three consecutive days decreases the carbonyl content in 1- and 18-month-old mice significantly. The present study thus suggests that the formation of protein carbonyls in the cerebral hemispheres of the aging mice can be prevented by the antioxidative effects of melatonin and ascorbic acid that could in turn be beneficial in having health benefits from age-related neurodegenerative diseases.

  19. The serum protein carbonyl content level in relation to exercise stress test

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    Titiporn Mekrungruangwong

    2012-01-01

    Full Text Available Background: Protein carbonyl (P is oxidatively-modified protein with diagnostic potential for acute myocardial infarction. However, many findings indicated the elevation of serum PC content level related to exercise, which could cause false positive results and limiting the specificity for acute coronary syndrome diagnosis. This study aims to evaluate the level of serum protein carbonyl content in healthy volunteers subjected to exercise stress test (EST. Materials and Methods: Serum from healthy volunteers was collected 5-10 min before performing EST and 1 hour after the EST was achieved. The serum was collected, and the serum PC content level was determined by spectrophotometric DNPH assay. Results: The serum PC content level after exercise stress test was significantly higher than that of before performing EST (0.373 ± 0.05 nM/mg vs. 0.275 ± 0.02 nM/mg, P < 0.0001. The results demonstrated that in both male and female, serum PC content level after EST was significantly higher than that of before performing EST (0.29 ± 0.03 nM/mg vs. 0.36 ± 0.05 nM/mg P < 0.0001 in male, 0.27 ± 0.02 nM/mg vs. 0.38 ± 0.06 nM/mg P < 0.0001 in female, respectively. Conclusions: This study demonstrated that exercise stress test could result in non-specificity and false positive increasing in serum PC content level in healthy subjects, which may cause misinterpretation when using PC as cardiac marker, especially in patients, who underwent exercise stress test or patients who performing heavy physical activities.

  20. Mitochondrial ascorbate-glutathione cycle and proteomic analysis of carbonylated proteins during tomato (Solanum lycopersicum) fruit ripening.

    Science.gov (United States)

    López-Vidal, O; Camejo, D; Rivera-Cabrera, F; Konigsberg, M; Villa-Hernández, J M; Mendoza-Espinoza, J A; Pérez-Flores, L J; Sevilla, F; Jiménez, A; Díaz de León-Sánchez, F

    2016-03-01

    In non-photosynthetic tissues, mitochondria are the main source of energy and of reactive oxygen species. Accumulation of high levels of these species in the cell causes damage to macromolecules including several proteins and induces changes in different metabolic processes. Fruit ripening has been characterized as an oxidative phenomenon; therefore, control of reactive oxygen species levels by mitochondrial antioxidants plays a crucial role on this process. In this work, ascorbate-glutathione cycle components, hydrogen peroxide levels and the proteomic profile of carbonylated proteins were analyzed in mitochondria isolated from tomato (Solanum lycopersicum) fruit at two ripening stages. A significant increase on most ascorbate-glutathione cycle components and on carbonylated proteins was observed in mitochondria from breaker to light red stage. Enzymes and proteins involved in diverse cellular and mitochondrial metabolic pathways were identified among the carbonylated proteins. These results suggest that protein carbonylation is a post-translational modification involved in tomato fruit ripening regulation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. The Behaviour of Protein Carbonyls in Newborns with Birth Respiratory Distress and Asphyxia

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    Gabriela ZAHARIE

    2009-12-01

    Full Text Available Objective: A prospective study was carried out in premature newborns with respiratory distress syndrome (RDS and asphyxia at birth in order to identify and analyze the effects of RDS on proteins. Material and Methods: Protein peroxidation was studied using the Reznick spectrophotometric method. The study group included 14 premature newborns with respiratory distress and asphyxia at birth. The control group included 13 newborns that were born on term, eutrophic and healthy. The determinations were carried out using venous blood. Statistical data analyses were performed using Statistica software. The comparisons between groups (study and controls were performed by applying parametric and non-parametric tests according with the type of distribution. Results: Statistically significant correlations were found between the value of protein carbonyls (PC and the weight of premature newborns in the case group (p < 0.05, as well as between the PC value and the presence of respiratory distress due to surfactant deficiency in the study group. The average PC value in the study group was higher in the third day as compared with the first day. The PC value was significantly higher in the control group as compared with study group. Conclusion: The results of our study revealed that the respiratory distress in the premature newborn and oxygen therapy stimulate the peroxidation of proteins.

  2. Protein carbonylation and water-holding capacity of pork subjected to frozen storage: effect of muscle type, premincing, and packaging.

    Science.gov (United States)

    Estévez, Mario; Ventanas, Sonia; Heinonen, Marina; Puolanne, Eero

    2011-05-25

    The present work aimed to study the relationship between protein carbonylation and the loss of water-holding capacity (WHC) occurring during frozen storage of porcine muscles. Pork samples corresponding to two different muscle types, glycolytic M. longissimus dorsi (LD) and oxidative M. psoas major (PM), and subjected to two different premincing, minced (MINCED) and intact pork pieces (INTACT), and packaging, vacuum packaged (VACUUM) and packed in oxygen-permeable bags (OXYGEN), procedures were frozen (-18 °C/12 weeks) and analyzed at sampling times upon thawing (weeks 1, 4, 8, and 12) for the relative amount of specific protein carbonyls, α-aminoadipic and γ-glutamic semialdehydes (AAS and GGS, respectively), and their ability to hold water using two different techniques. The formation of protein carbonyls occurred concomitantly with the loss of WHC, and both phenomena were found to be more intense in LD muscles and in MINCED and OXYGEN pork samples. The loss in WHC was from ca. 10 to 30% in 12 weeks, depending on the method of determination. Plausible mechanisms by which protein carbonylation may decrease the WHC of pork samples are thoroughly discussed in the present paper. Besides the likely impact of protein carbonylation in the water-myofibrillar protein relationships, the implication of AAS and GGS in further reactions including plausible cross-linking would explain the decrease of these semialdehydes by the end of frozen storage and would reinforce their liability in the loss of WHC of porcine muscles. The exact nature of these reactions, however, should be investigated in further studies.

  3. Photoaffinity Labeling of Plasma Proteins

    Directory of Open Access Journals (Sweden)

    Masaki Otagiri

    2013-11-01

    Full Text Available Photoaffinity labeling is a powerful technique for identifying a target protein. A high degree of labeling specificity can be achieved with this method in comparison to chemical labeling. Human serum albumin (HSA and α1-acid glycoprotein (AGP are two plasma proteins that bind a variety of endogenous and exogenous substances. The ligand binding mechanism of these two proteins is complex. Fatty acids, which are known to be transported in plasma by HSA, cause conformational changes and participate in allosteric ligand binding to HSA. HSA undergoes an N-B transition, a conformational change at alkaline pH, that has been reported to result in increased ligand binding. Attempts have been made to investigate the impact of fatty acids and the N-B transition on ligand binding in HSA using ketoprofen and flunitrazepam as photolabeling agents. Meanwhile, plasma AGP is a mixture of genetic variants of the protein. The photolabeling of AGP with flunitrazepam has been utilized to shed light on the topology of the protein ligand binding site. Furthermore, a review of photoaffinity labeling performed on other major plasma proteins will also be discussed. Using a photoreactive natural ligand as a photolabeling agent to identify target protein in the plasma would reduce non-specific labeling.

  4. Plasma protein oxidation and total antioxidant power in premenstrual syndrome

    Institute of Scientific and Technical Information of China (English)

    Eans Tara Tuladhar; Anjali Rao

    2010-01-01

    Objective:To explore whether oxidative stress has any role inpremenstrual syndrome (PMS). Methods: Female volunteers suffering from PMS , in the age group of 20-24 years were compared to their asymptomatic normomennorhoeic counterparts in follicular phase and late luteal phase for ferric reducing antioxidant power of plasma(FRAP), plasma protein thiols(PPT) and protein carbonyls(PPC) levels.Results:There was no significant change in FRAP and PPC levels in controls andPMS groups but PPT decreased significantly in luteal phase ofPMS (P< 0.05) when compared to follicular phase.Conclusions:Estrogen and progesterone, might be responsible for a healthy antioxidant profile inPMS. However, a marked decrease inPPT in luteal phase of PMS group may be due to pro-oxidant nature of estrogen-active in this phase of PMS leading to consumption of the sacrificial antioxidant-protein thiol.

  5. Direct 13C-detection for carbonyl relaxation studies of protein dynamics

    Science.gov (United States)

    Pasat, Gabriela; Zintsmaster, John S.; Peng, Jeffrey W.

    2008-08-01

    We describe a method that uses direct 13C-detection for measuring rotating-frame carbonyl ( 13CO) relaxation rates to describe protein functional dynamics. Key advantages of method include the following: (i) unique access to 13CO groups that lack a scalar-coupled 15N- 1H group; (ii) insensitivity to 15N/ 1H exchange-broadening that can derail 1H-detected 15N and HNCO methods; (iii) avoidance of artifacts caused by incomplete water suppression. We demonstrate the approach for both backbone and side-chain 13CO groups. Accuracy of the 13C-detected results is supported by their agreement with those obtained from established HNCO-based approaches. Critically, we show that the 13C-detection approach provides access to the 13CO groups of functionally important residues that are invisible via 1H-detected HNCO methods because of exchange-broadening. Hence, the 13C-based method fills gaps inherent in canonical 1H-detected relaxation experiments, and thus provides a novel complementary tool for NMR studies of biomolecular flexibility.

  6. Novel effects of FCCP [carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone] on amyloid precursor protein processing.

    Science.gov (United States)

    Connop, B P; Thies, R L; Beyreuther, K; Ida, N; Reiner, P B

    1999-04-01

    Amyloidogenic processing of the beta-amyloid precursor protein (APP) has been implicated in the pathology of Alzheimer's disease. Because it has been suggested that catabolic processing of the APP holoprotein occurs in acidic intracellular compartments, we studied the effects of the protonophore carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP) and the H+-ATPase inhibitor bafilomycin A1 on APP catabolism in human embryonic kidney 293 cells expressing either wild-type or "Swedish" mutant APP. Unlike bafilomycin A1, which inhibits beta-amyloid production in cells expressing mutant but not wild-type APP, FCCP inhibited beta-amyloid production in both cell types. Moreover, the effects of FCCP were independent of alterations in total cellular APP levels or APP maturation, and the concentrations used did not alter either cellular ATP levels or cell viability. Bafilomycin A1, which had no effect on beta-amyloid production in wild-type cells, inhibited endocytosis of fluorescent transferrin, whereas concentrations of FCCP that inhibited beta-amyloid production in these cells had no effect on endosomal function. Thus, in wild-type-expressing cells it appears that the beta-amyloid peptide is not produced in the classically defined endosome. Although bafilomycin A1 decreased beta-amyloid release from cells expressing mutant APP but not wild-type APP, it altered lysosomal function in both cell types, suggesting that in normal cells beta-amyloid is not produced in the lysosome. Although inhibition of beta-amyloid production by bafilomycin A1 in mutant cells may occur via changes in endosomal/lysosomal pH, our data suggest that FCCP inhibits wild-type beta-amyloid production by acting on a bafilomycin A1-insensitive acidic compartment that is distinct from either the endosome or the lysosome.

  7. Redox Proteomic Profiling of Specifically Carbonylated Proteins in the Serum of Triple Transgenic Alzheimer’s Disease Mice

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    Liming Shen

    2016-04-01

    Full Text Available Oxidative stress is a key event in the onset and progression of neurodegenerative diseases, including Alzheimer’s disease (AD. To investigate the role of oxidative stress in AD and to search for potential biomarkers in peripheral blood, serums were collected in this study from the 3-, 6-, and 12-month-old triple transgenic AD mice (3×Tg-AD mice and the age- and sex-matched non-transgenic (non-Tg littermates. The serum oxidized proteins were quantified by slot-blot analysis and enzyme-linked immunosorbent assay (ELISA to investigate the total levels of serum protein carbonyl groups. Western blotting, in conjunction with two-dimensional gel electrophoresis (2D-Oxyblot, was employed to identify and quantify the specifically-carbonylated proteins in the serum of 3×Tg-AD mice. The results showed that the levels of serum protein carbonyls were increased in the three month old 3×Tg-AD mice compared with the non-Tg control mice, whereas no significant differences were observed in the six and 12 months old AD mice, suggesting that oxidative stress is an early event in AD progression. With the application of 2D-Oxyblot analysis, (immunoglobin Ig gamma-2B chain C region (IGH-3, Ig lambda-2 chain C region (IGLC2, Ig kappa chain C region (IGKC, and Ig kappa chain V-V region HP R16.7 were identified as significantly oxidized proteins compared with the control. Among them IGH-3 and IGKC were validated via immunoprecipitation and Western blot analysis. Identification of oxidized proteins in the serums of 3×Tg-AD mice can not only reveal potential roles of those proteins in the pathogenesis of AD but also provide potential biomarkers of AD at the early stage.

  8. Quantifying millisecond time-scale exchange in proteins by CPMG relaxation dispersion NMR spectroscopy of side-chain carbonyl groups

    Energy Technology Data Exchange (ETDEWEB)

    Hansen, Alexandar L.; Kay, Lewis E., E-mail: kay@pound.med.utoronto.ca [University of Toronto, Departments of Molecular Genetics, Biochemistry and Chemistry (Canada)

    2011-08-15

    A new pulse sequence is presented for the measurement of relaxation dispersion profiles quantifying millisecond time-scale exchange dynamics of side-chain carbonyl groups in uniformly {sup 13}C labeled proteins. The methodology has been tested using the 87-residue colicin E7 immunity protein, Im7, which is known to fold via a partially structured low populated intermediate that interconverts with the folded, ground state on the millisecond time-scale. Comparison of exchange parameters extracted for this folding 'reaction' using the present methodology with those obtained from more 'traditional' {sup 15}N and backbone carbonyl probes establishes the utility of the approach. The extracted excited state side-chain carbonyl chemical shifts indicate that the Asx/Glx side-chains are predominantly unstructured in the Im7 folding intermediate. However, several crucial salt-bridges that exist in the native structure appear to be already formed in the excited state, either in part or in full. This information, in concert with that obtained from existing backbone and side-chain methyl relaxation dispersion experiments, will ultimately facilitate a detailed description of the structure of the Im7 folding intermediate.

  9. Nitric oxide enhances desiccation tolerance of recalcitrant Antiaris toxicaria seeds via protein S-nitrosylation and carbonylation.

    Directory of Open Access Journals (Sweden)

    Xuegui Bai

    Full Text Available The viability of recalcitrant seeds is lost following stress from either drying or freezing. Reactive oxygen species (ROS resulting from uncontrolled metabolic activity are likely responsible for seed sensitivity to drying. Nitric oxide (NO and the ascorbate-glutathione cycle can be used for the detoxification of ROS, but their roles in the seed response to desiccation remain poorly understood. Here, we report that desiccation induces rapid accumulation of H(2O(2, which blocks recalcitrant Antiaris toxicaria seed germination; however, pretreatment with NO increases the activity of antioxidant ascorbate-glutathione pathway enzymes and metabolites, diminishes H(2O(2 production and assuages the inhibitory effects of desiccation on seed germination. Desiccation increases the protein carbonylation levels and reduces protein S-nitrosylation of these antioxidant enzymes; these effects can be reversed with NO treatment. Antioxidant protein S-nitrosylation levels can be further increased by the application of S-nitrosoglutathione reductase inhibitors, which further enhances NO-induced seed germination rates after desiccation and reduces desiccation-induced H(2O(2 accumulation. These findings suggest that NO reinforces recalcitrant seed desiccation tolerance by regulating antioxidant enzyme activities to stabilize H(2O(2 accumulation at an appropriate concentration. During this process, protein carbonylation and S-nitrosylation patterns are used as a specific molecular switch to control antioxidant enzyme activities.

  10. Nitric oxide enhances desiccation tolerance of recalcitrant Antiaris toxicaria seeds via protein S-nitrosylation and carbonylation.

    Science.gov (United States)

    Bai, Xuegui; Yang, Liming; Tian, Meihua; Chen, Jinhui; Shi, Jisen; Yang, Yongping; Hu, Xiangyang

    2011-01-01

    The viability of recalcitrant seeds is lost following stress from either drying or freezing. Reactive oxygen species (ROS) resulting from uncontrolled metabolic activity are likely responsible for seed sensitivity to drying. Nitric oxide (NO) and the ascorbate-glutathione cycle can be used for the detoxification of ROS, but their roles in the seed response to desiccation remain poorly understood. Here, we report that desiccation induces rapid accumulation of H(2)O(2), which blocks recalcitrant Antiaris toxicaria seed germination; however, pretreatment with NO increases the activity of antioxidant ascorbate-glutathione pathway enzymes and metabolites, diminishes H(2)O(2) production and assuages the inhibitory effects of desiccation on seed germination. Desiccation increases the protein carbonylation levels and reduces protein S-nitrosylation of these antioxidant enzymes; these effects can be reversed with NO treatment. Antioxidant protein S-nitrosylation levels can be further increased by the application of S-nitrosoglutathione reductase inhibitors, which further enhances NO-induced seed germination rates after desiccation and reduces desiccation-induced H(2)O(2) accumulation. These findings suggest that NO reinforces recalcitrant seed desiccation tolerance by regulating antioxidant enzyme activities to stabilize H(2)O(2) accumulation at an appropriate concentration. During this process, protein carbonylation and S-nitrosylation patterns are used as a specific molecular switch to control antioxidant enzyme activities.

  11. iCar-PseCp: identify carbonylation sites in proteins by Monte Carlo sampling and incorporating sequence coupled effects into general PseAAC

    Science.gov (United States)

    Jia, Jianhua; Liu, Zi; Xiao, Xuan; Liu, Bingxiang; Chou, Kuo-Chen

    2016-01-01

    Carbonylation is a posttranslational modification (PTM or PTLM), where a carbonyl group is added to lysine (K), proline (P), arginine (R), and threonine (T) residue of a protein molecule. Carbonylation plays an important role in orchestrating various biological processes but it is also associated with many diseases such as diabetes, chronic lung disease, Parkinson's disease, Alzheimer's disease, chronic renal failure, and sepsis. Therefore, from the angles of both basic research and drug development, we are facing a challenging problem: for an uncharacterized protein sequence containing many residues of K, P, R, or T, which ones can be carbonylated, and which ones cannot? To address this problem, we have developed a predictor called iCar-PseCp by incorporating the sequence-coupled information into the general pseudo amino acid composition, and balancing out skewed training dataset by Monte Carlo sampling to expand positive subset. Rigorous target cross-validations on a same set of carbonylation-known proteins indicated that the new predictor remarkably outperformed its existing counterparts. For the convenience of most experimental scientists, a user-friendly web-server for iCar-PseCp has been established at http://www.jci-bioinfo.cn/iCar-PseCp, by which users can easily obtain their desired results without the need to go through the complicated mathematical equations involved. It has not escaped our notice that the formulation and approach presented here can also be used to analyze many other problems in computational proteomics. PMID:27153555

  12. A NMR experiment for simultaneous correlations of valine and leucine/isoleucine methyls with carbonyl chemical shifts in proteins.

    Science.gov (United States)

    Tugarinov, Vitali; Venditti, Vincenzo; Marius Clore, G

    2014-01-01

    A methyl-detected 'out-and-back' NMR experiment for obtaining simultaneous correlations of methyl resonances of valine and isoleucine/leucine residues with backbone carbonyl chemical shifts, SIM-HMCM(CGCBCA)CO, is described. The developed pulse-scheme serves the purpose of convenience in recording a single data set for all Ile(δ1), Leu(δ) and Val(γ) (ILV) methyl positions instead of acquiring two separate spectra selective for valine or leucine/isoleucine residues. The SIM-HMCM(CGCBCA)CO experiment can be used for ILV methyl assignments in moderately sized protein systems (up to ~100 kDa) where the backbone chemical shifts of (13)C(α), (13)Cβ and (13)CO are known from prior NMR studies and where some losses in sensitivity can be tolerated for the sake of an overall reduction in NMR acquisition time.

  13. Dormancy removal of apple seeds by cold stratification is associated with fluctuation in H2O2, NO production and protein carbonylation level.

    Science.gov (United States)

    Dębska, Karolina; Krasuska, Urszula; Budnicka, Katarzyna; Bogatek, Renata; Gniazdowska, Agnieszka

    2013-03-15

    Reactive oxygen (ROS) and nitrogen (RNS) species play a signaling role in seed dormancy alleviation and germination. Their action may be described by the oxidative/nitrosative "window/door". ROS accumulation in embryos could lead to oxidative modification of protein through carbonylation. Mature apple (Malus domestica Borkh.) seeds are dormant and do not germinate. Their dormancy may be overcome by 70-90 days long cold stratification. The aim of this work was to analyze the relationship between germinability of embryos isolated from cold (5°C) or warm (25°C) stratified apple seeds and ROS or nitric oxide (NO) production and accumulation of protein carbonyl groups. A biphasic pattern of variation in H2O2 concentration in the embryos during cold stratification was detected. H2O2 content increased markedly after 7 days of seeds imbibition at 5°C. After an additional two months of cold stratification, the H2O2 concentration in embryos reached the maximum. NO production by the embryos was low during entire period of stratification, but increased significantly in germination sensu stricto (i.e. phase II of the germination process). The highest content of protein carbonyl groups was detected after 6 weeks of cold stratification treatment. Fluctuation of H2O2 and protein carbonylation seems to play a pivotal role in seed dormancy alleviation by cold stratification, while NO appears to be necessary for seed germination.

  14. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins

    DEFF Research Database (Denmark)

    Weber, Daniela; Davies, Michael J.; Grune, Tilman

    2015-01-01

    different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used...

  15. Aldo-keto reductase family 1 B10 protein detoxifies dietary and lipid-derived alpha, beta-unsaturated carbonyls at physiological levels

    Energy Technology Data Exchange (ETDEWEB)

    Zhong, Linlin [Department of Medical Microbiology, Immunology, and Cell Biology, SimmonsCooper Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL 62794-9626 (United States); Department of Neurobiology and Anatomy, China Medical University, Shenyang 110001 (China); Liu, Ziwen [Department of Medical Microbiology, Immunology, and Cell Biology, SimmonsCooper Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL 62794-9626 (United States); Department of Surgery, Peking Union Medical College Hospital, Beijing 100730 (China); Yan, Ruilan [Department of Medical Microbiology, Immunology, and Cell Biology, SimmonsCooper Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL 62794-9626 (United States); Johnson, Stephen [Carbon Dynamics Institute, LLC, 2835 via Verde Drive, Springfield, IL 62703-4325 (United States); Zhao, Yupei [Department of Surgery, Peking Union Medical College Hospital, Beijing 100730 (China); Fang, Xiubin [Department of Neurobiology and Anatomy, China Medical University, Shenyang 110001 (China); Cao, Deliang, E-mail: dcao@siumed.edu [Department of Medical Microbiology, Immunology, and Cell Biology, SimmonsCooper Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL 62794-9626 (United States)

    2009-09-18

    Alpha, beta-unsaturated carbonyls are highly reactive mutagens and carcinogens to which humans are exposed on a daily basis. This study demonstrates that aldo-keto reductase family 1 member B10 (AKR1B10) is a critical protein in detoxifying dietary and lipid-derived unsaturated carbonyls. Purified AKR1B10 recombinant protein efficiently catalyzed the reduction to less toxic alcohol forms of crotonaldehyde at 0.90 {mu}M, 4-hydroxynonenal (HNE) at 0.10 {mu}M, trans-2-hexanal at 0.10 {mu}M, and trans-2,4-hexadienal at 0.05 {mu}M, the concentrations at or lower than physiological exposures. Ectopically expressed AKR1B10 in 293T cells eliminated immediately HNE at 1 (subtoxic) or 5 {mu}M (toxic) by converting to 1,4-dihydroxynonene, protecting the cells from HNE toxicity. AKR1B10 protein also showed strong enzymatic activity toward glutathione-conjugated carbonyls. Taken together, our study results suggest that AKR1B10 specifically expressed in the intestine is physiologically important in protecting the host cell against dietary and lipid-derived cytotoxic carbonyls.

  16. Low intensity training of mdx mice reduces carbonylation and increases expression levels of proteins involved in energy metabolism and muscle contraction.

    Science.gov (United States)

    Hyzewicz, Janek; Tanihata, Jun; Kuraoka, Mutsuki; Ito, Naoki; Miyagoe-Suzuki, Yuko; Takeda, Shin'ichi

    2015-05-01

    High intensity training induces muscle damage in dystrophin-deficient mdx mice, an animal model for Duchenne muscular dystrophy. However, low intensity training (LIT) rescues the mdx phenotype and even reduces the level of protein carbonylation, a marker of oxidative damage. Until now, beneficial effects of LIT were mainly assessed at the physiological level. We investigated the effects of LIT at the molecular level on 8-week-old wild-type and mdx muscle using 2D Western blot and protein-protein interaction analysis. We found that the fast isoforms of troponin T and myosin binding protein C as well as glycogen phosphorylase were overcarbonylated and downregulated in mdx muscle. Some of the mitochondrial enzymes of the citric acid cycle were overcarbonylated, whereas some proteins of the respiratory chain were downregulated. Of functional importance, ATP synthase was only partially assembled, as revealed by Blue Native PAGE analysis. LIT decreased the carbonylation level and increased the expression of fast isoforms of troponin T and of myosin binding protein C, and glycogen phosphorylase. In addition, it increased the expression of aconitate hydratase and NADH dehydrogenase, and fully restored the ATP synthase complex. Our study demonstrates that the benefits of LIT are associated with lowered oxidative damage as revealed by carbonylation and higher expression of proteins involved in energy metabolism and muscle contraction. Potentially, these results will help to design therapies for DMD based on exercise mimicking drugs.

  17. Protein carbonyl: An oxidative stress marker in gingival crevicular fluid in healthy, gingivitis, and chronic periodontitis subjects

    Directory of Open Access Journals (Sweden)

    Avani R Pradeep

    2013-01-01

    Full Text Available Background: A defined role for reactive oxygen species (ROS in the tissue destruction that characterizes periodontitis has been described. Protein carbonyl (PC is the most widely used biomarker for oxidative damage to proteins, and reflects cellular damage induced by multiple forms of ROS. The purpose of this study is to determine the presence of PC in gingival crevicular fluid (GCF in healthy, gingivitis, and chronic periodontitis (CP subjects and to find an association, if any. Materials and Methods: A total number of 75 subjects (38 males and 37 females were selected based on their clinical parameters into three groups: Group 1 (25 healthy subjects, Group 2 (25 gingivitis subjects, and Group 3 (25 CP subjects. GCF samples were collected to estimate the levels of PC. Results: The PC concentration in GCF was highest in subjects with CP as compared to gingivitis and healthy subjects and a significant association was observed between GCF PC levels and all periodontal parameters. Conclusion: There was an increase in PC levels in GCF as the disease process progressed from healthy to gingivitis and CP, suggesting a role for increased oxidative stress in CP.

  18. Analysis of protein carbonylation-pitfalls and promise in commonly used methods

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, Adelina; Wojdyla, K; Nedić, O

    2014-01-01

    Oxidation of proteins has received a lot of attention in the last decades due to the fact that they have been shown to accumulate and to be implicated in the progression and the pathophysiology of several diseases such as Alzheimer, coronary heart diseases, etc. This has also resulted in the fact...... that research scientists are becoming more eager to be able to measure accurately the level of oxidized protein in biological materials, and to determine the precise site of the oxidative attack on the protein, in order to get insights into the molecular mechanisms involved in the progression of diseases...

  19. Redox regulation of protein damage in plasma.

    Science.gov (United States)

    Griffiths, Helen R; Dias, Irundika H K; Willetts, Rachel S; Devitt, Andrew

    2014-01-01

    The presence and concentrations of modified proteins circulating in plasma depend on rates of protein synthesis, modification and clearance. In early studies, the proteins most frequently analysed for damage were those which were more abundant in plasma (e.g. albumin and immunoglobulins) which exist at up to 10 orders of magnitude higher concentrations than other plasma proteins e.g. cytokines. However, advances in analytical techniques using mass spectrometry and immuno-affinity purification methods, have facilitated analysis of less abundant, modified proteins and the nature of modifications at specific sites is now being characterised. The damaging reactive species that cause protein modifications in plasma principally arise from reactive oxygen species (ROS) produced by NADPH oxidases (NOX), nitric oxide synthases (NOS) and oxygenase activities; reactive nitrogen species (RNS) from myeloperoxidase (MPO) and NOS activities; and hypochlorous acid from MPO. Secondary damage to proteins may be caused by oxidized lipids and glucose autooxidation. In this review, we focus on redox regulatory control of those enzymes and processes which control protein maturation during synthesis, produce reactive species, repair and remove damaged plasma proteins. We have highlighted the potential for alterations in the extracellular redox compartment to regulate intracellular redox state and, conversely, for intracellular oxidative stress to alter the cellular secretome and composition of extracellular vesicles. Through secreted, redox-active regulatory molecules, changes in redox state may be transmitted to distant sites.

  20. Herbal adaptogens combined with protein fractions from bovine colostrum and hen egg yolk reduce liver TNF-α expression and protein carbonylation in Western diet feeding in rats.

    Science.gov (United States)

    Mobley, C Brooks; Toedebusch, Ryan G; Lockwood, Christopher M; Heese, Alexander J; Zhu, Conan; Krieger, Anna E; Cruthirds, Clayton L; Hofheins, John C; Company, Joseph M; Wiedmeyer, Charles E; Kim, Dae Y; Booth, Frank W; Roberts, Michael D

    2014-01-01

    We examined if a purported anti-inflammatory supplement (AF) abrogated Western-diet (WD)-induced liver pathology in rats. AF contained: 1) protein concentrates from bovine colostrum and avian egg yolk; 2) herbal adaptogens and antioxidants; and 3) acetyl-L-carnitine. Nine month-old male Brown Norway rats were allowed ad libitum access to WD for 41-43 days and randomly assigned to WD + AF feeding twice daily for the last 31-33 days (n = 8), or WD and water-placebo feeding twice daily for the last 31-33 days (n = 8). Rats fed a low-fat/low-sucrose diet (CTL, n = 6) for 41-43 days and administered a water-placebo twice daily for the last 31-33 days were also studied. Twenty-four hours following the last gavage-feed, liver samples were analyzed for: a) select mRNAs (via RT-PCR) as well as genome-wide mRNA expression patterns (via RNA-seq); b) lipid deposition; and, c) protein carbonyl and total antioxidant capacity (TAC). Serum was also examined for TAC, 8-isoprostane and clinical chemistry markers. WD + AF rats experienced a reduction in liver Tnf-α mRNA (-2.8-fold, p < 0.01). Serum and liver TAC was lower in WD + AF versus WD and CTL rats (p < 0.05), likely due to exogenous antioxidant ingredients provided through AF as evidenced by a tendency for mitochondrial SOD2 mRNA to increase in WD + AF versus CTL rats (p = 0.07). Liver fat deposition nor liver protein carbonyl content differed between WD + AF versus WD rats, although liver protein carbonyls tended to be lower in WD + AF versus CTL rats (p = 0.08). RNA-seq revealed that 19 liver mRNAs differed between WD + AF versus WD when both groups were compared with CTL rats (+/- 1.5-fold, p < 0.01). Bioinformatics suggest that AF prevented WD-induced alterations in select genes related to the transport and metabolism of carbohydrates in favor of select genes related to lipid transport and metabolism. Finally, serum clinical safety markers and liver pathology

  1. The effect of N-stearoylethanolamine on cholesterol content, fatty acid composition and protein carbonylation level in rats with alimentary obesity-induced insulin resistance

    Directory of Open Access Journals (Sweden)

    O. V. Onopchenko

    2014-12-01

    Full Text Available The effect of N-stearoylethanolamine (NSE on liver free fatty acid composition, cholesterol content and carbonylated protein level in rats with obesity-induced insulin resistance (IR was studied in the work. The experimental insulin resistance was induced by prolonged high fat diet (58% of energy derived from fat for 6 months combined with one injection of low-dose (15 mg/kg of streptozotocin. The lipid assay showed a rise in liver free cholesterol content and a significant reduction in cholesterol esters level. Analyzing liver fatty acid composition, a decrease in polyunsaturated of fatty acid (PUFA level and an increase in monounsaturated fatty acid (MUFA content was found. Fatty acid imbalance with high content of MUFA was associated with elevated level of protein carbonylation. The NSE administration (50 mg/kg of body weight for 2 weeks decreased free cholesterol content, increased cholesterol esters level and reduced free oleic fatty acid content in the liver of rats with IR. The effect of NSE on lipid imbalance led to a decrease in protein carbonylation level that may result in improvement of transmembrane protein function under obesity-induced insulin resistance state.

  2. Effects of organic contaminants in reactive oxygen species, protein carbonylation and DNA damage on digestive gland and haemolymph of land snails.

    Science.gov (United States)

    Itziou, A; Kaloyianni, M; Dimitriadis, V K

    2011-10-01

    The present study focused on early responses of land snails Eobania vermiculata to organic environmental contaminants, by investigating the use of a newly-established method for the measurement of protein carbonylation as a new biomarker of terrestrial pollution, as well as by measuring the ROS production and the DNA damage. Land snails were exposed to different concentrations of chlorpyrifos, parathion-methyl or PAHs in vivo or in vitro in the laboratory. The susceptibility of exposed snails was increased in relation to oxidative stress induced by contaminants tested. A statistically significant increase in ROS production, protein carbonylation and DNA damage was revealed in the snails treated with pollutants, compared to the untreated ones. The results indicated the effectiveness of measuring ROS production and DNA damage and reinforce the application of the present ELISA method in organic terrestrial pollution biomonitoring studies.

  3. Lack of FTSH4 Protease Affects Protein Carbonylation, Mitochondrial Morphology, and Phospholipid Content in Mitochondria of Arabidopsis: New Insights into a Complex Interplay.

    Science.gov (United States)

    Smakowska, Elwira; Skibior-Blaszczyk, Renata; Czarna, Malgorzata; Kolodziejczak, Marta; Kwasniak-Owczarek, Malgorzata; Parys, Katarzyna; Funk, Christiane; Janska, Hanna

    2016-08-01

    FTSH4 is one of the inner membrane-embedded ATP-dependent metalloproteases in mitochondria of Arabidopsis (Arabidopsis thaliana). In mutants impaired to express FTSH4, carbonylated proteins accumulated and leaf morphology was altered when grown under a short-day photoperiod, at 22°C, and a long-day photoperiod, at 30°C. To provide better insight into the function of FTSH4, we compared the mitochondrial proteomes and oxyproteomes of two ftsh4 mutants and wild-type plants grown under conditions inducing the phenotypic alterations. Numerous proteins from various submitochondrial compartments were observed to be carbonylated in the ftsh4 mutants, indicating a widespread oxidative stress. One of the reasons for the accumulation of carbonylated proteins in ftsh4 was the limited ATP-dependent proteolytic capacity of ftsh4 mitochondria, arising from insufficient ATP amount, probably as a result of an impaired oxidative phosphorylation (OXPHOS), especially complex V. In ftsh4, we further observed giant, spherical mitochondria coexisting among normal ones. Both effects, the increased number of abnormal mitochondria and the decreased stability/activity of the OXPHOS complexes, were probably caused by the lower amount of the mitochondrial membrane phospholipid cardiolipin. We postulate that the reduced cardiolipin content in ftsh4 mitochondria leads to perturbations within the OXPHOS complexes, generating more reactive oxygen species and less ATP, and to the deregulation of mitochondrial dynamics, causing in consequence the accumulation of oxidative damage.

  4. Determination of protein-carbonyls and ubiquitin-mediated proteolysis as biomarkers of oxidative-stress in bivalvia and anthozoa

    Energy Technology Data Exchange (ETDEWEB)

    Walker, Stephen Thomas

    2002-07-01

    This study describes the investigation of biomarkers of oxidative-stress in the bivalves Mytilus edulis and Dosinia lupinus, together with host and symbiont tissues of the scleractinian Anthozoa Agaricia agaricites. The biomarkers used were assay of total (via spectrophotometry) and individual (via Western blotting; Oxyblot kit) protein-carbonyls (PC=Os) and content of ubiquitin protein conjugates (UPC) via Western blotting (Bivalvia and Anthozoa) and immunohistochemistry (Anthozoa only). Additional assays for Bivalvia were Trolox equivalent antioxidant capacity (TEAC); and post {gamma}-irradiation survival rates. Experimental stressors for Bivalvia were increased seawater temperature, H{sub 2}O{sub 2} and {sup 60}Co {gamma}-radiation (latter two were used in vivo and in vitro). Comparisons of clean and polluted marine sites are included. Stressors used for Anthozoa were increased solar irradiation concomitant with elevated seawater temperature. Results and conclusions were as follows: individual samples showed considerable variation, pooling of samples improved consistency. Controls for both biomarkers had detectable background levels in each phylum, against which relatively small differences were assessed. In M. edulis, no measurable differences in PC=Os could be determined when elevated seawater temperature or dilute H{sub 2}O{sub 2} (<30% v/v) stressors were used, nor with between-site comparisons. Concentrated H{sub 2}O{sub 2} (30% v/v) produced a small difference. {sup 60}Co {gamma}-radiation produced clearer differences via Oxyblot and spectrophotometric assays. Comparison of four different tissues from the two bivalves found considerable species-specific and tissue-specific differences. Post-irradiation mortality between species was significantly different (<0.001), D. lupinus was more susceptible than M. edulis. TEAC values generally showed a decrease following irradiation (except for digestive gland). UPCs were clearly different between tissues and

  5. Carbonyl J Acid Derivatives Block Protein Priming of Hepadnaviral P Protein and DNA-Dependent DNA Synthesis Activity of Hepadnaviral Nucleocapsids

    Science.gov (United States)

    Wang, Yong-Xiang; Wen, Yu-Mei

    2012-01-01

    Current treatments for chronic hepatitis B are effective in only a fraction of patients. All approved directly antiviral agents are nucleos(t)ide analogs (NAs) that target the DNA polymerase activity of the hepatitis B virus (HBV) P protein; resistance and cross-resistance may limit their long-term applicability. P protein is an unusual reverse transcriptase that initiates reverse transcription by protein priming, by which a Tyr residue in the unique terminal protein domain acts as an acceptor of the first DNA nucleotide. Priming requires P protein binding to the ε stem-loop on the pregenomic RNA (pgRNA) template. This interaction also mediates pgRNA encapsidation and thus provides a particularly attractive target for intervention. Exploiting in vitro priming systems available for duck HBV (DHBV) but not HBV, we demonstrate that naphthylureas of the carbonyl J acid family, in particular KM-1, potently suppress protein priming by targeting P protein and interfering with the formation of P-DHBV ε initiation complexes. Quantitative evaluation revealed a significant increase in complex stability during maturation, yet even primed complexes remained sensitive to KM-1 concentrations below 10 μM. Furthermore, KM-1 inhibited the DNA-dependent DNA polymerase activity of both DHBV and HBV nucleocapsids, including from a lamivudine-resistant variant, directly demonstrating the sensitivity of human HBV to the compound. Activity against viral replication in cells was low, likely due to low intracellular availability. KM-1 is thus not yet a drug candidate, but its distinct mechanism of action suggests that it is a highly useful lead for developing improved, therapeutically applicable derivatives. PMID:22787212

  6. Products of DNA, protein and lipid oxidative damage in relation to vitamin C plasma concentration.

    Science.gov (United States)

    Krajcovicová-Kudlácková, M; Dusinská, M; Valachovicová, M; Blazícek, P; Pauková, V

    2006-01-01

    Oxidative stress plays an important role in the pathogenesis of numerous chronic age-related free radical-induced diseases. Improved antioxidant status minimizes oxidative damage to DNA, proteins, lipids and other biomolecules. Diet-derived antioxidants such as vitamin C, vitamin E, carotenoids and related plant pigments are important in antioxidative defense and maintaining health. The results of long-term epidemiological and clinical studies suggest that protective vitamin C plasma concentration for minimum risk of free radical disease is higher than 50 micromol/l. Products of oxidative damage to DNA (DNA strand breaks with oxidized purines and pyrimidines), proteins (carbonyls) and lipids (conjugated dienes of fatty acids, malondialdehyde) were estimated in a group of apparently healthy adult non-smoking population in dependence on different vitamin C plasma concentrations. Under conditions of protective plasma vitamin C concentrations (>50 micromol/l) significantly lower values of DNA, protein and lipid oxidative damage were found in comparison with the vitamin C-deficient group (fruit and vegetable consumption (leading to higher vitamin C intake and higher vitamin C plasma concentrations) on oxidation of DNA, proteins and lipids is also expressed by an inverse significant correlation between plasma vitamin C and products of oxidative damage. The results suggest an important role of higher and frequent consumption of protective food (fruit, vegetables, vegetable oils, nuts, seeds and cereal grains) in prevention of free radical disease.

  7. Redox regulation of protein damage in plasma

    Directory of Open Access Journals (Sweden)

    Helen R. Griffiths

    2014-01-01

    In this review, we focus on redox regulatory control of those enzymes and processes which control protein maturation during synthesis, produce reactive species, repair and remove damaged plasma proteins. We have highlighted the potential for alterations in the extracellular redox compartment to regulate intracellular redox state and, conversely, for intracellular oxidative stress to alter the cellular secretome and composition of extracellular vesicles. Through secreted, redox-active regulatory molecules, changes in redox state may be transmitted to distant sites.

  8. Plasma protein haptoglobin modulates renal iron loading

    DEFF Research Database (Denmark)

    Fagoonee, Sharmila; Gburek, Jakub; Hirsch, Emilio

    2005-01-01

    Haptoglobin is the plasma protein with the highest binding affinity for hemoglobin. The strength of hemoglobin binding and the existence of a specific receptor for the haptoglobin-hemoglobin complex in the monocyte/macrophage system clearly suggest that haptoglobin may have a crucial role in heme...

  9. May modifications of human plasma proteins stimulated by homocysteine and its thiolactone induce changes of hemostatic function of plasma in vitro?

    Science.gov (United States)

    Olas, Beata; Kołodziejczyk, Joanna; Malinowska, Joanna

    2010-06-01

    Homocysteine (Hcys) may be implicated in different diseases, especially in cardiovascular illnesses. The most reactive form of Hcys is its cyclic thioester-homocysteine thiolactone (HTL), which is formed in plasma and represents up to 0.29% of plasma total Hcys. Recently, it has been observed that Hcys and HTL may modify plasma proteins, including albumin, hemoglobin or fibrinogen, but the role of this process is not yet well known. The aim of our study in vitro was to investigate the modifications of human plasma total proteins after incubation with the reduced form of Hcys in concentrations 10-100 micromol/l, and HTL in concentrations 1-0.1 micromol/l, which correspond to levels found in human plasma during hyperhomocysteinemia in vivo. The aim of our study was also to explain the effects of Hcys and HTL on coagulation activity of human plasma. We showed that in model system in vitro Hcys and HTL change the level of thiol, amino and carbonyl groups in plasma total proteins. Moreover, our studies reported that not only Hcys (10-100 micromol/l), but also HTL (at lower concentrations than Hcys) modulates the coagulation properties of human plasma.

  10. Age-related differences in plasma proteins: how plasma proteins change from neonates to adults.

    Directory of Open Access Journals (Sweden)

    Vera Ignjatovic

    Full Text Available The incidence of major diseases such as cardiovascular disease, thrombosis and cancer increases with age and is the major cause of mortality world-wide, with neonates and children somehow protected from such diseases of ageing. We hypothesized that there are major developmental differences in plasma proteins and that these contribute to age-related changes in the incidence of major diseases. We evaluated the human plasma proteome in healthy neonates, children and adults using the 2D-DIGE approach. We demonstrate significant changes in number and abundance of up to 100 protein spots that have marked differences in during the transition of the plasma proteome from neonate and child through to adult. These proteins are known to be involved in numerous physiological processes such as iron transport and homeostasis, immune response, haemostasis and apoptosis, amongst others. Importantly, we determined that the proteins that are differentially expressed with age are not the same proteins that are differentially expressed with gender and that the degree of phosphorylation of plasma proteins also changes with age. Given the multi-functionality of these proteins in human physiology, understanding the differences in the plasma proteome in neonates and children compared to adults will make a major contribution to our understanding of developmental biology in humans.

  11. Proteins of the canine seminal plasma

    Directory of Open Access Journals (Sweden)

    Annice Aquino-Cortez

    2016-05-01

    Full Text Available ABSTRACT: Studies have been performed to identify the proteins present in canine seminal plasma (SP and relate them to sperm quality as well as to discover molecular markers of reproductive tract diseases. There is evidence that heparin-binding proteins, zinc-binding proteins, and lactoferrin as well as the matrix metalloproteinase, superoxide dismutase, catalase, and glutathione peroxidase enzymes are associated with canine sperm quality. Other studies indicate that prolactin and enzymes like arginine esterase, acid phosphatase, and alkaline phosphatase could be successfully used as biomarkers of reproductive disorders. Thus, the present literature review aims to address aspects related to proteins of the canine SP, their influence on fertility, and their importance as biomarkers of reproductive disorders.

  12. Association of plasma proteins at multiple stages of glycation and antioxidant status with erythrocyte oxidative stress in patients with type 2 diabetes.

    Science.gov (United States)

    Tupe, R S; Diwan, A G; Mittal, V D; Narayanam, P S; Mahajan, K B

    2014-01-01

    This study examines the individual stages of plasma protein glycation, antioxidant status and their association with erythrocyte oxidative stress in patients with type 2 diabetes mellitus (T2DM). Study was carried out on blood from 70 patients with T2DM and 40 healthy age- and gender-matched volunteers. Biomarkers of plasma protein glycation (fructosamine, protein carbonyls, advanced glycation end products [AGEs], amyloid), antioxidant status (thiols, total antioxidant capacity and erythrocyte oxidative parameters), osmotic fragility, lipid peroxidation (LPO), reduced glutathione (GSH) and catalase were determined. Plasma glycation markers were higher in T2DM patients than in healthy volunteers: fructosamine 578 vs. 525 micromol/mL; carbonyl 21.23 vs. 18.84 nmol/mg protein (P glycation markers, positive correlations were evident between fructosamine and amyloid (r = 0.350, P glycation markers showed negative correlation with plasma antioxidant status while positive correlation was demonstrated between erythrocytes fragility and AGEs and amyloid. Erythrocyte LPO levels correlated positively with amyloid. These data suggest that increased levels of multiple plasma protein glycation products in T2DM patients play a key role in reduced plasma antioxidant status and amplified erythrocyte oxidative damage.

  13. Identification of fibrin clot-bound plasma proteins

    NARCIS (Netherlands)

    S. Talens (Simone); F.W.G. Leebeek (Frank); J.A.A. Demmers (Jeroen); D.C. Rijken (Dingeman)

    2012-01-01

    textabstractSeveral proteins are known to bind to a fibrin network and to change clot properties or function. In this study we aimed to get an overview of fibrin clot-bound plasma proteins. A plasma clot was formed by adding thrombin, CaCl2 and aprotinin to citrated platelet-poor plasma and unbound

  14. Zeolite Nanoparticles for Selective Sorption of Plasma Proteins

    NARCIS (Netherlands)

    Rahimi, Mehran; Ng, E. -P.; Bakhtiari, K.; Vinciguerra, M.; Ahmad, H. Ali; Awala, H.; Mintova, S.; Daghighi, M.; Rostami, F. Bakhshandeh; de Vries, Marcel; Motazacker, M. M.; Peppelenbosch, M. P.; Mahmoudi, M.; Rezaee, F.

    2015-01-01

    The affinity of zeolite nanoparticles (diameter of 8-12 nm) possessing high surface area and high pore volume towards human plasma proteins has been investigated. The protein composition (corona) of zeolite nanoparticles has been shown to be more dependent on the plasma protein concentrations and

  15. Zeolite Nanoparticles for Selective Sorption of Plasma Proteins

    NARCIS (Netherlands)

    Rahimi, Mehran; Ng, E. -P.; Bakhtiari, K.; Vinciguerra, M.; Ahmad, H. Ali; Awala, H.; Mintova, S.; Daghighi, M.; Rostami, F. Bakhshandeh; de Vries, Marcel; Motazacker, M. M.; Peppelenbosch, M. P.; Mahmoudi, M.; Rezaee, F.

    2015-01-01

    The affinity of zeolite nanoparticles (diameter of 8-12 nm) possessing high surface area and high pore volume towards human plasma proteins has been investigated. The protein composition (corona) of zeolite nanoparticles has been shown to be more dependent on the plasma protein concentrations and th

  16. Zeolite Nanoparticles for Selective Sorption of Plasma Proteins

    NARCIS (Netherlands)

    M. Rahimi (Mehran); E.-P. Ng; K. Bakhtiari (Kamran); M. Vinciguerra (Manlio); H.A. Ahmad (H. Ali); H. Awala; S. Mintova; M. Daghighi (Mojtaba); F. Bakhshandeh Rostami; M. de Vries (Marieke); M.M. Motazacker (Mohammad); M.P. Peppelenbosch (Maikel); M. Mahmoudi; F. Rezaee (Farhad)

    2015-01-01

    textabstractThe affinity of zeolite nanoparticles (diameter of 8-12 nm) possessing high surface area and high pore volume towards human plasma proteins has been investigated. The protein composition (corona) of zeolite nanoparticles has been shown to be more dependent on the plasma protein concentra

  17. Augmented cardiac formation of oxidatively-induced carbonylated proteins accompanies the increased functional severity of post-myocardial infarction heart failure in the setting of type 1 diabetes mellitus.

    Science.gov (United States)

    Dennis, Kathleen E; Hill, Salisha; Rose, Kristie L; Sampson, Uchechukwu K A; Hill, Michael F

    2013-01-01

    Heart failure (HF) is a dominant cause for the higher mortality of diabetics after myocardial infarction (MI). In the present investigation, we have discovered that higher levels of oxidative stress (OS)-induced carbonylated proteins accompany worsening post-MI HF in the presence of type 1 diabetes. These findings provide a mechanistic link between amplified OS and exacerbation of post-infarction HF in diabetes. Type 1 diabetes mellitus (DM) patients surviving myocardial infarction (MI) manifest an increased incidence of subsequent heart failure (HF). We have previously shown that after MI, type 1 DM is associated with accentuated myocardial oxidative stress (OS) and concomitant worsening of left ventricular (LV) function. However, the precise mechanisms whereby type 1 DM-enhanced OS adversely affects HF after MI remain obscure. As carbonylation of proteins is an irreversible post-translational modification induced only by OS that often leads to the loss of function, we analyzed protein-bound carbonyls in the surviving LV myocardium of MI and DM+MI rats in relation to residual LV function. Type 1 DM was induced in rats via administration of streptozotocin. Two weeks after induction of type 1 DM, MI was produced in DM and non-DM rats by coronary artery ligation. Residual LV function and remodeling was assessed at 4 weeks post-MI by echocardiography. Myocardial carbonylated proteins were detected through OxyBlot analysis, and identified by mass spectrometry. Compared with MI rats, DM+MI rats exhibited significantly poorer residual LV systolic function and elevated wet to dry weight ratios of the lungs. Protein carbonyl content in cardiac tissue and isolated heart mitochondria of DM+MI rats was 20% and 48% higher, respectively, versus MI rats. Anti-oxidative enzymes and fatty acid utilization proteins were among the carbonylated protein candidates identified. These findings implicate myocardial protein carbonylation as part of the molecular pathophysiology of

  18. The 82-plex plasma protein signature that predicts increasing inflammation

    DEFF Research Database (Denmark)

    Tepel, Martin; Beck, Hans C; Tan, Qihua;

    2015-01-01

    transplant recipients and quantified 359 plasma proteins simultaneously using nano-Liquid-Chromatography-Tandem Mass-Spectrometry in individual samples and plasma C-reactive protein on the index day and the next day. Next-day C-reactive protein increased in 59 patients whereas it decreased in 32 patients......The objective of the study was to define the specific plasma protein signature that predicts the increase of the inflammation marker C-reactive protein from index day to next-day using proteome analysis and novel bioinformatics tools. We performed a prospective study of 91 incident kidney....... The prediction model selected and validated 82 plasma proteins which determined increased next-day C-reactive protein (area under receiver-operator-characteristics curve, 0.772; 95% confidence interval, 0.669 to 0.876; P protein signature (P 

  19. Plasma and Plasma Protein Product Transfusion: A Canadian Blood Services Centre for Innovation Symposium.

    Science.gov (United States)

    Zeller, Michelle P; Al-Habsi, Khalid S; Golder, Mia; Walsh, Geraldine M; Sheffield, William P

    2015-07-01

    Plasma obtained via whole blood donation processing or via apheresis technology can either be transfused directly to patients or pooled and fractionated into plasma protein products that are concentrates of 1 or more purified plasma protein. The evidence base supporting clinical efficacy in most of the indications for which plasma is transfused is weak, whereas high-quality evidence supports the efficacy of plasma protein products in at least some of the clinical settings in which they are used. Transfusable plasma utilization remains composed in part of applications that fall outside of clinical practice guidelines. Plasma contains all of the soluble coagulation factors and is frequently transfused in efforts to restore or reinforce patient hemostasis. The biochemical complexities of coagulation have in recent years been rationalized in newer cell-based models that supplement the cascade hypothesis. Efforts to normalize widely used clinical hemostasis screening test values by plasma transfusion are thought to be misplaced, but superior rapid tests have been slow to emerge. The advent of non-vitamin K-dependent oral anticoagulants has brought new challenges to clinical laboratories in plasma testing and to clinicians needing to reverse non-vitamin K-dependent oral anticoagulants urgently. Current plasma-related controversies include prophylactic plasma transfusion before invasive procedures, plasma vs prothrombin complex concentrates for urgent warfarin reversal, and the utility of increased ratios of plasma to red blood cell units transfused in massive transfusion protocols. The first recombinant plasma protein products to reach the clinic were recombinant hemophilia treatment products, and these donor-free equivalents to factors VIII and IX are now being supplemented with novel products whose circulatory half-lives have been increased by chemical modification or genetic fusion. Achieving optimal plasma utilization is an ongoing challenge in the interconnected

  20. Identification of fibrin clot-bound plasma proteins.

    Directory of Open Access Journals (Sweden)

    Simone Talens

    Full Text Available Several proteins are known to bind to a fibrin network and to change clot properties or function. In this study we aimed to get an overview of fibrin clot-bound plasma proteins. A plasma clot was formed by adding thrombin, CaCl(2 and aprotinin to citrated platelet-poor plasma and unbound proteins were washed away with Tris-buffered saline. Non-covalently bound proteins were extracted, separated with 2D gel electrophoresis and visualized with Sypro Ruby. Excised protein spots were analyzed with mass spectrometry. The identity of the proteins was verified by checking the mass of the protein, and, if necessary, by Western blot analysis. Next to established fibrin-binding proteins we identified several novel fibrin clot-bound plasma proteins, including α(2-macroglobulin, carboxypeptidase N, α(1-antitrypsin, haptoglobin, serum amyloid P, and the apolipoproteins A-I, E, J, and A-IV. The latter six proteins are associated with high-density lipoprotein particles. In addition we showed that high-density lipoprotein associated proteins were also present in fibrinogen preparations purified from plasma. Most plasma proteins in a fibrin clot can be classified into three groups according to either blood coagulation, protease inhibition or high-density lipoprotein metabolism. The presence of high-density lipoprotein in clots might point to a role in hemostasis.

  1. Poststroke depression as a factor adversely affecting the level of oxidative damage to plasma proteins during a brain stroke.

    Science.gov (United States)

    Cichoń, Natalia; Bijak, Michał; Miller, Elżbieta; Niwald, Marta; Saluk, Joanna

    2015-01-01

    Poststroke depression, the second most serious psychosomatic complication after brain stroke, leads to delay of the rehabilitation process and is associated with an increased disability and cognitive impairment along with increase in term mortality. Research into the biochemical changes in depression is still insufficiently described. The aim of our study was therefore to evaluate the possible association between plasma protein oxidative/nitrative damages and the development of poststroke depression. We evaluated oxidative/nitrative modifications of specific proteins by measurement of 3-nitrotyrosine and carbonyl groups levels using ELISA test. Additionally, we checked differences in proteins thiol groups by spectrophotometric assay based on reaction between DTNB and thiols. We also evaluated catalase activity in erythrocytes measured as ability to decompose H2O2. Correlation analysis was performed using Spearman's rank. We observed significant (P stroke patients compared to healthy group. Our research shows that oxidative damage of proteins is correlated with the degree of poststroke depression, while nitrative changes do not show any relationship. We demonstrate a positive correlation between the concentration of carbonyl groups and the Geriatric Depression Scale and a negative correlation between the degree of depression and the concentration of -SH groups or catalase activity.

  2. Poststroke Depression as a Factor Adversely Affecting the Level of Oxidative Damage to Plasma Proteins during a Brain Stroke

    Directory of Open Access Journals (Sweden)

    Natalia Cichoń

    2015-01-01

    Full Text Available Poststroke depression, the second most serious psychosomatic complication after brain stroke, leads to delay of the rehabilitation process and is associated with an increased disability and cognitive impairment along with increase in term mortality. Research into the biochemical changes in depression is still insufficiently described. The aim of our study was therefore to evaluate the possible association between plasma protein oxidative/nitrative damages and the development of poststroke depression. We evaluated oxidative/nitrative modifications of specific proteins by measurement of 3-nitrotyrosine and carbonyl groups levels using ELISA test. Additionally, we checked differences in proteins thiol groups by spectrophotometric assay based on reaction between DTNB and thiols. We also evaluated catalase activity in erythrocytes measured as ability to decompose H2O2. Correlation analysis was performed using Spearman’s rank. We observed significant (P<0.001 differences in all oxidative/nitrative stress parameters in brain stroke patients compared to healthy group. Our research shows that oxidative damage of proteins is correlated with the degree of poststroke depression, while nitrative changes do not show any relationship. We demonstrate a positive correlation between the concentration of carbonyl groups and the Geriatric Depression Scale and a negative correlation between the degree of depression and the concentration of -SH groups or catalase activity.

  3. Plasma PIVKA proteins in rabbits given warfarin.

    Science.gov (United States)

    Zivelin, A; Rao, L V; Rapaport, S I

    1996-06-01

    The presence of partially carboxylated forms of the vitamin K dependent coagulation factors (PIVKA) was evaluated in the plasma of rabbits treated with warfarin. Excess antigen over activity as measured in rabbit specific assays was taken as evidence for PIVKA. Our data confirm a previous report of the absence of plasma PIVKA prothrombin. In contrast, plasma PIVKA factors VII, IX, and X were demonstrable. A striking excess of plasma factor IX antigen over activity was measured and a large fraction of the factor IX antigen persisted in the plasma after its adsorption with barium citrate.

  4. POLY(N-VINYLPYRROLIDONE)-MODIFIED SURFACES REPEL PLASMA PROTEIN ADSORPTION

    Institute of Scientific and Technical Information of China (English)

    Xiao-li Liu; Zhao-qiang Wu; Dan Li; Hong Chen

    2012-01-01

    The present work aimed to study the interaction between plasma proteins and PVP-modified surfaces under more complex protein conditions.In the competitive adsorption of fibrinogen (Fg) and human serum albumin (HSA),the modified surfaces showed preferential adsorption of HSA.In 100% plasma,the amount of Fg adsorbed onto PVP-modified surfaces was as low as 10 ng/cm2,suggesting the excellent protein resistance properties of the modified surfaces.In addition,immunoblots of proteins eluted from the modified surfaces after plasma contact confirmed that PVP-modified surfaces can repel most plasma proteins,especially proteins that play important roles in the process of blood coagulation.

  5. Comparative changes in plasma protein concentration, hematocrit and plasma volume during exercise, bedrest and + Gz acceleration.

    Science.gov (United States)

    Van Beaumont, W.; Greenleaf, J. E.

    1972-01-01

    Discussion of experiments which indicate that under conditions of a constant red cell volume the proportional changes in hematocrit and plasma volume during exercise are never equal. On the basis of direct measurements and calculated changes of plasma volume it is concluded that during maximal exercise there is a small loss of protein from the plasma. It is clear that changes in content of blood constituents can only be evaluated correctly after determination of changes in plasma volume.

  6. Cold Atmospheric Plasma Manipulation of Proteins in Food Systems

    DEFF Research Database (Denmark)

    Tolouie, Haniye; Hashemi, Maryam; Mohammadifar, Mohammad Amin

    2017-01-01

    Plasma processing has been getting a lot of attention in recent applications as a novel, eco-friendly, and highly efficient approach. Cold plasma has mostly been used to reduce microbial counts in foodstuff and biological materials, as well as in different levels of packaging, particularly in cases...... where there is thermal sensitivity. As it is a very recent application, the impact of cold plasma treatment has been studied on the protein structures of food and pharmaceutical systems, as well as in the packaging industry. Proteins, as a food constituent, play a remarkable role in the techno...... of plasma on the conformation and function of proteins with food origin, especially enzymes and allergens, as well as protein-made packaging films. In enzyme manipulation with plasma, deactivation has been reported to be either partial or complete. In addition, an activity increase has been observed in some...

  7. 50Hz Extremely Low Frequency Electromagnetic Fields Enhance Protein Carbonyl Groups Content in Cancer Cells: Effects on Proteasomal Systems

    Directory of Open Access Journals (Sweden)

    A. M. Eleuteri

    2009-01-01

    Full Text Available Electromagnetic fields are an assessed cause of prolonging free radicals lifespan. This study was carried out to investigate the influence of extremely low frequency electromagnetic fields on protein oxidation and on the 20S proteasome functionality, the complex responsible for the degradation of oxidized proteins. Caco 2 cells were exposed, for 24–72 hours, to 1 mT, 50 Hz electromagnetic fields. The treatment induced a time-dependent increase both in cell growth and in protein oxidation, more evident in the presence of TPA, while no changes in cell viability were detected. Exposing the cells to 50 Hz electromagnetic fields caused a global activation of the 20S proteasome catalytic components, particularly evident at 72 hours exposure and in the presence of TPA. The finding that EGCG, a natural antioxidant compound, counteracted the field-related pro-oxidant effects demonstrates that the increased proteasome activity was due to an enhancement in intracellular free radicals.

  8. Glycosylation of hemoglobin and plasma proteins in petrochemical plant workers

    Energy Technology Data Exchange (ETDEWEB)

    Unrug, A.; Tomaszewski, L.

    1985-01-01

    The concentration of glycosylated hemoglobin and (plasma) proteins has been measured in 111 workers of 6 MZRiP departments in Plock and in 54 healthy people. In all subjects the mean concentrations of glycosylated hemoglobin and glycosylated plasma proteins have been in so called wide range of normal values. Significant shifts of glycosylated Hb concentrations have been found in two departments--those of ethylenederivatives and distillation. The concentration of glycosylated plasma proteins has been elevated only in workers of the Catalytic Processes Department.

  9. Optimized biotin-hydrazide enrichment and mass spectrometry analysis of peptide carbonyls

    DEFF Research Database (Denmark)

    Havelund, Jesper F.; Wojdyla, K; Jensen, O. N.;

    Irreversible cell damage through protein carbonylation is the result of reaction with reactive oxygen species (ROS) and has been coupled to many diseases. The precise molecular consequences of protein carbonylation, however, are still not clear. The localization of the carbonylated amino acid is ...

  10. Relative quantification of several plasma proteins during liver transplantation surgery.

    Science.gov (United States)

    Parviainen, Ville; Joenväärä, Sakari; Tukiainen, Eija; Ilmakunnas, Minna; Isoniemi, Helena; Renkonen, Risto

    2011-01-01

    Plasma proteome is widely used in studying changes occurring in human body during disease or other disturbances. Immunological methods are commonly used in such studies. In recent years, mass spectrometry has gained popularity in high-throughput analysis of plasma proteins. In this study, we tested whether mass spectrometry and iTRAQ-based protein quantification might be used in proteomic analysis of human plasma during liver transplantation surgery to characterize changes in protein abundances occurring during early graft reperfusion. We sampled blood from systemic circulation as well as blood entering and exiting the liver. After immunodepletion of six high-abundant plasma proteins, trypsin digestion, iTRAQ labeling, and cation-exchange fractionation, the peptides were analyzed by reverse phase nano-LC-MS/MS. In total, 72 proteins were identified of which 31 could be quantified in all patient specimens collected. Of these 31 proteins, ten, mostly medium-to-high abundance plasma proteins with a concentration range of 50-2000 mg/L, displayed relative abundance change of more than 10%. The changes in protein abundance observed in this study allow further research on the role of several proteins in ischemia-reperfusion injury during liver transplantation and possibly in other surgery.

  11. Relative Quantification of Several Plasma Proteins during Liver Transplantation Surgery

    Directory of Open Access Journals (Sweden)

    Ville Parviainen

    2011-01-01

    Full Text Available Plasma proteome is widely used in studying changes occurring in human body during disease or other disturbances. Immunological methods are commonly used in such studies. In recent years, mass spectrometry has gained popularity in high-throughput analysis of plasma proteins. In this study, we tested whether mass spectrometry and iTRAQ-based protein quantification might be used in proteomic analysis of human plasma during liver transplantation surgery to characterize changes in protein abundances occurring during early graft reperfusion. We sampled blood from systemic circulation as well as blood entering and exiting the liver. After immunodepletion of six high-abundant plasma proteins, trypsin digestion, iTRAQ labeling, and cation-exchange fractionation, the peptides were analyzed by reverse phase nano-LC-MS/MS. In total, 72 proteins were identified of which 31 could be quantified in all patient specimens collected. Of these 31 proteins, ten, mostly medium-to-high abundance plasma proteins with a concentration range of 50–2000 mg/L, displayed relative abundance change of more than 10%. The changes in protein abundance observed in this study allow further research on the role of several proteins in ischemia-reperfusion injury during liver transplantation and possibly in other surgery.

  12. Microdomains of SNARE proteins in the plasma membrane

    NARCIS (Netherlands)

    Bogaart, G. van den; Lang, T.; Jahn, R.

    2013-01-01

    Exocytosis is catalyzed by the engagement of SNARE proteins embedded in the plasma membrane with complementary SNAREs in the membrane of trafficking vesicles undergoing exocytosis. In most cells studied so far, SNAREs are not randomly distributed across the plasma membrane but are clustered and

  13. QSARs for Plasma Protein Binding: Source Data and Predictions

    Data.gov (United States)

    U.S. Environmental Protection Agency — The dataset has all of the information used to create and evaluate 3 independent QSAR models for the fraction of a chemical unbound by plasma protein (Fub) for...

  14. Transport proteins of the plant plasma membrane

    Science.gov (United States)

    Assmann, S. M.; Haubrick, L. L.; Evans, M. L. (Principal Investigator)

    1996-01-01

    Recently developed molecular and genetic approaches have enabled the identification and functional characterization of novel genes encoding ion channels, ion carriers, and water channels of the plant plasma membrane.

  15. RELATIONSHIP BETWEEN ANTIOXIDANT POWER OF PLASMA WITH LIPID PEROXIDE FORMATION IN PLASMA AND LIVER DAMAGES CAUSED BY OVERDOSE OF VITAMIN K1 IN ADULT AND WEANLING RATS

    Directory of Open Access Journals (Sweden)

    H. Ansari Hadipour

    2003-08-01

    Full Text Available In this study the plasma levels of lipid peroxidation (LP products, protein carbonyls and antioxidant capacity of plasma as judged by ferric reducing ability of plasma (FRAP assay were compared in adult and weanling rats treated with vitamin K1 phylloquinone.

  16. Zeolite Nanoparticles for Selective Sorption of Plasma Proteins.

    Science.gov (United States)

    Rahimi, M; Ng, E-P; Bakhtiari, K; Vinciguerra, M; Ali Ahmad, H; Awala, H; Mintova, S; Daghighi, M; Bakhshandeh Rostami, F; de Vries, M; Motazacker, M M; Peppelenbosch, M P; Mahmoudi, M; Rezaee, F

    2015-11-30

    The affinity of zeolite nanoparticles (diameter of 8-12 nm) possessing high surface area and high pore volume towards human plasma proteins has been investigated. The protein composition (corona) of zeolite nanoparticles has been shown to be more dependent on the plasma protein concentrations and the type of zeolites than zeolite nanoparticles concentration. The number of proteins present in the corona of zeolite nanoparticles at 100% plasma (in vivo state) is less than with 10% plasma exposure. This could be due to a competition between the proteins to occupy the corona of the zeolite nanoparticles. Moreover, a high selective adsorption for apolipoprotein C-III (APOC-III) and fibrinogen on the zeolite nanoparticles at high plasma concentration (100%) was observed. While the zeolite nanoparticles exposed to low plasma concentration (10%) exhibited a high selective adsorption for immunoglobulin gamma (i.e. IGHG1, IGHG2 and IGHG4) proteins. The zeolite nanoparticles can potentially be used for selectively capture of APOC-III in order to reduce the activation of lipoprotein lipase inhibition during hypertriglyceridemia treatment. The zeolite nanoparticles can be adapted to hemophilic patients (hemophilia A (F-VIII deficient) and hemophilia B (F-IX deficient)) with a risk of bleeding, and thus might be potentially used in combination with the existing therapy.

  17. Zeolite Nanoparticles for Selective Sorption of Plasma Proteins

    Science.gov (United States)

    Rahimi, M.; Ng, E.-P.; Bakhtiari, K.; Vinciguerra, M.; Ahmad, H. Ali; Awala, H.; Mintova, S.; Daghighi, M.; Bakhshandeh Rostami, F.; de Vries, M.; Motazacker, M. M.; Peppelenbosch, M. P.; Mahmoudi, M.; Rezaee, F.

    2015-11-01

    The affinity of zeolite nanoparticles (diameter of 8-12 nm) possessing high surface area and high pore volume towards human plasma proteins has been investigated. The protein composition (corona) of zeolite nanoparticles has been shown to be more dependent on the plasma protein concentrations and the type of zeolites than zeolite nanoparticles concentration. The number of proteins present in the corona of zeolite nanoparticles at 100% plasma (in vivo state) is less than with 10% plasma exposure. This could be due to a competition between the proteins to occupy the corona of the zeolite nanoparticles. Moreover, a high selective adsorption for apolipoprotein C-III (APOC-III) and fibrinogen on the zeolite nanoparticles at high plasma concentration (100%) was observed. While the zeolite nanoparticles exposed to low plasma concentration (10%) exhibited a high selective adsorption for immunoglobulin gamma (i.e. IGHG1, IGHG2 and IGHG4) proteins. The zeolite nanoparticles can potentially be used for selectively capture of APOC-III in order to reduce the activation of lipoprotein lipase inhibition during hypertriglyceridemia treatment. The zeolite nanoparticles can be adapted to hemophilic patients (hemophilia A (F-VIII deficient) and hemophilia B (F-IX deficient)) with a risk of bleeding, and thus might be potentially used in combination with the existing therapy.

  18. Differential plasma protein binding to metal oxide nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Deng, Zhou J; Mortimer, Gysell; Minchin, Rodney F [School of Biomedical Sciences, University of Queensland, Brisbane, QLD 4072 (Australia); Schiller, Tara; Musumeci, Anthony; Martin, Darren, E-mail: r.minchin@uq.edu.a [Australian Institute for Bioengineering and Nanotechnology, University of Queensland, Brisbane, QLD 4072 (Australia)

    2009-11-11

    Nanoparticles rapidly interact with the proteins present in biological fluids, such as blood. The proteins that are adsorbed onto the surface potentially dictate the biokinetics of the nanomaterials and their fate in vivo. Using nanoparticles with different sizes and surface characteristics, studies have reported the effects of physicochemical properties on the composition of adsorbed plasma proteins. However, to date, few studies have been conducted focusing on the nanoparticles that are commonly exposed to the general public, such as the metal oxides. Using previously established ultracentrifugation approaches, two-dimensional gel electrophoresis and mass spectrometry, the current study investigated the binding of human plasma proteins to commercially available titanium dioxide, silicon dioxide and zinc oxide nanoparticles. We found that, despite these particles having similar surface charges in buffer, they bound different plasma proteins. For TiO{sub 2}, the shape of the nanoparticles was also an important determinant of protein binding. Agglomeration in water was observed for all of the nanoparticles and both TiO{sub 2} and ZnO further agglomerated in biological media. This led to an increase in the amount and number of different proteins bound to these nanoparticles. Proteins with important biological functions were identified, including immunoglobulins, lipoproteins, acute-phase proteins and proteins involved in complement pathways and coagulation. These results provide important insights into which human plasma proteins bind to particular metal oxide nanoparticles. Because protein absorption to nanoparticles may determine their interaction with cells and tissues in vivo, understanding how and why plasma proteins are adsorbed to these particles may be important for understanding their biological responses.

  19. Interaction of plasma proteins with commercial protein repellent polyvinyl chloride (PVC): a word of caution.

    Science.gov (United States)

    De Somer, F; Van Landschoot, A; Van Nooten, G; Delanghe, J

    2008-07-01

    Protein adsorption onto polymers remains a problem. In recent years, several protein-repellent PVC tubings have been developed. Although several studies report the interaction between plasma coagulation proteins and PVC, few address the interaction with other plasma proteins. Two commercial brands of untreated medical grade PVC tubing, phosphorylcholine-coated PVC tubing, triblock-copolymer (polycaprolactone-polydimethylsiloxane-polycaprolactone)-treated PVC tubing and poly-2-methoxyethylacrylate (PMEA)-coated tubing were exposed for 60 minutes to human plasma. A broad spectrum of plasma proteins was found on all tubing. The adsorbed albumin to total protein ratio is lower than the similar ratio in plasma while alpha1 and alpha2 globulins are over-represented in the protein spectrum. On PMEA tubing, not only alpha globulins, but also beta and gamma globulins, are found in high concentrations in the adsorbed protein. PMEA tubing and uncoated PVC tubing of brand B had a higher amount of protein adsorbed compared against all other tubing (p < 0.05). There were no statistical differences in protein adsorption between the triblock-copolymer-treated tubing, the phosphorylcholine-coated tubing and the uncoated PVC tubing of brand A. The average thickness of the protein layer was 23 nm. Plasma protein adsorption still exists on uncoated and protein-repellent tubing and can initiate a systemic inflammatory reaction.

  20. Protein Adsorption on Various Plasma-Treated Polyethylene Terephthalate Substrates

    Directory of Open Access Journals (Sweden)

    Karin Stana-Kleinschek

    2013-10-01

    Full Text Available Protein adhesion and cell response to plasma-treated polymer surfaces were studied. The polymer polyethylene terephthalate (PET was treated in either an oxygen plasma to make the surface hydrophilic, or a tetrafluoromethane CF4 plasma to make the surface hydrophobic. The plasma source was radiofrequency (RF discharge. The adsorption of albumin and other proteins from a cell-culture medium onto these surfaces was studied using a quartz crystal microbalance (QCM, X-ray photoelectron spectroscopy (XPS and atomic force microscopy (AFM. The cellular response to plasma-treated surfaces was studied as well using an MTT assay and scanning electron microscopy (SEM. The fastest adsorption rate was found on the hydrophilic oxygen plasma-treated sample, and the lowest was found on the pristine untreated sample. Additionally, the amount of adsorbed proteins was higher for the oxygen-plasma-treated surface, and the adsorbed layer was more viscoelastic. In addition, cell adhesion studies support this finding because the best cell adhesion was observed on oxygen-plasma-treated substrates.

  1. The dynamics of plant plasma membrane proteins: PINs and beyond.

    Science.gov (United States)

    Luschnig, Christian; Vert, Grégory

    2014-08-01

    Plants are permanently situated in a fixed location and thus are well adapted to sense and respond to environmental stimuli and developmental cues. At the cellular level, several of these responses require delicate adjustments that affect the activity and steady-state levels of plasma membrane proteins. These adjustments involve both vesicular transport to the plasma membrane and protein internalization via endocytic sorting. A substantial part of our current knowledge of plant plasma membrane protein sorting is based on studies of PIN-FORMED (PIN) auxin transport proteins, which are found at distinct plasma membrane domains and have been implicated in directional efflux of the plant hormone auxin. Here, we discuss the mechanisms involved in establishing such polar protein distributions, focusing on PINs and other key plant plasma membrane proteins, and we highlight the pathways that allow for dynamic adjustments in protein distribution and turnover, which together constitute a versatile framework that underlies the remarkable capabilities of plants to adjust growth and development in their ever-changing environment.

  2. Comparative Plasma Protein Profiling of Hemoglobin H Disease

    Directory of Open Access Journals (Sweden)

    Kamonlak Leecharoenkiat

    2014-01-01

    Full Text Available HbH and HbH-constant spring (HbH-CS are the most common forms of α-thalassemia detected in the Thai population. The accumulation of excess β globin chains in these diseases results in increased red cell hemolysis, and patients with HbH-CS normally have a more severe clinical presentation than patients with HbH disease. This study aimed to detect alterations in the expression of plasma proteins of HbH and HbH-CS patients as compared to normal plasma. Platelet poor plasma was separated from HbH and HbH-CS and normal subjects and differential plasma proteins were detected using two-dimensional gel electrophoresis and identified using LC/MS/MS. A total of 14 differentially expressed proteins were detected of which 5 proteins were upregulated and 9 were downregulated. Most of the differentially expressed proteins are liver secreted proteins involved in hemolysis, oxidative stress response, and hemoglobin degradation. Seven proteins were found to be differentially expressed between HbH and HbH-CS. Levels of haptoglobin, a hemoglobin scavenging protein, were significantly increased in HbH patients as compared to HbH-CS patients. The identification of differentially expressed proteins may lead to a better understanding of the biological events underlying the clinical presentation of HbH and HbH-CS patients and can have application as hemolytic markers or severity predictors.

  3. Characterization of oxidative carbonylation on recombinant monoclonal antibodies.

    Science.gov (United States)

    Yang, Yi; Stella, Cinzia; Wang, Weiru; Schöneich, Christian; Gennaro, Lynn

    2014-05-20

    In the biotechnology industry, oxidative carbonylation as a post-translational modification of protein pharmaceuticals has not been studied in detail. Using Quality by Design (QbD) principles, understanding the impact of oxidative carbonylation on product quality of protein pharmaceuticals, particularly from a site-specific perspective, is critical. However, comprehensive identification of carbonylation sites has so far remained a very difficult analytical challenge for the industry. In this paper, we report for the first time the identification of specific carbonylation sites on recombinant monoclonal antibodies with a new analytical approach via derivatization with Girard's Reagent T (GRT) and subsequent peptide mapping with high-resolution mass spectrometry. Enhanced ionization efficiency and high quality MS(2) data resulted from GRT derivatization were observed as key benefits of this approach, which enabled direct identification of carbonylation sites without any fractionation or affinity enrichment steps. A simple data filtering process was also incorporated to significantly reduce false positive assignments. Sensitivity and efficiency of this approach were demonstrated by identification of carbonylation sites on both unstressed and oxidized antibody bulk drug substances. The applicability of this approach was further demonstrated by identification of 14 common carbonylation sites on three highly similar IgG1s. Our approach represents a significant improvement to the existing analytical methodologies and facilitates extended characterization of oxidative carbonylation on recombinant monoclonal antibodies and potentially other protein pharmaceuticals in the biotechnology industry.

  4. Organization and Dynamics of Receptor Proteins in a Plasma Membrane.

    Science.gov (United States)

    Koldsø, Heidi; Sansom, Mark S P

    2015-11-25

    The interactions of membrane proteins are influenced by their lipid environment, with key lipid species able to regulate membrane protein function. Advances in high-resolution microscopy can reveal the organization and dynamics of proteins and lipids within living cells at resolutions membranes of in vivo-like complexity. We explore the dynamics of proteins and lipids in crowded and complex plasma membrane models, thereby closing the gap in length and complexity between computations and experiments. Our simulations provide insights into the mutual interplay between lipids and proteins in determining mesoscale (20-100 nm) fluctuations of the bilayer, and in enabling oligomerization and clustering of membrane proteins.

  5. Regulation of IGF binding protein proteolysis by pregnancy-associated plasma protein-ARegulation of IGF binding protein proteolysis by pregnancy-associated plasma protein-A

    DEFF Research Database (Denmark)

    Gaidamauskas, Ervinas

    During his PhD studies, Ervinas Gaidamauskas researched the proteins pregnancy-associated plasma protein-A (PAPP-A) and its homologue PAPP-A2 in vitro. As suggested by its name, PAPP-A plays an important role in pregnancy and fetal development. Additionally, recent studies indicate a newly...

  6. Regulation of IGF binding protein proteolysis by pregnancy-associated plasma protein-ARegulation of IGF binding protein proteolysis by pregnancy-associated plasma protein-A

    DEFF Research Database (Denmark)

    Gaidamauskas, Ervinas

    During his PhD studies, Ervinas Gaidamauskas researched the proteins pregnancy-associated plasma protein-A (PAPP-A) and its homologue PAPP-A2 in vitro. As suggested by its name, PAPP-A plays an important role in pregnancy and fetal development. Additionally, recent studies indicate a newly...

  7. Hypochlorite-induced oxidation of proteins in plasma

    DEFF Research Database (Denmark)

    Hawkins, C L; Davies, Michael Jonathan

    1999-01-01

    Activated phagocyte cells generate hypochlorite (HOCl) via the release of H2O2 and the enzyme myeloperoxidase. Plasma proteins are major targets for HOCl, although little information is available about the mechanism(s) of oxidation. In this study the reaction of HOCl (at least 50 microM) with dil......Activated phagocyte cells generate hypochlorite (HOCl) via the release of H2O2 and the enzyme myeloperoxidase. Plasma proteins are major targets for HOCl, although little information is available about the mechanism(s) of oxidation. In this study the reaction of HOCl (at least 50 micro......M) with diluted fresh human plasma has been shown to generate material that oxidizes 5-thio-2-nitrobenzoic acid; these oxidants are believed to be chloramines formed from the reaction of HOCl with protein amine groups. Chloramines have also been detected with isolated plasma proteins treated with HOCl. In both....... These results are consistent with protein-derived chloramines, and the radicals derived from them, as contributing agents in HOCl-induced plasma protein oxidation....

  8. The Short-Term Effects of Soybean Intake on Oxidative and Carbonyl Stress in Men and Women

    Directory of Open Access Journals (Sweden)

    Lukáč Halčák

    2013-05-01

    Full Text Available Beyond other beneficial effects, a soy-rich diet has been shown to reduce the risk of cardiovascular diseases and diabetic complications. Reduction of oxidative and carbonyl stress has been proposed as the underlying mechanism, but the evidence for this is lacking. The aim of our study was to evaluate the effects of short-term increased soy intake on oxidative and carbonyl stress parameters in young volunteers. Young healthy probands (omnivores of both genders (55 women, 33 men were given soybeans (2 g/kg bodyweight daily for one week. Markers of oxidative and carbonyl stress were measured in plasma at the beginning and at the end of one week soybean intake and after another week of a wash-out period. Total antioxidant capacity was increased by soybean intake in both genders. This led to decreased levels of advanced oxidation protein products in women, but not in men. On the contrary, in men, soybean intake increased lipoperoxidation. No effects on carbonyl stress markers (advanced glycation end products-specific fluorescence and fructosamine were found. Soybean intake has gender-specific effects on oxidative stress in young healthy probands potentially due to divergent action and metabolism of phytoestrogens in men and women. Effects of soybean intake on carbonyl stress should be evaluated in longer studies.

  9. Lowering of plasma phospholipid transfer protein activity by acute hyperglycaemia-induced hyperinsulinaemia in healthy men

    NARCIS (Netherlands)

    vanTol, A; Ligtenberg, JJM; Riemens, SC; vanHaeften, TW; Dullaart, RPF

    1997-01-01

    Human plasma contains two lipid transfer proteins involved in the remodelling of plasma lipoproteins: cholesteryl ester transfer protein (CETP) and phospholipid transfer protein (PLTP). CETP mediates the transfer/exchange of cholesterylesters, triglycerides and phospholipids between high-density lip

  10. Assessing of plasma protein denaturation induced by exposure to cadmium, electromagnetic fields and their combined actions on rat.

    Science.gov (United States)

    Hassan, Nahed S; Abdelkawi, Salwa A

    2014-06-01

    In our environment, we have numerous chances to be exposed to not only electromagnetic fields (EMFs) but also many chemicals containing mutagens. Therefore, the aim of this study was to estimate whether rat's exposure to cadmium and/or EMFs could cause oxidative damage to molecular structure of proteins and whether and to what extent the effects of co-exposure differ from those observed under the treatment with each exposure alone. Thirty-two rats were divided into four groups. Group 1 was termed as control, group 2 was treated with cadmium (3.0 mg/Kg), group 3 was exposed to EMF (10 mT/h/day) and group 4 was treated with cadmium and exposed to EMF. Protein carbonyls (PCO) in the plasma as a marker of oxidative protein damage and total oxidant status (TOS), as well as electrical conductivity and SDS electrophoresis to estimate changes in molecular structure of protein, were determined. The exposure to Cd and/or EMF led to oxidative protein damage (increased PCO and TOS) accomplished by increased stress of electrical charges on the surface of the protein molecule (increased electrical conductivity) and changes in the molecular structure of protein. The effects were more pronounced after treatment with both Cd and EMF than at the treatment with each exposure alone. The serious damage to proteins at the co-exposure to Cd and EMF seems to be due to the interference of the EMF with the toxic activity of cadmium. This work concluded that combined exposure to Cd and EMFs might increase the risk of plasma damage via enhancing free radical generation and protein oxidation.

  11. Plasma concentrations of four pregnancy proteins in complications of pregnancy.

    Science.gov (United States)

    Lin, T M; Halbert, S P; Spellacy, W N; Berne, B H

    1977-08-01

    Toxemia of pregnancy was associated with an elevation of the pregnancy-associated plasma protein (PAPP)-A concentration, as compared to the level in normal pregnancy in the last month of gestation. The other pregnancy proteins measured were not altered in toxemia. In twin pregnancies, the PAPP-A, PAPP-C, and human placental lactogen levels were all increased, particularly PAPP-A. On the other hand, pregnancy zone protein was not affected by twinning. Pregnancy with diabetes showed normal levels of these proteins.

  12. Fish Proteins as Targets of Ferrous-Catalyzed Oxidation: Identification of Protein Carbonyls by Fluorescent Labeling on Two-Dimensional Gels and MALDI-TOF/TOF Mass Spectrometry

    DEFF Research Database (Denmark)

    Pazos, Manuel; da Rocha, Angela Pereira; Roepstorff, Peter

    2011-01-01

    Protein oxidation in fish meat is considered to affect negatively the muscle texture. An important source of free radicals taking part in this process is Fenton's reaction dependent on ferrous ions present in the tissue. The aim of this study was to investigate the susceptibility of cod muscle...... dehydrogenase, creatine kinase, and enolase were the sarcoplasmic proteins most vulnerable to ferrous-catalyzed oxidation. Moreover, NDK, phosphoglycerate mutase, and GAPDH were identified in several spots differing by their pI, and those forms showed different susceptibilities to metal-catalyzed oxidation...

  13. Spectrophotometric and Refractometric Determination of Total Protein in Avian Plasma

    Directory of Open Access Journals (Sweden)

    Rodica Căpriță

    2013-10-01

    Full Text Available The aim of this study was to compare the total protein values obtained in heparin plasma of chickens by a spectrophotometric technique (biuret method, and the values obtained on the same day in the same samples by refractometry. The results obtained by refractometry (average value 2.638±0.153g% were higher than those obtained by the spectrophotometric method (average value 2.441±0.181g%. There was a low correlation (r = 0.6709 between the total protein values, determined with both methods. Protein is the major determinant of plasma refractive index, but glucose contributes too. The refractometric method is not recommended in chickens for the determination of total protein, because avian blood glucose concentration averages about twice than in mammalian blood.

  14. Stereoselective binding of chiral drugs to plasma proteins

    Institute of Scientific and Technical Information of China (English)

    Qi SHEN; Lu WANG; Hui ZHOU; Hui-di JIANG; Lu-shan YU; Su ZENG

    2013-01-01

    Chiral drugs show distinct biochemical and pharmacological behaviors in the human body.The binding of chiral drugs to plasma proteins usually exhibits stereoselectivity,which has a far-reaching influence on their pharmacological activities and pharmacokinetic profiles.In this review,the stereoselective binding of chiral drugs to human serum albumin (HSA),α1-acid glycoprotein (AGP)and lipoprotein,three most important proteins in human plasma,are detailed.Furthermore,the application of AGP variants and recombinant fragments of HSA for studying enantiomer binding properties is also discussed.Apart from the stereoselectivity of enantiomer-protein binding,enantiomer-enantiomer interactions that may induce allosteric effects are also described.Additionally,the techniques and methods used to determine drug-protein binding parameters are briefly reviewed.

  15. [Determination of plasma protein binding rate of arctiin and arctigenin with ultrafiltration].

    Science.gov (United States)

    Han, Xue-Ying; Wang, Wei; Tan, Ri-Qiu; Dou, De-Qiang

    2013-02-01

    To determine the plasma protein binding rate of arctiin and arctigenin. The ultrafiltration combined with HPLC was employed to determine the plasma protein binding rate of arctiin and arctigenin as well as rat plasma and healthy human plasma proteins. The plasma protein binding rate of arctiin with rat plasma at the concentrations of 64. 29, 32.14, 16.07 mg x L(-1) were (71.2 +/- 2.0)%, (73.4 +/- 0.61)%, (78.2 +/- 1.9)%, respectively; while the plasma protein binding rate of arctiin with healthy human plasma at the above concentrations were (64.8 +/- 3.1)%, (64.5 +/- 2.5)%, (77.5 +/- 1.7)%, respectively. The plasma protein binding rate of arctigenin with rat plasma at the concentrations of 77.42, 38.71, 19.36 mg x L(-1) were (96.7 +/- 0.41)%, (96.8 +/- 1.6)%, (97.3 +/- 0.46)%, respectively; while the plasma protein binding rate of arctigenin with normal human plasma at the above concentrations were (94.7 +/- 3.1)%, (96.8 +/- 1.6)%, (97.9 +/- 1.3)%, respectively. The binding rate of arctiin with rat plasma protein was moderate, which is slightly higher than the binding rate of arctiin with healthy human plasma protein. The plasma protein binding rates of arctigenin with both rat plasma and healthy human plasma are very high.

  16. Use of refractometry for determination of psittacine plasma protein concentration.

    Science.gov (United States)

    Cray, Carolyn; Rodriguez, Marilyn; Arheart, Kristopher L

    2008-12-01

    Previous studies have demonstrated both poor and good correlation of total protein concentrations in various avian species using refractometry and biuret methodologies. The purpose of the current study was to compare these 2 techniques of total protein determination using plasma samples from several psittacine species and to determine the effect of cholesterol and other solutes on refractometry results. Total protein concentration in heparinized plasma samples without visible lipemia was analyzed by refractometry and an automated biuret method on a dry reagent analyzer (Ortho 250). Cholesterol, glucose, and uric acid concentrations were measured using the same analyzer. Results were compared using Deming regression analysis, Bland-Altman bias plots, and Spearman's rank correlation. Correlation coefficients (r) for total protein results by refractometry and biuret methods were 0.49 in African grey parrots (n=28), 0.77 in Amazon parrots (20), 0.57 in cockatiels (20), 0.73 in cockatoos (36), 0.86 in conures (20), and 0.93 in macaws (38) (Prefractometry in Amazon parrots, conures, and macaws (n=25 each, PRefractometry can be used to accurately measure total protein concentration in nonlipemic plasma samples from some psittacine species. Method and species-specific reference intervals should be used in the interpretation of total protein values.

  17. NMR Studies of Some Plasma Proteins.

    Science.gov (United States)

    Lawrence, Mark P.

    Available from UMI in association with The British Library. Requires signed TDF. The work reported in this thesis consists of a study of the solution structure of a domain of protein structure found in some of the enzymes involved in blood coagulation. These domains, known as kringles, are of between 78 and 82 residues and contain three conserved disulphide bridges in their primary sequence. The study attempts to elucidate the nature of the lysine-binding site of the fourth kringle of human plasminogen to probe its physiological action, and a theory is developed to explain the overall fold of the protein in terms of its physiological role. The protein structure is found to contain only one small region of secondary structure, an antiparallel beta-sheet of about 8 residues, which provides the support for the binding site. The binding site itself consists of a hydrophobic channel provided by the aromatic residues at positions 61, 63, 71 and 73 in the beta-sheet and a negatively charged site at one end of this channel provided by the aspartic acid residues at positions 54 and 56. The beta-sheet appears to become more tightly defined on binding the kringle with alpha,omega -amino acids which are analogues of lysine and exhibit known anti-fibrinolytic properties. The rest of the solution structure appears to be less clearly defined and relies mainly on the three disulphide bridges and some rather isolated hydrogen bonding for maintenance of the fold. An explanation for this structure with a rigid binding site and a more flexible region for the remainder of the domain is proposed. Shorter studies are reported on the second kringle of bovine prothrombin and the first of human plasminogen which suggest strongly that the kringle fold is conserved.

  18. Plasma protein characteristics of long-term hemodialysis survivors.

    Directory of Open Access Journals (Sweden)

    Yao-Ping Lin

    Full Text Available Hemodialysis (HD patients are under recurrent circulatory stress, and hemodialysis has a high mortality rate. The characteristics of plasma proteomes in patients surviving long-term HD remain obscure, as well as the potential biomarkers in predicting prognoses. This study reports the proteome analyses of patient plasma from non-diabetic long-term HD (LHD, dialysis vintage 14.9±4.1 years, n = 6 and the age/sex/uremic etiology-comparable short-term HD (SHD, dialysis vintage 5.3±2.9 years, n = 6 using 2-DE and mass spectrometry. In addition, a 4-year longitudinal follow-up of 60 non-diabetic HD patients was subsequently conducted to analyze the baseline plasma proteins by ELISA in predicting prognosis. Compared to the SHD, the LHD survivors had increased plasma vitamin D binding proteins (DBP and decreased clusterin, apolipoprotein A-IV, haptoglobin, hemopexin, complement factors B and H, and altered isoforms of α1-antitrypsin and fibrinogen gamma. During the 45.7±15 months for follow-up of the 60 HD patient cases, 16 patients died. Kaplan-Meier analysis demonstrated that HD patients with the lowest tertile of the baseline plasma DBP level have a significantly higher mortality rate. Multivariate Cox regression analysis further indicated that DBP is an independent predictor of mortality. In summary, the altered plasma proteins in LHD implicated accelerated atherosclerosis, defective antioxidative activity, increased inflammation/infection, and organ dysfunction. Furthermore, lower baseline plasma DBP in HD patients is related to mortality. The results suggest that the proteomic approach could help discover the potential biomarker in HD prognoses.

  19. Sardine protein diet increases plasma glucagon-like peptide-1 levels and prevents tissue oxidative stress in rats fed a high-fructose diet.

    Science.gov (United States)

    Madani, Zohra; Sener, Abdullah; Malaisse, Willy J; Dalila, Ait Yahia

    2015-11-01

    The current study investigated whether sardine protein mitigates the adverse effects of fructose on plasma glucagon‑like peptide-1 (GLP-1) and oxidative stress in rats. Rats were fed casein (C) or sardine protein (S) with or without high‑fructose (HF) for 2 months. Plasma glucose, insulin, GLP‑1, lipid and protein oxidation and antioxidant enzymes were assayed. HF rats developed obesity, hyperglycemia, hyperinsulinemia, insulin resistance and oxidative stress despite reduced energy and food intakes. High plasma creatinine and uric acid levels, in addition to albuminuria were observed in the HF groups. The S‑HF diet reduced plasma glucose, insulin, creatinine, uric acid and homeostasis model assessment‑insulin resistance index levels, however increased GLP‑1 levels compared with the C‑HF diet. Hydroperoxides were reduced in the liver, kidney, heart and muscle of S‑HF fed rats compared with C‑HF fed rats. A reduction in liver, kidney and heart carbonyls was observed in S‑HF fed rats compared with C‑HF fed rats. Reduced levels of nitric oxide (NO) were detected in the liver, kidney and heart of the S‑HF fed rats compared with C‑HF fed rats. The S diet compared with the C diet reduced levels of liver hydroperoxides, heart carbonyls and kidney NO. The S‑HF diet compared with the C‑HF diet increased the levels of liver and kidney superoxide dismutase, liver and muscle catalase, liver, heart and muscle glutathione peroxidase and liver ascorbic acid. The S diet prevented and reversed insulin resistance and oxidative stress, and may have benefits in patients with metabolic syndrome.

  20. Adsorption of proteins from plasma at polyester non-wovens.

    Science.gov (United States)

    Klomp, A J; Engbers, G H; Mol, J; Terlingen, J G; Feijen, J

    1999-07-01

    Polyester non-wovens in filters for the removal of leukocytes from platelet concentrates (PCs) must be platelet compatible. In PC filtration, the adsorption of proteins at the plasma-non-woven interface can be of great importance with respect to the yield of platelets. Unmodified and radio frequency glow discharge (RFGD) treated poly(ethylene terephthalate) non-woven (NW-PET) and two commercial surface-modified non-wovens were contacted with human plasma. Protein desorption by sodium dodecyl sulphate (SDS) was evaluated by X-ray photoelectron spectroscopy (XPS). The desorbed proteins were characterized by gel electrophoresis and immunoblotting. Compared to the commercial surface-modified non-wovens, unmodified and RFGD-treated NW-PETs adsorbed a relatively high amount of protein. Significantly more protein was removed from the hydrophobic NW-PET by SDS than from the hydrophilic RFGD-treated non-wovens. RFGD treatment of NW-PET reduces the reversibility of protein adsorption. Less albumin and fibrinogen were removed from the RFGD-treated non-wovens than from NW-PET. In addition, a large amount of histidine-rich glycoprotein was removed from RFGD-treated non-wovens, but not from NW-PET. The different behaviour of RFGFD-treated non-wovens towards protein adsorption is probably caused by differences in the chemical reactivity of the non-woven surfaces.

  1. Disproportional changes in hematocrit, plasma volume, and proteins during exercise and bed rest.

    Science.gov (United States)

    Van Beaumont, W.; Greenleaf, J. E.; Juhos, L.

    1972-01-01

    The interrelationships between the changes in plasma volume, hematocrit, and plasma proteins during muscular exercise and bed rest were investigated. Proportionally, the changes in hematocrit are always smaller than the changes in plasma volume. For this reason changes in the concentration of blood constituents can only be quantitated on the basis of plasma volume changes. During short periods of intensive exercise, there was a small loss of plasma proteins. With prolonged submaximal exercise there was a net gain in plasma protein, which contributes to stabilization of the vascular volume. Prolonged bed rest induced hypoproteinemia; this loss of plasma protein probably plays an important role in recumbency hypovolemia.

  2. Influence of carbonyl stress on rheological alterations of blood materials and decarbonylation effect of glutathione

    Institute of Scientific and Technical Information of China (English)

    彭密军; 蔡建光; 贺洪; 龚萍; 李国林; 汤婷; 朱泽瑞; 印大中

    2008-01-01

    The effects of various toxic carbonyls such as malondialdehyde(MDA),a secondary product of lipid peroxidation,and other aldehydes on rheological parameters and their relationship with aging-associated alterations were studied.Both MDA and glutaraldehyde(Glu) in different concentrations significantly increase viscosity,plastic viscosity and yield stress of human plasma and erythrocyte suspensions.MDA(20 mmol/L) reduces sharply the typical fluorescence of proteins(excitation 280 nm/emission 350 nm),and produces age pigment-like fluorescence with a strong emission peak at 460 nm when excites at 395 nm by only being incubated for some hours.In contrast,Glu decreases merely the fluorescence of proteins without producing age pigment-like fluorescence.These data suggest interestingly that the MDA-induced gradual protein cross linking seems to form from different mechanisms compared to the fast rheological changes of blood materials which may take place either in acute and chronic diseases or during aging.On the other hand,MDA induces various deleterious alterations of erythrocytes whereas glutathione(GSH) inhibits the MDA-related carbonyl stress in a concentration-dependent manner.The results indicate that carbonyl-amino reaction exists in the blood widely and GSH has the ability to interrupt or reverse this reaction in a certain way.It implies that carbonyl stress may be one of the important factors in blood stasis and suggests a theoretical and practical approach in anti-stresses and anti-aging.

  3. Radioimmunoassay for pregnancy-associated plasma protein A

    Energy Technology Data Exchange (ETDEWEB)

    Sinosich, M.J. (Royal North Shore Hospital, Sidney, New South Wales, Australia); Teisner, B.; Folkerson, J.; Saunders, D.M.; Grudzinskas, J.G.

    1982-01-01

    A specific and highly sensitive radioimmunoassay for determination of pregnancy-associated plasma protein A in human serum is described. The minimum detection limit for this protein was 2.9 ..mu..g/L. The within- and between-assay coefficients of variation were 4.0 and 4.5%, respectively. The circulating protein was detected within 32 days of conception in eight normal pregnancies and within 21 days in a twin pregnancy. Circulating concentrations in the mother at term were consistently higher (10-fold) than in matched amniotic fluid; none was detected in the umbilical circulation. This protein was also detected in the circulation of patients with hydatidiform mole. This assay will permit investigations into the clinical evaluation of measurements of the protein during early pregnancy and trophoblastic disease.

  4. Haptoglobin-related protein is a high-affinity hemoglobin-binding plasma protein

    DEFF Research Database (Denmark)

    Nielsen, Marianne Jensby; Petersen, Steen Vang; Jacobsen, Christian

    2006-01-01

    Haptoglobin-related protein (Hpr) is a primate-specific plasma protein associated with apolipoprotein L-I (apoL-I)-containing high-density lipoprotein (HDL) particles shown to be a part of the innate immune defense. Despite the assumption hitherto that Hpr does not bind to hemoglobin, the present...

  5. Plasma membrane microdomains regulate turnover of transport proteins in yeast

    Science.gov (United States)

    Grossmann, Guido; Malinsky, Jan; Stahlschmidt, Wiebke; Loibl, Martin; Weig-Meckl, Ina; Frommer, Wolf B.; Opekarová, Miroslava; Tanner, Widmar

    2008-01-01

    In this study, we investigate whether the stable segregation of proteins and lipids within the yeast plasma membrane serves a particular biological function. We show that 21 proteins cluster within or associate with the ergosterol-rich membrane compartment of Can1 (MCC). However, proteins of the endocytic machinery are excluded from MCC. In a screen, we identified 28 genes affecting MCC appearance and found that genes involved in lipid biosynthesis and vesicle transport are significantly overrepresented. Deletion of Pil1, a component of eisosomes, or of Nce102, an integral membrane protein of MCC, results in the dissipation of all MCC markers. These deletion mutants also show accelerated endocytosis of MCC-resident permeases Can1 and Fur4. Our data suggest that release from MCC makes these proteins accessible to the endocytic machinery. Addition of arginine to wild-type cells leads to a similar redistribution and increased turnover of Can1. Thus, MCC represents a protective area within the plasma membrane to control turnover of transport proteins. PMID:19064668

  6. Do plasma proteins distinguish between liposomes of varying charge density?

    KAUST Repository

    Capriotti, Anna Laura

    2012-03-01

    Cationic liposomes (CLs) are one of the most employed nonviral nanovector systems in gene therapy. However, their transfection efficiency is strongly affected by interactions with plasma components, that lead to the formation of a "protein corona" onto CL surface. The interactions between nanoparticles entering the body and biomolecules have an essential role for their biodistribution. Because the knowledge of proteins adsorbed onto vector surface could be useful in the screening of new, more efficient and more biocompatible liposomal formulations, the behavior of three CLs with different membrane charge densities was investigated. The proteins of the three coronas were identified by nano-liquid chromatography-tandem mass spectrometry, and quantified with label-free spectral counting strategy. Fibrinogen displayed higher association with CLs with high membrane charge density, while apolipoproteins and C4b-binding protein with CLs with low membrane charge density. These results are discussed in terms of the different lipid compositions of CLs and may have a deep biological impact for in vivo applications. Surface charge of nanoparticles is emerging as a relevant factor determining the corona composition after interaction with plasma proteins. Remarkably, it is also shown that the charge of the protein corona formed around CLs is strongly related to their membrane charge density. © 2012 Elsevier B.V.

  7. Markers of oxidative/nitrative damage of plasma proteins correlated with EDSS and BDI scores in patients with secondary progressive multiple sclerosis.

    Science.gov (United States)

    Morel, Agnieszka; Bijak, Michał; Niwald, Marta; Miller, Elżbieta; Saluk, Joanna

    2017-05-19

    Objectives The objective of the present study was to evaluate oxidative/nitrative stress in the plasma of 50 patients suffering from the secondary progressive course of multiple sclerosis (MS), and to verify its correlation with physical and mental disability as assessed by the Expanded Disability Status Scale (EDSS), and the Beck Depression Inventory (BDI). Methods Oxidative and nitrative damage to proteins was determined by the level of carbonyl groups and 3-nitrotyrosine using ELISA test. Based on the reaction with Ellman's reagent, we estimated the concentration of oxidized thiol groups. Additionally, we measured the level of lipid peroxidation. Results In plasma drawn from MS patients, we observed a significantly higher level of 3-NT (92%; P EDSS and BDI. Negative correlations were observed between concentration of -SH groups and EDSS and BDI. Conclusion Our results indicate that impaired red-ox balance can significantly promote neurodegeneration in secondary progressive MS.

  8. Dietary zinc depletion and repletion affects plasma proteins: an analysis of the plasma proteome.

    Science.gov (United States)

    Grider, Arthur; Wickwire, Kathie; Ho, Emily; Chung, Carolyn S; King, Janet

    2013-02-01

    Zinc (Zn) deficiency is a problem world-wide. Current methods for assessing Zn status are limited to measuring plasma or serum Zn within populations suspected of deficiency. Despite the high prevalence of Zn deficiency in the human population there are no methods currently available for sensitively assessing Zn status among individuals. The purpose of this research was to utilize a proteomic approach using two-dimensional gel electrophoresis (2DE) and mass spectrometry to identify protein biomarkers that were sensitive to changes in dietary Zn levels in humans. Proteomic analysis was performed in human plasma samples (n = 6) obtained from healthy adult male subjects that completed a dietary Zn depletion/repletion protocol, current dietary zinc intake has a greater effect on fractional zinc absorption than does longer term zinc consumption in healthy adult men. Chung et al. (Am J Clin Nutr 87 (5):1224-1229, 2008). After a 13 day Zn acclimatization period where subjects consumed a Zn-adequate diet, the male subjects consumed a marginal Zn-depleted diet for 42 days followed by consumption of a Zn-repleted diet for 28 days. The samples at baseline, end of depletion and end of repletion were pre-fractionated through immuno-affinity columns to remove 14 highly abundant proteins, and each fraction separated by 2DE. Following staining by colloidal Coomassie blue and densitometric analysis, three proteins were identified by mass spectrometry as affected by changes in dietary Zn. Fibrin β and chain E, fragment double D were observed in the plasma protein fraction that remained bound to the immunoaffinity column. An unnamed protein that was related to immunoglobulins was observed in the immunodepleted plasma fraction. Fibrin β increased two-fold following the Zn depletion period and decreased to baseline values following the Zn repletion period; this protein may serve as a viable biomarker for Zn status in the future.

  9. Increased capillary permeability for plasma proteins in oral contraceptive users.

    Science.gov (United States)

    Tollan, A; Kvenild, K; Strand, H; Oian, P; Maltau, J M

    1992-05-01

    The transcapillary fluid balance was examined in eleven women before administration of a monophasic oral contraceptive (desogestrel 0.15 mg, ethinylestradiol 0.03 mg), and after three and six months of use. The interstitial colloid osmotic pressure was measured by the "wick" method, and the interstitial hydrostatic pressure by the "wick-in-needle" method in subcutaneous tissue on thorax and leg. During the six-month observation period, the following changes were observed: Plasma colloid osmotic pressure decreased (mean 1.8 mmHg, p = 0.047), as well as serum albumin (mean 5.1 g/l, p = 0.0006), total protein concentration (mean 2.8 g/l, p = 0.0006), hemoglobin (mean 0.5 g/dl, p = 0.014) and hematocrit (mean 1.8%, p = 0.047). Blood pressure and body weight remained unchanged, but foot volume showed a significant increase. The colloid osmotic pressure gradient (plasma-interstitium) was significantly reduced. The results indicate an increase in plasma volume in addition to an increased capillary permeability to plasma proteins during oral contraceptive use. We suggest that the observed changes in transcapillary fluid balance is caused by the estrogen component of the oral contraceptive pill.

  10. A Protein Extract from Chicken Reduces Plasma Homocysteine in Rats

    Directory of Open Access Journals (Sweden)

    Vegard Lysne

    2015-06-01

    Full Text Available The present study aimed to evaluate effects of a water-soluble protein fraction of chicken (CP, with a low methionine/glycine ratio, on plasma homocysteine and metabolites related to homocysteine metabolism. Male Wistar rats were fed either a control diet with 20% w/w casein as the protein source, or an experimental diet where 6, 14 or 20% w/w of the casein was replaced with the same amount of CP for four weeks. Rats fed CP had reduced plasma total homocysteine level and markedly increased levels of the choline pathway metabolites betaine, dimethylglycine, sarcosine, glycine and serine, as well as the transsulfuration pathway metabolites cystathionine and cysteine. Hepatic mRNA level of enzymes involved in homocysteine remethylation, methionine synthase and betaine-homocysteine S-methyltransferase, were unchanged, whereas cystathionine gamma-lyase of the transsulfuration pathway was increased in the CP treated rats. Plasma concentrations of vitamin B2, folate, cobalamin, and the B-6 catabolite pyridoxic acid were increased in the 20% CP-treated rats. In conclusion, the CP diet was associated with lower plasma homocysteine concentration and higher levels of serine, choline oxidation and transsulfuration metabolites compared to a casein diet. The status of related B-vitamins was also affected by CP.

  11. Expression of Recombinant Pregnancy-associated Plasma Protein-A

    Institute of Scientific and Technical Information of China (English)

    CHENG; Bin-yan; LI; Zi-ying; ZHANG; Xue-feng; LIU; Yi-bing

    2013-01-01

    Pregnancy-associated plasma protein-A(PAPP-A)is producted by the syntrophoblast tissue of the placenta and decidual cells.It belongs to macromolecular glycoprotein.PAPP-A is a sensitive serum marker of Down’s syndrome and has clinical valuable in the early identification of acute coronary syndrome(ACS).According to the structure of PAPP-A,PAPP-A DNA is divided into five segments(S1-S5)for

  12. Spectrophotometric and Refractometric Determination of Total Protein in Avian Plasma

    OpenAIRE

    2013-01-01

    The aim of this study was to compare the total protein values obtained in heparin plasma of chickens by a spectrophotometric technique (biuret method), and the values obtained on the same day in the same samples by refractometry. The results obtained by refractometry (average value 2.638±0.153g%) were higher than those obtained by the spectrophotometric method (average value 2.441±0.181g%). There was a low correlation (r = 0.6709) between the total protein values, determined with both methods...

  13. Effects of experimentally increased protein supply to postpartum dairy cows on plasma protein synthesis, rumen tissue proliferation, and immune homeostasis

    DEFF Research Database (Denmark)

    Larsen, Mogens; Røntved, Christine Maria; Theil, Peter Kappel

    2017-01-01

    enrichment in arterial plasma free Phe, total plasma proteins, and albumin after 3, 5, and 7 h of jugular ring[13C]Phe infusion. Plasma volume was determined at +4 and +29 DRTC by dilution of a [125I]BSA dose. Synthesis rate of tissue protein in biopsied rumen papillae was determined by measuring [13C...

  14. DNA damage by carbonyl stress in human skin cells

    Energy Technology Data Exchange (ETDEWEB)

    Roberts, Michael J.; Wondrak, Georg T.; Laurean, Daniel Cervantes; Jacobson, Myron K.; Jacobson, Elaine L

    2003-01-28

    Reactive carbonyl species (RCS) are potent mediators of cellular carbonyl stress originating from endogenous chemical processes such as lipid peroxidation and glycation. Skin deterioration as observed in photoaging and diabetes has been linked to accumulative protein damage from glycation, but the effects of carbonyl stress on skin cell genomic integrity are ill defined. In this study, the genotoxic effects of acute carbonyl stress on HaCaT keratinocytes and CF3 fibroblasts were assessed. Administration of the {alpha}-dicarbonyl compounds glyoxal and methylglyoxal as physiologically relevant RCS inhibited skin cell proliferation, led to intra-cellular protein glycation as evidenced by the accumulation of N{sup {epsilon}}-(carboxymethyl)-L-lysine (CML) in histones, and caused extensive DNA strand cleavage as assessed by the comet assay. These effects were prevented by treatment with the carbonyl scavenger D-penicillamine. Both glyoxal and methylglyoxal damaged DNA in intact cells. Glyoxal caused DNA strand breaks while methylglyoxal produced extensive DNA-protein cross-linking as evidenced by pronounced nuclear condensation and total suppression of comet formation. Glycation by glyoxal and methylglyoxal resulted in histone cross-linking in vitro and induced oxygen-dependent cleavage of plasmid DNA, which was partly suppressed by the hydroxyl scavenger mannitol. We suggest that a chemical mechanism of cellular DNA damage by carbonyl stress occurs in which histone glycoxidation is followed by reactive oxygen induced DNA stand breaks. The genotoxic potential of RCS in cultured skin cells and its suppression by a carbonyl scavenger as described in this study have implications for skin damage and carcinogenesis and its prevention by agents selective for carbonyl stress.

  15. Cancer associated proteins in blood plasma: Determining normal variation.

    Science.gov (United States)

    Stenemo, Markus; Teleman, Johan; Sjöström, Martin; Grubb, Gabriel; Malmström, Erik; Malmström, Johan; Niméus, Emma

    2016-07-01

    Protein biomarkers have the potential to improve diagnosis, stratification of patients into treatment cohorts, follow disease progression and treatment response. One distinct group of potential biomarkers comprises proteins which have been linked to cancer, known as cancer associated proteins (CAPs). We determined the normal variation of 86 CAPs in 72 individual plasma samples collected from ten individuals using SRM mass spectrometry. Samples were collected weekly during 5 weeks from ten volunteers and over one day at nine fixed time points from three volunteers. We determined the degree of the normal variation depending on interpersonal variation, variation due to time of day, and variation over weeks and observed that the variation dependent on the time of day appeared to be the most important. Subdivision of the proteins resulted in two predominant protein groups containing 21 proteins with relatively high variation in all three factors (day, week and individual), and 22 proteins with relatively low variation in all factors. We present a strategy for prioritizing biomarker candidates for future studies based on stratification over their normal variation and have made all data publicly available. Our findings can be used to improve selection of biomarker candidates in future studies and to determine which proteins are most suitable depending on study design.

  16. High throughput assay for evaluation of reactive carbonyl scavenging capacity

    Directory of Open Access Journals (Sweden)

    N. Vidal

    2014-01-01

    Full Text Available Many carbonyl species from either lipid peroxidation or glycoxidation are extremely reactive and can disrupt the function of proteins and enzymes. 4-hydroxynonenal and methylglyoxal are the most abundant and toxic lipid-derived reactive carbonyl species. The presence of these toxics leads to carbonyl stress and cause a significant amount of macromolecular damages in several diseases. Much evidence indicates trapping of reactive carbonyl intermediates may be a useful strategy for inhibiting or decreasing carbonyl stress-associated pathologies. There is no rapid and convenient analytical method available for the assessment of direct carbonyl scavenging capacity, and a very limited number of carbonyl scavengers have been identified to date, their therapeutic potential being highlighted only recently. In this context, we have developed a new and rapid sensitive fluorimetric method for the assessment of reactive carbonyl scavengers without involvement glycoxidation systems. Efficacy of various thiol- and non-thiol-carbonyl scavenger pharmacophores was tested both using this screening assay adapted to 96-well microplates and in cultured cells. The scavenging effects on the formation of Advanced Glycation End-product of Bovine Serum Albumin formed with methylglyoxal, 4-hydroxynonenal and glucose-glycated as molecular models were also examined. Low molecular mass thiols with an α-amino-β-mercaptoethane structure showed the highest degree of inhibitory activity toward both α,β-unsaturated aldehydes and dicarbonyls. Cysteine and cysteamine have the best scavenging ability toward methylglyoxal. WR-1065 which is currently approved for clinical use as a protective agent against radiation and renal toxicity was identified as the best inhibitor of 4-hydroxynonenal.

  17. Carbonyl Compounds in the Gas Phase of Cigarette Mainstream Smoke and Their Pharmacological Properties.

    Science.gov (United States)

    Horinouchi, Takahiro; Higashi, Tsunehito; Mazaki, Yuichi; Miwa, Soichi

    2016-01-01

    Cigarette mainstream smoke is composed of gas and tar phases and contains >4000 chemical constituents, including nicotine and tar. The substances in the gas phase but not in the tar phase can pass through the airway epithelial barrier, enter the systemic circulation via the pulmonary circulation, and increase systemic oxidative damage, leading to the development of cigarette smoking-related diseases such as atherosclerosis. Recently, we identified some stable carbonyl compounds, including acrolein (ACR) and methyl vinyl ketone (MVK), as major cytotoxic factors in nicotine- and tar-free cigarette smoke extract (CSE) of the gas phase. CSE, ACR, and MVK induce protein kinase C (PKC)-dependent activation of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) and subsequent generation of reactive oxygen species (ROS) via NOX, causing plasma membrane damage and cell apoptosis. CSE, ACR, and MVK also trigger carbonylation of PKC, which is an irreversible oxidative modification. Cell damage and PKC carbonylation in response to treatment with CSE, ACR, or MVK are abolished by thiol-containing antioxidants such as N-acetyl-L-cysteine and reduced glutathione. Thus pharmacological modulation of PKC and NOX activities and the trapping of ROS are potential strategies for the prevention of diseases related to cigarette smoking.

  18. QSAR Models for the Prediction of Plasma Protein Binding

    Directory of Open Access Journals (Sweden)

    Zeshan Amin

    2013-02-01

    Full Text Available Introduction: The prediction of plasma protein binding (ppb is of paramount importance in the pharmacokinetics characterization of drugs, as it causes significant changes in volume of distribution, clearance and drug half life. This study utilized Quantitative Structure – Activity Relationships (QSAR for the prediction of plasma protein binding. Methods: Protein binding values for 794 compounds were collated from literature. The data was partitioned into a training set of 662 compounds and an external validation set of 132 compounds. Physicochemical and molecular descriptors were calculated for each compound using ACD labs/logD, MOE (Chemical Computing Group and Symyx QSAR software packages. Several data mining tools were employed for the construction of models. These included stepwise regression analysis, Classification and Regression Trees (CART, Boosted trees and Random Forest. Results: Several predictive models were identified; however, one model in particular produced significantly superior prediction accuracy for the external validation set as measured using mean absolute error and correlation coefficient. The selected model was a boosted regression tree model which had the mean absolute error for training set of 13.25 and for validation set of 14.96. Conclusion: Plasma protein binding can be modeled using simple regression trees or multiple linear regressions with reasonable model accuracies. These interpretable models were able to identify the governing molecular factors for a high ppb that included hydrophobicity, van der Waals surface area parameters, and aromaticity. On the other hand, the more complicated ensemble method of boosted regression trees produced the most accurate ppb estimations for the external validation set.

  19. Interactions between plasma proteins and naturally occurring polyphenols.

    Science.gov (United States)

    Li, Min; Hagerman, Ann E

    2013-05-01

    The plant natural products known as polyphenols are found at micronutrient levels in fruits, vegetables, and plant-based beverages such as wine, tea, coffee and cocoa. Consumption of a fruit- and vegetable-rich diet, the "Mediterranean diet", has been epidemiologically related to health benefits especially for chronic diseases including diabetes, cardiovascular disease, and Alzheimer's disease. The abundance of polyphenols in plant-rich diets, and the potent bioactivities of polyphenols, provide indirect evidence for a role for polyphenols in maintaining good health. However, molecular mechanisms for therapeutic or preventative activity have not been demonstrated in vivo. We summarize the chemical classes of natural polyphenols, their bioactivities and bioavailability and metabolism. Because many polyphenols bind protein, we focus on the potential of protein binding to mediate the health-related effects of polyphenols. We discuss interactions with plasma proteins as the first target organ past the digestive tract for these orally-ingested compounds.

  20. 49 CFR 173.198 - Nickel carbonyl.

    Science.gov (United States)

    2010-10-01

    ... 49 Transportation 2 2010-10-01 2010-10-01 false Nickel carbonyl. 173.198 Section 173.198... Nickel carbonyl. (a) Nickel carbonyl must be packed in specification steel or nickel cylinders as prescribed for any compressed gas except acetylene. A cylinder used exclusively for nickel carbonyl may...

  1. Changes in total plasma content of electrolytes and proteins with maximal exercise.

    Science.gov (United States)

    Van Beaumont, W.; Strand, J. C.; Petrofsky, J. S.; Hipskind, S. G.; Greenleaf, J. E.

    1973-01-01

    To determine to what extent the increases in concentration of plasma proteins and electrolytes with short maximal work were a result of hemoconcentration, the changes in plasma volume and total content of the plasma constituents were simultaneously evaluated. The results obtained from six human subjects indicated that in comparison to preexercise values there was a net decrease in total content of plasma protein, sodium, and chloride in the first 2 min of the postexercise period, due primarily to a significant loss (13-15%) of plasma fluid. The total plasma potassium content was increased immediately after exercise but was significantly below the preexercise plasma content after 2 min of recovery.

  2. Serum Copper and Plasma Protein Status in Normal Pregnancy

    Directory of Open Access Journals (Sweden)

    Nushrat Noor, Nasim Jahan, Nayma Sultana

    2012-12-01

    Full Text Available AbstractBackground: Gradual alteration of serum copper and some plasma protein levels may occur with advancement of pregnancy, which is associated with increased maternal and infant morbidity and mortality.Objective: To observe serum copper and plasma protein levels in normal pregnant women of different trimesters in order to find out their nutritional status.Methods: This cross sectional study was carried out in the Department of Physiology, Sir Salimullah Medical College (SSMC, Dhaka, between 1st January 2010 and December 2010. Ninety normal pregnant women of different trimesters with age 20-30 years were included in the study group. They were selected from Out Patient Department of Obstetrics and Gynaecology, SSMC. Age matched 30 non-pregnant women were taken as control. Serum copper level was measured by Spectrophotometric method, serum total protein and albumin levels were estimated by standard method. Statistical analysis was done by one way ANOVA, Bonferroni and Pearson’s correlation coefficient test as applicable.Results: Serum Cu levels were significantly higher in all trimesters of pregnant women compared to control. Again, this value was significantly higher in 3rd trimester than that of in 1st and 2nd trimester and also in 2nd trimester than that of in 1st trimester. In addition, mean serum total protein level was significantly lower in 3rd trimester than control but no statistically significant difference was observed among different trimesters. Again, mean serum albumin level was significantly lower in 2nd and 3rd trimester than 1st trimester and control. In addition, serum Cu concentration showed significant positive correlation with different trimesters of gestation.Conclusion: This study reveals that hypercupremia along with hypoproteinemia occur in pregnant women from 1st to 3rd trimester of gestation. This gradual alteration of micro and macronutrients become more profound with advancement of pregnancy.

  3. Shuttling of G protein subunits between the plasma membrane and intracellular membranes.

    Science.gov (United States)

    Chisari, Mariangela; Saini, Deepak Kumar; Kalyanaraman, Vani; Gautam, Narasimhan

    2007-08-17

    Heterotrimeric G proteins (alphabetagamma) mediate the majority of signaling pathways in mammalian cells. It is long held that G protein function is localized to the plasma membrane. Here we examined the spatiotemporal dynamics of G protein localization using fluorescence recovery after photobleaching, fluorescence loss in photobleaching, and a photoswitchable fluorescent protein, Dronpa. Unexpectedly, G protein subunits shuttle rapidly (t1/2 bromopalmitate. Thus, contrary to present thought, G proteins do not reside permanently on the plasma membrane but are constantly testing the cytoplasmic surfaces of the plasma membrane and endomembranes to maintain G protein pools in intracellular membranes to establish direct communication between receptors and endomembranes.

  4. Effect of Addition of Concentrated Proteins and Seminal Plasma Low Molecular Weight Proteins in Freezing and Thawing of Equine Semen

    Directory of Open Access Journals (Sweden)

    Bruno Fagundes

    2011-07-01

    Full Text Available Difficulties in obtaining equine frozen semen with potential fertility are recognized. This study was designed to investigate the effect of seminal plasma on frozen/thawing of eight stallion semen from different breed using the following treatments: Seminal plasma with ten-fold concentrated proteins with molecular weight above 10 kDa on frozen extender; Part of seminal plasma with proteins under 10 kDa on frozen extender; Conventional freezing, using whole seminal plasma on frozen extender. Using the parameter of 30% of seminal motility post-thawing as index of good freezability, it was verified an increased percentage of stallions that presented good freezability when semen was frozen with seminal plasma containing ten-fold concentrated proteins with molecular weight above 10 kDa on frozen extender. These results, suggested the use of seminal plasma concentrated proteins from own stallion to freezing/thawing semen.

  5. Ionic protein-lipid interaction at the plasma membrane: what can the charge do?

    Science.gov (United States)

    Li, Lunyi; Shi, Xiaoshan; Guo, Xingdong; Li, Hua; Xu, Chenqi

    2014-03-01

    Phospholipids are the major components of cell membranes, but they have functional roles beyond forming lipid bilayers. In particular, acidic phospholipids form microdomains in the plasma membrane and can ionically interact with proteins via polybasic sequences, which can have functional consequences for the protein. The list of proteins regulated by ionic protein-lipid interaction has been quickly expanding, and now includes membrane proteins, cytoplasmic soluble proteins, and viral proteins. Here we review how acidic phospholipids in the plasma membrane regulate protein structure and function via ionic interactions, and how Ca(2+) regulates ionic protein-lipid interactions via direct and indirect mechanisms.

  6. Synthesis and in vivo evaluation of the putative breast cancer resistance protein inhibitor [{sup 11}C]methyl 4-((4-(2-(6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolin-2-yl)ethyl)phenyl) amino-carbonyl)-2-(quinoline-2-carbonylamino)benzoate

    Energy Technology Data Exchange (ETDEWEB)

    Mairinger, Severin [Department of Medicinal Chemistry, University of Vienna, 1090 Vienna (Austria); Molecular Medicine, AIT Austrian Institute of Technology GmbH, 2444 Seibersdorf (Austria); Department of Clinical Pharmacology, Medical University of Vienna, Vienna (Austria); Langer, Oliver, E-mail: oliver.langer@ait.ac.a [Molecular Medicine, AIT Austrian Institute of Technology GmbH, 2444 Seibersdorf (Austria); Department of Clinical Pharmacology, Medical University of Vienna, Vienna (Austria); Kuntner, Claudia; Wanek, Thomas [Molecular Medicine, AIT Austrian Institute of Technology GmbH, 2444 Seibersdorf (Austria); Bankstahl, Jens P.; Bankstahl, Marion [Department of Pharmacology, Toxicology and Pharmacy, University of Veterinary Medicine Hanover (Germany); Stanek, Johann [Department of Clinical Pharmacology, Medical University of Vienna, Vienna (Austria); Doerner, Bernd [Department of Medicinal Chemistry, University of Vienna, 1090 Vienna (Austria); Bauer, Florian [Department of Medicinal Chemistry, University of Vienna, 1090 Vienna (Austria); Department of Clinical Pharmacology, Medical University of Vienna, Vienna (Austria); Baumgartner, Christoph [2nd Neurological Department, General Hospital Hietzing with Neurological Center Rosenhuegel, Vienna (Austria); Loescher, Wolfgang [Department of Pharmacology, Toxicology and Pharmacy, University of Veterinary Medicine Hanover (Germany); Erker, Thomas, E-mail: thomas.erker@univie.ac.a [Department of Medicinal Chemistry, University of Vienna, 1090 Vienna (Austria); Mueller, Markus [Department of Clinical Pharmacology, Medical University of Vienna, Vienna (Austria)

    2010-07-15

    Introduction: The multidrug efflux transporter breast cancer resistance protein (BCRP) is highly expressed in the blood-brain barrier (BBB), where it limits brain entry of a broad range of endogenous and exogenous substrates. Methyl 4-((4-(2-(6,7-dimethoxy-1,2,3,4-tetrahydroisoquinolin-2-yl)ethyl)phenyl) amino-carbonyl)-2-(quinoline-2-carbonylamino)benzoate (1) is a recently discovered BCRP-selective inhibitor, which is structurally derived from the potent P-glycoprotein (P-gp) inhibitor tariquidar. The aim of this study was to develop a new PET tracer based on 1 to map BCRP expression levels in vivo. Methods: Compound 1 was labelled with {sup 11}C in its methyl ester function by reaction of the corresponding carboxylic acid 2 with [{sup 11}C]methyl triflate. Positron emission tomography (PET) imaging of [{sup 11}C]-1 was performed in wild-type, Mdr1a/b{sup (-/-)}, Bcrp1{sup (-/-)} and Mdr1a/b{sup (-/-)}Bcrp1{sup (-/-)} mice (n=3 per mouse type) and radiotracer metabolism was assessed in plasma and brain. Results: Brain-to-plasma ratios of unchanged [{sup 11}C]-1 were 4.8- and 10.3-fold higher in Mdr1a/b{sup (-/-)} and in Mdr1a/b{sup (-/-)}Bcrp1{sup (-/-)} mice, respectively, as compared to wild-type animals, but only modestly increased in Bcrp1{sup (-/-)} mice. [{sup 11}C]-1 was rapidly metabolized in vivo giving rise to a polar radiometabolite which was taken up into brain tissue. Conclusion: Our data suggest that [{sup 11}C]-1 preferably interacts with P-gp rather than BCRP at the murine BBB which questions its reported in vitro BCRP selectivity. Consequently, [{sup 11}C]-1 appears to be unsuitable as a PET tracer to map cerebral BCRP expression.

  7. Identification of calcium-binding proteins associated with the human sperm plasma membrane

    National Research Council Canada - National Science Library

    Naaby-Hansen, Soren; Diekman, Alan; Shetty, Jagathpala; Flickinger, Charles J; Westbrook, Anne; Herr, John C

    2010-01-01

    The precise composition of the human sperm plasma membrane, the molecular interactions that define domain specific functions, and the regulation of membrane associated proteins during the capacitation...

  8. Aberrant Glycosylation of Plasma Proteins in Severe Preeclampsia Promotes Monocyte Adhesion

    Science.gov (United States)

    Kazanjian, Avedis A.; Tinnemore, Deborah; Gafken, Philip R.; Ogata, Yuko; Napolitano, Peter G.; Stallings, Jonathan D.; Ippolito, Danielle L.

    2014-01-01

    Glycosylation of plasma proteins increases during pregnancy. Our objectives were to investigate an anti-inflammatory role of these proteins in normal pregnancies and determine whether aberrant protein glycosylation promotes monocyte adhesion in preeclampsia. Plasma was prospectively collected from nonpregnant controls and nulliparous patients in all 3 trimesters. Patients were divided into cohorts based on the applicable postpartum diagnosis. U937 monocytes were preconditioned with enzymatically deglycosylated plasma, and monocyte adhesion to endothelial cell monolayers was quantified by spectrophotometry. Plasma from nonpregnant controls, first trimester normotensives, and first trimester patients with mild preeclampsia inhibited monocyte–endothelial cell adhesion (P < .05), but plasma from first trimester patients with severe preeclampsia and second and third trimester normotensives did not. Deglycosylating plasma proteins significantly increased adhesion in all the cohorts. These results support a role of plasma glycoprotein interaction in monocyte–endothelial cell adhesion and could suggest a novel therapeutic target for severe preeclampsia. PMID:23757314

  9. Quantitative analysis of plasma proteins in whole blood-derived fresh frozen plasma prepared with three pathogen reduction technologies.

    Science.gov (United States)

    Larrea, Luis; Ortiz-de-Salazar, María-Isabel; Martínez, Patricia; Roig, Roberto

    2015-06-01

    Several plasma pathogen reduction technologies (PRT) are currently available. We evaluated three plasma PRT processes: Cerus Amotosalen (AM), Terumo BCT riboflavin (RB) and Macopharma methylene blue (MB). RB treatment resulted in the shortest overall processing time and in the smallest volume loss (1%) and MB treatment in the largest volume loss (8%). MB treatment retained the highest concentrations of factors II, VII, X, IX, Protein C, and Antithrombin and the AM products of factor V and XI. Each PRT process evaluated offered distinct advantages such as procedural simplicity and volume retention (RB) and overall plasma protein retention (MB).

  10. A Method to Site-Specifically Identify and Quantitate Carbonyl End Products of Protein Oxidation Using Oxidation-Dependent Element Coded Affinity Tags (O-ECAT) and NanoLiquid Chromatography Fourier Transform Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Lee, S; Young, N L; Whetstone, P A; Cheal, S M; Benner, W H; Lebrilla, C B; Meares, C F

    2005-08-25

    Protein oxidation is linked to cellular stress, aging, and disease. Protein oxidations that result in reactive species are of particular interest, since these reactive oxidation products may react with other proteins or biomolecules in an unmediated and irreversible fashion, providing a potential marker for a variety of disease mechanisms. We have developed a novel system to identify and quantitate, relative to other states, the sites of oxidation on a given protein. A specially designed Oxidation-dependent carbonyl-specific Element-Coded Affinity Mass Tag (O-ECAT), AOD, ((S)-2-(4-(2-aminooxy)-acetamido)-benzyl)-1, 4, 7, 10-tetraazacyclododecane-N, N', N'', N'''-tetraacetic acid, is used to covalently tag the residues of a protein oxidized to aldehyde or keto end products. After proteolysis, the resulting AOD-tagged peptides are affinity purified, and analyzed by nanoLC-FTICR-MS, which provides high specificity in extracting co-eluting AOD mass pairs with a unique mass difference and affords relative quantitation based on isotopic ratios. Using this methodology, we have mapped the surface oxidation sites on a model protein, recombinant human serum albumin (rHSA) in its native form (as purchased) and after FeEDTA oxidation. A variety of modified amino acid residues including lysine, arginine, proline, histidine, threonine, aspartic and glutamic acids, were found to be oxidized to aldehyde and keto end products. The sensitivity of this methodology is shown by the number of peptides identified, twenty peptides on the native protein and twenty-nine after surface oxidation using FeEDTA and ascorbate. All identified peptides map to the surface of the HSA crystal structure validating this method for identifying oxidized amino acids on protein surfaces. In relative quantitation experiments between FeEDTA oxidation and native protein oxidation, identified sites showed different relative propensities towards oxidation independent of amino acid

  11. Binding patterns of seminal plasma plasma proteins on bovine epididymal and ejaculated sperm membrane

    Directory of Open Access Journals (Sweden)

    C.E.A. Souza

    2011-06-01

    Full Text Available The present study was designed to investigate the topographical distribution of seminal plasma (SP proteins on epididymal and ejaculated bovine sperm. Using immunocytochemistry and confocal microscopy the binding patterns of bovine SP proteins BSP-A3, albumin, transferrin, prostaglandin D-synthase (PGDS and nucleobindin in ejaculated and cauda epididymal sperm from adult bulls were evaluated. Experiments were performed using sperm from 5 males. Data showed a positive signal, only detected for anti-PGDS, in the acrosomal cap of epididymal and ejaculated sperm. In ejaculated sperm, a very weak signal for nucleobindin 2 in the midpiece and equatorial regions was detected, using the anti-rat nucleobindin. BSP-A3 was detected on all sperm regions studied, with a more evidenced signal in acrosome and midpiece. However, no binding was detected for albumin or transferrin in neither epididymal nor ejaculated sperm. In conclusion, PGDS, BSP-A3 and nucleobindin interact directly with bovine sperm, with specific topographic distribution. These findings may add to the knowledge of how these proteins modulate sperm functions, thus providing fundamental support for studies designed to evaluate how they influence sperm functions.

  12. Characterization of auxin-binding proteins from zucchini plasma membrane

    Science.gov (United States)

    Hicks, G. R.; Rice, M. S.; Lomax, T. L.

    1993-01-01

    We have previously identified two auxin-binding polypeptides in plasma membrane (PM) preparations from zucchini (Cucurbita pepo L.) (Hicks et al. 1989, Proc. Natl. Acad. Sci. USA 86, 4948-4952). These polypeptides have molecular weights of 40 kDa and 42 kDa and label specifically with the photoaffinity auxin analog 5-N3-7-3H-IAA (azido-IAA). Azido-IAA permits both the covalent and radioactive tagging of auxin-binding proteins and has allowed us to characterize further the 40-kDa and 42-kDa polypeptides, including the nature of their attachment to the PM, their relationship to each other, and their potential function. The azido-IAA-labeled polypeptides remain in the pelleted membrane fraction following high-salt and detergent washes, which indicates a tight and possibly integral association with the PM. Two-dimensional electrophoresis of partially purified azido-IAA-labeled protein demonstrates that, in addition to the major isoforms of the 40-kDa and 42-kDa polypeptides, which possess isoelectric points (pIs) of 8.2 and 7.2, respectively, several less abundant isoforms that display unique pIs are apparent at both molecular masses. Tryptic and chymotryptic digestion of the auxin-binding proteins indicates that the 40-kDa and 42-kDa polypeptides are closely related or are modifications of the same polypeptide. Phase extraction with the nonionic detergent Triton X-114 results in partitioning of the azido-IAA-labeled polypeptides into the aqueous (hydrophilic) phase. This apparently paradoxical behavior is also exhibited by certain integral membrane proteins that aggregate to form channels. The results of gel filtration indicate that the auxin-binding proteins do indeed aggregate strongly and that the polypeptides associate to form a dimer or multimeric complex in vivo. These characteristics are consistent with the hypothesis that the 40-kDa and 42-kDa polypeptides are subunits of a multimeric integral membrane protein which has an auxin-binding site, and which may

  13. Identification of frog photoreceptor plasma and disk membrane proteins by radioiodination

    Energy Technology Data Exchange (ETDEWEB)

    Witt, P.L.; Bownds, M.D.

    1987-03-24

    Several functions have been identified for the plasma membrane of the rod outer segment, including control of light-dependent changes in sodium conductance and a sodium-calcium exchange mechanism. However, little is known about its constituent proteins. Intact rod outer segments substantially free of contaminants were prepared in the dark and purified on a density gradient of Percoll. Surface proteins were then labeled by lactoperoxidase-catalyzed radioiodination, and intact rod outer segments were reisolated. Membrane proteins were identified by polyacrylamide gel electrophoresis and autoradiography. The surface proteins labeled included rhodopsin, the major membrane protein, and 12 other proteins. To compare the protein composition of plasma membrane with that of the internal disk membrane, purified rod outer segments were lysed by hypotonic disruption or freeze-thawing, and plasma plus disk membranes were radioiodinated. In these membrane preparations, rhodopsin was the major iodinated constituent, with 12 other proteins also labeled. Autoradiographic evidence indicated some differences in protein composition between disk and plasma membranes. A quantitative comparison of the two samples showed that labeling of two proteins, 24 kilodaltons (kDa) and 13 kDa, was enriched in the plasma membrane, while labeling of a 220-kDa protein was enriched in the disk membrane. These plasma membrane proteins may be associated with important functions such as the light-sensitive conductance and the sodium-calcium exchanger.

  14. Inhibition of platelet (/sup 3/H)- imipramine binding by human plasma protein fractions

    Energy Technology Data Exchange (ETDEWEB)

    Strijewski, A.; Chudzik, J.; Tang, S.W.

    1988-01-01

    Inhibition of high-affinity (/sup 3/H)-imipramine binding to platelet membranes by human plasma fractions and isolated plasma proteins was investigated. Several plasma proteins were found to contribute to the observed apparent inhibition and this contribution was assessed in terms of inhibitor units. Alpha/sub 1/ acid glycoprotein, high density and low density lipoprotein, IgG and ..cap alpha../sub 1/-antitrypsin were identified as effective non-specific inhibitors. Alpha-1-acid glycoprotein was confirmed to be the most potent plasma protein inhibitor. Cohn fractions were evaluated for the presence of the postulated endocoid of (/sup 3/H)-imipramine binding site.

  15. Blood plasma proteins and protein fractions in roe deer Capreolus capreolus L.

    Directory of Open Access Journals (Sweden)

    Dorota CYGAN-SZCZEGIELNIAK

    2015-09-01

    Full Text Available The aim of the research was to investigate some selected biochemical blood parameters in roe deer (Capreolus capreolus L.. The experiment covered 15 from 2 to 3-year-old bucks from Kuyavian-Pomeranian Voivodeship. The animals were shot by individual hunters on the shooting grounds during the hunting season of 2008/2009 (in the accordance with the Journal of Laws No 48. The material for the research was blood plasma obtained after centrifuging full, nonhemolyzed blood. The blood was collected from the zygomatic vein directly to the test tubes with EDTA and transported in cooling conditions to the laboratory. After transporting the samples of blood to a certified analytical laboratory, the following elements of the obtained blood plasma were examined: ceruloplasmin . using turbidimetric method; transferrin . using immunoturbimetric method; troponin- using a third generation assay on an Elecsys; total protein, albumin, globulin . using spectrophotometric method and total iron . using colorimetric method. The results were statistically analyzed, i.e. the correlation between the parameters was measured by means of Pearsonfs correlation coefficient. The analysis of the results revealed a number of statistically significant relations between the parameters under the investigation, especially among the compounds directly responsible for metabolism of iron and copper. A statistically important positive correlation was observed between ceruloplasmin and ferritin (r = 0.563; P.0.05 and a negative one between transferrin and troponin (r = -0.609; P.0.05. Moreover, the content of transferrin . an iron-binding protein . was 0.17 g/l, while the concentration of iron was 58 ƒĘmol/l. The content of ceruloplasmin . a protein responsible for metabolism of copper . was very low (0.036 g/l. The level of proteins in the blood plasma of the animals under the research was approximately 72 g/l, with the share of albumins about 46%. The albumin-globulin ratio was 0.86.

  16. Plasma levels of soluble endothelial cell protein C receptor in patients with Wegener's granulomatosis

    NARCIS (Netherlands)

    Boomsma, MM; Stearns-Kurosawa, DJ; Stegeman, CA; Raschi, E; Meroni, PL; Kurosawa, S; Tervaert, JWC

    Elevated soluble thrombomodulin (sTM) levels are an accepted marker of endothelial damage. The physiological significance of plasma endothelial protein C receptor (sEPCR) levels is not known. To assess the relevance of this plasma protein in Wegener's granulomatosis (WG), sEPCR levels were measured

  17. Interaction between La(III) and proteins on the plasma membrane of horseradish

    Science.gov (United States)

    Yang, Guang-Mei; Chu, Yun-Xia; Lv, Xiao-Fen; Zhou, Qing; Huang, Xiao-Hua

    2012-06-01

    Lanthanum (La) is an important rare earth element in the ecological environment of plant. The proteins on the plasma membrane control the transport of molecules into and out of cell. It is very important to investigate the effect of La(III) on the proteins on the plasma membrane in the plant cell. In the present work, the interaction between La(III) and proteins on the plasma membrane of horseradish was investigated using optimization of the fluorescence microscopy and fluorescence spectroscopy. It is found that the fluorescence of the complex system of protoplasts and 1-aniline Kenai-8-sulfonic acid in horseradish treated with the low concentration of La(III) is increased compared with that of the control horseradish. The opposite effect is observed in horseradish treated with the high concentration of La(III). These results indicated that the low concentration of La(III) can interact with the proteins on the plasma membrane of horseradish, causing the improvement in the structure of proteins on the plasma membrane. The high concentration of La(III) can also interact with the proteins on the plasma membrane of horseradish, leading to the destruction of the structure of proteins on the plasma membrane. We demonstrate that the proteins on the plasma membrane are the targets of La(III) action on plant cell.

  18. Estimation of protein content in the plasma of young chickens by a refractometric method.

    Science.gov (United States)

    Morgan, G W; Thaxton, P; Edens, F W

    1975-07-01

    This study was conducted to evaluate a refractometric method for determination of protein content of chicken plasma. Comparison of the results obtained with the refractometric and the Lowry methods indicated that refractometry, when used with due caution in a typical laboratory situation, provided a simple, fast, inexpensive and valid method for determining the protein content of plasma from young chickens.

  19. Multiplexed LC-MS/MS analysis of horse plasma proteins to study doping in sport.

    Science.gov (United States)

    Barton, Chris; Beck, Paul; Kay, Richard; Teale, Phil; Roberts, Jane

    2009-06-01

    The development of protein biomarkers for the indirect detection of doping in horse is a potential solution to doping threats such as gene and protein doping. A method for biomarker candidate discovery in horse plasma is presented using targeted analysis of proteotypic peptides from horse proteins. These peptides were first identified in a novel list of the abundant proteins in horse plasma. To monitor these peptides, an LC-MS/MS method using multiple reaction monitoring was developed to study the quantity of 49 proteins in horse plasma in a single run. The method was optimised and validated, and then applied to a population of race-horses to study protein variance within a population. The method was finally applied to longitudinal time courses of horse plasma collected after administration of an anabolic steroid to demonstrate utility for hypothesis-driven discovery of doping biomarker candidates.

  20. Separation of proteins from human plasma by sample displacement chromatography in hydrophobic interaction mode

    OpenAIRE

    Josic, Djuro; Breen, Lucas; Clifton, James; Gajdosik, Martina Srajer; Gaso-Sokac, Dajana; Rucevic, Marijana; Müller, Egbert

    2012-01-01

    Sample displacement chromatography (SDC) in reversed-phase and ion-exchange modes was introduced approximately twenty years ago. This method was first used for the preparative purification of peptides and proteins. Recently, SDC in ion-exchange mode was also successfully used for enrichment of low abundance proteins from human plasma. In this paper, the use of SDC for the separation of plasma proteins in hydrophobic interaction mode is demonstrated. By use of two or more columns coupled in se...

  1. Biofield-effect protein-sensor: Plasma functionalization of polyaniline, protein immobilization, and sensing mechanism

    Science.gov (United States)

    Cho, Chae-Ryong; Lee, Hyun-Uk; Ahn, Kyun; Jeong, Se-Young; Choi, Jun-Hee; Kim, Jinwoo; Cho, Jiung

    2014-06-01

    We report the fabrication of a biofield-effect protein-sensor (BioFEP) based on atmospheric-pressure plasma (AP) treatment of a conducting polyaniline (PANI) film. Successive H2 and O2 AP (OHAP) treatment generated dominant hydrophilic -OH and O=CO- functional groups on the PANI film surface, which served as strong binding sites to immobilize bovine serum albumin (BSA) protein molecules. The output current changes of the BioFEP as a function of BSA concentration were obtained. The resistance of the OHAP surface could be sensitively increased from 2.5 × 108 Ω to 2.0 × 1012 Ω with increasing BSA concentrations in the range of 0.025-4 μg/ml. The results suggest that the method is a simple and cost-effective tool to determine the concentration of BSA by measuring electrical resistance.

  2. Effect of anticoagulants and glucose on refractometric estimation of protein in canine and rabbit plasma.

    Science.gov (United States)

    Dubin, S; Hunt, P

    1978-10-01

    The effect of ethylenediaminetetraacetate (EDTA) compounds on the refractometric estimation of plasma protein concentration was attributed largely to osmotic fluid shifts, as reflected in changes in hematocrit, and to addition of total solids to the plasma. With H4EDTA, these two mechanisms were additive and caused increased plasma protein readings of significant magnitude even at recommended (1--2 mg/ml) anticoagulant concentrations. For the potassium and sodium salts, the two mechanisms were partly compensatory, which ameliorated the effect at 1--2 mg/ml concentration. At higher concentrations, such as might occur if a blood collecting tube were incompletely filled, all of the EDTA compounds caused technically significant over-estimation of plasma protein. When dextrose (d-glucose) was added in-vitro to canine blood, in amounts analogous to clinical hyperglycemia, the effect upon plasma protein estimation was minimal.

  3. A high confidence, manually validated human blood plasma protein reference set

    DEFF Research Database (Denmark)

    Schenk, Susann; Schoenhals, Gary J; de Souza, Gustavo

    2008-01-01

    sources, including the HUPO PPP dataset. CONCLUSION: Superior instrumentation combined with rigorous validation criteria gave rise to a set of 697 plasma proteins in which we have very high confidence, demonstrated by an exceptionally low false peptide identification rate of 0.29%.......BACKGROUND: The immense diagnostic potential of human plasma has prompted great interest and effort in cataloging its contents, exemplified by the Human Proteome Organization (HUPO) Plasma Proteome Project (PPP) pilot project. Due to challenges in obtaining a reliable blood plasma protein list...

  4. Measurement of carbonyl chemical shifts of excited protein states by relaxation dispersion NMR spectroscopy: comparison between uniformly and selectively {sup 13}C labeled samples

    Energy Technology Data Exchange (ETDEWEB)

    Lundstroem, Patrik; Hansen, D. Flemming; Kay, Lewis E. [University of Toronto, Department of Medical Genetics (Canada)], E-mail: kay@pound.med.utoronto.ca

    2008-09-15

    Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy has emerged as a powerful method for quantifying chemical shifts of excited protein states. For many applications of the technique that involve the measurement of relaxation rates of carbon magnetization it is necessary to prepare samples with isolated {sup 13}C spins so that experiments do not suffer from magnetization transfer between coupled carbon spins that would otherwise occur during the CPMG pulse train. In the case of {sup 13}CO experiments however the large separation between {sup 13}CO and {sup 13}C{sup {alpha}} chemical shifts offers hope that robust {sup 13}CO dispersion profiles can be recorded on uniformly {sup 13}C labeled samples, leading to the extraction of accurate {sup 13}CO chemical shifts of the invisible, excited state. Here we compare such chemical shifts recorded on samples that are selectively labeled, prepared using [1-{sup 13}C]-pyruvate and NaH{sup 13}CO{sub 3,} or uniformly labeled, generated from {sup 13}C-glucose. Very similar {sup 13}CO chemical shifts are obtained from analysis of CPMG experiments recorded on both samples, and comparison with chemical shifts measured using a second approach establishes that the shifts measured from relaxation dispersion are very accurate.

  5. Soluble Proteins Form Film by the Treatment of Low Temperature Plasma

    Science.gov (United States)

    Ikehara, Sanae; Sakakita, Hajime; Ishikawa, Kenji; Akimoto, Yoshihiro; Nakanishi, Hayao; Shimizu, Nobuyuki; Hori, Masaru; Ikehara, Yuzuru

    2015-09-01

    It has been pointed out that low temperature plasma in atmosphere was feasible to use for hemostasis without heat injury. Indeed, earlier studies demonstrated that low temperature plasma played an important role to stimulate platelets to aggregate and turned on the proteolytic activities of coagulation factors, resulting in the acceleration of the natural blood coagulation process. On the other hands, our developed equips could immediately form clots upon the contact with plasma flair, while the histological appearance was different from natural coagulation. Based on these findings in formed clots, we sought to determine if plasma flair supplied by our devices was capable of forming film using a series of soluble proteins Following plasma treatment, films were formed from bovine serum albumin, and the other plasma proteins at physiological concentration. Analysis of trans-electron microscope demonstrated that plasma treatment generated small protein particles and made them fuse to be larger aggregations The combined results demonstrated that plasma are capable of aggregating soluble proteins and that platelets and coagulation factors are not necessary for plasma induced blood coagulation. Supported in part by Grants-in-Aid for Scientific Research on Priority Area (21590454, 24590498, and 24108006 to Y. I.).

  6. Hypochlorite-induced damage to plasma and proteins: formation of nitrogen-centred radicals and their role in protein oxidation

    Energy Technology Data Exchange (ETDEWEB)

    Hawkins, C.L.; Davies, M.J. [Heart Research Institute, Camperdown, NSW (Australia)

    1998-12-31

    The respiratory burst of activated phagocyte cells results in the generation of hypochlorite (HOCl) via the release of the hydrogen peroxide and the enzyme myeloperoxidase. Little information is available about the mechanisms and intermediates involved in these reactions. Electron paramagnetic resonance (EPR) spectroscopy with spin trapping has been employed to identify radicals formed in fresh human plasma and isolated proteins and peptides on treatment with HOCI. Reaction of plasma with HOCI in the presence of a spin trap gives broad, anisotropic radical adducts consistent with the formation of large, slowly-tumbling, protein-derived radicals. The identity of the plasma-derived radical adducts was investigated further by the incubation of the pre-formed adducts with the non-specific proteolytic enzyme pronase. This treatment gave sharper, signals consistent with the release of more mobile, low-molecular-weight spin adducts from the initial protein-derived adducts. The hyperfine couplings of these sharper signals are characteristic of the formation of nitrogen-centred radical adducts. Similar or identical species are observed on treatment with isolated human serum albumin, suggesting that this is a major site of HOCI-induced oxidation. Reaction of HOCI-treated plasma or isolated proteins/peptides with excess methionine eliminates radical formation, consistent with lysine-derived chloramines (via homolysis or heterolysis of N-CI bonds) being the radical source. The effect of HOCI on the structural integrity of the plasma proteins was investigated by SDS-PAGE. It was demonstrated that incubation of HOCI-treated plasma or proteins, after removal of excess oxidant, resulted in a time- and HOCI-dependent fragmentation of the proteins. No evidence was obtained for the presence of either discrete fragments or aggregated material. This suggests that the reaction of HOCI with plasma proteins results in the formation of a large number of random fragments. Treatment with

  7. Meal composition and plasma amino acid ratios: Effect of various proteins or carbohydrates, and of various protein concentrations

    Science.gov (United States)

    Yokogoshi, Hidehiko; Wurtman, Richard J.

    1986-01-01

    The effects of meals containing various proteins and carbohydrates, and of those containing various proportions of protein (0 percent to 20 percent of a meal, by weight) or of carbohydrate (0 percent to 75 percent), on plasma levels of certain large neutral amino acids (LNAA) in rats previously fasted for 19 hours were examined. Also the plasma tryptophan ratios (the ratio of the plasma trytophan concentration to the summed concentrations of the other large neutral amino acids) and other plasma amino acid ratios were calculated. (The plasma tryptophan ratio has been shown to determine brain tryptophan levels and, thereby, to affect the synthesis and release of the neurotransmitter serotonin). A meal containing 70 percent to 75 percent of an insulin-secreting carbohydrate (dextrose or dextrin) increased plasma insulin levels and the tryptophan ratio; those containing 0 percent or 25 percent carbohydrate failed to do so. Addition of as little as 5 percent casein to a 70 percent carbohydrate meal fully blocked the increase in the plasma tryptophan ratio without affecting the secretion of insulin - probably by contributing much larger quantities of the other LNAA than of tryptophan to the blood. Dietary proteins differed in their ability to suppress the carbohydrate-induced rise in the plasma tryptophan ratio. Addition of 10 percent casein, peanut meal, or gelatin fully blocked this increase, but lactalbumin failed to do so, and egg white did so only partially. (Consumption of the 10 percent gelatin meal also produced a major reduction in the plasma tyrosine ratio, and may thereby have affected brain tyrosine levels and catecholamine synthesis.) These observations suggest that serotonin-releasing neurons in brains of fasted rats are capable of distinguishing (by their metabolic effects) between meals poor in protein but rich in carbohydrates that elicit insulin secretion, and all other meals. The changes in brain serotonin caused by carbohydrate-rich, protein

  8. Meal composition and plasma amino acid ratios: Effect of various proteins or carbohydrates, and of various protein concentrations

    Science.gov (United States)

    Yokogoshi, Hidehiko; Wurtman, Richard J.

    1986-01-01

    The effects of meals containing various proteins and carbohydrates, and of those containing various proportions of protein (0 percent to 20 percent of a meal, by weight) or of carbohydrate (0 percent to 75 percent), on plasma levels of certain large neutral amino acids (LNAA) in rats previously fasted for 19 hours were examined. Also the plasma tryptophan ratios (the ratio of the plasma trytophan concentration to the summed concentrations of the other large neutral amino acids) and other plasma amino acid ratios were calculated. (The plasma tryptophan ratio has been shown to determine brain tryptophan levels and, thereby, to affect the synthesis and release of the neurotransmitter serotonin). A meal containing 70 percent to 75 percent of an insulin-secreting carbohydrate (dextrose or dextrin) increased plasma insulin levels and the tryptophan ratio; those containing 0 percent or 25 percent carbohydrate failed to do so. Addition of as little as 5 percent casein to a 70 percent carbohydrate meal fully blocked the increase in the plasma tryptophan ratio without affecting the secretion of insulin - probably by contributing much larger quantities of the other LNAA than of tryptophan to the blood. Dietary proteins differed in their ability to suppress the carbohydrate-induced rise in the plasma tryptophan ratio. Addition of 10 percent casein, peanut meal, or gelatin fully blocked this increase, but lactalbumin failed to do so, and egg white did so only partially. (Consumption of the 10 percent gelatin meal also produced a major reduction in the plasma tyrosine ratio, and may thereby have affected brain tyrosine levels and catecholamine synthesis.) These observations suggest that serotonin-releasing neurons in brains of fasted rats are capable of distinguishing (by their metabolic effects) between meals poor in protein but rich in carbohydrates that elicit insulin secretion, and all other meals. The changes in brain serotonin caused by carbohydrate-rich, protein

  9. Depletion of abundant plasma proteins by poly(N-isopropylacrylamide-acrylic acid) hydrogel particles

    DEFF Research Database (Denmark)

    Such-Sanmartín, Gerard; Ventura-Espejo, Estela; Jensen, Ole N

    2014-01-01

    at higher efficiency than low abundance proteins, which are enriched in the supernatants, whereas (2) hydrogel particles incubated with high concentrations of plasma capture and irreversibly trap abundant proteins. During the elution step, irreversibly trapped proteins remain captured while low abundance...... (SRM) liquid chromatography (LC)-MS/MS. This novel use of hydrogel particles opens new perspectives for biomarker analysis based on mass spectrometry....

  10. Seldi-tof MS Profiling of Plasma Proteins in Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Shao-Pai Wu

    2006-03-01

    Conclusion: This study clearly demonstrates that the combined technology of SELDI-TOF MS and artificial intelligence is effective in distinguishing protein expression between normal and ovarian cancer plasma. The identified protein peaks may be candidate proteins for early detection of ovarian cancer or evaluation of therapeutic response.

  11. Kinetics of the Double Carbonylation of Benzylchloride

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    It is a multi-phase-catalyzed reaction to produce calcium phenylpyruvate by double carbonylation of benzylchloride. Based on the analysis of the reaction mechanism, a kinetic model of the carbonylation reaction was obtained. The model was verified through experiments in which the diffusion effect was neglected with the appropriate operation manner. But it is inevitable that the carbonylation process is controlled by diffusion as the autoclave scaling up.

  12. HIP2: An online database of human plasma proteins from healthy individuals

    Directory of Open Access Journals (Sweden)

    Shen Changyu

    2008-04-01

    Full Text Available Abstract Background With the introduction of increasingly powerful mass spectrometry (MS techniques for clinical research, several recent large-scale MS proteomics studies have sought to characterize the entire human plasma proteome with a general objective for identifying thousands of proteins leaked from tissues in the circulating blood. Understanding the basic constituents, diversity, and variability of the human plasma proteome is essential to the development of sensitive molecular diagnosis and treatment monitoring solutions for future biomedical applications. Biomedical researchers today, however, do not have an integrated online resource in which they can search for plasma proteins collected from different mass spectrometry platforms, experimental protocols, and search software for healthy individuals. The lack of such a resource for comparisons has made it difficult to interpret proteomics profile changes in patients' plasma and to design protein biomarker discovery experiments. Description To aid future protein biomarker studies of disease and health from human plasma, we developed an online database, HIP2 (Healthy Human Individual's Integrated Plasma Proteome. The current version contains 12,787 protein entries linked to 86,831 peptide entries identified using different MS platforms. Conclusion This web-based database will be useful to biomedical researchers involved in biomarker discovery research. This database has been developed to be the comprehensive collection of healthy human plasma proteins, and has protein data captured in a relational database schema built to contain mappings of supporting peptide evidence from several high-quality and high-throughput mass-spectrometry (MS experimental data sets. Users can search for plasma protein/peptide annotations, peptide/protein alignments, and experimental/sample conditions with options for filter-based retrieval to achieve greater analytical power for discovery and validation.

  13. Transition metal catalyzed carbonylation reactions carbonylative activation of C-X bonds

    CERN Document Server

    Beller, Matthias

    2014-01-01

    This book provides students and researchers in organic synthesis with a detailed discussion of carbonylation from the basics through to applications. It discusses the past, present and future of carbonylation reactions.

  14. Purification of Pregnancy-associated Plasma Protein-A and Preparation of Its Antibodies

    Institute of Scientific and Technical Information of China (English)

    CHENG; Bin-yan; LI; Zi-ying; YUAN; Zhi-gang; ZHANG; Xue-feng; LIU; Yi-bing

    2013-01-01

    Pregnancy-associated plasma protein-A(PAPP-A)is isolated from the plasma of pregnant women.It is producted by the syntrophoblast tissue of the placenta and decidual cells.PAPP-A belongs to macromolecular glycoprotein.As a sensitive serum marker,the decreased PAPP-A levels during the first

  15. Detection of boar sperm plasma membrane protein using Rhodamine 640; implications for cryobiology and physiology

    Science.gov (United States)

    Rhodamine 640 (R640) was used to detect changes in boar sperm plasma membrane protein (PMP) during cryopreservation; a poorly understood phenomenon. The protocol was adapted for boar sperm so that semen samples (n = 17) could be analyzed for PMP (R640 positive) and plasma membrane integrity (PMI; Y...

  16. Monitoring the native phosphorylation state of plasma membrane proteins from a single mouse cerebellum

    DEFF Research Database (Denmark)

    Schindler, J.; Ye, J. Y.; Jensen, Ole Nørregaard

    2013-01-01

    Neuronal processing in the cerebellum involves the phosphorylation and dephosphorylation of various plasma membrane proteins such as AMPA or NMDA receptors. Despite the importance of changes in phosphorylation pattern, no global phospho-proteome analysis has yet been performed. As plasma membrane...

  17. Early Diagnosis of Intestinal Ischemia Using Urinary and Plasma Fatty Acid Binding Proteins

    NARCIS (Netherlands)

    Thuijls, Geertje; van Wijck, Kim; Grootjans, Joep; Derikx, Joep P. M.; van Bijnen, Annemarie A.; Heineman, Erik; Dejong, Cornelis H. C.; Buurman, Wim A.; Poeze, Martijn

    2011-01-01

    Objective: This study aims at improving diagnosis of intestinal ischemia, by measuring plasma and urinary fatty acid binding protein (FABP) levels. Methods: Fifty consecutive patients suspected of intestinal ischemia were included and blood and urine were sampled at time of suspicion. Plasma and uri

  18. One-step isolation of plasma membrane proteins using magnetic beads with immobilized concanavalin A

    DEFF Research Database (Denmark)

    Lee, Yu-Chen; Block, Gregory; Chen, Huiwen;

    2008-01-01

    We have developed a simple method for isolating and purifying plasma membrane proteins from various cell types. This one-step affinity-chromatography method uses the property of the lectin concanavalin A (ConA) and the technique of magnetic bead separation to obtain highly purified plasma membran...

  19. Automation of plasma protein binding assay using rapid equilibrium dialysis device and Tecan workstation.

    Science.gov (United States)

    Ye, Zhengqi; Zetterberg, Craig; Gao, Hong

    2017-03-14

    Binding of drug molecules to plasma proteins is an important parameter in assessing drug ADME properties. Plasma protein binding (PPB) assays are routinely performed during drug discovery and development. A fully automated PPB assay was developed using rapid equilibrium dialysis (RED) device and Tecan workstation coupled to an automated incubator. The PPB assay was carried out in unsealed RED plates which allowed the assay to be fully automated. The plasma pH was maintained at 7.4 during the 6-h dialysis under 2% CO2 condition. The samples were extracted with acetonitrile and analyzed by liquid chromatography tandem mass spectrometry. The percent bound results of 10 commercial drugs in plasma protein binding were very similar between the automated and manual assays, and were comparable to literature values. The automated assay increases laboratory productivity and is applicable to high-throughput screening of drug protein binding in drug discovery.

  20. Proteomic profiling of human plasma exosomes identifies PPARgamma as an exosome-associated protein.

    Science.gov (United States)

    Looze, Christopher; Yui, David; Leung, Lester; Ingham, Matthew; Kaler, Maryann; Yao, Xianglan; Wu, Wells W; Shen, Rong-Fong; Daniels, Mathew P; Levine, Stewart J

    2009-01-16

    Exosomes are nanovesicles that are released from cells as a mechanism of cell-free intercellular communication. Only a limited number of proteins have been identified from the plasma exosome proteome. Here, we developed a multi-step fractionation scheme incorporating gel exclusion chromatography, rate zonal centrifugation through continuous sucrose gradients, and high-speed centrifugation to purify exosomes from human plasma. Exosome-associated proteins were separated by SDS-PAGE and 66 proteins were identified by LC-MS/MS, which included both cellular and extracellular proteins. Furthermore, we identified and characterized peroxisome proliferator-activated receptor-gamma (PPARgamma), a nuclear receptor that regulates adipocyte differentiation and proliferation, as well as immune and inflammatory cell functions, as a novel component of plasma-derived exosomes. Given the important role of exosomes as intercellular messengers, the discovery of PPARgamma as a component of human plasma exosomes identifies a potential new pathway for the paracrine transfer of nuclear receptors.

  1. Study of decomposing carbonyl slag

    Institute of Scientific and Technical Information of China (English)

    CHEN Ai-liang; SUN Pei-mei; ZHAO Zhong-wei; LI Hong-gui; CHEN Xing-yu

    2006-01-01

    A new technology was put forward to deal with the carbonyl slag at low acidity and low oxygen pressure in the kettle.With the orthogonal experiments for analyzing the sequence of four factors and some single factor experiments for the best conditions. The best conditions are used for extracting nickel, cobalt and copper and enriching precious metals: the cupric ion concentration is 5 g/L; and pH=6; the sulfur coefficient is 1.4; the oxygen pressure is 0.08 MPa; the time bubbling oxygen is 20 min;the ratio of liquid to solid is 8:1; the leaching time is 2 h; the heating time is 2.5 h. The leaching rates of nickel and cobalt are more than 98% and that of copper is above 97%. Nickel and cobalt can be separated efficiently from copper and precious metals from the carbonyl slag. Moreover, its leaching liquor has less copper. Nickel and cobalt can be reclaimed only once. During the whole process,the leaching rates of Au and Ag are more than 99.9%, while other precious metals are still in the residue without any loss.

  2. Plasma proteins production and excretion in diabetic nephropathy in ...

    African Journals Online (AJOL)

    Journal of African Association of Physiological Sciences ... Subjects, materials, and methods: Plasma albumin, and fibrinogen ... Results: A direct relationship was found between albuminuria and albumin concentration (r=0.59, p<0.05).

  3. Biochemical and cytogenetic studies of Poecilia from eastern México. I. Comparative microelectrophoresis of plasma proteins of seven species

    OpenAIRE

    Balsano, J. S.; Rasch, Ellen M.

    2016-01-01

    Over 2000 fisch plasmas from six species of Poecilia were collected from 33 populations in eastern Mexico and one from western Mexico. These plasmas were electrophoretically separated in 7.5% polyacrylamide gel which was stained for specific enzymes or total protein. Identiflcations of albumin band mobilities were verified by mixing isoaliquots of test plasmas with plasmas of known standards and by comparing test plasmas with plasmas from F1 hybrid progreny of known parentage.In the latipinna...

  4. Spectrophotometric determination of total proteins in blood plasma: a comparative study among dye-binding methods

    OpenAIRE

    Dimas Augusto Morozin Zaia; Fábio Rangel Marques; Cássia Thaïs Bussamra Vieira Zaia

    2005-01-01

    A comparative study between the biuret method (standard method for total proteins) and spectrophotometric methods using dyes (Bradford, 3',3",5',5"-tetrabromophenolphthalein ethyl ester-TBPEE, and erythrosin-B) was carried out for the determination of total proteins in blood plasma from rats. Bradford method showed the highest sensitivity for proteins and biuret method showed the lowest. For all the methods, the absorbance for different proteins (BSA, casein, and egg albumin) was measured and...

  5. Nanoparticle size matters in the formation of plasma protein coronas on Fe3O4 nanoparticles.

    Science.gov (United States)

    Hu, Zhengyan; Zhang, Hongyan; Zhang, Yi; Wu, Ren'an; Zou, Hanfa

    2014-09-01

    When nanoparticles (NPs) enter into biological systems, proteins would interact with NPs to form the protein corona that can critically impact the biological identity of the nanomaterial. Owing to their fundamental scientific interest and potential applications, Fe3O4 NPs of different sizes have been developed for applications in cell separation and protein separation and as contrast agents in magnetic resonance imaging (MRI), etc. Here, we investigated whether nanoparticle size affects the formation of protein coronas around Fe3O4 NPs. Both the identification and quantification results demonstrated that particle size does play an important role in the formation of plasma protein coronas on Fe3O4 NPs; it not only influenced the protein composition of the formed plasma protein corona but also affected the abundances of the plasma proteins within the coronas. Understanding the different binding profiles of human plasma proteins on Fe3O4 NPs of different sizes would facilitate the exploration of the bio-distributions and biological fates of Fe3O4 NPs in biological systems.

  6. The role of cholesteryl ester transfer protein and phospholipid transfer protein in the remodeling of plasma high-density lipoproteins.

    Science.gov (United States)

    Lagrost, L

    1997-08-01

    Recent studies demonstrated that alterations in the size distribution of high-density lipoproteins (HDLs) constitute reliable markers for the risk of coronary artery disease. These observations suggested that the determination of the size distribution of HDL subpopulations by using polyacrylamide gradient gel electrophoresis might constitute an effective tool in clinical practice for the detection of patients with elevated risk. During the last decade, concordant observations revealed that all the HDL subpopulations are metabolically interrelated, and their relative abundances are dependent on the activity of several plasma factors, among them the cholesteryl ester transfer protein (CETP) and the phospholipid transfer protein (PLTP). As reviewed in the present article, although both CETP and PLTP can promote the size redistribution or conversion of HDL, the two plasma lipid transfer proteins can alter differently the plasma HDL distribution profile through distinct mechanisms. (Trends Cardiovasc Med 1997;7:218-224). © 1997, Elsevier Science Inc.

  7. Platelet adhesion onto wettability gradient surfaces in the absence and presence of plasma proteins.

    Science.gov (United States)

    Lee, J H; Lee, H B

    1998-08-01

    A wettability gradient was prepared on lowdensity polyethylene (PE) sheets by treating them in air with a corona from a knife-type electrode the power of which increased gradually along the sample length. The PE surfaces oxidized gradually with the increasing corona power and a wettability gradient was created on the surfaces, as evidenced by the measurement of water contact angles, Fourier transform infrared spectroscopy in the attenuated total reflectance mode, and electron spectroscopy for chemical analysis. The wettability gradient surfaces prepared were used to investigate the adhesion behavior of platelets in the absence and presence of plasma proteins in terms of the surface hydrophilicity/hydrophobicity of polymeric materials. The platelets adhered to the wettability gradient surfaces along the sample length were counted and examined by scanning electron microscopy (SEM). It was observed that the platelet adhesion in the absence of plasma proteins increased gradually as the surface wettability increased along the sample length. The platelets adhered to the hydrophilic positions of the gradient surface also were more activated (possessed more pseudo pods as examined by SEM) than on the more hydrophobic ones. However, platelet adhesion in the presence of plasma proteins decreased gradually with the increasing surface wettability; the platelets adhered to the surface also were more activated on the hydrophobic positions of the gradient surface. This result is closely related to plasma protein adsorption on the surface. Plasma protein adsorption on the wettability gradient surface increased with the increasing surface wettability. More plasma protein adsorption on the hydrophilic positions of the gradient surface caused less platelet adhesion, probably due to platelet adhesion inhibiting proteins, such as high-molecular-weight kininogen, which preferably adsorbs onto the surface by the so-called Vroman effect. It seems that both the presence of plasma proteins

  8. Temperature-Induced Protein Conformational Changes in Barley Root Plasma Membrane-Enriched Microsomes: II. Intrinsic Protein Fluorescence.

    Science.gov (United States)

    Caldwell, C R

    1987-07-01

    The membrane-bound proteins of barley (Hordeum vulgare L. cv Conquest) root plasma membrane-enriched microsomes displayed fluorescence typical of protein-associated trytophan residues. The protein fluorescence intensity was sensitive to variations in sample temperature. The temperature-induced decline in protein fluorescence intensity was nonlinear with slope discontinuities at about 12 and 32 degrees C. Detergents at levels above their critical micelle concentration enhanced protein fluorescence. Glutaraldehyde reduced protein fluorescence. Protein fluorescence polarization increased at temperatures above 30 degrees C. Both the rate of tryptophan photoionization and the fluorescence intensity of the photoionization products suggested alterations in membrane protein conformation between 12 and 32 degrees C. The quenching of the intrinsic protein fluorescence by acrylamide and potassium iodide indicated changes in accessibility of the extrinsic agents to the protein tryptophan residues beginning at about 14 degrees C. The results indicate thermally induced changes in the dynamics of the membrane proteins over the temperature range of 12 to 32 degrees C which could account for the complex temperature dependence of the barley root plasma membrane ATPase.

  9. Direct protein introduction into plant cells using a multi-gas plasma jet.

    Science.gov (United States)

    Yanagawa, Yuki; Kawano, Hiroaki; Kobayashi, Tomohiro; Miyahara, Hidekazu; Okino, Akitoshi; Mitsuhara, Ichiro

    2017-01-01

    Protein introduction into cells is more difficult in plants than in mammalian cells, although it was reported that protein introduction was successful in shoot apical meristem and leaves only together with a cell-penetrating peptide. In this study, we tried to introduce superfolder green fluorescent protein (sGFP)-fused to adenylate cyclase as a reporter protein without a cell-penetrating peptide into the cells of tobacco leaves by treatment with atmospheric non-thermal plasmas. For this purpose, CO2 or N2 plasma was generated using a multi-gas plasma jet. Confocal microscopy indicated that sGFP signals were observed inside of leaf cells after treatment with CO2 or N2 plasma without substantial damage. In addition, the amount of cyclic adenosine monophosphate (cAMP) formed by the catalytic enzyme adenylate cyclase, which requires cellular calmodulin for its activity, was significantly increased in leaves treated with CO2 or N2 plasma, also indicating the introduction of sGFP-fused adenylate cyclase into the cells. These results suggested that treatment with CO2 or N2 plasma could be a useful technique for protein introduction into plant tissues.

  10. The carbonyl scavenger carnosine ameliorates dyslipidaemia and renal function in Zucker obese rats

    Science.gov (United States)

    Aldini, Giancarlo; Orioli, Marica; Rossoni, Giuseppe; Savi, Federica; Braidotti, Paola; Vistoli, Giulio; Yeum, Kyung-Jin; Negrisoli, Gianpaolo; Carini, Marina

    2011-01-01

    Abstract The metabolic syndrome is a risk factor that increases the risk for development of renal and vascular complications. This study addresses the effects of chronic administration of the endogenous dipeptide carnosine (β-alanyl-L-histidine, L-CAR) and of its enantiomer (β-alanyl-D-histidine, D-CAR) on hyperlipidaemia, hypertension, advanced glycation end products, advanced lipoxidation end products formation and development of nephropathy in the non-diabetic, Zucker obese rat. The Zucker rats received a daily dose of L-CAR or D-CAR (30 mg/kg in drinking water) for 24 weeks. Systolic blood pressure was recorded monthly. At the end of the treatment, plasma levels of triglycerides, total cholesterol, glucose, insulin, creatinine and urinary levels of total protein, albumin and creatinine were measured. Several indices of oxidative/carbonyl stress were also measured in plasma, urine and renal tissue. We found that both L- and D-CAR greatly reduced obese-related diseases in obese Zucker rat, by significantly restraining the development of dyslipidaemia, hypertension and renal injury, as demonstrated by both urinary parameters and electron microscopy examinations of renal tissue. Because the protective effect elicited by L- and D-CAR was almost superimposable, we conclude that the pharmacological action of L-CAR is not due to a pro-histaminic effect (D-CAR is not a precursor of histidine, since it is stable to peptidic hydrolysis), and prompted us to propose that some of the biological effects can be mediated by a direct carbonyl quenching mechanism. PMID:20518851

  11. The carbonyl scavenger carnosine ameliorates dyslipidaemia and renal function in Zucker obese rats.

    Science.gov (United States)

    Aldini, Giancarlo; Orioli, Marica; Rossoni, Giuseppe; Savi, Federica; Braidotti, Paola; Vistoli, Giulio; Yeum, Kyung-Jin; Negrisoli, Gianpaolo; Carini, Marina

    2011-06-01

    The metabolic syndrome is a risk factor that increases the risk for development of renal and vascular complications. This study addresses the effects of chronic administration of the endogenous dipeptide carnosine (β-alanyl-L-histidine, L-CAR) and of its enantiomer (β-alanyl-D-histidine, D-CAR) on hyperlipidaemia, hypertension, advanced glycation end products, advanced lipoxidation end products formation and development of nephropathy in the non-diabetic, Zucker obese rat. The Zucker rats received a daily dose of L-CAR or D-CAR (30 mg/kg in drinking water) for 24 weeks. Systolic blood pressure was recorded monthly. At the end of the treatment, plasma levels of triglycerides, total cholesterol, glucose, insulin, creatinine and urinary levels of total protein, albumin and creatinine were measured. Several indices of oxidative/carbonyl stress were also measured in plasma, urine and renal tissue. We found that both L- and D-CAR greatly reduced obese-related diseases in obese Zucker rat, by significantly restraining the development of dyslipidaemia, hypertension and renal injury, as demonstrated by both urinary parameters and electron microscopy examinations of renal tissue. Because the protective effect elicited by L- and D-CAR was almost superimposable, we conclude that the pharmacological action of L-CAR is not due to a pro-histaminic effect (D-CAR is not a precursor of histidine, since it is stable to peptidic hydrolysis), and prompted us to propose that some of the biological effects can be mediated by a direct carbonyl quenching mechanism.

  12. Surface decorated platinum carbonyl clusters

    Science.gov (United States)

    Ciabatti, Iacopo; Femoni, Cristina; Iapalucci, Maria Carmela; Longoni, Giuliano; Zacchini, Stefano; Zarra, Salvatore

    2012-06-01

    Four molecular Pt-carbonyl clusters decorated by Cd-Br fragments, i.e., [Pt13(CO)12{Cd5(μ-Br)5Br2(dmf)3}2]2- (1), [Pt19(CO)17{Cd5(μ-Br)5Br3(Me2CO)2}{Cd5(μ-Br)5Br(Me2CO)4}]2- (2), [H2Pt26(CO)20(CdBr)12]8- (3) and [H4Pt26(CO)20(CdBr)12(PtBr)x]6- (4) (x = 0-2), have been obtained from the reactions between [Pt3n(CO)6n]2- (n = 2-6) and CdBr2.H2O in dmf at 120 °C. The structures of these molecular clusters with diameters of 1.5-2 nm have been determined by X-ray crystallography. Both 1 and 2 are composed of icosahedral or bis-icosahedral Pt-CO cores decorated on the surface by Cd-Br motifs, whereas 3 and 4 display a cubic close packed Pt26Cd12 metal frame decorated by CO and Br ligands. An oversimplified and unifying approach to interpret the electron count of these surface decorated platinum carbonyl clusters is suggested, and extended to other low-valent organometallic clusters and Au-thiolate nanoclusters.Four molecular Pt-carbonyl clusters decorated by Cd-Br fragments, i.e., [Pt13(CO)12{Cd5(μ-Br)5Br2(dmf)3}2]2- (1), [Pt19(CO)17{Cd5(μ-Br)5Br3(Me2CO)2}{Cd5(μ-Br)5Br(Me2CO)4}]2- (2), [H2Pt26(CO)20(CdBr)12]8- (3) and [H4Pt26(CO)20(CdBr)12(PtBr)x]6- (4) (x = 0-2), have been obtained from the reactions between [Pt3n(CO)6n]2- (n = 2-6) and CdBr2.H2O in dmf at 120 °C. The structures of these molecular clusters with diameters of 1.5-2 nm have been determined by X-ray crystallography. Both 1 and 2 are composed of icosahedral or bis-icosahedral Pt-CO cores decorated on the surface by Cd-Br motifs, whereas 3 and 4 display a cubic close packed Pt26Cd12 metal frame decorated by CO and Br ligands. An oversimplified and unifying approach to interpret the electron count of these surface decorated platinum carbonyl clusters is suggested, and extended to other low-valent organometallic clusters and Au-thiolate nanoclusters. CCDC 867747 and 867748. For crystallographic data in CIF or other electronic format see DOI: 10.1039/c2nr30400g

  13. A method for studies on interactions between a gold-based drug and plasma proteins based on capillary electrophoresis with inductively coupled plasma mass spectrometry detection

    DEFF Research Database (Denmark)

    Nguyen, Tam T T N; Østergaard, Jesper; Gammelgaard, Bente

    2015-01-01

    An analytical method based on capillary electrophoresis (CE) and inductively coupled plasma mass spectrometry (ICP-MS) detection was developed for studies on the interaction of gold-containing drugs and plasma proteins using auranofin as example. A detection limit of 18 ng/mL of auranofin...... was the major auranofin-interacting protein in plasma. The CE-ICP-MS method is proposed as a novel approach for kinetic studies of the interactions between gold-based drugs and plasma proteins. Graphical Abstract Development of a CE-ICP-MS based method allows for studies on interaction of the gold containing...

  14. A comprehensive analysis of the Streptococcus pyogenes and human plasma protein interaction network.

    Science.gov (United States)

    Sjöholm, Kristoffer; Karlsson, Christofer; Linder, Adam; Malmström, Johan

    2014-07-01

    Streptococcus pyogenes is a major human bacterial pathogen responsible for severe and invasive disease associated with high mortality rates. The bacterium interacts with several human blood plasma proteins and clarifying these interactions and their biological consequences will help to explain the progression from mild to severe infections. In this study, we used a combination of mass spectrometry (MS) based techniques to comprehensively quantify the components of the S. pyogenes-plasma protein interaction network. From an initial list of 181 interacting human plasma proteins defined using liquid chromatography (LC)-MS/MS analysis we further subdivided the interacting protein list using selected reaction monitoring (SRM) depending on the level of enrichment and protein concentration on the bacterial surface. The combination of MS methods revealed several previously characterized interactions between the S. pyogenes surface and human plasma along with many more, so far uncharacterised, possible plasma protein interactions with S. pyogenes. In follow-up experiments, the combination of MS techniques was applied to study differences in protein binding to a S. pyogenes wild type strain and an isogenic mutant lacking several important virulence factors, and a unique pair of invasive and non-invasive S. pyogenes isolates from the same patient. Comparing the plasma protein-binding properties of the wild type and the mutant and the invasive and non-invasive S. pyogenes bacteria revealed considerable differences, underlining the significance of these protein interactions. The results also demonstrate the power of the developed mass spectrometry method to investigate host-microbial relationships with a large proteomics depth and high quantitative accuracy.

  15. Effects of garcinol and guttiferone K isolated from Garcinia cambogia on oxidative/nitrative modifications in blood platelets and plasma.

    Science.gov (United States)

    Kolodziejczyk, Joanna; Masullo, Milena; Olas, Beata; Piacente, Sonia; Wachowicz, Barbara

    2009-11-01

    The effects of garcinol and guttiferone K, two polyisoprenylated benzophenones occurring a food plant called Garcinia cambogia, on oxidative/nitrative protein damage (determined by parameters such as levels of protein carbonyl groups and nitrotyrosine residues) in human blood platelets and plasma after treatment with peroxynitrite (ONOO(-)) were studied in vitro. We also investigated the effects of garcinol and guttiferone K on lipid peroxidation in blood platelets and plasma induced by ONOO(-) (100 microM). Exposure of blood platelets or plasma to peroxynitrite (100 microM) resulted in an increased level of carbonyl groups and nitrotyrosine residues in proteins, and an increase of lipid peroxidation measured by the level of thiobarbituric acid reactive species (TBARS). In the presence of garcinol and guttiferone K (0.1-25 microg/ml), a distinct reduction in the formation of carbonyl groups in plasma and platelet proteins together with the decrease of TBARS caused by 100 microM peroxynitrite, was observed. However, garcinol and guttiferone K did not inhibit plasma and platelet protein nitration induced by peroxynitrite. Polyisoprenylated benzophenones present in human diet such as garcinol or guttiferone K in vitro have protective effects against lipid and protein oxidation and may have some promising effects in vivo because they are good antioxidants in the tested models in vitro. Garcinol and guttiferone K can be also useful as protecting factors against diseases associated with oxidative stress.

  16. Isolation and characterization of gelatin-binding proteins from goat seminal plasma

    Directory of Open Access Journals (Sweden)

    Lazure Claude

    2003-04-01

    Full Text Available Abstract A family of proteins designated BSP-A1, BSP-A2, BSP-A3 and BSP-30 kDa (collectively called BSP proteins for Bovine Seminal Plasma proteins constitute the major protein fraction in the bull seminal plasma. These proteins interact with choline phospholipids on the sperm surface and play a role in the membrane stabilization (decapacitation and destabilization (capacitation process. Homologous proteins have been isolated from boar and stallion seminal plasma. In the current study we report the isolation and preliminary characterization of homologous proteins from goat seminal plasma. Frozen semen (-80°C was thawed and centrifuged to remove sperm. The proteins in the supernatant were precipitated by the addition of cold ethanol. The precipitates were dissolved in ammonium bicarbonate and lyophilised. The lyophilised proteins were dissolved in phosphate buffer and loaded onto a gelatin-agarose column, which was previously equilibrated with the same buffer. The column was successively washed with phosphate buffer, with phosphate buffer saline and with 0.5 M urea in phosphate buffer saline to remove unadsorbed proteins, and the adsorbed proteins were eluted with 5 M urea in phosphate buffer saline. Analysis of pooled, dialysed and lyophilised gelatin-agarose adsorbed protein fraction by SDS-PAGE indicated the presence of four protein bands that were designated GSP-14 kDa, GSP-15 kDa, GSP-20 kDa and GSP-22 kDa (GSP, Goat Seminal Plasma proteins. Heparin-affinity chromatography was then used for the separation of GSP-20 and -22 kDa from GSP-14 and -15 kDa. Finally, HPLC separation permitted further isolation of each one from the other. Amino acid sequence analysis of these proteins indicated that they are homologous to BSP proteins. In addition, these BSP homologs bind to hen's egg-yolk low-density lipoproteins. These results together with our previous data indicate that BSP family proteins are ubiquitous in mammalian seminal plasma, exist in

  17. Function of plasma membrane microdomain-associated proteins during legume nodulation.

    Science.gov (United States)

    Qiao, Zhenzhen; Libault, Marc

    2017-08-17

    Plasma membrane microdomains are plasma membrane sub-compartments enriched in sphingolipids and sterols, and composed by a specific set of proteins. They are involved in recognizing signal molecules, transducing these signals, and controlling endocytosis and exocytosis processes. In a recent study, applying biochemical and microscopic methods, we characterized the soybean GmFWL1 protein, a major regulator of soybean nodulation, as a new membrane microdomain-associated protein. Interestingly, upon rhizobia inoculation of the soybean root system, GmFWL1 and one of its interacting partners, GmFLOT2/4, both translocate to the root hair cell tip, the primary site of interaction and infection between soybean and Rhizobium. The role of GmFWL1 as a plasma membrane microdomain-associated protein is also supported by immunoprecipitation assays performed on soybean nodules, which revealed 178 GmFWL1 protein partners including a large number of microdomain-associated proteins such as GmFLOT2/4. In this addendum, we provide additional information about the identity of the soybean proteins repetitively identified as GmFWL1 protein partners. Their function is discussed especially in regard to plant-microbe interactions and microbial symbiosis. This addendum will provide new insights in the role of plasma membrane microdomains in regulating legume nodulation.

  18. Evaluation of the refractometric method for the determination of total protein in avian plasma or serum.

    Science.gov (United States)

    Lumeij, J T; de Bruijne, J J

    1985-07-01

    Serum total protein concentrations in pigeon blood determined with the biuret method (TPB-se) were compared with total protein concentrations in plasma (TPR-pl) and serum (TPR-se) obtained by estimation from refractive index. The refractometric method consistently yielded higher values (Prefractometric method for determination of TP in pigeon blood is not recommended.

  19. Interaction between a plasma membrane-localized ankyrin-repeat protein ITN1 and a nuclear protein RTV1

    Energy Technology Data Exchange (ETDEWEB)

    Sakamoto, Hikaru [Department of Bioproduction, Faculty of Bioindustry, Tokyo University of Agriculture, 196 Yasaka, Abashiri-shi, Hokkaido 093-2422 (Japan); Sakata, Keiko; Kusumi, Kensuke [Department of Biology, Faculty of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Kojima, Mikiko; Sakakibara, Hitoshi [RIKEN Plant Science Center, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045 (Japan); Iba, Koh, E-mail: koibascb@kyushu-u.org [Department of Biology, Faculty of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan)

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer ITN1, a plasma membrane ankyrin protein, interacts with a nuclear DNA-binding protein RTV1. Black-Right-Pointing-Pointer The nuclear transport of RTV1 is partially inhibited by interaction with ITN1. Black-Right-Pointing-Pointer RTV1 can promote the nuclear localization of ITN1. Black-Right-Pointing-Pointer Both overexpression of RTV1 and the lack of ITN1 increase salicylic acids sensitivity in plants. -- Abstract: The increased tolerance to NaCl 1 (ITN1) protein is a plasma membrane (PM)-localized protein involved in responses to NaCl stress in Arabidopsis. The predicted structure of ITN1 is composed of multiple transmembrane regions and an ankyrin-repeat domain that is known to mediate protein-protein interactions. To elucidate the molecular functions of ITN1, we searched for interacting partners using a yeast two-hybrid assay, and a nuclear-localized DNA-binding protein, RTV1, was identified as a candidate. Bimolecular fluorescence complementation analysis revealed that RTV1 interacted with ITN1 at the PM and nuclei in vivo. RTV1 tagged with red fluorescent protein localized to nuclei and ITN1 tagged with green fluorescent protein localized to PM; however, both proteins localized to both nuclei and the PM when co-expressed. These findings suggest that RTV1 and ITN1 regulate the subcellular localization of each other.

  20. Differential protein expression in seminal plasma from fertile and infertile males

    Science.gov (United States)

    Cadavid J, Angela P.; Alvarez, Angela; Markert, Udo R.; Maya, Walter Cardona

    2014-01-01

    AIM: The aim of this study was to analyze human seminal plasma proteins in association with male fertility status using the proteomic mass spectrometry technology Surface-Enhanced Laser Desorption Ionization Time-of-Flight (SELDI-TOF-MS). MATERIALS AND METHODS: Semen analysis was performed using conventional methods. Protein profiles of the seminal plasma were obtained by SELDI-TOF mass spectrometry over a strong anion exchanger, ProteinChip® Q10 array. RESULTS AND CONCLUSION: We found statistically significant differences in motility and sperm count between fertile and infertile men. In addition, we observed ten seminal proteins that are significantly up-regulated in the infertile group. In conclusion, comparison of seminal plasma proteome in fertile and infertile men provides new aspects in the physiology of male fertility and might help in identifying novel markers of male infertility. PMID:25395747

  1. Pathogen inactivation in fresh frozen plasma using riboflavin and ultraviolet light: Effects on plasma proteins and coagulation factor VIII

    Directory of Open Access Journals (Sweden)

    Stanojković Zoran

    2011-01-01

    Full Text Available Background/Aim. Riboflavin (vitamin B2 activated by ultraviolet (UV light, produces active oxygen which damages cell membrane and prevents replication of the carrier of diseases (viruses, bacteria, protozoa in all blood products. The aim of this study was to establish the influence of the process of photo inactivation in pathogens using riboflavin and UV rays on the concentration of coagulation factor VIII:C (FVIII:C and proteins in plasma that were treated before freezing. Methods. The examination included 20 units of plasma, separated from whole blood donated by voluntary blood donors around 6 hours from the moment of collection. The units were pooled and separated in to two groups: one consisted of 10 control units and the other of 10 experimental units. Experimental units of the plasma were treated by riboflavin (35 mL and UV rays (6.24 J/mL, 265-370 nm on Mirasol aparature (Caridian BCT Biotechnologies, USA in approximate duration of 6 minutes. Furthermore, 35 mL of saline solution was added to the control plasma. One sample for examining was taken from the control plasma (KG and two residual were taken from experimental plasma after the addition of riboflavin either before (EG1 or post illumination (EG2. Results. Comparing the mean values of FVIII:C (% we noticed statistically significantly higher level in the EG1 group than in the EG2 group (65.00 ± 4.52 vs 63.20 ± 4.73; t = 4.323, p = 0.002, while between the KG and experimental groups (EG1 and EG2 there was no statistically significant difference in the concentration of FVIII:C. There was a statistically significant decrease of albumin concentration (g/L in the EG2 group comparing to the KG (33.35 ± 0.94 vs 31.94 ± 0.84; t = 3.534, p = 0.002, but there was no mentioned difference in albumin concentration between the KG and the EG1, so as between the EG1 and the EG2. Conclusion. Plasma inactivated by riboflavin and UV rays (Mirasol PRT sistem, Caridian BCT, USA keeps all the

  2. Type 2 diabetes mellitus is associated with differential effects on plasma cholesteryl ester transfer protein and phospholipid transfer protein activities and concentrations

    NARCIS (Netherlands)

    Dullaart, RPF; De Vries, R; Scheek, L; Borggreve, SE; Van Gent, T; Dallinga-Thie, GM; Ito, M; Nagano, M; Sluiter, WJ; Hattori, H; Van Tol, A

    2004-01-01

    Background: Human plasma contains two lipid transfer proteins, cholesteryl ester transfer protein (CETP) and phospholipid transfer protein (PLTP), which are crucial in reverse cholesterol transport. Methods: Plasma CETP and PLTP activity levels and concentrations in 16 type 2 diabetic patients and 1

  3. RNAi-mediated downregulation of poplar plasma membrane intrinsic proteins (PIPs) changes plasma membrane proteome composition and affects leaf physiology.

    Science.gov (United States)

    Bi, Zhen; Merl-Pham, Juliane; Uehlein, Norbert; Zimmer, Ina; Mühlhans, Stefanie; Aichler, Michaela; Walch, Axel Karl; Kaldenhoff, Ralf; Palme, Klaus; Schnitzler, Jörg-Peter; Block, Katja

    2015-10-14

    Plasma membrane intrinsic proteins (PIPs) are one subfamily of aquaporins that mediate the transmembrane transport of water. To reveal their function in poplar, we generated transgenic poplar plants in which the translation of PIP genes was downregulated by RNA interference investigated these plants with a comprehensive leaf plasma membrane proteome and physiome analysis. First, inhibition of PIP synthesis strongly altered the leaf plasma membrane protein composition. Strikingly, several signaling components and transporters involved in the regulation of stomatal movement were differentially regulated in transgenic poplars. Furthermore, hormonal crosstalk related to abscisic acid, auxin and brassinosteroids was altered, in addition to cell wall biosynthesis/cutinization, the organization of cellular structures and membrane trafficking. A physiological analysis confirmed the proteomic results. The leaves had wider opened stomata and higher net CO2 assimilation and transpiration rates as well as greater mesophyll conductance for CO2 (gm) and leaf hydraulic conductance (Kleaf). Based on these results, we conclude that PIP proteins not only play essential roles in whole leaf water and CO2 flux but have important roles in the regulation of stomatal movement.

  4. One-step isolation of plasma membrane proteins using magnetic beads with immobilized concanavalin A1

    Science.gov (United States)

    Lee, Yu-Chen; Block, Gregory; Chen, Huiwen; Folch-Puy, Emma; Foronjy, Robert; Jalili, Roxana; Jendresen, Christian Bille; Kimura, Masashi; Kraft, Edward; Lindemose, Søren; Lu, Jin; McLain, Teri; Nutt, Leta; Ramon-Garcia, Santiago; Smith, Joseph; Spivak, Aaron; Wang, Michael L.; Zanic, Marija; Lin, Sue-Hwa

    2008-01-01

    We have developed a simple method for isolating and purifying plasma membrane proteins from various cell types. This one-step affinity-chromatography method uses the property of the lectin concanavalin A (ConA) and the technique of magnetic-bead separation to obtain highly purified plasma membrane proteins from crude membrane preparations or cell lines. ConA is immobilized onto magnetic beads by binding biotinylated ConA to streptavidin magnetic beads. When these ConA magnetic beads were used to enrich plasma membranes from a crude membrane preparation, this procedure resulted in 3.7-fold enrichment of plasma membrane marker 5′-nucleotidase activity with 70% recovery of the activity in the crude membrane fraction of rat liver. In agreement with the results of 5′-nucleotidase activity, immunoblotting with antibodies specific for a rat liver plasma membrane protein, CEACAM1, indicated that CEACAM1 was enriched about threefold relative to that of the original membranes. In similar experiments, this method produced 13-fold enrichment of 5′-nucleotidase activity with 45% recovery of the activity from a total cell lysate of PC-3 cells and 7.1-fold enrichment of 5′-nucleotidase activity with 33% recovery of the activity from a total cell lysate of HeLa cells. These results suggest that this one-step purification method can be used to isolate total plasma membrane proteins from tissue or cells for the identification of membrane biomarkers. PMID:18765283

  5. A highly stable nonbiofouling surface with well-packed grafted zwitterionic polysulfobetaine for plasma protein repulsion.

    Science.gov (United States)

    Chang, Yung; Liao, Shih-Chieh; Higuchi, Akon; Ruaan, Ruoh-Chyu; Chu, Chih-Wei; Chen, Wen-Yih

    2008-05-20

    An ideal nonbiofouling surface for biomedical applications requires both high-efficient antifouling characteristics in relation to biological components and long-term material stability from biological systems. In this study we demonstrate the performance and stability of an antifouling surface with grafted zwitterionic sulfobetaine methacrylate (SBMA). The SBMA was grafted from a bromide-covered gold surface via surface-initiated atom transfer radical polymerization to form well-packed polymer brushes. Plasma protein adsorption on poly(sulfobetaine methacrylate) (polySBMA) grafted surfaces was measured with a surface plasmon resonance sensor. It is revealed that an excellent stable nonbiofouling surface with grafted polySBMA can be performed with a cycling test of the adsorption of three model proteins in a wide range of various salt types, buffer compositions, solution pH levels, and temperatures. This work also demonstrates the adsorption of plasma proteins and the adhesion of platelets from human blood plasma on the polySBMA grafted surface. It was found that the polySBMA grafted surface effectively reduces the plasma protein adsorption from platelet-poor plasma solution to a level superior to that of adsorption on a surface terminated with tetra(ethylene glycol). The adhesion and activation of platelets from platelet-rich plasma solution were not observed on the polySBMA grafted surface. This work further concludes that a surface with good hemocompatibility can be achieved by the well-packed surface-grafted polySBMA brushes.

  6. Proteomic identification of novel differentiation plasma protein markers in hypobaric hypoxia-induced rat model.

    Directory of Open Access Journals (Sweden)

    Yasmin Ahmad

    Full Text Available BACKGROUND: Hypobaric hypoxia causes complex changes in the expression of genes, including stress related genes and corresponding proteins that are necessary to maintain homeostasis. Whereas most prior studies focused on single proteins, newer methods allowing the simultaneous study of many proteins could lead to a better understanding of complex and dynamic changes that occur during the hypobaric hypoxia. METHODS: In this study we investigated the temporal plasma protein alterations of rat induced by hypobaric hypoxia at a simulated altitude of 7620 m (25,000 ft, 282 mm Hg in a hypobaric chamber. Total plasma proteins collected at different time points (0, 6, 12 and 24 h, separated by two-dimensional electrophoresis (2-DE and identified using matrix assisted laser desorption ionization time of flight (MALDI-TOF/TOF. Biological processes that were enriched in the plasma proteins during hypobaric hypoxia were identified using Gene Ontology (GO analysis. According to their properties and obvious alterations during hypobaric hypoxia, changes of plasma concentrations of Ttr, Prdx-2, Gpx -3, Apo A-I, Hp, Apo-E, Fetub and Nme were selected to be validated by Western blot analysis. RESULTS: Bioinformatics analysis of 25 differentially expressed proteins showed that 23 had corresponding candidates in the database. The expression patterns of the eight selected proteins observed by Western blot were in agreement with 2-DE results, thus confirming the reliability of the proteomic analysis. Most of the proteins identified are related to cellular defense mechanisms involving anti-inflammatory and antioxidant activity. Their presence reflects the consequence of serial cascades initiated by hypobaric hypoxia. CONCLUSION/SIGNIFICANCE: This study provides information about the plasma proteome changes induced in response to hypobaric hypoxia and thus identification of the candidate proteins which can act as novel biomarkers.

  7. Proteomic Profiling of Nonenzymatically Glycated Proteins in Human Plasma and Erythrocyte Membrane

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Qibin; Tang, Ning; Schepmoes, Athena A.; Phillips, Lawrence S.; Smith, Richard D.; Metz, Thomas O.

    2008-05-01

    Non-enzymatic glycation of peptides and proteins by D-glucose has important implications in the pathogenesis of diabetes mellitus, particularly in the development of diabetic complications. In this report, a thorough proteomic profiling of glycated proteins was attempted by using phenylboronate affinity chromatography to enrich glycated proteins and glycated, tryptic peptides from human plasma and erythrocyte membranes. Enriched peptides were subsequently analyzed by liquid chromatography coupled with electron transfer dissociation tandem mass spectrometry, and 76 and 31 proteins were confidently identified as glycated from human plasma and erythrocyte membrane, respectively. It was observed that most of the glycated proteins can be identified in samples from individuals with normal glucose tolerance, although samples from individuals with impaired glucose tolerance and type 2 diabetes mellitus have slightly higher numbers of glycated proteins and more glycation sites identified.

  8. Direct Capture of Functional Proteins from Mammalian Plasma Membranes into Nanodiscs.

    Science.gov (United States)

    Roy, Jahnabi; Pondenis, Holly; Fan, Timothy M; Das, Aditi

    2015-10-20

    Mammalian plasma membrane proteins make up the largest class of drug targets yet are difficult to study in a cell free system because of their intransigent nature. Herein, we perform direct encapsulation of plasma membrane proteins derived from mammalian cells into a functional nanodisc library. Peptide fingerprinting was used to analyze the proteome of the incorporated proteins in nanodiscs and to further demonstrate that the lipid composition of the nanodiscs directly affects the class of protein that is incorporated. Furthermore, the functionality of the incorporated membrane proteome was evaluated by measuring the activity of membrane proteins: Na(+)/K(+)-ATPase and receptor tyrosine kinases. This work is the first report of the successful establishment and characterization of a cell free functional library of mammalian membrane proteins into nanodiscs.

  9. [carbonyl-C-11]desmethyl-WAY-100635 (DWAY) is a potent and selective radioligand for central 5-HT1A receptors in vitro and in vivo

    NARCIS (Netherlands)

    Pike, VW; Halldin, C; McCarron, JA; Lundkvist, C; Hirani, E; Olsson, H; Hume, SP; Karlsson, P; Osman, S; Swahn, CG; Hall, H; Wikstrom, H; Mensonidas, M; Poole, KG; Farde, L

    1998-01-01

    [carbonyl-C-11]Desmethyl-WAY-100635 (DWAY) is possibly a low-level metabolite appearing in plasma after intravenous administration of [carbonyl(11)C]WAY-100635 to human subjects for positron emission tomographic (PET) imaging of brain 5-HT1A receptors. In this study we set out to assess the ability

  10. Semen quality and concentration of soluble proteins in the seminal plasma of Alpine bucks Semen quality and concentration of soluble proteins in the seminal plasma of Alpine bucks

    Directory of Open Access Journals (Sweden)

    Simone Eliza Facione Guimarães

    2010-06-01

    Full Text Available It was aimed to study the in vitro seminal quality analyzed by complementary tests and to compare them with physical, morphological and biochemical aspects of male goat semen of the Alpine breed. This experiment took place at the Federal University of Viçosa, situated at 20º45’ S latitude and 42º51’ W longitude, Southwest of Brazil. It was done during the summer months of January and February, and three adult male goats of the Alpine breed were used in intensive conditions. The semen was collected by artificial vagina method. In all semen samples (45 ejaculates, after the physical and morphological analysis, the hiposmotic test was done. In 24 ejaculates, it were done thermo-resistance test, and in 21 ejaculates it were determined the concentration of total soluble proteins in seminal plasma. The male goats presented difference in the semen physical and morphological aspects, in the hiposmotic test and thermo-resistance test, but they did not presented difference in total soluble proteins concentration in seminal plasma. Results of the slow thermo-resistance test and hiposmotic test were positively correlated (r = 0.60. It was concluded, according to our results, that the concentration of total soluble proteins in seminal plasma can not be used as a parameter to predict the seminal quality of Alpine bucks.It was aimed to study the in vitro seminal quality analyzed by complementary tests and to compare them with physical, morphological and biochemical aspects of male goat semen of the Alpine breed. This experiment took place at the Federal University of Viçosa, situated at 20º45’ S latitude and 42º51’ W longitude, Southwest of Brazil. It was done during the summer months of January and February, and three adult male goats of the Alpine breed were used in intensive conditions. The semen was collected by artificial vagina method. In all semen samples (45 ejaculates, after the physical and morphological analysis, the hiposmotic test

  11. Effect of whey protein on plasma amino acids in diabetic mice.

    Science.gov (United States)

    Han, Ting; Cai, Donglian; Geng, Shanshan; Wang, Ying; Zhen, Hui; Wu, Peiying

    2013-12-01

    The aim of this study was to investigate the effect of whey protein on plasma amino acid levels in a mouse model of type II diabetes, using high-performance liquid chromatography (HPLC). The composition and content of amino acids in the whey proteins were analyzed using HPLC. Type I and type II diabetic mouse models were prepared using streptozotocin (STZ) and normal mice were used as a control. The ICR mice in each group were then randomly divided into four subgroups, to which 0, 10, 20 and 40% whey protein, respectively, was administered for four weeks. Changes in the plasma amino acid levels were observed in each group. The proportions of leucine, isoleucine and valine in the whey proteins were 14.40, 5.93 and 5.32% of the total amino acids, respectively, that is, the branched-chain amino acid content was 25.65%. The levels of branched-chain amino acids increased in the plasma of the normal and model mice following the administration of whey proteins by gavage and the amino acid levels increased as the concentration of the administered protein increased. In addition, the branched-chain amino acid levels in the blood of the model mice were higher than those in the normal mice. The levels of plasma amino acids in diabetic mice increased following gavage with whey protein, which is rich in branched-chain amino acids.

  12. A rapid and simple assay for growth hormone-binding protein activity in human plasma.

    Science.gov (United States)

    Baumann, G; Shaw, M A; Amburn, K

    1988-12-01

    The newly discovered circulating growth hormone binding proteins dictate a re-evaluation of the state of GH in plasma in health and disease as the binding proteins are known to affect GH metabolism and action. We describe a rapid and simple GH-binding assay that allows determination of free and complexed plasma GH, as well as GH-binding protein activity as an index of GH-binding protein levels, with relative ease. The method is based on incubation of plasma with 125I-GH and separation of bound from free GH on small DEAE-cellulose columns; it can be used on a large scale for routine determinations. The results obtained by this method are comparable to those obtained with the previously used slow and more cumbersome gel filtration technique. Initial data obtained in normal subjects and certain disease states show that the bound fraction of plasma GH is similar in men, women and children, is unaffected by pregnancy or acute infection, but is marginally decreased in liver cirrhosis. In acromegaly, binding protein activity also appears normal when allowance is made for partial saturation of the binding proteins by the high prevailing GH levels. The technique we describe should facilitate investigations of normal and abnormal regulation of the GH binding proteins.

  13. A pilot study of muscle plasma protein changes after exercise

    DEFF Research Database (Denmark)

    Dahlqvist, Julia R; Voss, Line G; Lauridsen, Thomas

    2014-01-01

    INTRODUCTION: Creatine kinase (CK) and myoglobin (Mb) do not possess all good qualities as biomarkers of skeletal muscle damage. We investigated the utility of troponin I (TnI) and telethonin (Tcap) as markers and examined their temporal profiles after skeletal muscle damage. METHODS: Plasma...... profiles were measured before and after exercise in 3 groups: subjects affected by either Becker muscular dystrophy or McArdle disease, and healthy subjects. RESULTS: Mb and TnI appeared early in the blood, and the increase of TnI was only observed in patients with muscle disease. The CK increase was more...... delayed in plasma. Tcap was not detectable at any time. CONCLUSIONS: Our results suggest that TnI is a marker of more severe damage signifying sarcomeric damage, and it could therefore be an important supplement to CK and Mb in clinical practice. Tcap is not useful as a marker for skeletal muscle damage....

  14. New functional assays to selectively quantify the activated protein C- and tissue factor pathway inhibitor-cofactor activities of protein S in plasma.

    Science.gov (United States)

    Alshaikh, N A; Rosing, J; Thomassen, M C L G D; Castoldi, E; Simioni, P; Hackeng, T M

    2017-02-17

    Essentials Protein S is a cofactor of activated protein C (APC) and tissue factor pathway inhibitor (TFPI). There are no assays to quantify separate APC and TFPI cofactor activities of protein S in plasma. We developed assays to measure the APC- and TFPI-cofactor activities of protein S in plasma. The assays were sensitive to protein S deficiency, and not affected by the Factor V Leiden mutation.

  15. Blood carbonyl protein measurement as a specific oxidative stress biomarker after intestinal reperfusion in rats Dosagem da proteína carbonilada sanguínea como biomarcador específico do estresse oxidativo após reperfusão intestinal em ratos

    Directory of Open Access Journals (Sweden)

    Márcio José Jamel

    2010-02-01

    Full Text Available PURPOSE: An experimental study was performed to investigate the use of protein carbonyl group as a specific biological marker for oxidative stress in a rat model of intestinal ischaemia-reperfusion. METHODS: Twenty four male Wistar rats were randomly distributed into three groups with eight animals each: Group 1 - Control group; Group 2 - Sham; Group 3 - Intestinal ischaemia by clamping ileal branches of the superior mesenteric artery for one hour, followed by another hour of reperfusion. Blood samples were taken in order to analyze the protein carbonyl level by Slot blotting assay. RESULTS: In group 3 a significant increase of protein carbonyl level was observed if compared to the homogenous levels of groups 1 and 2. CONCLUSION: From the results it may be concluded that the protein carbonylation may be used as a specific marker for measuring oxidative stress in rat intestinal reperfusion model.OBJETIVO: Realizou-se um estudo experimental com a finalidade de investigar o uso da proteína carbonilada como um marcador biológico específico do estresse oxidativo em um modelo de isquemia e reperfusão intestinal, em ratos. MÉTODOS: Vinte e quarto ratos da linhagem Wistar, machos foram distribuídos, aleatoriamente, em três grupos compostos por oito animais cada: Grupo 1 - Controle; Grupo 2 - Simulação e Grupo 3 - Submetido à isquemia, mediante clampeamento de ramos ileais da artéria mesentérica superior por uma hora, seguida de reperfusão, por igual período. Amostras sanguíneas obtidas foram utilizadas para analise dos níveis de proteína carbonilada, através do método Slot blotting. RESULTADOS: No grupo 3 houve uma elevação significante da concentração de proteína carbonilada sérica se comparada aos níveis sanguíneos homogêneos encontrados nos grupos 1 e 2. CONCLUSÃO: Fundamentado nos resultados é possível concluir que, a carbonilação protéica pode ser utilizada como um marcador específico para a mensuração do

  16. Plasma proteins as biomarkers of the aging process.

    Science.gov (United States)

    Vranckx, R; Savu, L; Lambert, N; de Conchard, G V; Grosse, R; Mourey, M S; Corman, B

    1995-02-01

    This study was designed to characterize the rat serum proteins as biomarkers of the normal aging process. Crossed immunoelectrophoresis or electroimmunodiffusion quantitation of proteins was performed in rats aged 6, 12, 24, and 30 mo. Selection of healthy animals was based on confrontation of crossed immunoelectrophoresis patterns with those of experimentally inflamed young adults and with individual anatomopathological data. Convergence of inflammatory patterns and severe histological lesions was the exclusion criterion. Senescence-induced decrease was demonstrated for eight proteins [negative senescence reactants (SRs-)] and increase for six proteins [positive SRs (SRs+)]. Most SRs belonged to the class of proteins responsive to acute inflammation [acute phase reactants (APRs)]. One SR+, the thyroxine-binding globulin, a high-affinity thyroid hormone binder, emerged as a particularly reliable senescence biomarker, showing the highest aging-related variation (8-fold increase from 6 to 30 mo) and not belonging to the APR class. Chronic treatment with perindopril, an angiotensin I-converting enzyme inhibitor used in heart and renal disease therapy, significantly enhanced thyroxine-binding capacity, possibly by preventing age-related alterations of serum lipids. Serum protein patterns prove valuable both as indexes for selecting aging animals free from superimposed pathologies and as parameters of senescence-induced changes in protein biosynthesis.

  17. Whey Protein Delays Gastric Emptying and Suppresses Plasma Fatty Acids and Their Metabolites Compared to Casein, Gluten, and Fish Protein

    DEFF Research Database (Denmark)

    Stanstrup, Jan; Schou, Simon S; Holmer-Jensen, Jens

    2014-01-01

    Whey protein has been demonstrated to improve fasting lipid and insulin response in overweight and obese individuals. To establish new hypotheses for this effect and to investigate the impact of stomach emptying, we compared plasma profiles after intake of whey isolate (WI), casein, gluten (GLU...

  18. Protein retention on plasma-treated hierarchical nanoscale gold-silver platform

    Science.gov (United States)

    Fang, Jinghua; Levchenko, Igor; Mai-Prochnow, Anne; Keidar, Michael; Cvelbar, Uros; Filipic, Gregor; Han, Zhao Jun; Ostrikov, Kostya (Ken)

    2015-08-01

    Dense arrays of gold-supported silver nanowires of about 100 nm in diameter grown directly in the channels of nanoporous aluminium oxide membrane were fabricated and tested as a novel platform for the immobilization and retention of BSA proteins in the microbial-protective environments. Additional treatment of the silver nanowires using low-temperature plasmas in the inductively-coupled plasma reactor and an atmospheric-pressure plasma jet have demonstrated that the morphology of the nanowire array can be controlled and the amount of the retained protein may be increased due to the plasma effect. A combination of the neutral gold sublayer with the antimicrobial properties of silver nanowires could significantly enhance the efficiency of the platforms used in various biotechnological processes.

  19. Rational use of plasma protein and tissue binding data in drug design.

    Science.gov (United States)

    Liu, Xingrong; Wright, Matthew; Hop, Cornelis E C A

    2014-10-23

    It is a commonly accepted assumption that only unbound drug molecules are available to interact with their targets. Therefore, one of the objectives in drug design is to optimize the compound structure to increase in vivo unbound drug concentration. In this review, theoretical analyses and experimental observations are presented to illustrate that low plasma protein binding does not necessarily lead to high in vivo unbound plasma concentration. Similarly, low brain tissue binding does not lead to high in vivo unbound brain tissue concentration. Instead, low intrinsic clearance leads to high in vivo unbound plasma concentration, and low efflux transport activity at the blood-brain barrier leads to high unbound brain concentration. Plasma protein and brain tissue binding are very important parameters in understanding pharmacokinetics, pharmacodynamics, and toxicities of drugs, but these parameters should not be targeted for optimization in drug design.

  20. Protein retention on plasma-treated hierarchical nanoscale gold-silver platform

    Science.gov (United States)

    Fang, Jinghua; Levchenko, Igor; Mai-Prochnow, Anne; Keidar, Michael; Cvelbar, Uros; Filipic, Gregor; Han, Zhao Jun; Ostrikov, Kostya (Ken)

    2015-01-01

    Dense arrays of gold-supported silver nanowires of about 100 nm in diameter grown directly in the channels of nanoporous aluminium oxide membrane were fabricated and tested as a novel platform for the immobilization and retention of BSA proteins in the microbial-protective environments. Additional treatment of the silver nanowires using low-temperature plasmas in the inductively-coupled plasma reactor and an atmospheric-pressure plasma jet have demonstrated that the morphology of the nanowire array can be controlled and the amount of the retained protein may be increased due to the plasma effect. A combination of the neutral gold sublayer with the antimicrobial properties of silver nanowires could significantly enhance the efficiency of the platforms used in various biotechnological processes. PMID:26307515

  1. Large-scale inference of protein tissue origin in gram-positive sepsis plasma using quantitative targeted proteomics.

    Science.gov (United States)

    Malmström, Erik; Kilsgård, Ola; Hauri, Simon; Smeds, Emanuel; Herwald, Heiko; Malmström, Lars; Malmström, Johan

    2016-01-06

    The plasma proteome is highly dynamic and variable, composed of proteins derived from surrounding tissues and cells. To investigate the complex processes that control the composition of the plasma proteome, we developed a mass spectrometry-based proteomics strategy to infer the origin of proteins detected in murine plasma. The strategy relies on the construction of a comprehensive protein tissue atlas from cells and highly vascularized organs using shotgun mass spectrometry. The protein tissue atlas was transformed to a spectral library for highly reproducible quantification of tissue-specific proteins directly in plasma using SWATH-like data-independent mass spectrometry analysis. We show that the method can determine drastic changes of tissue-specific protein profiles in blood plasma from mouse animal models with sepsis. The strategy can be extended to several other species advancing our understanding of the complex processes that contribute to the plasma proteome dynamics.

  2. Characterization of Plasma Membrane Proteins from Ovarian Cancer Cells Using Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    David L. Springer

    2004-01-01

    Full Text Available To determine how the repertoire of plasma membrane proteins change with disease state, specifically related to cancer, several methods for preparation of plasma membrane proteins were evaluated. Cultured cells derived from stage IV ovarian tumors were grown to 90% confluence and harvested in buffer containing CHAPS detergent. This preparation was centrifuged at low speed to remove insoluble cellular debris resulting in a crude homogenate. Glycosylated proteins in the crude homogenate were selectively enriched using lectin affinity chromatography. The crude homogenate and the lectin purified sample were prepared for mass spectrometric evaluation. The general procedure for protein identification began with trypsin digestion of protein fractions followed by separation by reversed phase liquid chromatography that was coupled directly to a conventional tandem mass spectrometer (i.e. LCQ ion trap. Mass and fragmentation data for the peptides were searched against a human proteome data base using the informatics program SEQUEST. Using this procedure 398 proteins were identified with high confidence, including receptors, membrane-associated ligands, proteases, phosphatases, as well as structural and adhesion proteins. Results indicate that lectin chromatography provides a select subset of proteins and that the number and quality of the identifications improve as does the confidence of the protein identifications for this subset. These results represent the first step in development of methods to separate and successfully identify plasma membrane proteins from advanced ovarian cancer cells. Further characterization of plasma membrane proteins will contribute to our understanding of the mechanisms underlying progression of this deadly disease and may lead to new targeted interventions as well as new biomarkers for diagnosis.

  3. Prognostic value of plasma C-reactive protein in the evaluation of paraquat poisoning patients

    Institute of Scientific and Technical Information of China (English)

    Zong NingΔ; Yu-Long BaiΔ; Hua Lu; Kang-Lin Mo

    2015-01-01

    Objective:To investigate the prognostic value of plasma C-reactive protein (CRP) level in patients with paraquat poisoning. Methods: This study included 162 patients with paraquat poisoning. The data of plasma paraquat,CRP level and arterial blood gas were analyzed. Cox regression analysis was applied to evaluate the risk factors of prognosis. Receiver operating characteristics curve analysis and area under curve were used to calculate the predictive power of significant variable. Differences in patient survival were determined using the Kaplan-Meier method and a log-rank test. Results:PlasmaCRP level was significantly increased in non-survival patients compared with survival patients (P Conclusions: These results suggest that plasmaCRP level is distinct increased in patients with paraquat poisoning, and the plasmaCRP level may be useful for the prediction of prognosis in paraquat poisoning.

  4. Lectin receptor kinases participate in protein-protein interactions to mediate plasma membrane-cell wall adhesions in Arabidopsis.

    Science.gov (United States)

    Gouget, Anne; Senchou, Virginie; Govers, Francine; Sanson, Arnaud; Barre, Annick; Rougé, Pierre; Pont-Lezica, Rafael; Canut, Hervé

    2006-01-01

    Interactions between plant cell walls and plasma membranes are essential for cells to function properly, but the molecules that mediate the structural continuity between wall and membrane are unknown. Some of these interactions, which are visualized upon tissue plasmolysis in Arabidopsis (Arabidopsis thaliana), are disrupted by the RGD (arginine-glycine-aspartic acid) tripeptide sequence, a characteristic cell adhesion motif in mammals. In planta induced-O (IPI-O) is an RGD-containing protein from the plant pathogen Phytophthora infestans that can disrupt cell wall-plasma membrane adhesions through its RGD motif. To identify peptide sequences that specifically bind the RGD motif of the IPI-O protein and potentially play a role in receptor recognition, we screened a heptamer peptide library displayed in a filamentous phage and selected two peptides acting as inhibitors of the plasma membrane RGD-binding activity of Arabidopsis. Moreover, the two peptides also disrupted cell wall-plasma membrane adhesions. Sequence comparison of the RGD-binding peptides with the Arabidopsis proteome revealed 12 proteins containing amino acid sequences in their extracellular domains common with the two RGD-binding peptides. Eight belong to the receptor-like kinase family, four of which have a lectin-like extracellular domain. The lectin domain of one of these, At5g60300, recognized the RGD motif both in peptides and proteins. These results imply that lectin receptor kinases are involved in protein-protein interactions with RGD-containing proteins as potential ligands, and play a structural and signaling role at the plant cell surfaces.

  5. Lectin Receptor Kinases Participate in Protein-Protein Interactions to Mediate Plasma Membrane-Cell Wall Adhesions in Arabidopsis1

    Science.gov (United States)

    Gouget, Anne; Senchou, Virginie; Govers, Francine; Sanson, Arnaud; Barre, Annick; Rougé, Pierre; Pont-Lezica, Rafael; Canut, Hervé

    2006-01-01

    Interactions between plant cell walls and plasma membranes are essential for cells to function properly, but the molecules that mediate the structural continuity between wall and membrane are unknown. Some of these interactions, which are visualized upon tissue plasmolysis in Arabidopsis (Arabidopsis thaliana), are disrupted by the RGD (arginine-glycine-aspartic acid) tripeptide sequence, a characteristic cell adhesion motif in mammals. In planta induced-O (IPI-O) is an RGD-containing protein from the plant pathogen Phytophthora infestans that can disrupt cell wall-plasma membrane adhesions through its RGD motif. To identify peptide sequences that specifically bind the RGD motif of the IPI-O protein and potentially play a role in receptor recognition, we screened a heptamer peptide library displayed in a filamentous phage and selected two peptides acting as inhibitors of the plasma membrane RGD-binding activity of Arabidopsis. Moreover, the two peptides also disrupted cell wall-plasma membrane adhesions. Sequence comparison of the RGD-binding peptides with the Arabidopsis proteome revealed 12 proteins containing amino acid sequences in their extracellular domains common with the two RGD-binding peptides. Eight belong to the receptor-like kinase family, four of which have a lectin-like extracellular domain. The lectin domain of one of these, At5g60300, recognized the RGD motif both in peptides and proteins. These results imply that lectin receptor kinases are involved in protein-protein interactions with RGD-containing proteins as potential ligands, and play a structural and signaling role at the plant cell surfaces. PMID:16361528

  6. Oxidative Stress and Carbonyl Lesions in Ulcerative Colitis and Associated Colorectal Cancer

    Science.gov (United States)

    Wang, Zhiqi; Li, Sai; Cao, Yu; Tian, Xuefei; Zeng, Rong; Liao, Duan-Fang; Cao, Deliang

    2016-01-01

    Oxidative stress has long been known as a pathogenic factor of ulcerative colitis (UC) and colitis-associated colorectal cancer (CAC), but the effects of secondary carbonyl lesions receive less emphasis. In inflammatory conditions, reactive oxygen species (ROS), such as superoxide anion free radical (O2 ∙−), hydrogen peroxide (H2O2), and hydroxyl radical (HO∙), are produced at high levels and accumulated to cause oxidative stress (OS). In oxidative status, accumulated ROS can cause protein dysfunction and DNA damage, leading to gene mutations and cell death. Accumulated ROS could also act as chemical messengers to activate signaling pathways, such as NF-κB and p38 MAPK, to affect cell proliferation, differentiation, and apoptosis. More importantly, electrophilic carbonyl compounds produced by lipid peroxidation may function as secondary pathogenic factors, causing further protein and membrane lesions. This may in turn exaggerate oxidative stress, forming a vicious cycle. Electrophilic carbonyls could also cause DNA mutations and breaks, driving malignant progression of UC. The secondary lesions caused by carbonyl compounds may be exceptionally important in the case of host carbonyl defensive system deficit, such as aldo-keto reductase 1B10 deficiency. This review article updates the current understanding of oxidative stress and carbonyl lesions in the development and progression of UC and CAC. PMID:26823956

  7. Oxidative Stress and Carbonyl Lesions in Ulcerative Colitis and Associated Colorectal Cancer

    Directory of Open Access Journals (Sweden)

    Zhiqi Wang

    2016-01-01

    Full Text Available Oxidative stress has long been known as a pathogenic factor of ulcerative colitis (UC and colitis-associated colorectal cancer (CAC, but the effects of secondary carbonyl lesions receive less emphasis. In inflammatory conditions, reactive oxygen species (ROS, such as superoxide anion free radical (O2∙-, hydrogen peroxide (H2O2, and hydroxyl radical (HO∙, are produced at high levels and accumulated to cause oxidative stress (OS. In oxidative status, accumulated ROS can cause protein dysfunction and DNA damage, leading to gene mutations and cell death. Accumulated ROS could also act as chemical messengers to activate signaling pathways, such as NF-κB and p38 MAPK, to affect cell proliferation, differentiation, and apoptosis. More importantly, electrophilic carbonyl compounds produced by lipid peroxidation may function as secondary pathogenic factors, causing further protein and membrane lesions. This may in turn exaggerate oxidative stress, forming a vicious cycle. Electrophilic carbonyls could also cause DNA mutations and breaks, driving malignant progression of UC. The secondary lesions caused by carbonyl compounds may be exceptionally important in the case of host carbonyl defensive system deficit, such as aldo-keto reductase 1B10 deficiency. This review article updates the current understanding of oxidative stress and carbonyl lesions in the development and progression of UC and CAC.

  8. Relationship between the plasma levels of neurodegenerative proteins and motor subtypes of Parkinson's disease.

    Science.gov (United States)

    Ding, Jian; Zhang, Jiejin; Wang, Xixi; Zhang, Li; Jiang, Siming; Yuan, Yongsheng; Li, Junyi; Zhu, Lin; Zhang, Kezhong

    2017-03-01

    The aim of our study is to examine the plasma levels of the four kinds of neurodegenerative proteins in plasma: α-syn, T-tau, P-tau181, and Aβ-42 in Parkinson's disease (PD) and to evaluate the relationship between their plasma levels and PD motor subtypes. 84 patients with PD were enrolled in our study, and finally, 73 of them were classified into the tremor-dominant subtype (TD) and the postural instability gait difficulty subtype (PIGD). Their motor performance was evaluated by a series of clinical assessments: Freezing of Gait Questionnaire (FOGQ), Timed Up and Go (TUGs), Tinetti balance, and Tinetti gait. Plasma levels of these proteins were measured by enzyme-linked immunosorbent assay (ELISA). The plasma level of α-syn was significantly higher in PD patients when compared to controls (p = 0.004), and significantly higher in the PIGD group when compared to the TD group (p = 0.03). While the plasma level of Aβ-42 was significantly lower in PD patients than in controls (p = 0.002), and significantly lower in the PIGD group than in the TD group (p = 0.05). In PD patients, the plasma level of α-syn (r = -0.355, p score, even after performing multiple linear regression (p = 0.002). While the plasma level of Aβ-42 (r = -0.261, p score and remained correlate when performed multiple linear regression (p = 0.005). The patients with PIGD subtype are characterized with a lower level of plasma Aβ-42 and a higher plasma level of α-syn, which may be used as biomarkers for diagnosis and progression of the subtypes of PD.

  9. Pregnancy-associated plasma protein-A and the vulnerable plaque

    DEFF Research Database (Denmark)

    Jespersen, Camilla H B; Vestergaard, Kirstine R; Schou, Morten

    2014-01-01

    For more than a decade, pregnancy-associated plasma protein-A (PAPP-A) has been examined for its relation to acute coronary syndrome (ACS) and the vulnerable plaque. This review summarizes the current knowledge of plasma PAPP-A in relation to nonpregnant individuals focusing on patients with ACS,......, discusses its use as a possible biomarker for diagnosis and prognosis in ACS, briefly describes the challenges in different assay technologies and describes the effect of heparin administration on PAPP-A concentrations in plasma....

  10. Micro patterning of cell and protein non-adhesive plasma polymerized coatings for biochip applications

    DEFF Research Database (Denmark)

    Bouaidat, Salim; Berendsen, C.; Thomsen, P.;

    2004-01-01

    Micro scale patterning of bioactive surfaces is desirable for numerous biochip applications. Polyethyleneoxide-like (PEO-like) coating with non-fouling functionality has been deposited using low frequency AC plasma polymerization. The non-fouling properties of the coating were tested with human...... cells ( HeLa) and fluorescence labeled proteins (isothiocyanate-labeled bovine serum albumin, i.e. FITC-BSA). The PEO-like coatings were fabricated by plasma polymerization of 12-crown-4 (ppCrown) with plasma polymerized hexene (ppHexene) as adhesion layer. The coatings were micro patterned using...

  11. Reduced protein oxidation in Wistar rats supplemented with marine ω3 PUFAs.

    Science.gov (United States)

    Méndez, Lucía; Pazos, Manuel; Gallardo, José M; Torres, Josep L; Pérez-Jiménez, Jara; Nogués, Rosa; Romeu, Marta; Medina, Isabel

    2013-02-01

    The potential effects of various dietary eicosapentaenoic acid (EPA; 20:5) and docosahexaenoic acid (DHA; 22:6) ratios (1:1, 2:1, and 1:2, respectively) on protein redox states from plasma, kidney, skeletal muscle, and liver were investigated in Wistar rats. Dietary fish oil groups were compared with animals fed soybean and linseed oils, vegetable oils enriched in ω6 linoleic acid (LA; 18:2) and ω3 α-linolenic acid (ALA; 18:3), respectively. Fish oil treatments were effective at reducing the level of total fatty acids in plasma and enriching the plasmatic free fatty acid fraction and erythrocyte membranes in EPA and DHA. A proteomic approach consisting of fluorescein 5-thiosemicarbazide (FTSC) labeling of protein carbonyls, FTSC intensity visualization on 1-DE or 2-DE gels, and protein identification by MS/MS was used for the protein oxidation assessment. Albumin was found to be the most carbonylated protein in plasma for all dietary groups, and its oxidation level was significantly modulated by dietary interventions. Supplementation with an equal EPA:DHA ratio (1:1) showed the lowest oxidation score for plasma albumin, followed in increasing order of carbonylation by 1:2 proteins and cytosolic proteins from kidney and liver also indicated a protective effect on proteins for the fish oil treatments, the 1:1 ratio exhibiting the lowest protein oxidation scores. The effect of fish oil treatments at reducing carbonylation on specific proteins from plasma (albumin), skeletal muscle (actin), and liver (albumin, argininosuccinate synthetase, 3-α-hydroxysteroid dehydrogenase) was remarkable. This investigation highlights the efficiency of dietary fish oil at reducing in vivo oxidative damage of proteins compared to oils enriched in the 18-carbon polyunsaturated fatty acids ω3 ALA and ω6 LA, and such antioxidant activity may differ among different fish oil sources because of variations in EPA/DHA content.

  12. Valproic acid: in vitro plasma protein binding and interaction with phenytoin.

    Science.gov (United States)

    Cramer, J A; Mattson, R H

    1979-01-01

    Because valproic acid (VPA) is highly bound to plasma protein, several variables affecting binding will significantly alter the quantity of free drug which is pharmacologically active. Therefore, total VPA plasma concentrations do not reflect the therapeutic strength of the drug in tissue. We have performed equilibrium dialysis and ultrafiltration studies of VPA binding to plasma protein. The converging data in these in vitro studies indicate a clinically significant alteration in the percent of free VPA when total drug concentration exceeds 80 micrograms/ml. Saturation of drug binding sites probably occurs in this range. At 20--60 micrograms/ml VPA there is 5% free drug, with a significant increase to 8% free at 80 micrograms/ml; free drug increases to over 20% at 145 micrograms/ml total VPA. Human plasma, which is low in albumin, has twice the quantity of free VPA as normal plasma (10 versus 5% free). The clinical evidence of interaction between VPA and phenytoin is confirmed in vitro by the increase in the free fraction of both drugs. VPA binding decreases by 3--6%, while phenytoin binding decreases 5--6% as both drugs reach high plasma concentrations. When appropriate, laboratory reports should be available defining concentration of free drug in plasma for optimal interpretation of drug concetrations relative to clinical effects.

  13. Effects of plant proteins on postprandial, free plasma amino acid concentrations in rainbow trout (Oncorhynchus mykiss)

    DEFF Research Database (Denmark)

    Larsen, Bodil Katrine; Dalsgaard, Anne Johanne Tang; Pedersen, Per Bovbjerg

    2012-01-01

    proteins from wheat, peas, field beans, sunflower and soybean. Blood samples were obtained from the caudal vein of 7 fish in each dietary treatment group prior to feeding, as well as: 2, 4, 6, 8, 12, 24, 48 and 72 h after feeding (sampling 7 new fish at each time point), and plasma amino acid......Postprandial patterns in plasma free amino acid concentrations were investigated in juvenile rainbow trout (Oncorhynchus mykiss) fed either a fish meal based diet (FM) or a diet (VEG) where 59% of fish meal protein (corresponding to 46% of total dietary protein) was replaced by a matrix of plant...... concentrations were subsequently measured by HPLC. Nutrient digestibility and ammonia excretion of the two experimental diets were measured in a parallel experiment using a modified Guelph setup. Results showed that the appearance of most amino acids (essential and non-essential) in the plasma was delayed...

  14. Biomarkers of Oxidative Stress Study V: Ozone exposure of rats and its effect on lipids, proteins and DNA in plasma and urine

    Science.gov (United States)

    Kadiiska, Maria B.; Basu, Samar; Brot, Nathan; Cooper, Christopher; Csallany, A. Saari; Davies, Michael J.; George, Magdalene M.; Murray, Dennis M.; Roberts, L. Jackson; Shigenaga, Mark K.; Sohal, Rajindar S.; Stocker, Roland; Van Thiel, David H.; Wiswedel, Ingrid; Hatch, Gary E.; Mason, Ronald P.

    2014-01-01

    Ozone exposure effect on free radical-catalyzed oxidation products of lipids, proteins and DNA in the plasma and urine of rats was studied as a continuation of the international Biomarker of Oxidative Stress Study (BOSS) sponsored by NIEHS/NIH. The goal was to identify a biomarker for ozone-induced oxidative stress and to assess whether inconsistent results often reported in the literature might be due to the limitations of the available methods for measuring the various types of oxidative products. The time and dose-dependent effects of ozone exposure on rat plasma lipid hydroperoxides, malondialdehyde, F2-isoprostanes, protein carbonyls, methionine oxidation, tyrosine- and phenylalanine oxidation products, as well as urinary malondialdehyde and F2-isoprostanes were investigated with various techniques. The criterion used to recognize a marker in the model of ozone exposure was that a significant effect could be identified and measured in a biological fluid seen at both doses at more than one time point. No statistically significant differences between the experimental and control groups at either ozone dose and time point studied could be identified in this study. Tissue samples were not included. Despite all the work accomplished in the BOSS study of ozone, no available product of oxidation in biological fluid has yet met the required criteria of being a biomarker. The current negative findings as a consequence of ozone exposure are of great importance, because they document that in complex systems, as the present in vivo experiment, the assays used may not provide meaningful data of ozone oxidation, especially in human studies. PMID:23608465

  15. Temperature-Induced Protein Conformational Changes in Barley Root Plasma Membrane-Enriched Microsomes

    Science.gov (United States)

    Caldwell, Charles R.

    1987-01-01

    The membrane-bound proteins of barley (Hordeum vulgare L. cv Conquest) root plasma membrane-enriched microsomes displayed fluorescence typical of protein-associated trytophan residues. The protein fluorescence intensity was sensitive to variations in sample temperature. The temperature-induced decline in protein fluorescence intensity was nonlinear with slope discontinuities at about 12 and 32°C. Detergents at levels above their critical micelle concentration enhanced protein fluorescence. Glutaraldehyde reduced protein fluorescence. Protein fluorescence polarization increased at temperatures above 30°C. Both the rate of tryptophan photoionization and the fluorescence intensity of the photoionization products suggested alterations in membrane protein conformation between 12 and 32°C. The quenching of the intrinsic protein fluorescence by acrylamide and potassium iodide indicated changes in accessibility of the extrinsic agents to the protein tryptophan residues beginning at about 14°C. The results indicate thermally induced changes in the dynamics of the membrane proteins over the temperature range of 12 to 32°C which could account for the complex temperature dependence of the barley root plasma membrane ATPase. PMID:16665545

  16. Localization of porcine seminal plasma (PSP) proteins in the boar reproductive tract and spermatozoa.

    Science.gov (United States)

    Manásková, P; Jonáková, V

    2008-06-01

    Spermadhesins are proteins containing a characteristic CUB domain, originally isolated from seminal plasma and ejaculated spermatozoa in domestic animals. Boar spermadhesins are multifunctional proteins exhibiting ligand-binding abilities with various endogenous ligands present in the male and female reproductive tracts and may play a role in the reproduction process. Porcine spermadhesins (AQN, AWN, PSP protein families) are secreted mainly by the seminal vesicles, but their mRNAs have been found also in the cauda epididymis and prostate. Unlike AQN and AWN spermadhesins, localization of porcine seminal plasma (PSP) proteins in the boar reproductive tract has not been completely resolved. This work has focused on PSP protein expression and localization in the boar reproductive organs and on spermatozoa. Using specific rabbit polyclonal antibodies (anti-PSP I and anti-PSP II), PSP I and PSP II proteins were immunodetected in tissue extracts and in secretory tissues of cauda epididymis, prostate, seminal vesicles and Cowper's glands on the blots and by an indirect immunofluorescence technique, respectively. Moreover, the ability of PSP proteins to bind to epididymal spermatozoa indicated their presence on cauda epididymal and ejaculated spermatozoa. Porcine seminal plasma proteins bind to the sperm surface at ejaculation and may modulate several aspects of sperm activity during reproduction. PSP proteins are produced not only by seminal vesicles and prostate, but also by epididymis. However, their prospective role in sperm epididymal maturation is not clear. Further characterization of seminal plasma protein forms expressed in the individual reproductive organs will help to understand their subsequent role in the reproduction process.

  17. Influence of dietary fish proteins on plasma and liver cholesterol concentrations in rats.

    Science.gov (United States)

    Zhang, X; Beynen, A C

    1993-05-01

    The effects of amount and type of dietary fish proteins on plasma and liver cholesterol concentrations were evaluated in female rats. The isonitrogenous diets used contained 10 g cholesterol/kg and were carefully balanced for residual fat, cholesterol, Ca, Mg and P in the protein preparations. Cod meal, soya-bean protein or casein was incorporated into the diets as the only source of dietary protein at three levels: either 24, 48 or 72 g N/kg diet. Extra protein was added to the diet at the expense of the glucose component. In a second experiment soya-bean protein, casein, cod meal, whiting meal or plaice meal was added to the diet at a level of 24 g N/kg. When compared with casein, cod meal and soya-bean protein decreased plasma and liver cholesterol concentrations. A further cholesterol-lowering effect was achieved by increasing the proportion of either soya-bean protein or cod meal in the diet. Substitution of casein for glucose did not influence plasma and liver cholesterol concentrations. Plaice meal in the diet produced lower group mean plasma cholesterol concentrations than did whiting meal. In rats fed on the diet containing plaice meal, liver cholesterol concentrations were significantly lower than those in their counterparts fed on either cod meal or whiting meal. The present study demonstrates that different fish proteins in the diet have different effects on cholesterol metabolism and that the cholesterol-influencing properties of cod meal can be enhanced by the incorporation of higher proportions of this protein in the diet.

  18. Trifluoromethylation of Carbonyl Compounds with Sodium Trifluoroacetate

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    In the presence of copper (Ⅰ) halide as catalyst, a variety of carbonyl compounds could be trifluoromethylated with sodium trifluoroacetate to give the corresponding alcohols in moderate to high yields.

  19. Total plasma protein in very preterm babies: prognostic value and comparison with illness severity scores.

    Directory of Open Access Journals (Sweden)

    Silvia Iacobelli

    Full Text Available OBJECTIVE: We aimed to investigate the predictive value for severe adverse outcome of plasma protein measurements on day one of life in very preterm infants and to compare total plasma protein levels with the validated illness severity scores CRIB, CRIB-II, SNAP-II and SNAPPE-II, regarding their predictive ability for severe adverse outcome. METHODS: We analyzed a cohort of infants born at 24-31 weeks gestation, admitted to the tertiary intensive care unit of a university hospital over 10.5 years. The outcome measure was "severe adverse outcome" defined as death before discharge or severe neurological injury on cranial ultrasound. The adjusted odd ratio (aOR and 95% confidence interval (95% CI of severe adverse outcome for hypoproteinemia (total plasma protein level <40 g/L was calculated by univariate and multivariate analyses. Calibration (Hosmer-Lemeshow goodness-of-fit was performed and the predictive ability for severe adverse outcome was assessed for total plasma protein and compared with CRIB, CRIB-II, SNAP-II and SNAPPE-II, by calculating receiver operating characteristic (ROC curves and their associated area under the curve (AUC. RESULTS: 761 infants were studied: 14.4% died and 4.1% survived with severe cerebral ultrasound findings. The aOR of severe adverse outcome for hypoproteinemia was 6.1 (95% CI 3.8-9.9. The rank order for variables, as assessed by AUCs and 95% CIs, in predicting outcome was: total plasma protein [0.849 (0.821-0.873], SNAPPE-II [0.822 (0.792-0.848], CRIB [0.821 (0.792-0.848], SNAP-II [0.810 (0.780-0.837] and CRIB-II [0.803 (0.772-0.830]. Total plasma protein predicted severe adverse outcome significantly better than CRIB-II and SNAP-II (both p<0.05. Calibration for total plasma protein was very good. CONCLUSIONS: Early hypoproteinemia has prognostic value for severe adverse outcome in very preterm, sick infants. Total plasma protein has a predictive performance comparable with CRIB and SNAPPE-II and greater than

  20. Prognostic value of plasma C-reactive protein in the evaluation of paraquat poisoning patients简

    Institute of Scientific and Technical Information of China (English)

    Zong; Ning; Yu-Long; Bai; Hua; Lu; Kang-Lin; Mo

    2015-01-01

    Objective: To investigate the prognostic value of plasma C-reactive protein(CRP) level in patients with paraquat poisoning.Methods: This study included 162 patients with paraquat poisoning. The data of plasma paraquat, CRP level and arterial blood gas were analyzed. Cox regression analysis was applied to evaluate the risk factors of prognosis. Receiver operating characteristics curve analysis and area under curve were used to calculate the predictive power of significant variable. Differences in patient survival were determined using the Kaplan–Meier method and a log-rank test.Results: Plasma CRP level was significantly increased in non-survival patients compared with survival patients(P < 0.05), and positively correlated with plasma paraquat level(P < 0.05). Cox regression analysis revealed that plasma CRP level was an independent prognostic marker of mortality within 30 days. The receiver operating characteristics curve analysis indicated that area under curve of plasma CRP level was0.867(95% CI: 0.81–0.93), and the cut-off value was 18 mg/L, and patients with CRP level over this value had a poor survival time compared with those with less than this value.Conclusions: These results suggest that plasma CRP level is distinct increased in patients with paraquat poisoning, and the plasma CRP level may be useful for the prediction of prognosis in paraquat poisoning.

  1. Ab Initio MO Studies on the Reaction Mechanism for Carbonyl Insertion Catalyzed by Carbonyl Cobalt Complex

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Ab initio method, under the effective core potential(ECP) approximation at HF/LANL2DZ level, has been employed to study the reaction mechanism of the carbonyl insertion of olefin hydroformylation catalyzed by a carbonyl cobalt HCo(CO)3. The two reaction paths have been discussed. The calculated potential energy barriers for the carbonyl migration and the ethyl group migration are 105.0 kJ/mol and 39.17 kJ/mol, respectively. The results indicate that the reaction path via ethyl migration is more energetically favorable than that via carbonyl insertion.

  2. Arabidopsis protein kinase PKS5 inhibits the plasma membrane H+ -ATPase by preventing interaction with 14-3-3 protein

    DEFF Research Database (Denmark)

    Fuglsang, Anja Thoe; Guo, Yan; Cuin, Tracey A.

    2007-01-01

    that an Arabidopsis thaliana Ser/Thr protein kinase, PKS5, is a negative regulator of the plasma membrane proton pump (PM Hþ-ATPase). Loss-of-function pks5 mutant plants are more tolerant of high external pH due to extrusion of protons to the extracellular space. PKS5 phosphorylates the PM Hþ-ATPase AHA2 at a novel......Regulation of the trans-plasma membrane pH gradient is an important part of plant responses to several hormonal and environmental cues, including auxin, blue light, and fungal elicitors. However, little is known about the signaling components that mediate this regulation. Here, we report...

  3. Adsorption of proteins from plasma at polyester non-wovens

    NARCIS (Netherlands)

    Klomp, A.J.A.; Engbers, G.H.M.; Mol, J.; Terlingen, J.G.A.; Feijen, J.

    1999-01-01

    Polyester non-wovens in filters for the removal of leukocytes from platelet concentrates (PCs) must be platelet compatible. In PC filtration, the adsorption of proteins at the plasma–non-woven interface can be of great importance with respect to the yield of platelets. Unmodified and radio frequency

  4. Plasma Membrane Protein Ubiquitylation and Degradation as Determinants of Positional Growth in Plants

    Institute of Scientific and Technical Information of China (English)

    Barbara Korbei; Christian Luschnig

    2013-01-01

    Being sessile organisms, plants evolved an unparalleled plasticity in their post-embryonic development, allowing them to adapt and fine-tune their vital parameters to an ever-changing environment. Cross-talk between plants and their environment requires tight regulation of information exchange at the plasma membrane (PM). Plasma membrane proteins mediate such communication, by sensing variations in nutrient availability, external cues as well as by controlled solute transport across the membrane border. Localiza-tion and steady-state levels are essential for PM protein function and ongoing research identified cis- and trans-acting determinants, involved in control of plant PM protein localization and turnover. In this overview, we summarize recent progress in our understanding of plant PM protein sorting and degradation via ubiquitylation, a post-translational and reversible modification of proteins. We highlight characterized components of the machinery involved in sorting of ubiquitylated PM proteins and discuss consequences of protein ubiquitylation on fate of selected PM proteins. Specifically, we focus on the role of ubiquitylation and PM protein degradation in the regulation of polar auxin transport (PAT). We combine this regulatory circuit with further aspects of PM protein sorting control, to address the interplay of events that might control PAT and polarized growth in higher plants.

  5. Cytochemical Demonstration of Oxidative Damage in Alzheimer Disease by Immunochemical Enhancement of the Carbonyl Reaction with 2,4-Dinitrophenylhydrazine

    National Research Council Canada - National Science Library

    Smith, Mark A; Sayre, Lawrence M; Anderson, Vernon E; Harris, Peggy L.R; Beal, M. Flint; Kowall, Neil; Perry, George

    1998-01-01

    ...-bound carbonyl reactivity. When this technique was applied to tissues from cases of Alzheimer disease, in which oxidative events including lipoperoxidative, glycoxidative, and other oxidative protein modifications have been...

  6. Improved detection specificity for plasma proteins by targeting cysteine-containing peptides with photo-SRM.

    Science.gov (United States)

    Enjalbert, Quentin; Girod, Marion; Simon, Romain; Jeudy, Jérémy; Chirot, Fabien; Salvador, Arnaud; Antoine, Rodolphe; Dugourd, Philippe; Lemoine, Jérôme

    2013-03-01

    Targeted mass spectrometry using selected reaction monitoring (SRM) has emerged as an alternative to immunoassays for protein quantification owing to faster development time and higher multiplexing capability. However, the SRM strategy is faced with the high complexity of peptide mixtures after trypsin digestion of whole plasma or the cellular proteome that most of the time causes contamination, irremediably, by interfering compounds in the transition channels monitored. This problem becomes increasingly acute when the targeted protein is present at a low concentration. In this work, the merit of laser-induced photo-dissociation in the visible region at 473 nm implemented in an hybrid quadrupole linear ion-trap mass spectrometer (photo-SRM) was evaluated for detection specificity of cysteine-containing peptides in a group of plasma proteins after tagging with a dabcyl chromophore. Compared with conventional SRM, photo-SRM chromatograms have improved detection specificity for most of peptides monitored. Comparison of the signals obtained for the best proteotypic peptides in SRM mode and those recorded by photo-SRM of cysteine-containing peptides for the same proteins reveals either increased (up to 10-fold) or similar signal to photo-SRM detection. Finally, photo-SRM has extended response linearity across a calibration plot obtained by diluting human plasma in rat plasma, down to the lowest concentrations. Hence, photo-SRM may advantageously complement conventional SRM in assay of proteins in complex biological matrices.

  7. Plasma protein fractions in healthy blood donors quantitated by an automated multicapillary electrophoresis system.

    Science.gov (United States)

    Larsson, Anders; Hansson, Lars-Olof

    2006-09-01

    During the last decade, capillary electrophoresis (CE) has emerged as an important alternative to traditional analysis of serum and plasma proteins by agarose or celluloseacetate electrophoresis. CE analysis of plasma proteins can now be fully automated and also includes bar-code identification of samples, preseparation steps, and direct post-separation quantitation of individual peaks, which permits short assay times and high throughput. For laboratory work, it is important to have reference values from healthy individuals. Therefore, plasma samples from 156 healthy blood donors (79 females and 77 males) have been analyzed with the Capillarys instrument and the new high resolution buffer, which yields higher resolution than the beta1-beta2+ buffer. Albumin concentrations in samples are measured using nephelometry in order to assign protein concentrations to each peak. The 2.5 and 97.5 percentiles for both the percentages of different peaks and the protein concentrations in the peaks are calculated according to the recommendations of the International Federation of Clinical Chemistry on the statistical treatment of reference values. The Capillarys instrument is a reliable system for plasma protein analysis, combining advantages of full automation with high analytical performances and throughput.

  8. Atmospheric pressure plasma polymers for tuned QCM detection of protein adhesion.

    Science.gov (United States)

    Rusu, G B; Asandulesa, M; Topala, I; Pohoata, V; Dumitrascu, N; Barboiu, M

    2014-03-15

    Our efforts have been concentrated in preparing plasma polymeric thin layers at atmospheric pressure grown on Quartz Crystal Microbalance-QCM electrodes for which the non-specific absorption of proteins can be efficiently modulated, tuned and used for QCM biosensing and quantification. Plasma polymerization reaction at atmospheric pressure has been used as a simple and viable method for the preparation of QCM bioactive surfaces, featuring variable protein binding properties. Polyethyleneglycol (ppEG), polystyrene (ppST) and poly(ethyleneglycol-styrene) (ppST-EG) thin-layers have been grown on QCM electrodes. These layers were characterized by Atomic Force Microscopy (AFM), Contact angle measurements, Fourier transform infrared (FTIR) and X-ray photoelectron spectroscopy (XPS). The plasma ppST QCM electrodes present a higher adsorption of Concanavalin A (ConA) and Bovine Serum Albumin (BSA) proteins when compared with the commercial coated polystyrene (ppST) ones. The minimum adsorption was found for ppEG, surface, known by their protein anti-fouling properties. The amount of adsorbed proteins can be tuned by the introduction of PEG precursors in the plasma discharge during the preparation of ppST polymers. © 2013 Elsevier B.V. All rights reserved.

  9. Study of Plasma Malondialdehyde, Troponin I and C - Reactive protein in Acute Coronary Syndromes Patients

    Directory of Open Access Journals (Sweden)

    S. Shams

    2006-04-01

    Full Text Available Introduction & Objective: Ischemic injury of endothelium is associated with prostaglandin synthesis and platelet adhesion and aggregation, which may be associated with the release of aldehydes such as malondialdehyde (MDA. C-reactive protein and cardiac troponin I have been proposed as diagnostic markers of acute coronary syndromes. In this study, we compared the usefulness of plasma MDA as a marker of acute coronary syndromes with that of C-reactive protein and troponin I.Material & Methods: The study population contained 50 patients with unstable angina and 50 patients with acute myocardial infarction admitted to the hearth department of the Ekbatan Hospital of Hamadan. The subjects were matched according to age and sex. Total cholesterol, LDL and HDL cholesterol, triglycerides, plasma MDA, troponin I and C-reactive protein levels were determined in all patients. Results: Results showed that the plasma MDA levels were significantly higher in patients with acute myocardial infarction than in individuals with unstable angina (P<0.001 and were associated with increased levels of troponin I and C-reactive protein (P<0.001.Conclusion: The combination of the plasma MDA levels, which reflect endothelial injury, and troponin I and C-reactive protein levels may allow better discrimination in acute coronary syndromes patients.

  10. Protein profile of the seminal plasma of collared peccaries (Pecari tajacu Linnaeus, 1758).

    Science.gov (United States)

    Santos, E A A; Sousa, P C; Martins, J A M; Moreira, R A; Monteiro-Moreira, A C O; Moreno, F B M B; Oliveira, M F; Moura, A A; Silva, A R

    2014-06-01

    This study was conducted to characterize the major proteins of the peccary seminal plasma, based on the semen samples collected from nine adult and reproductively sound animals. Our approach included the use of two-dimensional electrophoresis followed by Coomassie blue staining and analysis of polypeptide maps with PDQuest Software (Bio-Rad). Proteins were identified by tandem mass spectrometry (LC-MS/MS). We detected 179 protein spots per gel and 98 spots were identified by mass spectrometry, corresponding to 23 different proteins. The combined intensity of those spots accounted for 56.2±6% of the intensities of all spots and 60.9% of the intensities of spots presented in every protein map. Protein spots identified as clusterin represented 19.7±8.3% of the integrated optical densities of all spots detected in the seminal plasma maps. There was a negative association (r=-0.87; P<0.05) between the intensity of a clusterin spot and the percentage of sperm with functional membrane. Spermadhesin porcine seminal plasma protein 1 and bodhesin 2 comprised 5.4±1.9 and 8.8±3.9% of the total intensity of all spots respectively. Many proteins appeared in a polymorphic pattern, such as clusterin (27 spots), epididymal secretory glutathione peroxidase (ten spots), inter-α-trypsin inhibitor (12 spots), and IgG-binding protein (ten spots), among others. In conclusion, we presently describe the major seminal plasma proteome of the peccary, which exhibits a distinct high expression of clusterin isoforms. Knowledge of wild species reproductive biology is crucial for an understanding of their survival strategies and adaptation in a changing environment.

  11. Maternal Low Quality Protein Diet Alters Plasma Amino Acid Concentrations of Weaning Rats

    Directory of Open Access Journals (Sweden)

    Arzu Kabasakal Cetin

    2015-12-01

    Full Text Available Several studies have indicated the influence of a maternal low protein diet on the fetus. However, the effect of a maternal low quality protein diet on fetal growth and development is largely unknown. Wistar rats (11 weeks old were mated and maintained on either a chow diet with 20% casein (n = 6 as the control group (C, or a low quality protein diet with 20% wheat gluten (n = 7 as the experimental group (WG through gestation and lactation. Maternal body weights were similar in both groups throughout the study. Birth weights were not influenced by maternal diet and offspring body weights during lactation were similar between the groups. Offspring’s plasma amino acid profiles showed that plasma methionine, glutamine and lysine were significantly lower and aspartic acid, ornithine and glycine-proline were significantly higher in the WG. Plant based protein comprises an important part of protein intake in developing countries. It is well-known that these diets can be inadequate in terms of essential amino acids. The current study shows differential effects of a maternal low quality protein diet on the offspring’s plasma amino acids. Future studies will examine further aspects of the influence of maternal low quality protein diets on fetal growth and development.

  12. Resolving mixed mechanisms of protein subdiffusion at the T cell plasma membrane

    Science.gov (United States)

    Golan, Yonatan; Sherman, Eilon

    2017-06-01

    The plasma membrane is a complex medium where transmembrane proteins diffuse and interact to facilitate cell function. Membrane protein mobility is affected by multiple mechanisms, including crowding, trapping, medium elasticity and structure, thus limiting our ability to distinguish them in intact cells. Here we characterize the mobility and organization of a short transmembrane protein at the plasma membrane of live T cells, using single particle tracking and photoactivated-localization microscopy. Protein mobility is highly heterogeneous, subdiffusive and ergodic-like. Using mobility characteristics, we segment individual trajectories into subpopulations with distinct Gaussian step-size distributions. Particles of low-to-medium mobility consist of clusters, diffusing in a viscoelastic and fractal-like medium and are enriched at the centre of the cell footprint. Particles of high mobility undergo weak confinement and are more evenly distributed. This study presents a methodological approach to resolve simultaneous mixed subdiffusion mechanisms acting on polydispersed samples and complex media such as cell membranes.

  13. Interaction of Globular Plasma Proteins with Water-Soluble CdSe Quantum Dots.

    Science.gov (United States)

    Pathak, Jyotsana; Rawat, Kamla; Sanwlani, Shilpa; Bohidar, H B

    2015-06-08

    The interactions between water-soluble semiconductor quantum dots [hydrophilic 3-mercaptopropionic acid (MPA)-coated CdSe] and three globular plasma proteins, namely, bovine serum albumin (BSA), β-lactoglobulin (β-Lg) and human serum albumin (HSA), are investigated. Acidic residues of protein molecules form electrostatic interactions with these quantum dots (QDs). To determine the stoichiometry of proteins bound to QDs, we used dynamic light scattering (DLS) and zeta potential techniques. Fluorescence resonance energy transfer (FRET) experiments revealed energy transfer from tryptophan residues in the proteins to the QD particles. Quenching of the intrinsic fluorescence of protein molecules was noticed during this binding process (hierarchy HSA<β-Lg protein molecules). Upon binding with QD particles, the protein molecules underwent substantial conformational changes at the secondary-structure level (50 % helicity lost), due to loss in hydration.

  14. Elevation of plasma phospholipid transfer protein increases the risk of atherosclerosis despite lower apolipoprotein B-containing lipoproteins.

    NARCIS (Netherlands)

    J. Lie (Jessica); M.P.G. de Crom (Rini); T. van Gent (Teus); M.J. van Haperen (Rien); L. Scheek (Leo); F. Sadeghi-Niaraki (Farah); A. van Tol (Arie)

    2004-01-01

    textabstractPlasma phospholipid transfer protein (PLTP) transfers phospholipids between lipoproteins and mediates HDL conversion. PLTP-overexpressing mice have increased atherosclerosis. However, mice do not express cholesteryl ester transfer protein (CETP), which is involved in

  15. Membrane potential governs lateral segregation of plasma membrane proteins and lipids in yeast.

    Science.gov (United States)

    Grossmann, Guido; Opekarová, Miroslava; Malinsky, Jan; Weig-Meckl, Ina; Tanner, Widmar

    2007-01-10

    The plasma membrane potential is mainly considered as the driving force for ion and nutrient translocation. Using the yeast Saccharomyces cerevisiae as a model organism, we have discovered a novel role of the membrane potential in the organization of the plasma membrane. Within the yeast plasma membrane, two non-overlapping sub-compartments can be visualized. The first one, represented by a network-like structure, is occupied by the proton ATPase, Pma1, and the second one, forming 300-nm patches, houses a number of proton symporters (Can1, Fur4, Tat2 and HUP1) and Sur7, a component of the recently described eisosomes. Evidence is presented that sterols, the main lipid constituent of the plasma membrane, also accumulate within the patchy compartment. It is documented that this compartmentation is highly dependent on the energization of the membrane. Plasma membrane depolarization causes reversible dispersion of the H(+)-symporters, not however of the Sur7 protein. Mitochondrial mutants, affected in plasma membrane energization, show a significantly lower degree of membrane protein segregation. In accordance with these observations, depolarized membranes also considerably change their physical properties (detergent sensitivity).

  16. Plasma levels of osteocalcin and retinol binding protein-4 in patients with medullary thyroid carcinoma

    Directory of Open Access Journals (Sweden)

    Jabar Lotfi

    2014-04-01

    Conclusion: According to difference between plasma levels of osteocalcin and retinol binding protein-4 in patients suffered of medullary thyroid carcinoma comparison with normal subjects, it can be said that, probably medullary thyroid carcinoma has effect on bone and adipose tissue metabolism, so osteocalcin and retinol binding protein-4 hormones have potential to be used for confirmation of diagnosis or following treatment of medullary thyroid carcinoma.

  17. Reticulomics: Protein-Protein Interaction Studies with Two Plasmodesmata-Localized Reticulon Family Proteins Identify Binding Partners Enriched at Plasmodesmata, Endoplasmic Reticulum, and the Plasma Membrane.

    Science.gov (United States)

    Kriechbaumer, Verena; Botchway, Stanley W; Slade, Susan E; Knox, Kirsten; Frigerio, Lorenzo; Oparka, Karl; Hawes, Chris

    2015-11-01

    The endoplasmic reticulum (ER) is a ubiquitous organelle that plays roles in secretory protein production, folding, quality control, and lipid biosynthesis. The cortical ER in plants is pleomorphic and structured as a tubular network capable of morphing into flat cisternae, mainly at three-way junctions, and back to tubules. Plant reticulon family proteins (RTNLB) tubulate the ER by dimerization and oligomerization, creating localized ER membrane tensions that result in membrane curvature. Some RTNLB ER-shaping proteins are present in the plasmodesmata (PD) proteome and may contribute to the formation of the desmotubule, the axial ER-derived structure that traverses primary PD. Here, we investigate the binding partners of two PD-resident reticulon proteins, RTNLB3 and RTNLB6, that are located in primary PD at cytokinesis in tobacco (Nicotiana tabacum). Coimmunoprecipitation of green fluorescent protein-tagged RTNLB3 and RTNLB6 followed by mass spectrometry detected a high percentage of known PD-localized proteins as well as plasma membrane proteins with putative membrane-anchoring roles. Förster resonance energy transfer by fluorescence lifetime imaging microscopy assays revealed a highly significant interaction of the detected PD proteins with the bait RTNLB proteins. Our data suggest that RTNLB proteins, in addition to a role in ER modeling, may play important roles in linking the cortical ER to the plasma membrane.

  18. Decreased Bacterial Attachment and Protein Adsorption to Coatings Produced by Low Enegy Plasma Polymerization

    DEFF Research Database (Denmark)

    Andersen, T.E.; Kingshott, Peter; Benter, M.

    with a surface less prone to the adsorption of biological matter. In the current study two different hydrophilic nanoscale coatings were produced by low energy plasma polymerization [3] and investigated· f()rl()w ... pr()tein adsorption and bacterial attachment properties. Methods were setup to enable...... and Methods: Coatings: Plasma polymerized poly(vinyl pyrrolidone) (PP-PVP), poly(2-methoxyethyl methacrylate) (PPPMEA) or an inorganic oxide (10) coating were applied onto medical grade silicon rubber sheets (Silopren LSR 2050, Momentive Performance Materials Inc.). Plasma polymerization chamber......-coated crystals were then treated with one of the plasma polymerized coatings. Adsorption of fibrinogen, human serum albumin or immunoglobulin G was measured using a QCM-D instrument [5] (model E4, Q-Sense AB, Vastra Frolunda, Sweden) using a solution of 50llg/1 protein in PBS buffer. Results and Discussion: Our...

  19. Adsorbed plasma proteins modulate the effects of single-walled carbon nanotubes on neutrophils in blood.

    Science.gov (United States)

    Vlasova, Irina I; Mikhalchik, Elena V; Barinov, Nikolay A; Kostevich, Valeria A; Smolina, Natalia V; Klinov, Dmitry V; Sokolov, Alexey V

    2016-08-01

    Proteins adsorbed on a surface may affect the interaction of this surface with cells. Here, we studied the binding of human serum albumin (HSA), fibrinogen (FBG) and immunoglobulin G (IgG) to PEGylated single-walled carbon nanotubes (PEG-SWCNTs) and evaluated the impact of PEG-SWCNT treated by these proteins on neutrophils in whole blood samples. Measurements of adsorption parameters revealed tight binding of proteins to PEG-SWCNTs. AFM was employed to directly observe protein binding to sidewalls of PEG-SWCNTs. Fluorescein-labeled IgG was used to ascertain the stability of PEG-SWCNT-IgG complexes in plasma. In blood samples, all plasma proteins mitigated damage of neutrophils observed just after blood exposure to PEG-SWCNTs, while only treatment of PEG-SWCNTs with IgG resulted in dose- and time-dependent enhancement of CNT-induced neutrophil activation and in potentiation of oxidative stress. Our study demonstrates the ability of adsorbed plasma proteins to influence neutrophil response caused by PEG-SWCNTs in whole blood.

  20. Differential proteomics of human seminal plasma: A potential target for searching male infertility marker proteins.

    Science.gov (United States)

    Tomar, Anil Kumar; Sooch, Balwinder Singh; Singh, Sarman; Yadav, Savita

    2012-04-01

    The clinical fertility tests, available in the market, fail to define the exact cause of male infertility in almost half of the cases and point toward a crucial need of developing better ways of infertility investigations. The protein biomarkers may help us toward better understanding of unknown cases of male infertility that, in turn, can guide us to find better therapeutic solutions. Many clinical attempts have been made to identify biomarkers of male infertility in sperm proteome but only few studies have targeted seminal plasma. Human seminal plasma is a rich source of proteins that are essentially required for development of sperm and successful fertilization. This viewpoint article highlights the importance of human seminal plasma proteome in reproductive physiology and suggests that differential proteomics integrated with functional analysis may help us in searching potential biomarkers of male infertility.

  1. A cell-free assay to determine the stoichiometry of plasma membrane proteins.

    Science.gov (United States)

    Trigo, Cesar; Vivar, Juan P; Gonzalez, Carlos B; Brauchi, Sebastian

    2013-04-01

    Plasma membrane receptors, transporters, and ion channel molecules are often found as oligomeric structures that participate in signaling cascades essential for cell survival. Different states of protein oligomerization may play a role in functional control and allosteric regulation. Stochastic GFP-photobleaching (SGP) has emerged as an affordable and simple method to determine the stoichiometry of proteins at the plasma membrane. This non-invasive optical approach can be useful for total internal reflection of fluorescence microscopy (TIRFM), where signal-to-noise ratio is very high at the plasma membrane. Here, we report an alternative methodology implemented on a standard laser scanning confocal microscope (LSCM). The simplicity of our method will allow for its implementation in any epifluorescence microscope of choice.

  2. METHODS OF DETECTING PREGNANCY-ASSOCIATED PLASMA PROTEIN-A2 (PAPP-A2)

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention provides pregnancy associated plasma protein A2 (PAPP-A2), its nucleotide and amino acid sequences, antisense molecules to the nucleotide sequences which encode PAPP-A2, expression vectors for the production of purified PAPP-A2, antibodies capable of binding specifically...

  3. PREGNANCY-ASSOCIATED PLASMA PROTEIN-A2 (PAPP-A2) POLYNUCLEOTIDES

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention provides pregnancy associated plasma protein A2 (PAPP-A2), its nucleotide and amino acid sequences antisense molecules to the nucleotide sequences which encode PAPP-A2, expression vectors for the production of purified PAPP-A2, antibodies capable of binding specifically...

  4. Differential dissociation micromethod for the investigation of binding of metandrostenolone (Nerobol) to plasma proteins

    Energy Technology Data Exchange (ETDEWEB)

    Bojadzsieva, M.; Kocsar, L. (Orszagos Frederic Joliot-Curie Sugarbiologiai es Sugaregeszseguegyi Kutato Intezet, Budapest (Hungary)); Kremmer, T. (Orszagos Onkologiai Intezet, Budapest (Hungary))

    1985-01-01

    A micromethod was developed to determine the binding of anabolic steroids to plasma proteins. The new procedure combines precipitation with ammonium sulphate and differential dissociation. The binding parameters (association constant, specific binding capacity) are calculated on the basis of dissociation curves of sup(3)H-metandrostenolone from the precipitated sexual binding globuline.

  5. METHODS OF DETECTING PREGNANCY-ASSOCIATED PLASMA PROTEIN-A2 (PAPP-A2)

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention provides pregnancy associated plasma protein A2 (PAPP-A2), its nucleotide and amino acid sequences, antisense molecules to the nucleotide sequences which encode PAPP-A2, expression vectors for the production of purified PAPP-A2, antibodies capable of binding specifically...

  6. PLASMA PROTEIN PROFILING AS A HIGH THROUGHPUT TOOL FOR CHEMICAL SCREENING USING A SMALL FISH MODEL

    Science.gov (United States)

    Hudson, R. Tod, Michael J. Hemmer, Kimberly A. Salinas, Sherry S. Wilkinson, James Watts, James T. Winstead, Peggy S. Harris, Amy Kirkpatrick and Calvin C. Walker. In press. Plasma Protein Profiling as a High Throughput Tool for Chemical Screening Using a Small Fish Model (Abstra...

  7. Binding of von Willebrand factor and plasma proteins to the eggshell of Schistosoma mansoni

    NARCIS (Netherlands)

    Dewalick, Saskia; Hensbergen, Paul J; Bexkens, Michiel L; Grosserichter-Wagener, Christina; Hokke, Cornelis H; Deelder, André M; de Groot, Philip G; Tielens, Aloysius G M; van Hellemond, Jaap J

    2014-01-01

    Schistosoma mansoni eggs have to cross the endothelium and intestinal wall to leave the host and continue the life cycle. Mechanisms involved in this essential step are largely unknown. Here we describe direct binding to the S. mansoni eggshell of von Willebrand factor and other plasma proteins invo

  8. Binding of von Willebrand factor and plasma proteins to the eggshell of Schistosoma mansoni

    NARCIS (Netherlands)

    Dewalick, Saskia; Hensbergen, Paul J; Bexkens, Michiel L; Grosserichter-Wagener, Christina; Hokke, Cornelis H; Deelder, André M; de Groot, Philip G; Tielens, Aloysius G M; van Hellemond, Jaap J

    Schistosoma mansoni eggs have to cross the endothelium and intestinal wall to leave the host and continue the life cycle. Mechanisms involved in this essential step are largely unknown. Here we describe direct binding to the S. mansoni eggshell of von Willebrand factor and other plasma proteins

  9. Heterogeneity of Arabinogalactan-Proteins on the Plasma Membrane of Rose Cells.

    Science.gov (United States)

    Serpe, M. D.; Nothnagel, E. A.

    1996-11-01

    Arabinogalactan-proteins (AGPs) have been purified from the plasma membrane of suspension-cultured Paul's Scarlet rose (Rosa sp.) cells. The two most abundant and homogeneous plasma membrane AGP fractions were named plasma membrane AGP1 (PM-AGP1) and plasma membrane AGP2 (PM-AGP2) and had apparent molecular masses of 140 and 217 kD, respectively. Both PM-AGP1 and PM-AGP2 had [beta]-(1-3)-, [beta]-(1,6)-, and [beta]-(1,3,6)-galactopyranosyl residues, predominantly terminal [alpha]-arabinofuranosyl residues, and (1,4)- and terminal glucuronopyranosyl residues. The protein moieties of PM-AGP1 and PM-AGP2 were both rich in hydroxyproline, alanine, and serine, but differed in the abundance of hydroxyproline, which was 1.6 times higher in PM-AGP2 than in PM-AGP1. Another difference was the overall protein content, which was 3.7% (w/w) in PM-AGP1 and 15% in PM-AGP2. As judged by their behavior on reverse-phase chromatography, PM-AGP1 and PM-AGP2 were not more hydrophobic than AGPs from the cell wall or culture medium. In contrast, a minor plasma membrane AGP fraction eluted later on reverse-phase chromatography and was more negatively charged at pH 5 than either PM-AGP1 or PM-AGP2. The more negatively charged fraction contained molecules with a glycosyl composition characteristic of AGPs and included at least two different macromolecules. The results of this investigation indicate that Rosa plasma membrane contains at least four distinct AGPs or AGP-like molecules. These molecules differed from each other in size, charge, hydrophobicity, amino-acyl composition, and/or protein content.

  10. Determination of plasma protein synthesis index in liver diseases by means of /sup 75/Se-selenomethionine

    Energy Technology Data Exchange (ETDEWEB)

    Pirwitz, B. (Centrum Medyczne Ksztalcenia Podyplomowego, Warsaw (Poland))

    1979-01-01

    The investigations were carried out in 96 patients with chronic liver diseases and 44 controls. After intravenous administration of /sup 75/Se-selenomethionine plasma and plasma-protein radioactivity was measured at intervals of 2 hours. On the basis of these measurements the index of plasma protein synthesis rate was determined. It was found that the index of plasma protein synthesis in liver cirrhosis was significantly decreased in an overwhelming number of cases in relation to controls. On the other hand, in liver neoplasms this index was statistically significantly increased. It is possible that this fact will be used in future for differential diagnosis.

  11. Easy measurement of diffusion coefficients of EGFP-tagged plasma membrane proteins using k-space Image Correlation Spectroscopy

    DEFF Research Database (Denmark)

    Christensen, Eva Arnspang; Koffman, Jennifer Skaarup; Marlar, Saw

    2014-01-01

    Lateral diffusion and compartmentalization of plasma membrane proteins are tightly regulated in cells and thus, studying these processes will reveal new insights to plasma membrane protein function and regulation. Recently, k-Space Image Correlation Spectroscopy (kICS)1 was developed to enable ro...

  12. Pregnancy associated plasma protein-A (PAPP-A) is not a marker of the vulnerable atherosclerotic plaque

    DEFF Research Database (Denmark)

    Iversen, Kasper; Teisner, Ane; Dalager, Soren

    2011-01-01

    To investigate if pregnancy associated plasma protein-A (PAPP-A) was present in the vulnerable plaque, and if not, to find alternative hypothesis for the release of PAPP-A.......To investigate if pregnancy associated plasma protein-A (PAPP-A) was present in the vulnerable plaque, and if not, to find alternative hypothesis for the release of PAPP-A....

  13. Pregnancy-associated plasma protein-A, a marker for outcome in patients suspected for acute coronary syndrome

    DEFF Research Database (Denmark)

    Iversen, Kasper; Dalsgaard, Morten; Teisner, Ane S

    2010-01-01

    To examine if pregnancy-associated plasma protein-A (PAPP-A) in patients with chest pain, could identify patients at risk for death or myocardial infarction.......To examine if pregnancy-associated plasma protein-A (PAPP-A) in patients with chest pain, could identify patients at risk for death or myocardial infarction....

  14. Pregnancy-associated plasma protein-A, a marker for outcome in patients suspected for acute coronary syndrome

    DEFF Research Database (Denmark)

    Iversen, Kasper K; Dalsgaard, Morten; Teisner, Ane S;

    2010-01-01

    To examine if pregnancy-associated plasma protein-A (PAPP-A) in patients with chest pain, could identify patients at risk for death or myocardial infarction.......To examine if pregnancy-associated plasma protein-A (PAPP-A) in patients with chest pain, could identify patients at risk for death or myocardial infarction....

  15. Proteomic and carbonylation profile analysis of rat skeletal muscles following acute swimming exercise.

    Directory of Open Access Journals (Sweden)

    Francesca Magherini

    Full Text Available Previous studies by us and other groups characterized protein expression variation following long-term moderate training, whereas the effects of single bursts of exercise are less known. Making use of a proteomic approach, we investigated the effects of acute swimming exercise (ASE on protein expression and carbonylation patterns in two hind limb muscles: the Extensor Digitorum Longus (EDL and the Soleus, mostly composed of fast-twitch and slow-twitch fibres, respectively. Carbonylation is one of the most common oxidative modifications of proteins and a marker of oxidative stress. In fact, several studies suggest that physical activity and the consequent increase in oxygen consumption can lead to increase in reactive oxygen and nitrogen species (RONS production, hence the interest in examining the impact of RONS on skeletal muscle proteins following ASE. Results indicate that protein expression is unaffected by ASE in both muscle types. Unexpectedly, the protein carbonylation level was reduced following ASE. In particular, the analysis found 31 and 5 spots, in Soleus and EDL muscles respectively, whose carbonylation is reduced after ASE. Lipid peroxidation levels in Soleus were markedly reduced as well. Most of the decarbonylated proteins are involved either in the regulation of muscle contractions or in the regulation of energy metabolism. A number of hypotheses may be advanced to account for such results, which will be addressed in future studies.

  16. Proteome oxidative carbonylation during oxidative stress-induced premature senescence of WI-38 human fibroblasts

    DEFF Research Database (Denmark)

    Le Boulch, Marine; Ahmed, Emad K; Rogowska-Wrzesinska, Adelina

    2017-01-01

    Accumulation of oxidatively damaged proteins is a hallmark of cellular and organismal ageing, and is also a phenotypic feature shared by both replicative senescence and stress-induced premature senescence of human fibroblasts. Moreover, proteins that are building up as oxidized (i.e. the "Oxi......-proteome") during ageing and age-related diseases represent a restricted set of cellular proteins, indicating that certain proteins are more prone to oxidative carbonylation and subsequent intracellular accumulation. The occurrence of specific carbonylated proteins upon oxidative stress induced premature senescence...... of WI-38 human fibroblasts and their follow-up identification have been addressed in this study. Indeed, it was expected that the identification of these proteins would give insights into the mechanisms by which oxidatively damaged proteins could affect cellular function. Among these proteins, some...

  17. Coarse-grained model of adsorption of blood plasma proteins onto nanoparticles

    CERN Document Server

    Lopez, Hender

    2016-01-01

    We present a coarse-grained model for evaluation of interactions of globular proteins with nanoparticles. The protein molecules are represented by one bead per aminoacid and the nanoparticle by a homogeneous sphere that interacts with the aminoacids via a central force that depends on the nanoparticle size. The proposed methodology is used to predict the adsorption energies for six common human blood plasma proteins on hydrophobic charged or neutral nanoparticles of different sizes as well as the preferred orientation of the molecules upon adsorption. Our approach allows one to rank the proteins by their binding affinity to the nanoparticle, which can be used for predicting the composition of the NP-protein corona. The predicted ranking is in good agreement with known experimental data for protein adsorption on surfaces.

  18. Plasma membrane domains enriched in cortical endoplasmic reticulum function as membrane protein trafficking hubs.

    Science.gov (United States)

    Fox, Philip D; Haberkorn, Christopher J; Weigel, Aubrey V; Higgins, Jenny L; Akin, Elizabeth J; Kennedy, Matthew J; Krapf, Diego; Tamkun, Michael M

    2013-09-01

    In mammalian cells, the cortical endoplasmic reticulum (cER) is a network of tubules and cisterns that lie in close apposition to the plasma membrane (PM). We provide evidence that PM domains enriched in underlying cER function as trafficking hubs for insertion and removal of PM proteins in HEK 293 cells. By simultaneously visualizing cER and various transmembrane protein cargoes with total internal reflectance fluorescence microscopy, we demonstrate that the majority of exocytotic delivery events for a recycled membrane protein or for a membrane protein being delivered to the PM for the first time occur at regions enriched in cER. Likewise, we observed recurring clathrin clusters and functional endocytosis of PM proteins preferentially at the cER-enriched regions. Thus the cER network serves to organize the molecular machinery for both insertion and removal of cell surface proteins, highlighting a novel role for these unique cellular microdomains in membrane trafficking.

  19. Protein receptor-independent plasma membrane remodeling by HAMLET

    DEFF Research Database (Denmark)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.

    2015-01-01

    in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range...... of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a "receptor independent" transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET...... accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. Finally, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features...

  20. Protein receptor-independent plasma membrane remodeling by HAMLET

    DEFF Research Database (Denmark)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.;

    2015-01-01

    A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This "protein-centric" view is increasingly challenged by evidence for the involvement of specialized membrane domains...... in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range...... of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a "receptor independent" transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET...

  1. Heterogeneous interactome between Litopenaeus vannamei plasma proteins and Vibrio parahaemolyticus outer membrane proteins.

    Science.gov (United States)

    Liu, Xiang; She, Xin-Tao; Zhu, Qing-Feng; Li, Hui; Peng, Xuan-Xian

    2013-01-01

    A great loss has been suffered by microbial infectious diseases under intensive shrimp farming in recent years. In this background, the understanding of shrimp innate immunity becomes an importantly scientific issue, but little is known about the heterogeneous protein-protein interaction between pathogenic cells and hosts, which is a key step for the invading microbes to infect internet organs through bloodstream. In the present study, bacterial outer membrane (OM) protein array and pull-down approaches are used to isolate both Vibrio parahaemolyticus OM proteins that bind to shrimp serum proteins and the shrimp serum proteins that interact with bacterial cells, respectively. Three interacting shrimp serum proteins, hemocyanin, β-1,3-glucan binding protein and LV_HP_RA36F08r and thirty interacting OM proteins were determined. They form 63 heterogeneous protein-protein interactions. Nine out of the 30 OM proteins were randomly demonstrated to be up-regulated or down-regulated when bacterial cells were cultured with shrimp sera, indicating the biological significance of the network. The interesting findings uncover the complexity of struggle between host immunity and bacterial infection. Compared with our previous report on heterogeneous interactome between fish grill and bacterial OM proteins, the present study further extends the investigation from lower vertebrates to invertebrates and develops a bacterial OM protein array to identify the OM proteins bound with shrimp serum proteins, which elevates the frequencies of the bound OM proteins. Our results highlight the way to determine and understand the heterogeneous interaction between hosts and microbes.

  2. Capillary high-performance liquid chromatography/mass spectrometric analysis of proteins from affinity-purified plasma membrane.

    Science.gov (United States)

    Zhao, Yingxin; Zhang, Wei; White, Michael A; Zhao, Yingming

    2003-08-01

    Proteomics analysis of plasma membranes is a potentially powerful strategy for the discovery of proteins involved in membrane remodeling under diverse cellular environments and identification of disease-specific membrane markers. A key factor for successful analysis is the preparation of plasma membrane fractions with low contamination from subcellular organelles. Here we report the characterization of plasma membrane prepared by an affinity-purification method, which involves biotinylation of cell-surface proteins and subsequent affinity enrichment with strepavidin beads. Western blotting analysis showed this method was able to achieve a 1600-fold relative enrichment of plasma membrane versus mitochondria and a 400-fold relative enrichment versus endoplasmic reticulum, two major contaminants in plasma membrane fractions prepared by conventional ultracentrifugation methods. Capillary-HPLC/MS analysis of 30 microg of affinity-purified plasma membrane proteins led to the identification of 918 unique proteins, which include 16.4% integral plasma membrane proteins and 45.5% cytosol proteins (including 8.6% membrane-associated proteins). Notable among the identified membrane proteins include 30 members of ras superfamily, receptors (e.g., EGF receptor, integrins), and signaling molecules. The low number of endoplasmic reticulum and mitochondria proteins (approximately 3.3% of the total) suggests the plasma membrane preparation has minimum contamination from these organelles. Given the importance of integral membrane proteins for drug design and membrane-associated proteins in the regulation cellular behaviors, the described approach will help expedite the characterization of plasma membrane subproteomes, identify signaling molecules, and discover therapeutic membrane-protein targets in diseases.

  3. GPI-anchored proteins do not reside in ordered domains in the live cell plasma membrane

    Science.gov (United States)

    Sevcsik, Eva; Brameshuber, Mario; Fölser, Martin; Weghuber, Julian; Honigmann, Alf; Schütz, Gerhard J.

    2015-04-01

    The organization of proteins and lipids in the plasma membrane has been the subject of a long-lasting debate. Membrane rafts of higher lipid chain order were proposed to mediate protein interactions, but have thus far not been directly observed. Here we use protein micropatterning combined with single-molecule tracking to put current models to the test: we rearranged lipid-anchored raft proteins (glycosylphosphatidylinositol(GPI)-anchored-mGFP) directly in the live cell plasma membrane and measured the effect on the local membrane environment. Intriguingly, this treatment does neither nucleate the formation of an ordered membrane phase nor result in any enrichment of nanoscopic-ordered domains within the micropatterned regions. In contrast, we find that immobilized mGFP-GPIs behave as inert obstacles to the diffusion of other membrane constituents without influencing their membrane environment over distances beyond their physical size. Our results indicate that phase partitioning is not a fundamental element of protein organization in the plasma membrane.

  4. Prolonging the plasma circulation of proteins by nano-encapsulation with phosphorylcholine-based polymer

    Institute of Scientific and Technical Information of China (English)

    Linlin Zhang; Yang Liu; Gan Liu; Duo Xu; Sheng Liang; Xinyuan Zhu; Yunfeng Lu

    2016-01-01

    Short in vivo circulation is a major hindrance to the widespread adoption of protein therapeutics.Protein nanocapsules generated by encapsulating proteins with a thin layer of phosphorylcholine-based polymer via a two-step encapsulation process exhibited significantly prolonged plasma half-life.Furthermore,by constructing nanocapsules with similar sizes but different surface charges and chemistry,we demonstrated a generic strategy for prolonging the plasma half-life of therapeutic proteins.In an in vitro experiment,four types of bovine serum albumin (BSA) nanocapsules were incubated with fetal bovine serum (FBS) in phosphate buffer saline (PBS);the cell uptake by HeLa cells was monitored to systematically evaluate the characteristics of the surface chemistry during drculation.Single positron emission tomography-computed tomography (SPECT)was employed to allow real-time observation of the BSA nanoparticle distribution in vivo,as well as quantification of the plasma concentration after intravenous administration.This study offers a practical method for translating a broad range of proteins for clinical use.

  5. An early nodulin-like protein accumulates in the sieve element plasma membrane of Arabidopsis

    DEFF Research Database (Denmark)

    Khan, Junaid A.; Wang, Qi; Sjölund, Richard D.

    2007-01-01

    Membrane proteins within the sieve element-companion cell complex have essential roles in the physiological functioning of the phloem. The monoclonal antibody line RS6, selected from hybridomas raised against sieve elements isolated from California shield leaf (Streptanthus tortuosus; Brassicaceae......) tissue cultures, recognizes an antigen in the Arabidopsis (Arabidopsis thaliana) ecotype Columbia that is associated specifically with the plasma membrane of sieve elements, but not companion cells, and accumulates at the earliest stages of sieve element differentiation. The identity of the RS6 antigen...... from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both...

  6. The kinetics and mechanism of dissociation of metal carbonyls in high infrared laser fields

    Science.gov (United States)

    Langsam, Y.; Ronn, A. M.

    1981-01-01

    The method of laser induced dielectric breakdown has been used to study the dissociation of three metal carbonyls. Following non-resonant excitation by a TEA CO 2 laser, amorphous micro-metallic particles are deposited. The identity of the various species within a laser induced plasma for a series of related reactant molecules, has been determined using infrared and visible real time fluorescence techniques. Based upon the energetics of dissociation and the relative magnitudes of the rates of activation and deactivation for various collision partners of the plasma fluorescence, an energy transfer map as well as probable mechanisms are proposed. The kinetics of a variety of chemical reactions under plasma conditions are discussed in terms of elementary collision theory. It is shown that the internal vibration manifold of the metal carbonyls is equilibrated prior to the dissociation in agreement with the majority of laser induced multiphoton dissociation experiments.

  7. Prion removal capacity of plasma protein manufacturing processes: a data collection from PPTA member companies.

    Science.gov (United States)

    Cai, Kang; Gröner, Albrecht; Dichtelmüller, Herbert O; Fabbrizzi, Fabrizio; Flechsig, Eckhard; Gajardo, Rodrigo; von Hoegen, Ilka; Jorquera, Juan I; Kempf, Christoph; Kreil, Thomas R; Lee, Douglas C; Moscardini, Mila; Pölsler, Gerhard; Roth, Nathan J

    2013-09-01

    The variant Creutzfeldt-Jakob disease incidence peaked a decade ago and has since declined. Based on epidemiologic evidence, the causative agent, pathogenic prion, has not constituted a tangible contamination threat to large-scale manufacturing of human plasma-derived proteins. Nonetheless, manufacturers have studied the prion removal capabilities of various manufacturing steps to better understand product safety. Collectively analyzing the results could reveal experimental reproducibility and detect trends and mechanisms driving prion removal. Plasma Protein Therapeutics Association member companies collected more than 200 prion removal studies on plasma protein manufacturing steps, including precipitation, adsorption, chromatography, and filtration, as well as combined steps. The studies used a range of model spiking agents and bench-scale process replicas. The results were grouped based on key manufacturing variables to identify factors impacting removal. The log reduction values of a group are presented for comparison. Overall prion removal capacities evaluated by independent groups were in good agreement. The removal capacity evaluated using biochemical assays was consistent with prion infectivity removal measured by animal bioassays. Similar reduction values were observed for a given step using various spiking agents, except highly purified prion protein in some circumstances. Comparison between combined and single-step studies revealed complementary or overlapping removal mechanisms. Steps with high removal capacities represent the conditions where the physiochemical differences between prions and therapeutic proteins are most significant. The results support the intrinsic ability of certain plasma protein manufacturing steps to remove prions in case of an unlikely contamination, providing a safeguard to products. © 2012 American Association of Blood Banks.

  8. Whey protein supplementation increases methionine intake but not homocysteine plasma concentration in rats.

    Science.gov (United States)

    Deminice, Rafael; Comparotto, Hugo; Jordao, Alceu Afonso

    2015-01-01

    The purpose of this study was to examine the effects of whey protein supplementation on homocysteine (Hcy) metabolism and liver oxidative stress in rats. Twenty-four rats were divided into 3 groups (n = 8) to receive one of the following diets for 4 weeks: control diet (C), whey protein-composed diet (WP), and whey protein-supplemented diet (WPS). The C and WP diets consisted of AIN-93 with 20% casein and 20% whey protein as protein source, respectively. WPS was AIN-93 (20% casein) supplemented by the addition of 20% (w/w) whey protein. Four weeks of ingesting a WPS diet resulted in a significantly higher (P protein and methionine intakes. Although a significant increase (P protein products, known liver oxidative stress markers, were increased in the WPS group compared with the C group. In addition, no change in glutathione liver concentration was observed in any of the groups studied. In conclusion, whey protein supplementation increases methionine intake substantially; however, it does not change plasma Hcy concentrations. On the other hand, increased hepatic oxidative stress markers were observed in whey protein supplemented rats were probably due to high protein intake.

  9. Plasma urea nitrogen and progesterone concentrations and follicular dynamics in ewes fed proteins of different degradability

    Directory of Open Access Journals (Sweden)

    Gustavo Bianchi Lazarin

    2012-07-01

    Full Text Available The effects of overfeeding with protein of different degradability on body condition, plasma urea nitrogen and progesterone concentrations, ovulation number and follicular dynamics were assessed in Santa Ines ewes. Twelve ewes were assigned to a randomized block design according to body weight and received overfeeding with soybean meal or with corn gluten meal or maintenance diet for 28 days before ovulation and during the next estrous cycle. Blood samples were taken on days 7, 14, 21, and 28 after the beginning of treatments for analysis of plasma urea nitrogen and on days 3, 6, 9, 12, and 15 into the estrous cycle for analysis of plasma urea nitrogen and progesterone. Follicular dynamics was monitored daily by ultrasound during one estrous cycle. Dry matter and crude protein intake, weight gain, plasma urea nitrogen concentration before ovulation, number of ovulations, diameter of the largest follicle of the 1st and of the 2nd waves and the growth rate of the largest follicle of the 1st wave were higher in the ewes that received overfeeding. The growth rate of the largest follicle of the 3rd wave was higher in the ewes fed maintenance diet. The back fat thickness, plasma urea nitrogen before ovulation and progesterone concentrations, diameter of the largest follicle of the 2nd wave and growth rate of the largest follicle of the 3rd wave were higher in ewes that received overfeeding with soybean meal. The growth rate of the largest follicle of the 1st wave was higher in ewes that received overfeeding with corn gluten meal. Overfeeding with protein-rich feeds may increase the ovulation number and with soybean meal, it may be effective in increasing plasma progesterone concentration in ewes.

  10. Application of plasma-polymerized films for isoelectric focusing of proteins in a capillary electrophoresis chip.

    Science.gov (United States)

    Tsai, Shuo-Wen; Loughran, Michael; Hiratsuka, Atsunori; Yano, Kazuyoshi; Karube, Isao

    2003-03-01

    The first use of plasma polymerization technique to modify the surface of a glass chip for capillary isoelectric focusing (cIEF) of different proteins is reported. The electrophoresis separation channel was machined in Tempax glass chips with length 70 mm, 300 microm width and 100 microm depth. Acetonitrile and hexamethyldisiloxane monomers were used for plasma polymerization. In each case 100 nm plasma polymer films were coated onto the chip surface to reduce protein wall adsorption and minimize the electroosmotic flow. Applied voltages of 1000 V, 2000 V and 3000 V were used to separate mixtures of cytochrome c (pI 9.6), hemoglobin (pI 7.0) and phycocyanin (pI 4.65). Reproducible isoelectric focusing of each pI marker protein was observed in different coated capillaries at increasing concentration 2.22-5 microg microL(-1). Modification of the glass capillary with hydrophobic HMDS plasma polymerized films enabled rapid cIEF within 3 min. The separation efficiency of cytochrome c and phycocyanin in both acrylamide and HMDS coated capillaries corresponded to a plate number of 19600 which compares favourably with capillary electrophoresis of neurotransmitters with amperometric detection.

  11. Plasma Protein Profiles Differ Between Women Diagnosed with Cervical Intraepithelial Neoplasia (CIN 1 and 3

    Directory of Open Access Journals (Sweden)

    Edward E. Partridge

    2006-01-01

    Full Text Available Early detection of precancerous cells in the cervix and their clinical management is the main purpose of cervical cancer prevention and treatment programs. Cytological findings or testing for high risk (HR-human papillomavirus (HPV are inadequately sensitive for use in triage of women at high risk for cervical cancer. The current study is an exploratory study to identify candidate surface-enhanced laser desorption/ionization (SELDI time of flight (TOF mass spectrometry (MS protein profiles in plasma that may distinguish cervical intraepithelial neoplasia (CIN 3 from CIN 1 among women infected with HR-HPV. We evaluated the SELDI-TOF-MS plasma protein profiles of HR-HPV positive 32 women with CIN 3 (cases and 28 women with CIN1 (controls. Case-control status was kept blinded and triplicates of each sample and quality control plasma samples were randomized and after robotic sample preparations were run on WCX2 chips. After alignment of mass/charge (m-z values, an iterative method was used to develop a classifier on a training data set that had 28 cases and 22 controls. The classifier developed was used to classify the subjects in a test data set that has six cases and six controls. The classifier separated the cases from controls in the test set with 100% sensitivity and 100% specificity suggesting the possibility of using plasma SELDI protein profiles to identify women who are likely to have CIN 3 lesions.

  12. A Comparison of Blood Factor XII Autoactivation in Buffer, Protein Cocktail, Serum, and Plasma Solutions

    Science.gov (United States)

    Golas, Avantika; Yeh, Chyi-Huey Josh; Pitakjakpipop, Harit; Siedlecki, Christopher A.; Vogler, Erwin A.

    2012-01-01

    Activation of blood plasma coagulation in vitro by contact with material surfaces is demonstrably dependent on plasma-volume-to-activator-surface-area ratio. The only plausible explanation consistent with current understanding of coagulation-cascade biochemistry is that procoagulant stimulus arising from the activation complex of the intrinsic pathway is dependent on activator surface area. And yet, it is herein shown that activation of the blood zymogen factor XII (Hageman factor, FXII) dissolved in buffer, protein cocktail, heat-denatured serum, and FXI deficient plasma does not exhibit activator surface-area dependence. Instead, a highly-variable burst of procoagulant-enzyme yield is measured that exhibits no measurable kinetics, sensitivity to mixing, or solution-temperature dependence. Thus, FXII activation in both buffer and protein-containing solutions does not exhibit characteristics of a biochemical reaction but rather appears to be a “mechanochemical” reaction induced by FXII molecule interactions with hydrophilic activator particles that do not formally adsorb blood proteins from solution. Results of this study strongly suggest that activator surface-area dependence observed in contact activation of plasma coagulation does not solely arise at the FXII activation step of the intrinsic pathway. PMID:23117212

  13. Protein composition in human plasma after long-term orbital missions and in rodent plasma after spaceflights on biosatellites "Cosmos-1887" and "Cosmos-2044".

    Science.gov (United States)

    Larina, O N

    1991-02-01

    The two-dimensional plasma protein map of crewmembers of long-duration "Mir" expeditions obtained the day after the recovery shows a manifold increase in the content of several proteins normally seen in trace amounts. The emergence of several unusual protein spots occurs as well, some of them probably due to charge shifts provided by the events influencing posttranslational modification processes. By the 8 postflight day these phenomena were disappeared. In the "Cosmos-1887" biosatellite experiment, the plasma samples obtained two days after the landing as well as plasma of synchronous animals exhibited the higher fibrinogen levels when compared to those of vivarium animals. The protein consisting of a number of fractions with molecular weight of 50 to 60 kD and pI 5 to 6 had protein spots of similar size in flight and synchronous animals while in vivarium rats one of the spots was larger in size as opposed to the others. The plasma protein spectrum of flight and synchronous groups of animals in "Cosmos-1887" experiment where plasma samples were prepared in the period of time from 5 to 10 hours after spaceflight coincided with the pattern of vivarium animals. The data suggest that the protein changes described above develop during postflight period and accelerations, vibrations, readaptation to 1 G gravity, emotional stress could be the cause of these alterations.

  14. Ovulation-inducing factor: a protein component of llama seminal plasma

    Directory of Open Access Journals (Sweden)

    Huanca Wilfredo

    2010-05-01

    Full Text Available Abstract Background Previously, we documented the presence of ovulation-inducing factor (OIF in the seminal plasma of llamas and alpacas. The purpose of the study was to define the biochemical characteristics of the molecule(s in seminal plasma responsible for inducing ovulation. Methods In Experiment 1, llama seminal plasma was centrifuged using filtration devices with nominal molecular mass cut-offs of 30, 10 and 5 kDa. Female llamas (n = 9 per group were treated i.m. with whole seminal plasma (positive control, phosphate-buffered saline (negative control, or the fraction of seminal plasma equal or higher than 30 kDa, 10 to 30 kDa, 5 to 10 kDa, or Results In Experiment 1, all llamas in the equal or higher than 30 kDa and positive control groups ovulated (9/9 in each, but none ovulated in the other groups (P Conclusions We conclude that ovulation-inducing factor (OIF in llama seminal plasma is a protein molecule that is resistant to heat and enzymatic digestion with proteinase K, and has a molecular mass of approximately equal or higher than 30 kDa.

  15. Carbonyl compounds indoors in a changing climate

    Directory of Open Access Journals (Sweden)

    Brimblecombe Peter

    2012-03-01

    Full Text Available Abstract Background Formic acid, acetic acid and formaldehyde are important compounds in the indoor environment because of the potential for these acids to degrade calcareous materials (shells, eggs, tiles and geological specimens, paper and corrode or tarnish metals, especially copper and lead. Carbonyl sulfide tarnishes both silver and copper encouraging the formation of surface sulfides. Results Carbonyls are evolved more quickly at higher temperatures likely in the Cartoon Gallery at Knole, an important historic house near Sevenoaks in Kent, England where the study is focused. There is a potential for higher concentrations to accumulate. However, it may well be that in warmer climates they will be depleted more rapidly if ventilation increases. Conclusions Carbonyls are likely to have a greater impact in the future.

  16. Triton X-114 cloud point extraction to subfractionate blood plasma proteins for two-dimensional gel electrophoresis.

    Science.gov (United States)

    Jessen, Flemming; Wulff, Tune

    2015-09-15

    A simple and reproducible procedure for enrichment of a plasma protein subfraction suitable for two-dimensional polyacrylamide gel electrophoresis (2DE) was developed, using a Triton X-114-based cloud point extraction (CPE). Appropriate conditions for such a CPE procedure were found by SDS-PAGE to be a plasma protein concentration of about 10mg/ml in 3% (w/v) Triton X-114. 2DE of proteins obtained by CPE of 400 μl of human plasma revealed about 200 spots constituting a spot pattern very different from the pattern of total plasma. The CPE procedure only had a limited contribution to the technical variation. Identification of about 60 spots, representing only 22 proteins, revealed that several proteins in the obtained subfraction were present in more isoforms or modifications. Among these were apolipoproteins (A-1, D, E, L1, and M), haptoglobin-related protein, phosphatidylcholine-sterol acyltransferase, serum amyloid A, and serum paraoxonase/arylesterase 1, which are proteins of a hydrophobic nature, as in plasma they relate to lipoprotein particles. Thus, Triton X-114-based CPE is a simple plasma prefractionation tool, attractive for detailed 2DE studies of hydrophobic plasma proteins and their isoforms or modifications.

  17. Plasma membrane protein trafficking in plant-microbe interactions: a plant cell point of view

    Directory of Open Access Journals (Sweden)

    Nathalie eLeborgne-Castel

    2014-12-01

    Full Text Available In order to ensure their physiological and cellular functions, plasma membrane (PM proteins must be properly conveyed from their site of synthesis, i.e. the endoplasmic reticulum, to their final destination, the PM, through the secretory pathway. PM protein homeostasis also relies on recycling and/or degradation, two processes that are initiated by endocytosis. Vesicular membrane trafficking events to and from the PM have been shown to be altered when plant cells are exposed to mutualistic or pathogenic microbes. In this review, we will describe the fine-tune regulation of such alterations, and their consequence in PM protein activity. We will consider the formation of intracellular perimicrobial compartments, the PM protein trafficking machinery of the host, and the delivery or retrieval of signaling and transport proteins such as pattern-recognition receptors, producers of reactive oxygen species, and sugar transporters.

  18. Proteomic analysis identifies interleukin 11 regulated plasma membrane proteins in human endometrial epithelial cells in vitro

    Directory of Open Access Journals (Sweden)

    Stanton Peter G

    2011-05-01

    Full Text Available Abstract Background During the peri-implantation period, the embryo adheres to an adequately prepared or receptive endometrial surface epithelium. Abnormal embryo adhesion to the endometrium results in embryo implantation failure and infertility. Endometrial epithelial cell plasma membrane proteins critical in regulating adhesion may potentially be infertility biomarkers or targets for treating infertility. Interleukin (IL 11 regulates human endometrial epithelial cells (hEEC adhesion. Its production is abnormal in women with infertility. The objective of the study was to identify IL11 regulated plasma membrane proteins in hEEC in vitro using a proteomic approach. Methods Using a 2D-differential in-gel electrophoresis (DIGE electrophoresis combined with LCMS/MS mass spectrometry approach, we identified 20 unique plasma membrane proteins differentially regulated by IL11 in ECC-1 cells, a hEEC derived cell line. Two IL11 regulated proteins with known roles in cell adhesion, annexin A2 (ANXA2 and flotillin-1 (FLOT1, were validated by Western blot and immunocytochemistry in hEEC lines (ECC-1 and an additional cell line, Ishikawa and primary hEEC. Flotilin-1 was further validated by immunohistochemistry in human endometrium throughout the menstrual cycle (n = 6-8/cycle. Results 2D-DIGE analysis identified 4 spots that were significantly different between control and IL11 treated group. Of these 4 spots, there were 20 proteins that were identified with LCMS/MS. Two proteins; ANXA2 and FLOT1 were chosen for further analyses and have found to be significantly up-regulated following IL11 treatment. Western blot analysis showed a 2-fold and a 2.5-fold increase of ANXA2 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. Similarly, a 1.8-fold and a 2.3/2.4-fold increase was also observed for FLOT1 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. In vitro, IL11 induced stronger ANXA2 expression on cell surface of primary h

  19. Longitudinal changes in C-reactive protein, proform of eosinophil major basic protein, and pregnancy-associated plasma protein-A during weight changes in obese children

    DEFF Research Database (Denmark)

    Lausten-Thomsen, Ulrik; Gamborg, Michael; Bøjsøe, Christine

    2015-01-01

    BACKGROUND: Childhood obesity is associated with several complications, including cardiovascular comorbidity. Several biomarkers, such as high-sensitive C-reactive protein (hs-CRP), proform of eosinophil major basic protein (Pro-MBP) and pregnancy associated plasma protein-A (PAPP-A), have equally...... been linked to increased cardiovascular susceptibility. This study investigates these biomarkers during weight loss and regain in obese children. MATERIALS AND METHODS: A longitudinal study during a 12-week weight loss program with a 28 months follow-up was conducted. Anthropometrics and plasma......), and 2.70 (girls) were included. Ninety children completed the weight loss program and 68 children entered the follow-up program. Pro-MBP and PAPP-A, but not hs-CRP, exhibited individual-specific levels (tracking) during weight loss and regain. The PAPP-A/Pro-MBP correlation was strong, whereas the hs...

  20. Proteomic Analysis of Rice Plasma Membrane-associated Proteins in Response to Chitooligosaccharide Elicitors

    Institute of Scientific and Technical Information of China (English)

    Fang Chen; Qun Li; Zuhua He

    2007-01-01

    Chitooligomers or chitooligosaccharides (COS) are elicitors that bind to the plasma membrane (PM) and elicit various defense responses. However, the PM-bound proteins involved in elicitor-mediated plant defense responses still remain widely unknown. In order to get more information about PM proteins involved in rice defense responses, we conducted PM proteomic analysis of the rice suspension cells elicited by COS. A total of 14 up- or down-regulated protein spots were observed on 2-D gels of PM fractions at 12 h and 24 h after COS incubation. Of them, eight protein spots were successfully identified by MS (mass spectrography) and predicted to be associated to the PM and function in plant defense, including a putative PKN/PRK1 protein kinase, a putative pyruvate kinase isozyme G, a putative zinc finger protein, a putative MAR-binding protein MFP1, and a putative calcium-dependent protein kinase. Interestingly, a COS-induced pM5-like protein was identified for the first time in plants, which is a trans-membrane nodal modulator in transforming growth factor-β(TGFβ) signaling in vertebrates. We also identified two members of a rice polyprotein family, which were up-regulated by COS. Our study would provide a starting point for functionality of PM proteins in the rice basal defense.

  1. Systematic study of plasma and serum proteins in the pig; Etude systematique des proteines plasmatiques et seriques du porc

    Energy Technology Data Exchange (ETDEWEB)

    Daburon, F.; Nizza, P.; Hatchikian, C.; Schmidt, J.-P. [Commissariat a l' Energie Atomique (France)

    1966-07-01

    This work has been carried out in the framework of the determination of the physiological constants of a normal pig. The aim was to study the serum and plasma proteins of this animal species, the ultimate object being to discover whether the qualitative and quantitative changes in these proteins can make a significant contribution to the establishment of a biological dosimetry for irradiated pigs. The serum and plasma from a normal pig were analyzed first by various simple electrophoretic methods and then by immuno-electrophoresis. As a result of the particular characteristics of pig serum we have gradually been led to make numerous modifications to the techniques used for human serums or for those of small laboratory animals. Much careful work and patience were required in order to obtain reproducible results. (authors) [French] Ce travail se situe dans le cadre de la determination des constantes physiologiques du porc normal. il s'agissait de proceder a l'etude des proteines seriques et plasmatiques de cette espece animale, le but ulterieur etant de savoir si les modifications qualitatives et quantitatives de ces proteines pourront representer une contribution valable a l'etablissement d'une dosimetrie biologique chez le porc irradie. Le serum et le plasma du porc normal ont ete analyses d'abord par diverses methodes electrophoretiques simples puis par immunoelectrophorese. Les caracteristiques particulieres du serum de porc nous ont conduits a apporter progressivement de nombreuses modifications aux techniques utilisees pour des serums humains ou de petits animaux de laboratoire. L'obtention de resultats reproductible a exige beaucoup de patience et de minutie. (auteurs)

  2. Seminal plasma protein profiles of ejaculates obtained by internal artificial vagina and electroejaculation in Brahman bulls.

    Science.gov (United States)

    Rego, J P A; Moura, A A; Nouwens, A S; McGowan, M R; Boe-Hansen, G B

    2015-09-01

    The present study was conducted to investigate if differences exist in the seminal plasma protein profile from mature Brahman bulls using two methods of semen collection: internal artificial vagina (IAV) and electroejaculation (EEJ). Semen was collected four times from three bulls on the same day and parameters were assessed immediately post-collection. Seminal plasma proteins were evaluated by 2-D fluorescence difference gel electrophoresis and identified by mass spectrometry. Semen volume was greater (P < 0.05) for EEJ (4.6 ± 0.35 mL) than for IAV (1.86 ± 0.24 mL) but sperm concentration was greater in IAV (1505 ± 189 × 10(6) sperm/mL) than in EEJ samples (344 ± 87 × 10(6) sperm/mL). Sperm motility and the percentage of normal sperm were not different between treatments. Total concentration of seminal plasma proteins was greater for samples collected by IAV as compared to EEJ (19.3 ± 0.9 compared with 13.0 ± 1.8 mg/mL, P < 0.05; respectively). Based on 2-D gels, 22 spots had a greater volume (P < 0.05) in gels derived from IAV samples, corresponding to 21 proteins identified as transferrin, albumin, epididymal secretory glutathione peroxidase, among others. Thirty-three spots, corresponding to 26 proteins, had a greater volume (P < 0.05) in gels derived from EEJ samples. These proteins were identified as spermadhesin-1, Bovine Sperm Protin 1, 3 and 5 isoforms, angiogenin-1, alpha-1B-glycoprotein, clusterin, nucleobindin-1, cathepsins, spermadhesin Z13, annexins, among others. Thus, proteins in greater amounts in samples obtained by IAV and EEJ were mainly of epididymal origin and accessory sex glands, respectively. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Effect of Bovine Plasma Protein on Autolysis and Gelation of Protein Extracted from Giant Squid (Dosidicus gigas Mantle

    Directory of Open Access Journals (Sweden)

    Laura Raquel Marquez-Alvarez

    2015-01-01

    Full Text Available The effect of bovine plasma protein (BPP on the inhibition of autolytic activity and its effect on the gelling properties of a protein concentrate (PC obtained from jumbo squid (Dosidicus gigas mantle were investigated. Sols and gels were prepared from the PC by adding different amounts of BPP (0, 1, and 2%. Dynamic oscillatory measurements indicated that systems with 1% BPP had a higher elastic modulus (G′, in which hydrophobic interactions were favored. Concerning the technological and textural quality of the gels, BPP caused a greater water holding capacity (WHC, force, cohesiveness, and elasticity, probably due to improvement of the electrostatic and hydrophobic interactions during gel formation. Scanning electron microscopy (SEM allowed visualization of the formation of more rigid and ordered gels with less porosity when BPP was added. Therefore, the addition of BPP improved the gelling capacity of proteins extracted from giant squid.

  4. Joint associations of blood plasma proteins with overwinter survival of a large mammal.

    Science.gov (United States)

    Garnier, Romain; Cheung, Christopher K; Watt, Kathryn A; Pilkington, Jill G; Pemberton, Josephine M; Graham, Andrea L

    2017-02-01

    In many wild animal populations, hosts are at risk of parasites and malnutrition and resource costs of defence may be difficult to afford. We postulate that proteins, important in homeostasis and immunity, play a complex but central role in condition dependence and resource costs of mammalian immune defence. To test this, we measured plasma concentrations of albumin, total proteins. Self-reactive antibodies and parasite-specific IgG in female Soay sheep. Using a principal component analysis, we found a new metric of condition reflecting individual variation in acquisition, assimilation and/or recycling of plasma proteins that predicted overwinter survival. Controlling for this metric, an age-dependent trade-off between antibody titres and protein reserves emerged, indicating costs of mounting an antibody response: younger individuals survived best when prioritising immunity while older individuals fared better when maintaining high-protein nutritional plane. These findings suggest fascinating roles for protein acquisition and allocation in influencing survival in wild animal populations.

  5. Fetuin-B, a liver-derived plasma protein is essential for fertilization.

    Science.gov (United States)

    Dietzel, Eileen; Wessling, Jennifer; Floehr, Julia; Schäfer, Cora; Ensslen, Silke; Denecke, Bernd; Rösing, Benjamin; Neulen, Joseph; Veitinger, Thomas; Spehr, Marc; Tropartz, Tanja; Tolba, René; Renné, Thomas; Egert, Angela; Schorle, Hubert; Gottenbusch, Yuliya; Hildebrand, André; Yiallouros, Irene; Stöcker, Walter; Weiskirchen, Ralf; Jahnen-Dechent, Willi

    2013-04-15

    The zona pellucida (ZP) is a glycoprotein matrix surrounding mammalian oocytes. Upon fertilization, ZP hardening prevents sperm from binding to and penetrating the ZP. Here, we report that targeted gene deletion of the liver-derived plasma protein fetuin-B causes premature ZP hardening and, consequently, female infertility. Transplanting fetuin-B-deficient ovaries into wild-type recipients restores fertility, indicating that plasma fetuin-B is necessary and sufficient for fertilization. In vitro fertilization of oocytes from fetuin-B-deficient mice only worked after rendering the ZP penetrable by laser perforation. Mechanistically, fetuin-B sustains fertility by inhibiting ovastacin, a cortical granula protease known to trigger ZP hardening. Thus, plasma fetuin-B is necessary to restrain protease activity and thereby maintain ZP permeability until after gamete fusion. These results also show that premature ZP hardening can cause infertility in mice.

  6. Simulated gastrointestinal digestion, intestinal permeation and plasma protein interaction of white, green, and black tea polyphenols.

    Science.gov (United States)

    Tenore, Gian Carlo; Campiglia, Pietro; Giannetti, Daniela; Novellino, Ettore

    2015-02-15

    The gastrointestinal digestion, intestinal permeation, and plasma protein interaction of polyphenols from a single tea cultivar at different stages of processing (white, green, and black teas) were simulated. The salivary phase contained 74.8-99.5% of native polyphenols, suggesting potential bioavailability of significant amounts of antioxidants through the oral mucosal epithelium that might be gastric sensitive and/or poorly absorbed in the intestine. White tea had the highest content and provided the best intestinal bioaccessibility and bioavailability for catechins. Since most of native catechins were not absorbed, they were expected to accumulate in the intestinal lumen where a potential inhibition capacity of cellular glucose and cholesterol uptake was assumed. The permeated catechins (approximately, 2-15% of intestinal levels) significantly bound (about 37%) to plasma HDLs, suggesting a major role in cholesterol metabolism. White tea and its potential nutraceuticals could be effective in the regulation of plasma glucose and cholesterol levels.

  7. Increased fasting plasma acylation-stimulating protein concentrations in nephrotic syndrome.

    Science.gov (United States)

    Ozata, Metin; Oktenli, Cagatay; Gulec, Mustafa; Ozgurtas, Taner; Bulucu, Fatih; Caglar, Kayser; Bingol, Necati; Vural, Abdulgaffar; Ozdemir, I Caglayan

    2002-02-01

    Acylation-stimulating protein (ASP) is an adipocyte-derived protein that has recently been suggested to play an important role in the regulation of lipoprotein metabolism and triglyceride (TG) storage. ASP also appears to have a role in the regulation of energy balance. In addition to its role as a hormonal regulator of body weight and energy expenditure, leptin is now implicated as a regulatory molecule in lipid metabolism. However, little is known about the alterations in fasting plasma ASP and leptin concentrations in the nephrotic syndrome. As hyperlipidemia is one of the most striking manifestations of the nephrotic syndrome, we have investigated fasting plasma ASP and leptin levels and their relation to lipid levels in this syndrome. Twenty-five patients with untreated nephrotic syndrome and 25 age-, sex-, and body mass index-matched healthy controls were included in the study. Fasting plasma lipoproteins, TG, total cholesterol, lipoprotein(a), apolipoprotein AI (apoAI), apoB, urinary protein, plasma albumin, third component of complement (C3), ASP, and leptin levels were measured in both groups. Total cholesterol, TG, low and very low density lipoproteins, lipoprotein(a), apoB, and urinary protein levels were increased in the patient group, whereas plasma albumin, high density lipoprotein cholesterol, and apoAI levels were decreased compared with those in the control group (P Fasting ASP concentrations showed no correlation with body mass index, proteinuria, plasma albumin, leptin, or any lipid parameter in either group, but C3 levels (in patient group: r(s) = 0.92; P < 0.001; in control group: r(s) = 0.68; P < 0.001). Our findings showed that plasma ASP levels were significantly elevated, whereas leptin levels were normal in the nephrotic syndrome. Increased ASP levels in the setting of dyslipidemia in the nephrotic syndrome raise the possibility of an ASP receptor defect in adipocytes, which also suggests the existence of so-called ASP resistance. Moreover

  8. Diclofenac plasma protein binding: PK-PD modelling in cardiac patients submitted to cardiopulmonary bypass

    Directory of Open Access Journals (Sweden)

    Auler Jr. J.O.

    1997-01-01

    Full Text Available Twenty-four surgical patients of both sexes without cardiac, hepatic, renal or endocrine dysfunctions were divided into two groups: 10 cardiac surgical patients submitted to myocardial revascularization and cardiopulmonary bypass (CPB, 3 females and 7 males aged 65 ± 11 years, 74 ± 16 kg body weight, 166 ± 9 cm height and 1.80 ± 0.21 m2 body surface area (BSA, and control, 14 surgical patients not submitted to CPB, 11 female and 3 males aged 41 ± 14 years, 66 ± 14 kg body weight, 159 ± 9 cm height and 1.65 ± 0.16 m2 BSA (mean ± SD. Sodium diclofenac (1 mg/kg, im Voltaren 75® twice a day was administered to patients in the Recovery Unit 48 h after surgery. Venous blood samples were collected during a period of 0-12 h and analgesia was measured by the visual analogue scale (VAS during the same period. Plasma diclofenac levels were measured by high performance liquid chromatography. A two-compartment open model was applied to obtain the plasma decay curve and to estimate kinetic parameters. Plasma diclofenac protein binding decreased whereas free plasma diclofenac levels were increased five-fold in CPB patients. Data obtained for analgesia reported as the maximum effect (EMAX were: 25% VAS (CPB vs 10% VAS (control, P<0.05, median measured by the visual analogue scale where 100% is equivalent to the highest level of pain. To correlate the effect versus plasma diclofenac levels, the EMAX sigmoid model was applied. A prolongation of the mean residence time for maximum effect (MRTEMAX was observed without any change in lag-time in CPB in spite of the reduced analgesia reported for these patients, during the time-dose interval. In conclusion, the extent of plasma diclofenac protein binding was influenced by CPB with clinically relevant kinetic-dynamic consequences

  9. Phosphorylation-dependent Trafficking of Plasma Membrane Proteins in Animal and Plant Cells

    Institute of Scientific and Technical Information of China (English)

    Remko Offringa; and Fang Huang

    2013-01-01

    In both unicellular and multicellular organisms, transmembrane (TM) proteins are sorted to and retained at specific membrane domains by endomembrane trafficking mechanisms that recognize sorting signals in the these proteins. The trafficking and distribution of plasma membrane (PM)-localized TM proteins (PM proteins), especially of those PM proteins that show an asymmetric distribution over the PM, has received much attention, as their proper PM localization is crucial for elementary signaling and transport processes, and defects in their localization often lead to severe disease symptoms or developmental defects. The subcellular localization of PM proteins is dynamically regulated by post-translational modifications, such as phosphorylation and ubiquitination. These modificaitons mostly occur on sorting signals that are located in the larger cytosolic domains of the cargo proteins. Here we review the effects of phosphorylation of PM proteins on their trafficking, and present the key examples from the animal field that have been subject to studies for already several decades, such as that of aquaporin 2 and the epidermal growth factor receptor. Our knowledge on cargo trafficking in plants is largely based on studies of the family of PIN FORMED (PIN) carriers that mediate the efflux of the plant hormone auxin. We will review what is known on the subcellular distribution and trafficking of PIN proteins, with a focus on how this is modulated by phosphorylation, and identify and discuss analogies and differences in trafficking with the well-studied animal examples.

  10. Phosphorylation-dependent trafficking of plasma membrane proteins in animal and plant cells.

    Science.gov (United States)

    Offringa, Remko; Huang, Fang

    2013-09-01

    In both unicellular and multicellular organisms, transmembrane (TM) proteins are sorted to and retained at specific membrane domains by endomembrane trafficking mechanisms that recognize sorting signals in the these proteins. The trafficking and distribution of plasma membrane (PM)-localized TM proteins (PM proteins), especially of those PM proteins that show an asymmetric distribution over the PM, has received much attention, as their proper PM localization is crucial for elementary signaling and transport processes, and defects in their localization often lead to severe disease symptoms or developmental defects. The subcellular localization of PM proteins is dynamically regulated by post-translational modifications, such as phosphorylation and ubiquitination. These modificaitons mostly occur on sorting signals that are located in the larger cytosolic domains of the cargo proteins. Here we review the effects of phosphorylation of PM proteins on their trafficking, and present the key examples from the animal field that have been subject to studies for already several decades, such as that of aquaporin 2 and the epidermal growth factor receptor. Our knowledge on cargo trafficking in plants is largely based on studies of the family of PIN FORMED (PIN) carriers that mediate the efflux of the plant hormone auxin. We will review what is known on the subcellular distribution and trafficking of PIN proteins, with a focus on how this is modulated by phosphorylation, and identify and discuss analogies and differences in trafficking with the well-studied animal examples.

  11. Ion-exchange chromatography used to isolate a spermadhesin-related protein from domestic goat (Capra hircus) seminal plasma.

    Science.gov (United States)

    Teixeira, Dárcio Italo Alves; Melo, Luciana Magalhães; Gadelha, Carlos Alberto de Almeida; Cunha, Rodrigo Maranguape Silva da; Bloch, Carlos; Rádis-Baptista, Gandhi; Cavada, Benildo Sousa; Freitas, Vicente José de Figueirêdo

    2006-03-31

    Mammalian seminal plasma contains among others, proteins called spermadhesins, which are the major proteins of boar and stallion seminal plasma. These proteins appear to be involved in capacitation and sperm-egg interaction. Previously, we reported the presence of a protein related to spermadhesins in goat seminal plasma. In the present study, we have further characterized this protein, and we propose ion-exchange chromatography to isolate this seminal protein. Semen was obtained from four adult Saanen bucks. Seminal plasma was pooled, dialyzed against distilled water and freeze-dried. Lyophilized proteins were loaded onto an ion-exchange chromatography column. Dialyzed-lyophilized proteins from the main peak of DEAE-Sephacel were applied to a C2/C18 column coupled to an RP-HPLC system, and the eluted proteins were lyophilized for electrophoresis. The N-terminal was sequenced and amino acid sequence similarity was determined using CLUSTAL W. Additionally, proteins from DEAE-Sephacel chromatography step were dialyzed and submitted to a heparin-Sepharose high-performance liquid chromatography. Goat seminal plasma after ion-exchange chromatography yielded 6.47 +/- 0.63 mg (mean +/- SEM) of the major retained fraction. The protein was designated BSFP (buck seminal fluid protein). BSFP exhibited N-terminal sequence homology to boar, stallion and bull spermadhesins. BSFP showed no heparin-binding capabilities. These results together with our previous data indicate that goat seminal plasma contains a protein that is structurally related to proteins of the spermadhesin family. Finally, this protein can be efficiently isolated by ion-exchange and reverse-phase chromatography.

  12. Plasma treatment of paper for protein immobilization on paper-based chemiluminescence immunodevice.

    Science.gov (United States)

    Zhao, Mei; Li, Huifang; Liu, Wei; Guo, Yumei; Chu, Weiru

    2016-05-15

    A novel protein immobilization method based on plasma treatment of paper on the low-cost paper-based immunodevice was established in this work. By using a benchtop plasma cleaner, the paper microzone was treated by oxygen plasma treatment for 4 min and then the antibody can be directly immobilized on the paper surface. Aldehyde group was produced after the plasma treatment, which can be verified from the fourier transform infrared spectroscopy (FT-IR) spectra and x-ray photoelectron spectroscopy (XPS) spectra. By linked to aldehyde group, the antibody can be immobilized on the paper surface without any other pretreatment. A paper-based immunodevice was introduced here through this antibody immobilization method. With sandwich chemiluminescence (CL) immunoassay method, the paper-based immunodevice was successfully performed for carcinoembryonic antigen (CEA) detection in human serum with a linear range of 0.1-80.0 ng/mL. The detection limit was 0.03 ng/mL, which was 30 times lower than the clinical CEA level. Comparing to the other protein immobilization methods on paper-based device, this strategy was faster and simpler and had potential applications in point-of-care testing, public health and environmental monitoring.

  13. Fibrinogen γ' increases the sensitivity to activated protein C in normal and factor V Leiden plasma.

    Science.gov (United States)

    Omarova, Farida; Uitte de Willige, Shirley; Simioni, Paolo; Ariëns, Robert A S; Bertina, Rogier M; Rosing, Jan; Castoldi, Elisabetta

    2014-08-28

    Activated protein C (APC) resistance, often associated with the factor V (FV) Leiden mutation, is the most common risk factor for venous thrombosis. We observed increased APC resistance in carriers of fibrinogen γ gene (FGG) haplotype 2, which is associated with reduced levels of the alternatively spliced fibrinogen γ' chain. This finding prompted us to study the effects of fibrinogen and its γ' chain on APC resistance. Fibrinogen, and particularly the γA/γ' isoform, improved the response of plasma to added APC in the thrombin generation-based assay. Similarly, a synthetic peptide mimicking the C-terminus of the fibrinogen γ' chain, which binds thrombin and inhibits its activities, greatly increased the APC sensitivity of normal and FV Leiden plasma, likely due to its ability to inhibit thrombin-mediated activation of FV and FVIII. Although the fibrinogen γ' peptide also inhibited protein C activation by the thrombin/thrombomodulin complex, it still increased the sensitivity of plasma to endogenously formed APC when thrombin generation was measured in the presence of soluble thrombomodulin. We conclude that fibrinogen, and particularly fibrinogen γ', increases plasma APC sensitivity. The fibrinogen γ' peptide might form the basis for pharmacologic interventions to counteract APC resistance. © 2014 by The American Society of Hematology.

  14. Isolation and characterization of the plasma hyaluronan-binding protein (PHBP) gene (HABP2).

    Science.gov (United States)

    Sumiya, J; Asakawa, S; Tobe, T; Hashimoto, K; Saguchi, K; Choi-Miura, N H; Shimizu, Y; Minoshima, S; Shimizu, N; Tomita, M

    1997-11-01

    PHBP is a novel human plasma hyaluronan-binding protein that shows significant homology in amino acid sequence to hepatocyte growth factor activator. Two overlapping clones that encode the human plasma hyaluronan-binding protein (PHBP) gene (HABP2) were isolated and characterized. The PHBP gene spans 35 kb and is composed of 13 exons from 37 to 1,394 bp in size with consensus splice sites. The gene's regulatory sequences contain putative promoter elements, but no typical TATA box. Some exons of this gene showed significant similarities to those of coagulation factor XII, tissue-type plasminogen activator, and urokinase genes in nucleotide length and in intron phasing. We also report the chromosome mapping of this gene by fluorescence in situ hybridization (FISH) using a genomic DNA fragment as a probe. The PHBP gene (HABP2) was located on chromosome 10q25-q26.

  15. Cysteine-rich secretory protein 3 is a ligand of alpha1B-glycoprotein in human plasma

    DEFF Research Database (Denmark)

    Udby, Lene; Sørensen, Ole E; Pass, Jesper

    2004-01-01

    Human cysteine-rich secretory protein 3 (CRISP-3; also known as SGP28) belongs to a family of closely related proteins found in mammals and reptiles. Some mammalian CRISPs are known to be involved in the process of reproduction, whereas some of the CRISPs from reptiles are neurotoxin......-like substances found in lizard saliva or snake venom. Human CRISP-3 is present in exocrine secretions and in secretory granules of neutrophilic granulocytes and is believed to play a role in innate immunity. On the basis of the relatively high content of CRISP-3 in human plasma and the small size of the protein...... (28 kDa), we hypothesized that CRISP-3 in plasma was bound to another component. This was supported by size-exclusion chromatography and immunoprecipitation of plasma proteins. The binding partner was identified by mass spectrometry as alpha(1)B-glycoprotein (A1BG), which is a known plasma protein...

  16. Cargo proteins of plasma astrocyte-derived exosomes in Alzheimer's disease.

    Science.gov (United States)

    Goetzl, Edward J; Mustapic, Maja; Kapogiannis, Dimitrios; Eitan, Erez; Lobach, Irina V; Goetzl, Laura; Schwartz, Janice B; Miller, Bruce L

    2016-11-01

    Efficient intercellular transfer of RNAs, proteins, and lipids as protected exosomal cargo has been demonstrated in the CNS, but distinct physiologic and pathologic roles have not been well defined for this pathway. The capacity to isolate immunochemically human plasma neuron-derived exosomes (NDEs), containing neuron-specific cargo, has permitted characterization of CNS-derived exosomes in living humans. Constituents of the amyloid β-peptide (Aβ)42-generating system now are examined in 2 distinct sets of human neural cells by quantification in astrocyte-derived exosomes (ADEs) and NDEs, enriched separately from plasmas of patients with Alzheimer's disease (AD) or frontotemporal dementia (FTD) and matched cognitively normal controls. ADE levels of β-site amyloid precursor protein-cleaving enzyme 1 (BACE-1), γ-secretase, soluble Aβ42, soluble amyloid precursor protein (sAPP)β, sAPPα, glial-derived neurotrophic factor (GDNF), P-T181-tau, and P-S396-tau were significantly (3- to 20-fold) higher than levels in NDEs for patients and controls. BACE-1 levels also were a mean of 7-fold higher in ADEs than in NDEs from cultured rat type-specific neural cells. Levels of BACE-1 and sAPPβ were significantly higher and of GDNF significantly lower in ADEs of patients with AD than in those of controls, but not significantly different in patients with FTD than in controls. Abundant proteins of the Aβ42 peptide-generating system in ADEs may sustain levels in neurons. ADE cargo proteins may be useful for studies of mechanisms of cellular interactions and effects of BACE-1 inhibitors in AD.-Goetzl, E. J., Mustapic, M., Kapogiannis, D., Eitan, E., Lobach, I. V., Goetzl, L., Schwartz, J. B., Miller, B. L. Cargo proteins of plasma astrocyte-derived exosomes in Alzheimer's disease.

  17. Hypochlorite-induced oxidation of proteins in plasma: formation of chloramines and nitrogen-centred radicals and their role in protein fragmentation.

    Science.gov (United States)

    Hawkins, C L; Davies, M J

    1999-06-01

    Activated phagocyte cells generate hypochlorite (HOCl) via the release of H2O2 and the enzyme myeloperoxidase. Plasma proteins are major targets for HOCl, although little information is available about the mechanism(s) of oxidation. In this study the reaction of HOCl (at least 50 microM) with diluted fresh human plasma has been shown to generate material that oxidizes 5-thio-2-nitrobenzoic acid; these oxidants are believed to be chloramines formed from the reaction of HOCl with protein amine groups. Chloramines have also been detected with isolated plasma proteins treated with HOCl. In both cases chloramine formation accounts for approx. 20-30% of the added HOCl. These chloramines decompose in a time-dependent manner when incubated at 20 or 37 degrees C but not at 4 degrees C. Ascorbate and urate remove these chloramines in a time- and concentration-dependent manner, with the former being more efficient. The reaction of fresh diluted plasma with HOCl also gives rise to protein-derived nitrogen-centred radicals in a time- and HOCl-concentration-dependent manner; these have been detected by EPR spin trapping. Identical radicals have been detected with isolated HOCl-treated plasma proteins. Radical formation was inhibited by excess methionine, implicating protein-derived chloramines (probably from lysine side chains) as the radical source. Plasma protein fragmentation occurs in a time- and HOCl-concentration-dependent manner, as evidenced by the increased mobility of the EPR spin adducts, the detection of further radical species believed to be intermediates in protein degradation and the loss of the parent protein bands on SDS/PAGE. Fragmentation can be inhibited by methionine and other agents (ascorbate, urate, Trolox C or GSH) capable of removing chloramines and reactive radicals. These results are consistent with protein-derived chloramines, and the radicals derived from them, as contributing agents in HOCl-induced plasma protein oxidation.

  18. Osteoblast response to oxygen functionalised plasma polymer surfaces

    CERN Document Server

    Kelly, J M

    2001-01-01

    Thin organic films with oxygen-carbon functionalities were deposited from plasmas containing vapour of the small organic compounds: allyI alcohol, methyl vinyl ketone and acrylic acid with octadiene. Characterisation of the deposits was carried out using X-ray photoelectron spectroscopy, in conjunction with chemical derivatisation, and this showed that plasma polymers retained high levels of original monomer functionality when the plasmas were sustained at low power for a given monomer vapour flow rate. High levels of attachment of rat osteosarcoma (ROS 17/2.8) cells were observed on surfaces that had high concentrations of hydroxyl and carbonyl functionalities and intermediate concentrations of carboxyl functionality. Cells did not attach to the octadiene plasma polymer. Cell attachment to carboxyl and methyl functionalised self-assembled monolayers increased with increasing concentration of surface carboxyl groups. Adsorption of the extracellular matrix protein fibronectin to acrylic acid/octadiene plasma c...

  19. Plasma bactericidal/permeability-increasing protein concentrations in critically ill children with the sepsis syndrome.

    Science.gov (United States)

    Wong, H R; Doughty, L A; Wedel, N; White, M; Nelson, B J; Havrilla, N; Carcillo, J A

    1995-12-01

    Bactericidal/permeability-increasing protein (BPI) is a neutrophil azurophilic granule component that is bactericidal towards Gram-negative bacteria and inhibits lipopolysaccharide-mediated inflammatory responses. We conducted a prospective study to measure plasma BPI concentrations in 36 critically ill children with and without the sepsis syndrome. Plasma BPI concentrations ranged from 0.5 to 452 ng/ml. Patients with the sepsis syndrome had higher median plasma BPI concentrations than critically ill controls (5.1 vs. 1.8 ng/ml, P = 0.006). Patients with organ system failure had higher median plasma BPI concentrations than those with no organ system failure (4.5 vs. 1.3 ng/ml, P = 0.001). Plasma BPI concentrations were positively associated with pediatric risk of mortality score (P = 0.03, rs = 0.4). These data provide the first clinical insights regarding the role of endogenous BPI production in critically ill children and suggest that BPI may play an important role in host defenses.

  20. DCCD inhibits protein translocation into plasma membrane vesicles from Escherichia coli at two different steps.

    OpenAIRE

    1987-01-01

    In vitro translocation of periplasmic and outer membrane proteins into inverted plasma membrane vesicles from Escherichia coli was completely prevented by the H+-ATPase inhibitor N,N'-dicyclohexylcarbodiimide (DCCD). DCCD was inhibitory to both co- and post-translational translocations, suggesting an involvement of the H+-translocating F1F0-ATPase in either mode of transport. This was verified by (i) the dependence of efficient co-translational translocation upon a low salt, i.e. F1-containin...

  1. Altered Plasma Profile of Antioxidant Proteins as an Early Correlate of Pancreatic β Cell Dysfunction.

    Science.gov (United States)

    Kuo, Taiyi; Kim-Muller, Ja Young; McGraw, Timothy E; Accili, Domenico

    2016-04-29

    Insulin resistance and β cell dysfunction contribute to the pathogenesis of type 2 diabetes. Unlike insulin resistance, β cell dysfunction remains difficult to predict and monitor, because of the inaccessibility of the endocrine pancreas, the integrated relationship with insulin sensitivity, and the paracrine effects of incretins. The goal of our study was to survey the plasma response to a metabolic challenge in order to identify factors predictive of β cell dysfunction. To this end, we combined (i) the power of unbiased iTRAQ (isobaric tag for relative and absolute quantification) mass spectrometry with (ii) direct sampling of the portal vein following an intravenous glucose/arginine challenge (IVGATT) in (iii) mice with a genetic β cell defect. By so doing, we excluded the effects of peripheral insulin sensitivity as well as those of incretins on β cells, and focused on the first phase of insulin secretion to capture the early pathophysiology of β cell dysfunction. We compared plasma protein profiles with ex vivo islet secretome and transcriptome analyses. We detected changes to 418 plasma proteins in vivo, and detected changes to 262 proteins ex vivo The impairment of insulin secretion was associated with greater overall changes in the plasma response to IVGATT, possibly reflecting metabolic instability. Reduced levels of proteins regulating redox state and neuronal stress markers, as well as increased levels of coagulation factors, antedated the loss of insulin secretion in diabetic mice. These results suggest that a reduced complement of antioxidants in response to a mixed secretagogue challenge is an early correlate of future β cell failure.

  2. A Preliminary Study of Trace Elements in Plasma Protein by Gel Chromatography Combined with SXRF

    Institute of Scientific and Technical Information of China (English)

    NIANQINGLIU; DEFUCHEN; 等

    1999-01-01

    Fractions of plasma protein of male Kunming mice (body weight 24.2±0.3g),treated with Cisplatin i.p.injection in dose of 10mg/kg,were obtained by separation on Sephadex-G-50 columns,buffered with ammonium acetate to pH5.7,The XSRF experiments were performed at the BEPC(Beijing Electron Positron Collider)synchrotron radiation facility.The elements(Pt,S,Ca,Fe,Ni,Cu,Zn,Se,Br and Sr)in the fraction of the plasma proteins(>22KD) were assayed using highly sensitive SXRF.The relative concentrations of elements were calculated by a normalization of COmpton scattering intensity around 22 KeV,after the normalization for collecting time of X-ray spectrum and the counting of the ion chamber,and subracting the contribution of the polycarbonate film used for supporting the samples.The determination could prove that the element Pt in plasma was bound with macro-molecularprotein.Cu and S were present in the fraction of the protein in mice treated with Cisplatin and exhibited an increase,the ration of treated/control were 1.66±0.06 and 1.78±0.33 repectively,whereas Zn decreased to a ratio of 0.78±0.09,Our results are in agreement with others which showed that Cisplatin exposure leads to a marked loss of kidney copper,and a moderate rise in didney zinc.However,this work mainly focussed on the implementation of this analytical procedure,but not on the results of the investigations of the effect of Cisplatin on trace elements in plasma protein.

  3. Sex steroid binding proteins in the plasma of hatchling Chelonia mydas.

    Science.gov (United States)

    Ikonomopoulou, M P; Ibrahim, K; Bradley, A J

    2008-09-01

    Sex steroid binding proteins were identified in hatchling female and male Chelonia mydas by dialysis and steady-state gel electrophoresis when examined at 4 degrees C. A testosterone binding protein with high binding affinity (K (a) = 0.98 +/- 0.5 x 10(8) M(-1)) and low to moderate binding capacity (B (max) = 7.58 +/- 4.2 x 10(-5) M) was observed in male hatchlings. An oestradiol binding protein with high affinity (K (a) = 0.35 +/- 1.8 x 10(8) M(-1)) and low to moderate binding capacity (B (max) = 0.16 +/- 0.5 x 10(-4) M) was identified in female hatchlings. This study confirmed that sex steroid binding proteins (SSBPs) become inactivate in both sexes at 36 degrees C, the maximum body temperature of sea turtle hatchlings at emergence. The inactivation of SSBPs at this temperature indicates that sex steroid hormones circulate freely in the body of the green turtles and are biologically available in the blood plasma. This observation is consistent with female and male hatchling C. mydas having different physiological (hormonal) and developmental requirements around the time of emergence. Moreover, concurrently conducted competition studies showed that sex steroids including testosterone and oestradiol do compete for binding sites in both male and female C. mydas hatchling plasma. Competition also occurred between testosterone and dihydrotestosterone for binding sites in the male C. mydas plasma. However, competition studies in the plasma of female hatchling C. mydas demonstrate that oestrone does not compete with oestradiol for binding sites.

  4. Fluorescence Enhancement of Fluorescein Isothiocyanate-Labeled Protein A Caused by Affinity Binding with Immunoglobulin G in Bovine Plasma

    Directory of Open Access Journals (Sweden)

    Kiyotaka Sakai

    2009-10-01

    Full Text Available Fluorescence enhancement of fluorescein isothiocyanate-labeled protein A (FITC-protein A caused by the binding with immunoglobulin G (IgG in bovine plasma was studied. FITC-protein A was immobilized onto a glass surface by covalent bonds. An increase in fluorescence intensity was dependent on IgG concentration ranging from 20 to 78 μg/mL in both phosphate buffer saline and bovine plasma. This method requires no separation procedure, and the reaction time is less than 15 min. A fluorescence enhancement assay by the affinity binding of fluorescence-labeled reagent is thus available for the rapid determination of biomolecules in plasma.

  5. Interaction of gold and silver nanoparticles with human plasma: Analysis of protein corona reveals specific binding patterns.

    Science.gov (United States)

    Lai, Wenjia; Wang, Qingsong; Li, Lumeng; Hu, Zhiyuan; Chen, Jiankui; Fang, Qiaojun

    2017-04-01

    Determining how nanomaterials interact with plasma will assist in understanding their effects on the biological system. This work presents a systematic study of the protein corona formed from human plasma on 20nm silver and gold nanoparticles with three different surface modifications, including positive and negative surface charges. The results show that all nanoparticles, even those with positive surface modifications, acquire negative charges after interacting with plasma. Approximately 300 proteins are identified on the coronas, while 99 are commonly found on each nanomaterial. The 20 most abundant proteins account for over 80% of the total proteins abundance. Remarkably, the surface charge and core of the nanoparticles, as well as the isoelectric point of the plasma proteins, are found to play significant roles in determining the nanoparticle coronas. Albumin and globulins are present at levels of less than 2% on these nanoparticle coronas. Fibrinogen, which presents in the plasma but not in the serum, preferably binds to negatively charged gold nanoparticles. These observations demonstrate the specific plasma protein binding pattern of silver and gold nanoparticles, as well as the importance of the surface charge and core in determining the protein corona compositions. The potential downstream biological impacts of the corona proteins were also investigated.

  6. Fish protein hydrolysate elevates plasma bile acids and reduces visceral adipose tissue mass in rats

    DEFF Research Database (Denmark)

    Liaset, Bjørn; Madsen, Lise; Hao, Qin

    2009-01-01

    Conjugation of bile acids (BAs) to the amino acids taurine or glycine increases their solubility and promotes liver BA secretion. Supplementing diets with taurine or glycine modulates BA metabolism and enhances fecal BA excretion in rats. However, it is still unclear whether dietary proteins...... varying in taurine and glycine contents alter BA metabolism, and thereby modulate the recently discovered systemic effects of BAs. Here we show that rats fed a diet containing saithe fish protein hydrolysate (saithe FPH), rich in taurine and glycine, for 26 days had markedly elevated fasting plasma BA...

  7. [Aspergillus ochraceus myxomycetes produce extracellular proteinases--protein C activators of blood plasma].

    Science.gov (United States)

    Osmolovskiĭ, A A; Kreĭer, V G; Kurakov, A V; Baranova, N A; Egorov, N S

    2012-01-01

    Natural isolates of Aspergillus ochraceus myxomycetes from soil and plant remains from various regions have been screened. The isolated strains were characterized by similar cultural and morphological features and an identical nucleotide sequence in the ITS1-5,8S-ITS2 region of rDNA. The ability of the extracellular proteinases of A. ochraceus myxomycetes to activate protein C of blood plasma has been established. Differences are revealed in the accumulation of proteinases activating protein C and proteinases with thrombin- and plasmin-like activities in the growth dynamics of producers.

  8. Comparison of two different plasma surface-modification techniques for the covalent immobilization of protein monolayers.

    Science.gov (United States)

    Cifuentes, Anna; Borrós, Salvador

    2013-06-04

    The immobilization of biologically active species is crucial for the fabrication of smart bioactive surfaces. For this purpose, plasma polymerization is frequently used to modify the surface nature without affecting the bulk properties of the material. Thus, it is possible to create materials with surface functional groups that can promote the anchoring of all kinds of biomolecules. Different methodologies in protein immobilization have been developed in recent years, although some drawbacks are still not solved, such as the difficulties that some procedures involve and/or the denaturalization of the protein due to the immobilization process. In this work, two different strategies to covalently attach bovine serum albumin (BSA) protein are developed. Both techniques are compared in order to understand how the nature of the surface modification affects the conformation of the protein upon immobilization.

  9. [Mechanisms of human plasma proteins adsorption on the surface of perfluorocarbon emulsion stabilized with proxanol 268].

    Science.gov (United States)

    Zhalimov, V K; Sklifas, A N; Kukushkin, N I

    2012-01-01

    It has been shown that sorption of most proteins with the molecular weight lower than 200 kDa from human blood plasma on the surface of perfluorocarbon emulsion, stabilized with proxanol 268, is mainly based on hydrophobic interaction, whereas sorption of immunoglobulin G is mainly the result of electrostatic interaction. The removal of lipidic components from plasma leads to the increase of a total amount of adsorbed proteins by 35%. Particularly, when lipidic components are removed, sorption of apolipoprotein AI and immunoglobulin G is considerably bettered as well as sorption of other proteins with the molecular weight of about 50 and 60 kDa occurs. It has been out that apolipoprotein AI in the adsorbed condition loses its capability of tryptophan fluorescence, which might be probably determined by the quenching influence of the perfluorocarbon core of nanoparticle. We think that the findings obtained also indicates considerable conformational rearrangements of this protein during adsorption. It was shown, that the fluorescence of proteins with sorption on nanoparticles in emulsion based on the hydrophobic interaction, is completely or partially quenched.

  10. A direct, automated, immuno-turbidimetric assay of free protein S antigen in plasma.

    Science.gov (United States)

    Deffert, C; Esteve, F; Grimaux, M; Gouault-Heilmann, M

    2001-03-01

    A new, fully automated, one-step, immuno-turbidimetric assay of free protein S (fPS) in plasma (STA Liatest Free Protein S; Diagnostica Stago, Asnières, France) has been developed for STA analysers. This technique combines the advantages of a direct assay of fPS using two monoclonal antibodies, which specifically recognize fPS but not protein S (PS)-C4b-binding protein complexes, and the advantages of automation. The assay has good analytical performances, with intra- and inter-assay variation coefficients below 5% for normal values, and slightly higher for abnormal values. In a comparison study with a one-step enzyme-linked immunosorbent assay for fPS (Asserachrom Free Protein S; Diagnostica Stago), a correlation coefficient of 0.93 with a regression line close to 1 was found between the two techniques (n = 166 normal or PS-deficient plasma samples collected from healthy subjects and individuals with a personal or family history of thrombosis). This new technique is specific, reproducible, easy to perform, and provides a useful tool in the diagnosis of PS deficiency.

  11. Forward transport of proteins in the plasma membrane of migrating cerebellar granule cells.

    Science.gov (United States)

    Wang, Dong; She, Liang; Sui, Ya-nan; Yuan, Xiao-bing; Wen, Yunqing; Poo, Mu-ming

    2012-12-18

    Directional flow of membrane components has been detected at the leading front of fibroblasts and the growth cone of neuronal processes, but whether there exists global directional flow of plasma membrane components over the entire migrating neuron remains largely unknown. By analyzing the trajectories of antibody-coated single quantum dots (QDs) bound to two membrane proteins, overexpressed myc-tagged synaptic vesicle-associated membrane protein VAMP2 and endogenous neurotrophin receptor TrkB, we found that these two proteins exhibited net forward transport, which is superimposed upon Brownian motion, in both leading and trailing processes of migrating cerebellar granule cells in culture. Furthermore, no net directional transport of membrane proteins was observed in nonmigrating cells with either growing or stalling leading processes. Analysis of the correlation of motion direction between two QDs on the same process in migrating neurons also showed a higher frequency of correlated forward than rearward movements. Such correlated QD movements were markedly reduced in the presence of myosin II inhibitor blebbistatin,suggesting the involvement of myosin II-dependent active transport processes. Thus, a net forward transport of plasma membrane proteins exists in the leading and trailing processes of migrating neurons, in line with the translocation of the soma.

  12. Development of reduced fat minced meats using inulin and bovine plasma proteins as fat replacers.

    Science.gov (United States)

    Rodriguez Furlán, Laura T; Padilla, Antonio Pérez; Campderrós, Mercedes E

    2014-02-01

    This work deals with the effect of the addition of inulin and bovine plasma proteins as fat replacers, on the quality of minced meat. The proteins are obtained by ultrafiltration and freeze-drying. The following determinations were carried out: chemical composition, sensorial analysis (color, flavor, taste and consistency), emulsion stability and instrumental texture analysis of samples. The resulting formulations were compared with full-fat minced meat, as control. The results showed an increase of protein contents after fat replacement, while a fat reduction of 20-35% produced light products enriched with proteins and inulin as the functional ingredient. No change was observed in color, flavor, or taste among the samples. However, the sensory analysis showed that the combination of plasma protein (2.5%w/w) and inulin (2%w/w) had the best acceptability with respect to consistency, and had a lower fat drain from the emulsion. Texture profile analysis revealed that this formulation assimilated the control texture properties, being that this result is required for adequate consumer acceptance.

  13. The importance of selecting a proper biological milieu for protein corona analysis in vitro: Human plasma versus human serum.

    Science.gov (United States)

    Mirshafiee, Vahid; Kim, Raehyun; Mahmoudi, Morteza; Kraft, Mary L

    2016-06-01

    Nanoparticle (NP) exposure to biological fluids in the body results in protein binding to the NP surface, which forms a protein coating that is called the "protein corona". To simplify studies of protein-NP interactions and protein corona formation, NPs are incubated with biological solutions, such as human serum or human plasma, and the effects of this exposure are characterized in vitro. Yet, how NP exposure to these two different biological milieus affects protein corona composition and cell response has not been investigated. Here, we explore the differences between the protein coronas that form when NPs are incubated in human serum versus human plasma. NP characterization indicated that NPs that were exposed to human plasma had higher amounts of proteins bound to their surfaces, and were slightly larger in size than those exposed to human serum. In addition, significant differences in corona composition were also detected with gel electrophoresis and liquid chromatography-mass spectrometry/mass spectrometry, where a higher fraction of coagulation proteins and complement factors were found on the plasma-exposed NPs. Flow cytometry and confocal microscopy showed that the uptake of plasma-exposed NPs was higher than that of serum-exposed NPs by RAW 264.7 macrophage immune cells, but not by NIH 3T3 fibroblast cells. This difference is likely due to the elevated amounts of opsonins, such as fibrinogen, on the surfaces of the NPs exposed to plasma, but not serum, because these components trigger NP internalization by immune cells. As the human plasma better mimics the composition of the in vivo environment, namely blood, in vitro protein corona studies should employ human plasma, and not human serum, so the biological phenomena that is observed is more similar to that occurring in vivo.

  14. The effect of polycarboxylate shell of magnetite nanoparticles on protein corona formation in blood plasma

    Science.gov (United States)

    Szekeres, Márta; Tóth, Ildikó Y.; Turcu, R.; Tombácz, Etelka

    2017-04-01

    The development of protein corona around nanoparticles upon administration to the human body is responsible in a large part for their biodistribution, cell-internalization and toxicity or biocompatibility. We studied the influence of the chemical composition of polyelectrolyte shells (citric acid (CA) and poly(acrylic-co-maleic acid) (PAM)) of core-shell magnetite nanoparticles (MNPs) on the evolution of protein corona in human plasma (HP). The aggregation state and zeta potential of the particles were measured in the range of HP concentration between 1 and 80 (v/v)% 3 min and 20 h after dispersing the particles in HP diluted with Tris buffered saline. Naked MNPs aggregated in HP solution, but the carboxylated MNPs became stabilized colloidally at higher plasma concentrations. Significant differences were observed at low plasma concentration. CA@MNPs aggregated instantly while the hydrodynamic diameter of PAM@MNP increased only slightly at 1-3 v/v % HP concentrations. The observed differences in protein corona formation can be explained by the differences in the steric effects of the polycarboxylate shells. It is interesting that relatively small but systematic changes in zeta potential alter the aggregation state significantly.

  15. Effects of Aminoguanidine on Lipid and Protein Oxidation in Diabetic Rat Kidneys

    OpenAIRE

    Yavuz, Dilek Gogas; Küçükkaya, Belgin; Ersöz, H. önder; Yalçin, A. Süha; Emerk, Kaya; Akalin, Sema

    2002-01-01

    Nonenzymatic glycation of tissue and plasma proteins may stimulate the production of oxidant and carbonyl stress in diabetes. The aim of this study was to evaluate the effects of aminoguanidine (AG) on lipid peroxidation, protein oxidation and nitric oxide (NO) release in diabetic rat kidneys. After induction of diabetes with streptozotocin, female Wistar rats were divided into 2 groups. Group DAG (n=9) rats were given AG hydrogen carbonate (1 g/L) in drinking water and group D (n=8) was diab...

  16. Millimeter wave spectra of carbonyl cyanide ⋆

    Science.gov (United States)

    Bteich, S.B.; Tercero, B.; Cernicharo, J.; Motiyenko, R.A.; Margulès, L.; Guillemin, J.-C.

    2016-01-01

    Context More than 30 cyanide derivatives of simple organic molecules have been detected in the interstellar medium, but only one dicarbonitrile has been found and that very recently. There is still a lack of high-resolution spectroscopic data particularly for dinitriles derivatives. The carbonyl cyanide molecule is a new and interesting candidate for astrophysical detection. It could be formed by the reaction of CO and CN radicals, or by substitution of the hydrogen atom by a cyano group in cyanoformaldehyde, HC(=O)CN, that has already been detected in the interstellar medium. Aims The available data on the rotational spectrum of carbonyl cyanide is limited in terms of quantum number values and frequency range, and does not allow accurate extrapolation of the spectrum into the millimeter-wave range. To provide a firm basis for astrophysical detection of carbonyl cyanide we studied its millimeter-wave spectrum. Methods The rotational spectrum of carbonyl cyanide was measured in the frequency range 152 - 308 GHz and analyzed using Watson’s A- and S-reduction Hamiltonians. Results The ground and first excited state of v5 vibrational mode were assigned and analyzed. More than 1100 distinct frequency lines of the ground state were fitted to produce an accurate set of rotational and centrifugal distortion constants up to the eighth order. The frequency predictions based on these constants should be accurate enough for astrophysical searches in the frequency range up to 500 GHz and for transition involving energy levels with J ≤ 100 and Ka ≤ 42. Based on the results we searched for interstellar carbonyl cyanide in available observational data without success. Thus, we derived upper limits to its column density in different sources. PMID:27738349

  17. Remodeling of the plasma membrane in preparation for sperm-egg recognition: roles of acrosomal proteins.

    Science.gov (United States)

    Tanphaichitr, Nongnuj; Kongmanas, Kessiri; Kruevaisayawan, Hathairat; Saewu, Arpornrad; Sugeng, Clarissa; Fernandes, Jason; Souda, Puneet; Angel, Jonathan B; Faull, Kym F; Aitken, R John; Whitelegge, Julian; Hardy, Daniel; Berger, Trish; Baker, Mark

    2015-01-01

    The interaction of sperm with the egg's extracellular matrix, the zona pellucida (ZP) is the first step of the union between male and female gametes. The molecular mechanisms of this process have been studied for the past six decades with the results obtained being both interesting and confusing. In this article, we describe our recent work, which attempts to address two lines of questions from previous studies. First, because there are numerous ZP binding proteins reported by various researchers, how do these proteins act together in sperm-ZP interaction? Second, why do a number of acrosomal proteins have ZP affinity? Are they involved mainly in the initial sperm-ZP binding or rather in anchoring acrosome reacting/reacted spermatozoa to the ZP? Our studies reveal that a number of ZP binding proteins and chaperones, extracted from the anterior sperm head plasma membrane, coexist as high molecular weight (HMW) complexes, and that these complexes in capacitated spermatozoa have preferential ability to bind to the ZP. Zonadhesin (ZAN), known as an acrosomal protein with ZP affinity, is one of these proteins in the HMW complexes. Immunoprecipitation indicates that ZAN interacts with other acrosomal proteins, proacrosin/acrosin and sp32 (ACRBP), also present in the HMW complexes. Immunodetection of ZAN and proacrosin/acrosin on spermatozoa further indicates that both proteins traffic to the sperm head surface during capacitation where the sperm acrosomal matrix is still intact, and therefore they are likely involved in the initial sperm-ZP binding step.

  18. Spectrophotometric determination of total proteins in blood plasma: a comparative study among dye-binding methods

    Directory of Open Access Journals (Sweden)

    Dimas Augusto Morozin Zaia

    2005-05-01

    Full Text Available A comparative study between the biuret method (standard method for total proteins and spectrophotometric methods using dyes (Bradford, 3',3",5',5"-tetrabromophenolphthalein ethyl ester-TBPEE, and erythrosin-B was carried out for the determination of total proteins in blood plasma from rats. Bradford method showed the highest sensitivity for proteins and biuret method showed the lowest. For all the methods, the absorbance for different proteins (BSA, casein, and egg albumin was measured and Bradford method showed the lowest variation of absorbance. The concentration of total protein obtained by using Bradford method was not statistically different (p>0.05 from concentration of total protein obtained by the biuret method. But in regard to erythrosin-B and TBPEE methods the concentrations of total protein were statistically different (pA determinação de proteínas totais em plasma sangüíneo é importante em diversas áreas de pesquisa. Um estudo comparativo entre o método de biureto (método padrão para proteínas totais e diversos métodos que utilizam corantes (Bradford, tetrabromofenolftaleína etil éster-TBPEE, e eritrosina-B foi realizado para a determinação de proteínas totais em plasma sangüíneo de ratos. O método de Bradford mostrou a maior sensibilidade para proteínas e o de biureto a menor. Para todos os métodos, as absorbâncias para diferentes proteínas (BSA, caseína, e ovoalbumina foram medidas e o método de Bradford mostrou a menor variação da absorbância. Utilizando o método de Bradford a concentração de proteínas totais obtida não foi estatisticamente diferente (p>0.05 daquela obtida pelo método do biureto. Porém, para os métodos da eritrosina-B e TBPEE as concentrações de proteínas totais foram estatisticamente diferentes (p<0.05 da obtida pelo método de biureto. Portanto o método de Bradford pode ser utilizado no lugar do método de biureto para a determinação de proteínas totais em plasma sangüíneo.

  19. In silicio identification of glycosyl-phosphatidylinositol-anchored plasma-membrane and cell wall proteins of Saccharomyces cerevisiae.

    Science.gov (United States)

    Caro, L H; Tettelin, H; Vossen, J H; Ram, A F; van den Ende, H; Klis, F M

    1997-12-01

    Use of the Von Heijne algorithm allowed the identification of 686 open reading frames (ORFs) in the genome of Saccharomyces cerevisiae that encode proteins with a potential N-terminal signal sequence for entering the secretory pathway. On further analysis, 51 of these proteins contain a potential glycosyl-phosphatidylinositol (GPI)-attachment signal. Seven additional ORFs were found to belong to this group. Upon examination of the possible GPI-attachment sites, it was found that in yeast the most probable amino acids for GPI-attachment as asparagine and glycine. In yeast, GPI-proteins are found at the cell surface, either attached to the plasma-membrane or as an intrinsic part of the cell wall. It was noted that plasma-membrane GPI-proteins possess a dibasic residue motif just before their predicted GPI-attachment site. Based on this, and on homologies between proteins, families of plasma-membrane and cell wall proteins were assigned, revealing 20 potential plasma-membrane and 38 potential cell wall proteins. For members of three plasma-membrane protein families, a function has been described. On the other hand, most of the cell wall proteins seem to be structural components of the wall, responsive to different growth conditions. The GPI-attachment site of yeast slightly differs from mammalian cells. This might be of use in the development of anti-fungal drugs.

  20. Expression of Bovine Leukemia Virus Genome is Blocked by a Nonimmunoglobulin Protein in Plasma from Infected Cattle

    Science.gov (United States)

    Gupta, P.; Ferrer, J. F.

    1982-01-01

    Plasma of cattle infected with bovine leukemia virus contains a soluble factor that blocks the expression of the viral genome in cultured lymphocytes. The blocking factor is not present in plasma of bovine leukemia virus-free cattle or of cattle infected with common bovine viruses. Blocking of bovine leukemia virus expression by the plasma factor is reversible, and seems to be mediated by a nonimmunoglobulin protein molecule.

  1. ENO1 Protein Levels in the Tumor Tissues and Circulating Plasma Samples of Non-small Cell Lung Cancer Patients

    Directory of Open Access Journals (Sweden)

    Ying ZHANG

    2010-12-01

    Full Text Available Background and objective Proper tumor markers are useful to diagnosis, prognosis and treatment for lung cancer. The aim of this study is to examine the levels of alpha-enolase (ENO1 protein in the tumor tissues and peripheral plasma samples obtained from non-small cell lung cancer (NSCLC patients, and evaluate its potential clinical significance. Methods The ENO1 protein levels in the tumor tissues and corresponding normal tissues from 16 cases of lung squamous cell carcinoma were analyzed by Western blot. The ENO1 protein levels in the plasma samples from 42 healthy individuals, 34 patients with lung benign disease and 84 patients with NSCLC were measured by double antibody sandwich enzyme-linked immunosorbent assay. Results For 87.5% (14/16 of the patients with lung squamous cell carcinoma, the ENO1 protein level in the tumor tissues was higher than that in the corresponding normal lung tissues. The ENO1 protein level in the plasma of NSCLC patients was significantly higher than that in the plasma of healthy individuals (P=0.031 and patients with lung benign disease (P=0.019. Furthermore, the ENO1 protein level was significantly higher in the plasma of patients with lung adenocarcinoma than that of patients with lung squamous cell carcinoma. Conclusion The elevated levels of ENO1 protein in the tumor tissues and the plasma samples from NSCLC patients indicate ENO1 may be a candidate biomarker of lung cancer.

  2. Krill protein hydrolysate reduces plasma triacylglycerol level with concurrent increase in plasma bile acid level and hepatic fatty acid catabolism in high-fat fed mice

    Directory of Open Access Journals (Sweden)

    Marie S. Ramsvik

    2013-11-01

    Full Text Available Background: Krill powder, consisting of both lipids and proteins, has been reported to modulate hepatic lipid catabolism in animals. Fish protein hydrolysate diets have also been reported to affect lipid metabolism and to elevate bile acid (BA level in plasma. BA interacts with a number of nuclear receptors and thus affects a variety of signaling pathways, including very low density lipoprotein (VLDL secretion. The aim of the present study was to investigate whether a krill protein hydrolysate (KPH could affect lipid and BA metabolism in mice. Method: C57BL/6 mice were fed a high-fat (21%, w/w diet containing 20% crude protein (w/w as casein (control group or KPH for 6 weeks. Lipids and fatty acid composition were measured from plasma, enzyme activity and gene expression were analyzed from liver samples, and BA was measured from plasma. Results: The effect of dietary treatment with KPH resulted in reduced levels of plasma triacylglycerols (TAG and non-esterified fatty acids (NEFAs. The KPH treated mice had also a marked increased plasma BA concentration. The increased plasma BA level was associated with induction of genes related to membrane canalicular exporter proteins (Abcc2, Abcb4 and to BA exporters to blood (Abcc3 and Abcc4. Of note, we observed a 2-fold increased nuclear farnesoid X receptor (Fxr mRNA levels in the liver of mice fed KPH. We also observed increased activity of the nuclear peroxiosme proliferator-activated receptor alpha (PPARα target gene carnitine plamitoyltransferase 2 (CPT-2. Conclusion: The KPH diet showed to influence lipid and BA metabolism in high-fat fed mice. Moreover, increased mitochondrial fatty acid oxidation and elevation of BA concentration may regulate the plasma level of TAGs and NEFAs.

  3. Differential regulation of plasma proteins between members of a family with homozygous HbE and HbEβ-thalassemia

    Directory of Open Access Journals (Sweden)

    Suchismita Halder

    2014-09-01

    Full Text Available In this report we’ve compared the plasma protein profiles of 4 individuals in a family. Father and the younger son both are hemoglobin (Hb Eβ-thalassemic {Cod 26 (G-A/IVS 1- 5 (G-C}, but the father never requires transfusion, whereas the younger son requires monthly blood transfusion. Mother and the elder son are HbEE {Cod 26 (G-A/Cod 26 (GA} without any history of transfusion. Proteomic study was done on the plasma fraction of the blood following ammonium sulphate precipitation. Proteins were separated by 2D-gel electrophoresis, expression of proteins compared by densitometry and proteins identified by tandem MALDI mass spectrometry. Proteins responsible in hemolysis, hypercoagulation and hemoglobin scavenging have shown differential regulation, establishing the relation between the differences in the levels of plasma proteins with the progression of the disease phenotype, manifested in the extent of transfusion dependence of the patient.

  4. Seminal plasma proteins interacting with sperm surface revert capacitation indicators in frozen-thawed ram sperm.

    Science.gov (United States)

    Ledesma, Alba; Fernández-Alegre, Estela; Cano, Adriana; Hozbor, Federico; Martínez-Pastor, Felipe; Cesari, Andreína

    2016-10-01

    This study was conducted to evaluate the effects of interacting seminal plasma proteins (iSPP) obtained by AV or EE on frozen-thawed ram sperm in order to test the hypothesis whether this fraction could be sufficient to emulate the effect of complete seminal plasma (SP). Additionally, we evaluated whether these proteins have a differential effect between spermatozoa from high and low fertility rams and between breeding and non-breeding seasons. We assessed sperm motility, quality parameters (intracellular reactive oxygen species, membrane fluidity, plasma membrane permeability and mitochondrial activity) and capacitation status. The main findings from this work were: i) iSPP had no effect on sperm motility, whereas SP (AV or EE) addition produced the highest values of total motility (74.13±2.99 and 72.27±2.99 for AV and EE, respectively) and progressive motility (64.97±2.64 and 63.73±2.64 for AV and EE, respectively); ii) iSPP had no effect on sperm quality parameters (p>0.05), but whole SP improved all parameters evaluated. Moreover, SP collected by AV yielded significantly higher viability (44.60±2.87) and sperm with stable plasma membrane (44.56±2.49) comparing with the addition of SP collected by EE (35.80±2.47 and 36.67±1.71, respectively); iii) iSPP and SP collected by EE, but not by AV, reverted molecular signals of capacitation as protein tyrosine phosphorylation caused by freezing temperatures; iv) there were no effects of fertility or season in sperm quality parameters evaluated. This study demonstrated that, although the iSPP have a clear decapacitating effect, including the ability to revert cryo-capacitation indicators, they are not sufficient to emulate the effects of complete SP regarding sperm functional parameters.

  5. Specific alterations in plasma proteins during depressed, manic, and euthymic states of bipolar disorder

    Energy Technology Data Exchange (ETDEWEB)

    Song, Y.R. [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Wu, B. [Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Yang, Y.T.; Chen, J. [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Zhang, L.J.; Zhang, Z.W. [Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Shi, H.Y. [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Huang, C.L.; Pan, J.X. [Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China); Xie, P. [Department of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Chongqing Key Laboratory of Neurobiology, Chongqing (China); Institute of Neuroscience and the Collaborative Innovation Center for Brain Science, Chongqing Medical University, Chongqing (China)

    2015-09-08

    Bipolar disorder (BD) is a common psychiatric mood disorder affecting more than 1-2% of the general population of different European countries. Unfortunately, there is no objective laboratory-based test to aid BD diagnosis or monitor its progression, and little is known about the molecular basis of BD. Here, we performed a comparative proteomic study to identify differentially expressed plasma proteins in various BD mood states (depressed BD, manic BD, and euthymic BD) relative to healthy controls. A total of 10 euthymic BD, 20 depressed BD, 15 manic BD, and 20 demographically matched healthy control subjects were recruited. Seven high-abundance proteins were immunodepleted in plasma samples from the 4 experimental groups, which were then subjected to proteome-wide expression profiling by two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight tandem mass spectrometry. Proteomic results were validated by immunoblotting and bioinformatically analyzed using MetaCore. From a total of 32 proteins identified with 1.5-fold changes in expression compared with healthy controls, 16 proteins were perturbed in BD independent of mood state, while 16 proteins were specifically associated with particular BD mood states. Two mood-independent differential proteins, apolipoprotein (Apo) A1 and Apo L1, suggest that BD pathophysiology may be associated with early perturbations in lipid metabolism. Moreover, down-regulation of one mood-dependent protein, carbonic anhydrase 1 (CA-1), suggests it may be involved in the pathophysiology of depressive episodes in BD. Thus, BD pathophysiology may be associated with early perturbations in lipid metabolism that are independent of mood state, while CA-1 may be involved in the pathophysiology of depressive episodes.

  6. Plasma levels of the arterial wall protein fibulin-1 are associated with carotid-femoral pulse wave velocity

    DEFF Research Database (Denmark)

    Laugesen, Esben; Høyem, Pernille; Christiansen, Jens Sandahl;

    2013-01-01

    -associated extracellular matrix protein, fibulin-1, was recently found in higher concentrations in the arterial wall and in plasma in patients with long duration type 2 diabetes. Furthermore, plasma fibulin-1 independently predicted total mortality and was associated with pulse pressure, an indirect measure of arterial...

  7. Proteomic Characterization of Zinc-Binding Proteins of Canine Seminal Plasma.

    Science.gov (United States)

    Mogielnicka-Brzozowska, M; Kowalska, N; Fraser, L; Kordan, W

    2015-12-01

    The zinc-binding proteins (ZnBPs) of the seminal plasma are implicated in different processes related to sperm-egg fusion. The aim of this study was to characterize the ZnBPs of canine seminal plasma using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry. The ZnBPs were isolated from the ejaculates of five dogs by affinity chromatography and subjected to 2D-PAGE analysis. The acquired spots, detected across the gels, were analysed by mass spectrometry. Using 2D-PAGE analysis, it was shown that canine seminal plasma comprised about 46-57 zinc-binding polypeptides, with molecular mass ranging from 9.3 to 138.7 kDa and pI at pH 5.2-10.0. It was found that zinc-binding polypeptides of low molecular masses (9.3-19.0 kDa and pI at pH 6.1-10.0) were predominant in the seminal plasma, and seven polypeptides, with molecular masses ranging from 11.7 to 15.4 kDa and pI at pH 6.8-8.7, were characterized by high optical density values. In addition, analysis with mass spectrometry (LC-MS-MS/MS) revealed that the identified seven polypeptides are canine prostate-specific esterase (CPSE), which is the main proteolytic enzyme of the seminal plasma. The findings of this study indicate an important regulatory role of seminal plasma zinc ions in the functional activity of CPSE, which is of great significance for maintaining the normal function of canine prostate and the spermatozoa functions.

  8. Assessment of coronary reperfusion in patients with myocardial infarction using fatty acid binding protein concentrations in plasma

    NARCIS (Netherlands)

    M.J.M. de Groot; A.M.M. Muijtjens; M.L. Simoons (Maarten); W.T. Hermens (Wim); J.F.C. Glatz

    2001-01-01

    textabstractOBJECTIVE: To examine whether successful coronary reperfusion after thrombolytic treatment in patients with confirmed acute myocardial infarction can be diagnosed from the plasma marker fatty acid binding protein (FABP), for either acute clinical decision making or retrospective purposes

  9. Toxicity challenges in environmental chemicals: Prediction of human plasma protein binding through quantitative structure-activity relationship (QSAR) models

    Science.gov (United States)

    The present study explores the merit of utilizing available pharmaceutical data to construct a quantitative structure-activity relationship (QSAR) for prediction of the fraction of a chemical unbound to plasma protein (Fub) in environmentally relevant compounds. Independent model...

  10. Easy measurement of diffusion coefficients of EGFP-tagged plasma membrane proteins using k-space Image Correlation Spectroscopy

    DEFF Research Database (Denmark)

    Christensen, Eva Arnspang; Koffman, Jennifer Skaarup; Marlar, Saw

    2014-01-01

    Lateral diffusion and compartmentalization of plasma membrane proteins are tightly regulated in cells and thus, studying these processes will reveal new insights to plasma membrane protein function and regulation. Recently, k-Space Image Correlation Spectroscopy (kICS)1 was developed to enable...... to the correlation function yields the diffusion coefficient. This paper provides a step-by-step guide to the image analysis and measurement of diffusion coefficients via kICS. First, a high frame rate image sequence of a fluorescently labeled plasma membrane protein is acquired using a fluorescence microscope Then...... routine measurements of diffusion coefficients directly from images of fluorescently tagged plasma membrane proteins, that avoided systematic biases introduced by probe photophysics. Although the theoretical basis for the analysis is complex, the method can be implemented by nonexperts using a freely...

  11. Effects of sugar-sweetened beverages on plasma acylation stimulating protein, leptin, and adiponectin: Relationships with metabolic outcomes

    Science.gov (United States)

    OBJECTIVE: The effects of fructose and glucose consumption on plasma acylation stimulating protein (ASP), adiponectin, and leptin concentrations relative to energy intake, body weight, adiposity, circulating triglycerides, and insulin sensitivity were determined. DESIGN AND METHODS: Thirty two over...

  12. Supplemental dietary protein for grazing dairy cows: reproduction, condition loss, plasma metabolites, and insulin.

    Science.gov (United States)

    Chapa, A M; McCormick, M E; Fernandez, J M; French, D D; Ward, J D; Beatty, J F

    2001-04-01

    An experiment was conducted over a 2-yr period to investigate the influence of grain crude protein (CP) and rumen undegradable protein (RUP) concentration on reproduction and energy status of dairy cows grazing annual ryegrass (Lolium multiflorum) and oats (Avena sativa). Holstein cows (n = 122) were blocked by calving group [partum (0 d postpartum) vs. postpartum (41 +/- 19 d postpartum at study initiation)] and assigned to grain supplements containing high CP [22.8% of dry matter (DM)], moderate CP (16.6%), or moderate CP (16.2%)] supplemented with RUP from blood meal and corn gluten meal. Postpartum condition loss was greater and first-service pregnancy rate was lower for partum-group cows receiving high CP grain supplements compared with control cows receiving moderate CP supplements. The RUP supplements reduced grain consumption, increased days to first estrus, and reduced first-service pregnancy rate of partum-group cows. The reproduction of postpartum group cows was unaffected by protein supplements. Plasma urea nitrogen was higher for cows fed high CP diets, but plasma ammonia nitrogen, glycated hemoglobin, nonesterified fatty acids, beta-hydoxybutyrate, glucose, and insulin concentrations were similar to cows fed moderate CP. Excess postpartum condition loss, coupled with inconsistent protein supplement effects on days to first service and first-service pregnancy rate, suggest that energy deprivation may have contributed to the low fertility experienced by grazing cows in this study.

  13. Super-resolution imaging of plasma membrane proteins with click chemistry

    Directory of Open Access Journals (Sweden)

    Pablo Mateos-Gil

    2016-09-01

    Full Text Available Besides its function as a passive cell wall, the plasma membrane (PM serves as a platform for different physiological processes such as signal transduction and cell adhesion, determining the ability of cells to communicate with the exterior and form tissues. Therefore, the spatial distribution of PM components, and the molecular mechanisms underlying it, have important implications in various biological fields including cell development, neurobiology, and immunology. The existence of confined compartments in the plasma membrane that vary on many length scales from protein multimers to micrometer-size domains with different protein and lipid composition is today beyond all questions. As much as the physiology of cells is controlled by the spatial organization of PM components, the study of distribution, size and composition remains challenging. Visualization of the molecular distribution of PM components has been impeded mainly due to two problems: the specific labeling of lipids and proteins without perturbing their native distribution and the diffraction-limit of fluorescence microscopy restricting the resolution to about half the wavelength of light. Here, we present a bioorthogonal chemical reporter strategy based on click chemistry and metabolic labeling for efficient and specific visualization of PM proteins and glycans with organic fluorophores in combination with super-resolution fluorescence imaging by direct stochastic optical reconstruction microscopy (dSTORM with single-molecule sensitivity.

  14. Investigating the effect of an arterial hypertension drug on the structural properties of plasma protein.

    Science.gov (United States)

    Hassan, Natalia; Maldonado-Valderrama, Julia; Gunning, A Patrick; Morris, V J; Ruso, Juan M

    2011-10-15

    Propanolol is a betablocker drug used in the treatment of arterial hypertension related diseases. In order to achieve an optimal performance of this drug it is important to consider the possible interactions of propanolol with plasma proteins. In this work, we have used several experimental techniques to characterise the effect of addition of the betablocker propanolol on the properties of bovine plasma fibrinogen (FB). Differential scanning calorimeter (DSC), circular dichroism (CD), dynamic light scattering (DLS), surface tension techniques and atomic force microscopy (AFM) measurements have been combined to carry out a detailed physicochemical and surface characterization of the mixed system. As a result, DSC measurements show that propranolol can play two opposite roles, either acting as a structure stabilizer at low molar concentrations or as a structure destabilizer at higher concentrations, in different domains of fibrinogen. CD measurements have revealed that the effect of propanolol on the secondary structure of fibrinogen depends on the temperature and the drug concentration and the DLS analysis showed evidence for protein aggregation. Interestingly, surface tension measurements provided further evidence of the conformational change induced by propanolol on the secondary structure of FB by importantly increasing the surface tension of the system. Finally, AFM imaging of the fibrinogen system provided direct visualization of the protein structure in the presence of propanolol. Combination of these techniques has produced complementary information on the behavior of the mixed system, providing new insights into the structural properties of proteins with potential medical interest.

  15. Plasma protein concentrations in hypertriglyceridaemic subjects. Effect of clofibrate and comparison with normal subjects.

    Science.gov (United States)

    Ballantyne, F C; Morrison, B A; Ballantyne, D; Dryburgh, F J; Epenetos, A A

    1978-07-01

    Clofibrate, a widely used hypolipidaemic agent was given for twelve weeks to ten subjects with hypertriglyceridaemia. Its effect on lipoprotein-lipids and caeruloplasmin, IgA, IgM, alpha2-microglobulin and transferrin was assessed by comparing analyses at 4, 8 and 12 weeks on therapy with the means of values at two weeks before and at the start of treatment. The normal variation in plasma proteins was assessed in six healthy volunteers during the same period of time. On clofibrate, very low density lipoprotein (VLDL) cholesterol and triglyceride concentrations fell, but the concentrations of cholesterol in low density (LDL) and high density (HDL) lipoproteins showed no consistent change. Caeruloplasmin and IgM concentrations decreased significantly, IgA showed a limited falls (significant only at 8 weeks) and alpha2-macroglobulin did not change. The concentration of transferrin increased on therapy. No relationships were found between the falls in VLDL-lipid concentrations and the alterations in other plasma proteins. No significant variation occurred in the concentrations of lipids or proteins in the normal subjects during the period of study. The results indicate that clofibrate exerts general effects on protein metabolism.

  16. Super-Resolution Imaging of Plasma Membrane Proteins with Click Chemistry

    Science.gov (United States)

    Mateos-Gil, Pablo; Letschert, Sebastian; Doose, Sören; Sauer, Markus

    2016-01-01

    Besides its function as a passive cell wall, the plasma membrane (PM) serves as a platform for different physiological processes such as signal transduction and cell adhesion, determining the ability of cells to communicate with the exterior, and form tissues. Therefore, the spatial distribution of PM components, and the molecular mechanisms underlying it, have important implications in various biological fields including cell development, neurobiology, and immunology. The existence of confined compartments in the plasma membrane that vary on many length scales from protein multimers to micrometer-size domains with different protein and lipid composition is today beyond all questions. As much as the physiology of cells is controlled by the spatial organization of PM components, the study of distribution, size, and composition remains challenging. Visualization of the molecular distribution of PM components has been impeded mainly due to two problems: the specific labeling of lipids and proteins without perturbing their native distribution and the diffraction-limit of fluorescence microscopy restricting the resolution to about half the wavelength of light. Here, we present a bioorthogonal chemical reporter strategy based on click chemistry and metabolic labeling for efficient and specific visualization of PM proteins and glycans with organic fluorophores in combination with super-resolution fluorescence imaging by direct stochastic optical reconstruction microscopy (dSTORM) with single-molecule sensitivity. PMID:27668214

  17. Chemoselective small molecules that covalently modify one lysine in a non-enzyme protein in plasma

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Sungwook; Connelly, Stephen; Reixach, Natàlia; Wilson, Ian A.; Kelly, Jeffery W. (Scripps)

    2010-02-19

    A small molecule that could bind selectively to and then react chemoselectively with a non-enzyme protein in a complex biological fluid, such as blood, could have numerous practical applications. Herein, we report a family of designed stilbenes that selectively and covalently modify the prominent plasma protein transthyretin in preference to more than 4,000 other human plasma proteins. They react chemoselectively with only one of eight lysine {epsilon}-amino groups within transthyretin. The crystal structure confirms the expected binding orientation of the stilbene substructure and the anticipated conjugating amide bond. These covalent transthyretin kinetic stabilizers exhibit superior amyloid inhibition potency compared to their noncovalent counterparts, and they prevent cytotoxicity associated with amyloidogenesis. Though there are a few prodrugs that, upon metabolic activation, react with a cysteine residue inactivating a specific non-enzyme, we are unaware of designed small molecules that react with one lysine {epsilon}-amine within a specific non-enzyme protein in a complex biological fluid.

  18. Intake of Mung Bean Protein Isolate Reduces Plasma Triglyceride Level in Rats

    Directory of Open Access Journals (Sweden)

    Nobuhiko Tachibana

    2013-09-01

    Full Text Available ABSTRACTBackground: Mung bean is well known as a starch source, but the physiological effects of mung bean protein have received little attention. In this study, we isolated mung bean protein from de-starched mung bean solutions, and investigated its influence on lipid metabolism. Objective: The aim of this study is to clarify the influence of the lipid metabolism by consumption of mung bean protein isolate (MPIMethods: Diets containing either mung bean protein isolate (MPI or casein were fed to normal rats for 28 days.Results: Both groups ate the same amount of food, but the plasma triglyceride level, relative liver weight and liver lipid contents (cholesterol and triglyceride pool in the MPI group were significantly lower than in the casein group. In the MPI group, the expression of sterol regulatory-element binding factor 1 (SREBF1 mRNA in the liver was significantly different when compared with the casein group. The significantly lower levels of insulin and free fatty acids in the MPI-fed rats may be due to the regulation of genes related to lipid metabolism in the liver.Conclusions: These results suggest that MPI may improve the plasma lipid profile by normalizing insulin sensitivity.Keywords: mung bean, Vigna radiata L., 8S globulin, triglyceride, β-conglycinin, 7S globulin, insulin sensitivity, SREBF1

  19. Effects of Long-Term Storage Time and Original Sampling Month on Biobank Plasma Protein Concentrations.

    Science.gov (United States)

    Enroth, Stefan; Hallmans, Göran; Grankvist, Kjell; Gyllensten, Ulf

    2016-10-01

    The quality of clinical biobank samples is crucial to their value for life sciences research. A number of factors related to the collection and storage of samples may affect the biomolecular composition. We have studied the effect of long-time freezer storage, chronological age at sampling, season and month of the year and on the abundance levels of 108 proteins in 380 plasma samples collected from 106 Swedish women. Storage time affected 18 proteins and explained 4.8-34.9% of the observed variance. Chronological age at sample collection after adjustment for storage-time affected 70 proteins and explained 1.1-33.5% of the variance. Seasonal variation had an effect on 15 proteins and month (number of sun hours) affected 36 proteins and explained up to 4.5% of the variance after adjustment for storage-time and age. The results show that freezer storage time and collection date (month and season) exerted similar effect sizes as age on the protein abundance levels. This implies that information on the sample handling history, in particular storage time, should be regarded as equally prominent covariates as age or gender and need to be included in epidemiological studies involving protein levels.

  20. High throughput atmospheric pressure plasma-induced graft polymerization for identifying protein-resistant surfaces.

    Science.gov (United States)

    Gu, Minghao; Kilduff, James E; Belfort, Georges

    2012-02-01

    Three critical aspects of searching for and understanding how to find highly resistant surfaces to protein adhesion are addressed here with specific application to synthetic membrane filtration. They include the (i) discovery of a series of previously unreported monomers from a large library of monomers with high protein resistance and subsequent low fouling characteristics for membrane ultrafiltration of protein-containing fluids, (ii) development of a new approach to investigate protein-resistant mechanisms from structure-property relationships, and (iii) adaptation of a new surface modification method, called atmospheric pressure plasma-induced graft polymerization (APP), together with a high throughput platform (HTP), for low cost vacuum-free synthesis of anti-fouling membranes. Several new high-performing chemistries comprising two polyethylene glycol (PEG), two amines and one zwitterionic monomers were identified from a library (44 commercial monomers) of five different classes of monomers as strong protein-resistant monomers. Combining our analysis here, using the Hansen solubility parameters (HSP) approach, and data from the literature, we conclude that strong interactions with water (hydrogen bonding) and surface flexibility are necessary for producing the highest protein resistance. Superior protein-resistant surfaces and subsequent anti-fouling performance was obtained with the HTP-APP as compared with our earlier HTP-photo graft-induced polymerization (PGP).

  1. Independent mobility of proteins and lipids in the plasma membrane of Escherichia coli.

    Science.gov (United States)

    Nenninger, Anja; Mastroianni, Giulia; Robson, Alexander; Lenn, Tchern; Xue, Quan; Leake, Mark C; Mullineaux, Conrad W

    2014-06-01

    Fluidity is essential for many biological membrane functions. The basis for understanding membrane structure remains the classic Singer-Nicolson model, in which proteins are embedded within a fluid lipid bilayer and able to diffuse laterally within a sea of lipid. Here we report lipid and protein diffusion in the plasma membrane of live cells of the bacterium Escherichia coli, using Fluorescence Recovery after Photobleaching (FRAP) and Total Internal Reflection Fluorescence (TIRF) microscopy to measure lateral diffusion coefficients. Lipid and protein mobility within the membrane were probed by visualizing an artificial fluorescent lipid and a simple model membrane protein consisting of a single membrane-spanning alpha-helix with a Green Fluorescent Protein (GFP) tag on the cytoplasmic side. The effective viscosity of the lipid bilayer is strongly temperature-dependent, as indicated by changes in the lipid diffusion coefficient. Surprisingly, the mobility of the model protein was unaffected by changes in the effective viscosity of the bulk lipid, and TIRF microscopy indicates that it clusters in segregated, mobile domains. We suggest that this segregation profoundly influences the physical behaviour of the protein in the membrane, with strong implications for bacterial membrane function and bacterial physiology.

  2. Lipoperoxidation and Protein Oxidative Damage Exhibit Different Kinetics During Septic Shock

    Directory of Open Access Journals (Sweden)

    Max Andresen

    2008-01-01

    Full Text Available Septic shock (SS-related multiorgan dysfunction has been associated with oxidative damage, but little is known about the temporal damage profile and its relationship to severity. The present work investigated prospectively 21 SS patients. Blood samples were obtained at diagnosis, 24, 72 hours, day 7, and at 3 months. At admission, thiobarbituric acid reactive substances (TBARSs, plasma protein carbonyls, plasma protein methionine sulfoxide (MS, ferric/reducing antioxidant power (FRAP, total red blood cell glutathione (RBCG, uric acid (UA, and bilirrubin levels were increased (P<.05. Total radical—trapping antioxidant potential (TRAP and vitamin-E were similar to controls, and vitamin-C was decreased (P<.05. During evolution, TBARS and RBCG increased (P<.001, vitamin-E levels remained stable, whereas plasma protein carbonyls and MS, TRAP, vitamin-C, reduced glutathione, and UA levels decreased (P<.006. After 3 months, plasma protein carbonyls and MS persisted elevated. More severe patients exhibited higher TBARS, TRAP, FRAP, vitamin-C, UA, and bilirrubin levels. Our results suggest early and persistent oxidative stress during septic shock and a correlation between increasing levels of lipoperoxidation and sepsis severity.

  3. Why mammals more susceptible to the hepatotoxic microcystins than fish: evidences from plasma and albumin protein binding through equilibrium dialysis.

    Science.gov (United States)

    Zhang, Wei; Liang, Gaodao; Wu, Laiyan; Tuo, Xun; Wang, Wenjing; Chen, Jun; Xie, Ping

    2013-08-01

    To elucidate the interspecies variation of susceptibility to microcystins (MCs), fresh plasma and purified albumin from six kinds of mammals and fish were used in toxins-substances binding test. Protein contents in the test plasma were analyzed and the binding characteristics to MCs were compared. Two kinds of widely observed MCs, microcystin-LR (MC-LR) and microcystin-RR (MC-RR) were tested and data were collected through the method of equilibrium dialysis. It was found that total plasma protein and albumin content in mammals were nearly two times and four times higher than that in fish, respectively. In the test range of 0-100 μg/mL, binding rates of fish plasma to MCs were considered significant lower (p mammals. And human plasma demonstrated the highest binding rate in mammals. In all the test species, plasma protein binding rates of MC-RR were significantly higher than MC-LR (p 0.05). From the view of protein binding, it is concluded that both the variation of plasma protein composition and albumin binding characteristic could influence the existing form of MCs in circulation, change MCs utilization, alter MCs half-life and further contribute to the difference of susceptibility between mammals and fish.

  4. Elevated pre-treatment levels of plasma C-reactive protein are associated with poor prognosis after breast cancer

    DEFF Research Database (Denmark)

    Allin, Kristine H; Nordestgaard, Børge G; Flyger, Henrik;

    2011-01-01

    We examined whether plasma C-reactive protein (CRP) levels at the time of diagnosis of breast cancer are associated with overall survival, disease-free survival, death from breast cancer, and recurrence of breast cancer.......We examined whether plasma C-reactive protein (CRP) levels at the time of diagnosis of breast cancer are associated with overall survival, disease-free survival, death from breast cancer, and recurrence of breast cancer....

  5. Ubiquitin initiates sorting of Golgi and plasma membrane proteins into the vacuolar degradation pathway

    Directory of Open Access Journals (Sweden)

    Scheuring David

    2012-09-01

    Full Text Available Abstract Background In yeast and mammals, many plasma membrane (PM proteins destined for degradation are tagged with ubiquitin. These ubiquitinated proteins are internalized into clathrin-coated vesicles and are transported to early endosomal compartments. There, ubiquitinated proteins are sorted by the endosomal sorting complex required for transport (ESCRT machinery into the intraluminal vesicles of multivesicular endosomes. Degradation of these proteins occurs after endosomes fuse with lysosomes/lytic vacuoles to release their content into the lumen. In plants, some PM proteins, which cycle between the PM and endosomal compartments, have been found to be ubiquitinated, but it is unclear whether ubiquitin is sufficient to mediate internalization and thus acts as a primary sorting signal for the endocytic pathway. To test whether plants use ubiquitin as a signal for the degradation of membrane proteins, we have translationally fused ubiquitin to different fluorescent reporters for the plasma membrane and analyzed their transport. Results Ubiquitin-tagged PM reporters localized to endosomes and to the lumen of the lytic vacuole in tobacco mesophyll protoplasts and in tobacco epidermal cells. The internalization of these reporters was significantly reduced if clathrin-mediated endocytosis was inhibited by the coexpression of a mutant of the clathrin heavy chain, the clathrin hub. Surprisingly, a ubiquitin-tagged reporter for the Golgi was also transported into the lumen of the vacuole. Vacuolar delivery of the reporters was abolished upon inhibition of the ESCRT machinery, indicating that the vacuolar delivery of these reporters occurs via the endocytic transport route. Conclusions Ubiquitin acts as a sorting signal at different compartments in the endomembrane system to target membrane proteins into the vacuolar degradation pathway: If displayed at the PM, ubiquitin triggers internalization of PM reporters into the endocytic transport route

  6. Shotgun proteomics and network analysis between plasma membrane and extracellular matrix proteins from rat olfactory ensheathing cells.

    Science.gov (United States)

    Liu, Yisong; Teng, Xiaohua; Yang, Xiaoxu; Song, Qing; Lu, Rong; Xiong, Jixian; Liu, Bo; Zeng, Nianju; Zeng, Yu; Long, Jia; Cao, Rui; Lin, Yong; He, Quanze; Chen, Ping; Lu, Ming; Liang, Songping

    2010-01-01

    Olfactory ensheathing cells (OECs) are a special type of glial cells that have characteristics of both astrocytes and Schwann cells. Evidence suggests that the regenerative capacity of OECs is induced by soluble, secreted factors that influence their microenvironment. These factors may regulate OECs self-renewal and/or induce their capacity to augment spinal cord regeneration. Profiling of plasma membrane and extracellular matrix through a high-throughput expression proteomics approach was undertaken to identify plasma membrane and extracellular matrix proteins of OECs under serum-free conditions. 1D-shotgun proteomics followed with gene ontology (GO) analysis was used to screen proteins from primary culture rat OECs. Four hundred and seventy nonredundant plasma membrane proteins and 168 extracellular matrix proteins were identified, the majority of which were never before reported to be produced by OECs. Furthermore, plasma membrane and extracellular proteins were classified based on their protein-protein interaction predicted by STRING quantitatively integrates interaction data. The proteomic profiling of the OECs plasma membrane proteins and their connection with the secretome in serum-free culture conditions provides new insights into the nature of their in vivo microenvironmental niche. Proteomic analysis for the discovery of clinical biomarkers of OECs mechanism warrants further study.

  7. Intracellular protein transport to the thyrocyte plasma membrane: potential implications for thyroid physiology.

    Science.gov (United States)

    Arvan, P; Kim, P S; Kuliawat, R; Prabakaran, D; Muresan, Z; Yoo, S E; Abu Hossain, S

    1997-02-01

    We present a snapshot of developments in epithelial biology that may prove helpful in understanding cellular aspects of the machinery designed for the synthesis of thyroid hormones on the thyroglobulin precursor. The functional unit of the thyroid gland is the follicle, delimited by a monolayer of thyrocytes. Like the cells of most simple epithelia, thyrocytes exhibit specialization of the cell surface that confronts two different extracellular environments-apical and basolateral, which are separated by tight junctions. Specifically, the basolateral domain faces the interstitium/bloodstream, while the apical domain is in contact with the lumen that is the primary target for newly synthesized thyroglobulin secretion and also serves as a storage depot for previously secreted protein. Thyrocytes use their polarity in several important ways, such as for maintaining basolaterally located iodide uptake and T4 deiodination, as well apically located iodide efflux and iodination machinery. The mechanisms by which this organization is established, fall in large part under the more general cell biological problem of intracellular sorting and trafficking of different proteins en route to the cell surface. Nearly all exportable proteins begin their biological life after synthesis in an intracellular compartment known as the endoplasmic reticulum (ER), upon which different degrees of difficulty may be encountered during nascent polypeptide folding and initial export to the Golgi complex. In these initial stages, ER molecular chaperones can assist in monitoring protein folding and export while themselves remaining as resident proteins of the thyroid ER. After export from the ER, most subsequent sorting for protein delivery to apical or basolateral surfaces of thyrocytes occurs within another specialized intracellular compartment known as the trans-Golgi network. Targeting information encoded in secretory proteins and plasma membrane proteins can be exposed or buried at different

  8. Plasma soluble prion protein, a potential biomarker for sport-related concussions: a pilot study.

    Directory of Open Access Journals (Sweden)

    Nam Pham

    Full Text Available Sport-related mild traumatic brain injury (mTBI or concussion is a significant health concern to athletes with potential long-term consequences. The diagnosis of sport concussion and return to sport decision making is one of the greatest challenges facing health care clinicians working in sports. Blood biomarkers have recently demonstrated their potential in assisting the detection of brain injury particularly, in those cases with no obvious physical injury. We have recently discovered plasma soluble cellular prion protein (PrP(C as a potential reliable biomarker for blast induced TBI (bTBI in a rodent animal model. In order to explore the application of this novel TBI biomarker to sport-related concussion, we conducted a pilot study at the University of Saskatchewan (U of S by recruiting athlete and non-athlete 18 to 30 year-old students. Using a modified quantitative ELISA method, we first established normal values for the plasma soluble PrP(C in male and female students. The measured plasma soluble PrP(C in confirmed concussion cases demonstrated a significant elevation of this analyte in post-concussion samples. Data collected from our pilot study indicates that the plasma soluble PrP(C is a potential biomarker for sport-related concussion, which may be further developed into a clinical diagnostic tool to assist clinicians in the assessment of sport concussion and return-to-play decision making.

  9. Plasma soluble prion protein, a potential biomarker for sport-related concussions: a pilot study.

    Science.gov (United States)

    Pham, Nam; Akonasu, Hungbo; Shishkin, Rhonda; Taghibiglou, Changiz

    2015-01-01

    Sport-related mild traumatic brain injury (mTBI) or concussion is a significant health concern to athletes with potential long-term consequences. The diagnosis of sport concussion and return to sport decision making is one of the greatest challenges facing health care clinicians working in sports. Blood biomarkers have recently demonstrated their potential in assisting the detection of brain injury particularly, in those cases with no obvious physical injury. We have recently discovered plasma soluble cellular prion protein (PrP(C)) as a potential reliable biomarker for blast induced TBI (bTBI) in a rodent animal model. In order to explore the application of this novel TBI biomarker to sport-related concussion, we conducted a pilot study at the University of Saskatchewan (U of S) by recruiting athlete and non-athlete 18 to 30 year-old students. Using a modified quantitative ELISA method, we first established normal values for the plasma soluble PrP(C) in male and female students. The measured plasma soluble PrP(C) in confirmed concussion cases demonstrated a significant elevation of this analyte in post-concussion samples. Data collected from our pilot study indicates that the plasma soluble PrP(C) is a potential biomarker for sport-related concussion, which may be further developed into a clinical diagnostic tool to assist clinicians in the assessment of sport concussion and return-to-play decision making.

  10. Irreversible binding of an anticancer compound (BI-94) to plasma proteins.

    Science.gov (United States)

    Gautam, Nagsen; Thakare, Rhishikesh; Rana, Sandeep; Natarajan, Amarnath; Alnouti, Yazen

    2015-01-01

    1. We investigated the mechanisms responsible for the in vivo instability of a benzofurazan compound BI-94 (NSC228148) with potent anti-cancer activity. 2. BI-94 was stable in MeOH, water, and in various buffers at pHs 2.5-5, regardless of the buffer composition. In contrast, BI-94 was unstable in NaOH and at pHs 7-9, regardless of the buffer composition. BI-94 disappeared immediately after spiking into mice, rat, monkey, and human plasma. BI-94 stability in plasma can be only partially restored by acidifying it, which indicated other mechanisms in addition to pH for BI-94 instability in plasma. 3. BI-94 formed adducts with the trapping agents, glutathione (GSH) and N-acetylcysteine (NAC), in vivo and in vitro via nucleophilic aromatic substitution reaction. The kinetics of adduct formation showed that neutral or physiological pHs enhanced and accelerated GSH and NAC adduct formation with BI-94, whereas acidic pHs prevented it. Therefore, physiological pHs not only altered BI-94 chemical stability but also enhanced adduct formation with endogenous nucleophiles. In addition, adduct formation with human serum albumin-peptide 3 (HSA-T3) at the Cys34 position was demonstrated. 4. In conclusion, BI-94 was unstable at physiological conditions due to chemical instability and irreversible binding to plasma proteins.

  11. Reliability of plasma lipopolysaccharide-binding protein (LBP) from repeated measures in healthy adults.

    Science.gov (United States)

    Citronberg, Jessica S; Wilkens, Lynne R; Lim, Unhee; Hullar, Meredith A J; White, Emily; Newcomb, Polly A; Le Marchand, Loïc; Lampe, Johanna W

    2016-09-01

    Plasma lipopolysaccharide-binding protein (LBP), a measure of internal exposure to bacterial lipopolysaccharide, has been associated with several chronic conditions and may be a marker of chronic inflammation; however, no studies have examined the reliability of this biomarker in a healthy population. We examined the temporal reliability of LBP measured in archived samples from participants in two studies. In Study one, 60 healthy participants had blood drawn at two time points: baseline and follow-up (either three, six, or nine months). In Study two, 24 individuals had blood drawn three to four times over a seven-month period. We measured LBP in archived plasma by ELISA. Test-retest reliability was estimated by calculating the intraclass correlation coefficient (ICC). Plasma LBP concentrations showed moderate reliability in Study one (ICC 0.60, 95 % CI 0.43-0.75) and Study two (ICC 0.46, 95 % CI 0.26-0.69). Restricting the follow-up period improved reliability. In Study one, the reliability of LBP over a three-month period was 0.68 (95 % CI: 0.41-0.87). In Study two, the ICC of samples taken ≤seven days apart was 0.61 (95 % CI 0.29-0.86). Plasma LBP concentrations demonstrated moderate test-retest reliability in healthy individuals with reliability improving over a shorter follow-up period.

  12. Direct covalent coupling of proteins to nanostructured plasma polymers: a route to tunable cell adhesion

    Energy Technology Data Exchange (ETDEWEB)

    Melnichuk, Iurii, E-mail: iurii.melnichuk@gmail.com [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); Choukourov, Andrei, E-mail: choukourov@kmf.troja.mff.cuni.cz [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); Bilek, Marcela, E-mail: m.bilek@physics.usyd.edu.au [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); School of Physics, University of Sydney, NSW 2006 (Australia); Weiss, Anthony, E-mail: tony.weiss@sydney.edu.au [School of Molecular Bioscience, University of Sydney, NSW 2006 (Australia); Vandrovcová, Marta, E-mail: Marta.Vandrovcova@fgu.cas.cz [Institute of Physiology of Czech Academy of Science, Prague 14220 (Czech Republic); Bačáková, Lucie, E-mail: Lucie.Bacakova@fgu.cas.cz [Institute of Physiology of Czech Academy of Science, Prague 14220 (Czech Republic); Hanuš, Jan, E-mail: jan.hanus@gmail.com [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); Kousal, Jaroslav, E-mail: jarda@kmf.troja.mff.cuni.cz [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); Shelemin, Artem, E-mail: artem.shelemin@gmail.com [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); Solař, Pavel, E-mail: pawell.solar@seznam.cz [Charles University in Prague, Faculty of Mathematics and Physics, Department of Macromolecular Physics, Prague 18000 (Czech Republic); and others

    2015-10-01

    Highlights: • Flat and nanostructured interfaces were overcoated by hydrocarbon plasma polymer. • Linker-free covalent attachment of proteins to resultant surfaces was validated. • Ultra-thin hydrocarbon overcoat (<2 nm) secured prolonged effective binding. • Pre-adsorbed tropoelastin promoted proliferation of osteoblast-like MG-63 cells. • Nanostructured films were multi-affine and impeded cell adhesion. - Abstract: Flat and nanostructured thin films were fabricated by deposition of ultra-thin (<2 nm) layer of hydrocarbon plasma polymer over polished silicon and over a pattern of 8 nm-thick poly(ethylene) islands on silicon. Linker-free radical-based covalent binding of bovine serum albumin and tropoelastin was confirmed for both types of films. The binding capability of albumin was found to be stable over many days of ambient air storage time. Tropoelastin-mediated flat plasma polymers favored adhesion and proliferation of osteoblast-like MG-63 cells. Nanostructured plasma polymers were multi-affine and their hierarchical surface represented an additional barrier for cell attachment.

  13. Olympic boxing is associated with elevated levels of the neuronal protein tau in plasma.

    Science.gov (United States)

    Neselius, Sanna; Zetterberg, Henrik; Blennow, Kaj; Randall, Jeffrey; Wilson, David; Marcusson, Jan; Brisby, Helena

    2013-01-01

    The aim of this study was to investigate if olympic (amateur) boxing is associated with elevation of brain injury biomarkers in peripheral blood compared to controls. Thirty olympic boxers competing in at least 47 bouts were compared to 25 controls. Blood was collected from the controls at one occasion and from the boxers within 1-6 days after a bout and after a rest period of at least 14 days. Tau concentration in plasma was determined using a novel single molecule ELISA assay and S-100B, glial fibrillary acidic protein, brain-derived neurotrophic factor and amyloid β 1-42 were determined using standard immunoassays. None of the boxers had been knocked-out during the bout. Plasma-tau was significantly increased in the boxers after a bout compared to controls (mean ± SD, 2.46 ± 5.10 vs. 0.79 ± 0.961 ng L(-1), p = 0.038). The other brain injury markers did not differ between the groups. Plasma-tau decreased significantly in the boxers after a resting period compared to after a bout (p = 0.030). Olympic boxing is associated with elevation of tau in plasma. The repetitive minimal head injury in boxing may lead to axonal injuries that can be diagnosed with a blood test.

  14. PLASMA PROTEIN ELECTROPHORESIS AND SELECT ACUTE PHASE PROTEINS IN HEALTHY BONNETHEAD SHARKS (SPHYRNA TIBURO) UNDER MANAGED CARE.

    Science.gov (United States)

    Hyatt, Michael W; Field, Cara L; Clauss, Tonya M; Arheart, Kristopher L; Cray, Carolyn

    2016-12-01

    Preventative health care of elasmobranchs is an important but understudied field of aquatic veterinary medicine. Evaluation of inflammation through the acute phase response is a valuable tool in health assessments. To better assess the health of bonnethead sharks ( Sphyrna tiburo ) under managed care, normal reference intervals of protein electrophoresis (EPH) and the acute phase proteins, C-reactive protein (CRP) and haptoglobin (HP), were established. Blood was collected from wild caught, captive raised bonnethead sharks housed at public aquaria. Lithium heparinized plasma was either submitted fresh or stored at -80°C prior to submission. Electrophoresis identified protein fractions with migration characteristics similar to other animals with albumin, α-1 globulin, α-2 globulin, β globulin, and γ globulin. These fractions were classified as fractions 1-5 as fractional contents are unknown in this species. Commercial reagents for CRP and HP were validated for use in bonnethead sharks. Reference intervals were established using the robust method recommended by the American Society for Veterinary Clinical Pathology for the calculation of 90% reference intervals. Once established, the diagnostic and clinical applicability of these reference intervals was used to assess blood from individuals with known infectious diseases that resulted in systemic inflammation and eventual death. Unhealthy bonnethead sharks had significantly decreased fraction 2, fraction 3, and fraction 3:4 ratio and significantly increased fraction 5, CRP, and HP. These findings advance our understanding of elasmobranch acute phase inflammatory response and health and aid clinicians in the diagnosis of inflammatory disease in bonnethead sharks.

  15. Non-thermal dielectric-barrier discharge plasma damages human keratinocytes by inducing oxidative stress

    Science.gov (United States)

    KIM, KI CHEON; PIAO, MEI JING; HEWAGE, SUSARA RUWAN KUMARA MADDUMA; HAN, XIA; KANG, KYOUNG AH; JO, JIN OH; MOK, YOUNG SUN; SHIN, JENNIFER H.; PARK, YEUNSOO; YOO, SUK JAE; HYUN, JIN WON

    2016-01-01

    The aim of this study was to identify the mechanisms through which dielectric-barrier discharge plasma damages human keratinocytes (HaCaT cells) through the induction of oxidative stress. For this purpose, the cells were exposed to surface dielectric-barrier discharge plasma in 70% oxygen and 30% argon. We noted that cell viability was decreased following exposure of the cells to plasma in a time-dependent manner, as shown by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The levels of intracellular reactive oxygen species (ROS) were determined using 2′,7′-dichlorodihydro-fluorescein diacetate and dihydroethidium was used to monitor superoxide anion production. Plasma induced the generation of ROS, including superoxide anions, hydrogen peroxide and hydroxyl radicals. N-acetyl cysteine, which is an antioxidant, prevented the decrease in cell viability caused by exposure to plasma. ROS generated by exposure to plasma resulted in damage to various cellular components, including lipid membrane peroxidation, DNA breaks and protein carbonylation, which was detected by measuring the levels of 8-isoprostane and diphenyl-1-pyrenylphosphine assay, comet assay and protein carbonyl formation. These results suggest that plasma exerts cytotoxic effects by causing oxidative stress-induced damage to cellular components. PMID:26573561

  16. Plasma proteins as indices of response to nutritional therapy in cancer patients.

    Science.gov (United States)

    Ota, D M; Frasier, P; Guevara, J; Foulkes, M

    1985-07-01

    The use of plasma albumin (ALB), transferrin (TFN), prealbumin (TBPA), retinol-binding protein (RBP), triceps skin fold (TSF), and midarm muscle circumference (MAMC) as determinants of response to nutritional therapy (TPN) was investigated in 40 cancer patients during preoperative TPN. Thirty-one patients received 90% or more of their anabolic caloric requirement (Harris-Benedict equation) by means of TPN. During this study period (average 11.1 +/- 4.7 days) nutritional assessments were completed before TPN and on the last day of TPN before surgery. Average weight loss based on usual body wt (UBW) and ideal body wt (IBW) was 19 +/- 11% and 9 +/- 15%, respectively (not significant, NS). Weight loss (UBW) correlated with ALB (P less than 0.001), TBPA (P less than 0.005) and RBP (P less than 0.02) but did not correlate with TFN (P less than 0.06), TSF, and MAMC. Weight loss (IBW) correlated with TSF (P less than 0.01) and MAMC (P less than 0.03) but did not correlate with plasma protein (PP). During TPN the average percent increases for PP were 0.1% (ALB, NS), 20% (TFN, NS), 60% (TBPA, P less than 0.02), and 116% (RBP, P less than 0.005). These results suggest that plasma TBPA and RBP are significant parameters of response to short-term nutritional therapy in cancer patients.

  17. Plasma monocyte chemotactic protein-1 remains elevated after minimally invasive colorectal cancer resection.

    Science.gov (United States)

    Shantha Kumara, H M C; Myers, Elizabeth A; Herath, Sonali Ac; Jang, Joon Ho; Njoh, Linda; Yan, Xiaohong; Kirchoff, Daniel; Cekic, Vesna; Luchtefeld, Martin; Whelan, Richard L

    2014-10-15

    To investigate plasma Monocyte Chemotactic Protein-1 levels preoperatively in colorectal cancer (CRC) and benign patients and postoperatively after CRC resection. A plasma bank was screened for minimally invasive colorectal cancer resection (MICR) for CRC and benign disease (BEN) patients for whom preoperative, early postoperative, and 1 or more late postoperative samples (postoperative day 7-27) were available. Monocyte chemotactic protein-1 (MCP-1) levels (pg/mL) were determined via enzyme linked immuno-absorbent assay. One hundred and two CRC and 86 BEN patients were studied. The CRC patient's median preoperative MCP-1 level (283.1, CI: 256.0, 294.3) was higher than the BEN group level (227.5, CI: 200.2, 245.2; P = 0.0004). Vs CRC preoperative levels, elevated MCP-1 plasma levels were found on postoperative day 1 (446.3, CI: 418.0, 520.1), postoperative day 3 (342.7, CI: 320.4, 377.4), postoperative day 7-13 (326.5, CI: 299.4, 354.1), postoperative day 14-20 (361.6, CI: 287.8, 407.9), and postoperative day 21-27 (318.1, CI: 287.2, 371.6; P MICR for CRC, MCP-1 levels were elevated for 1 mo and may promote angiogenesis, cancer recurrence and metastasis.

  18. Interaction of Mason-Pfizer monkey virus matrix protein with plasma membrane.

    Directory of Open Access Journals (Sweden)

    Jan ePrchal

    2014-01-01

    Full Text Available Budding is the final step of the late phase of retroviral life cycle. It begins with the interaction of Gag precursor with plasma membrane through its N-terminal domain, the matrix protein. However, single generas of Retroviridae family differ in the way how they interact with plasma membrane. While in case of lentiviruses (e.g. human immunodeficiency virus (HIV the structural polyprotein precursor Gag interacts with cellular membrane prior to the assembly, betaretroviruses (Mason-Pfizer monkey virus (M-PMV first assemble their virus-like particles in the pericentriolar region of the infected cell and therefore, already assembled particles interact with the membrane. Although both these types of retroviruses use similar mechanism of the interaction of Gag with the membrane, the difference in the site of assembly leads to some differences in the mechanism of the interaction. Here we describe the interaction of M-PMV matrix protein with plasma membrane with emphasis on the structural aspects of the interaction with single phospholipids.

  19. Reactive Carbonyl Species In Vivo: Generation and Dual Biological Effects

    Directory of Open Access Journals (Sweden)

    Halyna M. Semchyshyn

    2014-01-01

    Full Text Available Reactive carbonyls are widespread species in living organisms and mainly known for their damaging effects. The most abundant reactive carbonyl species (RCS are derived from oxidation of carbohydrates, lipids, and amino acids. Chemical modification of proteins, nucleic acids, and aminophospholipids by RCS results in cytotoxicity and mutagenicity. In addition to their direct toxicity, modification of biomolecules by RCS gives rise to a multitude of adducts and cross links that are increasingly implicated in aging and pathology of a wide range of human diseases. Understanding of the relationship between metabolism of RCS and the development of pathological disorders and diseases may help to develop effective approaches to prevent a number of disorders and diseases. On the other hand, constant persistence of RCS in cells suggests that they perform some useful role in living organisms. The most beneficial effects of RCS are their establishment as regulators of cell signal transduction and gene expression. Since RCS can modulate different biological processes, new tools are required to decipher the precise mechanisms underlying dual effects of RCS.

  20. Dextrose-mediated aggregation of therapeutic monoclonal antibodies in human plasma: Implication of isoelectric precipitation of complement proteins.

    Science.gov (United States)

    Luo, Shen; Zhang, Baolin

    2015-01-01

    Many therapeutic monoclonal antibodies (mAbs) are clinically administered through intravenous infusion after mixing with a diluent, e.g., saline, 5% dextrose. Such a clinical setting increases the likelihood of interactions among mAb molecules, diluent, and plasma components, which may adversely affect product safety and efficacy. Avastin® (bevacizumab) and Herceptin® (trastuzumab), but not Remicade® (infliximab), were shown to undergo rapid aggregation upon dilution into 5% dextrose when mixed with human plasma in vitro; however, the biochemical pathways leading to the aggregation were not clearly defined. Here, we show that dextrose-mediated aggregation of Avastin or Herceptin in plasma involves isoelectric precipitation of complement proteins. Using mass spectrometry, we found that dextrose-induced insoluble aggregates were composed of mAb itself and multiple abundant plasma proteins, namely complement proteins C3, C4, factor H, fibronectin, and apolipoprotein. These plasma proteins, which are characterized by an isoelectronic point of 5.5-6.7, lost solubility at the resulting pH in the mixture with formulated Avastin (pH 6.2) and Herceptin (pH 6.0). Notably, switching formulation buffers for Avastin (pH 6.2) and Remicade (pH 7.2) reversed their aggregation profiles. Avastin formed little, if any, insoluble aggregates in dextrose-plasma upon raising the buffer pH to 7.2 or above. Furthermore, dextrose induced pH-dependent precipitation of plasma proteins, with massive insoluble aggregates being detected at pH 6.5-6.8. These data show that isoelectric precipitation of complement proteins is a prerequisite of dextrose-induced aggregation of mAb in human plasma. This finding highlights the importance of assessing the compatibility of a therapeutic mAb with diluent and human plasma during product development.

  1. Modifications of blood platelet proteins of patients with schizophrenia.

    Science.gov (United States)

    Dietrich-Muszalska, Anna; Olas, Beata

    2009-03-01

    Oxidative damage to lipids in plasma, blood platelets and neurons in patients with schizophrenia was described. The aim of our present study was to evaluate oxidative/nitrative modifications of blood platelets proteins by measurement the level of biomarkers of oxidative stress such as carbonyl groups, thiol groups and 3-nitrotyrosine in proteins in patients with schizophrenia and compare with a control group. Levels of carbonyl groups and 3-nitrotyrosine residues in platelet proteins were measured by ELISA and competition ELISA, respectively. The method with 5,5'-dithio-bis(2-nitro-benzoic acid) has been used to analyse thiol groups in platelet proteins. We demonstrated for the first time in platelet proteins from patients with schizophrenia a statistically significant increase of the level of biomarkers of oxidative/nitrative stress such as carbonyl groups or 3-nitrotyrosine; in schizophrenic patients the amount of thiol groups in platelet proteins was lower than in platelets from healthy subjects. Our results strongly indicate that in patients with schizophrenia reactive oxygen species and reactive nitrogen species induce not only peroxidation of lipids, but also may stimulate oxidative/nitrative modifications of platelet proteins. The consequence of these modifications may be the alteration of platelet protein structure and function.

  2. Oxidation of Reduced Sulfur Species: Carbonyl Sulfide

    DEFF Research Database (Denmark)

    Glarborg, Peter; Marshall, Paul

    2013-01-01

    A detailed chemical kinetic model for oxidation of carbonyl sulfide (OCS) has been developed, based on a critical evaluation of data from the literature. The mechanism has been validated against experimental results from batch reactors, flow reactors, and shock tubes. The model predicts satisfact......A detailed chemical kinetic model for oxidation of carbonyl sulfide (OCS) has been developed, based on a critical evaluation of data from the literature. The mechanism has been validated against experimental results from batch reactors, flow reactors, and shock tubes. The model predicts...... by the competition between chain‐branching and ‐propagating steps; modeling predictions are particularly sensitive to the branching fraction for the OCS + O reaction to form CO + SO or CO2 + S....

  3. Presence of membranous vesicles in cat seminal plasma: ultrastructural characteristics, protein profile and enzymatic activity.

    Science.gov (United States)

    Polisca, A; Troisi, A; Minelli, A; Bellezza, I; Fontbonne, A; Zelli, R

    2015-02-01

    This study sought to verify the presence of membranous vesicles in cat seminal plasma by means of transmission electron microscopy and to identify protein profile and some of the enzymatic activities associated with these particles. The transmission electron microscopy observations showed the existence of different sized vesicular membranous structures of more or less spherical shape. These vesicles were surrounded by single-, double- or multiple-layered laminar membranes. The vesicle diameters ranged from 16.3 to 387.4 nm, with a mean of 116.5 ± 70.7 nm. Enzyme activity determinations showed the presence of dipeptilpeptidase IV, aminopeptidase, alkaline and acid phosphatase. To our knowledge, this is the first report that identifies and characterizes the membranous vesicles in cat seminal plasma. However, further studies are necessary to identify the exact site of production of these membranous vesicles in the cat male genital tract and to determine their specific roles in the reproductive events of this species.

  4. Sulfur-based absolute quantification of proteins using isotope dilution inductively coupled plasma mass spectrometry

    Science.gov (United States)

    Lee, Hyun-Seok; Heun Kim, Sook; Jeong, Ji-Seon; Lee, Yong-Moon; Yim, Yong-Hyeon

    2015-10-01

    An element-based reductive approach provides an effective means of realizing International System of Units (SI) traceability for high-purity biological standards. Here, we develop an absolute protein quantification method using double isotope dilution (ID) inductively coupled plasma mass spectrometry (ICP-MS) combined with microwave-assisted acid digestion for the first time. We validated the method and applied it to certify the candidate protein certified reference material (CRM) of human growth hormone (hGH). The concentration of hGH was determined by analysing the total amount of sulfur in hGH. Next, the size-exclusion chromatography method was used with ICP-MS to characterize and quantify sulfur-containing impurities. By subtracting the contribution of sulfur-containing impurities from the total sulfur content in the hGH CRM, we obtained a SI-traceable certification value. The quantification result obtained with the present method based on sulfur analysis was in excellent agreement with the result determined via a well-established protein quantification method based on amino acid analysis using conventional acid hydrolysis combined with an ID liquid chromatography-tandem mass spectrometry. The element-based protein quantification method developed here can be generally used for SI-traceable absolute quantification of proteins, especially pure-protein standards.

  5. Determination of Dynamics of Plant Plasma Membrane Proteins with Fluorescence Recovery and Raster Image Correlation Spectroscopy.

    Science.gov (United States)

    Laňková, Martina; Humpolíčková, Jana; Vosolsobě, Stanislav; Cit, Zdeněk; Lacek, Jozef; Čovan, Martin; Čovanová, Milada; Hof, Martin; Petrášek, Jan

    2016-04-01

    A number of fluorescence microscopy techniques are described to study dynamics of fluorescently labeled proteins, lipids, nucleic acids, and whole organelles. However, for studies of plant plasma membrane (PM) proteins, the number of these techniques is still limited because of the high complexity of processes that determine the dynamics of PM proteins and the existence of cell wall. Here, we report on the usage of raster image correlation spectroscopy (RICS) for studies of integral PM proteins in suspension-cultured tobacco cells and show its potential in comparison with the more widely used fluorescence recovery after photobleaching method. For RICS, a set of microscopy images is obtained by single-photon confocal laser scanning microscopy (CLSM). Fluorescence fluctuations are subsequently correlated between individual pixels and the information on protein mobility are extracted using a model that considers processes generating the fluctuations such as diffusion and chemical binding reactions. As we show here using an example of two integral PM transporters of the plant hormone auxin, RICS uncovered their distinct short-distance lateral mobility within the PM that is dependent on cytoskeleton and sterol composition of the PM. RICS, which is routinely accessible on modern CLSM instruments, thus represents a valuable approach for studies of dynamics of PM proteins in plants.

  6. Extracellular matrix protein fibulin-1 plasma levels are associated with increased cardiovascular risk in chronic kidney disease

    DEFF Research Database (Denmark)

    Scholze, Alexandra

    INTRODUCTION AND AIMS: Fibulin-1 is one of the few extracellular matrix proteins present in blood in high concentrations. We aimed to define the relationship between plasma fibulin-1 levels and risk markers of cardiovascular disease in patients with chronic kidney disease. METHODS: Plasma fibulin-1...... of plasma fibulin-1. CONCLUSIONS: Increased plasma fibulin-1 levels were associated with impaired kidney function and diabetes. Fibulin-1 levels were also associated with hemodynamic cardiovascular risk markers. We conclude, that fibulin-1 is involved in the pathogenesis of cardiovascular disease observed...

  7. Relationship between dietary folate intake and plasma monocyte chemoattractant protein-1 and interleukin-8 in heart failure patients.

    Science.gov (United States)

    Chung, Hye Kyung; Kim, Oh Yoen; Lee, Hyeran; Do, Hyun Joo; Kim, Young Soon; Oh, Jaewon; Kang, Seok-Min; Shin, Min-Jeong

    2011-07-01

    This study aimed to examine the association of dietary vitamin intakes with plasma pro-inflammatory cytokine levels in Korean heart failure patients. Stable outpatients with heart failure were recruited and finally 91 patients were included. Dietary intakes were estimated by a developed semi-quantitative food frequency questionnaire. The simultaneous measurement of 17 cytokines was performed along with analysis of plasma C-reactive protein. Plasma C-reactive protein levels significantly correlated with dietary intakes of vitamin C (r = -0.30, pmonocyte chemoattractant protein-1 significantly correlated with dietary folate intake (r = -0.31, pmonocyte chemoattractant protein-1 (pmonocyte chemoattractant protein-1 and interleukin-8 which indicates dietary folate may have a potentially beneficial role in the prevention and treatment of heart failure.

  8. Increased plasma retinol binding protein 4 levels in patients with inflammatory cardiomyopathy.

    Science.gov (United States)

    Bobbert, Peter; Weithäuser, Alice; Andres, Janin; Bobbert, Thomas; Kühl, Uwe; Schultheiss, Heinz Peter; Rauch, Ursula; Skurk, Carsten

    2009-12-01

    Chronic heart failure (CHF) is associated with a higher risk for diabetes mellitus. Retinol binding protein 4 (RBP 4) is an adipose tissue-derived protein with pro-diabetogenic effects. A complete understanding of the association of CHF and insulin resistance remains elusive. The purpose of this study was to examine the relationship between CHF and diabetes mellitus. Plasma levels of RBP 4, insulin, and interleukins (IL) 2, 8, and 10, were assessed in patients with dilated cardiomyopathy (DCM, n = 53), dilated inflammatory cardiomyopathy (DCMi, n = 54), and controls (n = 20). In addition, a possible mechanism of RBP 4 regulation was examined in adipocytes in vitro. Plasma levels of RBP 4 and insulin were measured by a specific ELISA. Interleukin concentrations were obtained by multiplex ELISA. Cell culture with 3T3-L1 adipocytes was performed to measure RBP 4 mRNA expression after stimulation with IL-8. RBP 4 levels were significantly increased in patients with DCMi (52.95 +/- 20.42 microg/mL) compared with DCM (35.54 +/- 23.08 microg/mL) and the control group (27.3 +/- 18.51 microg/mL). RBP 4 was positively correlated with IL-8 (r=0.416, P < 0.05) in human plasma in patients with DCMi. Moreover, increased insulin resistance was observed in patients with DCMi compared with the control and DCM groups. In vitro, IL-8 induced a significant upregulation of RBP 4 mRNA expression in adipocytes. Elevated RBP 4 plasma concentrations, induced by IL-8, might be one mechanism leading to a higher incidence of diabetes in patients with DCMi.

  9. Carbonyl Compounds Generated from Electronic Cigarettes

    Directory of Open Access Journals (Sweden)

    Kanae Bekki

    2014-10-01

    Full Text Available Electronic cigarettes (e-cigarettes are advertised as being safer than tobacco cigarettes products as the chemical compounds inhaled from e-cigarettes are believed to be fewer and less toxic than those from tobacco cigarettes. Therefore, continuous careful monitoring and risk management of e-cigarettes should be implemented, with the aim of protecting and promoting public health worldwide. Moreover, basic scientific data are required for the regulation of e-cigarette. To date, there have been reports of many hazardous chemical compounds generated from e-cigarettes, particularly carbonyl compounds such as formaldehyde, acetaldehyde, acrolein, and glyoxal, which are often found in e-cigarette aerosols. These carbonyl compounds are incidentally generated by the oxidation of e-liquid (liquid in e-cigarette; glycerol and glycols when the liquid comes in contact with the heated nichrome wire. The compositions and concentrations of these compounds vary depending on the type of e-liquid and the battery voltage. In some cases, extremely high concentrations of these carbonyl compounds are generated, and may contribute to various health effects. Suppliers, risk management organizations, and users of e-cigarettes should be aware of this phenomenon.

  10. Carbonyl compounds generated from electronic cigarettes.

    Science.gov (United States)

    Bekki, Kanae; Uchiyama, Shigehisa; Ohta, Kazushi; Inaba, Yohei; Nakagome, Hideki; Kunugita, Naoki

    2014-10-28

    Electronic cigarettes (e-cigarettes) are advertised as being safer than tobacco cigarettes products as the chemical compounds inhaled from e-cigarettes are believed to be fewer and less toxic than those from tobacco cigarettes. Therefore, continuous careful monitoring and risk management of e-cigarettes should be implemented, with the aim of protecting and promoting public health worldwide. Moreover, basic scientific data are required for the regulation of e-cigarette. To date, there have been reports of many hazardous chemical compounds generated from e-cigarettes, particularly carbonyl compounds such as formaldehyde, acetaldehyde, acrolein, and glyoxal, which are often found in e-cigarette aerosols. These carbonyl compounds are incidentally generated by the oxidation of e-liquid (liquid in e-cigarette; glycerol and glycols) when the liquid comes in contact with the heated nichrome wire. The compositions and concentrations of these compounds vary depending on the type of e-liquid and the battery voltage. In some cases, extremely high concentrations of these carbonyl compounds are generated, and may contribute to various health effects. Suppliers, risk management organizations, and users of e-cigarettes should be aware of this phenomenon.

  11. Association of Plasma Heat Shock Protein 70, Interleukin 6, and Creatine Kinase Concentrations in a Healthy, Young Adult Population

    Directory of Open Access Journals (Sweden)

    Carmen Contreras-Sesvold

    2015-01-01

    Full Text Available Variations of baseline plasma concentrations of creatine kinase (CK, heat shock protein 70 (HSP70, and interleukin 6 (IL-6 have been reported. We report categorical associations which may influence these protein levels. Methods. Blood was harvested for DNA and plasma protein analysis from 567 adults. Mean protein levels of CK, HSP70, and IL-6 were compared by sex, ethnicity, genetic variants—CKMM Nco1 (rs1803285, HSPA1B +A1538G (rs1061581, and IL6 G-174C (rs1800795—self-reported history of exercise, oral contraceptive use, and dietary supplement use. Results. SNP major allele frequencies for CKMM, HSPA1B, and IL6 were 70% A, 57% A, and 60%. Mean CK statistically differed by sex, ethnicity, oral contraceptives, and caffeine. Plasma HSP70 differed by caffeine and protein. Mean IL-6 concentration differed by sex, ethnicity, and genotype. Plasma IL-6 was significantly lower (29% in males (1.92 ± 0.08 pg/mL and higher (29% among African Americans (2.85 ± 0.50 pg/mL relative to the others. IL6 G-174C GG genotype (2.23 ± 0.14 pg/mL was 19% greater than CG or CC genotypes. Conclusion. Differences in baseline CK and IL-6 plasma protein concentrations are associated with genetics, sex, ethnicity, and the use of oral contraceptives, caffeine, and protein supplements in this young and athletic population.

  12. Site-Specific Zwitterionic Polymer Conjugates of a Protein Have Long Plasma Circulation.

    Science.gov (United States)

    Bhattacharjee, Somnath; Liu, Wenge; Wang, Wei-Han; Weitzhandler, Isaac; Li, Xinghai; Qi, Yizhi; Liu, Jinyao; Pang, Yan; Hunt, Donald F; Chilkoti, Ashutosh

    2015-11-01

    Many proteins suffer from suboptimal pharmacokinetics (PK) that limit their utility as drugs. The efficient synthesis of polymer conjugates of protein drugs with tunable PK to optimize their in vivo efficacy is hence critical. We report here the first study of the in vivo behavior of a site-specific conjugate of a zwitterionic polymer and a protein. To synthesize the conjugate, we first installed an initiator for atom-transfer radical polymerization (ATRP) at the N terminus of myoglobin (Mb-N-Br). Subsequently, in situ ATRP was carried out in aqueous buffer to grow an amine-functionalized polymer from Mb-N-Br. The cationic polymer was further derivatized to two zwitterionic polymers by treating the amine groups of the cationic polymer with iodoacetic acid to obtain poly(carboxybetaine methacrylate) with a one-carbon spacer (PCBMA; C1 ), and sequentially with 3-iodopropionic acid and iodoacetic acid to obtain PCBMA(mix) with a mixture of C1 and C2 spacers. The Mb-N-PCBMA polymer conjugates had a longer in vivo plasma half-life than a PEG-like comb polymer conjugate of similar molecular weights (MW). The structure of the zwitterion plays a role in controlling the in vivo behavior of the conjugate, as the PCBMA conjugate with a C1 spacer had significantly longer plasma circulation than the conjugate with a mixture of C1 and C2 spacers.

  13. Intravenous delivery of hydrophobin-functionalized porous silicon nanoparticles: stability, plasma protein adsorption and biodistribution.

    Science.gov (United States)

    Sarparanta, Mirkka; Bimbo, Luis M; Rytkönen, Jussi; Mäkilä, Ermei; Laaksonen, Timo J; Laaksonen, Päivi; Nyman, Markus; Salonen, Jarno; Linder, Markus B; Hirvonen, Jouni; Santos, Hélder A; Airaksinen, Anu J

    2012-03-01

    Rapid immune recognition and subsequent elimination from the circulation hampers the use of many nanomaterials as carriers to targeted drug delivery and controlled release in the intravenous route. Here, we report the effect of a functional self-assembled protein coating on the intravenous biodistribution of (18)F-labeled thermally hydrocarbonized porous silicon (THCPSi) nanoparticles in rats. (18)F-Radiolabeling enables the sensitive and easy quantification of nanoparticles in tissues using radiometric methods and allows imaging of the nanoparticle biodistribution with positron emission tomography. Coating with Trichoderma reesei HFBII altered the hydrophobicity of (18)F-THCPSi nanoparticles and resulted in a pronounced change in the degree of plasma protein adsorption to the nanoparticle surface in vitro. The HFBII-THCPSi nanoparticles were biocompatible in RAW 264.7 macrophages and HepG2 liver cells making their intravenous administration feasible. In vivo, the distribution of the nanoparticles between the liver and spleen, the major mononuclear phagocyte system organs in the body, was altered compared to that of uncoated (18)F-THCPSi. Identification of the adsorbed proteins revealed that certain opsonins and apolipoproteins are enriched in HFBII-functionalized nanoparticles, whereas the adsorption of abundant plasma components such as serum albumin and fibrinogen is decreased.

  14. Plasma lactate, GH and GH-binding protein levels in exercise following BCAA supplementation in athletes.

    Science.gov (United States)

    De Palo, E F; Gatti, R; Cappellin, E; Schiraldi, C; De Palo, C B; Spinella, P

    2001-01-01

    Branched chain amino acids (BCAA) stimulate protein synthesis, and growth hormone (GH) is a mediator in this process. A pre-exercise BCAA ingestion increases muscle BCAA uptake and use. Therefore after one month of chronic BCAA treatment (0.2 gkg(-1) of body weight), the effects of a pre-exercise oral supplementation of BCAA (9.64 g) on the plasma lactate (La) were examined in triathletes, before and after 60 min of physical exercise (75% of VO2 max). The plasma levels of GH (pGH) and of growth hormone binding protein (pGHBP) were also studied. The end-exercise La of each athlete was higher than basal. Furthermore, after the chronic BCAA treatment, these end-exercise levels were lower than before this treatment (8.6+/-0.8 mmol L(-1) after vs 12.8+/-1.0 mmol L(-1) before treatment; p BCAA chronic treatment, this end-exercise pGHBP was 738+/-85 pmol L(-1) before vs 1691+/-555 pmol L(-1) after. pGH/pGHBP ratio was unchanged in each athlete and between the groups, but a tendency to increase was observed at end-exercise. The lower La at the end of an intense muscular exercise may reflect an improvement of BCAA use, due to the BCAA chronic treatment. The chronic BCAA effects on pGH and pGHBP might suggest an improvement of muscle activity through protein synthesis.

  15. Changes in plasma protein levels as an early indication of a bloodstream infection

    Science.gov (United States)

    Joenväärä, Sakari; Kaartinen, Johanna; Järvinen, Asko; Renkonen, Risto

    2017-01-01

    Blood culture is the primary diagnostic test performed in a suspicion of bloodstream infection to detect the presence of microorganisms and direct the treatment. However, blood culture is slow and time consuming method to detect blood stream infections or separate septic and/or bacteremic patients from others with less serious febrile disease. Plasma proteomics, despite its challenges, remains an important source for early biomarkers for systemic diseases and might show changes before direct evidence from bacteria can be obtained. We have performed a plasma proteomic analysis, simultaneously at the time of blood culture sampling from ten blood culture positive and ten blood culture negative patients, and quantified 172 proteins with two or more unique peptides. Principal components analysis, Orthogonal Projections to Latent Structures Discriminant Analysis (OPLS-DA) and ROC curve analysis were performed to select protein(s) features which can classify the two groups of samples. We propose a number of candidates which qualify as potential biomarkers to select the blood culture positive cases from negative ones. Pathway analysis by two methods revealed complement activation, phagocytosis pathway and alterations in lipid metabolism as enriched pathways which are relevant for the condition. Data are available via ProteomeXchange with identifier PXD005022. PMID:28235076

  16. The modulation of platelet adhesion and activation by chitosan through plasma and extracellular matrix proteins.

    Science.gov (United States)

    Lord, Megan S; Cheng, Bill; McCarthy, Simon J; Jung, MoonSun; Whitelock, John M

    2011-10-01

    Chitosan has been shown to promote initial wound closure events to prevent blood loss. Platelet adhesion and activation are crucial early events in these processes after traumatic bleeding leading to thrombus formation. Platelet adhesion to chitosan was found to be enhanced in the presence of adsorbed plasma and extracellular matrix proteins and was found to be primarily mediated by α(IIb)β(3) integrins, while α(2)β(1) integrins were found to be involved in platelet adhesion to collagen and perlecan. Platelets were found to be activated by chitosan, as shown by an increase in the expression of α(IIb)β(3) integrins and P-selectin, while the extent of activation was modulated by the presence of proteins including perlecan and fibrinogen. Collagen-coated chitosan was found to activate platelets to the same extent as either chitosan or collagen alone. These data support the role of plasma and extracellular matrix proteins in promoting chitosan mediated platelet adhesion and activation supporting the hypothesis that chitosan promotes wound healing via these interactions.

  17. Rapid Changes in Plasma Membrane Protein Phosphorylation during Initiation of Cell Wall Digestion 1

    Science.gov (United States)

    Blowers, David P.; Boss, Wendy F.; Trewavas, Anthony J.

    1988-01-01

    Plasma membrane vesicles from wild carrot cells grown in suspension culture were isolated by aqueous two-phase partitioning, and ATP-dependent phosphorylation was measured with [γ-32P]ATP in the presence and absence of calcium. Treatment of the carrot cells with the cell wall digestion enzymes, driselase, in a sorbitol osmoticum for 1.5 min altered the protein phosphorylation pattern compared to that of cells treated with sorbitol alone. Driselase treatment resulted in decreased phosphorylation of a band of Mr 80,000 which showed almost complete calcium dependence in the osmoticum treated cells; decreased phosphorylation of a band of Mr 15,000 which showed little calcium activation, and appearance of a new band of calcium-dependent phosphorylation at Mr 22,000. These effects appeared not to be due to nonspecific protease activity and neither in vivo nor in vitro exposure to driselase caused a significant loss of Coomassie blue-staining bands on the gels of the isolated plasma membranes. However, protein phosphorylation was decreased. Adding driselase to the in vitro reaction mixture caused a general decrease in the membrane protein phosphorylation either in the presence or absence of calcium which did not mimic the in vivo response. Cells labeled in vivo with inorganic 32P also showed a response to the Driselase treatment. An enzymically active driselase preparation was required for the observed responses. Images Fig. 1 Fig. 3 Fig. 4 Fig. 5 PMID:16665936

  18. Rapid Changes in Plasma Membrane Protein Phosphorylation during Initiation of Cell Wall Digestion.

    Science.gov (United States)

    Blowers, D P; Boss, W F; Trewavas, A J

    1988-02-01

    Plasma membrane vesicles from wild carrot cells grown in suspension culture were isolated by aqueous two-phase partitioning, and ATP-dependent phosphorylation was measured with [gamma-(32)P]ATP in the presence and absence of calcium. Treatment of the carrot cells with the cell wall digestion enzymes, driselase, in a sorbitol osmoticum for 1.5 min altered the protein phosphorylation pattern compared to that of cells treated with sorbitol alone. Driselase treatment resulted in decreased phosphorylation of a band of M(r) 80,000 which showed almost complete calcium dependence in the osmoticum treated cells; decreased phosphorylation of a band of M(r) 15,000 which showed little calcium activation, and appearance of a new band of calcium-dependent phosphorylation at M(r) 22,000. These effects appeared not to be due to nonspecific protease activity and neither in vivo nor in vitro exposure to driselase caused a significant loss of Coomassie blue-staining bands on the gels of the isolated plasma membranes. However, protein phosphorylation was decreased. Adding driselase to the in vitro reaction mixture caused a general decrease in the membrane protein phosphorylation either in the presence or absence of calcium which did not mimic the in vivo response. Cells labeled in vivo with inorganic (32)P also showed a response to the Driselase treatment. An enzymically active driselase preparation was required for the observed responses.

  19. Impaired cardiac SIRT1 activity by carbonyl stress contributes to aging-related ischemic intolerance.

    Directory of Open Access Journals (Sweden)

    Chunhu Gu

    Full Text Available Reactive aldehydes can initiate protein oxidative damage which may contribute to heart senescence. Sirtuin 1 (SIRT1 is considered to be a potential interventional target for I/R injury management in the elderly. We hypothesized that aldehyde mediated carbonyl stress increases susceptibility of aged hearts to ischemia/reperfusion (I/R injury, and elucidate the underlying mechanisms with a focus on SIRT1. Male C57BL/6 young (4-6 mo and aged (22-24 mo mice were subjected to myocardial I/R. Cardiac aldehyde dehydrogenase (ALDH2, SIRT1 activity and protein carbonyls were assessed. Our data revealed that aged heart exhibited increased endogenous aldehyde/carbonyl stress due to impaired ALDH2 activity concomitant with blunted SIRT1 activity (P<0.05. Exogenous toxic aldehydes (4-HNE exposure in isolated cardiomyocyte verified that aldehyde-induced carbonyl modification on SIRT1 impaired SIRT1 activity leading to worse hypoxia/reoxygenation (H/R injury, which could all be rescued by Alda-1 (ALDH2 activator (all P<0.05. However, SIRT1 inhibitor blocked the protective effect of Alda-1 on H/R cardiomyocyte. Interestingly, myocardial I/R leads to higher carbonylation but lower activity of SIRT1 in aged hearts than that seen in young hearts (P<0.05. The application of Alda-1 significantly reduced the carbonylation on SIRT1 and markedly improved the tolerance to in vivo I/R injury in aged hearts, but failed to protect Sirt1(+/- knockout mice against myocardial I/R injury. This was verified by Alda-1 treatment improved postischemic contractile function recovery in ex vivo perfused aged but not in Sirt1(+/- hearts. Thus, aldehyde/carbonyl stress is accelerated in aging heart. These results provide a new insight that impaired cardiac SIRT1 activity by carbonyl stress plays a critical role in the increased susceptibility of aged heart to I/R injury. ALDH2 activation can restore this aging-related myocardial ischemic intolerance.

  20. Increased plasma ghrelin suppresses insulin release in wethers fed with a high-protein diet.

    Science.gov (United States)

    Takahashi, T; Sato, K; Kato, S; Yonezawa, T; Kobayashi, Y; Ohtani, Y; Ohwada, S; Aso, H; Yamaguchi, T; Roh, S G; Katoh, K

    2014-06-01

    Ghrelin is a multifunctional peptide that promotes an increase of food intake and stimulates GH secretion. Ghrelin secretion is regulated by nutritional status and nutrients. Although a high-protein (HP) diet increases plasma ghrelin secretion in mammals, the mechanisms and the roles of the elevated ghrelin concentrations due to a HP diet have not been fully established. To clarify the roles of elevated acylated ghrelin upon intake of a HP diet, we investigated the regulation of ghrelin concentrations in plasma and tissues in wethers fed with either the HP diet or the control (CNT) diet for 14 days, and examined the action of the elevated plasma ghrelin by using a ghrelin-receptor antagonist. The HP diet gradually increased the plasma acylated-ghrelin concentrations, but the CNT diet did not. Although the GH concentrations did not vary significantly across the groups, an injection of ghrelin-receptor antagonist enhanced insulin levels in circulation in the HP diet group. In the fundus region of the stomach, the ghrelin levels did not differ between the HP and CNT diet groups, whereas ghrelin O-acyltransferase mRNA levels were higher in the group fed with HP diet than those of the CNT diet group were. These results indicate that the HP diet elevated the plasma ghrelin levels by increasing its synthesis; this elevation strongly suppresses the appearance of insulin in the circulation of wethers, but it is not involved in GH secretion. Overall, our findings indicate a role of endogenous ghrelin action in secretion of insulin, which acts as a regulator after the consumption of a HP diet.

  1. Maternal Plasma Retinol Binding Protein 4 in Acute Pyelonephritis during Pregnancy

    Science.gov (United States)

    Vaisbuch, Edi; Romero, Roberto; Mazaki-Tovi, Shali; Kusanovic, Juan Pedro; Chaiworapongsa, Tinnakorn; Dong, Zhong; Kim, Sun Kwon; Ogge, Giovanna; Gervasi, Maria Teresa; Hassan, Sonia S.

    2010-01-01

    Objective Adipokines have been implicated in metabolic regulation and the immune response thus providing a molecular mechanism for the interaction between these two systems. Retinol binding protein 4 (RBP4) is a novel adipokine that plays a role in the pathophysiology of obesity-induced insulin resistance, as well as in the modulation of inflammation. The aim of this study was to determine whether there are changes in maternal plasma concentrations of RBP4 in pregnant women with acute pyelonephritis. Study design This cross-sectional study included pregnant women in the following groups: 1) normal pregnancy (n=80); 2) pyelonephritis (n=39). Maternal plasma RBP4 concentrations were determined by enzyme-linked immunoassays. Non-parametric statistics were used for analyses. Results 1) The median maternal plasma RBP4 concentration was lower in patients with acute pyelonephritis than in those with a normal pregnancy (3709.6 ng/mL, IQR 2917.7-5484.2 vs. 9167.6 ng/mL, IQR 7496.1-10384.1, ppyelonephritis who had a positive blood culture and those with a negative culture (3285.3 ng/mL, IQR 2274.1-4741.1 vs. 3922.6 ng/mL, IQR 3126.8-5547.1, respectively, p=0.2); and 3) lower maternal plasma RBP4 concentrations were independently associated with pyelonephritis after adjustment for confounding factors. Conclusions In contrast to what has been reported in preeclampsia, acute pyelonephritis during pregnancy is associated with lower maternal plasma RBP4 concentrations than in normal pregnancy. This finding suggests that the acute maternal inflammatory process associated with pyelonephritis is fundamentally different from that of the chronic systemic inflammatory process suggested in preeclampsia, in which RBP4 concentrations were found to be elevated. PMID:20163326

  2. Plasma-treated polystyrene film that enhances binding efficiency for sensitive and label-free protein biosensing

    Science.gov (United States)

    Guo, Bihong; Li, Shaopeng; Song, Lusheng; Yang, Mo; Zhou, Wenfei; Tyagi, Deependra; Zhu, Jinsong

    2015-08-01

    A plasma-treated ultrathin polystyrene (PS) film surface was explored as a simple, robust, and low-cost surface chemistry solution for protein biosensing applications. This surface could dramatically improve the binding efficiency of the protein-protein interactions, which is defined as the binding signal per immobilized ligand. The PS-modified protein biosensor was readily fabricated by spin coating and plasma treatment. Various parameters for fabrication, including the concentration of the PS solution, rate of spin coating, and duration of plasma treatment, were systematically optimized based on the improvement of fluorescence signal yielded by the microfluidic network-aided fluorescence immunoassay. The performance of the label-free protein detection on the optimized surfaces was further evaluated by surface plasmon resonance imaging (SPRi). PS surfaces with optimal fabrication parameters exhibited up to an 620% enhancement of the protein binding response and approximately 210% of the protein binding per immobilized protein ligand compared with a self-assembled monolayer (SAM) surface of 11-mercapto undecanoic acid (MUA). The relationship between the fabrication parameters used and changes to the surface chemistry and the morphological properties were characterized with atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FTIR). It was revealed that the morphological changes observed in the plasma-treated PS film were the dominant factor for the improvement of the protein bioassay performance, rather than the chemical changes.

  3. Colorectal resection is associated with persistent proangiogenic plasma protein changes: postoperative plasma stimulates in vitro endothelial cell growth, migration, and invasion.

    Science.gov (United States)

    Kumara, H M C Shantha; Shantha Kumara, H M C; Feingold, Daniel; Kalady, Matthew; Dujovny, Nadav; Senagore, Anthony; Hyman, Neil; Cekic, Vesna; Whelan, Richard L

    2009-06-01

    Plasma vascular endothelial growth factor (VEGF) levels are elevated for weeks after minimally invasive colorectal resection (MICR). Decreased plasma angiopoietin-(Ang) 1 and increased Ang-2 levels have been noted on postoperative days (POD) 1 and 3. These proangiogenic changes may stimulate tumor growth postoperatively (postop). This study's purpose was to track plasma VEGF, Ang-1, and Ang-2 levels for 4 to 8 weeks after MICR for cancer and to assess the impact of preoperative (preop) and postop plasma on in vitro endothelial cell (EC) behavior. Blood samples from 105 MICR patients were taken preop, on POD 5 and at varying time points for 2 months. Samples from 7 day time blocks after POD 5 were bundled to permit statistical analysis. Plasma protein levels were measured via enzyme-linked immunosorbent assay. In vitro EC branch point formation, EC invasion, and EC migration assays were carried out with preop, POD 7 to 13 and 14 to 20 plasma. The t test and Bonferonni correction was used. VEGF levels were significantly elevated on POD 5 and 7 to 13; lesser increases were noted on POD 14 to 20 and 21 to 27. Ang-2 levels were significantly increased at all time points postop. No significant Ang-1 changes were noted. When compared to preop EC culture results, there was significantly more EC branch point formation, EC invasion, and EC migration assays noted with POD 7 to 13 and POD 14 to 20 plasma. MICR is associated with proangiogenic plasma changes for 2 to 4 weeks and plasma from POD 7 to 13 and 14 to 20 stimulated EC growth, invasion, and migration. Postop plasma may stimulate the growth of residual tumor.

  4. Plasma-treated polystyrene film that enhances binding efficiency for sensitive and label-free protein biosensing

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Bihong [National Center for NanoScience and Technology, No. 11 Beiyitiao, Zhongguancun, Beijing 100190 (China); Li, Shaopeng [National Center for NanoScience and Technology, No. 11 Beiyitiao, Zhongguancun, Beijing 100190 (China); Department of Chemistry, Tsinghua University, Beijing 100084 (China); Song, Lusheng [National Center for NanoScience and Technology, No. 11 Beiyitiao, Zhongguancun, Beijing 100190 (China); Yang, Mo; Zhou, Wenfei; Tyagi, Deependra [National Center for NanoScience and Technology, No. 11 Beiyitiao, Zhongguancun, Beijing 100190 (China); University of Chinese Academy of Sciences, Yuquan Rd., 19(A), Beijing 100049 (China); Zhu, Jinsong, E-mail: jizhu88@gmail.com [National Center for NanoScience and Technology, No. 11 Beiyitiao, Zhongguancun, Beijing 100190 (China)

    2015-08-01

    Highlights: • A simple and robust plasma-treated ultrathin polystyrene film surface was developed for protein biosensing. • The surface was optimized by evaluating up to 120 types of fabrication parameters with high-throughput analytical methods. • The optimized surface showed a 620% improvement of the protein detection signal and 210% protein binding per immobilized protein ligand compared with a self-assembled monolayer surface. - Abstract: A plasma-treated ultrathin polystyrene (PS) film surface was explored as a simple, robust, and low-cost surface chemistry solution for protein biosensing applications. This surface could dramatically improve the binding efficiency of the protein–protein interactions, which is defined as the binding signal per immobilized ligand. The PS-modified protein biosensor was readily fabricated by spin coating and plasma treatment. Various parameters for fabrication, including the concentration of the PS solution, rate of spin coating, and duration of plasma treatment, were systematically optimized based on the improvement of fluorescence signal yielded by the microfluidic network-aided fluorescence immunoassay. The performance of the label-free protein detection on the optimized surfaces was further evaluated by surface plasmon resonance imaging (SPRi). PS surfaces with optimal fabrication parameters exhibited up to an 620% enhancement of the protein binding response and approximately 210% of the protein binding per immobilized protein ligand compared with a self-assembled monolayer (SAM) surface of 11-mercapto undecanoic acid (MUA). The relationship between the fabrication parameters used and changes to the surface chemistry and the morphological properties were characterized with atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FTIR). It was revealed that the morphological changes observed in the plasma-treated PS film were the dominant factor for the

  5. Boar seminal plasma exosomes: effect on sperm function and protein identification by sequencing.

    Science.gov (United States)

    Piehl, Lidia L; Fischman, M Laura; Hellman, Ulf; Cisale, Humberto; Miranda, Patricia V

    2013-04-15

    Mammalian seminal plasma contains membranous vesicles (exosomes), with a high content of cholesterol and sphingomyelin and a complex protein composition. Their physiological role is uncertain because sperm stabilization and activation effects have been reported. To analyze a putative modulatory role for semen exosomes on sperm activity in the boar, the effects of these vesicles on several sperm functional parameters were examined. Additionally, boar exosome proteins were sequenced and their incorporation into sperm was explored. Boar sperm were incubated under conditions that induce capacitation, manifested as increased tyrosine phosphorylation, cholesterol loss and greater fluidity in apical membranes, and the ability to undergo the lysophosphatidylcholine-induced acrosome reaction. After establishing this cluster of capacitation-dependent functional parameters, the effect produced by exosomes when present during or after sperm capacitation was analyzed. Exosomes inhibited the capacitation-dependent cholesterol efflux and fluidity increase in apical membranes, and the disappearance of a 14-kD phosphorylated polypeptide. In contrast, the acrosome reaction (spontaneous and lysophosphatidylcholine-induced) was not affected, and sperm binding to the oocyte zona pellucida was reduced only when vesicles were present during gamete coincubation. Liposomes with a lipid composition similar to that present in exosomes mimicked these effects, except the one on zona pellucida binding. Interaction between exosomes and sperm was confirmed by transfer of aminopeptidase activity. In addition, the major exosome protein, identified as actin, appeared to associate with sperm after coincubation. Exosome composition had a predominance for structural proteins (actin, plastin, ezrin, and condensin), enzymes, and several porcine seminal plasma-specific polypeptides (e.g., spermadhesins). Transfer of proteins from exosome to sperm and their ability to block cholesterol efflux supports a

  6. Lipid-protein interactions in plasma membranes of fiber cells isolated from the human eye lens.

    Science.gov (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K

    2014-03-01

    The protein content in human lens membranes is extremely high, increases with age, and is higher in the nucleus as compared with the cortex, which should strongly affect the organization and properties of the lipid bilayer portion of intact membranes. To assess these effects, the intact cortical and nuclear fiber cell plasma membranes isolated from human lenses from 41- to 60-year-old donors were studied using electron paramagnetic resonance spin-labeling methods. Results were compared with those obtained for lens lipid membranes prepared from total lipid extracts from human eyes of the same age group [Mainali, L., Raguz, M., O'Brien, W. J., and Subczynski, W. K. (2013) Biochim. Biophys. Acta]. Differences were considered to be mainly due to the effect of membrane proteins. The lipid-bilayer portions of intact membranes were significantly less fluid than lipid bilayers of lens lipid membranes, prepared without proteins. The intact membranes were found to contain three distinct lipid environments termed the bulk lipid domain, boundary lipid domain, and trapped lipid domain. However, the cholesterol bilayer domain, which was detected in cortical and nuclear lens lipid membranes, was not detected in intact membranes. The relative amounts of bulk and trapped lipids were evaluated. The amount of lipids in domains uniquely formed due to the presence of membrane proteins was greater in nuclear membranes than in cortical membranes. Thus, it is evident that the rigidity of nuclear membranes is greater than that of cortical membranes. Also the permeability coefficients for oxygen measured in domains of nuclear membranes were significantly lower than appropriate coefficients measured in cortical membranes. Relationships between the organization of lipids into lipid domains in fiber cells plasma membranes and the organization of membrane proteins are discussed.

  7. Intrinsic stability of Brassicaceae plasma membrane in relation to changes in proteins and lipids as a response to salinity.

    Science.gov (United States)

    Chalbi, Najla; Martínez-Ballesta, Ma Carmen; Youssef, Nabil Ben; Carvajal, Micaela

    2015-03-01

    Changes in plasma membrane lipids, such as sterols and fatty acids, have been observed as a result of salt stress. These alterations, together with modification of the plasma membrane protein profile, confer changes in the physical properties of the membrane to be taken into account for biotechnological uses. In our experiments, the relationship between lipids and proteins in three different Brassicaceae species differing in salinity tolerance (Brassica oleracea, B. napus and Cakile maritima) and the final plasma membrane stability were studied. The observed changes in the sterol (mainly an increase in sitosterol) and fatty acid composition (increase in RUFA) in each species led to physical adaptation of the plasma membrane to salt stress. The in vitro vesicles stability was higher in the less tolerant (B. oleracea) plants together with low lipoxygenase activity. These results indicate that the proteins/lipids ratio and lipid composition is an important aspect to take into account for the use of natural vesicles in plant biotechnology.

  8. Plasma protein corona modulates the vascular wall interaction of drug carriers in a material and donor specific manner.

    Directory of Open Access Journals (Sweden)

    Daniel J Sobczynski

    Full Text Available The nanoscale plasma protein interaction with intravenously injected particulate carrier systems is known to modulate their organ distribution and clearance from the bloodstream. However, the role of this plasma protein interaction in prescribing the adhesion of carriers to the vascular wall remains relatively unknown. Here, we show that the adhesion of vascular-targeted poly(lactide-co-glycolic-acid (PLGA spheres to endothelial cells is significantly inhibited in human blood flow, with up to 90% reduction in adhesion observed relative to adhesion in simple buffer flow, depending on the particle size and the magnitude and pattern of blood flow. This reduced PLGA adhesion in blood flow is linked to the adsorption of certain high molecular weight plasma proteins on PLGA and is donor specific, where large reductions in particle adhesion in blood flow (>80% relative to buffer is seen with ∼60% of unique donor bloods while others exhibit moderate to no reductions. The depletion of high molecular weight immunoglobulins from plasma is shown to successfully restore PLGA vascular wall adhesion. The observed plasma protein effect on PLGA is likely due to material characteristics since the effect is not replicated with polystyrene or silica spheres. These particles effectively adhere to the endothelium at a higher level in blood over buffer flow. Overall, understanding how distinct plasma proteins modulate the vascular wall interaction of vascular-targeted carriers of different material characteristics would allow for the design of highly functional delivery vehicles for the treatment of many serious human diseases.

  9. Plasma IgG autoantibody against actin-related protein 3 in liver fluke Opisthorchis viverrini infection.

    Science.gov (United States)

    Rucksaken, R; Haonon, O; Pinlaor, P; Pairojkul, C; Roytrakul, S; Yongvanit, P; Selmi, C; Pinlaor, S

    2015-07-01

    Opisthorchiasis secondary to Opisthorchis viverrini infection leads to cholangiocellular carcinoma through chronic inflammation of the bile ducts and possibly inducing autoimmunity. It was hypothesized that plasma autoantibodies directed against self-proteins are biomarkers for opisthorchiasis. Plasma from patients with opisthorchiasis was tested using proteins derived from immortalized cholangiocyte cell lines, and spots reacting with plasma were excised and subjected to LC-MS/MS. Seven protein spots were recognized by IgG autoantibodies, and the highest matching scored protein was actin-related protein 3 (ARP3). The antibody against ARP3 was tested in plasma from 55 O. viverrini-infected patients, 24 patients with others endemic parasitic infections and 17 healthy controls using Western blot and ELISA. Immunoreactivity against recombinant ARP3 was significantly more prevalent in opisthorchiasis compared to healthy controls at Western blotting and ELISA (P < 0.05). Plasma ARP3 autoantibody titres were also higher in opisthorchiasis compared to healthy individuals (P < 0.01) and other parasitic infections including Strongyloides stercoralis (P < 0.001), echinostome (P < 0.05), hookworms (P < 0.001) and Taenia spp. (P < 0.05). It was further characterized in that the ARP3 autoantibody titre had a sensitivity of 78.18% and specificity of 100% for opisthorchiasis. In conclusion, it may be suggested that plasma anti-ARP3 might represent a new diagnostic antibody for opisthorchiasis.

  10. In vitro plasma protein binding and aqueous aggregation behavior of astaxanthin dilysinate tetrahydrochloride.

    Science.gov (United States)

    Zsila, Ferenc; Fitos, Ilona; Bikádi, Zsolt; Simonyi, Miklós; Jackson, Henry L; Lockwood, Samuel F

    2004-11-01

    The tetrahydrochloride salt of astaxanthin di-L-lysinate (lys(2)AST) is a highly water-dispersible astaxanthin-amino acid conjugate, with an aqueous dispersibility of > or = 181.6 mg/mL. The statistical mixture of stereoisomers has been well characterized as an aqueous-phase superoxide anion scavenger, effective at micromolar (microM) concentrations. In the current study, the aqueous aggregation behavior and in vitro plasma protein binding [with fatty-acid-free human serum albumin (HSA) and alpha(1)-acid glycoprotein (AGP)] were investigated with a suite of techniques, including circular dichroism (CD) and UV-vis spectroscopy, ultrafiltration, competitive ligand displacement, and fluorescence quenching. Induced CD bands obtained in Ringer buffer solution of HSA demonstrated high affinity monomeric binding of the compound at low ligand per protein (L/P) ratios (in aqueous solution alone the carotenoid molecules formed card-pack aggregates). The binding constant ( approximately 10(6)M(-1)) and the binding stoichiometry (approximately 0.2 per albumin molecule) were calculated from CD titration data. CD displacement and ultrafiltration experiments performed with marker ligands of HSA indicated that the ligand binding occurred at a site distinct from the main drug binding sites of HSA (i.e., Sites I and II). At intermediate L/P ratios, both monomeric and aggregated ("chirally complexed") binding occurred simultaneously at distinct sites of the protein. At high L/P ratios, chiral complexation predominantly occurred on the asymmetric protein template. The tentative location of the chirally-complexed aggregation on the HSA template was identified as the large interdomain cleft of HSA, where carotenoid derivatives have been found to bind previously. Only weak binding to AGP was observed. These results suggest that parenteral use of this highly potent, water-dispersible astaxanthin-amino acid conjugate will result in plasma protein association, and plasma protein binding at

  11. Protein receptor-independent plasma membrane remodeling by HAMLET: a tumoricidal protein-lipid complex.

    Science.gov (United States)

    Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L; James, Ho C S; Rydström, Anna; Ngassam, Viviane N; Klausen, Thomas Kjær; Pedersen, Stine Falsig; Lam, Matti; Parikh, Atul N; Svanborg, Catharina

    2015-11-12

    A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This ''protein-centric" view is increasingly challenged by evidence for the involvement of specialized membrane domains in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a ''receptor independent" transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. Finally, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features suggest that HAMLET-induced curvature-dependent membrane conformations serve as surrogate receptors for initiating signal transduction cascades, ultimately leading to cell death.

  12. Microfiltration platform for continuous blood plasma protein extraction from whole blood during cardiac surgery.

    Science.gov (United States)

    Aran, Kiana; Fok, Alex; Sasso, Lawrence A; Kamdar, Neal; Guan, Yulong; Sun, Qi; Ündar, Akif; Zahn, Jeffrey D

    2011-09-07

    This report describes the design, fabrication, and testing of a cross-flow filtration microdevice, for the continuous extraction of blood plasma from a circulating whole blood sample in a clinically relevant environment to assist in continuous monitoring of a patient's inflammatory response during cardiac surgeries involving cardiopulmonary bypass (CPB) procedures (about 400,000 adult and 20,000 pediatric patients in the United States per year). The microfiltration system consists of a two-compartment mass exchanger with two aligned sets of PDMS microchannels, separated by a porous polycarbonate (PCTE) membrane. Using this microdevice, blood plasma has been continuously separated from blood cells in a real-time manner with no evidence of bio-fouling or cell lysis. The technology is designed to continuously extract plasma containing diagnostic plasma proteins such as complements and cytokines using a significantly smaller blood volume as compared to traditional blood collection techniques. The microfiltration device has been tested using a simulated CPB circulation loop primed with donor human blood, in a manner identical to a clinical surgical setup, to collect plasma fractions in order to study the effects of CPB system components and circulation on immune activation during extracorporeal circulatory support. The microdevice, with 200 nm membrane pore size, was connected to a simulated CPB circuit, and was able to continuously extract ~15% pure plasma volume (100% cell-free) with high sampling frequencies which could be analyzed directly following collection with no need to further centrifuge or modify the fraction. Less than 2.5 ml total plasma volume was collected over a 4 h sampling period (less than one Vacutainer blood collection tube volume). The results tracked cytokine concentrations collected from both the reservoir and filtrate samples which were comparable to those from direct blood draws, indicating very high protein recovery of the microdevice

  13. Hypoxia facilitates neurogenic dural plasma protein extravasation in mice : a novel animal model for migraine pathophysiology

    OpenAIRE

    Anika Hunfeld; Daniel Segelcke; Ingo Bäcker; Badreddine Mecheri; Kathrin Hemmer; Elisabeth Dlugosch; Michael Andriske; Frank Paris; Xinran Zhu; Hermann Lübbert

    2015-01-01

    Migraine animal models generally mimic the onset of attacks and acute treatment processes. A guinea pig model used the application of meta-chlorophenylpiperazine (mCPP) to trigger immediate dural plasma protein extravasation (PPE) mediated by 5-HT2B receptors. This model has predictive value for antimigraine drugs but cannot explain the delayed onset of efficacy of 5-HT2B receptor antagonists when clinically used for migraine prophylaxis. We found that mCPP failed to induce dural PPE in mice....

  14. [Determination of capillary plasma C-reactive protein during therapy for acute infectious lung diseases].

    Science.gov (United States)

    Makarenko, V V; Vavilikhina, N F; Kastrikina, T N; El'chaninova, S A

    2011-06-01

    Changes in the concentration of C-reactive protein (CRP), leukocytes, erythrocyte sedimentation rate, and differential blood count were comparatively estimated in the treatment of 66 infants (aged 1.12 +/- 0.95 years) with acute infectious lung diseases. There was a high correlation between capillary plasma and venous serum CRP concentrations. On the first day of effective antibiotic therapy, there was a significant decrease in CRP levels; the sensitivity and specificity were 96 and 94%, respectively. Thus, measurement of capillary blood CRP is an accessible and informative tool to monitor therapy for infectious lung diseases in infants.

  15. Regulatory effect of porcine plasma protein hydrolysates on pasting and gelatinization action of corn starch.

    Science.gov (United States)

    Kong, Baohua; Niu, Haili; Sun, Fangda; Han, Jianchun; Liu, Qian

    2016-01-01

    The objective of this study was to investigate the regulatory effect of porcine plasma protein hydrolysates (PPPH) on the physicochemical, pasting, and gelatinization properties of corn starch (CS). The results showed that the solubility of CS markedly increased, whereas swelling power and gel penetration force decreased with increased PPPH concentration (Pgelatinization temperature as determined in differential scanning calorimetry (DSC) (Pgelatinization. Atomic force microscopy (AFM) images indicated that the blocklet sizes of gelatinized CS-PPPH mixtures were smaller and more uniform than native CS. The results proved that pasting and gelatinization abilities of CS can be effectively influenced by adding PPPH. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. [Interaction of FABP4 with plasma membrane proteins of endothelial cells].

    Science.gov (United States)

    Saavedra, Paula; Girona, Josefa; Aragonès, Gemma; Cabré, Anna; Guaita, Sandra; Heras, Mercedes; Masana, Lluís

    2015-01-01

    Fatty acid binding protein (FABP4) is an adipose tissue-secreted adipokine implicated in the regulation of the energetic metabolism and inflammation. High levels of circulating FABP4 have been described in people with obesity, atherogenic dyslipidemia, diabetes and metabolic syndrome. Recent studies have demonstrated that FABP4 could have a direct effect on peripheral tissues and, specifically, on vascular function. It is still unknown how the interaction between FABP4 and the endothelial cells is produced to prompt these effects on vascular function. The objective of this work is studying the interaction between FABP4 and the plasma membrane proteins of endothelial cells. HUVEC cells were incubated with and without FABP4 (100 ng/ml) for 5 minutes. Immunolocalization of FABP4 was studied by confocal microscopy. The results showed that FABP4 colocalizates with CD31, a membrane protein marker. A strategy which combines 6XHistidine-tag FABP4 (FABP4-His), incubations with or without FABP4-His (100 ng/ml), formaldehyde cross-linking, cellular membrane protein extraction and western blot, was designed to study the FABP4 interactions with membrane proteins of HUVECs. The results showed different western blot profiles depending of the incubation with or without FABP4-His. The immunoblot revelead three covalent protein complexes of about 108, 77 and 33 kDa containing FAPB4 and its putative receptor. The existence of a specific binding protein complex able to bind FABP4 to endothelial cells is supported by these results. The obtained results will permit us advance in the molecular knowledge of FABP4 effects as well as use this protein and its receptor as therapeutic target to prevent cardiovascular. Copyright © 2014 Sociedad Española de Arteriosclerosis. Published by Elsevier España. All rights reserved.

  17. A physiologically based pharmacokinetic model to predict the pharmacokinetics of highly protein-bound drugs and the impact of errors in plasma protein binding.

    Science.gov (United States)

    Ye, Min; Nagar, Swati; Korzekwa, Ken

    2016-04-01

    Predicting the pharmacokinetics of highly protein-bound drugs is difficult. Also, since historical plasma protein binding data were often collected using unbuffered plasma, the resulting inaccurate binding data could contribute to incorrect predictions. This study uses a generic physiologically based pharmacokinetic (PBPK) model to predict human plasma concentration-time profiles for 22 highly protein-bound drugs. Tissue distribution was estimated from in vitro drug lipophilicity data, plasma protein binding and the blood: plasma ratio. Clearance was predicted with a well-stirred liver model. Underestimated hepatic clearance for acidic and neutral compounds was corrected by an empirical scaling factor. Predicted values (pharmacokinetic parameters, plasma concentration-time profile) were compared with observed data to evaluate the model accuracy. Of the 22 drugs, less than a 2-fold error was obtained for the terminal elimination half-life (t1/2 , 100% of drugs), peak plasma concentration (Cmax , 100%), area under the plasma concentration-time curve (AUC0-t , 95.4%), clearance (CLh , 95.4%), mean residence time (MRT, 95.4%) and steady state volume (Vss , 90.9%). The impact of fup errors on CLh and Vss prediction was evaluated. Errors in fup resulted in proportional errors in clearance prediction for low-clearance compounds, and in Vss prediction for high-volume neutral drugs. For high-volume basic drugs, errors in fup did not propagate to errors in Vss prediction. This is due to the cancellation of errors in the calculations for tissue partitioning of basic drugs. Overall, plasma profiles were well simulated with the present PBPK model. Copyright © 2016 John Wiley & Sons, Ltd.

  18. Proteins involved in the Vroman effect during exposure of human blood plasma to glass and polyethylene

    NARCIS (Netherlands)

    Turbill, P.; Beugeling, T.; Poot, A.A.

    1996-01-01

    The amounts of fibrinogen adsorbed to glass from various human blood plasmas have been measured as a function of time. The plasmas were 11 single donor plasmas, pooled plasma, a single donor high molecular weight kininogen (HMWK)-deficient plasma and HMWK-deficient plasma, which had been reconstitut

  19. Plasma membrane calcium ATPase proteins as novel regulators of signal transduction pathways

    Institute of Scientific and Technical Information of China (English)

    Mary; Louisa; Holton; Michael; Emerson; Ludwig; Neyses; Angel; L; Armesilla

    2010-01-01

    Emerging evidence suggests that plasma membrane calcium ATPases (PMCAs) play a key role as regulators of calcium-triggered signal transduction pathways via interaction with partner proteins. PMCAs regulate these pathways by targeting specific proteins to cellular sub-domains where the levels of intracellular freecalcium are kept low by the calcium ejection properties of PMCAs. According to this model, PMCAs have been shown to interact functionally with the calcium-sensitive proteins neuronal nitric oxide synthase, calmodulindependent serine protein kinase, calcineurin and endothelial nitric oxidase synthase. Transgenic animals with altered expression of PMCAs are being used to evaluate the physiological significance of these interactions. To date, PMCA interactions with calcium-dependent partner proteins have been demonstrated to play a crucial role in the pathophysiology of the cardiovascular system via regulation of the nitric oxide and calcineurin/nuclear factor of activated T cells pathways. This new evidence suggests that PMCAs play a more sophisticated role than the mere ejection of calcium from the cells, by acting as modulators of signaling transduction pathways.

  20. Proteomic analysis of seminal plasma from asthenozoospermia patients reveals proteins that affect oxidative stress responses and semen quality.

    Science.gov (United States)

    Wang, Jun; Wang, Jian; Zhang, Hua-Rong; Shi, Hui-Juan; Ma, Duan; Zhao, Hong-Xin; Lin, Biaoyang; Li, Run-Sheng

    2009-07-01

    Asthenozoospermia (AS) is a common cause of human male infertility. In one study, more than 80% of the samples from infertile men had reduced sperm motility. Seminal plasma is a mixture of secretions from the testis, epididymis and several male accessory glands, including the prostate, seminal vesicles and Cowper's gland. Studies have shown that seminal plasma contains proteins that are important for sperm motility. To further explore the pathophysiological character of AS, we separated the seminal plasma proteins from AS patients and healthy donors using sodium dodecyl sulfate polyacrylamide gel electrophoresis and in-gel digestion, and then subjected the proteins to liquid chromatography-mass spectrometry (LC-MS/MS) analysis. A total of 741 proteins were identified in the seminal plasma, with a false discovery rate of 3.3%. Using spectral counting, we found that 45 proteins were threefold upregulated and 56 proteins were threefold downregulated in the AS group when compared with the control. Most of these proteins originated from the epididymis and prostate. This study identified a rich source of biomarker candidates for male infertility and indicates that functional abnormalities of the epididymis and prostate can contribute to AS. We identified DJ-1-a protein that has been shown elsewhere to be involved in the control of oxidative stress (OS)-as a downregulated protein in AS seminal plasma. The levels of DJ-1 in AS seminal plasma were about half of those in the control samples. In addition, the levels of reactive oxygen species were 3.3-fold higher in the AS samples than in the controls. Taken together, these data suggest that downregulation of DJ-1 is involved in OS in semen, and therefore affects the quality of the semen.

  1. Carbonyl Emissions from Gasoline and Diesel Motor Vehicles

    Energy Technology Data Exchange (ETDEWEB)

    Destaillats, Hugo; Jakober, Chris A.; Robert, Michael A.; Riddle, Sarah G.; Destaillats, Hugo; Charles, M. Judith; Green, Peter G.; Kleeman, Michael J.

    2007-12-01

    Carbonyls from gasoline powered light-duty vehicles (LDVs) and heavy-duty diesel powered vehicles (HDDVs) operated on chassis dynamometers were measured using an annular denuder-quartz filter-polyurethane foam sampler with O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine derivatization and chromatography-mass spectrometry analyses. Two internal standards were utilized based on carbonyl recovery, 4-fluorobenzaldehyde forcarbonyls and 6-fluoro-4-chromanone for>_C8 compounds. Gas- and particle-phase emissions for 39 aliphatic and 20 aromatic carbonyls ranged from 0.1 ? 2000 ?g/L fuel for LDVs and 1.8 - 27000 mu g/L fuel for HDDVs. Gas-phase species accounted for 81-95percent of the total carbonyls from LDVs and 86-88percent from HDDVs. Particulate carbonyls emitted from a HDDV under realistic driving conditions were similar to concentrations measured in a diesel particulate matter (PM) standard reference material. Carbonyls accounted for 19percent of particulate organic carbon (POC) emissions from low-emission LDVs and 37percent of POC emissions from three-way catalyst equipped LDVs. This identifies carbonyls as one of the largest classes of compounds in LDV PM emissions. The carbonyl fraction of HDDV POC was lower, 3.3-3.9percent depending upon operational conditions. Partitioning analysis indicates the carbonyls had not achieved equilibrium between the gas- and particle-phase under the dilution factors of 126-584 used in the current study.

  2. Characterisation of polystyrene coatings after plasma immersion ion implantation and adsorption of protein

    CERN Document Server

    Dekker, S; Steel, B; Bilek, M M M; McKenzie, D R; James, M

    2012-01-01

    A polystyrene film spun onto polished silicon substrates was implanted with either nitrogen or argon ions using plasma immersion ion implantation (PIII) and subsequently investigated by X-ray and neutron reflectometry, UV-VIS and FTIR ellipsometry, as well as by FTIR and Raman spectroscopy. The depth profile of the densified carbon structures resulting from the ion collision cascades in the polystyrene coating are clearly observed by both X-ray and neutron reflectometry. Argon ions produce a higher density modified layer at a shallower depth than nitrogen ions. The thickness measured for these graded layers agrees with the expected depths of ion implantation as calculated by SRIM. The sensitivity of X-ray and neutron reflectometry allows resolution of density and hydrogen content gradients within the graphitized layers. The treated layers were found to covalently immobilized protein directly from solution. The tropoelastin protein monolayers immobilized on the surface were characterized. Tropoelastin remained...

  3. Mapping of 79 loci for 83 plasma protein biomarkers in cardiovascular disease

    DEFF Research Database (Denmark)

    Folkersen, Lasse Westergaard; Fauman, Eric; Sabater-Lleal, Maria

    2017-01-01

    Recent advances in highly multiplexed immunoassays have allowed systematic large-scale measurement of hundreds of plasma proteins in large cohort studies. In combination with genotyping, such studies offer the prospect to 1) identify mechanisms involved with regulation of protein expression...... identified 79 genome-wide significant (pmining, manual curation, and network-based methods incorporating information on expression quantitative trait loci (eQTL), we...... propose plausible causal mechanisms for 25 trans-acting loci, including a potential post-translational regulation of stem cell factor by matrix metalloproteinase 9 and receptor-ligand pairs such as RANK-RANK ligand. Using public GWA study data, we further evaluate all 79 loci for their causal effect...

  4. An early nodulin-like protein accumulates in the sieve element plasma membrane of Arabidopsis

    DEFF Research Database (Denmark)

    Khan, Junaid A.; Wang, Qi; Sjölund, Richard D.

    2007-01-01

    that have a similar overall domain structure of an amino-terminal signal peptide, plastocyanin-like copper-binding domain, proline/serine-rich domain, and carboxy-terminal hydrophobic domain. The amino- and carboxy-terminal domains of the 21.5-kD sieve element-specific ENOD are posttranslationally cleaved...... from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both...... floral and vegetative tissues, the sieve element-specific ENOD is expressed only within the phloem. Members of the ENOD subfamily of the cupredoxin superfamily do not appear to bind copper and have unknown functions. Phenotypic analysis of homozygous T-DNA insertion mutants for the gene At3g20570 shows...

  5. Plasma and liver acetaminophen-protein adduct levels in mice after acetaminophen treatment: Dose–response, mechanisms, and clinical implications

    Energy Technology Data Exchange (ETDEWEB)

    McGill, Mitchell R.; Lebofsky, Margitta [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Norris, Hye-Ryun K.; Slawson, Matthew H. [Center for Human Toxicology, University of Utah, Salt Lake City, UT (United States); Bajt, Mary Lynn; Xie, Yuchao; Williams, C. David [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Wilkins, Diana G.; Rollins, Douglas E. [Center for Human Toxicology, University of Utah, Salt Lake City, UT (United States); Jaeschke, Hartmut, E-mail: hjaeschke@kumc.edu [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States)

    2013-06-15

    At therapeutic doses, acetaminophen (APAP) is a safe and effective analgesic. However, overdose of APAP is the principal cause of acute liver failure in the West. Binding of the reactive metabolite of APAP (NAPQI) to proteins is thought to be the initiating event in the mechanism of hepatotoxicity. Early work suggested that APAP-protein binding could not occur without glutathione (GSH) depletion, and likely only at toxic doses. Moreover, it was found that protein-derived APAP-cysteine could only be detected in serum after the onset of liver injury. On this basis, it was recently proposed that serum APAP-cysteine could be used as diagnostic marker of APAP overdose. However, comprehensive dose–response and time course studies have not yet been done. Furthermore, the effects of co-morbidities on this parameter have not been investigated. We treated groups of mice with APAP at multiple doses and measured liver GSH and both liver and plasma APAP-protein adducts at various timepoints. Our results show that protein binding can occur without much loss of GSH. Importantly, the data confirm earlier work that showed that protein-derived APAP-cysteine can appear in plasma without liver injury. Experiments performed in vitro suggest that this may involve multiple mechanisms, including secretion of adducted proteins and diffusion of NAPQI directly into plasma. Induction of liver necrosis through ischemia–reperfusion significantly increased the plasma concentration of protein-derived APAP-cysteine after a subtoxic dose of APAP. While our data generally support the measurement of serum APAP-protein adducts in the clinic, caution is suggested in the interpretation of this parameter. - Highlights: • Extensive GSH depletion is not required for APAP-protein binding in the liver. • APAP-protein adducts appear in plasma at subtoxic doses. • Proteins are adducted in the cell and secreted out. • Coincidental liver injury increases plasma APAP-protein adducts at subtoxic doses

  6. Etching of polymers, proteins and bacterial spores by atmospheric pressure DBD plasma in air

    Science.gov (United States)

    Kuzminova, A.; Kretková, T.; Kylián, O.; Hanuš, J.; Khalakhan, I.; Prukner, V.; Doležalová, E.; Šimek, M.; Biederman, H.

    2017-04-01

    Many studies proved that non-equilibrium discharges generated at atmospheric pressure are highly effective for the bio-decontamination of surfaces of various materials. One of the key processes that leads to a desired result is plasma etching and thus the evaluation of etching rates of organic materials is of high importance. However, the comparison of reported results is rather difficult if impossible as different authors use diverse sources of atmospheric plasma that are operated at significantly different operational parameters. Therefore, we report here on the systematic study of the etching of nine different common polymers that mimic the different structures of more complicated biological systems, bovine serum albumin (BSA) selected as the model protein and spores of Bacillus subtilis taken as a representative of highly resistant micro-organisms. The treatment of these materials was performed by means of atmospheric pressure dielectric barrier discharge (DBD) sustained in open air at constant conditions. All tested polymers, BSA and spores, were readily etched by DBD plasma. However, the measured etching rates were found to be dependent on the chemical structure of treated materials, namely on the presence of oxygen in the structure of polymers.

  7. Advanced oxidation protein products in plasma: stability during storage and correlation with other clinical characteristics.

    Science.gov (United States)

    Matteucci, E; Biasci, E; Giampietro, O

    2001-12-01

    Proteins are susceptible to free radical damage. We measured advanced oxidation protein products (AOPP) in the plasma of 56 hospitalised patients. Concentrations of AOPP were expressed as chloramine-T equivalents by measuring absorbance in acidic conditions at 340 nm in the presence of potassium iodide. We also determined erythrocyte sedimentation rate (ESR), circulating urea, creatinine, glucose, uric acid, electrolytes, lipids, total proteins and fractions and fibrinogen. Twenty-four samples were processed both immediately and after 7, 15, 30, 90, 180 and 438 days of storage at both at -20 degrees C and -80 degrees C (aliquots were frozen and thawed only once) to evaluate AOPP stability. The remaining 32 samples were also processed for thiobarbituric-acid-reactive substances (TBARS). Mean AOPP concentration in all 56 patients was 48.3+/-37.2 microM. Mean basal concentration of AOPP in the 24 plasma samples (55.0+/-47.1 microM) showed no significant change at each intermediate determination, yet significantly increased after 438 days of storage both at -80 degrees C (96.6+/-83.2, p<0.01) and, markedly, at -20 degrees C (171.3+/-94.6, p<0.001). TBARS concentration was 1.59+/-0.65 micromol/l. Multiple regression analysis evidenced that AOPP concentration was positively correlated (multiple r=0.62, p<0.001) with serum urea and triglycerides, but negatively correlated with patient age (indeed, serum albumin and total proteins decreased with increasing age, r=0.3, p<0.05). TBARS concentration was associated with ESR and serum glucose (multiple r=0.73, p<0.001), yet positively with AOPP (r=0.39, simple p<0.05). We conclude that AOPP remain stable during sample storage both at -20 degrees C and -80 degrees C for 6 months. Renal failure and hypertriglyceridemia probably enhance the in vivo process of AOPP formation. Oxidative damage as measured by TBARS may be increased because of exposure to hyperglycemia causing nonenzymatic glycation of plasma proteins.

  8. PARTIAL PURIFICATION AND IMMUNO-BIOCHEMICAL CHARACTERISATION OF FERTILITY ASSOCIATED PROTEIN OF KARAN FRIES BULL SEMINAL PLASMA

    Directory of Open Access Journals (Sweden)

    Brajesh Raman

    2014-06-01

    Full Text Available The objective of the present study was detection, isolation, partial purification and immunobiochemical characterization of fertility associated protein in the seminal plasma of high prolific Karan fries bull. Seminal plasma of Karan Fries bull was partially purified by gel filtration chromatography and analyzed by 10% SDS-PAGE for their polypeptide profile. PAGE analysis revealed major band of 55 kDa, and 26 kDa. Hyperimmune serum was raised in rabbit against crude seminal plasma protein. Single precipitin line was observed in DID test when each of the partially purified 26 kDa and 55 kDa proteins were reacted with hyperimmune serum. These proteins were also found to be immunoreactive against hyperimmune serum in Western blot technique.

  9. Heme protein-induced tubular cytoresistance: expression at the plasma membrane level.

    Science.gov (United States)

    Zager, R A

    1995-05-01

    Following experimental rhabdomyolysis, animals become resistant to heme protein-induced acute renal failure (ARF). The goals of this study were to: (a) ascertain whether this resistance, previously documented only in vivo, is expressed directly at the proximal tubular cell level; (b) determine whether heme proteinuria (vs. other consequences of rhabdomyolysis) is its trigger; and (c) ascertain some of its subcellular determinants. Rats were injected with a borderline toxic dose of glycerol and 24 hours later proximal tubular segments (PTS) were isolated for study. Their vulnerability to diverse forms of injury (FeSO4-induced oxidant stress, hypoxia, Ca2+ ionophore, cytochalasin D, PLA2) was compared to that found in normal PTS. Post-glycerol PTS manifested significant resistance to each insult (decreased lactate dehydrogenase +/- N-acetyl-beta-D-glucosaminidase release). Protection against FeSO4 was virtually complete and it was associated with a 50% decrease in membrane lipid peroxidation. No decrease in hydroxyl radical generation was noted during the FeSO4 challenge (salicylate trap assessment), suggesting a primary increase in membrane resistance to attack. That PLA2 addition caused less deacylation, plasma membrane enzyme (alanine aminopeptidase) release, and LDH leakage from post-glycerol versus normal tubules supported this hypothesis. To test whether cytoresistance was specifically triggered by heme proteins (vs. being a non-specific filtered protein effect, or a result of endotoxin cascade activation), rats were injected with purified myoglobin, non-heme containing filterable proteins, or endotoxin. Only myoglobin induced cytoresistance. In vivo heme oxygenase inhibition (tin-protoporphyrin) did not block the emergence of cytoresistance and it was expressed despite Na,K-ATPase inhibition (ouabain) or cytoskeletal disruption (cytochalasin D). In vivo heat shock failed to protect. In conclusion, (1) rhabdomyolysis induces broad based proximal tubular

  10. Plasma protein-binding parameters of prednisolone in immune disease patients receiving long-term prednisone therapy.

    Science.gov (United States)

    Wagner, J G; Wexler, D; Ağabeyoğlu, I T; Bergstrom, R F; Sakmar, E; Kay, D R

    1981-04-01

    Prednisone and prednisolone bind in plasma to albumin and transcortin. In am attempt to determine whether prednisone side effects and/or type of disease correlated with prednisolone plasma protein binding, multiple plasma samples from 17 patients (three asthma, eight SLE, three RA, two PSS, one PAN) receiving long-term prednisone therapy were monitored during an interval between two prednisone doses. Prednisolone plasma protein binding was nonlinear and exhibited large intrapatient and interpatient variability. For the group, mean association constants of the prednisolone-albumin complex and the prednisolone-transcortin complex were 2.3 X 10(3) M-1 and 2.9 X 10(7) M-1, with coefficients of variation of 82% and 127%, respectively. SLE patients tended to have lower mean prednisolone association constants for albumin and transcortin than did other patients. The presence of corticosteroid side effects did not correlate with prednisolone plasma protein-binding parameters. The wide range of prednisolone free fraction noted in plasma from patients who achieved comparable total prednisolone plasma concentrations implies that administration of a uniform prednisone dose will not lead to a predictable clinical response.

  11. Informing the Human Plasma Protein Binding of Environmental Chemicals by Machine Learning in the Pharmaceutical Space: Applicability Domain and Limits of Predictability

    Science.gov (United States)

    The free fraction of a xenobiotic in plasma (Fub) is an important determinant of chemical adsorption, distribution, metabolism, elimination, and toxicity, yet experimental plasma protein binding data is scarce for environmentally relevant chemicals. The presented work explores th...

  12. Informing the Human Plasma Protein Binding of Environmental Chemicals by Machine Learning in the Pharmaceutical Space: Applicability Domain and Limits of Predictability

    Science.gov (United States)

    The free fraction of a xenobiotic in plasma (Fub) is an important determinant of chemical adsorption, distribution, metabolism, elimination, and toxicity, yet experimental plasma protein binding data is scarce for environmentally relevant chemicals. The presented work explores th...

  13. Platelet adhesion and plasma protein adsorption control of collagen surfaces by He{sup +} ion implantation

    Energy Technology Data Exchange (ETDEWEB)

    Kurotobi, K. E-mail: kurotobi@postman.riken.go.jp; Suzuki, Y.; Nakajima, H.; Suzuki, H.; Iwaki, M

    2003-05-01

    He{sup +} ion implanted collagen-coated tubes with a fluence of 1 x 10{sup 14} ions/cm{sup 2} were exhibited antithrombogenicity. To investigate the mechanisms of antithrombogenicity of these samples, plasma protein adsorption assay and platelet adhesion experiments were performed. The adsorption of fibrinogen (Fg) and von Willebrand factor (vWf) was minimum on the He{sup +} ion implanted collagen with a fluence of 1 x 10{sup 14} ions/cm{sup 2}. Platelet adhesion (using platelet rich plasma) was inhibited on the He{sup +} ion implanted collagen with a fluence of 1 x 10{sup 14} ions/cm{sup 2} and was accelerated on the untreated collagen and ion implanted collagen with fluences of 1 x 10{sup 13}, 1 x 10{sup 15} and 1 x 10{sup 16} ions/cm{sup 2}. Platelet activation with washed platelets was observed on untreated collagen and He{sup +} ion implanted collagen with a fluence of 1 x 10{sup 14} ions/cm{sup 2} and was inhibited with fluences of 1 x 10{sup 13}, 1 x 10{sup 15} and 1 x 10{sup 16} ions/cm{sup 2}. Generally, platelets can react with a specific ligand inside the collagen (GFOGER sequence). The results of platelets adhesion experiments using washed platelets indicated that there were no ligands such as GFOGER on the He{sup +} ion implanted collagen over a fluence of 1 x 10{sup 13} ions/cm{sup 2}. On the 1 x 10{sup 14} ions/cm{sup 2} implanted collagen, no platelet activation was observed due to the influence of plasma proteins. >From the above, it is concluded that the decrease of adsorbed Fg and vWf caused the antithrombogenicity of He{sup +} ion implanted collagen with a fluence of 1 x 10{sup 14} ions/cm{sup 2} and that plasma protein adsorption took an important role repairing the graft surface.

  14. Effects of storage time on total protein and globulin concentrations in bovine fresh frozen plasma obtained for transfusion.

    Science.gov (United States)

    Proverbio, D; Spada, E; Baggiani, L; Bagnagatti De Giorgi, G; Roggero, N; Belloli, A; Pravettoni, D; Perego, R

    2015-01-01

    To evaluate the effects of storage conditions on total protein (TP) and globulin fractions in fresh frozen bovine plasma units prepared and stored for transfusion, TP and globulin fractions were evaluated in fresh plasma and at 1 month and 6 and 12 months after blood collection in plasma stored at -20°C. Significant differences in concentrations were found in the median concentration of total protein (P=0.0336), between 0 months and 1 month (P=0.0108), 0 and 6 months (P=0.0023), and 0 and 12 months (P=0.0027), in mean concentration (g/dL) of albumin (P=0.0394), between 0 months and 1 month (P=0.0131), 0 and 6 months (P=0.0035), and 0 and 12 months (P=0.0038), and beta-2 fraction (P=0.0401), between 0 and 6 months (P=0.0401) and 0 and 12 months (P=0.0230). This study suggests that total gamma globulin concentration in bovine frozen plasma is stable for 12 months at -20°C. Total protein, ALB, and beta-2 fraction have significantly different concentrations (g/dL) when compared to prestorage. This study has shown IgG protein fraction stability in bovine fresh frozen plasma collected for transfusion; therefore, bovine fresh frozen plasma seems to be suitable for the treatment of hypogammaglobulinemia (failure of passive transfer) in calves when stored for 12 months at -20°C.

  15. High precision quantification of human plasma proteins using the automated SISCAPA Immuno-MS workflow.

    Science.gov (United States)

    Razavi, Morteza; Leigh Anderson, N; Pope, Matthew E; Yip, Richard; Pearson, Terry W

    2016-09-25

    Efficient robotic workflows for trypsin digestion of human plasma and subsequent antibody-mediated peptide enrichment (the SISCAPA method) were developed with the goal of improving assay precision and throughput for multiplexed protein biomarker quantification. First, an 'addition only' tryptic digestion protocol was simplified from classical methods, eliminating the need for sample cleanup, while improving reproducibility, scalability and cost. Second, methods were developed to allow multiplexed enrichment and quantification of peptide surrogates of protein biomarkers representing a very broad range of concentrations and widely different molecular masses in human plasma. The total workflow coefficients of variation (including the 3 sequential steps of digestion, SISCAPA peptide enrichment and mass spectrometric analysis) for 5 proteotypic peptides measured in 6 replicates of each of 6 different samples repeated over 6 days averaged 3.4% within-run and 4.3% across all runs. An experiment to identify sources of variation in the workflow demonstrated that MRM measurement and tryptic digestion steps each had average CVs of ∼2.7%. Because of the high purity of the peptide analytes enriched by antibody capture, the liquid chromatography step is minimized and in some cases eliminated altogether, enabling throughput levels consistent with requirements of large biomarker and clinical studies.

  16. Decreased dietary protein or energy intake and plasma growth hormone levels of the pregnant pig, its fetuses and developing progeny.

    Science.gov (United States)

    Atinmo, T; Baldijrao, C; Pond, W G; Barnes, R H

    1976-07-01

    The effects of low protein diets on plasma growth hormone were studied in pregnant pigs, fetuses and the developing progeny. Pregnant pigs were fed 18%, 3% or 0.5% protein diet throughout the gestation period. At 10, 13 and 15 week of gestation, fetuses were removed from the uterus after the dam had been bled to death. Plasma samples were used for growth hormone determinations. In a second experiment, 2-day old pigs from another set of pregnant pigs fed the diet containing 18%, 3% or 0.5% protein during gestation were cross-fostered to control nursing dams and weaned at 4 weeks of age to a standard diet. Plasma obtained at regular intervals was used for growth hormone determination. Plasma growth hormone was significantly higher in dams fed 0.5% protein after week 13 of gestation. High growth hormone (ten times the dam GH level) was observed in all fetuses irrespective of maternal dietary manipulation. Offspring of severely protein deprived pits (0.5% protein) had significantly elevated growth hormone levels up to 12 weeks of age in spite of cross fostering to a control dam after birth. The data suggest that there is little or no effect of maternal protein restriction on fetal growth hormone levels but the persistent high growth hormone levels in the progeny of severely malnourished pigs indicate a possible impairment of the production, release or catabolism of growth hormone and/or its releasing factor.

  17. Sperm nuclear DNA fragmentation rate is associated with differential protein expression and enriched functions in human seminal plasma.

    Science.gov (United States)

    Intasqui, Paula; Camargo, Mariana; Del Giudice, Paula T; Spaine, Deborah M; Carvalho, Valdemir M; Cardozo, Karina H M; Zylbersztejn, Daniel S; Bertolla, Ricardo P

    2013-10-01

    To analyse the proteomic profile of seminal plasma with the aim of identifying the proteins and post-genomic pathways associated with sperm DNA fragmentation. A cross-sectional study including 89 subjects from a human reproduction service was carried out. All semen samples were assessed for sperm DNA fragmentation using a comet assay. Results from 60 sperm were analysed using Komet 6.0.1 software and the 'Olive tail moment' variable was used to stratify these into low and high sperm DNA fragmentation groups. Seminal plasma proteins from the two groups were pooled and used for proteomic analysis. Quantitative data were used for functional enrichment studies. Seventy-two proteins were identified or quantified in seminal plasma. Of these, nine were differentially expressed in the low group and 21 in the high group. Forty-two proteins were conserved between these groups. Functional enrichment analysis indicated that sperm DNA fragmentation was related to functions such as lipoprotein particle remodelling and regulation, fatty acid binding and immune response. Proteins found exclusively in the low group may be involved in correcting spermatogenesis and/or improving sperm function. Proteins in the high group were associated with increased innate immune response, sperm motility and/or maturation and inhibition of mitochondrial apoptosis. Protein expression and post-genomic pathways of seminal plasma differ according to the rate of sperm DNA integrity. © 2013 The Authors. BJU International © 2013 BJU International.

  18. Ram seminal plasma proteins contribute to sperm capacitation and modulate sperm-zona pellucida interaction.

    Science.gov (United States)

    Luna, C; Colás, C; Casao, A; Serrano, E; Domingo, J; Pérez-Pé, R; Cebrián-Pérez, J A; Muiño-Blanco, T

    2015-03-01

    Incubation of ram spermatozoa in capacitating conditions with cAMP-elevating agents promotes a progressive time-dependent increase in the capacitated sperm subpopulation. In this study, the fertilizing capacity of ram spermatozoa (ability to bind to the zona pellucida, ZBA rate) capacitated in these conditions was determined. The results showed an increase (P ram spermatozoa. Likewise, the presence of two seminal plasma (SP) proteins able to protect sperm against cold shock (RSVP14 and RSVP20) was evidenced in both SP and the ram sperm surface, and their influence in the fertilizing ability of spermatozoa capacitated in basal medium or with cAMP-elevating agents was determined. The results verified that RSVP14 and RSVP20 act as decapacitating factors given that their addition to SP-free sperm samples previously to capacitation maintained high proportions of the noncapacitated sperm pattern with no increase in protein tyrosine phosphorylation. However, the obtained ZBA rate in the high-cAMP-containing samples was increased in the presence of RSVP20 (P < 0.05). These findings would indicate that the stimulating effect exerted by this protein on the sperm-oocyte binding occurs downstream from the cAMP generation and that the mechanisms by which RSVP20 promotes the zona pellucida binding might be independent of protein tyrosine phosphorylation.

  19. Binding of plasma proteins to titanium dioxide nanotubes with different diameters.

    Science.gov (United States)

    Kulkarni, Mukta; Flašker, Ajda; Lokar, Maruša; Mrak-Poljšak, Katjuša; Mazare, Anca; Artenjak, Andrej; Čučnik, Saša; Kralj, Slavko; Velikonja, Aljaž; Schmuki, Patrik; Kralj-Iglič, Veronika; Sodin-Semrl, Snezna; Iglič, Aleš

    2015-01-01

    Titanium and titanium alloys are considered to be one of the most applicable materials in medical devices because of their suitable properties, most importantly high corrosion resistance and the specific combination of strength with biocompatibility. In order to improve the biocompatibility of titanium surfaces, the current report initially focuses on specifying the topography of titanium dioxide (TiO2) nanotubes (NTs) by electrochemical anodization. The zeta potential (ζ-potential) of NTs showed a negative value and confirmed the agreement between the measured and theoretically predicted dependence of ζ-potential on salt concentration, whereby the absolute value of ζ-potential diminished with increasing salt concentrations. We investigated binding of various plasma proteins with different sizes and charges using the bicinchoninic acid assay and immunofluorescence microscopy. Results showed effective and comparatively higher protein binding to NTs with 100 nm diameters (compared to 50 or 15 nm). We also showed a dose-dependent effect of serum amyloid A protein binding to NTs. These results and theoretical calculations of total available surface area for binding of proteins indicate that the largest surface area (also considering the NT lengths) is available for 100 nm NTs, with decreasing surface area for 50 and 15 nm NTs. These current investigations will have an impact on increasing the binding ability of biomedical devices in the body leading to increased durability of biomedical devices.

  20. Preferential transfer of certain plasma membrane proteins onto T and B cells by trogocytosis.

    Directory of Open Access Journals (Sweden)

    Sandrine Daubeuf

    Full Text Available T and B cells capture antigens via membrane fragments of antigen presenting cells (APC in a process termed trogocytosis. Whether (and how a preferential transfer of some APC components occurs during trogocytosis is still largely unknown. We analyzed the transfer onto murine T and B cells of a large panel of fluorescent proteins with different intra-cellular localizations in the APC or various types of anchors in the plasma membrane (PM. Only the latter were transferred by trogocytosis, albeit with different efficiencies. Unexpectedly, proteins anchored to the PM's cytoplasmic face, or recruited to it via interaction with phosphinositides, were more efficiently transferred than those facing the outside of the cell. For proteins spanning the PM's whole width, transfer efficiency was found to vary quite substantially, with tetraspanins, CD4 and FcRgamma found among the most efficiently transferred proteins. We exploited our findings to set immunodiagnostic assays based on the capture of preferentially transferred components onto T or B cells. The preferential transfer documented here should prove useful in deciphering the cellular structures involved in trogocytosis.

  1. Vacuolar Sorting Receptor (VSR) Proteins Reach the Plasma Membrane in Germinating Pollen Tubes

    Institute of Scientific and Technical Information of China (English)

    Hao Wang; Xiao-Hong Zhuang; Stefan Hillmer; David G. Robinson; Li-Wen Jiang

    2011-01-01

    Vacuolar sorting receptors (VSRs) are type I integral membrane proteins that mediate the vacuolar transport of soluble cargo proteins via prevacuolar compartments (PVCs) in plants.Confocal immunofluorescent and immunogold Electron Microscope (EM) studies have localized VSRs to PVCs or multivesicular bodies (MVBs) and trans-Golgi network (TGN) in various plant cell types,including suspension culture cells,root cells,developing and germinating seeds.Here,we provide evidence that VSRs reach plasma membrane (PM) in growing pollen tubes.Both immunofluorescent and immunogold EM studies with specific VSR antibodies show that,in addition to the previously demonstrated PVC/MVB localization,VSRs also localize to PM in lily and tobacco pollen tubes prepared from chemical fixation or high-pressure freezing/frozen substitution.Such a PM localization suggests an additional role of VSR proteins in mediating protein transport to PM and endocytosis in growing pollen tubes.Using a high-speed Spinning Disc Confocal Microscope,the possible fusion between VSR-positive PVC organelles and the PM was also observed in living tobacco pollen tubes transiently expressing the PVC reporter GFP-VSR.In contrast,the lack of a prominent PM localization of GFP-VSR in living pollen tubes may be due to the highly dynamic situation of vesicular transport in this fast-growing cell type.

  2. Aldehyde dehydrogenase 2 activation in aged heart improves the autophagy by reducing the carbonyl modification on SIRT1.

    Science.gov (United States)

    Wu, Bing; Yu, Lu; Wang, Yishi; Wang, Hongtao; Li, Chen; Yin, Yue; Yang, Jingrun; Wang, Zhifa; Zheng, Qiangsun; Ma, Heng

    2016-01-19

    Cardiac aging is characterized by accumulation of damaged proteins and decline of autophagic efficiency. Here, by forestalling SIRT1 carbonylated inactivation in aged heart, we determined the benefits of activation of aldehyde dehydrogenase 2 (ALDH2) on the autophagy. In this study, the ALDH2 KO mice progressively developed age-related heart dysfunction and showed reduction in the life span, which strongly suggests that ALDH2 ablation leads to cardiac aging. What's more, aged hearts displayed a significant decrease ALDH2 activity, resulting in accumulation of 4-HNE-protein adducts and protein carbonyls, impairment in the autophagy flux, and, consequently, deteriorated cardiac function after starvation. Sustained Alda-1 (selective ALDH2 activator) treatment increased cardiac ALDH2 activity and abrogated these effects. Using SIRT1 deficient heterozygous (Sirt1+/-) mice, we found that SIRT1 was necessary for ALDH2 activation-induced autophagy. We further demonstrated that ALDH2 activation attenuated SIRT1 carbonylation and improved SIRT1 activity, thereby increasing the deacetylation of nuclear LC3 and FoxO1. Sequentially, ALDH2 enhanced SIRT1 regulates LC3-Atg7 interaction and FoxO1 increased Rab7 expression, which were both necessary and sufficient for restoring autophagy flux. These results highlight that both accumulation of proteotoxic carbonyl stress linkage with autophagy decline contribute to heart senescence. ALDH2 activation is adequate to improve the autophagy flux by reducing the carbonyl modification on SIRT1, which in turn plays an important role in maintaining cardiac health during aging.

  3. Protein secretion is required for pregnancy-associated plasma protein-A to promote lung cancer growth in vivo.

    Directory of Open Access Journals (Sweden)

    Hong Pan

    Full Text Available Pregnancy-associated plasma protein-A (PAPPA has been reported to regulate the activity of insulin-like growth factor (IGF signal pathway through proteolytic degradation of IGF binding proteins (IGFBPs thereby increasing the local concentration of free IGFs available to receptors. In this study we found that PAPPA is secreted from two out of seven lung cancer cell lines examined. None of immortalized normal bronchial epithelial cells (HBE tested secrets PAPPA. There is no correlation between expression level and secretion of PAPPA in these cells. A cell line over-expressing PAPPA accompanied with secretion shows no notable changes in proliferation under cell culture conditions in vitro, but displays significantly augmentation of tumor growth in vivo in a xenograft model. In contrast, a cell line over-expressing PAPPA without secretion exhibits reduction of tumor growth both in vitro and in vivo. Down-regulation of PAPPA expression and secretion by RNAi knockdown decreases tumor growth after implanted in vivo. The tumor promoting activity of PAPPA appears to be mediated mainly through augmentation of the IGF signaling pathway as indicated by notable increases in downstream Akt kinase phosphorylation in tumor samples. Our results indicate that PAPPA secretion may play an important role in lung cancer growth and progression.

  4. Cholesterol and F-actin are required for clustering of recycling synaptic vesicle proteins in the presynaptic plasma membrane.

    Science.gov (United States)

    Dason, Jeffrey S; Smith, Alex J; Marin, Leo; Charlton, Milton P

    2014-02-15

    Synaptic vesicles (SVs) and their proteins must be recycled for sustained synaptic transmission. We tested the hypothesis that SV cholesterol is required for proper sorting of SV proteins during recycling in live presynaptic terminals. We used the reversible block of endocytosis in the Drosophila temperature-sensitive dynamin mutant shibire-ts1 to trap exocytosed SV proteins, and then examined the effect of experimental treatments on the distribution of these proteins within the presynaptic plasma membrane by confocal microscopy. SV proteins synaptotagmin, vglut and csp were clustered following SV trapping in control experiments but dispersed in samples treated with the cholesterol chelator methyl-β-cyclodextrin to extract SV cholesterol. There was accumulation of phosphatidylinositol (4,5)-bisphosphate (PIP2) in presynaptic terminals following SV trapping and this was reduced following SV cholesterol extraction. Reduced PIP2 accumulation was associated with disrupted accumulation of actin in presynaptic terminals. Similar to vesicular cholesterol extraction, disruption of actin by latrunculin A after SV proteins had been trapped on the plasma membrane resulted in the dispersal of SV proteins and prevented recovery of synaptic transmission due to impaired endocytosis following relief of the endocytic block. Our results demonstrate that vesicular cholesterol is required for aggregation of exocytosed SV proteins in the presynaptic plasma membrane and are consistent with a mechanism involving regulation of PIP2 accumulation and local actin polymerization by cholesterol. Thus, alteration of membrane or SV lipids may affect the ability of synapses to undergo sustained synaptic transmission by compromising the recycling of SV proteins.

  5. Acute inhalation toxicity of carbonyl sulfide

    Energy Technology Data Exchange (ETDEWEB)

    Benson, J.M.; Hahn, F.F.; Barr, E.B. [and others

    1995-12-01

    Carbonyl sulfide (COS), a colorless gas, is a side product of industrial procedures sure as coal hydrogenation and gasification. It is structurally related to and is a metabolite of carbon disulfide. COS is metabolized in the body by carbonic anhydrase to hydrogen sulfide (H{sub 2}S), which is thought to be responsible for COS toxicity. No threshold limit value for COS has been established. Results of these studies indicate COS (with an LC{sub 50} of 590 ppm) is slightly less acutely toxic than H{sub 2}S (LC{sub 50} of 440 ppm).

  6. Leukocyte-Reduced Platelet-Rich Plasma Alters Protein Expression of Adipose Tissue-Derived Mesenchymal Stem Cells.

    Science.gov (United States)

    Loibl, Markus; Lang, Siegmund; Hanke, Alexander; Herrmann, Marietta; Huber, Michaela; Brockhoff, Gero; Klein, Silvan; Nerlich, Michael; Angele, Peter; Prantl, Lukas; Gehmert, Sebastian

    2016-08-01

    Application of platelet-rich plasma and stem cells has become important in regenerative medicine. Recent literature supports the use of platelet-rich plasma as a cell culture media supplement to stimulate proliferation of adipose tissue-derived mesenchymal stem cells. The underlying mechanism of proliferation stimulation by platelet-rich plasma has not been investigated so far. Adipose tissue-derived mesenchymal stem cells were cultured in α-minimal essential medium supplemented with platelet-rich plasma or fetal calf serum. Cell proliferation was assessed with cell cycle kinetics using flow cytometric analyses after 48 hours. Differences in proteome expression of the adipose tissue-derived mesenchymal stem cells were analyzed using a reverse-phase protein array to quantify 214 proteins. Complementary Ingenuity Pathways Analysis and gene set enrichment analysis were performed using protein data, and confirmed by Western blot analysis. A higher percentage of adipose tissue-derived mesenchymal stem cells in the S phase in the presence of platelet-rich plasma advocates the proliferation stimulation. Ingenuity Pathways Analysis and gene set enrichment analysis confirm the involvement of the selected proteins in the process of cell growth and proliferation. Ingenuity Pathways Analysis revealed a participation in the top-ranked canonical pathways PI3K/AKT, PTEN, ILK, and IGF-1. Gene set enrichment analysis identified the authors' protein set as being part of significantly regulated protein sets with the focus on cell cycle, metabolism, and the Kyoto Encyclopedia of Genes and Genomes transforming growth factor-β signaling pathway. The present study provides evidence that platelet-rich plasma stimulates proliferation and induces a unique change in the proteomic profile of adipose tissue-derived mesenchymal stem cells. The interpretation of altered expression of regulatory proteins represents a step forward toward achieving good manufacturing practice-compliant criteria

  7. Pollution survey of carbonyl compounds in train air

    Institute of Scientific and Technical Information of China (English)

    LU Hao; ZHU Lizhong

    2007-01-01

    The train iS an important vehicle in China,but its air quality has important impacts on passengers' health.In this work,pollution from carbonyl compounds was measured in the air of six trains.The obiectives of this work were to investigate carbonyl compound levels in selected air from trains,identify their emission sources,and assess the intake of carbonyl compounds for passengers.The methods for sampling and analyzing 10 carbonyl compounds such as formaldehyde,acetaldehyde,acrolein,acetone,propionalde hyde,crotonaldehyde,butyraldehyde,benzaldehyde,cyclo hexanone,and valeraldehyde in indoor air were proposed with the sampling efficiency,recovery,and detection limit being 92%-100%,91%-104%,and 0.26-0.82 ng/m3,respec tively.It was indicated that the total concentrations of carbonyl compounds were 0.159-0.2828 mg/m3 with the average concentration of 0.2330 mg/m3.The average concen trations of formaldehyde.acetaldehyde and acetone were 0.0922,0.0499,and 0.0580 mg/m3,accounting for 42.6%,21.4%.and 24.9% of the total concentrations of carbonyl compounds,respectively.The carbonyl compounds probably came from woodwork and cigarette smoking.The intake of carbonyl compounds for the passengers was approximately 0.043-0.076 mg/h.The carbonyl compounds in train air could be harmful to human health.

  8. Plasma Levels of Monocyte Chemotactic Protein 3 and Beta-Nerve Growth Factor Increase with Amnestic Mild Cognitive Impairment

    Institute of Scientific and Technical Information of China (English)

    Kang Soo Lee; Ji Hyung Chung; Kyung Hye Lee; Min-Jeong Shin; Byoung Hoon Oh; Soo Hyung Lee; Chang Hyung Hong

    2009-01-01

    A number of studies have investigated peripheral inflammatory indices, including plasma cytokines and related molecules according to subtypes of dementia, but not in mild cognitive impairment (MCI). In this study, we used multiplex cytokine assay to assess the plasma levels of 22 cytokines in patients with MCI subtyped as amnestic and non-amnestic, according to cognitive features. When comparing the levels of plasma growth factors, chemokines and cytokines, plasma levels of monocyte chemotactic protein 3 (MCP-3), and beta-nerve growth factor (β-NGF) in these two groups, they were found to be significantly higher in amnestic MCI patients than in non-amnestic MCI patients, after adjusting for age and gender. This suggests that plasma MCP-3 and β-NGF may be useful in differentiating subtypes of MCI. Cellular & Molecular Immunology.

  9. Effect of hydrogen atoms on the structures of trinuclear metal carbonyl clusters: trinuclear manganese carbonyl hydrides.

    Science.gov (United States)

    Liu, Xian-mei; Wang, Chao-yang; Li, Qian-shu; Xie, Yaoming; King, R Bruce; Schaefer, Henry F

    2009-05-18

    The structures of the trinuclear manganese carbonyl hydrides H(3)Mn(3)(CO)(n) (n = 12, 11, 10, 9) have been investigated by density functional theory (DFT). Optimization of H(3)Mn(3)(CO)(12) gives the experimentally known structure in which all carbonyl groups are terminal and each edge of a central Mn(3) equilateral triangle is bridged by a single hydrogen atom. This structure establishes the canonical distance 3.11 A for the Mn-Mn single bond satisfying the 18-electron rule. The central triangular (mu-H)(3)Mn(3) unit is retained in the lowest energy structure of H(3)Mn(3)(CO)(11), which may thus be derived from the H(3)Mn(3)(CO)(12) structure by removal of a carbonyl group with concurrent conversion of one of the remaining carbonyl groups into a semibridging carbonyl group to fill the resulting hole. The potential energy surface of H(3)Mn(3)(CO)(10) is relatively complicated with six singlet and five triplet structures. One of the lower energy H(3)Mn(3)(CO)(10) structures has one of the hydrogen atoms bridging the entire Mn(3) triangle and the other two hydrogen atoms bridging Mn-Mn edges. This H(3)Mn(3)(CO)(10) structure achieves the favored 18-electron configuration with a very short MnMn triple bond of 2.36 A. The other low energy H(3)Mn(3)(CO)(10) structure retains the (mu-H)(3)Mn(3) core of H(3)Mn(3)(CO)(12) but has a unique six-electron donor eta(2)-mu(3) carbonyl group bridging the entire Mn(3) triangle similar to the unique carbonyl group in the known compound Cp(3)Nb(3)(CO)(6)(eta(2)-mu(3)-CO). For H(3)Mn(3)(CO)(9) a structure with a central (mu(3)-H)(2)Mn(3) trigonal bipyramid lies >20 kcal/mol below any of the other structures. Triplet structures were found for the unsaturated H(3)Mn(3)(CO)(n) (n = 11, 10, 9) systems but at significantly higher energies than the lowest lying singlet structures.

  10. Effects of Chinese herbal medicine on plasma glucose, protein and energy metabolism in sheep

    Institute of Scientific and Technical Information of China (English)

    Xi Liang; Kyota Yamazaki; Mohammad Kamruzzaman; Xue Bi; Arvinda Panthee; Hiroaki Sano

    2014-01-01

    Background:The use of antibiotics in animal diets is facing negative feedback due to the hidden danger of drug residues to human health. Traditional Chinese herbal medicine has been used to replace antibiotics in the past two decades and played an increasingly important role in livestock production. The present study was carried out to assess the feeding effects of a traditional nourishing Chinese herbal medicine mixture on kinetics of plasma glucose, protein and energy metabolism in sheep. Ruminal fermentation characteristics were also determined. Methods:Four sheep were fed on either mixed hay (MH-diet) or MH-diet supplemented with 2%of Chinese herbal medicine (mixture of Astragalus root, Angelica root and Atractylodes rhizome;CHM-diet) over two 35-day periods using a crossover design. The turnover rate of plasma glucose was measured with an isotope dilution method using [U-13C]glucose. The rates of plasma leucine turnover and leucine oxidation, whole body protein synthesis (WBPS) and metabolic heat production were measured using the [1-13C]leucine dilution and open circuit calorimetry. Results:Body weight gain of sheep was higher (P=0.03) for CHM-diet than for MH-diet. Rumen pH was lower (P=0.02), concentration of rumen total volatile fatty acid tended to be higher (P=0.05) and acetate was higher (P=0.04) for CHM-diet than for MH-diet. Turnover rates of plasma glucose and leucine did not differ between diets. Oxidation rate of leucine tended to be higher (P=0.06) for CHM-diet than for MH-diet, but the WBPS did not differ between diets. Metabolic heat production tended to be greater (P=0.05) for CHM-diet than for MH-diet. Conclusions:The sheep fed on CHM-diet had a higher body weight gain and showed positive impacts on rumen fermentation and energy metabolism without resulting in any adverse response. Therefore, these results suggested that the Chinese herbal medicine mixture should be considered as a potential feed additive for sheep.

  11. Hydration and hydrogen bonding of carbonyls in dimyristoyl-phosphatidylcholine bilayer.

    Science.gov (United States)

    Volkov, Victor V; Nuti, Francesca; Takaoka, Yuji; Chelli, Riccardo; Papini, Anna Maria; Righini, Roberto

    2006-07-26

    We combine two-color ultrafast infrared spectroscopy and molecular dynamics simulation to investigate the hydration of carbonyl moieties in a dimyristoyl-phosphatidylcholine bilayer. Excitation with femtosecond infrared pulses of the OD stretching mode of heavy water produces a time dependent change of the absorption band of the phospholipid carbonyl groups. This intermolecular vibrational coupling affects the entire C=O band, thus suggesting that the optical inhomogeneity of the infrared response of carbonyl in phospholipid membranes cannot be attributed to the variance in hydration. Both the experimental and the theoretical results demonstrate that sn-1 carbonyl has a higher propensity to form hydrogen bonds with water in comparison to sn-2. The time-resolved experiment allows following the evolution of the system from a nonequilibrium localization of energy in the OD stretching mode to a thermally equilibrated condition and provides the characteristic time constants of the process. The approach opens a new opportunity for investigation of intermolecular structural relations in complex systems, like membranes, polymers, proteins, and glasses.

  12. Source of dietary protein influences kinetics of plasma gut regulatory peptide concentration in response to feeding in preruminant calves.

    Science.gov (United States)

    le Huërou-Luron, I; Gestin, M; Le Dréan, G; Romé, V; Bernard, C; Chayvialle, J A; Guilloteau, P

    1998-03-01

    The kinetics of the peripheral plasma concentrations of eight gut regulatory peptides were examined in response to feeding in preruminant calves. Two experiments were carried out in animals fed milk substitutes either based on milk protein (control diet) or in which casein had been replaced by hydrolyzed fish (fish diet in experiment 1) or whey (whey diet in experiment 2) protein concentrate. In contrast to the control diet, the latter two did not coagulate within the abomasum. No variation was observed in plasma concentrations of gut regulatory peptides during 1-1.4 hr before the morning meal regardless of the nature of the dietary protein. With the control diet, the meal was followed by an increase in cholecystokinin, gastrin and gastric inhibitory polypeptide and a fall in secretin, vasoactive intestinal polypeptide and motilin, whereas no significant change was observed for somatostatin and pancreatic polypeptide. The replacement of casein by protein substitutes did not greatly modify the pattern of plasma responses to feeding, but the prefeeding and postfeeding levels were highly affected. We conclude that the most important characteristic influencing plasma gut peptide concentrations is the ability of dietary protein to clot in the abomasum, consequently determining the pattern of gastric emptying, and that variations appear depending on the origin of protein substitutes in relation to the duodenal content and mainly to the digesta pH.

  13. EFFECT OF PROTEINE CONTENT IN BOAR SEMINAL PLASMA ON THE SPERM MOTILITY IN DILUTED SEMEN STORED FOR 3 DAYS

    Directory of Open Access Journals (Sweden)

    Jelena Apić

    2015-02-01

    Full Text Available Recently, it was frequently demonstrated that fertility of sows after artificially inseminated is lower than after mating. This is associated with a reduced fertilization capacity of overdiluted insemination doses. The aim of this study was to investigate the sperm motility in the semen samples, forming from the ejaculates with high or low protein content, stored in vitro on 17oC for 3 days. Progressive motility was significantly higher (p<0.01 in the ejaculates with high, compared to the ejaculates with low protein content (82% vs. 76%. After 3 days of storage, in the1:4 dilution proportion, the average progressive motility was significantly (p<0.01 decreased in relation to this value in native semen from the boars with high (82% to64%, as well from the boars with low protein content in seminal plasma (76% to48%. However, the average diluted semen progressive motility was significantly greater (p<0.01 in the boars with high (64%, compared to the boars with low protein content in seminal plasma (48%. The number of good diluted semen samples (≥65% progressive motility, was also significantly (p<0.01 greater in the boars with high (41%, compared to the boars with low protein content in seminal plasma (12%. These results show that seminal plasma proteins play an important role in maintaining the sperm progressive motility of diluted semen in vitro stored for 3 days.

  14. Hepatocyte cytotoxicity induced by hydroperoxide (oxidative stress model) or glyoxal (carbonylation model): prevention by bioactive nut extracts or catechins.

    Science.gov (United States)

    Banach, Monica S; Dong, Qiang; O'Brien, Peter J

    2009-03-16

    Carbonyl and oxidative stress play important roles in the development of diabetic complications and have been shown to be augmented by various natural compounds and pharmacological agents. Nuts are a rich source of bioactive compounds and antioxidants and various beneficial health effects of nuts have been reported. This study was conducted to evaluate the cytoprotectiveness of various nut extracts and bioactive compounds found in nuts for decreasing cytotoxicity, lipid peroxidation and protein carbonylation in cell toxicity models of diabetes-related carbonyl (glyoxal) and oxidative stress (hydroperoxide). Methanol, ethyl acetate or water were used to prepare crude hazelnut and walnut extracts, which were then used to screen for in vitro cytoprotection of freshly isolated rat hepatocytes against these toxins. The order of protection by nut extracts against hydroperoxide induced cell death was: walnut methanolic extract>walnut aqueous extract>lipophilic walnut extract>hazelnut aqueous extract>hazelnut methanolic extract whereas the lipophilic hazelnut extract did not protect against cell death. The order of protection against lipid peroxidation was the same except for the hazelnut methanolic extract, which prevented lipid peroxidation better than the hazelnut aqueous extract. Catechin, epicatechin and epigallocatechin gallate (EGCG) were investigated for possible protective effects against carbonyl stress cell death and protein carbonylation in hepatocytes. Catechin protected against glyoxal induced cell death and protein carbonylation, and even elicited protection when added to hepatocytes 30 min after the addition of glyoxal. When catechin and epicatechin were compared for protectiveness against glyoxal induced carbonyl stress in hepatocytes, epicatechin protected more effectively than catechin against cell death and protein carbonylation at 120 min. Both compounds also elicited better protection when premixed with glyoxal before addition to hepatocytes, compared

  15. Effect of taurine and caffeine on plasma c-reactive protein and calcium in Wistar rats.

    Science.gov (United States)

    Owoyele, B V; Oyewole, A L; Biliaminu, S A; Alashi, Y

    2015-09-01

    Caffeine is a component of several beverages such as coffee and tea. It has been shown to possess psychoactive properties because it increases alertness, energy and ability to concentrate at moderate doses. Taurine on the other hand, is an amino acid which has the capacity to promote neural development, osmoregulation and neuroprotection. There is paucity of information on the effect of the combined administration of taurine and caffeine on C-reactive protein (CRP)--a marker of inflammation and plasma calcium level in rats. The present study was designed to investigate the effects of combined taurine and caffeine on the plasma level of CRP, Ca2+ as well as the effect of nifedipine on calcium level. Fifty four rats weighing 120-140 g were used for these studies. The animals were divided into nine groups consisting of six animals each. Group 1 was treated with 10 m/kg of normal saline, Groups 2 and 3 were given 100 mg/kg and 200 mg/kg of taurine respectively, groups 4 and 5 received 7.5 mg/kg and 15 mg/kg of caffeine respectively while group 6 was administered taurine (200 mg/kg) and caffeine (15 mg/kg), groups 7 and 8 were treated with taurine (200 mg/kg) plus nifedipine (10 mg/kg) and taurine (200 mg/kg)plus furosemide (20 mg/kg) respectively while group 9 was given taurine plu caffeine plus nifdipine plus furosemide. Treatment was done once daily for 21 days and blood was finally collected via cardiac puncture for the assay of CRP and calcium while the animals were under anaesthesia. The results showed that CRP was significantly decreased in five of the treated groups compared with the control with the exception of the group treated with taurine alone (Group 2), and that treated with combined taurine and caffeine (Group 6). The Ca2+ level of groups treated with caffeine (11.70 ± 0.29 mg/dL) and taurine with caffeine (11.64 ± 0.15 mg/dL) were significantly (p taurine and nifedipine (Group 7) led to significant (p taurine can boost plasma calcium level and

  16. Application of epifluorescence scanning for monitoring the efficacy of protein removal by RF gas-plasma decontamination

    Energy Technology Data Exchange (ETDEWEB)

    Baxter, Helen C; Richardson, Patricia R; Campbell, Gaynor A; Jones, Anita C; Baxter, Robert L [School of Chemistry, Joseph Black Chemistry Building, University of Edinburgh, West Mains Road, Edinburgh EH9 3JJ (United Kingdom); Kovalev, Valeri I; Maier, Robert; Barton, James S [School of Engineering and Physical Science, Heriot-Watt University, Edinburgh EH14 4AS (United Kingdom); DeLarge, Greg [Plasma Etch Inc, 3522 Arrowhead Drive, Carson City, NV 89706 (United States); Casey, Mark [Sterile Services Department, Royal Infirmary of Edinburgh, Edinburgh EH16 4AS (United Kingdom)], E-mail: r.baxter@ed.ac.uk

    2009-11-15

    The development of methods for measuring the efficiency of gas-plasma decontamination has lagged far behind application. An approach to measuring the efficiency of protein removal from solid surfaces using fluorescein-labelled bovine serum albumin and epifluorescence scanning (EFSCAN) is described. A method for fluorescently labelling proteins, which are adsorbed and denatured on metal surfaces, has been developed. Both approaches have been used to evaluate the efficiency of radio frequency (RF) gas-plasma decontamination protocols. Examples with 'real' surgical instruments demonstrate that an argon-oxygen RF gas-plasma treatment can routinely reduce the protein load by about three orders of magnitude beyond that achieved by current decontamination methods.

  17. Carbonyl sulfide: No remedy for global warming

    Science.gov (United States)

    Taubman, Steven J.; Kasting, James F.

    1995-04-01

    The enhancement of the stratospheric aerosol layer caused by the eruption of Mt. Pinatubo (June 15, 1991), and the subsequent cooling of the earth's lower atmosphere [Dutton and Christy, 1992; Minnis et al., 1993] shows that stratospheric aerosols can have a strong effect on the earth's climate. This supports the notion that the intentional enhancement of the stratospheric aerosol layer through increased carbonyl sulfide (OCS) emissions might be an effective means for counteracting global warming. Through the use of a one-dimensional photochemical model, we investigate what effect such a program might have on global average stratospheric ozone. In addition, we consider the impact of enhanced OCS emissions on rainwater acidity and on the overall health of both plants and animals. We find that while the warming produced by a single CO2 doubling (1 to 4°C) might be offset with ozone losses of less than 5%, any attempt to use carbonyl sulfide as a permanent solution to global warming could result in depletion of global average ozone by 30% or more. We estimate that in order to achieve cooling of 4°C rainwater pH would fall to between 3.5 and 3.8. Finally, a 4°C cooling at the surface will require that ambient near ground OCS levels rise to above 10 ppmv which is probably greater than the safe exposure limit for humans. Thus, enhanced OCS emissions do not provide an environmentally acceptable solution to the problem of global warming.

  18. Functional diversity of HIV-1 envelope proteins expressed by contemporaneous plasma viruses

    Directory of Open Access Journals (Sweden)

    Clavel François

    2008-02-01

    Full Text Available Abstract Background Numerous studies have shown that viral quasi-species with genetically diverse envelope proteins (Env replicate simultaneously in patients infected with the human immunodeficiency virus type 1 (HIV-1. Less information is available concerning the extent that envelope sequence diversity translates into a diversity of phenotypic properties, including infectivity and resistance to entry inhibitors. Methods To study these questions, we isolated genetically distinct contemporaneous clonal viral populations from the plasma of 5 HIV-1 infected individuals (n = 70, and evaluated the infectivity of recombinant viruses expressing Env proteins from the clonal viruses in several target cells. The sensitivity to entry inhibitors (enfuvirtide, TAK-799, soluble CD4 and monoclonal antibodies (2G12, 48d, 2F5 was also evaluated for a subset of the recombinant viruses (n = 20. Results Even when comparisons were restricted to viruses with similar tropism, the infectivity for a given target cell of viruses carrying different Env proteins from the same patient varied over an approximately 10-fold range, and differences in their relative ability to infect different target cells were also observed. Variable region haplotypes associated with high and low infectivity could be identified for one patient. In addition, clones carrying unique mutations in V3 often displayed low infectivity. No correlation was observed between viral infectivity and sensitivity to inhibition by any of the six entry inhibitors evaluated, indicating that these properties can be dissociated. Significant inter-patient differences, independent of infectivity, were observed for the sensitivity of Env proteins to several entry inhibitors and their ability to infect different target cells. Conclusion These findings demonstrate the marked functional heterogeneity of HIV-1 Env proteins expressed by contemporaneous circulating viruses, and underscore the advantage of clonal analyses in

  19. Redox chemistry of the molecular interactions between tea catechins and human serum proteins under simulated hyperglycemic conditions.

    Science.gov (United States)

    Özyurt, Hazal; Luna, Carolina; Estévez, Mario

    2016-03-01

    Carbonylation is an irreversible modification in oxidized proteins that has been directly related to a number of health disorders including Type 2 diabetes. Dietary antioxidants have been proposed to counteract the oxidative stress occurring under hyperglycemic conditions. An understanding of the nature and consequences of the molecular interactions between phytochemicals and human plasma proteins is of utmost scientific interest. Three tea catechins namely epicatechin (EC), epigallocatechin (EGC) and epigallocatechin-3-gallate (EGCG) were tested for (i) their affinity to bind to human serum albumin (HSA) and human hemoglobin (HH) and (ii) their ability to inhibit tryptophan (Trp) depletion and for the formation of specific protein carbonyls and pentosidine in the aforementioned proteins. Both proteins (20 mg mL(-1)) were allowed to react with postprandial plasmatic concentrations of the catechins (EC: 0.7 μM, EGC: 1.8 μM, and EGCG: 0.7 μM) under simulated hyperglycemic conditions (12 mM glucose/0.2 mM Fe(3+)/37 °C/10 days). The three catechins were able to inhibit Trp oxidation and protein carbonylation in both plasma proteins. Some anti-glycation properties were linked to their binding affinities. The molecular interactions reported in the present study may explain the alleged beneficial effects of tea catechins against the redox impairment linked to hyperglycemic conditions.

  20. Identification of plasma proteins that are susceptible to thiol oxidation by hypochlorous acid and N-chloramines.

    Science.gov (United States)

    Summers, Fiona A; Morgan, Philip E; Davies, Michael J; Hawkins, Clare L

    2008-09-01

    Hypochlorous acid (HOCl), the major strong oxidant produced by myeloperoxidase, reacts readily with free amino groups to form N-chloramines. Although HOCl and N-chloramines play an important role in the human immune system by killing bacteria and invading pathogens, they have also been shown to cause damage to tissues, which is believed to contribute to a number of diseases. It has been shown previously that N-chloramines react more readily with protein thiols than with other targets in plasma, but the nature of the plasma thiol-containing proteins oxidized is unknown. In this study, the ability of N-chloramines to selectively oxidize thiol-containing plasma proteins was determined using the thiol-specific probe, 5-iodoacetamidofluorescein, combined with two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Experiments were performed with N-chloramines formed on Nalpha-acetyl-lysine, Nalpha-acetyl-histidine (HisCA), glycine, taurine, and ammonia. With the exception of HisCA, the N-chloramines were more efficient than HOCl at oxidizing plasma thiols. The thiol-containing plasma proteins alpha1-antitrypsin and transthyretin were found to be oxidized in addition to albumin, with this treatment resulting in the inactivation of alpha1-antitrypsin. A similar selectivity of reaction and extent of thiol oxidation were also observed with myeloperoxidase in the presence of hydrogen peroxide and chloride ions.

  1. Purification and identification of the fusicoccin binding protein from oat root plasma membrane

    Science.gov (United States)

    de Boer, A. H.; Watson, B. A.; Cleland, R. E.

    1989-01-01

    Fusicoccin (FC), a fungal phytotoxin, stimulates the H(+) -ATPase located in the plasma membrane (PM) of higher plants. The first event in the reaction chain leading to enhanced H(+) -efflux seems to be the binding of FC to a FC-binding protein (FCBP) in the PM. We solubilized 90% of the FCBP from oat (Avena sativa L. cv Victory) root PM in an active form with 1% octyl-glucoside. The FCBP was stabilized by the presence of protease inhibitors. The FCBP was purified by affinity chromatography using FC-linked adipic acid dihydrazide agarose (FC-AADA). Upon elution with 8 molar urea, two major protein bands on sodium dodecyl sulfate-polyaerylamide gel electrophoresis with molecular weights of 29,700 and 31,000 were obtained. Successive chromatography on BBAB Bio-Gel A, hexyl agarose, and FC-AADA resulted in the same two bands when the FC-AADA was eluted with sodium dodecyl sulfate. A direct correlation was made between 3H-FC-binding activity and the presence of the two protein bands. The stoichiometry of the 29,700 and 31,000 molecular weight bands was 1:2. This suggests that the FCBP occurs in the native form as a heterotrimer with an apparent molecular weight of approximately 92,000.

  2. Aggregation analysis of Con A binding proteins of human seminal plasma: a dynamic light scattering study.

    Science.gov (United States)

    Tomar, Anil Kumar; Sooch, Balwinder Singh; Singh, Sarman; Yadav, Savita

    2013-02-01

    Concanavalin A (Con A) binding fraction of human seminal plasma is vital as it shows decapacitating activity and contains proteins which have critical roles in fertility related processes. Con A binding proteins were isolated by lectin affinity chromatography. These proteins form high molecular weight aggregates at near physiological pH (7.0) as inferred by gel filtration. Aggregation analysis was performed by dynamic light scattering (DLS). DLS analysis was also performed at different pH values and in presence of various additives including NaCl, EDTA, cholesterol and sugars, such as d-glucose, d-fructose and d-mannose to identify their effect on aggregation size. The results indicate that degree of aggregation was highly reduced in presence of d-fructose, EDTA and at lower and higher pH values as depicted by lowering of hydrodynamic radii. This aggregation behaviour might be decisive for fertility related events with a suggestive role towards inhibition of premature capacitation.

  3. Myelin basic protein induces neuron-specific toxicity by directly damaging the neuronal plasma membrane.

    Directory of Open Access Journals (Sweden)

    Jie Zhang

    Full Text Available The central nervous system (CNS insults may cause massive demyelination and lead to the release of myelin-associated proteins including its major component myelin basic protein (MBP. MBP is reported to induce glial activation but its effect on neurons is still little known. Here we found that MBP specifically bound to the extracellular surface of the neuronal plasma membrane and induced neurotoxicity in vitro. This effect of MBP on neurons was basicity-dependent because the binding was blocked by acidic lipids and competed by other basic proteins. Further studies revealed that MBP induced damage to neuronal membrane integrity and function by depolarizing the resting membrane potential, increasing the permeability to cations and other molecules, and decreasing the membrane fluidity. At last, artificial liposome vesicle assay showed that MBP directly disturbed acidic lipid bilayer and resulted in increased membrane permeability. These results revealed that MBP induces neurotoxicity through its direct interaction with acidic components on the extracellular surface of neuronal membrane, which may suggest a possible contribution of MBP to the pathogenesis in the CNS disorders with myelin damage.

  4. Rapid changes in plasma membrane protein phosphorylation during initiation of cell wall digestion

    Energy Technology Data Exchange (ETDEWEB)

    Blowers, D.P.; Boss, W.F.; Trewavas, A.J. (Univ. of Edinburgh (England))

    1988-02-01

    Plasma membrane vesicles from wild carrot cells grown in suspension culture were isolated by aqueous two-phase partitioning, and ATP-dependent phosphorylation was measured with ({gamma}-{sup 32}P)ATP in the presence and absence of calcium. Treatment of the carrot cells with the cell wall digestion enzymes, driselase, in a sorbitol osmoticum for 1.5 min altered the protein phosphorylation pattern compared to that of cells treated with sorbitol alone. Driselase treatment resulted in decreased phosphorylation of a band of M{sub r} 80,000 which showed almost complete calcium dependence in the osmoticum treated cells; decreased phosphorylation of a band of M{sub r} 15,000 which showed little calcium activation, and appearance of a new band of calcium-dependent phosphorylation at M{sub r} 22,000. However, protein phosphorylation was decreased. Adding driselase to the in vitro reaction mixture caused a general decrease in the membrane protein phosphorylation either in the presence or absence of calcium which did not mimic the in vivo response. Cells labeled in vivo with inorganic {sup 32}P also showed a response to the Driselase treatment. An enzymically active driselas preparation was required for the observed responses.

  5. Plasma immersion ion implantation of polyurethane shape memory polymer: Surface properties and protein immobilization

    Science.gov (United States)

    Cheng, Xinying; Kondyurin, Alexey; Bao, Shisan; Bilek, Marcela M. M.; Ye, Lin

    2017-09-01

    Polyurethane-type shape memory polymers (SMPU) are promising biomedical implant materials due to their ability to recover to a predetermined shape from a temporary shape induced by thermal activation close to human body temperature and their advantageous mechanical properties including large recovery strains and low recovery stresses. Plasma Immersion Ion Implantation (PIII) is a surface modification process using energetic ions that generates radicals in polymer surfaces leading to carbonisation and oxidation and the ability to covalently immobilise proteins without the need for wet chemistry. Here we show that PIII treatment of SMPU significantly enhances its bioactivity making SMPU suitable for applications in permanent implantable biomedical devices. Scanning Electron Microscopy (SEM), contact angle measurements, surface energy measurements, attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) were used to characterise the PIII modified surface, including its after treatment aging kinetics and its capability to covalently immobilise protein