Biochemical Analysis (biochemical + analysis)

Distribution by Scientific Domains
Distribution within Life Sciences

Kinds of Biochemical Analysis

  • serum biochemical analysis

  • Selected Abstracts

    Effect of Age and Abomasal Puncture on Peritoneal Fluid, Hematology, and Serum Biochemical Analyses in Young Calves

    Luiz Claudio N. Mendes
    The goals of this study were to evaluate techniques for collection of peritoneal fluid from calves, establish reference ranges for fibrinogen in peritoneal fluid during the 1st month of life, and determine if abomasal puncture would alter peritoneal fluid or hematologic variables. Twenty-two healthy Holstein calves underwent 3 peritoneal fluid collections on day 1, day 15, and day 30 of age. Fibrinogen concentration in peritoneal fluid was 0.20 g/dL and 0.10 g/dL (P < .05) for day 1 and day 30, respectively, and 0.10 at day 15 (P > .05) for calves without abomasal puncture. Plasma fibrinogen concentration was 0.60 g/dL and 0.70 g/dL (P < .05) for days 15 and 30, respectively, in calves without abomasal puncture. There were no significant differences (P, .05) in peritoneal fluid and peripheral blood total protein and fibrinogen concentrations, specific gravity, total and differential cell count, or erythrocyte counts between calves with or without abomasal puncture. We concluded that the reference ranges established for fibrinogen and total protein concentration are important for accurate evaluation of peritoneal fluid in calves for further comparison with similar-aged animals with gastrointestinal-tract or abdominal-cavity disease. Additionally, accidental abomasal puncture does not alter values of fibrinogen, total protein, and nucleated cell count in peritoneal fluid and does not cause apparent clinical abnormalities. [source]

    Biochemical Analysis of Pericardial Fluid and Whole Blood in Dogs with Pericardial Effusion

    Armelle M. de Laforcade
    Studies evaluating pericardial fluid analysis in dogs to determine the etiology of pericardial effusions have yielded conflicting results. The purpose of this prospective study was to compare acid-base status, electrolyte concentrations, glucose, and lactate of pericardial fluid to peripheral blood from dogs with pericardial effusion and to compare these variables between dogs with neoplastic and nonneoplastic pericardial effusion. Acid-base status, electrolyte concentrations, glucose, hematocrit, urea nitrogen, and lactate concentrations were evaluated in peripheral blood samples and in pericardial effusion samples of 41 client-owned dogs with pericardial effusion. Common abnormal findings in the peripheral blood of dogs with pericardial effusion included hyperlactatemia (n = 38 [of 41]; 93%), hyponatremia (n = 25/41; 61%), hyperglycemia (n = 13/41; 32%), and hypermagnesemia (n = 13/41; 32%). Bicarbonate, sodium, ionized calcium, glucose, and hematocrit were all significantly lower in the pericardial fluid compared with peripheral blood, whereas lactate, chloride, and PCO2 were significantly higher in the pericardial fluid. When comparing the concentrations of variables in the pericardial fluid of dogs with neoplasia (n = 28) to those without neoplasia (n = 13), pH, bicarbonate, and chloride were significantly lower in dogs with neoplasia, whereas lactate, hematocrit, and urea nitrogen were significantly higher in the pericardial fluid of dogs with neoplasia. The difference between peripheral and pericardial glucose concentrations was significantly larger in dogs with neoplasia than in dogs without neoplasia. Although differences between variables in dogs with neoplastic and nonneoplastic pericardial effusion were documented, clinical relevance is likely limited by the degree of overlap between the 2 groups. [source]

    Extracellular signal-regulated kinase activation is required for consolidation and reconsolidation of memory at an early stage of ontogenesis

    Solène Languille
    Abstract The ability to form long-term memories exists very early during ontogeny; however, the properties of early memory processes, brain structures involved and underlying cellular mechanisms are poorly defined. Here, we examine the role of extracellular signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase/ERK signaling cascade, which is crucial for adult memory, in the consolidation and reconsolidation of an early memory using a conditioned taste aversion paradigm in 3-day-old rat pups. We show that intraperitoneal injection of SL327, the upstream mitogen-activated protein kinase kinase inhibitor, impairs both consolidation and reconsolidation of early memory, leaving short-term memory after acquisition and after reactivation intact. The amnesic effect of SL327 diminishes with increasing delays after acquisition and reactivation. Biochemical analyses revealed ERK hyperphosphorylation in the amygdala but not the hippocampus following acquisition, suggesting functional activation of the amygdala as early as post-natal day 3, although there was no clear evidence for amygdalar ERK activation after reactivation. These results indicate that, despite an immature brain, the basic properties of memory and at least some of the molecular mechanisms and brain structures implicated in aversion memory share a number of similarities with the adult and emerge very early during ontogeny. [source]

    Possible involvement of GABAergic modulation in the protective effect of gabapentin against immobilization stress-induced behavior alterations and oxidative damage in mice

    Anil Kumar
    Abstract Introduction Acute stress may be experienced in response to an immediate physical, emotional or psychological stimulus. Stress has been known to affect several brain activities and promote long-term changes in multiple neural systems. In the present study, we investigated the possible involvement of GABAergic modulation in the protective effect of gabapentin in acute immobilization-induced behavioral alterations and oxidative damage in mice. Materials and methods Mice were immobilized for periods of 6 h. Animals were divided into different groups, consisting of six in each. Various GABAergic modulators were administered either alone or in their combinations, 30 min before subjecting the animals for immobilization stress. Various behavioral tests (mirror chamber, actophotometer) followed by oxidative parameters (malondialdehyde level, glutathione, catalase, nitrite and protein) were assessed in animals. Results Six hours acute immobilization stress caused significant locomotor impairment, anxiety-like behavior in mice. Biochemical analyses also revealed an increase malondialdehyde, nitrite level and depletion of glutathione and catalase activity in 6 h stressed brains. Pretreatment with gabapentin (50 and 100 mg/kg, i.p.) significantly improved ambulatory movements, anti-anxiety effect (decreased time latency to enter in mirror chamber, increased number of entries and duration in mirror chamber) and antioxidative activity in stressed mice (P < 0.05). Further, picrotoxin (1.0 mg/kg) blocked and muscimol (0.05 mg/kg) potentiated the protective action of gabapentin (50 mg/kg). Results of both behavior as well as biochemical alterations in combination studies were significant as compared to their effect per se (P < 0.05). Conclusion Results of present study suggest GABAergic modulation might be involved in the protective effect of gabapentin against immobilization-induced behavior alteration and oxidative damage in mice. [source]

    Functional characterization of human nucleosome assembly protein 1-like proteins as histone chaperones

    GENES TO CELLS, Issue 1 2010
    Mitsuru Okuwaki
    Nucleosome Assembly Protein 1 (NAP1) is a highly conserved histone chaperone protein suspected to be involved in the dynamical regulation of the histone H2A-H2B hetero-dimer. However, the exact mechanism by which NAP1-like proteins act is currently unknown. In this work, we characterized the biochemical properties of two human NAP1-like proteins, hNAP1L1 and hNAP1L4, including a previously uncharacterized subtype, with the aim of determining their exact mechanistic role. Both hNAP1L1 and hNAP1L4 were found to be localized mainly to the cytoplasm and a minor population of them was suggested to be in the nucleus. Biochemical analyses demonstrated that both hNAP1L1 and hNAP1L4 mediated nucleosome formation. In addition, hNAP1L1 was shown to possess a significantly greater nucleosome disassembly activity than hNAP1L4, suggesting that hNAP1L1 and hNAP1L4 may play distinct roles in the regulation of histone dynamics. Building upon this initial discovery we also found that histone H2A-H2B and various histone H2A variants-H2B dimers were found to associate with both hNAP1L1 and hNAP1L4 in cell extracts. These results suggest that human NAP1-like proteins play overlapping roles in transport and deposition of histone H2A-H2B or H2A variants-H2B dimers on chromatin and nonoverlapping roles in nucleosome disassembly. [source]

    Mammalian septin Sept2 modulates the activity of GLAST, a glutamate transporter in astrocytes

    GENES TO CELLS, Issue 1 2004
    Nagatoki Kinoshita
    Sept2 is a member of the septin family of GTPases. Septins form filaments in a GTP-form dependent manner, and are involved in cytokinesis from yeast to mammals; however, some mammalian septins, including Sept2, are expressed in the brain, a tissue in which almost all the cells are postmitotic. Recently, some functions of mammalian septin other than cytokinesis such as vesicle transport have been reported. However, mammalian septin's physiological functions are still unclear. The present study revealed that Sept2 co-localizes with the astrocyte glutamate transporter GLAST in the Bergmann glial processes facing axons and synapses. Biochemical analyses demonstrated that Sept2 bound directly to the carboxy-terminal region of GLAST in a GDP-form dependent manner. Expression of constitutive GDP-form Sept2 mutant reduced the glutamate uptake activity of GLAST via internalization of GLAST from cell surface. Thus Sept2 may regulate GLAST-mediated glutamate uptake by astrocytes, which is important for appropriate transmitter signalling in the cerebellum. [source]

    Two conserved structural components, A-rich bulge and P4 XJ6/7 base-triples, in activating the group I ribozymes

    GENES TO CELLS, Issue 12 2002
    Yoshiya Ikawa
    Background: The A-rich bulge of the group I intron ribozyme, a highly conserved structural element in its P5 peripheral region, plays a significant role in activating the ribozyme. The bulge has been known to interact with the P4 stem forming P4 XJ6/7 base-triples in the conserved core. The base-triples by themselves have also been identified as a distinctive element responsible for enhancing the activity of the ribozyme. Results: A weakly active variant of the Tetrahymena ribozyme lacking the P5 extension was dramatically activated by the addition of an A-rich bulge at the peripheral region, or by replacement of the original P4 XJ6/7 base-triples in the core structure with more stabilized isosteric ones. Biochemical analyses showed that the two methods of activation affect the ribozyme differently. Conclusions: The long-range interaction between the A-rich bulge and P4 or additionally stabilized P4 XJ6/7 base-triples can contribute dramatically to activation of the Tetrahymena ribozyme. Both improve the kcat value, which represents the rate of the limiting step of the ribozyme reaction when its binding site is saturated with GTP. However, the bulge or the modified base-triples gave a moderate reduction or considerable increase, respectively, to the Km(GTP) value. [source]

    Levels of aspartate aminotransferase (AST) in saliva of patients with different periodontal conditions

    Ricardo de Toledo Cesco
    Abstract Objectives: The purpose of this study was to evaluate the relationship between aspartate aminotransferase (AST) levels in saliva measured by ReflotronÔ System of Diagnosis and periodontal condition indicated by Community Periodontal Index of Treatment Needs (CPITN). Material and methods: Fifteen patients were assigned to one of four groups C0, C1, C3 and C4, based on their largest CPITN code among the examined sites, totaling 60 participants. About 1.0 ml of non-stimulated saliva was collected from the individuals after a mouth rinse with water. Biochemical analyses of saliva samples were carried out using the proposed system in order to quantify their AST concentration. Results: There were no significant differences between levels (U/ml) of AST (median; interquartile range) from groups C0 (30.9; 14.7,41.7), C1 (30.3; 19.5,39.4) and C3 (35.1; 27.0,63.5). However, group C4 (106.2; 84.4,129.7) differed statistically from the others (p<0.001) and presented AST levels as high as 284.2 U/ml. Gingival bleeding and suppuration were observed in three individuals with concentrations higher than 125.0 U/ml. Conclusion: Levels of AST in saliva from patients presenting CPITN code 4 were higher than from patients coded lower and could be detected by the evaluated diagnostic system. Periodontal destruction such as periodontal pockets, gingival bleeding and suppuration seems to be related to higher AST levels in saliva. Zusammenfassung Ziele: Der Zweck der Studie war die Evaluation der Beziehung zwischen den Levels von Aspartataminotransferase (AST) im Speichel, die mit dem ReflotronÔ System gemessen wurden, und den parodontalen Bedingungen, die mit dem CPITN erfasst wurden. Material und Methoden: 15 Patienten wurden für eine der vier Gruppen C0, C1, C3 und C4 ausgesucht, was aufgrund ihres höchsten CPITN-Wertes unter den überprüften Flächen bei total 60 Teilnehmern geschah. Ungefähr 1.0 ml von nicht stimuliertem Speichel wurde von den Personen nach einer Mundspülung mit Wasser gesammelt. Die biochemischen Analysen der Speichelproben wurden unter Nutzung des vorgeschlagenen Systems durchgeführt, um die AST Konzentration zu bestimmen. Ergebnisse: Es gab keine signifikanten Differenzen zwischen den Levels (U/ml) von AST (Median, Streuung) bei den Gruppen C0 (30.9; 14.7,41.7), C1 (30.3; 19.5,39.4) und C3 (35.1; 27.0,63.5). Jedoch unterschied sich die Gruppe C4 (106.2; 84.4,129.7) signifikant von den anderen (p<0.001) und zeigte AST Level höher als 284.2 U/ml. Gingivale Blutung und Suppuration wurden bei drei Personen beobachtet mit Konzentrationen höher als 125.0 U/ml. Schlussfolgerung: Die Level von AST im Speichel von Patienten mit einem CPITN von 4 waren höher als bei den Patienten, deren CPITN niedriger war. Sie konnten mit dem evaluierten Diagnostiksystem entdeckt werden. Parodontale Destruktion wie parodontale Taschen, gingivale Blutung und Suppuration scheinen zu höheren AST Level im Speichel in Beziehung zu stehen. Résumé Objectifs: Cette étude se propose d'évaluer la relation entre les niveaux d'aspartate aminotransferase (AST) dans la salive, mesuré par le ReflotronÔ System of Diagnosis et la condition parodontale déterminée par le CPITN. Matériel & méthodes: 15 patients (60 au total) furent répartis dans un des 4 groupes C0, C1, C3 et C4, sur la base de leur plus grand code CPITN parmi les sites examinés. Environ 1.0 ml de salive non stimulée fut prélevé après rinçage à l'eau. Des analyses biochimiques des échantillons salivaires furent réalisées avec le système proposé afin de quantifier la concentration en AST. Résultats: Il n'y avait pas de différences significatives entre les niveaux (U/ml) d'AST (median; interquartile range) entre les groupes C0 (30.9; 14.7,41.7), C1 (30.3; 19.5,39.4) et C3 (35.1; 27.0,63.5). Cependant, le groupe C4 (106.2; 84.4,129.7) présentait une différence significative par rapport aux autres (p<0.001) avec des niveaux d'AST allant jusqu'à 284.2 U/ml. Le saignement gingival et la suppuration s'observaient chez trois individus avec des concentrations au dessus de 125.0 U/ml. Conclusion: Les niveaux d'AST dans la salive de patients au CPITN code 4 étaient plus importants que ceux des patients au code CPITN inférieur et pouvaient être détectés par le système de diagnostique évalué. Des destructions parodontales comme des poches parodontales, des saignements gingivaux et des suppurations semblent en relation avec des niveaux plus grands d'AST dans la salive. [source]

    Thermal tolerance and metabolic physiology among redband trout populations in south-eastern Oregon

    K. J. Rodnick
    Streamside measurements of critical thermal maxima (Tcrit), swimming performance (Ucrit), and routine (Rr) and maximum (Rmax) metabolic rates were performed on three populations of genetically distinct redband trout Oncorhynchus mykiss in the high-desert region of south-eastern Oregon. The Tcrit values (29·4 ± 0·1° C) for small (40,140 g) redband trout from the three streams, and large (400,1400 g) redband trout at Bridge Creek were not different, and were comparable to published values for other salmonids. At high water temperatures (24,28° C), large fish incurred higher metabolic costs and were more thermally sensitive than small fish. Ucrit(3·6 ± 0·1 LF s,1), Rr(200 ± 13 mg O2 kg,0·830 h,1) and metabolic power (533 ± 22 mg O2 kg,0·882 h,1) were not significantly different between populations of small redband trout at 24° C. Rmax and metabolic power, however, were higher than previous measurements for rainbow trout at these temperatures. Fish from Bridge Creek had a 30% lower minimum total cost of transport (Cmin), exhibited a lower refusal rate, and had smaller hearts than fish at 12-mile or Rock Creeks. In contrast, no differences in Ucrit or metabolism were observed between the two size classes of redband trout, although Cmin was significantly lower for large fish at all swimming speeds. Biochemical analyses revealed that fish from 12-mile Creek, which had the highest refusal rate (36%), were moderately hyperkalemic and had substantially lower circulating levels of free fatty acids, triglycerides and albumin. Aerobic and anaerobic enzyme activities in axial white muscle, however, were not different between populations, and morphological features were similar. Results of this study: 1) suggest that the physiological mechanisms that determine Tcrit in salmonids are highly conserved; 2) show that adult (large) redband trout are more susceptible to the negative affects of elevated temperatures than small redband trout; 3) demonstrate that swimming efficiency can vary considerably between redband trout populations; 4) suggest that metabolic energy stores correlate positively with swimming behaviour of redband trout at high water temperatures; 5) question the use of Tcrit for assessing physiological function and defining thermal habitat requirements of stream-dwelling salmonids like the redband trout. [source]

    The zinc-finger protein ZFR is critical for Staufen 2 isoform specific nucleocytoplasmic shuttling in neurons

    George Elvira
    Abstract In mammalian neurons, transport and translation of mRNA to individual potentiated synapses is believed to occur via a heterogeneous population of RNA granules. To identify components of Staufen2-containing granules, we used the yeast two-hybrid system. A mouse fetal cDNA library was screened with the N-terminal fragment of Staufen2 as bait. ZFR, a three zinc finger protein, was identified as an interacting protein. Confocal microscopy showed that ZFR, although mainly nuclear, was also found in the somatodendritic compartment of primary hippocampal neurons where it localized as granule-like structures. Co-localization with Staufen2 was observed in several granules. Biochemical analyses (immunoprecipitation, cell fractionation) further confirmed the ZFR/Staufen2 association. ZFR was shown to interact with at least the Staufen262 isoform, but not with Staufen1. ZFR also co-fractionated with ribosomes and Staufen259 and Staufen252 in a sucrose gradient. Interestingly, knockdown expression of ZFR through RNA interference in neurons relocated specifically the Staufen262, but not the Staufen259, isoform to the nucleus. Our results demonstrate that ZFR is a native component of Staufen2-containing granules and likely plays its role during early steps of RNA transport and localization. They also suggest that one of these roles may be linked to Staufen262 -containing RNA granule formation in the nucleus and/or to their nucleo-cytoplasmic shuttling. [source]

    Molecular mechanisms of mild and moderate hemophilia A

    M. Jacquemin
    Summary., Mutations responsible for mild/moderate hemophilia A were extensively characterized over the last 15 years and more than 200 mutations have been identified. However, most of the molecular mechanisms responsible for the reduced factor (F)VIII levels in patients' plasma were determined only recently. Recent progresses in the study of the FVIII molecule three-dimensional structure provided a major insight for understanding molecular events leading to mild/moderate hemophilia A. This allowed prediction of mutations impairing FVIII folding and intracellular processing, which result in reduced FVIII secretion. Mutations potentially slowing down FVIII activation by thrombin were also identified. A number of mutations were also predicted to result in altered stability of activated FVIII. Biochemical analyses allowed identification of mutations reducing FVIII production. Mutations impairing FVIII stability in plasma, by reducing FVIII binding to von Willebrand factor (VWF) were also characterized. Defects in FVIII activity, notably slow activation by thrombin, or abnormal interaction with FIXa, were also recently demonstrated. Biochemical analysis of FVIII variants provided information regarding the structure/function relationship of the FVIII molecule and validated predictions of the three-dimensional structure of the molecule. These observations also contributed to explain the discrepant activities recorded for some FVIII variants using different types of FVIII assays. Altogether, the study of the biochemical properties of FVIII variants and the evaluation of the effects of mutations in three-dimensional models of FVIII identified molecular mechanisms potentially explaining reduced FVIII levels for a majority of patients with mild/moderate hemophilia A. It is expected that these studies will improve diagnosis and treatment of this disease. [source]

    Resistance to ACCase-inhibiting herbicides and isoproturon in UK populations of Lolium multiflorum: mechanisms of resistance and implications for control

    Kay M Cocker
    Abstract Herbicide-resistant Lolium multiflorum (Italian rye-grass) was first reported in the UK in 1993 and had been confirmed on 25 farms by 1999. In this study, resistance to five herbicides belonging to the aryloxyphenoxypropionate, cyclohexanedione and phenyl-urea classes was determined in six populations of L multiflorum from the UK under glasshouse and simulated field conditions. Glasshouse conditions tended to exaggerate the degree of resistance, but experiments performed in both environments detected resistance in four populations of L multiflorum. Four populations (Essex A1, Lincs A1, Wilts B1, Yorks A2) were resistant to diclofop-methyl, fluazifop-P-butyl, tralkoxydim and partially resistant to isoproturon, but only the population from Yorkshire (Yorks A2) showed resistance to cycloxydim. Biochemical analyses of acetyl coenzyme A carboxylase (ACCase) activity, oxygen consumption by thylakoids, diclofop metabolism and glutathione S -transferase activity showed that, in three of the resistant populations, an enhanced rate of herbicide metabolism conferred resistance. This is the first report world-wide of an enhanced metabolism mechanism of diclofop resistance in L multiflorum. In the Yorks A2 population, an insensitive ACCase was detected (target-site resistance) which also conferred cross-resistance to all of the other ACCase inhibitors investigated. © 2001 Society of Chemical Industry [source]

    Neuroprotective effects of Tanshinone IIA on permanent focal cerebral ischemia in mice,

    Kenan Dong
    Abstract The objective of this study was to evaluate whether Tanshinone IIA (TSA) was neuroprotective in permanent focal cerebral ischemia and to determine the possible mechanisms of its neuroprotection. Mice were subjected to permanent middle cerebral artery occlusion. The neuroprotection of TSA was investigated with respect to neurological deficit scores and infarct volume. Biochemical analyses for malondialdehyde (MDA) content and superoxide dismutase (SOD) activity in serum, and nitric oxide (NO) content and the inducible nitric oxide synthase (iNOS) activity in brain tissue were performed at 24 h after ischemia. Immunohistochemistry was used to measure the expression of iNOS. In vitro, the effects of TSA were tested in the cultured astrocytes exposed to hydrogen dioxide (H2O2). TSA (5, 10 and 20 mg/kg, i.p.) significantly reduced the infarct volume and improve neurological deficit. TSA also significantly increased the activity of SOD after 24 h of ischemia and decreased the MDA level, NO content, and iNOS expression. In vitro, the translocation of NF- ,B was inhibited by TSA and the survival rate of astrocytes was markedly increased and the NO production was decreased. In conclusion, these results illustrated that TSA protected the brain from ischemic injury by suppressing the oxidative stress and the radical-mediated inflammatory insult. Copyright © 2008 John Wiley & Sons, Ltd. [source]

    Modulation of peripheral B cell tolerance by CD72 in a murine model

    ARTHRITIS & RHEUMATISM, Issue 10 2008
    Daniel Hsieh-Hsin Li
    Objective B cells play a dominant role in the pathogenesis of several autoimmune diseases, including systemic lupus erythematosus. It is not well understood how B cell signaling contributes to autoantibody production. The goal of this study was to elucidate the role of CD72 in modulating B cell receptor (BCR),mediated tolerogenic signaling and peripheral B cell tolerance. Methods A mouse model utilizing hen egg lysozyme (HEL) "anergic" B cells was studied. CD72-deficient mice carrying the BCR-specific IgHEL and/or soluble HEL (sHEL) transgenes were generated by breeding IgHEL -transgenic MD4 mice and/or sHEL -transgenic ML5 mice with congenic, CD72-deficient C57BL/6J mice. Normal and anergic B cells were isolated for analyses of B cell signaling. Aged wild-type and CD72-deficient mice were also examined for autoimmune phenomena. Results In the absence of CD72, anergic B cells inappropriately proliferated and survived in response to stimulation with self antigen. Biochemical analyses indicated that in anergic B cells, CD72 dominantly down-regulated BCR signaling to limit the antigen-induced elevation in [Ca2+]i and the activation of NFATc1, NF-,B, MAPK, and Akt. Mechanistically, CD72 was associated with, and regulated, the molecular adaptor Cbl-b in anergic B cells, suggesting that Cbl-b may play a role in mediating the negative effects of CD72 on BCR signaling. Moreover, in aged CD72-deficient mice, spontaneous production of antinuclear and anti,double-stranded DNA autoantibodies and features of lupus-like autoimmune disease were observed. Conclusion CD72 is required to maintain B cell anergy and functions as a regulator of peripheral B cell tolerance. Thus, altered CD72 expression may play a role during the development of systemic lupus erythematosus. [source]

    Effects of antioxidant stobadine on protein carbonylation, advanced oxidation protein products and reductive capacity of liver in streptozotocin-diabetic rats: Role of oxidative/nitrosative stress

    BIOFACTORS, Issue 3 2007
    Ahmet Cumao
    Background: Increased oxidative/nitrosative stress is important in the pathogenesis of diabetic complications, and the protective effects of antioxidants are a topic of intense research. The purpose of this study was to investigate whether a pyridoindole antioxidant stobadine (STB) have a protective effect on tissue oxidative protein damage represented by the parameters such as protein carbonylation (PC), protein thiol (P-SH), total thiol (T-SH) and non-protein thiol (Np-SH), nitrotyrosine (3-NT), and advanced oxidation protein products (AOPP) in streptozotocin-diabetic rats. Methods: Diabetes was induced in male Wistar rats by intraperitonal injection of streptozotocin (55 mg/kg). Some of the non-diabetic (control) and diabetic rats treated with STB (24.7 mg/kg/day) during 16 weeks, and the effects on blood glucose, PC, AOPP, 3-NT, P-SH, T-SH and Np-SH were studied. Biomarkers were assayed by enzyme-linked immunosorbent assay (ELISA) or by colorimetric methods. Results: Administration of stobadine to diabetic animals lowered elevated blood glucose levels by ,16% relative to untreated diabetic rats. Although stobadine decreased blood glucose, poor glycemic control was maintained in stobadine treated diabetic rats during the treatment period. Biochemical analyses of liver proteins showed significant diminution of sulfhydryl groups, P-SH, T-SH, Np-SH, and elevation of carbonyl groups in diabetic animals in comparison to healthy controls. As a biomarker of nitrosative stress, 3-NT levels did not significantly change by diabetes induction or by stobadine treatment when compared to control animals. However, the treatment with stobadine resulted in a significant decrease in PC, AOPP levels and normalized P-SH, T-SH, Np-SH groups in liver of diabetic animals. [source]

    A mutant of the green alga Dunaliella salina constitutively accumulates zeaxanthin under all growth conditions

    EonSeon Jin
    Abstract A novel mutant (zea1) of the halotolerant unicellular green alga Dunaliella salina is impaired in the zeaxanthin epoxidation reaction, thereby lacking a number of the ,-branch xanthophylls. HPLC analysis revealed that the zea1 mutant lacks neoxanthin (N), violaxanthin (V) and antheraxanthin (A) but constitutively accumulates zeaxanthin (Z). Under low-light physiological growth conditions, the zea1 (6 mg Z per g dry weight or 8 × 10,16 mol Z/cell) had a substantially higher Z content than the wild type (0.2 mg Z per g dry weight or 0.5 × 10,16 mol Z/cell). Lack of N, V, and A did not affect photosynthesis or growth of the zea1 strain. Biochemical analyses suggested that Z constitutively and quantitatively substitutes for N, V, and A in the zea1 strain. This mutant is discussed in terms of its commercial value and potential utilization by the algal biotechnology industry for the production of zeaxanthin, a high-value bioproduct. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 115,124, 2003. [source]

    Interallelic complementation provides genetic evidence for the multimeric organization of the Phycomyces blakesleeanus phytoene dehydrogenase

    FEBS JOURNAL, Issue 3 2002
    Catalina Sanz
    The Phycomyces blakesleeanus wild-type is yellow, because it accumulates ,-carotene as the main carotenoid. A new carotenoid mutant of this fungus (A486) was isolated, after treatment with ethyl methane sulfonate (EMS), showing a whitish coloration. It accumulates large amounts of phytoene, small quantities of phytofluene, ,-carotene and neurosporene, in decreasing amounts, and traces of ,-carotene. This phenotype indicates that it carries a leaky mutation affecting the enzyme phytoene dehydrogenase (EC 1.3.-.-), which is specified by the gene carB. Biochemical analysis of heterokaryons showed that mutant A486 complements two previously characterized carB mutants, C5 (carB10) and S442 (carB401). Sequence analysis of the carB gene genomic copy from these three strains revealed that they are all altered in the gene carB, giving information about the nature of the mutation in each carB mutant allele. The interallelic complementation provides evidence for the multimeric organization of the P. blakesleeanus phytoene dehydrogenase. [source]

    Involvement of thiaminase II encoded by the THI20 gene in thiamin salvage of Saccharomyces cerevisiae

    FEMS YEAST RESEARCH, Issue 2 2008
    Mari Onozuka
    Abstract The physiological significance of thiaminase II, which catalyzes the hydrolysis of thiamin, has remained elusive for several decades. The C-terminal domains of THI20 family proteins (THI20/21/22) and the whole region of PET18 gene product of Saccharomyces cerevisiae are homologous to bacterial thiaminase II. On the other hand, the N-terminal domains of THI20 and THI21 encode 2-methyl-4-amino-5-hydroxymethylpyrimidine kinase and 2-methyl-4-amino-5-hydroxymethylpyrimidine phosphate kinase involved in the thiamin synthetic pathway. In this study, it was first indicated that the C-terminal domains of the THI20 family and PET18 are not required for de novo thiamin synthesis in S. cerevisiae, using a quadruple deletion strain expressing the N-terminal domain of THI20. Biochemical analysis using cell-free extracts and recombinant proteins demonstrated that yeast thiaminase II activity is exclusively encoded by THI20. It appeared that Thi20p has an affinity for the pyrimidine moiety of thiamin, and HMP produced by the thiaminase II activity is immediately phosphorylated. Thi20p was found to participate in the formation of thiamin from two synthetic antagonists, pyrithiamin and oxythiamin, by hydrolyzing both antagonists and phosphorylating HMP to give HMP pyrophosphate. Furthermore, 2-methyl-4-amino-5-aminomethylpyrimidine, a presumed naturally occurring thiamin precursor, was effectively converted to HMP by incubation with Thi20p. It is proposed that the thiaminase II activity of Thi20p is involved in the thiamin salvage pathway by catalyzing the hydrolysis of HMP precursors in S. cerevisiae. [source]

    Protection of corticospinal tract neurons after dorsal spinal cord transection and engraftment of olfactory ensheathing cells

    GLIA, Issue 4 2006
    Masanori Sasaki
    Abstract Transplantation of olfactory ensheathing cells (OECs) into the damaged rat spinal cord leads to directed elongative axonal regeneration and improved functional outcome. OECs are known to produce a number of neurotrophic molecules. To explore the possibility that OECs are neuroprotective for injured corticospinal tract (CST) neurons, we transplanted OECs into the dorsal transected spinal cord (T9) and examined primary motor cortex (M1) to assess apoptosis and neuronal loss at 1 and 4 weeks post-transplantation. The number of apoptotic cortical neurons was reduced at 1 week, and the extent of neuronal loss was reduced at 4 weeks. Biochemical analysis indicated an increase in BDNF levels in the spinal cord injury zone after OEC transplantation at 1 week. The transplanted OECs associated longitudinally with axons at 4 weeks. Thus, OEC transplantation into the injured spinal cord has distant neuroprotective effects on descending cortical projection neurons. © 2005 Wiley-Liss, Inc. [source]

    The growth respiration component in eddy CO2 flux from a Quercus ilex mediterranean forest

    S. Rambal
    Abstract Ecosystem respiration, arising from soil decomposition as well as from plant maintenance and growth, has been shown to be the most important component of carbon exchange in most terrestrial ecosystems. The goal of this study was to estimate the growth component of whole-ecosystem respiration in a Mediterranean evergreen oak (Quercus ilex) forest over the course of 3 years. Ecosystem respiration (Reco) was determined from night-time carbon dioxide flux (Fc) using eddy correlation when friction velocity (u*) was greater than 0.35 m s,1 We postulated that growth respiration could be evaluated as a residual after removing modeled base Reco from whole-ecosystem Reco during periods when growth was most likely occurring. We observed that the model deviated from the night-time Fc -based Reco during the period from early February to early July with the largest discrepancies occurring at the end of May, coinciding with budburst when active aboveground growth and radial growth increment are greatest. The highest growth respiration rates were observed in 2001 with daily fluxes reaching up to 4 g C m,2. The cumulative growth respiration for the entire growth period gave total carbon losses of 170, 208, and 142 g C m,2 for 1999, 2001, and 2002, respectively. Biochemical analysis of soluble carbohydrates, starch, cellulose, hemicellulose, proteins, lignin, and lipids for leaves and stems allowed calculation of the total construction costs of the different growth components, which yielded values of 154, 200, and 150 g C for 3 years, respectively, corresponding well to estimated growth respiration. Estimates of both leaf and stem growth showed very large interannual variation, although average growth respiration coefficients and average yield of growth processes were fairly constant over the 3 years and close to literature values. The time course of the growth respiration may be explained by the growth pattern of leaves and stems and by cambial activity. This approach has potential applications for interpreting the effects of climate variation, disturbances, and management practices on growth and ecosystem respiration. [source]

    m.6267G>A: a recurrent mutation in the human mitochondrial DNA that reduces cytochrome c oxidase activity and is associated with tumors,

    HUMAN MUTATION, Issue 6 2006
    M. Esther Gallardo
    Abstract Complete sequencing of the mitochondrial genome of 13 cell lines derived from a variety of human cancers revealed nine novel mitochondrial DNA (mtDNA) variations. One of them, m.6267G>A, is a recurrent mutation that introduces the Ala122Thr substitution in the mitochondrially encoded cytochrome c oxidase I (MT-CO1): p.MT-CO1: Ala122Thr (GenBank: NP_536845.1). Biochemical analysis of the original cell lines and the transmitochondrial cybrids generated by transferring mitochondrial DNAs to a common nuclear background, indicate that cytochrome c oxidase (COX) activity, respiration, and growth in galactose are impaired by the m.6267G>A mutation. This mutation, found twice in the cancer cell lines included in this study, has been also encountered in one out of 63 breast cancer samples, one out of 64 colon cancer samples, one out of 260 prostate cancer samples, and in one out of 15 pancreatic cancer cell lines. In all instances the m.6267G>A mutation was associated to different mtDNA haplogroups. These findings, contrast with the extremely low frequency of the m.6267G>A mutation in the normal population (1:2264) and its apparent absence in other pathologies, strongly suggesting that the m.6267G>A missense mutation is a recurrent mutation specifically associated with cancer. Hum Mutat 27(6), 575,582, 2006. © 2006 Wiley-Liss, Inc. [source]

    Biochemical and mutational analyses of the cathepsin c gene (CTSC) in three North American families with Papillon Lefèvre syndrome

    HUMAN MUTATION, Issue 1 2002
    Y. Zhang
    Abstract Papillon Lefèvre syndrome (PLS) is an autosomal recessive disorder characterized by palmoplantar hyperkeratosis and severe periodontitis. The disease is caused by mutations in the cathepsin C gene (CTSC) that maps to chromosome 11q14. CTSC gene mutations associated with PLS have been correlated with significantly decreased enzyme activity. Mutational analysis of the CTSC gene in three North American families segregating PLS identified four mutations, including a novel mutation p.G139R. All mutations were associated with dramatically reduced CTSC protease enzyme activity. A homozygous c.96T>G transversion resulting in a p.Y32X change was present in a Mexican PLS proband, while one Caucasian PLS proband was a compound heterozygote for the p.Y32X and p.R272P (c.815G>C) mutations. The other Caucasian PLS proband was a compound heterozygote for c.415G>A transition and c.1141delC mutations that resulted in a p.G139R and a frameshift and premature termination (p.L381fsX393), respectively. The c.415G>A was not present in more than 300 controls, suggesting it is not a CTSC polymorphism. Biochemical analysis demonstrated almost no detectable CTSC activity in leukocytes of all three probands. These mutations altered restriction enzyme sites in the highly conserved CTSC gene. Sequence analysis of CTSC exon 3 confirmed the previously reported p.T153I polymorphism in 4 of the 5 ethnically diverse populations studied. © 2002 Wiley-Liss, Inc. [source]


    ABSTRACT Processing techniques like cooking and freezing exhibited significant (P < 0.001) reduction in the bacterial load of cuttlefish, Sepia pharaonis, and marine crab, Portunus pelagicus. Raw cuttle fish had 2.4 × 107 cfu/g which on cooking reduced to 9.7 × 106 cfu/g. Freezing reduced the bacterial load further as cooked frozen product had only 9.9 × 104 cfu/g. Similarly, raw crab had 2.6 × 107 cfu/g which on cooking reduced to 6.5 × 106 cfu/g. A further reduction in bacterial load was seen after freezing as cooked frozen crab exhibited only 7.3 × 104 cfu/g. Escherichia coli and Staphylococcus aureus were present in the limit of acceptability for fish and fish products. Salmonella typhimurium and Vibrio cholerae were absent even in raw stage. Biochemical analysis performed on stored frozen products of cuttle fish and crab exhibited a significant (P , 0.05) increase in bacterial spoilage and rancidity with increasing days of storage. Total volatile base nitrogen, trimethylamine, thiobarbituric acid and free fatty acid contents in frozen products of cuttle fish and crab increased significantly with 120 days of frozen storage. [source]

    Characterization of VR1 within the BMBF-Leitproject: ,Molecular Pain Research'

    R. Jostock
    The vanilloid receptor VR1 is a ligand, heat and proton gated ion channel, expressed predominantly by primary sensory neurons. We show the molecular characterization of VR1 and its involvement in nociceptive behavior. Biochemical analysis of VR1 showed glycosylation at N604 and the predicted tetrameric structure. Reduced pH potentiated the gating of the receptor by NADA and anandamide in recombinant VR1. Acidification could sensitize VR1 and lead to hyperalgesia. Therefore, the VR1 antagonist capsazepine was tested in several animal models. Capsazepine reduced formalin induced nocifensive behavior and CFA induced mechanical hyperalgesia, and was antiallodynic and antihyperalgesic in animal models of neuropathic pain. VR1 antisense oligonucleotides inhibited VR1 expression in vitro and reduced tactile allodynia in vivo. In conclusion, we could provide evidence for a role of VR1 in inflammatory and neuropathic pain pathways. [source]

    Assessment of acid-base status of cats with naturally occurring chronic renal failure

    J. Elliott
    Metabolic acidosis is reported to be a common complication of feline chronic renal failure (CRF) but acid-base status of feline patients with this disease is rarely assessed by general practitioners. A cross-sectional study involving 59 cases of naturally occurring feline CRF was conducted to determine the prevalence of acid-base disturbances. Cases were categorised on the basis of their plasma creatinine concentrations as mild, moderate or severe. A group of 27 clinically healthy, age-matched cats was assessed for comparison. A low venous blood pH (<7·270) was found in 10 of the 19 severe cases (52·6 per cent), three of the 20 moderate cases (15 per cent) and none of the 20 mild cases. Acidaemia was associated with an increased anion gap contributed to by both low plasma bicarbonate and low chloride ion concentrations. Biochemical analysis of urine samples showed urine pH to decrease with increasing severity of renal failure. Urinary loss of bicarbonate was not associated with the occurrence of acidaemia and there was a tendency for urinary ammonium ion excretion to decrease as the severity of renal failure increased. Cats with naturally occurring CRF do not show plasma biochemical evidence of acid-base disturbances until the disease is advanced. [source]

    Molecular mechanisms of mild and moderate hemophilia A

    M. Jacquemin
    Summary., Mutations responsible for mild/moderate hemophilia A were extensively characterized over the last 15 years and more than 200 mutations have been identified. However, most of the molecular mechanisms responsible for the reduced factor (F)VIII levels in patients' plasma were determined only recently. Recent progresses in the study of the FVIII molecule three-dimensional structure provided a major insight for understanding molecular events leading to mild/moderate hemophilia A. This allowed prediction of mutations impairing FVIII folding and intracellular processing, which result in reduced FVIII secretion. Mutations potentially slowing down FVIII activation by thrombin were also identified. A number of mutations were also predicted to result in altered stability of activated FVIII. Biochemical analyses allowed identification of mutations reducing FVIII production. Mutations impairing FVIII stability in plasma, by reducing FVIII binding to von Willebrand factor (VWF) were also characterized. Defects in FVIII activity, notably slow activation by thrombin, or abnormal interaction with FIXa, were also recently demonstrated. Biochemical analysis of FVIII variants provided information regarding the structure/function relationship of the FVIII molecule and validated predictions of the three-dimensional structure of the molecule. These observations also contributed to explain the discrepant activities recorded for some FVIII variants using different types of FVIII assays. Altogether, the study of the biochemical properties of FVIII variants and the evaluation of the effects of mutations in three-dimensional models of FVIII identified molecular mechanisms potentially explaining reduced FVIII levels for a majority of patients with mild/moderate hemophilia A. It is expected that these studies will improve diagnosis and treatment of this disease. [source]

    Disruption of msl3 abolishes the synthesis of mycolipanoic and mycolipenic acids required for polyacyltrehalose synthesis in Mycobacterium tuberculosis H37Rv and causes cell aggregation

    Vinod S. Dubey
    Summary Cell wall lipids of Mycobacterium tuberculosis containing multiple methylbranched fatty acids play critical roles in pathogenesis and thus offer targets for new antimycobacterial drugs. Mycocerosic acid synthase gene (mas) encodes the enzyme that produces one class of such acids. Seven mas -like genes (msls) were identified in the genome. One of them, msl3, originally annotated as two separate genes, pks 3 and pks 4, is now shown to constitute a single open reading frame, which encodes a 220.3 kDa protein. Msl3 was disrupted using a phage mediated delivery system and the gene replacement in the mutant was confirmed by polymerase chain reaction analysis of the flanking regions of the introduced disrupted gene and by Southern analysis. Biochemical analysis showed that the msl3 mutant does not produce mycolipanoic acids and mycolipenic (phthienoic) acids, the major constituents of polyacyl trehaloses and thus lacks this cell wall lipid, but synthesizes all of the other classes of lipids. The absence of the major acyl chains that anchor the surface-exposed acyltrehaloses causes a novel growth morphology; the cells stick to each other, most probably via the intercellular interaction between the exposed hydrophobic cell surfaces, manifesting a bead-like growth morphology without affecting the overall growth rate. [source]

    Protective effect of quercetin against ICV colchicine-induced cognitive dysfunctions and oxidative damage in rats

    Anil Kumar
    Abstract Intracerebroventricular (i.c.v.) administration of colchicine, a microtubule-disrupting agent, causes cognitive dysfunction and oxidative stress. The present study was designed to investigate the protective effects of quercetin against colchicine-induced memory impairment and oxidative damage in rats. An i.c.v. cannula was implanted in the lateral ventricle of male Wistar rats. Colchicine was administered at dose of 15 µg/rat. Morris water maze and plus-maze performance tests were used to assess memory tasks. Various biochemical parameters such as lipid peroxidation, reduced glutathione, nitrite level, acetylcholinesterase and proteins were also assessed. Central administration of colchicine (15 µg/rat) showed poor retention of memory. Chronic treatment with quercetin (20 and 40 mg/kg, p.o.) twice daily for a period of 25 days beginning 4 days prior to colchicine injection significantly improved the colchicine-induced cognitive impairment. Biochemical analysis revealed that i.c.v. colchicine injection significantly increased lipid peroxidation, nitrite and depleted reduced glutathione activity in the brains of rats. Chronic administration of quercetin significantly attenuated elevated lipid peroxidation and restored the depleted reduced glutathione, acetylcholinesterase activity and nitrite activity. The results of the present study clearly indicated that quercetin has a neuroprotective effect against colchicine-induced cognitive dysfunctions and oxidative damage. This article was published online on 3 November 2008. An error was subsequently identified. This notice is included in the online and print version to indicate that both have been corrected. [24 November 2008] Copyright © 2008 John Wiley & Sons, Ltd. [source]

    A 10-kDa acyl-CoA-binding protein (ACBP) from Brassica napus enhances acyl exchange between acyl-CoA and phosphatidylcholine

    Olga P. Yurchenko
    Summary The gene encoding a 10-kDa acyl-CoA-binding protein (ACBP) from Brassica napus was over-expressed in developing seeds of Arabidopsis thaliana. Biochemical analysis of T2 and T3 A. thaliana seeds revealed a significant increase in polyunsaturated fatty acids (FAs) (18:2cis,9,12 and 18:3cis,9,12,15) at the expense of very long monounsaturated FA (20:1cis,11) and saturated FAs. In vitro assays demonstrated that recombinant B. napus ACBP (rBnACBP) strongly increases the formation of phosphatidylcholine (PC) in the absence of added lysophosphatidylcholine in microsomes from ,YOR175c yeast expressing A. thaliana lysophosphatidylcholine acyltransferase (AthLPCAT) cDNA or in microsomes from microspore-derived cell suspension cultures of B. napus L. cv. Jet Neuf. rBnACBP or bovine serum albumin (BSA) were also shown to be crucial for AthLPCAT to catalyse the transfer of acyl group from PC into acyl-CoA in vitro. These data suggest that the cytosolic 10-kDa ACBP has an effect on the equilibrium between metabolically active acyl pools (acyl-CoA and phospholipid pools) involved in FA modifications and triacylglycerol bioassembly in plants. Over-expression of ACBP during seed development may represent a useful biotechnological approach for altering the FA composition of seed oil. [source]

    X-Linked dominant chondrodysplasia punctata: prenatal diagnosis and autopsy findings

    PRENATAL DIAGNOSIS, Issue 13 2006
    Shalini Umranikar
    Abstract Objective To report our experience of the prenatal diagnosis of X-linked dominant chondrodysplasia punctata (CDPX2) and highlight its variable phenotypic presentation. Methods We report the sonographic features of three female fetuses affected with CDPX2. The ultrasound, radiographic and pathological findings were compared. Results Family 1: Two affected pregnancies, both terminated. Fetus 1: Presented with epiphyseal stippling involving the vertebrae, upper and lower limbs, asymmetric shortening of the long bones and flat facial profile. Fetus 2: Prenatal findings included premature epiphyseal stippling, paravertebral cartilaginous calcific foci, mild shortening of the long bones and flat facies. Mutation analysis of the mother and both fetuses revealed mutation in the emopamil-binding protein (EBP) gene. Family 2: Prenatal sonography showed scattered epiphyseal stippling, minimal vertebral segmentation anomalies, mild asymmetric limb shortening and flat facies. Female infant delivered at 39 weeks of gestation. Biochemical analysis in all three fetuses showed increased levels of serum 8(9)-cholestenol consistent with delta (8), delta (7)-isomerase deficiency and CDPX2. Conclusion Prenatal diagnosis of CDPX2 is difficult because of marked phenotypic variation. Epiphyseal stippling, ectopic paravertebral calcifications, asymmetric shortening of long bones and dysmorphic flattened facies are crucial for prenatal diagnosis. DNA analysis of the CDPX2 gene and biochemical determination of the serum 8(9)-cholestenol level are important for diagnosis, especially if future pregnancies are planned. Copyright © 2006 John Wiley & Sons, Ltd. [source]