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Sample records for creatine kinase activities

  1. Creatine kinase activity is associated with blood pressure

    NARCIS (Netherlands)

    Brewster, Lizzy M.; Mairuhu, Gideon; Bindraban, Navin R.; Koopmans, Richard P.; Clark, Joseph F.; van Montfrans, Gert A.

    2006-01-01

    BACKGROUND: We previously hypothesized that high activity of creatine kinase, the central regulatory enzyme of energy metabolism, facilitates the development of high blood pressure. Creatine kinase rapidly provides adenosine triphosphate to highly energy-demanding processes, including cardiovascular

  2. Creatine kinase activity in dogs with experimentally induced acute inflammation

    Directory of Open Access Journals (Sweden)

    Dimitrinka Zapryanova

    2013-01-01

    Full Text Available The main purpose of this study was to investigate the effect of acute inflammation on total creatine kinase (CK activity in dogs. In these animals, CK is an enzyme found predominantly in skeletal muscle and significantly elevated serum activity is largely associated with muscle damage. Plasma increases in dogs are associated with cell membrane leakage and will therefore be seen in any condition associated with muscular inflammation. The study was induced in 15 mongrel male dogs (n=9 in experimental group and n=6 in control group at the age of two years and body weight 12-15 kg. The inflammation was reproduced by inoculation of 2 ml turpentine oil subcutaneously in lumbar region. The plasma activity of creatine kinase was evaluated at 0, 6, 24, 48, 72 hours after inoculation and on days 7, 14 and 21 by a kit from Hospitex Diagnostics. In the experimental group, the plasma concentrations of the CK-activity were increased at the 48th hour (97.48±6.92 U/L and remained significantly higher (p<0.05 at the 72 hour (97.43±2.93 U/L compared to the control group (77.08±5.27 U/L. The results of this study suggest that the evaluation of creatine kinase in dogs with experimentally induced acute inflammation has a limited diagnostic value. It was observed that the creatine kinase activity is slightly affected by the experimentally induced acute inflammation in dogs.

  3. Creatine-creatine phosphate shuttle modeled as two-compartment system at different levels of creatine kinase activity

    DEFF Research Database (Denmark)

    Fedosov, Sergey

    1994-01-01

    In order to characterize ADP-ATP and creatine-creatine phosphate (Cr-CrP) shuttles a minimal mathematical model with two compartments and cyclic turnover of matter was designed. The 'mitochondrial' compartment contained 'ATP-synthase' and 'mitochondrial ereatine kinase' (mitCK). The 'cytoplasmic......' compartment consisted of 'ATPase', 'cytoplasmic creatine kinase' (cytCK) and an 'ADP-binding structure'. The exchange of metabolites between these compartments was limited. Different levels of cytCK and mitCK expression as welt as different exchange rate constants between the compartments were assigned...

  4. Evaluation of Creatine Kinase Activity and Inorganic Phosphate ...

    African Journals Online (AJOL)

    subjects presenting with major VOC. Keywords: Serum creatine kinase activity, Serum inorganic phosphate concentration, Sickle cell disease,. Steady state, Vaso‑occlusive crisis. Original Article. Address for correspondence: Dr. John C Aneke,. Department of Hematology,. Nnamdi Azikiwe University Teaching. Hospital ...

  5. Serum creatine kinase and lactate dehydrogenase activities in ...

    African Journals Online (AJOL)

    ... in thyroid function are common endocrine disorders affecting 5-10% of individuals over ... Key words: Hyperthyroidism, hypothyroidism, lactate dehydrogenase, serum creatine kinase ... individuals depends on age, race, lean body mass and physical activity. ... measured by radioimmunoassay on AXSYM System (Abbott.

  6. Evaluation of Creatine Kinase Activity and Inorganic Phosphate ...

    African Journals Online (AJOL)

    Background: Biochemical parameters vary in subjects with different hemoglobin phenotypes, compared with normal controls. Aim: The aim was to evaluate serum creatine kinase (CK) activity and inorganic phosphate concentrations in Nigerian adults with homozygous and heterozygous hemoglobin phenotypes. Subjects ...

  7. 21 CFR 862.1215 - Creatine phosphokinase/creatine kinase or isoenzymes test system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Creatine phosphokinase/creatine kinase or... Clinical Chemistry Test Systems § 862.1215 Creatine phosphokinase/creatine kinase or isoenzymes test system. (a) Identification. A creatine phosphokinase/creatine kinase or isoenzymes test system is a device...

  8. Serum creatine kinase and lactate dehydrogenase activities in ...

    African Journals Online (AJOL)

    Background and Objectives: There is the recognition of a pattern of elevations of serum enzymes in hyperthyroid and hypothyroid patients. The aims of this study were to determine the activities of serum creatine kinase (CK) and lactate deydrogenase (LDH) in thyroid disorders, and to evaluate the relationship between CK, ...

  9. Exclusion of acute myocardial infarction. The value of measuring creatine kinase slope

    NARCIS (Netherlands)

    Bakker, A. J.; Koelemay, M. J.; van Vlies, B.; Gorgels, J. P.; Smits, R.; Tijssen, J. G.; Haagen, F. D.

    1995-01-01

    For the exclusion (and diagnosis) of acute myocardial infarction, we studied timed sequential (slope) measurements of creatine kinase and creatine kinase-MB catalytic activity concentration, creatine kinase-MB mass concentration, troponin T and myoglobin, using data from 242 patients consecutively

  10. Immunological measurements on the disappearance of creatine kinase MM from the circulation

    International Nuclear Information System (INIS)

    Wevers, R.A.; Landeghem, A.A.J. van; Mul-Steinbusch, M.W.F.J.; Bijdendijk, J.G.; Weerts, P.; Kloeg, P.; Soons, J.B.J.

    1983-01-01

    Both a two-site immunoradiometric assay and a two-site enzyme-linked immunosorbent assay for creatine kinase MM are described. Linearity, reproducibility and cross-reactivity of the assays are satisfactory. Creatine kinase MM incubated in a pH-controlled serum matrix loses its activity, but has its antigenic determinants affected as well. Of all the techniques used, only the immunoradiometric assay is capable of measuring part of the inactivated enzyme. Measurements with this assay on the sera of patients after a myocardial infarction show identical results for enzyme activity and creatine kinase protein quantity. The in vitro disappearance rate of creatine kinase activity is slow compared with the in vivo half-life of the enzyme. These two observations lead to the conclusion that creatine kinase is removed from the circulation long before it is inactivated in the blood stream. (Auth.)

  11. Complete inhibition of creatine kinase in isolated perfused rat hearts

    International Nuclear Information System (INIS)

    Fossel, E.T.; Hoefeler, H.

    1987-01-01

    Transient exposure of an isolated isovolumic perfused rat heart to low concentrations (0.5 mM) of perfusate-born iodoacetamide resulted in complete inhibition of creatine kinase and partial inhibition of glyceraldehyde-3-phosphate dehydrogenase in the heart. At low levels of developed pressure, hearts maintained mechanical function, ATP, and creatine phosphate levels at control values. However, iodoacetamide-inhibited hearts were unable to maintain control values of end diastolic pressure or peak systolic pressure as work load increased. Global ischemia resulted in loss of all ATP without loss of creatine phosphate, indicating lack of active creatine kinase. These results indicate that isovolumic perfused rat hearts are able to maintain normal function and normal levels of high-energy phosphates without active creatine kinase at low levels of developed pressure. 31 P-NMR of the heart was carried out

  12. Creatine kinase and creatine kinase subunit-B in coronary sinus blood in pacing-induced angina pectoris

    DEFF Research Database (Denmark)

    Bagger, J P; Ingerslev, J; Heinsvig, E M

    1982-01-01

    In nine out of 10 patients with angiographic documented coronary artery disease, pacing-induced angina pectoris provoked myocardial production of lactate, whereas no significant release of either creatine kinase or creatine kinase subunit-B to coronary sinus and peripheral venous blood could...

  13. Two-site immunoradiometric assay for the MB isoenzyme of creatine kinase

    Energy Technology Data Exchange (ETDEWEB)

    Willson, V J.C.; Jones, H M; Thompson, R J [Cambridge Univ. (UK). Clinical School

    1981-06-18

    A two-site immunoradiometric assay for myocardial creatine kinase MB isoenzyme is described. The method utilizes immobilized anti-human creatine kinase BB antibodies and /sup 125/I-labelled anti-human creatine kinase MM antibodies and can specifically detect creatine kinase MB in the presence of approximately 1000-fold excess of creatine kinase MM or BB. Native kinase MB prepared from human heart and creatine kinase MB prepared by hybridisation of purified human creatine kinase MM and creatine kinase BB appeared to react identically in the assay. Serum estimations on patients with suspected myocardial infarction correlated with the presence of MB band on electrophoresis but preliminary results suggest that the two-site immunoradiometric assay may be more sensitive.

  14. Immunological measurements on the disappearance of creatine kinase MM from the circulation. [Immunoradiometric assay

    Energy Technology Data Exchange (ETDEWEB)

    Wevers, R A; van Landeghem, A A.J.; Mul-Steinbusch, M W.F.J.; Bijdendijk, J G; Weerts, P; Kloeg, P; Soons, J B.J. [Rijksuniversiteit Utrecht (Netherlands)

    1983-07-15

    Both a two-site immunoradiometric assay and a two-site enzyme-linked immunosorbent assay for creatine kinase MM are described. Linearity, reproducibility and cross-reactivity of the assays are satisfactory. Creatine kinase MM incubated in a pH-controlled serum matrix loses its activity, but has its antigenic determinants affected as well. Of all the techniques used, only the immunoradiometric assay is capable of measuring part of the inactivated enzyme. Measurements with this assay on the sera of patients after a myocardial infarction show identical results for enzyme activity and creatine kinase protein quantity. The in vitro disappearance rate of creatine kinase activity is slow compared with the in vivo half-life of the enzyme. These two observations lead to the conclusion that creatine kinase is removed from the circulation long before it is inactivated in the blood stream.

  15. The B isozyme creatine kinase is active as a fusion protein in Escherichia coli

    International Nuclear Information System (INIS)

    Koretsky, A.P.; Traxler, B.A.

    1989-01-01

    A cDNA encoding the B isozyme of creatine kinase CK B has been expressed in Escherichia coli from a fusion with lacZ carried by λgtll. Western blots indicate that a stable polypeptide with the appropriate mobility for the Β-galactosidase-creatine kinase Β-gal-CK B ) fusion protein cross-reacts with both Β-gal and CK B antiserum. No significant CK activity is detected in control E. coli; however, extracts from cells containing the λgtll-CK B construct have a CK activity of 1.54j0.07 μmol/min per mg protein. The fusion protein appears to provide this activity bacause immunoprecipitation of protein with Β-gal antiserum leads to a loss of CK activity from extracts. That the enzyme is active in vivo was demonstrated by detection of a phosphocreatine (PCr) peak in the 31 P NMR spectrum from E. coli grown on medium supplemented with creatine. As in mammalian brain and muscle, the PCr peak detected was sensitive to the energy status of the E. coli. (author). 17 refs.; 3 figs.; 1 tab

  16. Level of hamstrings damage depending on force-generating capacity and creatine kinase activity

    OpenAIRE

    Carmona, Gerard; Alomar, Xavier; Mendiguchia, Jurdan; Serrano, David; Padullés, Josep Maria; Nescolarde Selva, Lexa Digna; Rodas Font, Gil; Cusso Calabuig, Roser; Guerrero, M.; Idoate, F.; Balius, Ramon; Cadefau, Joan

    2014-01-01

    The aim of the present study was to categorize the eccentric exercise-induced hamstrings damage by using easy measurable markers such as force-generating capacity and serum creatine kinase activity Peer Reviewed

  17. A two-site immunoradiometric assay for the MB isoenzyme of creatine kinase

    International Nuclear Information System (INIS)

    Willson, V.J.C.; Jones, H.M.; Thompson, R.J.

    1981-01-01

    A two-site immunoradiometric assay for myocardial creatine kinase MB isoenzyme is described. The method utilizes immobilized anti-human creatine kinase BB antibodies and 125 I-labelled anti-human creatine kinase MM antibodies and can specifically detect creatine kinase MB in the presence of approximately 1000-fold excess of creatine kinase MM or BB. Native kinase MB prepared from human heart and creatine kinase MB prepared by hybridisation of purified human creatine kinase MM and creatine kinase BB appeared to react identically in the assay. Serum estimations on patients with suspected myocardial infarction correlated with the presence of MB band on electrophoresis but preliminary results suggest that the two-site immunoradiometric assay may be more sensitive. (Auth.)

  18. Dependence of myosin-ATPase on structure bound creatine kinase in cardiac myfibrils from rainbow trout and freshwater turtle

    DEFF Research Database (Denmark)

    Haagensen, L.; Jensen, D.H.; Gesser, Hans

    2008-01-01

    The influence of myofibrillar creatine kinase on the myosin-ATPase activity was examined in cardiac ventricular myofibrils isolated from rainbow trout (Oncorhynchus mykiss) and freshwater turtle (Trachemys scripta). The ATPase rate was assessed by recording the rephosphorylation of ADP by the pyr......The influence of myofibrillar creatine kinase on the myosin-ATPase activity was examined in cardiac ventricular myofibrils isolated from rainbow trout (Oncorhynchus mykiss) and freshwater turtle (Trachemys scripta). The ATPase rate was assessed by recording the rephosphorylation of ADP...... by the pyruvate kinase reaction alone or together with the amount of creatine formed, when myofibrillar bound creatine kinase was activated with phosphocreatine. The steady-state concentration of ADP in the solution was varied through the activity of pyruvate kinase added to the solution. For rainbow trout...... myofibrils at a high pyruvate kinase activity, creatine kinase competed for ADP but did not influence the total ATPase activity. When the ADP concentration was elevated within the physiological range by lowering the pyruvate kinase activity, creatine kinase competed efficiently and increased the ATPase...

  19. Purification and characterization of creatine kinase isozymes from the nurse shark Ginglymostoma cirratum.

    Science.gov (United States)

    Gray, K A; Grossman, S H; Summers, D D

    1986-01-01

    Creatine kinase from nurse shark brain and muscle has been purified to apparent homogeneity. In contrast to creatine kinases from most other vertebrate species, the muscle isozyme and the brain isozyme from nurse shark migrate closely in electrophoresis and, unusually, the muscle isozyme is anodal to the brain isozyme. The isoelectric points are 5.3 and 6.2 for the muscle and brain isozymes, respectively. The purified brain preparation also contains a second active protein with pI 6.0. The amino acid content of the muscle isozyme is compared with other isozymes of creatine kinase using the Metzger Difference Index as an estimation of compositional relatedness. All comparisons show a high degree of compositional similarity including arginine kinase from lobster muscle. The muscle isozyme is marginally more resistant to temperature inactivation than the brain isozyme; the muscle protein does not exhibit unusual stability towards high concentrations of urea. Kinetic analysis of the muscle isozyme reveals Michaelis constants of 1.6 mM MgATP, 12 mM creatine, 1.2 mM MgADP and 50 mM creatine phosphate. Dissociation constants for the same substrate from the binary and ternary enzyme-substrate complex do not differ significantly, indicating limited cooperatively in substrate binding. Enzyme activity is inhibited by small planar anions, most severely by nitrate. Shark muscle creatine kinase hybridizes in vitro with rabbit muscle or monkey brain creatine kinase; shark brain isozyme hybridizes with monkey brain or rabbit brain creatine kinase. Shark muscle and shark brain isozymes, under a wide range of conditions, failed to produce a detectable hybrid.

  20. Assessment of creatine kinase and lactate dehydrogenase activities ...

    African Journals Online (AJOL)

    Ina bid to investigate the influence of menopausal on coronary heart disease, plasma creatine kinase (CK) and lactate dehydrogenase (LDH) enzymes were analysed on a prospective cohort of 100 women attending Irrua Specialist Teaching Hospital (ISTH), Irrua, Edo state-Nigeria. They were divided into two groups; ...

  1. Histochemical demonstration of creatine kinase activity using polyvinyl alcohol and auxiliary enzymes

    NARCIS (Netherlands)

    Frederiks, W. M.; Marx, F.; van Noorden, C. J.

    1987-01-01

    Creatine kinase activity (EC 2.7.3.2.) has been demonstrated in myocardium and skeletal muscle from rats by a method based on the incubation of cryostat sections with a polyvinyl alcohol-containing medium and the use of auxiliary enzymes. Hexokinase and glucose-6-phosphate dehydrogenase were spread

  2. Proteinase K processing of rabbit muscle creatine kinase

    DEFF Research Database (Denmark)

    Leydier, C; Andersen, Jens S.; Couthon, F

    1997-01-01

    Proteinase K cleaves selectively both cytosolic and mitochondrial isoforms of creatine kinase leading to the appearance of two fragments, a large N-terminal one (K1) and a small C-terminal peptide (K2) which remain associated together. The loss of enzymatic activity correlates with the extent...... of monomer cleavage. N-terminal sequencing of the K2 fragments from rabbit cytosolic and pig mitochondrial creatine kinase shows that these peptides begin with A328 and A324, respectively. Electrospray ionization mass spectrometry demonstrates that K2 peptide is composed of 53 residues (A328-K380). However...

  3. Creatine supplementation: effects on blood creatine kinase activity responses to resistance exercise and creatine kinase activity measurement

    Directory of Open Access Journals (Sweden)

    Marco Machado

    2009-12-01

    Full Text Available The purpose of this study was to determine the effects of creatine supplementation and exercise on the integrity of muscle fiber, as well as the effect of the supplementation on the creatine kinase (CK assay measurement. Forty-nine sedentary individuals participated in a double-blind study and were divided into two groups: C (n=26 received 4x5-day packages of 0.6 g.kg-1 of body weight contained 50% of creatine + 50% of dextrose, and P (n=23 received packages containing only dextrose. On the first day the groups performed a 1RM test for bench press, seated row, leg extension, leg curl and leg press. On D7 they received the supplements. On the fourteenth day, they performed a training session of five exercises, each in three sets of ten repetitions at 75% of 1RM. Blood was collected before (D14 and after the exercise session (D15. Differing levels of blood creatine were tested to determine the influence on the assay measurements of CK. ANOVA and Tukey's post-hoc tests were used to compare groups and different times of study protocol (PO objetivo do presente estudo foi determinar o efeito da suplementação de creatina e do exercício na integridade das fibras musculares e, também, o efeito da suplementação na técnica de mensuração da atividade da creatina kinase (CK. Quarenta e nove sedentários participaram de um estudo duplo-cego e foram divididos em dois grupos: C (n=26 que receberam 4x5 dias embalagens com 0,6 g.kg-1 de massa corporal com 50% de creatina + 50% de dextrose, e P (n=23 que receberam embalagens contendo apenas dextrose. No primeiro dia, eles realizaram o teste de 1RM para os exercícios supino reto, remada sentada, cadeira extensora, mesa flexora, e leg press. No D7 receberam os suplementos. No décimo quarto dia eles realizaram uma sessão de treinos com os cinco exercícios, cada um com 3x10 repetições a 75% de 1RM. Sangue foi coletado antes (D14 e depois da sessão de exercícios (D15. Diferentes concentrações de

  4. Quantitative estimation of infarct size by simultaneous dual radionuclide single photon emission computed tomography: comparison with peak serum creatine kinase activity

    International Nuclear Information System (INIS)

    Kawaguchi, K.; Sone, T.; Tsuboi, H.; Sassa, H.; Okumura, K.; Hashimoto, H.; Ito, T.; Satake, T.

    1991-01-01

    To test the hypothesis that simultaneous dual energy single photon emission computed tomography (SPECT) with technetium-99m (99mTc) pyrophosphate and thallium-201 (201TI) can provide an accurate estimate of the size of myocardial infarction and to assess the correlation between infarct size and peak serum creatine kinase activity, 165 patients with acute myocardial infarction underwent SPECT 3.2 +/- 1.3 (SD) days after the onset of acute myocardial infarction. In the present study, the difference in the intensity of 99mTc-pyrophosphate accumulation was assumed to be attributable to difference in the volume of infarcted myocardium, and the infarct volume was corrected by the ratio of the myocardial activity to the osseous activity to quantify the intensity of 99mTc-pyrophosphate accumulation. The correlation of measured infarct volume with peak serum creatine kinase activity was significant (r = 0.60, p less than 0.01). There was also a significant linear correlation between the corrected infarct volume and peak serum creatine kinase activity (r = 0.71, p less than 0.01). Subgroup analysis showed a high correlation between corrected volume and peak creatine kinase activity in patients with anterior infarctions (r = 0.75, p less than 0.01) but a poor correlation in patients with inferior or posterior infarctions (r = 0.50, p less than 0.01). In both the early reperfusion and the no reperfusion groups, a good correlation was found between corrected infarct volume and peak serum creatine kinase activity (r = 0.76 and r = 0.76, respectively; p less than 0.01)

  5. Serum creatine kinase isoenzymes in children with osteogenesis imperfecta.

    Science.gov (United States)

    D'Eufemia, P; Finocchiaro, R; Zambrano, A; Lodato, V; Celli, L; Finocchiaro, S; Persiani, P; Turchetti, A; Celli, M

    2017-01-01

    This study evaluates serum creatine kinase isoenzyme activity in children with osteogenesis imperfecta to determine its usefulness as a biochemical marker during treatment with bisphosphonate. The changes of creatine kinase (CK) isoenzyme activity during and after discontinuation therapy were observed. These results could be useful in addressing over-treatment risk prevention. The brain isoenzyme of creatine kinase (CKbb) is highly expressed in mature osteoclasts during osteoclastogenesis, thus plays an important role in bone resorption. We previously identified high serum CKbb levels in 18 children with osteogenesis imperfect (OI) type 1 treated for 1 year with bisphosphonate (neridronate). In the present study, serum CK isoenzymes were evaluated in the same children with continuous versus discontinued neridronate treatment over a further 2-year follow-up period. This study included 18 children with OI type 1, 12 with continued (group A) and 6 with ceased (group B) neridronate treatment. Auxological data, serum biochemical markers of bone metabolism, bone mineral density z-score, and serum total CK and isoenzyme activities were determined in both groups. Serum CKbb was progressively and significantly increased in group A (p < 0.004) but rapidly decreased to undetectable levels in group B. In both groups, the cardiac muscle creatine kinase isoenzyme (CKmb) showed a marked decrease, while serum C-terminal telopeptide (CTx) levels were almost unchanged. This study provides evidence of the cumulative effect of neridronate administration in increasing serum CKbb levels and the reversible effect after its discontinuation. This approach could be employed for verifying the usefulness of serum CKbb as a biochemical marker in patients receiving prolonged bisphosphonate treatment. Moreover, the decreased serum CKmb levels suggest a systemic effect of these drugs.

  6. Creatine kinase isozyme expression in embryonic chicken heart

    NARCIS (Netherlands)

    Lamers, W. H.; Geerts, W. J.; Moorman, A. F.; Dottin, R. P.

    1989-01-01

    The distribution pattern of creatine kinase (EC 2.7.3.2) isozymes in developing chicken heart was studied by immunohistochemistry. Creatine kinase M, which is absent from adult heart, is transiently expressed between 4 and 11 days of incubation. During that period, numerous muscular cells in the

  7. Boehringer immunoinhibition procedure for creatine kinase-MB evaluated and compared with column ion-exchange chromatography

    NARCIS (Netherlands)

    ter Welle, H. F.; Baartscheer, T.; Fiolet, J. W.

    1983-01-01

    In determination of creatine kinase isoenzyme MB (CK-MB), the Boehringer immunoinhibition method gives a high and variable blank activity as compared with column-chromatography. Thus a correction must be applied. Furthermore, a second correction of 1% of total creatine kinase activity is necessary

  8. Improved radioimmunoassay for creatine kinase isoenzymes in plasma

    International Nuclear Information System (INIS)

    Ritter, C.S.; Mumm, S.R.; Roberts, R.

    1981-01-01

    We describe convenient and relatively rapid procedures for purifying creatine kinase isoenzymes MM, BB, and MB, and their use in an improved radioimmunoassay for creatine kinase isoenzymes in plasma. The modifications include use of: (a) BB with a specific activity of 400 kU/G, which can be labeled with a specific radioactivity of 20 Ci/g; (b) albumin-free purified MB as inhibitor; (c) antiserum to MB creatine kinase; and (d) a second-antibody technique that necessitates only a 15-min incubation. The radioimmunoassay for MB has a sensitivity of 0.2 μg/L (80 mU/L) and a CV of <5%. Plasma MB average 22 (SD 12) μg/L in 200 normal subjects; 24 (SD 12) μg/L in 200 patients with chest pain without infarction; and 23 (SD 7) μg/L in 43 patients with renal disease, whether measured before or after dialysis. Peak values for plasma MB averaged 191 (SD 86) μg/L in 325 patients with documented myocardial infarction; BB was negligible. Extensive clinical experience indicates the radioimmunoassay to be suitably rapid, highly sensitive, and reliable as a diagnostic assay for MB on plasma

  9. Creatine Kinase Activity Weakly Correlates to Volume Completed Following Upper Body Resistance Exercise

    Science.gov (United States)

    Machado, Marco; Willardson, Jeffrey M.; Silva, Dailson P.; Frigulha, Italo C.; Koch, Alexander J.; Souza, Sergio C.

    2012-01-01

    In the current study, we examined the relationship between serum creatine kinase (CK) activity following upper body resistance exercise with a 1- or 3-min rest between sets. Twenty men performed two sessions, each consisting of four sets with a 10-repetition maximum load. The results demonstrated significantly greater volume for the 3-min…

  10. Activation of sea urchin sperm motility is accompanied by an increase in the creatine kinase exchange flux

    NARCIS (Netherlands)

    Dorsten, van F.A.; Wyss, M.; Wallimann, T.; Nicolaij, K.

    1997-01-01

    The kinetics of the creatine kinase (CK) reaction were studied in suspensions of quiescent and active, intact sea-urchin spermatozoa in artificial seawater, using 31P-NMR magnetization transfer. In inactive sperm, no CK-mediated exchange flux was detected, whereas in activated motile sperm, the

  11. Hypoxia decreases creatine uptake in cardiomyocytes, while creatine supplementation enhances HIF activation.

    Science.gov (United States)

    Santacruz, Lucia; Arciniegas, Antonio Jose Luis; Darrabie, Marcus; Mantilla, Jose G; Baron, Rebecca M; Bowles, Dawn E; Mishra, Rajashree; Jacobs, Danny O

    2017-08-01

    Creatine (Cr), phosphocreatine (PCr), and creatine kinases (CK) comprise an energy shuttle linking ATP production in mitochondria with cellular consumption sites. Myocytes cannot synthesize Cr: these cells depend on uptake across the cell membrane by a specialized creatine transporter (CrT) to maintain intracellular Cr levels. Hypoxia interferes with energy metabolism, including the activity of the creatine energy shuttle, and therefore affects intracellular ATP and PCr levels. Here, we report that exposing cultured cardiomyocytes to low oxygen levels rapidly diminishes Cr transport by decreasing V max and K m Pharmacological activation of AMP-activated kinase (AMPK) abrogated the reduction in Cr transport caused by hypoxia. Cr supplementation increases ATP and PCr content in cardiomyocytes subjected to hypoxia, while also significantly augmenting the cellular adaptive response to hypoxia mediated by HIF-1 activation. Our results indicate that: (1) hypoxia reduces Cr transport in cardiomyocytes in culture, (2) the cytoprotective effects of Cr supplementation are related to enhanced adaptive physiological responses to hypoxia mediated by HIF-1, and (3) Cr supplementation increases the cellular ATP and PCr content in RNCMs exposed to hypoxia. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

  12. Structural Changes of Creatine Kinase upon Substrate Binding

    OpenAIRE

    Forstner, Michael; Kriechbaum, Manfred; Laggner, Peter; Wallimann, Theo

    1998-01-01

    Small-angle x-ray scattering was used to investigate structural changes upon binding of individual substrates or a transition state analog complex (TSAC; Mg-ADP, creatine, and KNO3) to creatine kinase (CK) isoenzymes (dimeric muscle-type (M)-CK and octameric mitochondrial (Mi)-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occurred upon binding of Mg-nucleotide or TSAC. The radius of gyration of Mi-CK was reduced from 55.6 A (free enzym...

  13. The Influence of Red Fruit Oil on Creatin Kinase Level at Maximum Physical Activity

    Science.gov (United States)

    Apollo Sinaga, Fajar; Hotliber Purba, Pangondian

    2018-03-01

    Heavy physical activities can cause the oxidative stress which resulting in muscle damage with an indicator of elevated levels of Creatin Kinase (CK) enzyme. The oxidative stress can be prevented or reduced by antioxidant supplementation. One of natural resources which contain antioxidant is Red Fruit (Pandanus conoideus) Oil (RFO). This study aims to see the effect of Red Fruit Oil on Creatin Kinase (CK) level at maximum physical activity. This study is an experimental research by using the design of randomized control group pretest-posttest. This study was using 24 male mice divided into four groups, the control group was given aquadest, the treatment groups P1, P2, and P3 were given the RFO orally of 0.15 ml/kgBW, 0.3 ml/kgBW, and 0.6 ml/kgBW, respectively, for a month. The level of CK was checked for all groups at the beginning of study and after the maximum physical activity. The obtained data were then tested statistically by using t-test and ANOVA. The result shows the RFO supplementation during exercise decreased the CK level in P1, P2, and P3 groups with p<0.05, and the higher RFO dosage resulted in decreased CK level at p<0.05. The conclusion of this study is the Red Fruit Oil could decrease the level of CK at maximum physical activity.

  14. Radioimmunoassay of inactive creatine kinase B protein in human plasma

    Energy Technology Data Exchange (ETDEWEB)

    Burnam, M H; Shell, W E [California Univ., Los Angeles (USA). School of Medicine

    1981-08-27

    The authors describe a rapid, sensitive radioimmunoassay for enzymatically inactive creatine kinase B protein (CK-Bi) in plasma. /sup 125/I-CK-Bi of high specific activity and good stability was prepared by oxidant-based iodination. A 12-minute first antibody incubation was used. Bound and free antigen were separated by a second antibody system. Large excesses of purified CK-MM from human skeletal muscle did not react in the assay. Cross reactivity to CK-MB purified from the plasma of patients with acute myocardial infarction was negligible. The 95th percentile of plasma CK-Bi in 150 adults was 145 ..mu..g equivalents/ml. Within-assay and between-assay precision ranged from 5% to 9% and 6% to 10%, respectively. Evidence is presented indicating that the assay measures inactive creatine kinase B protein, a protein not measured by current assay systems dependent on biological activity.

  15. Radioimmunoassay of inactive creatine kinase B protein in human plasma

    International Nuclear Information System (INIS)

    Burnam, M.H.; Shell, W.E.

    1981-01-01

    The authors describe a rapid, sensitive radioimmunoassay for enzymatically inactive creatine kinase B protein (CK-Bi) in plasma. 125 I-CK-Bi of high specific activity and good stability was prepared by oxidant-based iodination. A 12-minute first antibody incubation was used. Bound and free antigen were separated by a second antibody system. Large excesses of purified CK-MM from human skeletal muscle did not react in the assay. Cross reactivity to CK-MB purified from the plasma of patients with acute myocardial infarction was negligible. The 95th percentile of plasma CK-Bi in 150 adults was 145 μg equivalents/ml. Within-assay and between-assay precision ranged from 5% to 9% and 6% to 10%, respectively. Evidence is presented indicating that the assay measures inactive creatine kinase B protein, a protein not measured by current assay systems dependent on biological activity. (Auth.)

  16. The Influence of Whole-Body Vibration on Creatine Kinase Activity and Jumping Performance in Young Basketball Players

    Science.gov (United States)

    Fachina, Rafael; da Silva, Antônio; Falcão, William; Montagner, Paulo; Borin, João; Minozzo, Fábio; Falcão, Diego; Vancini, Rodrigo; Poston, Brach; de Lira, Claudio

    2013-01-01

    Purpose: To quantify creatine kinase (CK) activity changes across time following an acute bout of whole-body vibration (WBV) and determine the association between changes in CK activity and jumping performance. Method: Twenty-six elite young basketball players were assigned to 3 groups: 36-Hz and 46-Hz vibration groups (G36 and G46, respectively)…

  17. Determination of total creatine kinase activity in blood serum using an amperometric biosensor based on glucose oxidase and hexokinase.

    Science.gov (United States)

    Kucherenko, I S; Soldatkin, O O; Lagarde, F; Jaffrezic-Renault, N; Dzyadevych, S V; Soldatkin, A P

    2015-11-01

    Creatine kinase (CK: adenosine-5-triphosphate-creatine phosphotransferase) is an important enzyme of muscle cells; the presence of a large amount of the enzyme in blood serum is a biomarker of muscular injuries, such as acute myocardial infarction. This work describes a bi-enzyme (glucose oxidase and hexokinase based) biosensor for rapid and convenient determination of CK activity by measuring the rate of ATP production by this enzyme. Simultaneously the biosensor determines glucose concentration in the sample. Platinum disk electrodes were used as amperometric transducers. Glucose oxidase and hexokinase were co-immobilized via cross-linking with BSA by glutaraldehyde and served as a biorecognition element of the biosensor. The biosensor work at different concentrations of CK substrates (ADP and creatine phosphate) was investigated; optimal concentration of ADP was 1mM, and creatine phosphate - 10 mM. The reproducibility of the biosensor responses to glucose, ATP and CK during a day was tested (relative standard deviation of 15 responses to glucose was 2%, to ATP - 6%, to CK - 7-18% depending on concentration of the CK). Total time of CK analysis was 10 min. The measurements of creatine kinase in blood serum samples were carried out (at 20-fold sample dilution). Twentyfold dilution of serum samples was chosen as optimal for CK determination. The biosensor could distinguish healthy and ill people and evaluate the level of CK increase. Thus, the biosensor can be used as a test-system for CK analysis in blood serum or serve as a component of multibiosensors for determination of important blood substances. Determination of activity of other kinases by the developed biosensor is also possible for research purposes. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. AMP kinase expression and activity in human skeletal muscle: effects of immobilization, retraining, and creatine supplementation

    DEFF Research Database (Denmark)

    Eijnde, Bert O.; Derave, Wim; Wojtaszewski, Jørgen

    2005-01-01

    The effects of leg immobilization and retraining in combination with oral creatine intake on muscle AMP-activated protein kinase (AMPK) protein expression and phosphorylation status were investigated. A double-blind trial was performed in young healthy volunteers (n = 22). A cast immobilized...... the right leg for 2 wk, whereafter the knee-extensor muscles of that leg were retrained for 6 wk. Half of the subjects received creatine monohydrate throughout the study (Cr; from 15 g down to 2.5 g daily), and the others ingested placebo (P; maltodextrin). Before and after immobilization and retraining...... that immobilization-induced muscle inactivity for 2 wk does not alter AMPK a1-, a2-, and ß2-subunit expression or a-AMPK phosphorylation status. Furthermore, the present observations indicate that AMPK probably is not implicated in the previously reported beneficial effects of oral creatine supplementation on muscle...

  19. Radiation inactivation method provides evidence that membrane-bound mitochondrial creatine kinase is an oligomer

    International Nuclear Information System (INIS)

    Quemeneur, E.; Eichenberger, D.; Goldschmidt, D.; Vial, C.; Beauregard, G.; Potier, M.

    1988-01-01

    Lyophilized suspensions of rabbit heart mitochondria have been irradiated with varying doses of gamma rays. Mitochondrial creatine kinase activity was inactivated exponentially with a radiation inactivation size of 352 or 377 kDa depending upon the initial medium. These values are in good agreement with the molecular mass previously deduced from by permeation experiments: 357 kDa. This is the first direct evidence showing that the native form of mitochondrial creatine kinase is associated to the inner membrane as an oligomer, very likely an octamer

  20. Creatine Kinase Activity in Patients with Diabetes Mellitus Type I and Type II

    Directory of Open Access Journals (Sweden)

    Adlija Jevrić-Čaušević

    2006-08-01

    Full Text Available Diabetes mellitus can be looked upon as an array of diseases, all of which exhibit common symptoms. While pathogenesis of IDDM (insulin dependant diabetes mellitus is well understood, the same is not true for diabetes mellitus type II. In the latter case, relative contribution of the two factors (insulin resistance or decreased insulin secretion varies individually, being highly increased in peripheral tissues and strictly dependant on insulin for glucose uptake. Moreover, in patients with diabetes mellitus type II, disbalance at the level of regulation of glucose metabolism as well as lipid metabolism has been noted in skeletal muscles. It is normal to assume that in this type of diabetes, these changes are reflected at the level of total activity of enzyme creatine kinase. This experimental work was performed on a group of 80 regular patients of Sarajevo General Hospital. Forty of those patients were classified as patients with diabetes type I and forty as patients with diabetes type II. Each group of patients was carefully chosen and constituted of equal number of males and females. The same was applied for adequate controls. Concentration of glucose was determined for each patient with GOD method, while activity of creatine kinase was determined with CK-NAC activated kit. Statistical analysis of the results was performed with SPSS software for Windows. Obtained results point out highly expressed differences in enzyme activity between two populations examined. Changes in enzyme activity are more expressed in patients with diabetes type II. Positive correlation between concentration of glucose and serum activity of the enzyme is seen in both categories of diabetic patients which is not the case for the patients in control group. At the same time, correlation between age and type of diabetes does exist . This is not followed at the level of enzyme activity or concentration of glucose.

  1. Radioimmunoassay of creatine kinase BB isoenzyme

    International Nuclear Information System (INIS)

    Geng Jianguo

    1988-01-01

    A radioimmunoassay of creatine kinase BB isoenzyme (CK-BB) was developed by using CK-BB purified from human brain. The CK-BB antiserum was raised by immunizing rabbite and 125 I-CK-BB iodinated with Bolton-Hunter reagent. The affinity constant was 3.0 x 10 9 mol/L. No cross reactions with creatine kinase MM isoenzyme and neuron-specific enolase were found. The measuring range was 3.5 x 10 -8 ∼ 1.2 x 10 -5 mmol/L, the average recovery rate 97.5%, with the inter and intrassay CV 3.1% and 12%, respectively. The average serum CK-BB concentration in 83 normal persons was 1.5 x 10 -7 +- 8.1 x 10 -8 mmol/L, quite different from the values of acute myocardial infarction (5.2 x 10 -6 +- 1.2 x 10 -4 mmol/L, n = 28) and cerebral vascular accident (8.4 x 10 -4 +- 5.0 x 10 -4 mmol/L, n = 10)

  2. Radioimmunoassay measurement of creatine kinase BB in the serum of schizophrenic patients

    Energy Technology Data Exchange (ETDEWEB)

    Lerner, M H; Friedhoff, A J [New York Univ., NY (USA). Medical Center

    1980-10-23

    Brain type creatine kinase (BB) isoenzyme was measured using a highly sensitive and specific radioimmunoassay procedure (limit of detection, 1 ..mu..g/l of sample) in two schizophrenic populations, an acute non-medicated group consisting of 35 subjects and a chronic group of 15 subjects. Since the assay can also measure the B subunit of MB isoenzyme, patients were selected so as to exclude subjects with possible heart, kidney or other ailments which might result in an increased serum creatine kinase B subunit. Both the acute schizophrenics (3.0 +- 0.23) x S.E.M. and the chronic schizophrenics (2.9 +- 0.33) had serum levels of creatine kinase BB similar to those of controls (2.8 +- 0.21) and non-cardiac patients (3.5 +- 0.58). Patients having myocardial infarction or neurovascular surgery had elevated creatine kinase B subunit. Similar but much less sensitive quantitative results were obtained using agarose multizonal electrophoresis.

  3. Radioimmunoassay measurement of creatine kinase BB in the serum of schizophrenic patients

    International Nuclear Information System (INIS)

    Lerner, M.H.; Friedhoff, A.J.

    1980-01-01

    Brain type creatine kinase (BB) isoenzyme was measured using a highly sensitive and specific radioimmunoassay procedure (limit of detection, 1 μg/l of sample) in two schizophrenic populations, an acute non-medicated group consisting of 35 subjects and a chronic group of 15 subjects. Since the assay can also measure the B subunit of MB isoenzyme, patients were selected so as to exclude subjects with possible heart, kidney or other ailments which might result in an increased serum creatine kinase B subunit. Both the acute schizophrenics (3.0 +- 0.23) x S.E.M. and the chronic schizophrenics (2.9 +- 0.33) had serum levels of creatine kinase BB similar to those of controls (2.8 +- 0.21) and non-cardiac patients (3.5 +- 0.58). Patients having myocardial infarction or neurovascular surgery had elevated creatine kinase B subunit. Similar but much less sensitive quantitative results were obtained using agarose multizonal electrophoresis. (Auth.)

  4. Creatine kinase response to high-intensity aerobic exercise in adult-onset muscular dystrophy

    DEFF Research Database (Denmark)

    Andersen, Søren P; Sveen, Marie-Louise; Hansen, Regitze S

    2013-01-01

    We investigated the effect of high-intensity exercise on plasma creatine kinase (CK) in patients with muscular dystrophies.......We investigated the effect of high-intensity exercise on plasma creatine kinase (CK) in patients with muscular dystrophies....

  5. Hydrogen peroxide (H2O2) irreversibly inactivates creatine kinase from Pelodiscus sinensis by targeting the active site cysteine.

    Science.gov (United States)

    Wang, Wei; Lee, Jinhyuk; Hao, Hao; Park, Yong-Doo; Qian, Guo-Ying

    2017-12-01

    Creatine kinase (EC 2.7.3.2, CK) plays an important role in cellular energy metabolism and homeostasis by catalysing the transfer of phosphate between ATP and creatine phosphate. In this study, we investigated the effects of H 2 O 2 on PSCKM (muscle type creatine kinase from Pelodiscus sinensis) by the integrating method between enzyme kinetics and docking simulations. We found that H 2 O 2 strongly inactivated PSCKM (IC 50 =0.25mM) in a first-order kinetic process, and targeted the active site cysteine directly. A conformational study showed that H 2 O 2 did not induce the tertiary structural changes in PSCKM with no extensive exposure of hydrophobic surfaces. Sequential docking simulations between PSCKM and H 2 O 2 indicated that H 2 O 2 interacts with the ADP binding region of the active site, consistent with experimental results that demonstrated H 2 O 2 -induced inactivation. Our study demonstrates the effect of H 2 O 2 on PSCKM enzymatic function and unfolding, and provides important insight into the changes undergone by this central metabolic enzyme in ectothermic animals in response to the environment. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Diagnostic value of creatine kinase activity in canine cerebrospinal fluid.

    Science.gov (United States)

    Ferreira, Alexandra

    2016-10-01

    This study aimed to determine whether creatine kinase (CK) activity in cerebrospinal fluid (CSF) has diagnostic value for various groups of neurological conditions or for different anatomical areas of the nervous system (NS). The age, breed, results of CSF analysis, and diagnosis of 578 canine patients presenting with various neurological conditions between January 2009 and February 2015 were retrospectively collected. The cases were divided according to anatomical areas of the nervous system, i.e., brain, spinal cord, and peripheral nervous system, and into groups according to the nature of the condition diagnosed: vascular, immune/inflammatory/infectious, traumatic, toxic, anomalous, metabolic, idiopathic, neoplastic, and degenerative. Statistical analysis showed that CSF-CK alone cannot be used as a diagnostic tool and that total proteins in the CSF and red blood cells (RBCs) do not have a significant relationship with the CSF-CK activity. CSF-CK did not have a diagnostic value for different disease groups or anatomical areas of the nervous system.

  7. Influence of noise and vibration upon creatine kinase activity in blood serum

    Energy Technology Data Exchange (ETDEWEB)

    Jonderko, G; Gabryel, A; Jonderko, K; Konca, A; Marcisz, C; Olak, Z; Szramek-Urbaniak, A

    1982-02-01

    The investigations comprised part of the workers employed in a factory of prefabricated concrete elements. The creatine kinase (CK) activity was determined before and after a day's work in the following groups of workers: I - exposed to noise and vibration exceeding the allowed norm, II - exposed to noise, performing the same hard physical work as group I and exposed to vibration not exceeding 79 dB in particular octave bands, III - practising physical work as group I and II but in environment free from noise and vibration, and a control group of persons not exposed to any of the tested factors. The obtained results indicate that the determination of the CK activity may serve as a test of exposure to vibration and noise.

  8. Radioimmunoassay of creatine kinase BB isoenzyme

    Energy Technology Data Exchange (ETDEWEB)

    Jianguo, Geng [Shanghai Medical Univ. (China). Zhongshan Hospital; and others

    1988-11-01

    A radioimmunoassay of creatine kinase BB isoenzyme (CK-BB) was developed by using CK-BB purified from human brain. The CK-BB antiserum was raised by immunizing rabbite and {sup 125}I-CK-BB iodinated with Bolton-Hunter reagent. The affinity constant was 3.0 x 10{sup 9} mol/L. No cross reactions with creatine kinase MM isoenzyme and neuron-specific enolase were found. The measuring range was 3.5 x 10{sup -8} {approx} 1.2 x 10{sup -5} mmol/L, the average recovery rate 97.5%, with the inter and intrassay CV 3.1% and 12%, respectively. The average serum CK-BB concentration in 83 normal persons was 1.5 x 10{sup -7} +- 8.1 x 10{sup -8} mmol/L, quite different from the values of acute myocardial infarction (5.2 x 10{sup -6} +- 1.2 x 10{sup -4} mmol/L, n = 28) and cerebral vascular accident (8.4 x 10{sup -4} +- 5.0 x 10{sup -4} mmol/L, n = 10).

  9. Variations in creatine kinase activity and reactive oxygen species levels are involved in capacitation of bovine spermatozoa.

    Science.gov (United States)

    Córdoba, M; Pintos, L; Beconi, M T

    2008-12-01

    The generation of reactive oxygen species (ROS) is associated with some factors such as oxidative substrate sources, mitochondrial function and NAD(P)H oxidase activity. In bovine spermatozoa, heparin capacitation produces a respiratory burst sensitive to diphenyleneiodonium (DPI). Creatine kinase (CK) is related to extramitochondrial ATP disponibility. Our purpose was to determine the variation in ROS level and its relation with NAD(P)H oxidase sensitive to DPI and CK participation, as factors involved in redox state and energy generation in capacitation. The chlortetracycline technique was used to evaluate capacitation. CK activity and ROS level were measured by spectrophotometry and spectrofluorometry respectively. The capacitation percentage was increased by heparin or quercetin treatment (P level as control (238.62 +/- 23.47 arbitrary units per 10(8) spermatozoa) (P > 0.05). CK activity decreased by 50% with heparin or quercetin (P level variations were observed in heparin- or quercetin-treated samples (P bovine spermatozoa, capacitation requires equilibrium between oxidative damage susceptibility and ROS levels. CK activity is associated with redox state variation and energy sources. In conclusion, capacitation induction depends on NADPH oxidase and the shuttle creatine-creatine phosphate, both sensitive to DPI.

  10. Rate equation for creatine kinase predicts the in vivo reaction velocity: 31P NMR surface coil studies in brain, heart, and skeletal muscle of the living rat

    International Nuclear Information System (INIS)

    Bittl, J.A.; DeLayre, J.; Ingwall, J.S.

    1987-01-01

    Brain, heart, and skeletal muscle contain four different creatine kinase isozymes and various concentrations of substrates for the creatine kinase reaction. To identify if the velocity of the creatine kinase reaction under cellular conditions is regulated by enzyme activity and substrate concentrations as predicted by the rate equation, the authors used 31 P NMR and spectrophotometric techniques to measure reaction velocity, enzyme content, isozyme distribution, and concentrations of substrates in brain, heart, and skeletal muscle of living rat under basal or resting conditions. The total tissue activity of creatine kinase in the direction of MgATP synthesis provided an estimate for V/sub max/ and exceeded the NMR-determined in vivo reaction velocities by an order of magnitude. The isozyme composition varied among the three tissues: >99% BB for brain; 14% MB, 61% MM, and 25% mitochondrial for heart; and 98% MM and 2% mitochondrial for skeletal muscle. The NMR-determined reaction velocities agreed with predicted values from the creatine kinase rate equation. The concentrations of free creatine and cytosolic MgADP, being less than or equal to the dissociation constants for each isozyme, were dominant terms in the creatine kinase rate equation for predicting the in vivo reaction velocity. Thus, they observed that the velocity of the creatine kinase reaction is regulated by total tissue enzyme activity and by the concentrations of creatine and MgADP in a manner that is independent of isozyme distribution

  11. The influence of noise and vibration upon creatine kinase activity in blood serum

    Energy Technology Data Exchange (ETDEWEB)

    Jonderko, G; Gabryel, A; Jonderko, K; Konca, A; Marcisz, C; Olak, Z; Szramek-Urbaniak, A

    1982-02-01

    The investigations comprised part of the workers employed in a factory of prefabricated concrete elements. The creatine kinase (CK) activity was determined before and after a day's work in the following groups of workers: These exposed to noise and vibration exceeding the allowed norm; those exposed to noise, performing the same hard physical work as group I and exposed to vibration not exceeding 79dB in particular octave bands, and those practising physical work a group I and II but in environment free from noise and vibration, and a control group of persons not exposed to any of the tested factors (vibration, noise, work). The obtained results indicate that the determination of the CK activity may serve as a test of exposure to vibration and noise.

  12. Frequency-domain lifetime fluorometry of double-labeled creatine kinase.

    Science.gov (United States)

    Gregor, M; Kubala, M; Amler, E; Mejsnar, J

    2003-01-01

    Myofibril-bound creatine kinase EC 2.7.3.2 (CK), a key enzyme of muscle energy metabolism, has been selected for studies of conformational changes that underlie the cellular control of enzyme activity. For fluorescence spectroscopy measurements, the CK molecule was double-labeled with IAF (5-iodoacetamidofluorescein) and ErITC (erythrosin 5'-isothiocyanate). Measurement of fluorescence resonance energy transfer (FRET) from fluorescein to erythrosin was used to obtain information about the donor-acceptor pair distance. Frequency-domain lifetime measurements evaluate the donor-acceptor distance in the native CK molecule as 7.8 nm. The Förster radius equals 5.3 nm with the resolution range from 0.2 to 1.0 nm. Erythrosin-fluorescein labeling (EFL) was tested for artificial conformational changes of the CK molecule with high-salt concentration treatment. The transition distance, defined by His-97 and Cys-283 and derived from a 3D model equals 0.766 nm for the open (inactive) form and 0.277 nm for the closed (reactive) form of the CK molecule. In this way, the resolution range of the used spectroscopy method is significant, concerning the difference of 0.489 nm. Nevertheless, the CK enzyme activity, assessed by the hexokinase-coupled assay, was diminished down to 1 % of the activity of the native enzyme. EFL is suitable for description of conformational behavior implied from the regulation of creatine kinase. However, the observed inhibition restricts EFL to studies of conformational changes during natural catalytic activity.

  13. Aeromonas caviae alters the cytosolic and mitochondrial creatine kinase activities in experimentally infected silver catfish: Impairment on renal bioenergetics.

    Science.gov (United States)

    Baldissera, Matheus D; Souza, Carine F; Júnior, Guerino B; Verdi, Camila Marina; Moreira, Karen L S; da Rocha, Maria Izabel U M; da Veiga, Marcelo L; Santos, Roberto C V; Vizzotto, Bruno S; Baldisserotto, Bernardo

    2017-09-01

    Cytosolic and mitochondrial creatine kinases (CK), through the creatine kinase-phosphocreatine (CK/PCr) system, provide a temporal and spatial energy buffer to maintain cellular energy homeostasis. However, the effects of bacterial infections on the kidney remain poorly understood and are limited only to histopathological analyses. Thus, the aim of this study was to investigate the involvement of cytosolic and mitochondrial CK activities in renal energetic homeostasis in silver catfish experimentally infected with Aeromonas caviae. Cytosolic CK activity decreased in infected animals, while mitochondrial CK activity increased compared to uninfected animals. Moreover, the activity of the sodium-potassium pump (Na + , K + -ATPase) decreased in infected animals compared to uninfected animals. Based on this evidence, it can be concluded that the inhibition of cytosolic CK activity by A. caviae causes an impairment on renal energy homeostasis through the depletion of adenosine triphosphate (ATP) levels. This contributes to the inhibition of Na + , K + -ATPase activity, although the mitochondrial CK activity acted in an attempt to restore the cytosolic ATP levels through a feedback mechanism. In summary, A. caviae infection causes a severe energetic imbalance in infected silver catfish, which may contribute to disease pathogenesis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Effects of combined irradiation and hyperthermia on creatine kinase activity in rat tissues

    Energy Technology Data Exchange (ETDEWEB)

    Matyushichev, V B; Taratukhin, V P; Shamratova, V G; Yuzhakova, G A [Leningradskij Gosudarstvennyj Univ. (USSR)

    1977-09-01

    The effects of different doses of beta- and X-irradiations on creatine kinase activity (CKA) in cerebrum and liver of rats were investigated under hyperthermia conditions. Beta irradiation was carried out at 2.50, 3.05, 4.45 or 7.40 Krad doses under heat load. After withdrawal of the animals from the chamber they were exposed to 25,50,100,250 or 400 R X-ray doses (for beta irradiation 2.50,2.50,3.05,4.45 or 7.40 Krad, respectively). In 5,12,19 and 26 days after exposure the animals were decapitated and CKA of aqueous extracts of cerebral hemispheres and livers were determined. It has been established, that the thermal stress decreases the radiosensitivity of cerebral CKA, but intensifies the irradiation consequences in respect to liver activity.

  15. Contraction-mediated glycogenolysis in mouse skeletal muscle lacking creatine kinase: the role of phosphorylase b activation.

    NARCIS (Netherlands)

    Katz, A.; Andersson, D.C.; Yu, J.; Norman, B.; Sandstrom, M.E.; Wieringa, B.; Westerblad, H.

    2003-01-01

    Skeletal muscle that is deficient in creatine kinase (CK-/-) exhibits accelerated glycogenolysis during contraction. Understanding this phenomenon could provide insight into the control of glycogenolysis during contraction. Therefore, glycogen breakdown was investigated in isolated extensor

  16. The tyrosyl residues in creatine kinase. Modification by iodine.

    Science.gov (United States)

    Fattoum, A; Kassab, R; Pradel, L A

    1975-10-20

    The effect of the iodination of tyrosyl residues in creatine kinase from rabbit muscle has been investigated at alkaline pH after reversible masking of the reactive thiol groups. The conversion of 4-5 tyrosyl residues to monoiodotyrosines as measured by spectrotitration and by radioactive iodine labelling resulted in almost total loss of enzymic activity. The modified enzyme was unable to bind its nucleotide substrates but no significant conformational change was revealed by optical rotatory dispersion or Stokes radius measurements. However, change in the reactivity of some non-essential thiol groups, presumably those located near the active thiol groups, was observed.

  17. Effects of acrylamide and acrylic acid on creatine kinase activity in the rat brain

    International Nuclear Information System (INIS)

    Kohriyama, Kazuaki; Matsuoka, Masato; Igisu, Hideki

    1994-01-01

    In vitro, both acrylamide and acrylic acid inhibited creatine kinase (CK) activity in rat brain homogenates, and acrylic acid was more potent than acrylamide. In vivo, however, when given i.p. 50 mg/kg per day for 8 days to rats, only acrylamide inhibited CK activity in the brain and caused apparent neurological signs. 14 C in the brain 24 h after the injection of 14 C-labelled chemicals was more than 7 times greater with acrylamide than with acrylic acid. The inhibition of CK activity by acrylamide varied in eight regions of the brain; from 54% in hypothalamus to 27% in cerebellar vermis. The regional difference of CK inhibition, however, did not agree well with either 14 C distribution or with the distribution in regions which appear clinically or pathologically vulnerable to acrylamide. (orig.)

  18. 31P NMR measurements of the ADP concentration in yeast cells genetically modified to express creatine kinase

    International Nuclear Information System (INIS)

    Brindle, K.; Braddock, P.; Fulton, S.

    1990-01-01

    Rabbit muscle creatine kinase has been introduced into the yeast Saccharomyces cerevisiae by transforming cells with a multicopy plasmid containing the coding sequence for the enzyme under the control of the yeast phosphoglycerate kinase promoter. The transformed cells showed creating kinase activities similar to those found in mammalian heart muscle. 31 P NMR measurements of the near-equilibrium concentrations of phosphocreatine and cellular pH together with measurements of the total extractable concentrations of phosphocreatine and creatine allowed calculation of the free ADP/ATP ratio in the cell. The calculated ratio of approximately 2 was considerably higher than the ratio of between 0.06 and 0.1 measured directly in cell extracts

  19. Chromatographic separation and continuously referenced, on-line monitoring of creatine kinase isoenzymes by use of an immobilized-enzyme microreactor

    International Nuclear Information System (INIS)

    Denton, M.S.; Bostick, W.D.; Dinsmore, S.R.; Mrochek, J.E.

    1978-01-01

    We describe a new concept in continuously referenced monitoring of the isoenzyme activities of creatine kinase (EC 2.7.3.2) after liquid-chromatographic separation. After separation on a diethylaminoethyl-Sephacel column, the three isoenzymes of creatine kinase undergo a series of upled enzyme reactions, ultimately resulting in the formation of ultraviolet-detectable NADPH. A major advantage of this detection system is the immobilized-enzyme microreactor (2 x 17 mm), which may be removed and stored refrigerated when not in use. A split-stream configuration allows self-blanking of endogenous ultraviolet-absorbing constituents in authentic sera samples, which would otherwise make definitive diagnosis and quantitation difficult or impossible. This detection system is applicable to the automated analysis of creatine kinase isoenzymes in the clinical laboratory. 5 figures; 42 references

  20. Structural changes of creatine kinase upon substrate binding.

    Science.gov (United States)

    Forstner, M; Kriechbaum, M; Laggner, P; Wallimann, T

    1998-08-01

    Small-angle x-ray scattering was used to investigate structural changes upon binding of individual substrates or a transition state analog complex (TSAC; Mg-ADP, creatine, and KNO3) to creatine kinase (CK) isoenzymes (dimeric muscle-type (M)-CK and octameric mitochondrial (Mi)-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occurred upon binding of Mg-nucleotide or TSAC. The radius of gyration of Mi-CK was reduced from 55.6 A (free enzyme) to 48.9 A (enzyme plus Mg-ATP) and to 48.2 A (enzyme plus TSAC). M-CK showed similar changes from 28.0 A (free enzyme) to 25.6 A (enzyme plus Mg-ATP) and to 25.5 A (enzyme plus TSAC). Creatine alone did not lead to significant changes in the radii of gyration, nor did free ATP or ADP. AK also showed a change of the radius of gyration from 21.5 A (free enzyme) to 19.7 A (enzyme plus Mg-ATP), whereas with arginine alone only a minor change could be observed. The primary change in structure as seen with monomeric AK seems to be a Mg-nucleotide-induced domain movement relative to each other, whereas the effect of substrate may be of local order only. In CK, however, additional movements have to be involved.

  1. Interpretation of acid α-glucosidase activity in creatine kinase elevation: A case of Becker muscular dystrophy.

    Science.gov (United States)

    Oitani, Yoshiki; Ishiyama, Akihiko; Kosuga, Motomichi; Iwasawa, Kentaro; Ogata, Ayako; Tanaka, Fumiko; Takeshita, Eri; Shimizu-Motohashi, Yuko; Komaki, Hirofumi; Nishino, Ichizo; Okuyama, Torayuki; Sasaki, Masayuki

    2018-05-16

    Diagnosis of Pompe disease is sometimes challenging because it exhibits clinical similarities to muscular dystrophy. We describe a case of Becker muscular dystrophy (BMD) with a remarkable reduction in activity of the acid α-glucosidase (GAA) enzyme, caused by a combination of pathogenic mutation and polymorphism variants resulting in pseudodeficiency in GAA. The three-year-old boy demonstrated asymptomatic creatine kinase elevation. Neither exon deletion nor duplication was detected on multiplex ligation-dependent probe amplification (MLPA) of DMD. GAA enzyme activity in both dried blood spots and lymphocytes was low, at 11.7% and 7.7% of normal, respectively. However, genetic analysis of GAA detected only heterozygosity for a nonsense mutation (c.118C > T, p.Arg40 ∗ ). Muscle pathology showed no glycogen deposits and no high acid phosphatase activity. Hematoxylin-eosin staining detected scattered regenerating fibers; the fibers were faint and patchy on immunochemistry staining of dystrophin. The amount of dystrophin protein was reduced to 11.8% of normal, on Western blotting analysis. Direct sequencing analysis of DMD revealed hemizygosity for a nonsense mutation (c.72G > A, p.Trp24 ∗ ). The boy was diagnosed with BMD, despite remarkable reduction in GAA activity; further, he demonstrated heterozygosity for [p.Gly576Ser; p.Glu689Lys] polymorphism variants that indicated pseudodeficiency on another allele in GAA. Pseudodeficiency alleles are detected in approximately 4% of the Asian population; these demonstrate low activity of acid α-glucosidase (GAA), similar to levels found in Pompe disease. Clinicians should be careful in their interpretations of pseudodeficiency alleles that complicate diagnosis in cases of elevated creatine kinase. Copyright © 2018 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

  2. Periodontal status and serum creatine kinase levels among young ...

    African Journals Online (AJOL)

    Objectives: It is hypothesized that soccer players with periodontal disease exhibit raised serum creatine kinase (CK) levels as compared to those without periodontal disease. We assessed the clinical gingival status and serum CK levels among young soccer players. Materials and Methods: Demographic data were ...

  3. The value of creatine kinase, estradiol and progesterone levels in early diagnosis of ectopic pregnancies: a prospective controlled study

    Directory of Open Access Journals (Sweden)

    Feride Mimaroğlu

    2010-06-01

    Full Text Available INTRODUCTION: To evaluate the role of serum creatine kinase, progesterone and estradiol as a biochemical marker in the early diagnosis of tubal pregnancy. MATERIAL-METHODS: A prospective controlled study was carried out on 44 women with first trimester pregnancy. First group (n=22 with tubal pregnancy formed the study group and second group (n=22 with normal intrauterine pregnancy was taken as controls. Serum beta hCG, creatine kinase, progesterone and estradiol levels in the two groups were compared. Surgical treatment had choosen as a treatment modality of ectopic pregnancy. RESULTS: The optimal cutoff value of creatine kinase to be used for the prediction of ectopic pregnancy was 45 IU/l, which resulted in a sensitivity of 86%, specificity of 31%, positive predictive value 55 % and negative predictive value 70 %. The same values for estradiol and progesterone were detected >225 pg/ml, 100 %, 68 %, 75%, 100 % and >13 ng/mL, 95 %, 81 %, % 84, % 97 in discriminating ectopic pregnancies. According to AUC levels there was a significant difference between estradiol-creatine kinase levels, progesterone-estradiol levels and progesterone–creatin kinase levels (p values 0.024, 0.0082, and 0.0001, respectively. CONCLUSION: Serum creatine kinase values appear to be a useful marker in the diagnosis of ectopic pregnancy.

  4. Changes in neutrophil count, creatine kinases and muscle soreness ...

    African Journals Online (AJOL)

    Objective. A primary objective was to examine circulating neutrophil count after repeated bouts of downhill running. An additional aim was to determine creatine kinase (CK) levels during the initial 12 hours, after repeated DHRs. Design. Eleven healthy, untrained Caucasian males performed 2 x 60 min bouts of DHR ...

  5. Periodontal status and serum creatine kinase levels among young ...

    African Journals Online (AJOL)

    2015-12-02

    Dec 2, 2015 ... Key words: Periodontal disease, serum creatine kinase, soccer players ... has also been reported that poor oral health status influences the quality of life of an individual ..... A short‑term longitudinal randomized case‑control study. Clin Oral ... crevicular fluid from chronic periodontitis patients before and after.

  6. The influence of individualizing physical loads on speed, creatine kinase activity and lactate dehydrogenase in football players.

    Directory of Open Access Journals (Sweden)

    2008-06-01

    Full Text Available Introduction: One of the most important training problems in: contemporary football is speed preparation of a player for the season and the ability of keeping it on the same, relatively high level throughout the starting period [1]. The main process used for re-synthesis ATP during single, short-lasting efforts of maximal intensity, is decomposition of phospho-creatine under the influence of creatine kinase enzyme. Physical loads imposed during speed trainings often exceed the possibility of producing energy from phosphogenic reserve through oxygen - lactate free processes, because the supply of phospho-creatine is used very quickly. In such circumstances the lacking energy is refilled through processes called oxygen free glicolise with the help of lactate dehydrogenase enzyme. The aim of the work was to answer the question:

  7. Changes of creatine kinase structure upon ligand binding as seen by small-angle scattering

    Science.gov (United States)

    Forstner, Michael; Kriechbaum, Manfred; Laggner, Peter; Wallimann, Theo

    1996-09-01

    Small-angle X-ray and neutron scattering have been used to investigate structural changes upon binding of individual substrates or a transition state analogue complex (TSAC), consisting of Mg-ADP, creatine and KNO 3 to creatine kinase isoenzymes (dimeric M-CK and octameric Mi-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occurred upon binding of Mg-ATP and TSAC, whereas creatine alone had only a small effect. In Mi-CK, the radius of gyration was reduced from 55.6 Å (free enzyme) to 48.9 Å (enzyme + Mg-ATP) and to 48.2 Å (enzyme + TSAC). The experiments performed with M-CK showed similar changes from 28.0 Å (free enzyme) to 25.6 Å (enzyme + Mg-ATP) and to 25.5 Å (enzyme + TSAC). Creatine alone did not lead to significant changes in the radii of gyration, nor did free ATP or ADP. AK showed the same behaviour: a change of the radius of gyration from 21.5 Å (free enzyme) to 19.7 Å (enzyme + MG-ATP), whereas with arginine alone only a minor change could be observed. The primary change in structure as seen with monomeric AK seems to be a magnesium-nucleotide induced domain movement relative to each other, whereas the effect of substrate may be of local order only. In creatine kinase, however, further movements must be involved in the large conformational change.

  8. Prognostic importance of troponin T and creatine kinase after elective angioplasty

    NARCIS (Netherlands)

    Nienhuis, Mark B.; Ottervanger, Jan Paul; Dikkeschei, Bert; Suryapranata, Harry; de Boer, Menko-Jan; Dambrink, Jan-Henk E.; Hoorntje, Jan C. A.; van't Hof, Arnoud W. J.; Gosselink, Marcel; Zijlstra, Felix

    2007-01-01

    Background: The prognostic importance of elevated cardiac enzymes after elective percutaneous coronary intervention has been debated. Therefore, we performed a prospective observational study to evaluate the prognostic value of postprocedural rise of troponin T and creatine kinase. Methods: Troponin

  9. Creatine kinase in the serum of patients with acute infections of the central nervous system

    DEFF Research Database (Denmark)

    Peterslund, N A; Heinsvig, E M; Christensen, K D

    1985-01-01

    Serum creatine kinase was assessed in 94 consecutive patients without convulsions admitted to hospital due to suspicion of infection of the central nervous system. No reliable discrimination between patients with aseptic and those with bacterial meningitis was obtained. Patients with bacterial...... of bacterial meningitis. The highest serum CK value found in patients with encephalitis was 725 U/l. Reference values for control patients with meningism were 16-269 U/1. In a subset of 9 patients creatine kinase isoenzyme analysis was performed. In all cases only muscle type (MM) isoenzyme was found...

  10. Monoclonal antibody FsC-47 against carp sperm creatine kinase

    Czech Academy of Sciences Publication Activity Database

    Koubek, Pavel; Elzeinová, Fatima; Šulc, Miroslav; Linhart, O.; Pěknicová, Jana

    2006-01-01

    Roč. 25, č. 3 (2006), s. 154-157 ISSN 1554-0014 R&D Projects: GA ČR(CZ) GA524/03/0178 Institutional research plan: CEZ:AV0Z50520514; CEZ:AV0Z50200510 Keywords : creatin kinase * monoclonal antibody * carp sperm Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.411, year: 2006

  11. Comparison of Creatine Kinase Activity and Myoglobin Blood Level in Acute Myocardial Infarction Patients

    Directory of Open Access Journals (Sweden)

    Sabaheta Hasić

    2008-02-01

    Full Text Available The aim of this prospective study was to evaluate and compare the relative increase of serum myoglobin level and total creatine kinase(CK activity in acute myocardial infarction (AMI patients (n=36. We measured serial changes in total CK activity and myoglobin serum level in three-time periods (6-9 hours, 24 hours and 6-7 days from chest pains onset. Myoglobin peaked during the first 6-9 hours but total CK reached its peak activity after 24 hours from AMI symptoms onset. Results of this study showed that as non-specific cardiac marker myoglobin had better sensitivity and earlier rise in serum than total CK activity in AMI patients. Rapid kinetic of myoglobin level is important for its utility as marker for re-infarction diagnosis. Early myoglobin increase in serum is important for early triage of AMI patients and early "ruling out" of AMI diagnosis if there is no evidence of its elevation in circulation.

  12. Measurement of the enzymes lactate dehydrogenase and creatine kinase using reflectance spectroscopy and reagent strips.

    OpenAIRE

    Stevens, J F; Tsang, W; Newall, R G

    1983-01-01

    Two new methods for the assay of total activities of lactate dehydrogenase and creatine kinase are described, in which the enzyme activities are measured from a solid-state reagent strip during a kinetic reaction, the reaction being monitored in the ultra-violet region of the spectrum by reflectance spectroscopy. The performances of these methods are evaluated, and compared to conventional "wet" chemistry methods. The solid-phase reagent methods demonstrated precision and accuracy acceptable ...

  13. Creatine kinase B-driven energy transfer in the brain is important for habituation and spatial learning behaviour, mossy fibre field size and determination of seizure susceptibility.

    NARCIS (Netherlands)

    Jost, C.R.; Zee, C.E.E.M. van der; Zandt, H.J.A. in t; Oerlemans, F.T.J.J.; Verheij, M.M.M.; Streijger, F.; Fransen, J.A.M.; Deursen, J.; Heerschap, A.; Cools, A.R.; Wieringa, B.

    2002-01-01

    Creatine kinases are important in maintaining cellular-energy homeostasis, and neuroprotective effects have been attributed to the administration of creatine and creatine-like compounds. Herein we examine whether ablation of the cytosolic brain-type creatine kinase (B-CK) in mice has detrimental

  14. Creatine kinase B-driven energy transfer in the brain is important for habituation and spatial learning behaviour, mossy fibre field size and determination of seizure susceptibility

    NARCIS (Netherlands)

    Jost, C.R.; Zee, C.E.E.M. van der; Zandt, H.J.A. in t; Oerlemans, F.T.J.J.; Verheij, M.M.M.; Streijger, F.; Fransen, J.A.M.; Heerschap, A.; Cools, A.R.; Wieringa, B.

    2002-01-01

    Creatine kinases are important in maintaining cellular-energy homeostasis, and neuroprotective effects have been attributed to the administration of creatine and creatine-like compounds. Herein we examine whether ablation of the cytosolic brain-type creatine kinase (B-CK) in mice has detrimental

  15. Increasing creatine kinase activity protects against hypoxia / reoxygenation injury but not against anthracycline toxicity in vitro.

    Directory of Open Access Journals (Sweden)

    Sevasti Zervou

    Full Text Available The creatine kinase (CK phosphagen system is fundamental to cellular energy homeostasis. Cardiomyocytes express three CK isoforms, namely the mitochondrial sarcomeric CKMT2 and the cytoplasmic CKM and CKB. We hypothesized that augmenting CK in vitro would preserve cell viability and function and sought to determine efficacy of the various isoforms. The open reading frame of each isoform was cloned into pcDNA3.1, followed by transfection and stable selection in human embryonic kidney cells (HEK293. CKMT2- CKM- and CKB-HEK293 cells had increased protein and total CK activity compared to non-transfected cells. Overexpressing any of the three CK isoforms reduced cell death in response to 18h hypoxia at 1% O2 followed by 2h re-oxygenation as assayed using propidium iodide: by 33% in CKMT2, 47% in CKM and 58% in CKB compared to non-transfected cells (P<0.05. Loading cells with creatine did not modify cell survival. Transient expression of CK isoforms in HL-1 cardiac cells elevated isoenzyme activity, but only CKMT2 over-expression protected against hypoxia (0.1% for 24h and reoxygenation demonstrating 25% less cell death compared to non-transfected control (P<0.01. The same cells were not protected from doxorubicin toxicity (250nM for 48h, in contrast to the positive control. These findings support increased CK activity as protection against ischaemia-reperfusion injury, in particular, protection via CKMT2 in a cardiac-relevant cell line, which merits further investigation in vivo.

  16. Similar mitochondrial activation kinetics in wild-type and creatine kinase-deficient fast-twitch muscle indicate significant Pi control of respiration

    NARCIS (Netherlands)

    Jeneson, J.A.L.; Veld, ter F.; Schmitz, J.P.J.; Meyer, R.A.; Hilbers, P.A.J.; Nicolay, K.

    2011-01-01

    Past simulations of oxidative ATP metabolism in skeletal muscle have predicted that elimination of the creatine kinase (CK) reaction should result in dramatically faster oxygen consumption dynamics during transitions in ATP turnover rate. This hypothesis was investigated. Oxygen consumption of

  17. Measurement of the enzymes lactate dehydrogenase and creatine kinase using reflectance spectroscopy and reagent strips.

    Science.gov (United States)

    Stevens, J F; Tsang, W; Newall, R G

    1983-01-01

    Two new methods for the assay of total activities of lactate dehydrogenase and creatine kinase are described, in which the enzyme activities are measured from a solid-state reagent strip during a kinetic reaction, the reaction being monitored in the ultra-violet region of the spectrum by reflectance spectroscopy. The performances of these methods are evaluated, and compared to conventional "wet" chemistry methods. The solid-phase reagent methods demonstrated precision and accuracy acceptable for diagnostic purposes, and were easy to use by trained operators. PMID:6655069

  18. Mitochondrial DNA depletion by ethidium bromide decreases neuronal mitochondrial creatine kinase: Implications for striatal energy metabolism.

    Science.gov (United States)

    Warren, Emily Booth; Aicher, Aidan Edward; Fessel, Joshua Patrick; Konradi, Christine

    2017-01-01

    Mitochondrial DNA (mtDNA), the discrete genome which encodes subunits of the mitochondrial respiratory chain, is present at highly variable copy numbers across cell types. Though severe mtDNA depletion dramatically reduces mitochondrial function, the impact of tissue-specific mtDNA reduction remains debated. Previously, our lab identified reduced mtDNA quantity in the putamen of Parkinson's Disease (PD) patients who had developed L-DOPA Induced Dyskinesia (LID), compared to PD patients who had not developed LID and healthy subjects. Here, we present the consequences of mtDNA depletion by ethidium bromide (EtBr) treatment on the bioenergetic function of primary cultured neurons, astrocytes and neuron-enriched cocultures from rat striatum. We report that EtBr inhibition of mtDNA replication and transcription consistently reduces mitochondrial oxygen consumption, and that neurons are significantly more sensitive to EtBr than astrocytes. EtBr also increases glycolytic activity in astrocytes, whereas in neurons it reduces the expression of mitochondrial creatine kinase mRNA and levels of phosphocreatine. Further, we show that mitochondrial creatine kinase mRNA is similarly downregulated in dyskinetic PD patients, compared to both non-dyskinetic PD patients and healthy subjects. Our data support a hypothesis that reduced striatal mtDNA contributes to energetic dysregulation in the dyskinetic striatum by destabilizing the energy buffering system of the phosphocreatine/creatine shuttle.

  19. Mitochondrial DNA depletion by ethidium bromide decreases neuronal mitochondrial creatine kinase: Implications for striatal energy metabolism.

    Directory of Open Access Journals (Sweden)

    Emily Booth Warren

    Full Text Available Mitochondrial DNA (mtDNA, the discrete genome which encodes subunits of the mitochondrial respiratory chain, is present at highly variable copy numbers across cell types. Though severe mtDNA depletion dramatically reduces mitochondrial function, the impact of tissue-specific mtDNA reduction remains debated. Previously, our lab identified reduced mtDNA quantity in the putamen of Parkinson's Disease (PD patients who had developed L-DOPA Induced Dyskinesia (LID, compared to PD patients who had not developed LID and healthy subjects. Here, we present the consequences of mtDNA depletion by ethidium bromide (EtBr treatment on the bioenergetic function of primary cultured neurons, astrocytes and neuron-enriched cocultures from rat striatum. We report that EtBr inhibition of mtDNA replication and transcription consistently reduces mitochondrial oxygen consumption, and that neurons are significantly more sensitive to EtBr than astrocytes. EtBr also increases glycolytic activity in astrocytes, whereas in neurons it reduces the expression of mitochondrial creatine kinase mRNA and levels of phosphocreatine. Further, we show that mitochondrial creatine kinase mRNA is similarly downregulated in dyskinetic PD patients, compared to both non-dyskinetic PD patients and healthy subjects. Our data support a hypothesis that reduced striatal mtDNA contributes to energetic dysregulation in the dyskinetic striatum by destabilizing the energy buffering system of the phosphocreatine/creatine shuttle.

  20. Chronic Hypoxia Enhances Expression and Activity of Mitochondrial Creatine Kinase and Hexokinase in the Rat Ventricular Myocardium

    Czech Academy of Sciences Publication Activity Database

    Wasková-Arnoštová, P.; Kašparová, D.; Elsnicová, B.; Novotný, J.; Neckář, Jan; Kolář, František; Žurmanová, J.

    2014-01-01

    Roč. 33, č. 2 (2014), s. 310-320 ISSN 1015-8987 R&D Projects: GA AV ČR(CZ) IAAX01110901; GA ČR(CZ) GAP303/12/1162 Grant - others:Univerzita Karlova(CZ) 349211; GA AV ČR(CZ) IAA601110908 Institutional support: RVO:67985823 Keywords : creatine kinase * hexokinase * normobaric hypoxia * left ventricle * right ventricle * mitochondria co-localization Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 2.875, year: 2014

  1. Effects on Energy Metabolism of Two Guanidine Molecules, (Boc)2 -Creatine and Metformin.

    Science.gov (United States)

    Garbati, Patrizia; Ravera, Silvia; Scarfì, Sonia; Salis, Annalisa; Rosano, Camillo; Poggi, Alessandro; Damonte, Gianluca; Millo, Enrico; Balestrino, Maurizio

    2017-09-01

    Several enzymes are involved in the energy production, becoming a possible target for new anti-cancer drugs. In this paper, we used biochemical and in silico studies to evaluate the effects of two guanidine molecules, (Boc) 2 -creatine and metformin, on creatine kinase, an enzyme involved in the regulation of intracellular energy levels. Our results show that both drugs inhibit creatine kinase activity; however, (Boc) 2 -creatine displays a competitive inhibition, while metformin acts with a non-competitive mechanism. Moreover, (Boc) 2 -creatine is able to inhibit the activity of hexokinase with a non-competitive mechanism. Considering that creatine kinase and hexokinase are involved in energy metabolism, we evaluated the effects of (Boc) 2 -creatine and metformin on the ATP/AMP ratio and on cellular proliferation in healthy fibroblasts, human breast cancer cells (MDA-MB-468), a human neuroblastoma cell line (SH-SY5Y), a human Hodgkin lymphoma cell line (KMH2). We found that healthy fibroblasts were only partially affected by (Boc) 2 -creatine, while both ATP/AMP ratio and viability of the three cancer cell lines were significantly decreased. By inhibiting both creatine kinase and hexokinase, (Boc) 2 -creatine appears as a promising new agent in anticancer treatment. Further research is needed to understand what types of cancer cells are most suitable to treatment by this new compound. J. Cell. Biochem. 118: 2700-2711, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  2. The Effects of Pre-slaughter Stress and Season on the Activity of Plasma Creatine Kinase and Mutton Quality from Different Sheep Breeds Slaughtered at a Smallholder Abattoir

    Directory of Open Access Journals (Sweden)

    A. Y. Chulayo

    2013-12-01

    Full Text Available The objective of the current study was to determine the effect of pre-slaughter stress, season and breed on the activity of plasma creatine kinase (CK and the quality of mutton. One hundred and seventy-three (173 castrated sheep from Dormer (DM, South African Mutton Merino (SAMM, Dorper (DP and Blackhead Persian (BP sheep breeds were used in the study. The animals were grouped according to age-groups as follows: Group 1 (6 to 8 months, Group 2 (9 to 12 months and Group 3 (13 to 16 months. Blood samples were collected during exsanguinations using disposable vacutainer tubes for CK analysis. Representative samples of the Muscularis longissimuss thoracis et. lumborum (LTL were collected from 84 castrated sheep, of different breeds (28 per breed 24 h after slaughter. The following physico-chemical characteristics of mutton were determined; meat pH (pH24, color (L*, a* and b*, thawing and cooking losses and Warner Braztler Shear Force (WBSF. The activity of plasma CK was significantly higher (p0.001 levels (1,358.6±191.08 of CK. South African Mutton Merino had higher values for pH24 (5.9±0.06, L* (34.2±0.97, b* (12.2±0.50 and WBSF (26.8±1.51 and Blackhead Persian had higher values (35.5±2.17 for cooking loss (CL% than the other breeds. Computed Principal Component Analyses (PCA on the activity of plasma CK and physico-chemical characteristics of mutton revealed no correlations between these variables. However, positive correlations were observed between pH24, L*, a*, b*, CL% and WBSF. Relationships between pre-slaughter stress, CK activity and physico-chemical characteristics of mutton were also observed. It was therefore concluded that although mutton quality and creatine kinase were not related, pre-slaughter stress, season and breed affected the activity of creatine kinase and mutton quality.

  3. Creatine for women: a review of the relationship between creatine and the reproductive cycle and female-specific benefits of creatine therapy.

    Science.gov (United States)

    Ellery, Stacey J; Walker, David W; Dickinson, Hayley

    2016-08-01

    The creatine/phosphocreatine/creatine kinase circuit is instrumental in regulating high-energy phosphate metabolism, and the maintenance of cellular energy turnover. The mechanisms by which creatine is able to buffer and regulate cellular energy balance, maintain acid-base balance, and reduce the effects of oxidative stress have led to a large number of studies into the use of creatine supplementation in exercise performance and to treat diseases associated with cellular energy depletion. Some of these studies have identified sex-specific responses to creatine supplementation, as such; there is the perception, that females might be less receptive to the benefits of creatine supplementation and therapy, compared to males. This review will describe the differences in male and female physique and physiology that may account for such differences, and discuss the apparent endocrine modulation of creatine metabolism in females. Hormone-driven changes to endogenous creatine synthesis, creatine transport and creatine kinase expression suggest that significant changes in this cellular energy circuit occur during specific stages of a female's reproductive life, including pregnancy and menopause. Recent studies suggest that creatine supplementation may be highly beneficial for women under certain conditions, such as depression. A greater understanding of these pathways, and the consequences of alterations to creatine bioavailability in females are needed to ensure that creatine is used to full advantage as a dietary supplement to optimize and enhance health outcomes for women.

  4. Disturbed energy metabolism and muscular dystrophy caused by pure creatine deficiency are reversible by creatine intake

    Science.gov (United States)

    Nabuurs, C I; Choe, C U; Veltien, A; Kan, H E; van Loon, L J C; Rodenburg, R J T; Matschke, J; Wieringa, B; Kemp, G J; Isbrandt, D; Heerschap, A

    2013-01-01

    Creatine (Cr) plays an important role in muscle energy homeostasis by its participation in the ATP–phosphocreatine phosphoryl exchange reaction mediated by creatine kinase. Given that the consequences of Cr depletion are incompletely understood, we assessed the morphological, metabolic and functional consequences of systemic depletion on skeletal muscle in a mouse model with deficiency of l-arginine:glycine amidinotransferase (AGAT−/−), which catalyses the first step of Cr biosynthesis. In vivo magnetic resonance spectroscopy showed a near-complete absence of Cr and phosphocreatine in resting hindlimb muscle of AGAT−/− mice. Compared with wild-type, the inorganic phosphate/β-ATP ratio was increased fourfold, while ATP levels were reduced by nearly half. Activities of proton-pumping respiratory chain enzymes were reduced, whereas F1F0-ATPase activity and overall mitochondrial content were increased. The Cr-deficient AGAT−/− mice had a reduced grip strength and suffered from severe muscle atrophy. Electron microscopy revealed increased amounts of intramyocellular lipid droplets and crystal formation within mitochondria of AGAT−/− muscle fibres. Ischaemia resulted in exacerbation of the decrease of pH and increased glycolytic ATP synthesis. Oral Cr administration led to rapid accumulation in skeletal muscle (faster than in brain) and reversed all the muscle abnormalities, revealing that the condition of the AGAT−/− mice can be switched between Cr deficient and normal simply by dietary manipulation. Systemic creatine depletion results in mitochondrial dysfunction and intracellular energy deficiency, as well as structural and physiological abnormalities. The consequences of AGAT deficiency are more pronounced than those of muscle-specific creatine kinase deficiency, which suggests a multifaceted involvement of creatine in muscle energy homeostasis in addition to its role in the phosphocreatine–creatine kinase system. PMID:23129796

  5. Nuclear Overhauser effect studies on the conformation of magnesium adenosine 5'-triphosphate bound to rabbit muscle creatine kinase

    International Nuclear Information System (INIS)

    Rosevear, P.R.; Powers, V.M.; Dowhan, D.; Mildvan, A.S.; Kenyon, G.L.

    1987-01-01

    Nuclear Overhauser effects were used to determine interproton distances on MgATP bound to rabbit muscle creatine kinase. The internuclear distances were used in a distance geometry program that objectively determines both the conformation of the bound MgATP and its uniqueness. Two classes of structures were found that satisfied the measured interproton distances. Both classes had the same anti glycosidic torsional angle (X = 78 +/- 10 0 ) but differed in their ribose ring puckers (O1'-endo or C4'-exo). The uniqueness of the glycosidic torsional angle is consistent with the preference of creatine kinase for adenine nucleotides. One of these conformations of MgATP bound to creatine kinase is indistinguishable from the conformation found for Co(NH 3 ) 4 ATP bound to the catalytic subunit of protein kinase, which also has a high specificity for adenine nucleotides. Distance geometry calculations also suggest that upper limit distances, when low enough (≤ 3.4 A), can be used instead of measured distances to define, within experimental error, the glycosidic torsional angle of bound nucleotides. However, this approach does not permit an evaluation of the ribose ring pucker

  6. Three-step preparation and purification of phosphorus-33-labeled creatine phosphate of high specific activity

    International Nuclear Information System (INIS)

    Savabi, F.; Geiger, P.J.; Bessman, S.P.

    1984-01-01

    Rabbit heart mitochondria were used as a source of enzymes for the synthesis of phosphorus-labeled creatine phosphate. This method is based on the coupled reaction between mitochondrial oxidative phosphorylation and mitochondrial-bound creatine kinase. It is possible to convert more than 90% of the inorganic phosphate (P/sub i/) to creatine phosphate. The method used only small amounts of adenine nucleotides which led to a product with only slight nucleotide contamination. This could be removed by activated charcoal extraction. For further purification, a method for the removal of residual P/sub i/ is described. 20 references

  7. Normal results of post-race thallium-201 myocardial perfusion imaging in marathon runners with elevated serum MB creatine kinase levels

    International Nuclear Information System (INIS)

    Siegel, A.J.; Silverman, L.M.; Holman, B.L.

    1985-01-01

    Elevated cardiac enzyme values in asymptomatic marathon runners after competition can arise from skeletal muscle through exertional rhabdomyolysis, silent injury to the myocardium, or a combined tissue source. Peak post-race levels of the MB isoenzyme of creatine kinase are similar to values in patients with acute myocardial infarction. Previously reported normal results of infarct-avid myocardial scintigraphy with technetium 99m pyrophosphate in runners after competition suggest a non-cardiac source but cannot exclude silent injury to the myocardium. Therefore, thallium 201 myocardial perfusion imaging was performed in runners immediately after competition together with determination of sequential cardiac enzyme levels. Among 15 runners tested, the average peak in serum MB creatine kinase 24 hours after the race was 128 IU/liter with a cumulative MB creatine kinase release of 117 IU/liter; these values are comparable to those in patients with acute transmural myocardial infarction. Thallium 201 myocardial scintigraphic results were normal in five runners randomly selected from those who volunteered for determination of sequential blood levels. It is concluded that elevations of serum MB creatine kinase in marathon runners arise from a skeletal muscle source and that thallium 201 myocardial scintigraphy is useful to assess runners for myocardial injury when clinical questions arise

  8. Mice lacking the UbCKmit isoform of creatine kinase reveal slower spatial learning acquisition, diminished exploration and habituation, and reduced acoustic startle reflex responses.

    NARCIS (Netherlands)

    Streijger, F.; Jost, C.R.; Oerlemans, F.T.J.J.; Ellenbroek, B.A.; Cools, A.R.; Wieringa, B.; Zee, C.E.E.M. van der

    2004-01-01

    Brain-type creatine kinases B-CK (cytosolic) and UbCKmit (mitochondrial) are considered important for the maintenance and distribution of cellular energy in the central nervous system. Previously, we have demonstrated an abnormal behavioral phenotype in mice lacking the B-CK creatine kinase isoform,

  9. Growth inhibition in response to estrogen withdrawal and tamoxifen therapy of human breast cancer xenografts evaluated by in vivo 31P magnetic resonance spectroscopy, creatine kinase activity, and apoptotic index

    DEFF Research Database (Denmark)

    Kristensen, C A; Kristjansen, P E; Brünner, N

    1995-01-01

    index, and creatine kinase (CK) activity. Tumors of each line were grown in ovariectomized nude mice during stimulation from a s.c. 17 beta-estradiol pellet. At a tumor size of approximately 350 mm3, the pellet was removed from one-half of the animals. The remaining one-half served as controls...

  10. Creatin-kinase elevation after accidental ingestion of almotriptan in an 18-month-old girl.

    Science.gov (United States)

    Castagno, E; Lupica, M; Viola, S; Savino, F; Miniero, R

    2014-02-01

    Few studies have been published to demonstrate tolerability and efficacy of almotriptan in adolescents and children with migraine, particularly in the first years of life, though preliminary results are favorable. We report the case of an 18-month-old infant with elevation of serum levels of creatin-kinase after the accidental ingestion of almotriptan. A previously healthy 18-month-old girl (weight: 13 kg) was admitted to our Department four hours after the accidental ingestion of 6.25 mg of almotriptan (0.48 mg/kg), without any specific symptom. The performed investigations showed high serum levels of creatin-kinase (CK) (527 IU/L; normal values: 24-170 IU/L). Transaminase, creatinine, aldolase, myoglobin and troponin T serum levels were normal. The electrocardiogram proved negative. Initial management consisted of parenteral rehydration with saline solution. CK levels lowered significantly at 12 hours (455 IU/L) and at 65 hours (188 IU/L) after the ingestion. No symptoms were observed before discharge and on follow-up.

  11. Augmentation of Creatine in the Heart.

    Science.gov (United States)

    Zervou, Sevasti; Whittington, Hannah J; Russell, Angela J; Lygate, Craig A

    2016-01-01

    Creatine is a principle component of the creatine kinase (CK) phosphagen system common to all vertebrates. It is found in excitable cells, such as cardiomyocytes, where it plays an important role in the buffering and transport of chemical energy to ensure that supply meets the dynamic demands of the heart. Multiple components of the CK system, including intracellular creatine levels, are reduced in heart failure, while ischaemia and hypoxia represent acute crises of energy provision. Elevation of myocardial creatine levels has therefore been suggested as potentially beneficial, however, achieving this goal is not trivial. This mini-review outlines the evidence in support of creatine elevation and critically examines the pharmacological approaches that are currently available. In particular, dietary creatine-supplementation does not sufficiently elevate creatine levels in the heart due to subsequent down-regulation of the plasma membrane creatine transporter (CrT). Attempts to increase passive diffusion and bypass the CrT, e.g. via creatine esters, have yet to be tested in the heart. However, studies in mice with genetic overexpression of the CrT demonstrate proof-of-principle that elevated creatine protects the heart from ischaemia-reperfusion injury. This suggests activation of the CrT as a major unmet pharmacological target. However, translation of this finding to the clinic will require a greater understanding of CrT regulation in health and disease and the development of small molecule activators.

  12. Creatine biosynthesis and transport in health and disease.

    Science.gov (United States)

    Joncquel-Chevalier Curt, Marie; Voicu, Pia-Manuela; Fontaine, Monique; Dessein, Anne-Frédérique; Porchet, Nicole; Mention-Mulliez, Karine; Dobbelaere, Dries; Soto-Ares, Gustavo; Cheillan, David; Vamecq, Joseph

    2015-12-01

    Creatine is physiologically provided equally by diet and by endogenous synthesis from arginine and glycine with successive involvements of arginine glycine amidinotransferase [AGAT] and guanidinoacetate methyl transferase [GAMT]. A specific plasma membrane transporter, creatine transporter [CRTR] (SLC6A8), further enables cells to incorporate creatine and through uptake of its precursor, guanidinoacetate, also directly contributes to creatine biosynthesis. Breakthrough in the role of creatine has arisen from studies on creatine deficiency disorders. Primary creatine disorders are inherited as autosomal recessive (mutations affecting GATM [for glycine-amidinotransferase, mitochondrial]) and GAMT genes) or X-linked (SLC6A8 gene) traits. They have highlighted the role of creatine in brain functions altered in patients (global developmental delay, intellectual disability, behavioral disorders). Creatine modulates GABAergic and glutamatergic cerebral pathways, presynaptic CRTR (SLC6A8) ensuring re-uptake of synaptic creatine. Secondary creatine disorders, addressing other genes, have stressed the extraordinary imbrication of creatine metabolism with many other cellular pathways. This high dependence on multiple pathways supports creatine as a cellular sensor, to cell methylation and energy status. Creatine biosynthesis consumes 40% of methyl groups produced as S-adenosylmethionine, and creatine uptake is controlled by AMP activated protein kinase, a ubiquitous sensor of energy depletion. Today, creatine is considered as a potential sensor of cell methylation and energy status, a neurotransmitter influencing key (GABAergic and glutamatergic) CNS neurotransmission, therapeutic agent with anaplerotic properties (towards creatine kinases [creatine-creatine phosphate cycle] and creatine neurotransmission), energetic and antioxidant compound (benefits in degenerative diseases through protection against energy depletion and oxidant species) with osmolyte behavior (retention of

  13. Radioimmunoassay of creatine kinase isoenzymes in human serum: isoenzyme MM

    International Nuclear Information System (INIS)

    Van Steirteghem, A.C.; Zweig, M.H.; Schechter, A.N.

    1978-01-01

    Measurement of the mass concentration of serum enzymes by radioimmunoassay provides direct quantitation of specific isoenzymes and may be less subject to some of the limitations of traditional assay procedures for enzymes. We describe the development of a sensitive and specific radioimmunoassay for the muscle isoenzyme of creatine kinase, CK-MM, in human serum. CK-MM, purified from human skeletal muscle, was used to raise high-titer antisera and for iodination by the Chloramine T method. The radioimmunoassay required 50 μl of sample, utilized a double-antibody separation method, and was completed in 24 h. Cross reactivity with CK-BB was virtually zero, 3 to 17 percent with CK-MB. The mass concentration of CK-MM in the serum of healthy subjects ranged from 36 to 1668 μg/liter and correlated closely with total CK enzymatic activity. Serum concentrations of CK-MM from casually selected patients correlated less well with total CK enzymatic activity, suggesting the existence of other CK isoenzymes or the presence of inactive forms

  14. Exchange rates of creatine kinase metabolites: feasibility of imaging creatine by chemical exchange saturation transfer MRI.

    Science.gov (United States)

    Haris, Mohammad; Nanga, Ravi Prakash Reddy; Singh, Anup; Cai, Kejia; Kogan, Feliks; Hariharan, Hari; Reddy, Ravinder

    2012-11-01

    Creatine (Cr), phosphocreatine (PCr) and adenosine-5-triphosphate (ATP) are major metabolites of the enzyme creatine kinase (CK). The exchange rate of amine protons of CK metabolites at physiological conditions has been limited. In the current study, the exchange rate and logarithmic dissociation constant (pKa) of amine protons of CK metabolites were calculated. Further, the chemical exchange saturation transfer effect (CEST) of amine protons of CK metabolites with bulk water was explored. At physiological temperature and pH, the exchange rate of amine protons in Cr was found to be 7-8 times higher than PCr and ATP. A higher exchange rate in Cr was associated with lower pKa value, suggesting faster dissociation of its amine protons compared to PCr and ATP. CEST MR imaging of these metabolites in vitro in phantoms displayed predominant CEST contrast from Cr and negligible contribution from PCr and ATP with the saturation pulse parameters used in the current study. These results provide a new method to perform high-resolution proton imaging of Cr without contamination from PCr. Potential applications of these finding are discussed. Copyright © 2012 John Wiley & Sons, Ltd.

  15. Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer

    Directory of Open Access Journals (Sweden)

    Tai Phillip WL

    2011-07-01

    Full Text Available Abstract Background Hundreds of genes, including muscle creatine kinase (MCK, are differentially expressed in fast- and slow-twitch muscle fibers, but the fiber type-specific regulatory mechanisms are not well understood. Results Modulatory region 1 (MR1 is a 1-kb regulatory region within MCK intron 1 that is highly active in terminally differentiating skeletal myocytes in vitro. A MCK small intronic enhancer (MCK-SIE containing a paired E-box/myocyte enhancer factor 2 (MEF2 regulatory motif resides within MR1. The SIE's transcriptional activity equals that of the extensively characterized 206-bp MCK 5'-enhancer, but the MCK-SIE is flanked by regions that can repress its activity via the individual and combined effects of about 15 different but highly conserved 9- to 24-bp sequences. ChIP and ChIP-Seq analyses indicate that the SIE and the MCK 5'-enhancer are occupied by MyoD, myogenin and MEF2. Many other E-boxes located within or immediately adjacent to intron 1 are not occupied by MyoD or myogenin. Transgenic analysis of a 6.5-kb MCK genomic fragment containing the 5'-enhancer and proximal promoter plus the 3.2-kb intron 1, with and without MR1, indicates that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers (types I and IIa, respectively, but is not required for expression in fast-twitch muscle fibers (types IIb and IId. Conclusions In this study, we discovered that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers and that MR1's positive transcriptional activity depends on a paired E-box MEF2 site motif within a SIE. This is the first study to delineate the DNA controls for MCK expression in different skeletal muscle fiber types.

  16. Rhabdomyolysis and acute kidney injury: creatine kinase as a prognostic marker and validation of the McMahon Score in a 10-year cohort: A retrospective observational evaluation.

    Science.gov (United States)

    Simpson, Joanna P; Taylor, Andrew; Sudhan, Nazneen; Menon, David K; Lavinio, Andrea

    2016-12-01

    High-volume fluid resuscitation and the administration of sodium bicarbonate and diuretics have a theoretical renoprotective role in patients at high risk of acute kidney injury (AKI) following rhabdomyolysis. Abnormally elevated creatine kinase has previously been used as a biological marker for the identification of patients at high risk of AKI following rhabdomyolysis. To assess the sensitivity and specificity of plasma creatine kinase (admission and peak values) for the prediction of AKI requiring renal replacement therapy (RRT) or of death in patients with confirmed rhabdomyolysis. To compare the diagnostic performance of creatine kinase with the McMahon score. Retrospective observational study. Data collection included McMahon and Acute Physiology and Chronic Health Evaluation II (APACHE II) scores; daily creatine kinase; daily creatinine and electrolytes; ICU length of stay and mortality. Neurosciences and Trauma Critical Care Unit (Cambridge, UK). In total, 232 adults with confirmed rhabdomyolysis (creatine kinase > 1000 Ul) admitted to Neurosciences and Trauma Critical Care Unit between 2002 and 2012. AKI, RRT and mortality. Forty-five (19%) patients developed AKI and 29 (12.5%) patients required RRT. Mortality was significantly higher in patients who developed AKI (62 vs. 18%, P rhabdomyolysis. Although a PEAK creatine kinase of at least 5000 Ul has sensitivity acceptable for screening purposes, this is often a delayed finding. A McMahon score of at least 6 calculated on admission allows for a more sensitive, specific and timely identification of patients who may benefit from high-volume fluid resuscitation.

  17. Contribution of creatine kinase MB mass concentration at admission to early diagnosis of acute myocardial infarction

    NARCIS (Netherlands)

    Bakker, A. J.; Gorgels, J. P.; van Vlies, B.; Koelemay, M. J.; Smits, R.; Tijssen, J. G.; Haagen, F. D.

    1994-01-01

    OBJECTIVE: To assess the diagnostic value at admission of creatine kinase MB mass concentration, alone or in combination with electrocardiographic changes, in suspected myocardial infarction. DESIGN: Prospective study of all consecutive patients admitted within 12 hours after onset of chest pain to

  18. Living Without Creatine: Unchanged Exercise Capacity and Response to Chronic Myocardial Infarction in Creatine-Deficient Mice

    Science.gov (United States)

    Lygate, Craig A.; Aksentijevic, Dunja; Dawson, Dana; Hove, Michiel ten; Phillips, Darci; de Bono, Joseph P.; Medway, Debra J.; Sebag-Montefiore, Liam; Hunyor, Imre; Channon, Keith M.; Clarke, Kieran; Zervou, Sevasti; Watkins, Hugh; Balaban, Robert S.; Neubauer, Stefan

    2014-01-01

    Rationale Creatine is thought to be involved in the spatial and temporal buffering of ATP in energetic organs such as heart and skeletal muscle. Creatine depletion affects force generation during maximal stimulation, while reduced levels of myocardial creatine are a hallmark of the failing heart, leading to the widely held view that creatine is important at high workloads and under conditions of pathological stress. Objective We therefore hypothesised that the consequences of creatine-deficiency in mice would be impaired running capacity, and exacerbation of heart failure following myocardial infarction. Methods and Results Surprisingly, mice with whole-body creatine deficiency due to knockout of the biosynthetic enzyme (guanidinoacetate N-methyltransferase – GAMT) voluntarily ran just as fast and as far as controls (>10km/night) and performed the same level of work when tested to exhaustion on a treadmill. Furthermore, survival following myocardial infarction was not altered, nor was subsequent LV remodelling and development of chronic heart failure exacerbated, as measured by 3D-echocardiography and invasive hemodynamics. These findings could not be accounted for by compensatory adaptations, with no differences detected between WT and GAMT−/− proteomes. Alternative phosphotransfer mechanisms were explored; adenylate kinase activity was unaltered, and although GAMT−/− hearts accumulated the creatine pre-cursor guanidinoacetate, this had negligible energy-transfer activity, while mitochondria retained near normal function. Conclusions Creatine-deficient mice show unaltered maximal exercise capacity and response to chronic myocardial infarction, and no obvious metabolic adaptations. Our results question the paradigm that creatine is essential for high workload and chronic stress responses in heart and skeletal muscle. PMID:23325497

  19. Quantitative determination of creatine kinase release from herring (Clupea harengus) spermatozoa induced by tributyltin.

    Science.gov (United States)

    Grzyb, Katarzyna; Rychłowski, Michał; Biegniewska, Anna; Skorkowski, Edward F

    2003-02-01

    Creatine kinase (CK, ATP creatine phosphotransferase, EC 2.7.3.2) is an enzyme participating in ATP regeneration, which is the primary source of energy in living organisms. We demonstrated that CK from herring spermatozoa has high activity ( approximately 452 micromol/min/g of fresh semen) and has a different electrophoretic mobility from isoenzymes present in skeletal muscle. In our study, we investigated toxic effect of tributyltin (TBT) on herring spermatozoa using a specific sperm viability kit to observe live and dead sperm cells with a confocal microscope. Treatment of herring spermatozoa with TBT caused a time-dependent decrease of viability: 35% nonviable cells with 5 microM TBT and more than 90% nonviable cells with 10 microM TBT after 6 h exposure. We also monitored CK release from damaged spermatozoa into surrounding medium containing different concentrations of TBT. The higher concentration of TBT was used the more CK release from spermatozoa was observed. We suggest that CK could be a good biomarker of sperm cell membranes degradation in the case when lactate dehydrogenase release from permeabilized cells is not possible for rapid determination of the effect of TBT.

  20. Local ATP generation by brain-type creatine kinase (CK-B facilitates cell motility.

    Directory of Open Access Journals (Sweden)

    Jan W P Kuiper

    Full Text Available BACKGROUND: Creatine Kinases (CK catalyze the reversible transfer of high-energy phosphate groups between ATP and phosphocreatine, thereby playing a storage and distribution role in cellular energetics. Brain-type CK (CK-B deficiency is coupled to loss of function in neural cell circuits, altered bone-remodeling by osteoclasts and complement-mediated phagocytotic activity of macrophages, processes sharing dependency on actomyosin dynamics. METHODOLOGY/PRINCIPAL FINDINGS: Here, we provide evidence for direct coupling between CK-B and actomyosin activities in cortical microdomains of astrocytes and fibroblasts during spreading and migration. CK-B transiently accumulates in membrane ruffles and ablation of CK-B activity affects spreading and migration performance. Complementation experiments in CK-B-deficient fibroblasts, using new strategies to force protein relocalization from cytosol to cortical sites at membranes, confirmed the contribution of compartmentalized CK-B to cell morphogenetic dynamics. CONCLUSION/SIGNIFICANCE: Our results provide evidence that local cytoskeletal dynamics during cell motility is coupled to on-site availability of ATP generated by CK-B.

  1. Creatine and creatine pyruvate reduce hypoxia-induced effects on phrenic nerve activity in the juvenile mouse respiratory system.

    Science.gov (United States)

    Scheer, Monika; Bischoff, Anna M; Kruzliak, Peter; Opatrilova, Radka; Bovell, Douglas; Büsselberg, Dietrich

    2016-08-01

    Adequate concentrations of ATP are required to preserve physiological cell functions and protect tissue from hypoxic damage. Decreased oxygen concentration results in ATP synthesis relying increasingly on the presence of phosphocreatine. The lack of ATP through hypoxic insult to neurons that generate or regulate respiratory function, would lead to the cessation of breathing (apnea). It is not clear whether creatine plays a role in maintaining respiratory phrenic nerve (PN) activity during hypoxic challenge. The aim of the study was to test the effects of exogenously applied creatine or creatine pyruvate in maintaining PN induced respiratory rhythm against the deleterious effects of severe hypoxic insult using Working Heart-Brainstem (WHB) preparations of juvenile Swiss type mice. WHB's were perfused with control perfusate or perfusate containing either creatine [100μM] or creatine pyruvate [100μM] prior to hypoxic challenge and PN activity recorded throughout. Results showed that severe hypoxic challenge resulted in an initial transient increase in PN activity, followed by a reduction in that activity leading to respiratory apnea. The results demonstrated that perfusing the WHB preparation with creatine or creatine pyruvate, significantly reduced the onset of apnea compared to control conditions, with creatine pyruvate being the more effective substance. Overall, creatine and creatine pyruvate each produced time-dependent degrees of protection against severe hypoxic-induced disturbances of PN activity. The underlying protective mechanisms are unknown and need further investigations. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Selective decrease of components of the creatine kinase system and ATP synthase complex in chronic Chagas disease cardiomyopathy.

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    Priscila Camillo Teixeira

    2011-06-01

    Full Text Available BACKGROUND: Chronic Chagas disease cardiomyopathy (CCC is an inflammatory dilated cardiomyopathy with a worse prognosis than other cardiomyopathies. CCC occurs in 30 % of individuals infected with Trypanosoma cruzi, endemic in Latin America. Heart failure is associated with impaired energy metabolism, which may be correlated to contractile dysfunction. We thus analyzed the myocardial gene and protein expression, as well as activity, of key mitochondrial enzymes related to ATP production, in myocardial samples of end-stage CCC, idiopathic dilated (IDC and ischemic (IC cardiomyopathies. METHODOLOGY/PRINCIPAL FINDINGS: Myocardium homogenates from CCC (N=5, IC (N=5 and IDC (N=5 patients, as well as from heart donors (N=5 were analyzed for protein and mRNA expression of mitochondrial creatine kinase (CKMit and muscular creatine kinase (CKM and ATP synthase subunits aplha and beta by immunoblotting and by real-time RT-PCR. Total myocardial CK activity was also assessed. Protein levels of CKM and CK activity were reduced in all three cardiomyopathy groups. However, total CK activity, as well as ATP synthase alpha chain protein levels, were significantly lower in CCC samples than IC and IDC samples. CCC myocardium displayed selective reduction of protein levels and activity of enzymes crucial for maintaining cytoplasmic ATP levels. CONCLUSIONS/SIGNIFICANCE: The selective impairment of the CK system may be associated to the loss of inotropic reserve observed in CCC. Reduction of ATP synthase alpha levels is consistent with a decrease in myocardial ATP generation through oxidative phosphorylation. Together, these results suggest that the energetic deficit is more intense in the myocardium of CCC patients than in the other tested dilated cardiomyopathies.

  3. The influence of whole-body vibration on creatine kinase activity and jumping performance in young basketball players.

    Science.gov (United States)

    Fachina, Rafael; da Silva, Antônio; Falcão, William; Montagner, Paulo; Borin, João; Minozzo, Fábio; Falcão, Diego; Vancini, Rodrigo; Poston, Brach; de Lira, Claudio

    2013-12-01

    To quantify creatine kinase (CK) activity changes across time following an acute bout of whole-body vibration (WBV) and determine the association between changes in CK activity and jumping performance. Twenty-six elite young basketball players were assigned to 3 groups: 36-Hz and 46-Hz vibration groups (G36 and G46, respectively) and a control group. The study quantified CK activity and jumping performance following an acute bout of WBV at 2 vibration frequencies. Both WBV groups performed a protocol that consisted of 10 sets of 60 s of WBV while standing on a vibration plate in a quarter-squat position. CK activity, countermovement jumps (CMJ), and squat jumps (SJ) were measured immediately before and 24 hr and 48 hr after WBV. In addition, CMJ and SJ were also measured 5 min after WBV. CK activity was statistically significantly increased 24 hr following WBV in G36 and G46. At 48 hr after WBV, CK activity was similar to baseline levels in G36 but remained statistically significantly above baseline levels in G46. The CMJ and SJ heights were statistically significantly decreased at 5 min following the protocol for both WBV groups. Overall, the changes in CK activity did not present a strong relationship with the changes in jump heights for any of the comparisons. These findings suggest that WBV protocols with such characteristics may not cause excessive muscle damage and may partly explain why many WBV training studies have failed to elicit increases in strength performance.

  4. Creatine kinase radioimmunoassay and isoenzyme electrophoresis compared in the diagnosis of acute myocardial infarction

    International Nuclear Information System (INIS)

    Homburger, H.A.; Jacob, G.L.

    1980-01-01

    We compared, in 116 patients, the relative usefulness of results of tests for creatine kinase B-isoenzymes, as measured by radioimmunoassay, and the MB isoenzyme, as measured by electrophoresis, in diagnosis of acute myocardial infarction. The radioimmunoassay was specific for isoenzymes of creatine kinase containing the B subunit. All patients with acute transmural infarcts had positive test results by both techniques, but concentrations of B-isoenzymes were more frequently above normal than were MB bands in the case of patients with acute subendocardial infarcts and in the case of all patients with acute myocardial infarcts from whom sera were collected more than 24 h after onset of chest pain. Concentrations of B-isoenzymes also were increased, even when MB bands were not electrophoretically detectable in specimens from several patients without documented acute myocardial infarcts. These abnormal results presumably were caused by increased concentrations of the BB isoenzyme in serum. Accordingly, an increased concentration of B-isoenzymes had less diagnostic specificity and predictive value for acute myocardial infarction than did a detectable MB band. Results of isoenzyme electrophoresis were more reliable for establishing this diagnosis, but the results of radioimmunoassay were more reliable for excluding it in patients with chest pain as the primary symptom

  5. Roles of the creatine kinase system and myoglobin in maintaining energetic state in the working heart

    Directory of Open Access Journals (Sweden)

    Beard Daniel A

    2009-02-01

    Full Text Available Abstract Background The heart is capable of maintaining contractile function despite a transient decrease in blood flow and increase in cardiac ATP demand during systole. This study analyzes a previously developed model of cardiac energetics and oxygen transport to understand the roles of the creatine kinase system and myoglobin in maintaining the ATP hydrolysis potential during beat-to-beat transient changes in blood flow and ATP hydrolysis rate. Results The theoretical investigation demonstrates that elimination of myoglobin only slightly increases the predicted range of oscillation of cardiac oxygenation level during beat-to-beat transients in blood flow and ATP utilization. In silico elimination of myoglobin has almost no impact on the cytoplasmic ATP hydrolysis potential (ΔGATPase. In contrast, disabling the creatine kinase system results in considerable oscillations of cytoplasmic ADP and ATP levels and seriously deteriorates the stability of ΔGATPase in the beating heart. Conclusion The CK system stabilizes ΔGATPase by both buffering ATP and ADP concentrations and enhancing the feedback signal of inorganic phosphate in regulating mitochondrial oxidative phosphorylation.

  6. the Effect of six weeks of high intensity interval training and zinc ‎supplement on serum ‎creatine kinase and uric acid levels in ‎futsal players ‎

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    Malihe Saeedy

    2017-01-01

    Full Text Available Background: strenuous training‎-induced reactive oxygen species is associated with several ‎chronic diseases‎ by damaging cell proteins and membrane lipids; it seems uric acid as a major ‎intracellular antioxidant could lower membranous lipid peroxidation and muscle damage. The aim ‎of this study was to examine the effect of six weeks of high-intensity interval training with and ‎‎without zinc ‎on serum Creatine Kinase and uric acid in female futsal players.‎ Methods: Thirty-two female futsal players were randomly divided into four groups: placebo, ‎Zinc, ‎HIT ‎and Zinc+HIT. All subjects had to attend futsal-specific training three sessions per ‎week. Zinc and ‎Placebo groups took ‎30 mg ‎day−1 of zinc gluconate or dextrose, respectively; ‎HIT groups accomplished high-intensity interval training contained 6 to 10 repetitions of a 30-‎second ‎running at 100% of VO2peak with a 4-minutes rest between efforts, during six weeks.‎ Results: After six weeks, Creatine Kinase ‎levels augmented insignificantly from 83.98 to 120.19‎ ‎‎(P=0.101 in ‎placebo, from 99.58 to 150.1(P=0.167 in HIT and from 81.07 to 107.90‎ ‎‎(P=0.152 ‎in HIT+Zinc group; while Creatine Kinase ‎levels increased significantly from 66.86 to ‎‎‎124.81(P=0.004 only in Zinc group. Uric acid levels increased in all groups (Placebo (P=1, Zinc ‎‎(P=‎0.317‎, HIT (P=‎0.157‎, ‎Zinc+HIT (P=1 insignificantly Conclusions: The findings indicated that ‎after six weeks, serum Creatine Kinase and uric acid ‎levels increased insignificantly in all groups; Creatine Kinase ‎levels augmented significantly, only ‎in Zinc group. Zinc as an antioxidant supplement could not decrease the muscle damage, and even ‎increased the serum Creatine Kinase as a marker of muscle damage, significantly

  7. A creatine-driven substrate cycle enhances energy expenditure and thermogenesis in beige fat.

    Science.gov (United States)

    Kazak, Lawrence; Chouchani, Edward T; Jedrychowski, Mark P; Erickson, Brian K; Shinoda, Kosaku; Cohen, Paul; Vetrivelan, Ramalingam; Lu, Gina Z; Laznik-Bogoslavski, Dina; Hasenfuss, Sebastian C; Kajimura, Shingo; Gygi, Steve P; Spiegelman, Bruce M

    2015-10-22

    Thermogenic brown and beige adipose tissues dissipate chemical energy as heat, and their thermogenic activities can combat obesity and diabetes. Herein the functional adaptations to cold of brown and beige adipose depots are examined using quantitative mitochondrial proteomics. We identify arginine/creatine metabolism as a beige adipose signature and demonstrate that creatine enhances respiration in beige-fat mitochondria when ADP is limiting. In murine beige fat, cold exposure stimulates mitochondrial creatine kinase activity and induces coordinated expression of genes associated with creatine metabolism. Pharmacological reduction of creatine levels decreases whole-body energy expenditure after administration of a β3-agonist and reduces beige and brown adipose metabolic rate. Genes of creatine metabolism are compensatorily induced when UCP1-dependent thermogenesis is ablated, and creatine reduction in Ucp1-deficient mice reduces core body temperature. These findings link a futile cycle of creatine metabolism to adipose tissue energy expenditure and thermal homeostasis. PAPERCLIP. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. A Creatine-Driven Substrate Cycle Enhances Energy Expenditure and Thermogenesis in Beige Fat

    Science.gov (United States)

    Kazak, Lawrence; Chouchani, Edward T.; Jedrychowski, Mark P.; Erickson, Brian K.; Shinoda, Kosaku; Cohen, Paul; Vetrivelan, Ramalingam; Lu, Gina Z.; Laznik-Bogoslavski, Dina; Hasenfuss, Sebastian C.; Kajimura, Shingo; Gygi, Steve P.; Spiegelman, Bruce M.

    2015-01-01

    SUMMARY Thermogenic brown and beige adipose tissues dissipate chemical energy as heat, and their thermogenic activities can combat obesity and diabetes. Herein the functional adaptations to cold of brown and beige adipose depots are examined using quantitative mitochondrial proteomics. We identify arginine/creatine metabolism as a beige adipose signature and demonstrate that creatine enhances respiration in beige fat mitochondria when ADP is limiting. In murine beige fat, cold exposure stimulates mitochondrial Creatine Kinase activity and induces coordinated expression of genes associated with creatine metabolism. Pharmacological reduction of creatine levels decreases whole body energy expenditure after administration of a β3-agonist and reduces the adipose metabolic rate. Genes of creatine metabolism are compensatorily induced when UCP1-dependent thermogenesis is ablated, and creatine reduction in Ucp1-deficient mice reduces core body temperature. These findings link a futile cycle of creatine metabolism to adipose tissue energy expenditure and thermal homeostasis. PMID:26496606

  9. Early detection of skeletal muscle injury by assay of creatine kinase MM isoforms in serum after acute exercise

    DEFF Research Database (Denmark)

    Apple, F. S.; Hellsten, Ylva; Clarkson, P. M.

    1988-01-01

    We could detect skeletal muscle injury early after an acute exercise bout by measuring creatine kinase (CK, EC 2.7.3.2) MM isoforms in serum. Eleven men performed 120 alternating-arm, eccentric (muscle lengthening) biceps contractions with the intensity of each contraction being 110% of maximal...

  10. Creatine and creatine analogues in hypertension and cardiovascular disease

    NARCIS (Netherlands)

    Horjus, Deborah L.; Oudman, Inge; van Montfrans, Gert A.; Brewster, Lizzy M.

    2011-01-01

    The creatine kinase system, the central regulatory system of cellular energy metabolism, provides ATP in situ at ATP-ases involved in ion transport and muscle contraction. Furthermore, the enzyme system provides relative protection from tissue ischaemia and acidosis. The system could therefore be a

  11. Interleukin-6, Creatine Kinase, and Antioxidant Enzyme Activities following Platelet-Rich Plasma Treatment on Muscle Injury: A Pilot Study

    Directory of Open Access Journals (Sweden)

    Lingling Lai

    2016-06-01

    Full Text Available The aim of this study was to investigate the effect of autologous platelet-rich plasma (PRP treatment alongside rehabilitation compared with rehabilitation alone on inflammatory cytokine (interleukin-6, IL-6, creatine kinase muscle type (CKM, and antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT following hamstring injury. This study was a randomised control trial. Participants diagnosed with grade-2 acute hamstring injury (n=16 were divided into 2 groups of PRP treatment with rehabilitation program (PRP-T and rehabilitation program (CON. Blood samples were collected at baseline, and 2 fortnightly for the various biochemical assessments. Participants were certified to have recovered upon fulfilling return to play (RTP criteria. Level of IL-6 and the activities of CKM, SOD, and CAT were measured. PRP-T group benefited from earlier time to RTP with significantly lower IL-6 level and CAT activity compared to CON group. There was no significant difference in CKM and SOD activities between the groups, though a trend of lower values in all variables was observed at week 4 compared to week 0. PRP treatment potentially improves muscle healing process by altering both the inflammatory and oxidative responses, hence hastens time to RTP. KEY WORDS:  Autologous, blood injection, rehabilitation, sports injury, hamstring injury

  12. Decreased creatine kinase is linked to diastolic dysfunction in rats with right heart failure induced by pulmonary artery hypertension

    NARCIS (Netherlands)

    Fowler, Ewan D.; Benoist, David; Drinkhill, Mark J.; Stones, Rachel; Helmes, Michiel; Wüst, Rob C. I.; Stienen, Ger J. M.; Steele, Derek S.; White, Ed

    2015-01-01

    Our objective was to investigate the role of creatine kinase in the contractile dysfunction of right ventricular failure caused by pulmonary artery hypertension. Pulmonary artery hypertension and right ventricular failure were induced in rats by monocrotaline and compared to saline-injected control

  13. Diagnostic value of serum creatine kinase-BB for acute meningitis in adults

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    Seyed Mohammad Alavi

    2017-01-01

    Full Text Available Objective: To find out an easy and feasible test instead of cerebrospinal fluid analysis for the diagnosis of acute meningitis. Methods: This cross-sectional study was conducted in 2013 in Ahvaz, a city located in the Southwest Iran including 75 patients with clinical diagnosis of fever, headache, vomiting and neck stiffness suspected to have acute meningitis based on cerebrospinal fluid analysis. In the beginning, the patients were divided into two categories as acute meningitis, and non-acute meningitis. Then, 5 L of blood was taken from each patient to determine serum creatine kinase isoenzyme-BB by using ELISA method. After that, the related data including demographics, clinical and laboratory results were analyzed by SPSS software version 16 using Chi-square test for qualitative variables and student’s t-test for quantitative variables. Results: Among the total 75 patients, 37 (49.3% were males and 38 (50.7% were females including 45 patients (60% with acute meningitis and 30 patients (40% without acute meningitis. On the other hand, CK-BB serum levels in acute meningitis and non-acute meningitis patients were 18.23 ± 7.56 and 2.67 ± 1.62, respectively, so significant difference was found between acute meningitis group and non-acute meningitis group (P < 0.000 1. Conclusions: Serum creatine kinase isoenzyme-BB test is a useful test to differentiate acute meningitis from non-acute meningitis among suspected cases of meningitis disease, so measuring the CK-BB serum level in Iran's health system with an expanded health setting especially in remote areas will be useful and helpful in prompt diagnosis and treatment of the acute meningitis.

  14. Linear Analysis of Autonomic Activity and Its Correlation with Creatine Kinase-MB in Overt Thyroid Dysfunctions.

    Science.gov (United States)

    Mavai, Manisha; Singh, Yogendra Raj; Gupta, R C; Mathur, Sandeep K; Bhandari, Bharti

    2018-04-01

    Autonomic activity may be deranged in thyroid dysfunctions and may lead to cardiovascular morbidity and mortality. Myopathy is a common manifestation in thyroid disorders and may be associated with raised serum creatine kinase (CK). We hypothesized that cardiovascular abnormality in thyroid dysfunction may manifest as raised CK-MB. This study was designed to investigate the correlation of CK and its isoform CK-MB with thyroid profile and linear parameters of heart rate variability (HRV). The study was conducted on 35 hypothyroid and hyperthyroid patients each, and 25 age-matched healthy controls. Autonomic activity was assessed by simple short term 5-min HRV. Biochemical evaluation of serum thyroid profile, CK-NAC and CK-MB were estimated in all the subjects. Our results demonstrated low HRV in hypo- as well as hyperthyroid patients. We observed significantly higher serum CK levels in hypothyroid patients when compared to hyperthyroids and controls. However, no significant differences were observed in CK-MB levels in the three groups. Significant positive correlation of CK with TSH and negative correlation with some HRV parameters (LF power, HF power, total power, SDNN, RMSSD) was observed in hypothyroid patients. Whereas correlation of CK-MB with thyroid profile as well as HRV parameters was non-significant in all the groups. Based on the CK and CK-MB findings and their correlation, we conclude that the cardiovascular changes seen in thyroid dysfunctions may primarily be due to autonomic imbalance without apparent cardiac muscle involvement. Whereas, raised CK levels indicate predominantly skeletal muscle involvement in hypothyroid patients.

  15. The Effect of Direct Current Transthoracic Countershock on Human Myocardial Cells Evidenced by Creatine Kinase and Lactic Dehydrogenase Isoenzymes.

    Science.gov (United States)

    1986-05-01

    however, fractionation of these enzymes will identify their specific source. Plasma levels of CK isoenzymes (CKMB) and LDH isoenzymes ( LDHI ) are most...damage--inferred by isoenzyme levels of CKMB and/or LDHI in the serum above normal levels (see definitions of creatine kinase and lactic dehydrogenase

  16. PUTATIVE CREATINE KINASE M-ISOFORM IN HUMAN SPERM IS IDENTIFIED AS THE 70-KILODALTON HEAT SHOCK PROTEIN HSPA2

    Science.gov (United States)

    THE PUTATIVE CREATINE KINASE M-ISOFORM IN HUMAN SPERM IS IDENTIFIED AS THE 70 kDa HEAT SHOCK PROTEIN HSPA2* Gabor Huszar1, Kathryn Stone2, David Dix3 and Lynne Vigue11The Sperm Physiology Laboratory, Department of Obstetrics and Gynecology, 2 W.M. Keck Foundatio...

  17. Mitochondrial affinity for ADP is twofold lower in creatine kinase knock-out muscles - Possible role in rescuing cellular energy homeostasis

    NARCIS (Netherlands)

    ter Veld, F; Jeneson, JAL; Nicolay, K

    Adaptations of the kinetic properties of mitochondria in striated muscle lacking cytosolic (M) and/or mitochondrial (Mi) creatine kinase (CK) isoforms in comparison to wild-type (WT) were investigated in vitro. Intact mitochondria were isolated from heart and gastrocnemius muscle of WT and single-

  18. NMR studies on 15N-labeled creatine (CR), creatinine (CRN), phosphocreatine (PCR), and phosphocreatinine (PCRN), and on barriers to rotation in creatine kinase-bound creatine in the enzymatic reaction

    International Nuclear Information System (INIS)

    Kenyon, G.L.; Reddick, R.E.

    1986-01-01

    Recently, the authors have synthesized 15 N-2-Cr, 15 N-3-Crn, 15 N-2-Crn, 15 N-3-PCrn, 15 N-3-PCr, and 15 N-2-PCr. 1 H, 15 N, 31 P NMR data show that Crn protonates exclusively at the non-methylated ring nitrogen, confirm that PCrn is phosphorylated at the exocyclic nitrogen, and demonstrate that the 31 P- 15 N one-bond coupling constant in 15 N-3-PCr is 18 Hz, not 3 Hz as previously reported by Brindle, K.M., Porteous, R. and Radda, G.K.. The authors have found that creatine kinase is capable of catalyzing the 14 N/ 15 N positional isotope exchange of 3- 15 N-PCr in the presence of MgADP, but not in its absence. Further, the exchange does not take place when labeled PCr is resynthesized exclusively from the ternary complex E X Cr X MgATP as opposed to either E X Cr or free Cr. This suggests that the enzyme both imparts an additional rotational barrier to creatine in the complex and catalyzes the transfer of phosphoryl group with essentially complete regiospecificity

  19. Creatine kinase BB and beta-2-microglobulin as markers of CNS metastases in patients with small-cell lung cancer

    DEFF Research Database (Denmark)

    Pedersen, A G; Bach, F W; Nissen, Mogens Holst

    1985-01-01

    Creatine kinase (CK) and its BB isoenzyme (CK-BB) were measured in CSF in 65 evaluable patients suspected of CNS metastases secondary to small-cell lung cancer (SCLC). In addition, CSF and plasma levels of beta-2-microglobulin (beta-2-m) were measured in a group of 73 evaluable patients. Of the 65...

  20. Phosphocreatine kinetics at the onset of contractions in skeletal muscle of MM creatine kinase knockout mice

    Science.gov (United States)

    Roman, Brian B.; Meyer, Ronald A.; Wiseman, Robert W.

    2002-01-01

    Phosphocreatine (PCr) depletion during isometric twitch stimulation at 5 Hz was measured by (31)P-NMR spectroscopy in gastrocnemius muscles of pentobarbital-anesthetized MM creatine kinase knockout (MMKO) vs. wild-type C57B (WT) mice. PCr depletion after 2 s of stimulation, estimated from the difference between spectra gated to times 200 ms and 140 s after 2-s bursts of contractions, was 2.2 +/- 0.6% of initial PCr in MMKO muscle vs. 9.7 +/- 1.6% in WT muscles (mean +/- SE, n = 7, P muscle after 2 s only if ADP-stimulated oxidative phosphorylation was included in the model. Taken together, the results suggest that cytoplasmic ADP more rapidly increases and oxidative phosphorylation is more rapidly activated at the onset of contractions in MMKO compared with WT muscles.

  1. Changes in Creatine Kinase and Hormones over the Course of an American Football Season.

    Science.gov (United States)

    Stone, J D; Kreutzer, A; Mata, J D; Nystrom, M G; Jagim, A R; Jones, M T; Oliver, J M

    2017-04-04

    The purpose of this study was to examine changes in creatine kinase and hormones over the course of an entire season of American football. A secondary purpose was to determine differences between starters and non-starters. Fasting blood samples were obtained from nineteen National Collegiate Athletic Association Division I (n = 19; 20 ± 1 years) football athletes over the course of a season beginning prior to the start of summer off-season conditioning (T1), before (T2) and after pre-season (T3) football camp, with remaining samples taken throughout the competitive season (T4-T8). A magnitude-based inference approach was used to define outcomes. Testosterone was higher in starters prior to the start of the season (T1, Effect Size [ES] = 0.8) and during pre-conference (T4; ES = 0.7). Post-Camp (T3) testosterone was lower in all players, though greater in starters (starters, 0.0%/0.3%/99.7%; non-starters, 0.2%/2.9%/96.9%). An increase cortisol relative to baseline (T1) was observed in starters early in season (T4, ES = 0.7; T5, ES = 0.5). Creatine kinase was elevated at all time-points in all athletes, with starters having higher circulating levels throughout season. These data demonstrate that changes in hormonal markers may be experienced over a season of football and differ by playing status. Differences between starters and non-starters may be indicative of greater damage and stress experienced by starters, which may result from a greater number of repetitions.

  2. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Oyku Gulmez; Ilyas Atar; Bülent Ozin; Mehmet Emin Korkmaz; Asli Atar; et al

    2008-01-01

    Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Asli Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in pat...

  3. Cardiac Troponin and Creatine Kinase Response to the Three Modes of Training (Running, Pedaling and Swimming in Young Girls

    Directory of Open Access Journals (Sweden)

    Abbas Saremi

    2016-04-01

    Full Text Available Abstract Background: Cardiac troponin T and creatine kinase are used as biological markers for cardiomyocytes and its levels in serum are used as indicators of myocardial cell injury. The purpose of this study was to compare the effects of 3 different training protocols (runing, swimming, and pedaling training on myocardial cell injury biomarkers in young girls. Materials and Methods: In this semi-experimental study with pretest–posttest design, ten healthy young girls (aged 23.0±1.6 y were selected in a convenience sampling way. The subjects performed three types of exercise in 7 days interval. Blood sample was assessed before and after the exercise sessions. Data were analyzed using t-test and analysis of variance. Results: Our results indicated that creatin kinase increased significantly after three types of exercise (p0.05. Conclusion: Our data suggest that intensive exercise is associated with cardiac damage in less trained girls and the type of exercise is determinants of the magnitude of myocardial injury biomarkers release.

  4. Exercise responses in patients with chronically high creatine kinase levels.

    Science.gov (United States)

    Cooper, Christopher B; Dolezal, Brett A; Neufeld, Eric V; Shieh, Perry; Jenner, John R; Riley, Marshall

    2017-08-01

    Elevated serum creatine kinase (CK) is often taken to reflect muscle disease, but many individuals have elevated CK without a specific diagnosis. How elevated CK reflects muscle metabolism during exercise is not known. Participants (46 men, 48 women) underwent incremental exercise testing to assess aerobic performance, cardiovascular response, and ventilatory response. Serum lactate, ammonia, and CK were measured at rest, 4 minutes into exercise, and 2 minutes into recovery. High-CK and control subjects demonstrated similar aerobic capacities and cardiovascular responses to incremental exercise. Those with CK ≥ 300 U/L exhibited significantly higher lactate and ammonia levels after maximal exercise, together with increased ventilatory responses, whereas those with CK ≥200 U/L but ≤ 300 U/L did not. We recommend measurement of lactate and ammonia profiles during a maximal incremental exercise protocol to help identify patients who warrant muscle biopsy to rule out myopathy. Muscle Nerve 56: 264-270, 2017. © 2016 Wiley Periodicals, Inc.

  5. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Gulmez, Oyku

    2008-01-01

    Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Aslı Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in pa...

  6. Extracellular creatine regulates creatine transport in rat and human muscle cells.

    OpenAIRE

    Loike, J D; Zalutsky, D L; Kaback, E; Miranda, A F; Silverstein, S C

    1988-01-01

    Muscle cells do not synthesize creatine; they take up exogenous creatine by specific Na+-dependent plasma membrane transporters. We found that extracellular creatine regulates the level of expression of these creatine transporters in L6 rat muscle cells. L6 myoblasts maintained for 24 hr in medium containing 1 mM creatine exhibited 1/3rd of the creatine transport activity of cells maintained for 24 hr in medium without creatine. Down-regulation of creatine transport was partially reversed whe...

  7. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Gulmez, Oyku; Atar, Ilyas; Ozin, B?lent; Korkmaz, Mehmet Emin; Atar, Asli; Aydinalp, Alp; Yildirir, Aylin; Muderrisoglu, Haldun

    2008-01-01

    Background: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in patients undergoing elective PCI. Methods: A total of 570 patients who underwent PCI were evaluated for CK-MB elevation. Patients who were on CCB therapy when admitted to the hospital constituted the CCB group. ...

  8. Serum creatine kinase elevations in ultramarathon runners at high altitude.

    Science.gov (United States)

    Magrini, Danielle; Khodaee, Morteza; San-Millán, Iñigo; Hew-Butler, Tamara; Provance, Aaron J

    2017-05-01

    Creatine kinase (CK) is a sensitive enzyme marker for muscle damage in athletes. Elevated CK levels have been reported in many endurance physical activities. The consequence and possible long-term sequela of the CK elevation in athletes is unknown. There is a paucity of literature stating actual numerical values of CK associated with competing in an ultramarathon with extreme environmental conditions. Our hypothesis was that the serum CK levels increase significantly as a result of running a 161 km ultramarathon at high altitude. This was a prospective observational study of participants of the Leadville 100 ultramarathon race in Leadville, Colorado at high altitude (2800-3840 m) in August 2014. We collected blood samples from sixty-four volunteer runners before and eighty-three runners immediately after the race. Out of 669 athletes who started the race, 352 successfully completed the race in less than the 30-hour cut-off time (52%). The majority of runners were male (84%). We were able to collect both pre- and post-race blood samples from 36 runners. Out of these 36 runners, the mean pre-race CK was increased from 126 ± 64 U/L to 14,569 ± 14,729 U/L (p athletes' age, BMI, or finishing time. Significant elevation of CK level occurs as a result of running ultramarathons. The majority of athletes with significantly elevated CK levels were asymptomatic and required no major medical attention.

  9. Radioimmunoassay of serum creatine kinase-BB as a tumour marker in breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, R J; Rubery, E D; Jones, H M [Addenbrooke' s Hospital, Cambridge (UK)

    1980-09-27

    Brain type creatine kinase-BB (CPK-BB) was measured by radioimmunoassay in the serum of 113 women with breast cancer and 354 female controls. 80% of women with metastatic breast cancer had levels above 3 ng/ml (control range 0.5-3.7 ng/ml); the highest level was 23 ng/ml. 60% of women with local disease but no evidence of distant metastases showed levels above 3 ng/ml, the highest being 9.0 ng/ml. Of women who had presented with stage I, II, or III disease and postoperatively had no evidence of persistent disease 30% had levels above 3 ng/ml. Serial measurements in 31 patients indicated that the serum CPK-BB correlated with clinical response to treatment.

  10. Radioimmunoassay of serum creatine kinase-BB as a tumour marker in breast cancer

    International Nuclear Information System (INIS)

    Thompson, R.J.; Rubery, E.D.; Jones, H.M.

    1980-01-01

    Brain type creatine kinase-BB (CPK-BB) was measured by radioimmunoassay in the serum of 113 women with breast cancer and 354 female controls. 80% of women with metastatic breast cancer had levels above 3 ng/ml (control range 0.5-3.7 ng/ml); the highest level was 23 ng/ml. 60% of women with local disease but no evidence of distant metastases showed levels above 3 ng/ml, the highest being 9.0 ng/ml. Of women who had presented with stage I, II, or III disease and postoperatively had no evidence of persistent disease 30% had levels above 3 ng/ml. Serial measurements in 31 patients indicated that the serum CPK-BB correlated with clinical response to treatment. (author)

  11. Slaughter value, meat quality, creatine kinase activity and cortisol levels in the blood serum of growing-finishing pigs slaughtered immediately after transport and after a rest period.

    Science.gov (United States)

    Smiecińska, K; Denaburski, J; Sobotka, W

    2011-01-01

    The experimental materials comprised 44 hybrid [female (Polish Large White x Polish Landrace) x male Duroc] growing-finishing pigs. The animals were randomly divided into two groups: 24 pigs were slaughtered immediately after transport and 20 pigs were slaughtered after a 24-hour rest period in the lairage. The meat content of pork carcasses, carcass dressing percentage, the proximate chemical composition, physicochemical and sensory properties of meat and shear force values were determined. Serum creatine kinase activity and cortisol levels were determined in blood samples collected before transport and during carcass bleeding. Pigs slaughtered immediately after transport, compared with those slaughtered after a 24-hour rest period, were characterized by a higher meat content of the carcass and a higher carcass dressing percentage. Pre-slaughter handling had no effect on pork quality. The incidence of normal-quality meat, partially PSE (pale, soft, exudative) meat and PSE meat was similar in both groups. Chemical analysis showed that the content of dry matter, total protein, fat and minerals in meat was comparable in both groups. As regards the functional properties of the pork, samples from the carcasses of pigs that had rested before slaughter had a higher contribution of the red color component. Meat from pigs slaughtered immediately after transport had more desirable sensory properties. Pre-slaughter resting had a significant effect on those analyzed physiological parameters which were found to be good indicators of pre-slaughter stress. Serum creatine kinase activity and cortisol levels were higher in blood samples collected after transport (during carcass bleeding) than in samples collected before transport, pointing to a strong stress response of animals to pre-slaughter treatment. The decrease in serum cortisol levels in blood samples collected during bleeding from the carcasses of pigs slaughtered after a 24-hour rest period, compared with samples

  12. Free creatine available to the creatine phosphate energy shuttle in isolated rat atria

    International Nuclear Information System (INIS)

    Savabi, F.

    1988-01-01

    To measure the actual percentage of intracellular free creatine participating in the process of energy transport, the incorporation of [1- 14 C]creatine into the free creatine and phosphocreatine (PCr) pools in spontaneously beating isolated rat atria, under various conditions, was examined. The atria were subjected to three consecutive periods, control, anoxia, and postanoxic recover, in medium containing tracers of [1- 14 C]creatine. The tissue content and specific activity of creatine and PCr were determined at the end of each period. The higher specific activity found for tissue PCr (1.87 times) than creatine, independent of the percentage of total intracellular creatine that was present as free creatine, provides evidence for the existence of two separate pools of free creatine. Analysis of the data shows that in the normal oxygenated state ∼ 9% of the total intracellular creatine is actually free to participate in the process of energy transport (shuttle pool). About 36% of the total creatine is bound to unknown intracellular components and the rest exists as PCr. The creatine that was taken up and the creatine that was released from the breakdown of PCr have much greater access to the site of phosphorylation than the rest of the intracellular creatine. A sharp increase in the specific activity of residual PCr on prolongation of anoxic time was also observed. This provides evidence for a nonhomogeneous pool of PCr, for the most recently formed (radioactive) PCr appeared to be hydrolyzed last

  13. Temporal changes in serum creatine kinase concentration and degree of muscle rigidity in 24 patients with neuroleptic malignant syndrome

    Directory of Open Access Journals (Sweden)

    Nisijima K

    2013-06-01

    Full Text Available Koichi Nisijima, Katutoshi ShiodaDepartment of Psychiatry, Jichi Medical University, Tochigi, JapanAbstract: Neuroleptic malignant syndrome (NMS is a dangerous adverse response to antipsychotic drugs. It is characterized by the four major clinical symptoms of hyperthermia, severe muscle rigidity, autonomic dysfunction, and altered mental state. Serum creatine kinase (CK elevation occurs in over 90% of NMS cases. In the present study, the detailed temporal changes in serum CK and degree of muscle rigidity, and the relationship between CK concentration and degree of muscle rigidity over the time course from fever onset, were evaluated in 24 affected patients. The results showed that serum CK peaked on day 2 after onset of fever and returned to within normal limits at day 12. Mild muscle rigidity was observed before the onset of fever in 17 of 24 cases (71%. Muscle rigidity was gradually exacerbated and worsened until day 4 after onset of fever. These findings confirm physicians' empirical understanding of serum CK concentrations and muscle rigidity in NMS based on data accumulated from numerous patients with the syndrome, and they indicate that serum CK may contribute to the early detection of NMS.Keywords: neuroleptic malignant syndrome, creatine kinase, muscle rigidity

  14. Mutation of cis-proline 207 in mitochondrial creatine kinase to alanine leads to increased acid stability.

    Science.gov (United States)

    Forstner, M; Müller, A; Rognan, D; Kriechbaum, M; Wallimann, T

    1998-07-01

    We show that the mutation of an uncharged residue far from the active site to another uncharged residue can have effects on the active site without disturbing the overall structure of the protein. Cis-proline 207 of mitochondrial creatine kinase was mutated to alanine. The mutant showed a decrease in the pH-optimum for ATP synthesis by 1.5 units while the maximum relative activity was lowered to 53% of the wild-type enzyme. In the direction of ATP consumption, the pH optimum was lowered by 1.3 units and the maximum relative activity was 49% of the wild-type enzyme. The enzyme kinetic parameters Km and Kd for the substrates did not change dramatically, indicating a largely unperturbed active site. Small-angle X-ray scattering was used to investigate the structural change concomitant with the mutation, yielding a scattering profile only slightly different from that of the wild-type enzyme. Neither the radius of gyration nor the molecular mass showed any significant differences, leading to the conclusion that quarternary organization and fold of the mutant and the wild-type enzymes were similar. Theoretical analysis suggests the most probable primary source of structural change to be a transition of residue 207 peptide bond torsional angle co from the cis to the trans configuration.

  15. Exploratory studies of the potential anti-cancer effects of creatine.

    Science.gov (United States)

    Campos-Ferraz, P L; Gualano, B; das Neves, W; Andrade, I T; Hangai, I; Pereira, R T S; Bezerra, R N; Deminice, R; Seelaender, M; Lancha, A H

    2016-08-01

    Two experiments were performed, in which male Wistar Walker 256 tumor-bearing rats were inoculated with 4 × 10(7) tumor cells subcutaneously and received either creatine (300 mg/kg body weight/day; CR) or placebo (water; PL) supplementation via intragastric gavage. In experiment 1, 50 rats were given PL (n = 22) or CR (n = 22) and a non-supplemented, non-inoculated group served as control CT (n = 6), for 40 days, and the survival rate and tumor mass were assessed. In experiment 2, 25 rats were given CR or PL for 15 days and sacrificed for biochemical analysis. Again, a non-supplemented, non-inoculated group served as control (CT; n = 6). Tumor and muscle creatine kinase (CK) activity and total creatine content, acidosis, inflammatory cytokines, and antioxidant capacity were assessed. Tumor growth was significantly reduced by approximately 30 % in CR when compared with PL (p = 0.03), although the survival rate was not significantly different between CR and PL (p = 0.65). Tumor creatine content tended to be higher in CR than PL (p = 0.096). Tumor CK activity in the cytosolic fraction was higher in CR than PL (p Creatine supplementation was able to slow tumor growth without affecting the overall survival rate, probably due to the re-establishment of the CK-creatine system in cancer cells, leading to attenuation in acidosis, inflammation, and oxidative stress. These findings support the role of creatine as a putative anti-cancer agent as well as help in expanding our knowledge on its potential mechanisms of action in malignancies.

  16. Myocardial Creatine Levels Do Not Influence Response to Acute Oxidative Stress in Isolated Perfused Heart

    Science.gov (United States)

    Aksentijević, Dunja; Zervou, Sevasti; Faller, Kiterie M. E.; McAndrew, Debra J.; Schneider, Jurgen E.; Neubauer, Stefan; Lygate, Craig A.

    2014-01-01

    Background Multiple studies suggest creatine mediates anti-oxidant activity in addition to its established role in cellular energy metabolism. The functional significance for the heart has yet to be established, but antioxidant activity could contribute to the cardioprotective effect of creatine in ischaemia/reperfusion injury. Objectives To determine whether intracellular creatine levels influence responses to acute reactive oxygen species (ROS) exposure in the intact beating heart. We hypothesised that mice with elevated creatine due to over-expression of the creatine transporter (CrT-OE) would be relatively protected, while mice with creatine-deficiency (GAMT KO) would fare worse. Methods and Results CrT-OE mice were pre-selected for creatine levels 20–100% above wild-type using in vivo 1H–MRS. Hearts were perfused in isovolumic Langendorff mode and cardiac function monitored throughout. After 20 min equilibration, hearts were perfused with either H2O2 0.5 µM (30 min), or the anti-neoplastic drug doxorubicin 15 µM (100 min). Protein carbonylation, creatine kinase isoenzyme activities and phospho-PKCδ expression were quantified in perfused hearts as markers of oxidative damage and apoptotic signalling. Wild-type hearts responded to ROS challenge with a profound decline in contractile function that was ameliorated by co-administration of catalase or dexrazoxane as positive controls. In contrast, the functional deterioration in CrT-OE and GAMT KO hearts was indistinguishable from wild-type controls, as was the extent of oxidative damage and apoptosis. Exogenous creatine supplementation also failed to protect hearts from doxorubicin-induced dysfunction. Conclusions Intracellular creatine levels do not influence the response to acute ROS challenge in the intact beating heart, arguing against creatine exerting (patho-)physiologically relevant anti-oxidant activity. PMID:25272153

  17. Effects of obesity on protein kinase C, brain creatine kinase, transcription, and autophagy in cochlea.

    Science.gov (United States)

    Hwang, Juen-Haur

    2017-06-01

    Diet-induced obesity (DIO) has been shown to exacerbate hearing degeneration via increased hypoxia, inflammatory responses, and cell loss via both caspase-dependent and caspase-independent apoptosis signaling pathways. This study aimed to investigate the effects of DIO on the mRNA expressions of protein kinase c-β (PKC-β), brain creatine kinase (CKB), transcription modification genes, and autophagy-related genes in the cochlea of CD/1 mice. Sixteen 4-week-old male CD/1 mice were randomly divided into 2 groups. For 16 weeks, the DIO group was fed a high fat diet (60% kcal fat) and the controls were fed a standard diet. Morphometry, biochemistry, auditory brainstem response thresholds, omental fat, and histopathology of the cochlea were compared. Results showed that body weight, body length, body-mass index, omental fat, plasma triglyceride, and auditory brainstem response thresholds were significantly elevated in the DIO group compared with those of the control group. The ratio of vessel wall thickness to radius in the stria vascularis was significantly higher in the DIO group. The cell densities in the spiral ganglion, but not in the spiral prominence, of the cochlea were significantly lower in the DIO group. The expression of histone deacetylation gene 1 (HDAC1) was significantly higher in the DIO group than the control group. However, the expressions of PKC-β, CKB, HDAC3, histone acetyltransferase gene (P300), lysosome-associated membrane protein 2 (Lamp2), and light chain 3 (Lc3) genes were not significantly different between two groups. These results suggest that DIO might exacerbate hearing degeneration possibly via increased HDAC1 gene expression in the cochlea of CD/1 mice.

  18. Cloning and characterization of the promoter regions from the parent and paralogobs creatine transporter genes

    NARCIS (Netherlands)

    Ndika, J.D.T.; Lusink, V.; Beaubrun, C.; Kanhai, W.; Martinez-Munoz, C.; Jakobs, C.A.J.M.; Salomons, G.S.

    2014-01-01

    Interconversion between phosphocreatine and creatine, catalyzed by creatine kinase is crucial in the supply of ATP to tissues with high energy demand. Creatine's importance has been established by its use as an ergogenic aid in sport, as well as the development of intellectual disability in patients

  19. Microcomputer Assisted Interpretative Reporting of Sequential Creatine Kinase (CK) and Lactate Dehydrogenase (LDH) Isoenzyme Determination

    Science.gov (United States)

    Talamo, Thomas S.; Losos, Frank J.; Mercer, Donald W.

    1984-01-01

    We have developed a microcomputer based system for interpretative reporting of creatine kinase (CK) and lactate dehydrogenase (LDH) isoenzyme studies. Patient demographic data and test results (total CK, CK-MB, LD-1, and LD-2) are entered manually through the keyboard. The test results are compared with normal range values and an interpretative report is generated. This report consists of all pertinent demographic information with a graphic display of up to 12 previous CK and LDH isoenzyme determinations. Diagnostic interpretative statements are printed beneath the graphic display following analysis of previously entered test results. The combination of graphic data display and interpretations based on analysis of up to 12 previous specimens provides useful and accurate information to the cardiologist.

  20. Spatial distribution of "tissue-specific" antigens in the developing human heart and skeletal muscle. I. An immunohistochemical analysis of creatine kinase isoenzyme expression patterns

    NARCIS (Netherlands)

    Wessels, A.; Vermeulen, J. L.; Virágh, S.; Kálmán, F.; Morris, G. E.; Man, N. T.; Lamers, W. H.; Moorman, A. F.

    1990-01-01

    Using monoclonal antibodies against the M and B subunit isoforms of creatine kinase (CK) we have investigated their distribution in developing human skeletal and cardiac muscle immunohistochemically. It is demonstrated that in skeletal muscle, a switch from CK-B to CK-M takes place around the week 8

  1. One- and two-dimensional chemical exchange nuclear magnetic resonance studies of the creatine kinase catalyzed reaction

    International Nuclear Information System (INIS)

    Gober, J.R.

    1988-01-01

    The equilibrium chemical exchange dynamics of the creatine kinase enzyme system were studied by one- and two-dimensional 31 P NMR techniques. Pseudo-first-order reaction rate constants were measured by the saturation transfer method under an array of experimental conditions of pH and temperature. Quantitative one-dimensional spectra were collected under the same conditions in order to calculate the forward and reverse reaction rates, the K eq , the hydrogen ion stoichiometry, and the standard thermodynamic functions. The pure absorption mode in four quadrant two-dimensional chemical exchange experiment was employed so that the complete kinetic matrix showing all of the chemical exchange process could be realized

  2. Pressure overload stimulated cardiac hypertrophy leads to a rapid decrease in the mRNA for creatine kinase

    International Nuclear Information System (INIS)

    Boheler, K.; Popovich, B.; Dillmann, W.H.

    1987-01-01

    Cardiac hypertrophy (CH) leads to a decrease in creatine kinase (CK) enzymatic activity. To determine if the mRNA for CK also decreases with CH, they performed the following studies. Cardiac RNA was isolated from rats subjected to either abdominal aortic stenosis (AS) or sham surgery. Through Northern blot analysis, total cardiac RNA was quantitated with a CK specific 32 P-labelled cDNA clone. At 3 and 8 days post-constriction, the mRNA for CK decreases by 54.6 +/- 7% and 65.3 +/- 18% respectively, whereas the heart weight increases by 19% and 37% relative to controls. Further studies indicate that CK mRNA also decreases by 41.8% in hypothyroid rats (Tx) but decreases by a total of 68.1% in Tx rats subjected to 8 days of AS. Pressure overload stimulated CH leads to a rapid decrease in CK mRNA in normal and Tx rats. This CK mRNA decrease may account for the decreased efficiency of contraction seen in CH

  3. 5' adenosine monophosphate-activated protein kinase, metabolism and exercise.

    Science.gov (United States)

    Aschenbach, William G; Sakamoto, Kei; Goodyear, Laurie J

    2004-01-01

    The 5' adenosine monophosphate-activated protein kinase (AMPK) is a member of a metabolite-sensing protein kinase family that functions as a metabolic 'fuel gauge' in skeletal muscle. AMPK is a ubiquitous heterotrimeric protein, consisting of an alpha catalytic, and beta and gamma regulatory subunits that exist in multiple isoforms and are all required for full enzymatic activity. During exercise, AMPK becomes activated in skeletal muscle in response to changes in cellular energy status (e.g. increased adenosine monophosphate [AMP]/adenosine triphosphate [ATP] and creatine/phosphocreatine ratios) in an intensity-dependent manner, and serves to inhibit ATP-consuming pathways, and activate pathways involved in carbohydrate and fatty-acid metabolism to restore ATP levels. Recent evidence shows that although AMPK plays this key metabolic role during acute bouts of exercise, it is also an important component of the adaptive response of skeletal muscles to endurance exercise training because of its ability to alter muscle fuel reserves and expression of several exercise-responsive genes. This review discusses the putative roles of AMPK in acute and chronic exercise responses, and suggests avenues for future AMPK research in exercise physiology and biochemistry.

  4. Creatine and creatine forms intended for sports nutrition.

    Science.gov (United States)

    Andres, Susanne; Ziegenhagen, Rainer; Trefflich, Iris; Pevny, Sophie; Schultrich, Katharina; Braun, Hans; Schänzer, Wilhelm; Hirsch-Ernst, Karen Ildico; Schäfer, Bernd; Lampen, Alfonso

    2017-06-01

    Creatine is a popular ergogenic supplement in sports nutrition. Yet, supplementation of creatine occasionally caused adverse effects such as gastrointestinal complaints, muscle cramps and an increase in body weight. Creatine monohydrate has already been evaluated by different competent authorities and several have come to the conclusion that a daily intake of 3 g creatine per person is unlikely to pose safety concerns, focusing on healthy adults with exclusion of pregnant and breastfeeding women. Possible vulnerable subgroups were also discussed in relation to the safety of creatine. The present review provides an up-to-date overview of the relevant information with special focus on human studies regarding the safety of creatine monohydrate and other marketed creatine forms, in particular creatine pyruvate, creatine citrate, creatine malate, creatine taurinate, creatine phosphate, creatine orotate, creatine ethyl ester, creatine pyroglutamate, creatine gluconate, and magnesium creatine chelate. Limited data are available with regard to the safety of the latter creatine forms. Considering an acceptable creatine intake of 3 g per day, most of the evaluated creatine forms are unlikely to pose safety concerns, however some safety concerns regarding a supplementary intake of creatine orotate, creatine phosphate, and magnesium creatine chelate are discussed here. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Early detection of response in small cell bronchogenic carcinoma by changes in serum concentrations of creatine kinase, neuron specific enolase, calcitonin, ACTH, serotonin and gastrin releasing peptide

    DEFF Research Database (Denmark)

    Bork, E; Hansen, M; Urdal, P

    1988-01-01

    Creatine kinase (CK-BB), neuron specific enolase (NSE), ACTH, calcitonin, serotonin and gastrin releasing peptide (GRP) were measured in serum or plasma before and immediately after initiation of treatment in patients with small cell lung cancer (SCC). Pretherapeutic elevated concentrations of CK...

  6. The effect of active and passive recovery on creatine kinase and C-reactive protein after an exercise session in football players

    Directory of Open Access Journals (Sweden)

    Mohammad Saeed Mostafavi Darani

    2018-01-01

    Full Text Available Aims: One of the important issues in the context of exercise physiology is apply the best recovery methods completely after intense physical activities. Recovery methods will decrease the risk of muscle damage and subsequent inflammation. The aim of this study is to investigate the recovery procedures on changes of creatine kinase (CK and C-reactive protein (CRP after an exercise session simulated in professional football players. Methods: Thirty Iran's Azadegan League football players were participated in this research (age: 22.4 ± 2.38, height: 179.1 ± 2.63 cm, weight: 68.5 ± 4.82 kg, body mass index: 21.5 ± 2.10 kg/m2. After exercise protocol, simulation team randomly divided the participants into three groups (n = 10 under an active recovery on land, floating in the cold water (10°C, and passive recovery for 12 min. The levels of serum CK and CRP were collected immediately, 24 h, and 48 h after the exercise protocol. Findings: The results showed a significant decrease in CK and CRP after training in cold-water immersion method comparing to the other methods (P < 0.05. In addition, the level of CRP was significantly less than passive recovery 48 h postexercise recovery (P < 0.05. Conclusion: The finding of this study shows that if recovery in cold water was used immediately after exercise, much better results are obtained in future periods. This means that immersion method in cold water probably leads to decreased signs of muscle soreness and inflammatory responses in male football players.

  7. Creatine metabolism and psychiatric disorders: Does creatine supplementation have therapeutic value?

    Science.gov (United States)

    Allen, Patricia J.

    2012-01-01

    Athletes, body builders, and military personnel use dietary creatine as an ergogenic aid to boost physical performance in sports involving short bursts of high-intensity muscle activity. Lesser known is the essential role creatine, a natural regulator of energy homeostasis, plays in brain function and development. Creatine supplementation has shown promise as a safe, effective, and tolerable adjunct to medication for the treatment of brain-related disorders linked with dysfunctional energy metabolism, such as Huntington’s Disease and Parkinson’s Disease. Impairments in creatine metabolism have also been implicated in the pathogenesis of psychiatric disorders, leaving clinicians, researchers and patients alike wondering if dietary creatine has therapeutic value for treating mental illness. The present review summarizes the neurobiology of the creatine-phosphocreatine circuit and its relation to psychological stress, schizophrenia, mood and anxiety disorders. While present knowledge of the role of creatine in cognitive and emotional processing is in its infancy, further research on this endogenous metabolite has the potential to advance our understanding of the biological bases of psychopathology and improve current therapeutic strategies. PMID:22465051

  8. Transcriptional activation of rat creatine kinase B by 17beta-estradiol in MCF-7 cells involves an estrogen responsive element and GC-rich sites.

    Science.gov (United States)

    Wang, F; Samudio, I; Safe, S

    2001-01-01

    The rat creatine kinase B (CKB) gene is induced by estrogen in the uterus, and constructs containing rat CKB gene promoter inserts are highly estrogen-responsive in cell culture. Analysis of the upstream -568 to -523 region of the promoter in HeLa cells has identified an imperfect palindromic estrogen response element (ERE) that is required for hormone inducibility. Analysis of the CKB gene promoter in MCF-7 breast cancer cells confirmed that pCKB7 (containing the -568 to -523 promoter insert) was estrogen-responsive in transient transfection studies. However, mutation and deletion analysis of this region of the promoter showed that two GC-rich sites and the concensus ERE were functional cis-elements that bound estrogen receptor alpha (ERalpha)/Sp1 and ERalpha proteins, respectively. The role of these elements was confirmed in gel mobility shift and chromatin immunoprecipitation assays and transfection studies in MDA-MB-231 and Schneider Drosophila SL-2 cells. These results show that transcriptional activation of CKB by estrogen is dependent, in part, on ERalpha/Sp1 action which is cell context-dependent. Copyright 2001 Wiley-Liss, Inc.

  9. Creatine metabolism and psychiatric disorders: Does creatine supplementation have therapeutic value?

    OpenAIRE

    Allen, Patricia J.

    2012-01-01

    Athletes, body builders, and military personnel use dietary creatine as an ergogenic aid to boost physical performance in sports involving short bursts of high-intensity muscle activity. Lesser known is the essential role creatine, a natural regulator of energy homeostasis, plays in brain function and development. Creatine supplementation has shown promise as a safe, effective, and tolerable adjunct to medication for the treatment of brain-related disorders linked with dysfunctional energy me...

  10. Creatine, Similar to Ketamine, Counteracts Depressive-Like Behavior Induced by Corticosterone via PI3K/Akt/mTOR Pathway.

    Science.gov (United States)

    Pazini, Francis L; Cunha, Mauricio P; Rosa, Julia M; Colla, André R S; Lieberknecht, Vicente; Oliveira, Ágatha; Rodrigues, Ana Lúcia S

    2016-12-01

    Ketamine has emerged as a novel strategy to treat refractory depression, producing rapid remission, but elicits some side effects that limit its use. In an attempt to investigate a safer compound that may afford an antidepressant effect similar to ketamine, this study examined the effects of the ergogenic compound creatine in a model of depression, and the involvement of phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway in its effect. In order to induce a depressive-like behavior, mice were administered with corticosterone (20 mg/kg, per os (p.o.)) for 21 days. This treatment increased immobility time in the tail suspension test (TST), an effect abolished by a single administration of creatine (10 mg/kg, p.o.) or ketamine (1 mg/kg, i.p.), but not by fluoxetine (10 mg/kg, p.o., conventional antidepressant). Treatment of mice with wortmannin (PI3K inhibitor, 0.1 μg/site, intracerebroventricular (i.c.v.)) or rapamycin (mTOR inhibitor, 0.2 nmol/site, i.c.v.) abolished the anti-immobility effect of creatine and ketamine. None of the treatments affected locomotor activity of mice. The immunocontents of p-mTOR, p-p70S6 kinase (p70S6K), and postsynaptic density-95 protein (PSD95) were increased by creatine and ketamine in corticosterone or vehicle-treated mice. Moreover, corticosterone-treated mice presented a decreased hippocampal brain-derived neurotrophic factor (BDNF) level, an effect abolished by creatine or ketamine. Altogether, the results indicate that creatine shares with ketamine the ability to acutely reverse the corticosterone-induced depressive-like behavior by a mechanism dependent on PI3K/AKT/mTOR pathway, and modulation of the synaptic protein PSD95 as well as BDNF in the hippocampus, indicating the relevance of targeting these proteins for the management of depressive disorders. Moreover, we suggest that creatine should be further investigated as a possible fast-acting antidepressant.

  11. Both Creatine and Its Product Phosphocreatine Reduce Oxidative Stress and Afford Neuroprotection in an In Vitro Parkinson’s Model

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    Mauricio Peña Cunha

    2014-10-01

    Full Text Available Creatine is the substrate for creatine kinase in the synthesis of phosphocreatine (PCr. This energetic system is endowed of antioxidant and neuroprotective properties and plays a pivotal role in brain energy homeostasis. The purpose of this study was to investigate the neuroprotective effect of creatine and PCr against 6-hydroxydopamine (6-OHDA-induced mitochondrial dysfunction and cell death in rat striatal slices, used as an in vitro Parkinson’s model. The possible involvement of the signaling pathway mediated by phosphatidylinositol-3 kinase (PI3K, protein kinase B (Akt, and glycogen synthase kinase-3β (GSK3β was also evaluated. Exposure of striatal slices to 6-OHDA caused a significant disruption of the cellular homeostasis measured as 3-(4,5 dimethylthiazol-2-yl-2,5-diphenyl-tetrazolium bromide reduction, lactate dehydrogenase release, and tyrosine hydroxylase levels. 6-OHDA exposure increased the levels of reactive oxygen species and thiobarbituric acid reactive substances production and decreased mitochondrial membrane potential in rat striatal slices. Furthermore, 6-OHDA decreased the phosphorylation of Akt (Serine473 and GSK3β (Serine9. Coincubation with 6-OHDA and creatine or PCr reduced the effects of 6-OHDA toxicity. The protective effect afforded by creatine or PCr against 6-OHDA-induced toxicity was reversed by the PI3K inhibitor LY294002. In conclusion, creatine and PCr minimize oxidative stress in striatum to afford neuroprotection of dopaminergic neurons.

  12. Both Creatine and Its Product Phosphocreatine Reduce Oxidative Stress and Afford Neuroprotection in an In Vitro Parkinson’s Model

    Science.gov (United States)

    Martín-de-Saavedra, Maria D.; Romero, Alejandro; Egea, Javier; Ludka, Fabiana K.; Tasca, Carla I.; Farina, Marcelo; Rodrigues, Ana Lúcia S.; López, Manuela G.

    2014-01-01

    Creatine is the substrate for creatine kinase in the synthesis of phosphocreatine (PCr). This energetic system is endowed of antioxidant and neuroprotective properties and plays a pivotal role in brain energy homeostasis. The purpose of this study was to investigate the neuroprotective effect of creatine and PCr against 6-hydroxydopamine (6-OHDA)-induced mitochondrial dysfunction and cell death in rat striatal slices, used as an in vitro Parkinson’s model. The possible involvement of the signaling pathway mediated by phosphatidylinositol-3 kinase (PI3K), protein kinase B (Akt), and glycogen synthase kinase-3β (GSK3β) was also evaluated. Exposure of striatal slices to 6-OHDA caused a significant disruption of the cellular homeostasis measured as 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide reduction, lactate dehydrogenase release, and tyrosine hydroxylase levels. 6-OHDA exposure increased the levels of reactive oxygen species and thiobarbituric acid reactive substances production and decreased mitochondrial membrane potential in rat striatal slices. Furthermore, 6-OHDA decreased the phosphorylation of Akt (Serine473) and GSK3β (Serine9). Coincubation with 6-OHDA and creatine or PCr reduced the effects of 6-OHDA toxicity. The protective effect afforded by creatine or PCr against 6-OHDA-induced toxicity was reversed by the PI3K inhibitor LY294002. In conclusion, creatine and PCr minimize oxidative stress in striatum to afford neuroprotection of dopaminergic neurons. PMID:25424428

  13. Sequential Events in the Irreversible Thermal Denaturation of Human Brain-Type Creatine Kinase by Spectroscopic Methods

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    Yan-Song Gao

    2010-06-01

    Full Text Available The non-cooperative or sequential events which occur during protein thermal denaturation are closely correlated with protein folding, stability, and physiological functions. In this research, the sequential events of human brain-type creatine kinase (hBBCK thermal denaturation were studied by differential scanning calorimetry (DSC, CD, and intrinsic fluorescence spectroscopy. DSC experiments revealed that the thermal denaturation of hBBCK was calorimetrically irreversible. The existence of several endothermic peaks suggested that the denaturation involved stepwise conformational changes, which were further verified by the discrepancy in the transition curves obtained from various spectroscopic probes. During heating, the disruption of the active site structure occurred prior to the secondary and tertiary structural changes. The thermal unfolding and aggregation of hBBCK was found to occur through sequential events. This is quite different from that of muscle-type CK (MMCK. The results herein suggest that BBCK and MMCK undergo quite dissimilar thermal unfolding pathways, although they are highly conserved in the primary and tertiary structures. A minor difference in structure might endow the isoenzymes dissimilar local stabilities in structure, which further contribute to isoenzyme-specific thermal stabilities.

  14. Role of quaternary structure in muscle creatine kinase stability: tryptophan 210 is important for dimer cohesion.

    Science.gov (United States)

    Perraut, C; Clottes, E; Leydier, C; Vial, C; Marcillat, O

    1998-07-01

    A mutant of the dimeric rabbit muscle creatine kinase (MM-CK) in which tryptophan 210 was replaced has been studied to assess the role of this residue in dimer cohesion and the importance of the dimeric state for the native enzyme stability. Wild-type protein equilibrium unfolding induced by guanidine hydrochloride occurs through intermediate states with formation of a molten globule and a premolten globule. Unlike the wild-type enzyme, the mutant inactivates at lower denaturant concentration and the loss of enzymatic activity is accompanied by the dissociation of the dimer into two apparently compact monomers. However, the Stokes radius of the monomer increases with denaturant concentration as determined by size exclusion chromatography, indicating that, upon monomerization, the protein structure is destabilized. Binding of 8-anilinonaphthalene-1-sulfonate shows that the dissociated monomer exposes hydrophobic patches at its surface, suggesting that it could be a molten globule. At higher denaturant concentrations, both wild-type and mutant follow similar denaturation pathways with formation of a premolten globule around 1.5-M guanidine, indicating that tryptophan 210 does not contribute to a large extent to the monomer conformational stability, which may be ensured in the dimeric state through quaternary interactions.

  15. Creatine in the central nervous system: From magnetic resonance spectroscopy to creatine deficiencies.

    Science.gov (United States)

    Rackayova, Veronika; Cudalbu, Cristina; Pouwels, Petra J W; Braissant, Olivier

    2017-07-15

    Creatine (Cr) is an important organic compound acting as intracellular high-energy phosphate shuttle and in energy storage. While located in most cells where it plays its main roles in energy metabolism and cytoprotection, Cr is highly concentrated in muscle and brain tissues, in which Cr also appears to act in osmoregulation and neurotransmission. This review discusses the basis of Cr metabolism, synthesis and transport within brain cells. The importance of Cr in brain function and the consequences of its impaired metabolism in primary and secondary Cr deficiencies are also discussed. Cr and phosphocreatine (PCr) in living systems can be well characterized using in vivo magnetic resonance spectroscopy (MRS). This review describes how 1 H MRS allows the measurement of Cr and PCr, and how 31 P MRS makes it possible to estimate the creatine kinase (CK) rate constant and so detect dynamic changes in the Cr/PCr/CK system. Absolute quantification by MRS using creatine as internal reference is also debated. The use of in vivo MRS to study brain Cr in a non-invasive way is presented, as well as its use in clinical and preclinical studies, including diagnosis and treatment follow-up in patients. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Creatine kinase and alpha-actin mRNA levels decrease in diabetic rat hearts

    International Nuclear Information System (INIS)

    Popovich, B.; Barrieux, A.; Dillmann, W.H.

    1987-01-01

    Diabetic cardiomyopathy is associated with cardiac atrophy and isoenzyme redistribution. To determine if tissue specific changes occur in mRNAs coding for α-actin and creatine kinase (CK), they performed RNA blot analysis. Total ventricular RNA from control (C) and 4 wk old diabetic (D) rats were hybridized with 32 P cDNA probes for α-actin and CK. A tissue independent cDNA probe, CHOA was also used. Signal intensity was quantified by photodensitometry. D CK mRNA was 47 +/- 16% lower in D vs C. Insulin increases CK mRNA by 20% at 1.5 hs, and completely reverses the deficit after 4 wks. D α-actin mRNA is 66 +/- 18% lower in D vs C. Insulin normalized α-actin mRNA by 5 hs. CHOA mRNA is unchanged in D vs C, but D + insulin CHOA mRNA is 30 +/- 2% lower than C. In rats with diabetic cardiomyopathy, muscle specific CK and α-actin mRNAs are decreased. Insulin treatment reverses these changes

  17. Delirium and High Creatine Kinase and Myoglobin Levels Related to Synthetic Cannabinoid Withdrawal

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    Ahmet Bulent Yazici

    2017-01-01

    Full Text Available Synthetic cannabinoids (SCs are included in a group of drugs called new psychoactive substances. Effects of SCs on the central nervous system are similar to other cannabinoids, but 2–100 times more potent than marijuana. Thus, addiction and withdrawal symptoms are more severe than natural cannabinoids. Withdrawal symptoms of SCs were reported in the literature previously. But there is no report about SC withdrawal delirium and its treatment. Several studies reported that agonists of CB1 receptors play a role in GABA and glutamatergic neurotransmission, which is similar to the effects of alcohol on GABA and glutamatergic receptors. Previous studies on alcohol delirium cases suggested that elevated creatine kinase (CK can be a marker of progress. This study reports delirium and high serum CK levels related to SC withdrawal and offers a treatment with benzodiazepine for them. We described two cases treated in our inpatient clinic about SC withdrawal with increase of serum CK level and other laboratory parameters. One of them demonstrated delirium symptoms and the other did not with early rapid treatment.

  18. Creatine biosynthesis and transport by the term human placenta.

    Science.gov (United States)

    Ellery, Stacey J; Della Gatta, Paul A; Bruce, Clinton R; Kowalski, Greg M; Davies-Tuck, Miranda; Mockler, Joanne C; Murthi, Padma; Walker, David W; Snow, Rod J; Dickinson, Hayley

    2017-04-01

    Creatine is an amino acid derivative that is involved in preserving ATP homeostasis. Previous studies suggest an important role for the creatine kinase circuit for placental ATP turnover. Creatine is obtained from both the diet and endogenous synthesis, usually along the renal-hepatic axis. However, some tissues with a high-energy demand have an inherent capacity to synthesise creatine. In this study, we determined if the term human placenta has the enzymatic machinary to synthesise creatine. Eleven placentae were collected following elective term caesarean section. Samples from the 4 quadrants of each placenta were either fixed in formalin or frozen. qPCR was used to determine the mRNA expression of the creatine synthesising enzymes arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT), and the creatine transporter (SLC6A8). Protein expression of AGAT and GAMT was quantified by Western blot, and observations of cell localisation of AGAT, GAMT and SLC6A8 made with immunohistochemistry. Synthesis of guanidinoacetate (GAA; creatine precursor) and creatine in placental homogenates was determined via GC-MS and HPLC, respectively. AGAT, GAMT and SLC6A8 mRNA and protein were detected in the human placenta. AGAT staining was identified in stromal and endothelial cells of the fetal capillaries. GAMT and SLC6A8 staining was localised to the syncytiotrophoblast of the fetal villi. Ex vivo, tissue homogenates produce both GAA (4.6 nmol mg protein -1 h -1 ) and creatine (52.8 nmol mg protein -1 h -1 ). The term human placenta has the capacity to synthesise creatine. These data present a new understanding of placental energy metabolism. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Cloning and characterization of the promoter regions from the parent and paralogous creatine transporter genes.

    Science.gov (United States)

    Ndika, Joseph D T; Lusink, Vera; Beaubrun, Claudine; Kanhai, Warsha; Martinez-Munoz, Cristina; Jakobs, Cornelis; Salomons, Gajja S

    2014-01-10

    Interconversion between phosphocreatine and creatine, catalyzed by creatine kinase is crucial in the supply of ATP to tissues with high energy demand. Creatine's importance has been established by its use as an ergogenic aid in sport, as well as the development of intellectual disability in patients with congenital creatine deficiency. Creatine biosynthesis is complemented by dietary creatine uptake. Intracellular transport of creatine is carried out by a creatine transporter protein (CT1/CRT/CRTR) encoded by the SLC6A8 gene. Most tissues express this gene, with highest levels detected in skeletal muscle and kidney. There are lower levels of the gene detected in colon, brain, heart, testis and prostate. The mechanism(s) by which this regulation occurs is still poorly understood. A duplicated unprocessed pseudogene of SLC6A8-SLC6A10P has been mapped to chromosome 16p11.2 (contains the entire SLC6A8 gene, plus 2293 bp of 5'flanking sequence and its entire 3'UTR). Expression of SLC6A10P has so far only been shown in human testis and brain. It is still unclear as to what is the function of SLC6A10P. In a patient with autism, a chromosomal breakpoint that intersects the 5'flanking region of SLC6A10P was identified; suggesting that SLC6A10P is a non-coding RNA involved in autism. Our aim was to investigate the presence of cis-acting factor(s) that regulate expression of the creatine transporter, as well as to determine if these factors are functionally conserved upstream of the creatine transporter pseudogene. Via gene-specific PCR, cloning and functional luciferase assays we identified a 1104 bp sequence proximal to the mRNA start site of the SLC6A8 gene with promoter activity in five cell types. The corresponding 5'flanking sequence (1050 bp) on the pseudogene also had promoter activity in all 5 cell lines. Surprisingly the pseudogene promoter was stronger than that of its parent gene in 4 of the cell lines tested. To the best of our knowledge, this is the first

  20. Individual analysis of creatine kinase concentration in Brazilian elite soccer players

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    Adriano Lima Alves

    2015-04-01

    Full Text Available OBJECTIVE: to determine the individual profile of blood concentration of creatine kinase CK in elite soccer players as well as to analyze the CK concentrations in different periods during the Professional Brazilian Championship. METHODS: resting CK of 17 soccer players was evaluated before the competition pre-season and after the matches 36 and 46 hours after the games CKGame for the individual blood CK. The Chi-square test was used to analyze the individual CK during the season. The competitive season was divided into three periods: initial, intermediate and final. The one-way ANOVA with repeated measurements followed by post hoc Student-Newman-Keuls test was used to compare the individual CK of each soccer player in each competitive period. The significance level was set at p<0.05. RESULTS: the highest frequency of individual CK was found in the second quartile 71 observations and the lowest frequency in the first 26 observations and the fourth quartile 40 observations compared to the expected number of 45.8 x2=22.21. CK concentrations were lower in the intermediate mean=66.99% and final mean=60.21% periods than in the initial period mean=89.33%. CONCLUSION: soccer players did not show elevated muscle damage and probably a muscle adaptation occurred in the competition, due to the reduction of CK concentrations observed.

  1. Over-expression of mitochondrial creatine kinase in the murine heart improves functional recovery and protects against injury following ischaemia-reperfusion.

    Science.gov (United States)

    Whittington, Hannah J; Ostrowski, Philip J; McAndrew, Debra J; Cao, Fang; Shaw, Andrew; Eykyn, Thomas R; Lake, Hannah; Tyler, Jack; Schneider, Jurgen E; Neubauer, Stefan; Zervou, Sevasti; Lygate, Craig A

    2018-03-02

    Mitochondrial creatine kinase (MtCK) couples ATP production via oxidative phosphorylation to phosphocreatine in the cytosol, which acts as a mobile energy store available for regeneration of ATP at times of high demand. We hypothesised that elevating MtCK would be beneficial in ischaemia-reperfusion (I/R) injury. Mice were created overexpressing the sarcomeric MtCK gene with αMHC promoter at the Rosa26 locus (MtCK-OE) and compared with wild-type (WT) littermates. MtCK activity was 27% higher than WT, with no change in other CK isoenzymes or creatine levels. Electron microscopy confirmed normal mitochondrial cell density and mitochondrial localisation of transgenic protein. Respiration in isolated mitochondria was unaltered and metabolomic analysis by 1H-NMR suggests that cellular metabolism was not grossly affected by transgene expression. There were no significant differences in cardiac structure or function under baseline conditions by cine-MRI or LV haemodynamics. In Langendorff-perfused hearts subjected to 20min ischaemia and 30 min reperfusion, MtCK-OE exhibited less ischaemic contracture and improved functional recovery (Rate pressure product 58% above WT; P < 0.001). These hearts had reduced myocardial infarct size, which was confirmed in vivo: 55±4% in WT vs 29±4% in MtCK-OE; P < 0.0001). Isolated cardiomyocytes from MtCK-OE hearts exhibited delayed opening of the mitochondrial permeability transition pore (mPTP) compared to WT, which was confirmed by reduced mitochondrial swelling in response to calcium. There was no detectable change in the structural integrity of the mitochondrial membrane. Modest elevation of MtCK activity in the heart does not adversely affect cellular metabolism, mitochondrial or in vivo cardiac function, but modifies mPTP opening to protect against I/R injury and improve functional recovery. Our findings support MtCK as a prime therapeutic target in myocardial ischaemia.

  2. Plasma Creatine Kinetics After Ingestion of Microencapsulated Creatine Monohydrate with Enhanced Stability in Aqueous Solutions.

    Science.gov (United States)

    Hone, Michelle; Kent, Robert M; Scotto di Palumbo, Alessandro; Bleiel, Sinead B; De Vito, Giuseppe; Egan, Brendan

    2017-07-04

    Creatine monohydrate represents one of the largest sports supplement markets. Enhancing creatine (CRE) stability in aqueous solutions, such as with microencapsulation, represents innovation potential. Ten physically active male volunteers were randomly assigned in a double-blind design to either placebo (PLA) (3-g maltodextrin; n = 5) or microencapsulated CRE (3-g creatine monohydrate; n = 5) conditions. Experimental conditions involved ingestion of the samples in a 70-mL ready-to-drink format. CRE was delivered in a novel microencapsulation matrix material consisting entirely of hydrolyzed milk protein. Three hours after ingestion, plasma creatine concentrations were unchanged during PLA, and averaged ∼45 μM. During CRE, plasma creatine concentration peaked after 30 min at 101.6 ± 14.9 μM (p creatine concentration gradually trended downwards but remained significantly elevated (∼50% above resting levels) 3 hr after ingestion. These results demonstrate that the microencapsulated form of creatine monohydrate reported herein remains bioavailable when delivered in aqueous conditions, and has potential utility in ready-to-drink formulations for creatine supplementation.

  3. Lean body mass and creatine kinase are associated with reduced inflammation in obesity.

    Science.gov (United States)

    Bekkelund, Svein I; Jorde, Rolf

    2017-11-01

    Obesity is associated with inflammation, but the role of lean mass and creatine kinase (CK) on the inflammatory process is less known. We investigated the associations between lean mass, CK and fat mass upon inflammatory parameters in an overweight and obese adult population. Body composition examined by dual-energy X-ray absorptiometry, high-sensitivity C-reactive protein (hs-CRP), erythrocyte sedimentation rate (ESR), CK and supplementary clinical parameters were measured in 454 overweight and obese individuals. This is a secondary analysis from a cohort of obese individuals treated with Vitamin D. Mean age was 47·6 ± 11·4 years and mean body mass index 34·6 ± 3·9 kg/m 2 . Lean mass correlated negatively with hs-CRP (r = -0·127, P = 0·042) and ESR (r = -0·381, P lean mass in the lower ESR quartile was significantly higher than in the upper quartile (P lean mass and CK in an overweight and obese population. Hypothetically, lean mass has a favourable effect on obesity-related inflammation, and CK may play a role as an inhibitor of inflammation in obesity. © 2017 Stichting European Society for Clinical Investigation Journal Foundation.

  4. Radioimmunoassay of serum creatine kinase BB as index of brain damage after head injury

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, J P; Jones, H M; Hitchcock, R; Adams, N; Thompson, R J [Addenbrooke' s Hospital, Cambridge (UK)

    1980-09-20

    Brain-type creatine kinase isoenzyme (CK-BB) was measured by radioimmunoassay in the serum of 54 patients with head injuries. CK-BB was not detectable in 476 out of 1006 controls, the remaining 530 normal samples containing a mean of 1.5 +- SDO.75 ..mu..g/l. The mean CK-BB concentrations in patients with mild, moderate, and fatal head injuries were all significantly higher than the control value (p<0.01 in each instance). Patients with serious head injury had serum concentrations many times the normal value, in two cases within 30 minutes after impact. Fatally injured patients continued to have high serum concentrations several days after injury. In less serious cases values approached normal within two or three days. Every patient with evidence of cerebral laceration, bruising, or swelling had a serum CK-BB concentration above normal. Raised concentrations were found in 14 out of 22 patients with concussion only. Thus the serum CK-BB concentration appears to be a sensitive index of brain damage and may prove useful in the management and follow-up of head-injured patients.

  5. Radioimmunoassay of serum creatine kinase BB as index of brain damage after head injury

    International Nuclear Information System (INIS)

    Phillips, J.P.; Jones, H.M.; Hitchcock, R.; Adams, N.; Thompson, R.J.

    1980-01-01

    Brain-type creatine kinase isoenzyme (CK-BB) was measured by radioimmunoassay in the serum of 54 patients with head injuries. CK-BB was not detectable in 476 out of 1006 controls, the remaining 530 normal samples containing a mean of 1.5 +- SDO.75 μg/l. The mean CK-BB concentrations in patients with mild, moderate, and fatal head injuries were all significantly higher than the control value (p<0.01 in each instance). Patients with serious head injury had serum concentrations many times the normal value, in two cases within 30 minutes after impact. Fatally injured patients continued to have high serum concentrations several days after injury. In less serious cases values approached normal within two or three days. Every patient with evidence of cerebral laceration, bruising, or swelling had a serum CK-BB concentration above normal. Raised concentrations were found in 14 out of 22 patients with concussion only. Thus the serum CK-BB concentration appears to be a sensitive index of brain damage and may prove useful in the management and follow-up of head-injured patients. (author)

  6. Molecular Characterization and Expression Analysis of Creatine Kinase Muscle (CK-M) Gene in Horse.

    Science.gov (United States)

    Do, Kyong-Tak; Cho, Hyun-Woo; Badrinath, Narayanasamy; Park, Jeong-Woong; Choi, Jae-Young; Chung, Young-Hwa; Lee, Hak-Kyo; Song, Ki-Duk; Cho, Byung-Wook

    2015-12-01

    Since ancient days, domestic horses have been closely associated with human civilization. Today, horse racing is an important industry. Various genes involved in energy production and muscle contraction are differentially regulated during a race. Among them, creatine kinase (CK) is well known for its regulation of energy preservation in animal cells. CK is an iso-enzyme, encoded by different genes and expressed in skeletal muscle, heart, brain and leucocytes. We confirmed that the expression of CK-M significantly increased in the blood after a 30 minute exercise period, while no considerable change was observed in skeletal muscle. Analysis of various tissues showed an ubiquitous expression of the CK-M gene in the horse; CK-M mRNA expression was predominant in the skeletal muscle and the cardiac muscle compared to other tissues. An evolutionary study by synonymous and non-synonymous single nucleotide polymorphism ratio of CK-M gene revealed a positive selection that was conserved in the horse. More studies are warranted in order to develop the expression of CK-M gene as a biomarker in blood of thoroughbred horses.

  7. Determinação das atividades séricas de creatina quinase, lactato desidrogenase e aspartato aminotransferase em eqüinos de diferentes categorias de atividade Determination of serum activities of creatine kinase, lactate dehydrogenase, and aspartate aminotransferase in horses of different activities classes

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    I.A. Câmara e Silva

    2007-02-01

    Full Text Available The creatine kinase (CK, lactate dehydrogenase (LDH, and aspartate aminotransferase (AST seric activities in horses of different activity classes (athlete, traction, and reproduction, were compared. Fifty-eight horses were alloted into three groups - group 1 with 20 athletes, "vaquejada" competitors; group 2 with 20 breeding horses; and group 3 with 18 draft horses, averaging 10 working hours daily. The average values for CK serum activity were 80.2, 83.9, and 94.4 U/l in groups 1, 2, and 3, respectively. Result of group 3 was significantly different from the other groups. The averages values for LDH were 102.5, 98.6, and 112.8 U/l in groups 1, 2, and 3, respectively, with no statistical difference between groups. The AST averages were 56.8, 33.0, and 50.1 U/l in groups 1, 2, and 3, respectively, with group 2 significantly differing from the others. Clinical biochemistry values of muscular function in horses varied according to activity category.

  8. Comparison of early myocardial technetium-99m pyrophosphate uptake to early peaking of creatine kinase and creatine kinase-MB as indicators of early reperfusion in acute myocardial infarction

    International Nuclear Information System (INIS)

    Kondo, M.; Yuzuki, Y.; Arai, H.; Shimizu, K.; Morikawa, M.; Shimono, Y.

    1987-01-01

    The value of technetium-99m pyrophosphate (Tc-99m-PYP) scintigraphy as an indicator of reperfusion 2.8 to 8 hours after the onset of symptoms of acute myocardial infarction was compared with the value of early peak creatine kinase (CK) and CK-MB release within 16 hours after the onset of symptoms. In 29 patients who received thrombolytic therapy, recanalization was seen (group 1) and in 7 it was not (group 2). In 23 patients (79%) in group 1 scintigraphic findings were positive and in all 7 in group 2 they were negative. In 15 patients (52%) in group 1 and 1 patient (14%) in group 2, CK reached its peak level within 16 hours. In 20 patients (69%) in group 1 and 3 (43%) in group 2 the CK-MB level reached a peak within 16 hours. The sensitivity, specificity and predictive accuracy of positive results of early Tc-99m-PYP scintigraphy in predicting the reperfusion were 79%, 100% and 83%. These values are significantly higher than or similar to those of early peaking of CK and CK-MB release. In contrast to measurements of enzyme release, reperfusion data for Tc-99m-PYP scintigraphy are available immediately after thrombolytic therapy. Therefore, early Tc-99m-PYP scintigraphy (3 to 8 hours after onset of symptoms) is valuable as a noninvasive technique for early diagnosis of reperfusion

  9. Creatine salts provide neuroprotection even after partial impairment of the creatine transporter.

    Science.gov (United States)

    Adriano, E; Garbati, P; Salis, A; Damonte, G; Millo, E; Balestrino, M

    2017-01-06

    Creatine, a compound that is critical for energy metabolism of nervous cells, crosses the blood-brain barrier (BBB) and the neuronal plasma membrane with difficulty, and only using its specific transporter. In the hereditary condition where the creatine transporter is defective (creatine transporter deficiency) there is no creatine in the brain, and administration of creatine is useless lacking the transporter. The disease is severe and incurable. Creatine-derived molecules that could cross BBB and plasma membrane independently of the transporter might be useful to cure this condition. Moreover, such molecules could be useful also in stroke and other brain ischemic conditions. In this paper, we investigated three creatine salts, creatine ascorbate, creatine gluconate and creatine glucose. Of these, creatine glucose was ineffective after transporter block with guanidine acetic acid (GPA) administration. Creatine ascorbate was not superior to creatine in increasing tissue creatine and phosphocreatine content after transporter impairment, however even after such impairment it delayed synaptic failure during anoxia. Finally, creatine gluconate was superior to creatine in increasing tissue content of creatine after transporter block and slowed down PS disappearance during anoxia, an effect that creatine did not have. These findings suggest that coupling creatine to molecules having a specific transporter may be a useful strategy in creatine transporter deficiency. In particular, creatine ascorbate has effects comparable to those of creatine in normal conditions, while being superior to it under conditions of missing or impaired creatine transporter. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  10. Correlation of Creatine Kinase Levels with Clinical Features and Survival in Amyotrophic Lateral Sclerosis

    Directory of Open Access Journals (Sweden)

    Hongfei Tai

    2017-07-01

    Full Text Available ObjectiveTo evaluate serum creatine kinase (CK levels of amyotrophic lateral sclerosis (ALS patients and to explore the relationship between CK levels and the clinical characteristics and survival prognosis of ALS patients.MethodsWe analyzed the CK levels of 185 ALS patients who underwent long-term follow-up. The relationship between CK levels and clinical features including sex, age, disease duration, site of onset, body mass index (BMI, serum creatinine (Cr, and spontaneous electromyographic activity was analyzed by univariate analysis and multiple linear regression. Kaplan–Meier and Cox proportional hazards models were used to explore whether CK levels were independently correlated with survival prognosis of ALS.ResultsBaseline serum CK was raised in 43% of participants. The median CK level was 160 U/L (range: 20–2,574 U/L, and 99% of patients had a baseline serum CK level less than 1,000 U/L. CK levels were significantly higher in male patients than in female patients [204 (169 versus 117 (111 U/L, p < 0.001] and in patients with limb onset ALS than with bulbar onset ALS (p < 0.001. CK levels were also correlated with serum Cr (p = 0.011 and the spontaneous potential score of electromyography (EMG (p = 0.037 but not correlated with age (p = 0.883, disease duration (p = 0.116, or BMI (p = 0.481. Log CK was independently correlated with survival of ALS patients (HR = 0.457, 95% confidence interval 0.221–0.947, p = 0.035 after adjusting for age, sex, site of onset, serum Cr, and BMI.ConclusionSerum CK levels of ALS patients were correlated with sex, site of onsite, serum Cr, and spontaneous activity in EMG. Serum CK could be an independent prognostic factor for survival of ALS patients.

  11. [Effect of low-intensity 900 MHz frequency electromagnetic radiation on rat liver and blood serum enzyme activities].

    Science.gov (United States)

    Nersesova, L S; Petrosian, M S; Gazariants, M G; Mkrtchian, Z S; Meliksetian, G O; Pogosian, L G; Akopian, Zh I

    2014-01-01

    The comparative analysis of the rat liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and purine nucleoside phosphorylase post-radiation activity levels after a total two-hour long single and fractional exposure of the animals to low-intensity 900 MHz frequency electromagnetic field showed that the most sensitive enzymes to the both schedules of radiation are the liver creatine kinase, as well as the blood serum creatine kinase and alkaline phosphatase. According to the comparative analysis of the dynamics of changes in the activity level of the liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase and purine nucleoside phosphorylase, both single and fractional radiation schedules do not affect the permeability of a hepatocyte cell membrane, but rather cause changes in their energetic metabolism. The correlation analysis of the post-radiation activity level changes of the investigated enzymes did not reveal a clear relationship between them. The dynamics of post-radiation changes in the activity of investigated enzyme levels following a single and short-term fractional schedules of radiation did not differ essentially.

  12. Use of plasma creatine kinase pharmacokinetics to estimate the amount of excercise-induced muscle damage in Beagles.

    Science.gov (United States)

    Chanoit, G P; Lefebvre, H P; Orcel, K; Laroute, V; Toutain, P L; Braun, J P

    2001-09-01

    To assess the effects of moderate exercise on plasma creatine kinase (CK) pharmacokinetics and to estimate exercise-induced muscle damage in dogs. 6 untrained adult Beagles. The study was divided into 3 phases. In phase 1, dogs ran for 1 hour at a speed of 9 km/h, and samples were used to determine the area under the plasma CK activity versus time curve (AUC) induced by exercise. In phases 2 and 3, pharmacokinetics of CK were calculated in dogs during exercise and at rest, respectively. Values for AUC and plasma clearance (CI) were used to estimate muscle damage. At rest, values for Cl, steady-state volume of distribution (Vdss), and mean retention time (MRT) were 0.32+/-0.02 ml/kg of body weight/min, 57+/-173 ml/kg, and 3.0+/-0.57 h, respectively. During exercise, Cl decreased significantly (0.26+/-0.03 ml/kg/min), MRT increased significantly, (4.4+/-0.97 h), and Vdss remained unchanged. Peak of plasma CK activity (151+/-58.8 U/L) was observed 3 hours after completion of exercise. Estimated equivalent amount of muscle corresponding to the quantity of CK released was 41+/-29.3 mg/kg. These results revealed that exercise had a minor effect on CK disposition and that the equivalent amount of muscle damaged by moderate exercise was negligible. This study illustrates the relevance for use of the minimally invasive and quantitative pharmacokinetic approach when estimating muscle damage.

  13. [Creatine kinase BB and lactate in the cerebrospinal fluid of neonates and infants with perinatal injuries of the CNS].

    Science.gov (United States)

    Alatyrtsev, V V; Iakunin, Iu A; Burkova, A S; Podkopaev, V N; Afonina, L G

    1989-01-01

    A study was made of the content of creatine kinase-BB (CK-BB) and lactate in cerebrospinal fluid (CSF) of 202 neonates and infants with perinatal CNS injuries. The relationship was found between the rise of the CK-BB content and the gravity of perinatal CNS injuries. The highest content of CK-BB in CSF was marked in neonates with cerebral disorders complicated by infectious and inflammatory diseases (pneumonia, sepsis). Within the first 5 days of life, the children of this group demonstrated the relationship between the content of CK-BB and lactate of CSF. The measurement of the content of CK-BB in CSF should be used for early diagnosis, assessment of the gravity and course of perinatal CNS injuries in neonates and in infants.

  14. Musculus soleus of rats at physical activity and L-carnitine and creatine phosphate effect

    Directory of Open Access Journals (Sweden)

    Irina A. Khutorskaya

    2017-09-01

    Full Text Available Introduction: The study of the effect of metabolic drugs on the histochemical characteristics of soleus muscle is relevant for solving the problem of providing the training process in Russia with non-doping drugs for safe correction of the consequences of intense physical activity in athletes. Materials and Methods: Dynamic physical activity in rats (n = 24 was simulated by swimming “to the limit” with weighting of 10 % of body weight (20 days, 1 time per day. The experimental animals were divided into four groups (6 animals each: № 1 – control, № 2 – swimming + isotonic NaCl solution, № 3 and № 4 – swimming + L-carnitine or creatine phosphate 100.0 mg/kg daily intraperitoneally. The object of the study was musculus soleus. Differentiation of muscle fibers was carried out by the intensity of histochemical activity of succinate dehydrogenase (SDG and alkaline stable adenosine triphosphate (ATP of myosin. The percentage of muscle fibers was evaluated and their diameter was defined by the direct morphometry. The obtained data were treated statistically by Student’s T-test. Results: Swimming of the animals “to the limit” do not affect the ratio of fibers with different phenotypes in the soleus muscle. This indicator is genetically determined and was not modified by L-carnitine and creatine phosphate. Dynamic physical activity promotes the development of hypertrophy of muscle fibers of various types. The investigated medicaments of the metabolic type either do not influence on the formation of exerciseinduced hypertrophy (predominantly creatine phosphate or reduce the intensity of the hypertrophic process (predominantly L-carnitine under dynamic physical activity. Discussion and Conclusions: The obtained data indicate L-carnitine and creatine phosphate do not have an anabolic effect. Taking into account the relevant data on ability of L-carnitine and creatine phosphate to effectively correct a negative effects of intensive

  15. syk kinase activation by a src kinase-initiated activation loop phosphorylation chain reaction

    Science.gov (United States)

    El-Hillal, O.; Kurosaki, T.; Yamamura, H.; Kinet, J.-P.; Scharenberg, A. M.

    1997-01-01

    Activation of the syk tyrosine kinase occurs almost immediately following engagement of many types of antigen receptors, including Fc receptors, but the mechanism through which syk is activated is currently unclear. Here we demonstrate that Fc receptor-induced syk activation occurs as the result of phosphorylation of the syk activation loop by both src family kinases and other molecules of activated syk, suggesting that syk activation occurs as the result of a src kinase-initiated activation loop phosphorylation chain reaction. This type of activation mechanism predicts that syk activation would exhibit exponential kinetics, providing a potential explanation for its rapid and robust activation by even weak antigen receptor stimuli. We propose that a similar mechanism may be responsible for generating rapid activation of other cytoplasmic tyrosine kinases, such as those of the Bruton tyrosine kinase/tec family, as well. PMID:9050880

  16. Neuroleptic malignant-like syndrome with a slight elevation of creatine-kinase levels and respiratory failure in a patient with Parkinson's disease

    Directory of Open Access Journals (Sweden)

    Wei L

    2014-02-01

    Full Text Available Li Wei,1,2 Yinghui Chen1,2 1Department of Neurology, Jinshan Hospital, 2Department of Neurology, Shanghai Medical College, Fudan University, Shanghai, People's Republic of China Abstract: Neuroleptic malignant-like syndrome (NMLS is a rare but catastrophic complication of drug treatment for Parkinson's disease (PD. Sudden withdrawal and abrupt reduction of antiparkinsonian drugs are major risk factors. Just as its name suggests, the clinical features of NMLS are similar to neuroleptic malignant syndrome, which is a dangerous adverse response to antipsychotic drugs. Both of these conditions can present with hyperthermia, marked muscle rigidity, altered consciousness, autonomic dysfunction, and elevated serum creatine-kinase (CK levels. However, we describe a special NMLS case with a slight elevation of CK levels and respiratory failure in the full course of her treatment. The patient, a 68-year-old woman with a 4-years history of Parkinson's disease, presented with hyperthermia and severe muscular rigidity. During the course of her treatment, her maximum temperature was extremely high (above 41°C. At the beginning, the diagnosis of NMLS secondary to dopamine decrease was difficult to make, because her initial blood examination revealed that her serum CK levels were mildly elevated and decreased to normal range rapidly. Although antiparkinsonian drugs and supportive treatment were applied, the patient developed an acute respiratory failure in the early course of treatment. This case report highlights that when confronted with Parkinson's patients with high body temperature and muscle rigidity, NMLS should be taken into consideration even if there is no CK elevation. Likewise, the need for supportive care is essential, because its complications are severe, even such as respiratory failure. Keywords: antiparkinsonian drugs, creatine kinase, parkinsonism–hyperpyrexia syndrome, respiratory failure

  17. Comparison of new forms of creatine in raising plasma creatine levels

    Directory of Open Access Journals (Sweden)

    Purpura Martin

    2007-11-01

    Full Text Available Abstract Background Previous research has shown that plasma creatine levels are influenced by extracellular concentrations of insulin and glucose as well as by the intracellular creatine concentration. However, the form of creatine administered does not appear to have any effect although specific data on this is lacking. This study examined whether the administration of three different forms of creatine had different effects on plasma creatine concentrations and pharmacokinetics. Methods Six healthy subjects (three female and three male subjects participated in the study. Each subject was assigned to ingest a single dose of isomolar amounts of creatine (4.4 g in the form of creatine monohydrate (CrM, tri-creatine citrate (CrC, or creatine pyruvate (CrPyr using a balanced cross-over design. Plasma concentration curves, determined over eight hours after ingestion, were subject to pharmacokinetic analysis and primary derived data were analyzed by repeated measures ANOVA. Results Mean peak concentrations and area under the curve (AUC were significantly higher with CrPyr (17 and 14%, respectively in comparison to CrM and CrC. Mean peak concentration and AUC were not significantly different between CrM and CrC. Despite the higher peak concentration with CrPyr there was no difference between the estimated velocity constants of absorption (ka or elimination (kel between the three treatments. There was no effect of treatment with CrPyr on the plasma pyruvate concentration. Conclusion The findings suggest that different forms of creatine result in slightly altered kinetics of plasma creatine absorption following ingestion of isomolar (with respect to creatine doses of CrM, CrC and CrPyr although differences in ka could not be detected due to the small number of blood samples taken during the absorption phase. Characteristically this resulted in higher plasma concentrations of creatine with CrPyr. Differences in bioavailability are thought to be unlikely

  18. ALTERATIONS IN BRAIN CREATINE CONCENTRATIONS UNDER LONG-TERM SOCIAL ISOLATION (EXPERIMENTAL STUDY).

    Science.gov (United States)

    Koshoridze, N; Kuchukashvili, Z; Menabde, K; Lekiashvili, Sh; Koshoridze, M

    2016-02-01

    Stress represents one of the main problems of modern humanity. This study was done for understanding more clearly alterations in creatine content of the brain under psycho-emotional stress induced by long-term social isolation. It was shown that under 30 days social isolation creatine amount in the brain was arisen, while decreasing concentrations of synthesizing enzymes (AGAT, GAMT) and creatine transporter protein (CrT). Another important point was that such changes were accompanied by down-regulation of creatine kinase (CK), therefore the enzyme's concentration was lowered. In addition, it was observed that content of phosphocreatine (PCr) and ATP were also reduced, thus indicating down-regulation of energy metabolism of brain that is really a crucial point for its normal functioning. To sum up the results it can be underlined that long-term social isolation has negative influence on energy metabolism of brain; and as a result reduce ATP content, while increase of free creatine concentration, supposedly maintaining maximal balance for ATP amount, but here must be also noted that up-regulated oxidative pathways might have impact on blood brain barrier, resulting on its permeability.

  19. Moderate elevation of intracellular creatine by targeting the creatine transporter protects mice from acute myocardial infarction

    Science.gov (United States)

    Lygate, Craig A.; Bohl, Steffen; ten Hove, Michiel; Faller, Kiterie M.E.; Ostrowski, Philip J.; Zervou, Sevasti; Medway, Debra J.; Aksentijevic, Dunja; Sebag-Montefiore, Liam; Wallis, Julie; Clarke, Kieran; Watkins, Hugh; Schneider, Jürgen E.; Neubauer, Stefan

    2012-01-01

    Aims Increasing energy storage capacity by elevating creatine and phosphocreatine (PCr) levels to increase ATP availability is an attractive concept for protecting against ischaemia and heart failure. However, testing this hypothesis has not been possible since oral creatine supplementation is ineffectual at elevating myocardial creatine levels. We therefore used mice overexpressing creatine transporter in the heart (CrT-OE) to test for the first time whether elevated creatine is beneficial in clinically relevant disease models of heart failure and ischaemia/reperfusion (I/R) injury. Methods and results CrT-OE mice were selected for left ventricular (LV) creatine 20–100% above wild-type values and subjected to acute and chronic coronary artery ligation. Increasing myocardial creatine up to 100% was not detrimental even in ageing CrT-OE. In chronic heart failure, creatine elevation was neither beneficial nor detrimental, with no effect on survival, LV remodelling or dysfunction. However, CrT-OE hearts were protected against I/R injury in vivo in a dose-dependent manner (average 27% less myocardial necrosis) and exhibited greatly improved functional recovery following ex vivo I/R (59% of baseline vs. 29%). Mechanisms contributing to ischaemic protection in CrT-OE hearts include elevated PCr and glycogen levels and improved energy reserve. Furthermore, creatine loading in HL-1 cells did not alter antioxidant defences, but delayed mitochondrial permeability transition pore opening in response to oxidative stress, suggesting an additional mechanism to prevent reperfusion injury. Conclusion Elevation of myocardial creatine by 20–100% reduced myocardial stunning and I/R injury via pleiotropic mechanisms, suggesting CrT activation as a novel, potentially translatable target for cardiac protection from ischaemia. PMID:22915766

  20. Radioimmunoassay of creatine kinase-B isoenzyme in human sera: results in patients with acute myocardial infarction

    International Nuclear Information System (INIS)

    Willerson, J.T.; Stone, M.J.; Ting, R.; Mukherjee, A.; Gomez-Sanchez, C.E.; Lewis, P.; Hersh, L.B.

    1977-01-01

    A radiommunoassay was developed to measure serum levels of the B isoenzyme of creatine kinase (ATP: creatine N-phosphotransferase, EC 2.7.3.2) (CPK) in order to evaluate the time course and frequency of MB isoenzyme elevation in patients with acute myocardial infarction. The method can identify as little as 0.2 ng of the B portion of the CPK-MB isoenzyme, does not significantly crossreact with CPK-MM isoenzyme, and is not affected by storage of serum at -20 0 . CPK isoenzyme containing B subunits was detected in 48 out of 51 sera from normal adults; serum levels in these individuals ranged between 1.2 and 12.5 ng/ml[mean +- SEM was 2.7 +- 0.30 ng/ml]. The mean serum level of CPK-B isoenzyme in a pool of sera obtained from 100 normal subjects was 2.9 +- 0.35 ng/ml; two patients with rhabdomyolysis that were studied had serum CPK-B isoenzyme levels of 2.5 and 3.5 ng/ml, respectively. In contrast, serum levels of the CPK-B isoenzyme were markedly elevated in sera from 18 patients with acute myocardial infarcts when obtained within 12 hr after hospital admission; the mean +- SEM concentration was 56 +- 7.8 ng/ml. We performed serial determinations on 14 patients with acute myocardial infarcts and demonstrated that maximal serum CPK-B levels occurred within the first 12 hr after admission and were lower thereafter. The serum concentration of B-containing CPK isoenzyme in 19 additional patients admitted with chest pain but without acute myocardial infarction was 3.4 +- 0.50 ng/ml. Thus, radioimmunoassay measurement of CPK-B isoenzyme appears to be a useful and sensitive test for the detection of acute myocardial infarcts in patients

  1. Creatine salts provide neuroprotection even after partial impairment of the creatine transporter

    OpenAIRE

    Adriano, E.; Garbati, P.; Salis, A.; Damonte, G.; Millo, E.; Balestrino, M.

    2017-01-01

    Creatine, a compound that is critical for energy metabolism of nervous cells, crosses the blood-brain barrier (BBB) and the neuronal plasma membrane with difficulty, and only using its specific transporter. In the hereditary condition where the creatine transporter is defective (creatine transporter deficiency) there is no creatine in the brain, and administration of creatine is useless lacking the transporter. The disease is severe and incurable. Creatine-derived molecules that could cross B...

  2. Guanidinoacetic acid versus creatine for improved brain and muscle creatine levels

    DEFF Research Database (Denmark)

    Ostojic, Sergej M; Ostojic, Jelena; Drid, Patrik

    2016-01-01

    In this randomized, double-blind, crossover trial, we evaluated whether 4-week supplementation with guanidinoacetic acid (GAA) is superior to creatine in facilitating creatine levels in healthy men (n = 5). GAA (3.0 g/day) resulted in a more powerful rise (up to 16.2%) in tissue creatine levels...... in vastus medialis muscle, middle-cerebellar peduncle, and paracentral grey matter, as compared with creatine (P creatine for improved bioenergetics in energy-demanding tissues....

  3. Creatine Use in Sports.

    Science.gov (United States)

    Butts, Jessica; Jacobs, Bret; Silvis, Matthew

    The use of creatine as a dietary supplement has become increasingly popular over the past several decades. Despite the popularity of creatine, questions remain with regard to dosing, effects on sports performance, and safety. PubMed was searched for articles published between 1980 and January 2017 using the terms creatine, creatine supplementation, sports performance, and dietary supplements. An additional Google search was performed to capture National Collegiate Athletic Association-specific creatine usage data and US dietary supplement and creatine sales. Clinical review. Level 4. Short-term use of creatine is considered safe and without significant adverse effects, although caution should be advised as the number of long-term studies is limited. Suggested dosing is variable, with many different regimens showing benefits. The safety of creatine supplementation has not been studied in children and adolescents. Currently, the scientific literature best supports creatine supplementation for increased performance in short-duration, maximal-intensity resistance training. While creatine appears to be safe and effective for particular settings, whether creatine supplementation leads to improved performance on the field of play remains unknown.

  4. Exceptionally High Creatine Kinase (CK) Levels in Multicausal and Complicated Rhabdomyolysis: A Case Report.

    Science.gov (United States)

    Luckoor, Pavan; Salehi, Mashal; Kunadu, Afua

    2017-07-04

    BACKGROUND Rhabdomyolysis is a syndrome caused by muscle breakdown. It can be caused by traumatic as well as non-traumatic factors such as drugs, toxins, and infections. Although it has been initially associated with only traumatic causes, non-traumatic causes now appear to be at least 5 times more frequent. In rhabdomyolysis, the CK levels can range anywhere from 10 000 to 200 000 or even higher. The higher the CK levels, the greater will be the renal damage and associated complications. We present the case of a patient with exceptionally massive rhabdomyolysis with unusually high CK levels (nearly 1 million) caused by combined etiologic factors and complicated with acute renal failure. CASE REPORT A 36-year-old African American male patient with no significant past medical history and a social history of cocaine and alcohol abuse presented with diarrhea and generalized weakness of 2 days' duration. He was found to be febrile, tachycardic, tachypneic, and hypoxic. The patient was subsequently intubated and admitted to the medical ICU. Laboratory work-up showed acute renal failure with deranged liver functions test results, and elevated creatine kinase of 701,400 U/L. CK levels were subsequently too high for the lab to quantify. Urine legionella testing was positive for L. pneumophilia serogroup 1 antigen and urine toxicology was positive for cocaine. The patient had a protracted course in the ICU. He was initially started on CVVH, and later received intermittent hemodialysis for about 1 month. CONCLUSIONS In the presence of multiple etiologic factors, rhabdomyolysis can be massive with resultant significant morbidity. Clinicians should have a high index of suspicion for rhabdomyolysis in the presence of multiple factors, as early recognition of this diseases is very important in the prevention and active management of life-threatening conditions.

  5. Dietary creatine supplementation during pregnancy: a study on the effects of creatine supplementation on creatine homeostasis and renal excretory function in spiny mice.

    Science.gov (United States)

    Ellery, Stacey J; LaRosa, Domenic A; Kett, Michelle M; Della Gatta, Paul A; Snow, Rod J; Walker, David W; Dickinson, Hayley

    2016-08-01

    Recent evidence obtained from a rodent model of birth asphyxia shows that supplementation of the maternal diet with creatine during pregnancy protects the neonate from multi-organ damage. However, the effect of increasing creatine intake on creatine homeostasis and biosynthesis in females, particularly during pregnancy, is unknown. This study assessed the impact of creatine supplementation on creatine homeostasis, body composition, capacity for de novo creatine synthesis and renal excretory function in non-pregnant and pregnant spiny mice. Mid-gestation pregnant and virgin spiny mice were fed normal chow or chow supplemented with 5 % w/w creatine for 18 days. Weight gain, urinary creatine and electrolyte excretion were assessed during supplementation. At post mortem, body composition was assessed by Dual-energy X-ray absorptiometry, or tissues were collected to assess creatine content and mRNA expression of the creatine synthesising enzymes arginine:glycine amidinotransferase (AGAT) and guanidinoacetate methyltransferase (GAMT) and the creatine transporter (CrT1). Protein expression of AGAT and GAMT was also assessed by Western blot. Key findings of this study include no changes in body weight or composition with creatine supplementation; increased urinary creatine excretion in supplemented spiny mice, with increased sodium (P < 0.001) and chloride (P < 0.05) excretion in pregnant dams after 3 days of supplementation; lowered renal AGAT mRNA (P < 0.001) and protein (P < 0.001) expressions, and lowered CrT1 mRNA expression in the kidney (P < 0.01) and brain (P < 0.001). Creatine supplementation had minimal impact on creatine homeostasis in either non-pregnant or pregnant spiny mice. Increasing maternal dietary creatine consumption could be a useful treatment for birth asphyxia.

  6. Clinical utility of a two-site immunoradiometric assay for creatine kinase-MB in the detection of perioperative myocardial infarction

    International Nuclear Information System (INIS)

    DePuey, E.G.; Aessopos, A.; Monroe, L.R.; Hall, R.J.; Thompson, W.L.; Sonnemaker, R.E.; Burdine, J.A.

    1983-01-01

    In 144 patients, creatine kinase MB was measured serially at 0, 8, 16, 24, 48 and 72 h using a two-site immunoradionmetric assay (IRMA). Cardiac enzymes were also measured, including SGOT, LDH, total CPK, and CK-MB by electrophoresis. The presence of perioperative myocardial infarction (poMI) was established in 24 patients by the appearance of new electrocardiographic Q waves and/or new wall motion abnormalities detected by radionuclide ventriculography. In patients without poMI, CK-MB (IRMA) was elevated at 0 to 8 h but decreased by 16 h. In patients with poMI, peak values occurred at 16 to 24 h. Using a threshold value of 8.5 EU/I, patients with poMI could be distinguished from those without with 97% accuracy (sensitivity = 88%, specificity = 99%). We conclude that the CK-MB (IRMA) can serve as a valuable postoperative screening tet for poMI

  7. Dietary creatine supplementation lowers hepatic triacylglycerol by increasing lipoprotein secretion in rats fed high-fat diet.

    Science.gov (United States)

    da Silva, Robin P; Leonard, Kelly-Ann; Jacobs, René L

    2017-12-01

    Recent studies have shown that dietary creatine supplementation can prevent lipid accumulation in the liver. Creatine is a small molecule that plays a large role in energy metabolism, but since the enzyme creatine kinase is not present in the liver, the classical role in energy metabolism does not hold in this tissue. Fat accumulation in the liver can lead to the development of nonalcoholic fatty liver disease (NAFLD), a progressive disease that is prevalent in humans. We have previously reported that creatine can directly influence lipid metabolism in cell culture to promote lipid secretion and oxidation. Our goal in the current study was to determine whether similar mechanisms that occur in cell culture were present in vivo. We also sought to determine whether dietary creatine supplementation could be effective in reversing steatosis. Sprague-Dawley rats were fed a high-fat diet or a high-fat diet supplemented with creatine for 5 weeks. We found that rats supplemented with creatine had significantly improved rates of lipoprotein secretion and alterations in mitochondrial function that were consistent with greater oxidative capacity. We also find that introducing creatine into a high-fat diet halted hepatic lipid accumulation in rats with fatty liver. Our results support our previous report that liver cells in culture with creatine secrete and oxidize more oleic acid, demonstrating that dietary creatine can effectively change hepatic lipid metabolism by increasing lipoprotein secretion and oxidation in vivo. Our data suggest that creatine might be an effective therapy for NAFLD. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Thermal stability and aggregation of creatine kinase from rabbit skeletal muscle. Effect of 2-hydroxypropyl-beta-cyclodextrin.

    Science.gov (United States)

    Maloletkina, Olga I; Markossian, Kira A; Belousova, Lyubov V; Kleimenov, Sergey Yu; Orlov, Victor N; Makeeva, Valentina F; Kurganov, Boris I

    2010-05-01

    Effect of 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CD) on thermal aggregation of creatine kinase from rabbit skeletal muscle (RMCK) at 48 degrees C has been studied using dynamic light scattering. An increase in the duration of the lag period on the kinetic curves of aggregation, registered as an increment of the light scattering intensity in time, has been observed in the presence of HP-beta-CD. It has been shown that the initial parts of the dependences of the hydrodynamic radius (R(h)) of the protein aggregates on time follow the exponential law. The reciprocal value of parameter t(2R) (t(2R) is the time interval over which the R(h) value is doubled) was used to characterize the rate of aggregation. A 10-fold decrease in the 1/t(2R) value was observed in the presence of 76mM HP-beta-CD. Judging from the data on the kinetics of RMCK inactivation and the data on differential scanning calorimetry of RMCK, HP-beta-CD does not affect the rate of RMCK unfolding.

  9. Effect on hemo-dialysis on concentration of lactate dehyrogenase, creatine kinase and a-amylase in renal failure

    International Nuclear Information System (INIS)

    Modawe, G. O. H.; Idris, O. F.

    2009-01-01

    This study was conducted to compare the concentration of plasma enzymes in chronic renal failure pre dialysis (group A) and post dialysis (group B), and evaluate the concentration of these enzymes between pre and post dialysis. The study was performed in 25 samples of Sudanese patients (chronic renal failure) and compared with 15 samples as the control groups. plasma samples were analyzed using spectrophotometric methods, plasma concentration of these enzymes showed increase in chronic renal failure pre and post dialysis. The mean values of lactate dehydrogenase pre dialysis was 259IU/L, post dialysis was 276IU/L, the mean of creatine kinase pre dialysis was 252IU/L, and post dialysis was 241 IU/L but the mean of amylase pre and post dialysis was the same 144 IU/L. This study showed that there was no difference of concentration of amylase enzyme, but the different in concentration of CK and LDH between pre and post dialysis during chronic renal failure depend on normal range of this enzyme in control groups.(Author)

  10. Role of creatine kinase isoenzymes on muscular and cardiorespiratory endurance: genetic and molecular evidence.

    Science.gov (United States)

    Echegaray, M; Rivera, M A

    2001-01-01

    The ability to perform well in activities that require muscular and cardiorespiratory endurance is a trait influenced, in a considerable part, by the genetic make-up of individuals. Early studies of performance and recent scans of the human genome have pointed at various candidate genes responsible for the heterogeneity of these phenotypes within the population. Among these are the genes for the various creatine kinase (CK) isoenzyme subunits. CK and phosphocreatine (PCr) form an important metabolic system for temporal and spatial energy buffering in cells with large variations in energy demand. The different CK isoenzyme subunits (CK-M and CK-B) are differentially expressed in the tissues of the body. Although CK-M is the predominant form in both skeletal and cardiac muscle, CK-B is expressed to a greater extent in heart than in skeletal muscle. Studies in humans and mice have shown that the expression of CK-B messenger RNA (mRNA) and the abundance and activity of the CK-MB dimer increase in response to cardiorespiratory endurance training. Increases in muscle tissue CK-B content can be energetically favourable because of its lower Michaelis constant (Km) for ADP. The activity of the mitochondrial isoform of CK (Scmit-CK) has also been significantly and positively correlated to oxidative capacity and to CK-MB activity in muscle. In mice where the CK-M gene has been knocked out, significant increases in fatigue resistance together with cellular adaptations increasing aerobic capacity have been observed. These observations have led to the notion that this enzyme may be responsible for fatigue under normal circumstances, most likely because of the local cell compartment increase in inorganic phosphate concentration. Studies where the Scmit-CK gene was knocked out have helped demonstrate that this isoenzyme is very important for the stimulation of aerobic respiration. Human studies of CK-M gene sequence variation have shown a significant association between a

  11. Macro creatine kinases: results of isoenzyme electrophoresis and differentiation of the immunoglobulin-bound type by radioassay

    International Nuclear Information System (INIS)

    Bohner, J.; Stein, W.; Steinhart, R.; Wurzburg, U.; Eggstein, M.

    1982-01-01

    In 2.9% of sera from 1253 unselected patients we detected two different types of macromolecular creatine kinases (CK;EC 2.7.3.2). One macro type was represented by immunoglobulin-linked CK; in sera containing macro CK-BB isoenzyme, 125 I-labeled CK-BB was bound with high affinity to the immunoglobulin fraction. Furthermore, during electrophoresis, macro CK-BB mostly migrated between CK-MB and CK-MM, and was fixed to Protein A from Staphlococcus aureus. We therefore propose radioelectrophoresis as a specific, highly sensitive, and simple method for detecting this type of macro CK. This form occurs predominantly in elderly women, is not correlated to any specific disease, and persists in blood over a long period of time. In contrast, a second type (macro-CK type 2) never bound radiolabeled CK isoenzymes, and was not adsorbed to protein A. Electrophoretic migration of this macro-CK type 2 was generally cathodic to CK-MM. We observed this type in severely ill patients, frequently those suffering from malignant tumors. Clinical observations and biochemical data suggest that macro-CK type 2 is of mitochondrial origin

  12. Influence of Arm-cranking on Changes in Plasma CK Activity after High Force Eccentric Exercise of the Elbow Flexors

    OpenAIRE

    Kazunori, NOSAKA; Kei, SAKAMOTO; Mike, NEWTON; Exercise and Sports Science, Graduate School of Integrated Science, Yokohama City University:School of Biomedical and Sports Science, Edith Cowan University; Exercise and Sports Science, Graduate School of Integrated Science, Yokohama City University; School of Biomedical and Sports Science, Edith Cowan University

    2002-01-01

    It was hypothesized that the time course of changes in plasma creatine kinase (CK) activity following eccentric exercise was influenced by rhythmical muscle contractions performed after eccentric exercise. This study examined whether arm-cranking (AC) alters the time course of changes in plasma creatine kinase (CK) activity after eccentric exercise of the elbow flexors (ECC). Six male students performed two bouts of ECC separated by 3 weeks, and AC (25watts, 2-hours) was performed immediately...

  13. Control of creatine metabolism by HIF is an endogenous mechanism of barrier regulation in colitis.

    Science.gov (United States)

    Glover, Louise E; Bowers, Brittelle E; Saeedi, Bejan; Ehrentraut, Stefan F; Campbell, Eric L; Bayless, Amanda J; Dobrinskikh, Evgenia; Kendrick, Agnieszka A; Kelly, Caleb J; Burgess, Adrianne; Miller, Lauren; Kominsky, Douglas J; Jedlicka, Paul; Colgan, Sean P

    2013-12-03

    Mucosal surfaces of the lower gastrointestinal tract are subject to frequent, pronounced fluctuations in oxygen tension, particularly during inflammation. Adaptive responses to hypoxia are orchestrated largely by the hypoxia-inducible transcription factors (HIFs). As HIF-1α and HIF-2α are coexpressed in mucosal epithelia that constitute the barrier between the lumen and the underlying immune milieu, we sought to define the discrete contribution of HIF-1 and HIF-2 transactivation pathways to intestinal epithelial cell homeostasis. The present study identifies creatine kinases (CKs), key metabolic enzymes for rapid ATP generation via the phosphocreatine-creatine kinase (PCr/CK) system, as a unique gene family that is coordinately regulated by HIF. Cytosolic CKs are expressed in a HIF-2-dependent manner in vitro and localize to apical intestinal epithelial cell adherens junctions, where they are critical for junction assembly and epithelial integrity. Supplementation with dietary creatine markedly ameliorated both disease severity and inflammatory responses in colitis models. Further, enzymes of the PCr/CK metabolic shuttle demonstrate dysregulated mucosal expression in a subset of ulcerative colitis and Crohn disease patients. These findings establish a role for HIF-regulated CK in epithelial homeostasis and reveal a fundamental link between cellular bioenergetics and mucosal barrier.

  14. Creatine supplementation and swim performance: a brief review.

    Science.gov (United States)

    Hopwood, Melissa J; Graham, Kenneth; Rooney, Kieron B

    2006-03-01

    Nutritional supplements are popular among athletes participating in a wide variety of sports. Creatine is one of the most commonly used dietary supplements, as it has been shown to be beneficial in improving performance during repeated bouts of high-intensity anaerobic activity. This review examines the specific effects of creatine supplementation on swimming performance, and considers the effects of creatine supplementation on various measures of power development in this population. Research performed on the effect of creatine supplementation on swimming performance indicates that whilst creatine supplementation is ineffective in improving performance during a single sprint swim, dietary creatine supplementation may benefit repeated interval swim set performance. Considering the relationship between sprint swimming performance and measurements of power, the effect of creatine supplementation on power development in swimmers has also been examined. When measured on a swim bench ergometer, power development does show some improvement following a creatine supplementation regime. How this improvement in power output transfers to performance in the pool is uncertain. Although some evidence exists to suggest a gender effect on the performance improvements seen in swimmers following creatine supplementation, the majority of research indicates that male and female swimmers respond equally to supplementation. A major limitation to previous research is the lack of consideration given to the possible stroke dependant effect of creatine supplementation on swimming performance. The majority of the research conducted to date has involved examination of the freestyle swimming stroke only. The potential for performance improvements in the breaststroke and butterfly swimming strokes is discussed, with regards to the biomechanical differences and differences in efficiency between these strokes and freestyle. Key PointsCreatine supplementation does not improve single sprint

  15. Plasma creatine kinase B correlates with injury severity and symptoms in professional boxers.

    Science.gov (United States)

    Kilianski, Joseph; Peeters, Sophie; Debad, Jeff; Mohmed, Joseph; Wolf, Steven E; Minei, Joseph P; Diaz-Arrastia, Ramon; Gatson, Joshua W

    2017-11-01

    Each year in the United States, approximately 1.7 million people sustain a traumatic brain injury (TBI). Of these TBI events, about 75 percent are characterized as being mild brain injuries. Immediately following TBI, a secondary brain damage persists for hours, days, and even months. Previously, detection of neuronal and glial biomarkers have proven to be useful to predict neurological outcomes. Here, we hypothesized that creatine kinase, brain (CKBB) is a sensitive biomarker for acute secondary brain injury in professional boxers. Blood (8cc) was collected from the boxing athletes (n=18) prior to and after competition (∼30min). The plasma levels of CKBB were measured using the Meso Scale Diagnostic (MSD) electrochemiluminescence (ECL) array-based multiplex format. Additional data such as number of blows to the head and symptom score (Rivermead Post Concussion Symptoms Questionnaire) were collected. At approximately 30min after the competition, the plasma levels of CKBB were significantly elevated in concussed professional boxers and correlated with the number of blows to the head and symptom scores. Additionally, receiver operating curve (ROC) analysis yielded a 77.8% sensitivity and a specificity of 82.4% with an area under the curve (AUC) of 90% for CKBB as an identifier of secondary brain injury within this population. This study describes the detection of CKBB as a brain biomarker to detect secondary brain injury in professional athletes that have experienced multiple high impact blows to the head. This acute biomarker may prove useful in monitoring secondary brain injury after injury. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. The role of dietary creatine.

    Science.gov (United States)

    Brosnan, Margaret E; Brosnan, John T

    2016-08-01

    The daily requirement of a 70-kg male for creatine is about 2 g; up to half of this may be obtained from a typical omnivorous diet, with the remainder being synthesized in the body Creatine is a carninutrient, which means that it is only available to adults via animal foodstuffs, principally skeletal muscle, or via supplements. Infants receive creatine in mother's milk or in milk-based formulas. Vegans and infants fed on soy-based formulas receive no dietary creatine. Plasma and muscle creatine levels are usually somewhat lower in vegetarians than in omnivores. Human intake of creatine was probably much higher in Paleolithic times than today; some groups with extreme diets, such as Greenland and Alaskan Inuit, ingest much more than is currently typical. Creatine is synthesized from three amino acids: arginine, glycine and methionine (as S-adenosylmethionine). Humans can synthesize sufficient creatine for normal function unless they have an inborn error in a creatine-synthetic enzyme or a problem with the supply of substrate amino acids. Carnivorous animals, such as lions and wolves, ingest much larger amounts of creatine than humans would. The gastrointestinal tract and the liver are exposed to dietary creatine in higher concentrations before it is assimilated by other tissues. In this regard, our observations that creatine supplementation can prevent hepatic steatosis (Deminice et al. J Nutr 141:1799-1804, 2011) in a rodent model may be a function of the route of dietary assimilation. Creatine supplementation has also been reported to improve the intestinal barrier function of the rodent suffering from inflammatory bowel disease.

  17. Creatine, energetic function, metabolism and supplementation effects on sports

    Directory of Open Access Journals (Sweden)

    Emerson Gimenes Bernardo da Silva

    2008-06-01

    Full Text Available The purpose of this work is to review the literature regarding creatine ingestion by athletes and physical activity enthusiasts, discussing its necessity and, if possible, predicting some consequences. In order to achieve this purpose it was necessary to study the relationship between the muscles energetic system and their regulation. It was also proved necessary to investigate the creatine cycle, its endogenous origin, its metabolizing and conversion into creatine-phosphate. A bibliography was used to collect information about the subject. The research lead to the following conclusions: diet supplementation with creatine leads to increased phosphocreatine levels in human muscles. However, new in vivo experiments are most desirable, because it is already known that creatine interferes with the regulation of some metabolic pathways.

  18. Proton NMR studies of creatine in human erythrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kuchel, P W; Chapman, B E [Sydney Univ. (Australia). Dept. of Biochemistry

    1983-09-01

    Proton spin-echo nuclear magnetic resonance spectroscopy was used to measure the relative levels of some metabolites in intact human erythrocytes that had been fractionated by density gradient centrifugation. Age dependent changes in the concentrations of free glycine, choline and ergothioneine were seen for the first time, while glutathione was essentially invariant. In addition, there was a 10-fold decrease in creatine levels from the youngest to oldest cells. This confirms earlier reports and provides a simple explanation for the variable creatine resonance intensities seen in spectra obtained from different erythrocyte samples prepared from the same donor. The different chemical shifts of the methylene resonances of creatine and creatine phosphate was demonstrated and hence confirmed that the bulk of the creatine in intact erythrocytes is not phosphorylated. The chemical shift difference enabled the monitoring of the creatine phosphokinase catalysed reaction in lysates to which the rabbit muscle enzyme had been added. This experiment indicated that the enzyme is not significantly inhibited by factors in the lysates, and introduced a new means of assaying the in situ activity of the enzyme.

  19. Proton NMR studies of creatine in human erythrocytes

    International Nuclear Information System (INIS)

    Kuchel, P.W.; Chapman, B.E.

    1983-01-01

    Proton spin-echo nuclear magnetic resonance spectroscopy was used to measure the relative levels of some metabolites in intact human erythrocytes that had been fractionated by density gradient centrifugation. Age dependent changes in the concentrations of free glycine, choline and ergothioneine were seen for the first time, while glutathione was essentially invariant. In addition, there was a 10-fold decrease in creatine levels from the youngest to oldest cells. This confirms earlier reports and provides a simple explanation for the variable creatine resonance intensities seen in spectra obtained from different erythrocyte samples prepared from the same donor. The different chemical shifts of the methylene resonances of creatine and creatine phosphate was demonstrated and hence confirmed that the bulk of the creatine in intact erythrocytes is not phosphorylated. The chemical shift difference enabled the monitoring of the creatine phosphokinase catalysed reaction in lysates to which the rabbit muscle enzyme had been added. This experiment indicated that the enzyme is not significantly inhibited by factors in the lysates, and introduced a new means of assaying the in situ activity of the enzyme. (author)

  20. Effects of Creatine Supplementation on Muscle Strength and Optimal Individual Post-Activation Potentiation Time of the Upper Body in Canoeists.

    Science.gov (United States)

    Wang, Chia-Chi; Lin, Shu-Cheng; Hsu, Shu-Ching; Yang, Ming-Ta; Chan, Kuei-Hui

    2017-10-27

    Creatine supplementation reduces the impact of muscle fatigue on post-activation potentiation (PAP) of the lower body, but its effects on the upper body remain unknown. This study examined the effects of creatine supplementation on muscle strength, explosive power, and optimal individual PAP time of the upper body during a set of complex training bouts in canoeists. Seventeen male high school canoeists performed a bench row for one repetition at maximum strength and conducted complex training bouts to determine the optimal individual timing of PAP and distance of overhead medicine ball throw before and after the supplementation. Subjects were assigned to a creatine or placebo group, and later consumed 20 g of creatine or carboxymethyl cellulose per day for six days. After supplementation, the maximal strength in the creatine group significantly increased ( p creatine group was significantly earlier than the pre-supplementation times ( p creatine supplementation increases maximal strength and shortens the optimal individual PAP time of the upper body in high school athletes, but has no effect on explosive power. Moreover, it was found that the recovery time between a bench row and an overhead medicine ball throw in a complex training bout is an individual phenomenon.

  1. Multisystem Disease, Including Eosinophilia and Progressive Hyper-Creatine-Kinase-emia over 10 Years, Suggests Mitochondrial Disorder

    Directory of Open Access Journals (Sweden)

    Josef Finsterer

    2017-04-01

    Full Text Available Background: Eosinophilia has not been reported as a manifestation of a mitochondrial disorder (MID. Here, we report a patient with clinical features suggesting a MID and permanent eosinophilia, multisystem disease, and progressive hyper-creatine-kinase (CK-emia for at least 10 years. Materials and Methods: Methods applied included a clinical exam, blood chemical investigations, electrophysiological investigations, imaging, and invasive cardiological investigations. The patient was repeatedly followed up over several years. He required replacement cardiac surgery. Results: In a 57-year-old male, eosinophilia was first detected at the age of 44 years and has remained almost constantly present until today. In addition to eosinophilia, he developed progressive hyper-CK-emia at the age of 47 years. His history was further positive for hepatopathy, hyperlipidemia, hypothyroidism, renal insufficiency, spontaneous Achilles tendon rupture, double vision, exercise intolerance, muscle aching, mild hypoacusis, sensory neuropathy, seizures, and mitral insufficiency/stenosis requiring valve replacement therapy, oral anticoagulation, and pacemaker implantation. Based on the multisystem nature of his abnormalities and permanent hyper-CK-emia, a MID was suspected. Conclusion: Eosinophilia can be associated with a MID with myopathy, possibly as a reaction to myofiber necrosis. If eosinophilia is associated with progressive hyper-CK-emia and multisystem disease, a MID should be suspected.

  2. CREATINE SUPPLEMENTATION AND SWIM PERFORMANCE: A BRIEF REVIEW

    Directory of Open Access Journals (Sweden)

    Melissa J. Hopwood

    2006-03-01

    Full Text Available Nutritional supplements are popular among athletes participating in a wide variety of sports. Creatine is one of the most commonly used dietary supplements, as it has been shown to be beneficial in improving performance during repeated bouts of high-intensity anaerobic activity. This review examines the specific effects of creatine supplementation on swimming performance, and considers the effects of creatine supplementation on various measures of power development in this population. Research performed on the effect of creatine supplementation on swimming performance indicates that whilst creatine supplementation is ineffective in improving performance during a single sprint swim, dietary creatine supplementation may benefit repeated interval swim set performance. Considering the relationship between sprint swimming performance and measurements of power, the effect of creatine supplementation on power development in swimmers has also been examined. When measured on a swim bench ergometer, power development does show some improvement following a creatine supplementation regime. How this improvement in power output transfers to performance in the pool is uncertain. Although some evidence exists to suggest a gender effect on the performance improvements seen in swimmers following creatine supplementation, the majority of research indicates that male and female swimmers respond equally to supplementation. A major limitation to previous research is the lack of consideration given to the possible stroke dependant effect of creatine supplementation on swimming performance. The majority of the research conducted to date has involved examination of the freestyle swimming stroke only. The potential for performance improvements in the breaststroke and butterfly swimming strokes is discussed, with regards to the biomechanical differences and differences in efficiency between these strokes and freestyle

  3. Does brain creatine content rely on exogenous creatine in healthy youth? A proof-of-principle study.

    Science.gov (United States)

    Merege-Filho, Carlos Alberto Abujabra; Otaduy, Maria Concepción Garcia; de Sá-Pinto, Ana Lúcia; de Oliveira, Maira Okada; de Souza Gonçalves, Lívia; Hayashi, Ana Paula Tanaka; Roschel, Hamilton; Pereira, Rosa Maria Rodrigues; Silva, Clovis Artur; Brucki, Sonia Maria Dozzi; da Costa Leite, Claudia; Gualano, Bruno

    2017-02-01

    It has been hypothesized that dietary creatine could influence cognitive performance by increasing brain creatine in developing individuals. This double-blind, randomized, placebo-controlled, proof-of-principle study aimed to investigate the effects of creatine supplementation on cognitive function and brain creatine content in healthy youth. The sample comprised 67 healthy participants aged 10 to 12 years. The participants were given creatine or placebo supplementation for 7 days. At baseline and after the intervention, participants undertook a battery of cognitive tests. In a random subsample of participants, brain creatine content was also assessed in the regions of left dorsolateral prefrontal cortex, left hippocampus, and occipital lobe by proton magnetic resonance spectroscopy (1H-MRS) technique. The scores obtained from verbal learning and executive functions tests did not significantly differ between groups at baseline or after the intervention (all p > 0.05). Creatine content was not significantly different between groups in left dorsolateral prefrontal cortex, left hippocampus, and occipital lobe (all p > 0.05). In conclusion, a 7-day creatine supplementation protocol did not elicit improvements in brain creatine content or cognitive performance in healthy youth, suggesting that this population mainly relies on brain creatine synthesis rather than exogenous creatine intake to maintain brain creatine homeostasis.

  4. Creatine phosphokinase test

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003503.htm Creatine phosphokinase test To use the sharing features on this page, please enable JavaScript. Creatine phosphokinase (CPK) is an enzyme in the body. ...

  5. Reference values for the creatine kinase response to professional Australian football match-play.

    Science.gov (United States)

    Inman, Luke A G; Rennie, Michael J; Watsford, Mark L; Gibbs, Nathan J; Green, James; Spurrs, Robert W

    2018-08-01

    Due to the importance of monitoring markers of muscle damage in high-level sport from a medical and athlete recovery perspective, this study aimed to determine the upper limits of normal (ULN) for post-match plasma creatine kinase (CK) in professional Australian footballers. Raw CK values were considered, along with intra-individual deviations from the season-mean. Case series. CK was collected between 36-48h following professional Australian football match-play. A total of 1565 samples from 62 players were assessed over three consecutive seasons. The ULN were determined for raw scores and as a percentage of each player's season-mean response. The ULN for raw CK, as determined by the 97.5th, 95th and 90th percentiles were 1715 (90%CI: 1605-1890), 1380 (90%CI: 1325-1475) and 1110 (90%CI: 1050-1170) UL -1 respectively. The ULN intra-individual response (97.5th percentile) was defined as a player's score being greater than 94% (90%CI: 84-102%) above their season-mean. Professional Australian football elicits a profound effect on the CK response. The values provide a reference tool for athletes competing at this level of competition. The novel method of representing the CK response as a percentage difference from an individuals' season-mean enables a superior comparative ability between CK responses and reduces the high CK responder bias that occurs when using raw scores alone. The data will assist medical and conditioning staff in excluding medical emergencies and also aid in individualising the prescription of training loads and recovery to optimise athlete performance and minimise further muscle damage. Copyright © 2018 Sports Medicine Australia. Published by Elsevier Ltd. All rights reserved.

  6. Magnesium-adenosine diphosphate binding sites in wild-type creatine kinase and in mutants: role of aromatic residues probed by Raman and infrared spectroscopies.

    Science.gov (United States)

    Hagemann, H; Marcillat, O; Buchet, R; Vial, C

    2000-08-08

    Two distinct methods were used to investigate the role of Trp residues during Mg-ADP binding to cytosolic creatine kinase (CK) from rabbit muscle: (1) Raman spectroscopy, which is very sensitive to the environment of aromatic side-chain residues, and (2) reaction-induced infrared difference spectroscopy (RIDS) and photolabile substrate (ADP[Et(PhNO(2))]), combined with site-directed mutagenesis on the four Trp residues of CK. Our Raman results indicated that the environment of Trp and of Tyr were not affected during Mg-ADP binding to CK. Analysis of RIDS of wild-type CK, inactive W227Y, and active W210,217,272Y mutants suggested that Trp227 was not involved in the stacking interactions. Results are consistent with Trp227 being essential to prevent water molecules from entering in the active site [as suggested by Gross, M., Furter-Graves, E. M., Wallimann, T., Eppenberger, H. M., and Furter, R. (1994) Protein Sci. 3, 1058-1068] and that another Trp could in addition help to steer the nucleotide in the binding site, although it is not essential for the activity of CK. Raman and infrared spectra indicated that Mg-ADP binding does not involve large secondary structure changes. Only 3-4 residues absorbing in the amide I region are directly implicated in the Mg-ADP binding (corresponding to secondary structure changes less than 1%), suggesting that movement of protein domains due to Mg-nucleotide binding do not promote large secondary structure changes.

  7. Protein kinase activity of phosphoinositide 3-kinase regulates cytokine-dependent cell survival.

    Directory of Open Access Journals (Sweden)

    Daniel Thomas

    Full Text Available The dual specificity protein/lipid kinase, phosphoinositide 3-kinase (PI3K, promotes growth factor-mediated cell survival and is frequently deregulated in cancer. However, in contrast to canonical lipid-kinase functions, the role of PI3K protein kinase activity in regulating cell survival is unknown. We have employed a novel approach to purify and pharmacologically profile protein kinases from primary human acute myeloid leukemia (AML cells that phosphorylate serine residues in the cytoplasmic portion of cytokine receptors to promote hemopoietic cell survival. We have isolated a kinase activity that is able to directly phosphorylate Ser585 in the cytoplasmic domain of the interleukin 3 (IL-3 and granulocyte macrophage colony stimulating factor (GM-CSF receptors and shown it to be PI3K. Physiological concentrations of cytokine in the picomolar range were sufficient for activating the protein kinase activity of PI3K leading to Ser585 phosphorylation and hemopoietic cell survival but did not activate PI3K lipid kinase signaling or promote proliferation. Blockade of PI3K lipid signaling by expression of the pleckstrin homology of Akt1 had no significant impact on the ability of picomolar concentrations of cytokine to promote hemopoietic cell survival. Furthermore, inducible expression of a mutant form of PI3K that is defective in lipid kinase activity but retains protein kinase activity was able to promote Ser585 phosphorylation and hemopoietic cell survival in the absence of cytokine. Blockade of p110α by RNA interference or multiple independent PI3K inhibitors not only blocked Ser585 phosphorylation in cytokine-dependent cells and primary human AML blasts, but also resulted in a block in survival signaling and cell death. Our findings demonstrate a new role for the protein kinase activity of PI3K in phosphorylating the cytoplasmic tail of the GM-CSF and IL-3 receptors to selectively regulate cell survival highlighting the importance of targeting

  8. Upregulation of the Creatine Transporter Slc6A8 by Klotho

    Directory of Open Access Journals (Sweden)

    Ahmad Almilaji

    2014-11-01

    Full Text Available Background/Aims: The transmembrane Klotho protein contributes to inhibition of 1,25(OH2D3 formation. The extracellular domain of Klotho protein could function as an enzyme with e.g. β-glucuronidase activity, be cleaved off and be released into blood and cerebrospinal fluid. Klotho regulates several cellular transporters. Klotho protein deficiency accelerates the appearance of age related disorders including neurodegeneration and muscle wasting and eventually leads to premature death. The main site of Klotho protein expression is the kidney. Klotho protein is also appreciably expressed in other tissues including chorioid plexus. The present study explored the effect of Klotho protein on the creatine transporter CreaT (Slc6A8, which participates in the maintenance of neuronal function and survival. Methods: To this end cRNA encoding Slc6A8 was injected into Xenopus oocytes with and without additional injection of cRNA encoding Klotho protein. Creatine transporter CreaT (Slc6A8 activity was estimated from creatine induced current determined by two-electrode voltage-clamp. Results: Coexpression of Klotho protein significantly increased creatine-induced current in Slc6A8 expressing Xenopus oocytes. Coexpression of Klotho protein delayed the decline of creatine induced current following inhibition of carrier insertion into the cell membrane by brefeldin A (5 µM. The increase of creatine induced current by coexpression of Klotho protein in Slc6A8 expressing Xenopus oocytes was reversed by β-glucuronidase inhibitor (DSAL. Similarly, treatment of Slc6A8 expressing Xenopus oocytes with recombinant human alpha Klotho protein significantly increased creatine induced current. Conclusion: Klotho protein up-regulates the activity of creatine transporter CreaT (Slc6A8 by stabilizing the carrier protein in the cell membrane, an effect requiring β-glucuronidase activity of Klotho protein.

  9. Chitin and stress induced protein kinase activation

    DEFF Research Database (Denmark)

    Kenchappa, Chandra Shekar; Azevedo da Silva, Raquel; Bressendorff, Simon

    2017-01-01

    The assays described here are pertinent to protein kinase studies in any plant. They include an immunoblot phosphorylation/activation assay and an in-gel activity assay for MAP kinases (MPKs) using the general protein kinase substrate myelin basic protein. They also include a novel in-gel peptide...... substrate assay for Snf1-related kinase family 2 members (SnRK2s). This kinase family-specific assay overcomes some limitations of in-gel assays and permits the identification of different types of kinase activities in total protein extracts....

  10. Engineering of kinase-based protein interacting devices: active expression of tyrosine kinase domains

    KAUST Repository

    Diaz Galicia, Miriam Escarlet

    2018-05-01

    Protein-protein interactions modulate cellular processes in health and disease. However, tracing weak or rare associations or dissociations of proteins is not a trivial task. Kinases are often regulated through interaction partners and, at the same time, themselves regulate cellular interaction networks. The use of kinase domains for creating a synthetic sensor device that reads low concentration protein-protein interactions and amplifies them to a higher concentration interaction which is then translated into a FRET (Fluorescence Resonance Energy Transfer) signal is here proposed. To this end, DNA constructs for interaction amplification (split kinases), positive controls (intact kinase domains), scaffolding proteins and phosphopeptide - SH2-domain modules for the reading of kinase activity were assembled and expression protocols for fusion proteins containing Lyn, Src, and Fak kinase domains in bacterial and in cell-free systems were optimized. Also, two non-overlapping methods for measuring the kinase activity of these proteins were stablished and, finally, a protein-fragment complementation assay with the split-kinase constructs was tested. In conclusion, it has been demonstrated that features such as codon optimization, vector design and expression conditions have an impact on the expression yield and activity of kinase-based proteins. Furthermore, it has been found that the defined PURE cell-free system is insufficient for the active expression of catalytic kinase domains. In contrast, the bacterial co-expression with phosphatases produced active kinase fusion proteins for two out of the three tested Tyrosine kinase domains.

  11. Comparative Molecular Dynamics Simulations of Mitogen-Activated Protein Kinase-Activated Protein Kinase 5

    Directory of Open Access Journals (Sweden)

    Inger Lindin

    2014-03-01

    Full Text Available The mitogen-activated protein kinase-activated protein kinase MK5 is a substrate of the mitogen-activated protein kinases p38, ERK3 and ERK4. Cell culture and animal studies have demonstrated that MK5 is involved in tumour suppression and promotion, embryogenesis, anxiety, cell motility and cell cycle regulation. In the present study, homology models of MK5 were used for molecular dynamics (MD simulations of: (1 MK5 alone; (2 MK5 in complex with an inhibitor; and (3 MK5 in complex with the interaction partner p38α. The calculations showed that the inhibitor occupied the active site and disrupted the intramolecular network of amino acids. However, intramolecular interactions consistent with an inactive protein kinase fold were not formed. MD with p38α showed that not only the p38 docking region, but also amino acids in the activation segment, αH helix, P-loop, regulatory phosphorylation region and the C-terminal of MK5 may be involved in forming a very stable MK5-p38α complex, and that p38α binding decreases the residual fluctuation of the MK5 model. Electrostatic Potential Surface (EPS calculations of MK5 and p38α showed that electrostatic interactions are important for recognition and binding.

  12. Creatine Supplementation Does Not Prevent the Development of Alcoholic Steatosis.

    Science.gov (United States)

    Ganesan, Murali; Feng, Dan; Barton, Ryan W; Thomes, Paul G; McVicker, Benita L; Tuma, Dean J; Osna, Natalia A; Kharbanda, Kusum K

    2016-11-01

    Alcohol-induced reduction in the hepatocellular S-adenosylmethionine (SAM):S-adenosylhomocysteine (SAH) ratio impairs the activities of many SAM-dependent methyltransferases. These impairments ultimately lead to the generation of several hallmark features of alcoholic liver injury including steatosis. Guanidinoacetate methyltransferase (GAMT) is an important enzyme that catalyzes the final reaction in the creatine biosynthetic process. The liver is a major site for creatine synthesis which places a substantial methylation burden on this organ as GAMT-mediated reactions consume as much as 40% of all the SAM-derived methyl groups. We hypothesized that dietary creatine supplementation could potentially spare SAM, preserve the hepatocellular SAM:SAH ratio, and thereby prevent the development of alcoholic steatosis and other consequences of impaired methylation reactions. For these studies, male Wistar rats were pair-fed the Lieber-DeCarli control or ethanol (EtOH) diet with or without 1% creatine supplementation. At the end of 4 to 5 weeks of feeding, relevant biochemical and histological analyses were performed. We observed that creatine supplementation neither prevented alcoholic steatosis nor attenuated the alcohol-induced impairments in proteasome activity. The lower hepatocellular SAM:SAH ratio seen in the EtOH-fed rats was also not normalized or SAM levels spared when these rats were fed the creatine-supplemented EtOH diet. However, a >10-fold increased level of creatine was observed in the liver, serum, and hearts of rats fed the creatine-supplemented diets. Overall, dietary creatine supplementation did not prevent alcoholic liver injury despite its known efficacy in preventing high-fat-diet-induced steatosis. Betaine, a promethylating agent that maintains the hepatocellular SAM:SAH, still remains our best option for treating alcoholic steatosis. Copyright © 2016 by the Research Society on Alcoholism.

  13. Effects of Creatine Supplementation on Muscle Strength and Optimal Individual Post-Activation Potentiation Time of the Upper Body in Canoeists

    Directory of Open Access Journals (Sweden)

    Chia-Chi Wang

    2017-10-01

    Full Text Available Creatine supplementation reduces the impact of muscle fatigue on post-activation potentiation (PAP of the lower body, but its effects on the upper body remain unknown. This study examined the effects of creatine supplementation on muscle strength, explosive power, and optimal individual PAP time of the upper body during a set of complex training bouts in canoeists. Seventeen male high school canoeists performed a bench row for one repetition at maximum strength and conducted complex training bouts to determine the optimal individual timing of PAP and distance of overhead medicine ball throw before and after the supplementation. Subjects were assigned to a creatine or placebo group, and later consumed 20 g of creatine or carboxymethyl cellulose per day for six days. After supplementation, the maximal strength in the creatine group significantly increased (p < 0.05. The optimal individual PAP time in the creatine group was significantly earlier than the pre-supplementation times (p < 0.05. There was no significant change in explosive power for either group. Our findings support the notion that creatine supplementation increases maximal strength and shortens the optimal individual PAP time of the upper body in high school athletes, but has no effect on explosive power. Moreover, it was found that the recovery time between a bench row and an overhead medicine ball throw in a complex training bout is an individual phenomenon.

  14. Glycogen synthase kinase 3β promotes liver innate immune activation by restraining AMP-activated protein kinase activation.

    Science.gov (United States)

    Zhou, Haoming; Wang, Han; Ni, Ming; Yue, Shi; Xia, Yongxiang; Busuttil, Ronald W; Kupiec-Weglinski, Jerzy W; Lu, Ling; Wang, Xuehao; Zhai, Yuan

    2018-02-13

    Glycogen synthase kinase 3β (Gsk3β [Gsk3b]) is a ubiquitously expressed kinase with distinctive functions in different types of cells. Although its roles in regulating innate immune activation and ischaemia and reperfusion injuries (IRIs) have been well documented, the underlying mechanisms remain ambiguous, in part because of the lack of cell-specific tools in vivo. We created a myeloid-specific Gsk3b knockout (KO) strain to study the function of Gsk3β in macrophages in a murine liver partial warm ischaemia model. Compared with controls, myeloid Gsk3b KO mice were protected from IRI, with diminished proinflammatory but enhanced anti-inflammatory immune responses in livers. In bone marrow-derived macrophages, Gsk3β deficiency resulted in an early reduction of Tnf gene transcription but sustained increase of Il10 gene transcription on Toll-like receptor 4 stimulation in vitro. These effects were associated with enhanced AMP-activated protein kinase (AMPK) activation, which led to an accelerated and higher level of induction of the novel innate immune negative regulator small heterodimer partner (SHP [Nr0b2]). The regulatory function of Gsk3β on AMPK activation and SHP induction was confirmed in wild-type bone marrow-derived macrophages with a Gsk3 inhibitor. Furthermore, we found that this immune regulatory mechanism was independent of Gsk3β Ser9 phosphorylation and the phosphoinositide 3-kinase-Akt signalling pathway. In vivo, myeloid Gsk3β deficiency facilitated SHP upregulation by ischaemia-reperfusion in liver macrophages. Treatment of Gsk3b KO mice with either AMPK inhibitor or SHP small interfering RNA before the onset of liver ischaemia restored liver proinflammatory immune activation and IRI in these otherwise protected hosts. Additionally, pharmacological activation of AMPK protected wild-type mice from liver IRI, with reduced proinflammatory immune activation. Inhibition of the AMPK-SHP pathway by liver ischaemia was demonstrated in tumour resection

  15. Serum concentrations of creatine kinase and of triglycerides during lactation in gilts bred older and in multiparous sows fed ad libitum

    Directory of Open Access Journals (Sweden)

    Nogueira R.H.G.

    2000-01-01

    Full Text Available The aim of this paper was to assess the possible variation in blood concentrations of creatine kinase (CK and triglycerides in gilts bred older in comparison with multiparous sows. Ten primiparous and ten Camborough multiparous sows from the fourth to seventh parities were used. Breeding age and weight of gilts averaged respectively 231 days and 149.5kg. All females were moved into individual farrowing crates and were managed under the same conditions. Blood samples were collected by puncturing the coccygeal artery on day 7 before expected farrowing, and on days 2, 7, 14, 21 of lactation and 2 days after weaning. No difference in triglycerides and CK serum concentrations between groups were observed. The CK levels were low before farrowing, increased substantially on days 2 and 7 and decreased toward the end of lactation. The concentrations of triglycerides were slightly high before the parturition, diminished on days 2, 7, 14 and 21 and increased after weaning.

  16. Whole body creatine and protein kinetics in healthy men and women: effects of creatine and amino acid supplementation.

    Science.gov (United States)

    Kalhan, Satish C; Gruca, Lourdes; Marczewski, Susan; Bennett, Carole; Kummitha, China

    2016-03-01

    Creatine kinetics were measured in young healthy subjects, eight males and seven females, age 20-30 years, after an overnight fast on creatine-free diet. Whole body turnover of glycine and its appearance in creatine was quantified using [1-(13)C] glycine and the rate of protein turnover was quantified using L-ring [(2)H5] phenylalanine. The creatine pool size was estimated by the dilution of a bolus [C(2)H3] creatine. Studies were repeated following a five days supplement creatine 21 g.day(-1) and following supplement amino acids 14.3 g day(-1). Creatine caused a ten-fold increase in the plasma concentration of creatine and a 50 % decrease in the concentration of guanidinoacetic acid. Plasma amino acids profile showed a significant decrease in glycine, glutamine, and taurine and a significant increase in citrulline, valine, lysine, and cysteine. There was a significant decrease in the rate of appearance of glycine, suggesting a decrease in de-novo synthesis (p = 0.006). The fractional and absolute rate of synthesis of creatine was significantly decreased by supplemental creatine. Amino acid supplement had no impact on any of the parameters. This is the first detailed analysis of creatine kinetics and the effects of creatine supplement in healthy young men and women. These methods can be applied for the analysis of creatine kinetics in different physiological states.

  17. Creatine Kinase and Lactate Dehydrogenase Responses After Different Resistance and Aerobic Exercise Protocols

    Directory of Open Access Journals (Sweden)

    Callegari Gustavo A.

    2017-08-01

    Full Text Available The aim of this study was to investigate the responses of creatine kinase (CK and lactate dehydrogenase (LDH after performing different resistance and aerobic exercise protocols. Twelve recreationally trained men (age, 23.2 ± 5.6 years; body mass, 84.3 ± 9.3 kg; body height, 178.9 ± 4.5 cm; and BMI, 26.3 ± 2.3 kg·m2 volunteered to participate in this study. All subjects were randomly assigned to four experimental protocols (crossover: (a aerobic training at 60% of VO2max, (b aerobic training at 80% of VO2max, (c a resistance exercise (RE session with a bi-set protocol, and (d an RE session with a multiple sets protocol. Blood samples were collected before, immediately after and 24 hours following the experimental protocols. After 24 hours, there was a significant increase in CK for the 80% of VO2max protocol vs. the bi-set RE session (p = 0.016. Immediately after the protocols, we observed a significant increase in LDH among certain groups compared to others, as follows: multiple sets RE session vs. 60% of VO2max, bi-set RE session vs. 60% of VO2max, multiple sets RE session vs. 80% of VO2max, and bi-set RE session vs. 80% of VO2max (p = 0.008, p = 0.013; p = 0.002, p = 0.004, respectively. In conclusion, aerobic exercise performed at 80% of VO2max appears to elevate plasma CK levels more than bi-set RE sessions. However, the bi-set and multiple sets RE sessions appeared to trigger greater levels of blood LDH compared to aerobic protocols performed at 60% and 80% of VO2max.

  18. The SH2 domain of Abl kinases regulates kinase autophosphorylation by controlling activation loop accessibility

    Science.gov (United States)

    Lamontanara, Allan Joaquim; Georgeon, Sandrine; Tria, Giancarlo; Svergun, Dmitri I.; Hantschel, Oliver

    2014-11-01

    The activity of protein kinases is regulated by multiple molecular mechanisms, and their disruption is a common driver of oncogenesis. A central and almost universal control element of protein kinase activity is the activation loop that utilizes both conformation and phosphorylation status to determine substrate access. In this study, we use recombinant Abl tyrosine kinases and conformation-specific kinase inhibitors to quantitatively analyse structural changes that occur after Abl activation. Allosteric SH2-kinase domain interactions were previously shown to be essential for the leukemogenesis caused by the Bcr-Abl oncoprotein. We find that these allosteric interactions switch the Abl activation loop from a closed to a fully open conformation. This enables the trans-autophosphorylation of the activation loop and requires prior phosphorylation of the SH2-kinase linker. Disruption of the SH2-kinase interaction abolishes activation loop phosphorylation. Our analysis provides a molecular mechanism for the SH2 domain-dependent activation of Abl that may also regulate other tyrosine kinases.

  19. Structural coupling of SH2-kinase domains links Fes and Abl substrate recognition and kinase activation.

    Science.gov (United States)

    Filippakopoulos, Panagis; Kofler, Michael; Hantschel, Oliver; Gish, Gerald D; Grebien, Florian; Salah, Eidarus; Neudecker, Philipp; Kay, Lewis E; Turk, Benjamin E; Superti-Furga, Giulio; Pawson, Tony; Knapp, Stefan

    2008-09-05

    The SH2 domain of cytoplasmic tyrosine kinases can enhance catalytic activity and substrate recognition, but the molecular mechanisms by which this is achieved are poorly understood. We have solved the structure of the prototypic SH2-kinase unit of the human Fes tyrosine kinase, which appears specialized for positive signaling. In its active conformation, the SH2 domain tightly interacts with the kinase N-terminal lobe and positions the kinase alphaC helix in an active configuration through essential packing and electrostatic interactions. This interaction is stabilized by ligand binding to the SH2 domain. Our data indicate that Fes kinase activation is closely coupled to substrate recognition through cooperative SH2-kinase-substrate interactions. Similarly, we find that the SH2 domain of the active Abl kinase stimulates catalytic activity and substrate phosphorylation through a distinct SH2-kinase interface. Thus, the SH2 and catalytic domains of active Fes and Abl pro-oncogenic kinases form integrated structures essential for effective tyrosine kinase signaling.

  20. The Regulation and Expression of the Creatine Transporter: A Brief Review of Creatine Supplementation in Humans and Animals

    OpenAIRE

    Schoch, Ryan D; Willoughby, Darryn; Greenwood, Mike

    2006-01-01

    Abstract Creatine monohydrate has become one of the most popular ergogenic sport supplements used today. It is a nonessential dietary compound that is both endogenously synthesized and naturally ingested through diet. Creatine ingested through supplementation has been observed to be absorbed into the muscle exclusively by means of a creatine transporter, CreaT1. The major rationale of creatine supplementation is to maximize the increase within the intracellular pool of total creatine (creatin...

  1. Cocoa Procyanidins Suppress Transformation by Inhibiting Mitogen-activated Protein Kinase Kinase*S⃞

    Science.gov (United States)

    Kang, Nam Joo; Lee, Ki Won; Lee, Dong Eun; Rogozin, Evgeny A.; Bode, Ann M.; Lee, Hyong Joo; Dong, Zigang

    2008-01-01

    Cocoa was shown to inhibit chemically induced carcinogenesis in animals and exert antioxidant activity in humans. However, the molecular mechanisms of the chemopreventive potential of cocoa and its active ingredient(s) remain unknown. Here we report that cocoa procyanidins inhibit neoplastic cell transformation by suppressing the kinase activity of mitogen-activated protein kinase kinase (MEK). A cocoa procyanidin fraction (CPF) and procyanidin B2 at 5 μg/ml and 40 μm, respectively, inhibited 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced neoplastic transformation of JB6 P+ mouse epidermal (JB6 P+) cells by 47 and 93%, respectively. The TPA-induced promoter activity and expression of cyclooxygenase-2, which is involved in tumor promotion and inflammation, were dose-dependently inhibited by CPF or procyanidin B2. The activation of activator protein-1 and nuclear factor-κB induced by TPA was also attenuated by CPF or procyanidin B2. The TPA-induced phosphorylation of MEK, extracellular signal-regulated kinase, and p90 ribosomal s6 kinase was suppressed by CPF or procyanidin B2. In vitro and ex vivo kinase assay data demonstrated that CPF or procyanidin B2 inhibited the kinase activity of MEK1 and directly bound with MEK1. CPF or procyanidin B2 suppressed JB6 P+ cell transformation induced by epidermal growth factor or H-Ras, both of which are known to be involved in MEK/ERK signal activation. In contrast, theobromine (up to 80 μm) had no effect on TPA-induced transformation, cyclooxygenase-2 expression, the transactivation of activator protein-1 or nuclear factor-κB, or MEK. Notably, procyanidin B2 exerted stronger inhibitory effects compared with PD098059 (a well known pharmacological inhibitor of MEK) on MEK1 activity and neoplastic cell transformation. PMID:18519570

  2. LPIN1 deficiency with severe recurrent rhabdomyolysis and persistent elevation of creatine kinase levels due to chromosome 2 maternal isodisomy

    Directory of Open Access Journals (Sweden)

    I.A. Meijer

    2015-12-01

    Full Text Available Fatty acid oxidation disorders and lipin-1 deficiency are the commonest genetic causes of rhabdomyolysis in children. We describe a lipin-1-deficient boy with recurrent, severe rhabdomyolytic episodes from the age of 4 years. Analysis of the LPIN1 gene that encodes lipin-1 revealed a novel homozygous frameshift mutation in exon 9, c.1381delC (p.Leu461SerfsX47, and complete uniparental isodisomy of maternal chromosome 2. This mutation is predicted to cause complete lipin-1 deficiency. The patient had six rhabdomyolytic crises, with creatine kinase (CK levels up to 300,000 U/L (normal, 30 to 200. Plasma CK remained elevated between crises. A treatment protocol was instituted, with early aggressive monitoring, hydration, electrolyte replacement and high caloric, high carbohydrate intake. The patient received dexamethasone during two crises, which was well-tolerated and in these episodes, peak CK values were lower than in preceding episodes. Studies of anti-inflammatory therapy may be indicated in lipin-1 deficiency.

  3. Creatine supplementation with specific view to exercise/sports performance: an update

    Directory of Open Access Journals (Sweden)

    Cooper Robert

    2012-07-01

    Full Text Available Abstract Creatine is one of the most popular and widely researched natural supplements. The majority of studies have focused on the effects of creatine monohydrate on performance and health; however, many other forms of creatine exist and are commercially available in the sports nutrition/supplement market. Regardless of the form, supplementation with creatine has regularly shown to increase strength, fat free mass, and muscle morphology with concurrent heavy resistance training more than resistance training alone. Creatine may be of benefit in other modes of exercise such as high-intensity sprints or endurance training. However, it appears that the effects of creatine diminish as the length of time spent exercising increases. Even though not all individuals respond similarly to creatine supplementation, it is generally accepted that its supplementation increases creatine storage and promotes a faster regeneration of adenosine triphosphate between high intensity exercises. These improved outcomes will increase performance and promote greater training adaptations. More recent research suggests that creatine supplementation in amounts of 0.1 g/kg of body weight combined with resistance training improves training adaptations at a cellular and sub-cellular level. Finally, although presently ingesting creatine as an oral supplement is considered safe and ethical, the perception of safety cannot be guaranteed, especially when administered for long period of time to different populations (athletes, sedentary, patient, active, young or elderly.

  4. Effects of Combined Creatine Plus Fenugreek Extract vs. Creatine Plus Carbohydrate Supplementation on Resistance Training Adaptations

    Science.gov (United States)

    Taylor, Lem; Poole, Chris; Pena, Earnest; Lewing, Morgan; Kreider, Richard; Foster, Cliffa; Wilborn, Colin

    2011-01-01

    The purpose of this study was to evaluate the effects of combined creatine and fenugreek extract supplementation on strength and body composition. Forty- seven resistance trained men were matched according to body weight to ingest either 70 g of a dextrose placebo (PL), 5 g creatine/70 g of dextrose (CRD) or 3.5 g creatine/900 mg fenugreek extract (CRF) and participate in a 4-d/wk periodized resistance-training program for 8-weeks. At 0, 4, and 8-weeks, subjects were tested on body composition, muscular strength and endurance, and anaerobic capacity. Statistical analyses utilized a separate 3X3 (condition [PL vs. CRD vs. CRF] x time [T1 vs. T2 vs. T3]) ANOVAs with repeated measures for all criterion variables (p ≤ 0.05). No group x time interaction effects or main effects (p > 0.05) were observed for any measures of body composition. CRF group showed significant increases in lean mass at T2 (p = 0.001) and T3 (p = 0.001). Bench press 1RM increased in PL group (p = 0.050) from T1-T3 and in CRD from T1-T2 (p = 0. 001) while remaining significant at T3 (p 0.05). In conclusion, creatine plus fenugreek extract supplementation had a significant impact on upper body strength and body composition as effectively as the combination of 5g of creatine with 70g of dextrose. Thus, the use of fenugreek with creatine supplementation may be an effective means for enhancing creatine uptake while eliminating the need for excessive amounts of simple carbohydrates. Key points Fenugreek plus creatine supplementation may be a new means of increasing creatine uptake. Creatine plus fenugreek seems to be just as effective as the classic creatine plus carbohydrate ingestion in terms of stimulating training adaptations. This is the first study to our knowledge that has combined fenugreek with creatine supplementation in conjunction with a resistance training program. PMID:24149869

  5. Contractions activate hormone-sensitive lipase in rat muscle by protein kinase C and mitogen-activated protein kinase

    DEFF Research Database (Denmark)

    Donsmark, Morten; Langfort, Jozef; Holm, Cecilia

    2003-01-01

    and contractions. Adrenaline acts via cAMP-dependent protein kinase (PKA). The signalling mediating the effect of contractions is unknown and was explored in this study. Incubated soleus muscles from 70 g male rats were electrically stimulated to perform repeated tetanic contractions for 5 min. The contraction......Intramuscular triacylglycerol is an important energy store and is also related to insulin resistance. The mobilization of fatty acids from this pool is probably regulated by hormone-sensitive lipase (HSL), which has recently been shown to exist in muscle and to be activated by both adrenaline......-induced activation of HSL was abolished by the protein kinase C (PKC) inhibitors bisindolylmaleimide I and calphostin C and reduced 50% by the mitogen-activated protein kinase kinase (MEK) inhibitor U0126, which also completely blocked extracellular signal-regulated kinase (ERK) 1 and 2 phosphorylation. None...

  6. Guanidinoacetic acid versus creatine for improved brain and muscle creatine levels: a superiority pilot trial in healthy men.

    Science.gov (United States)

    Ostojic, Sergej M; Ostojic, Jelena; Drid, Patrik; Vranes, Milan

    2016-09-01

    In this randomized, double-blind, crossover trial, we evaluated whether 4-week supplementation with guanidinoacetic acid (GAA) is superior to creatine in facilitating creatine levels in healthy men (n = 5). GAA (3.0 g/day) resulted in a more powerful rise (up to 16.2%) in tissue creatine levels in vastus medialis muscle, middle-cerebellar peduncle, and paracentral grey matter, as compared with creatine (P creatine for improved bioenergetics in energy-demanding tissues.

  7. Myxedema coma of both primary and secondary origin, with non-classic presentation and extremely elevated creatine kinase.

    Science.gov (United States)

    Benvenga, S; Squadrito, S; Saporito, F; Cimino, A; Arrigo, F; Trimarchi, F

    2000-09-01

    Myxedema coma is a rare, often fatal endocrine emergency that concerns elderly patients with long-standing primary hypothyroidism; myxedema coma of central origin is exceedingly rare. Here, we report a 37-year-old woman in whom classical symptoms of hypothyroidism had been absent. Six years earlier, she had severe obstetric hemorrhage and, shortly after, two subsequent episodes of pericardial effusion. On the day of admission, pericardiocentesis was performed for the third episode of pericardial effusion. Because of the subsequent grave arrhythmias and unconsciousness, she was transferred to our ICU. Prior to the endocrine consultation, a silent myocardial infarction had been suspected, based on the extremely high serum levels of creatine kinase (CK) and isoenzyme CK-MB. However, based on thyroid sonography, pituitary computed tomography, elevated titers of antithyroid antibodies and pituitary stimulation tests, the final diagnosis was myxedema coma of dual origin: an atrophic variant of Hashimoto's thyroiditis and post-necrotic pituitary atrophy (Sheehan syndrome). Substitutive therapy caused a prompt clinical amelioration and normalization of CK levels. Our patient is the first case of myxedema coma of double etiology, and illustrates how its presentation deviates markedly from the one endocrinologists and physicians at ICU are prepared to encounter. In addition, cardiac problems as those of our patient should not discourage from substitutive treatment (using L-thyroxine and the gastrointestinal route of absorption), if the age is relatively low.

  8. Regulation of energy metabolism during social interactions in rainbow trout: a role for AMP-activated protein kinase.

    Science.gov (United States)

    Gilmour, K M; Craig, P M; Dhillon, R S; Lau, G Y; Richards, J G

    2017-11-01

    Rainbow trout ( Oncorhynchus mykiss ) confined in pairs form social hierarchies in which subordinate fish typically experience fasting and high circulating cortisol levels, resulting in low growth rates. The present study investigated the role of AMP-activated protein kinase (AMPK) in mediating metabolic adjustments associated with social status in rainbow trout. After 3 days of social interaction, liver AMPK activity was significantly higher in subordinate than dominant or sham (fish handled in the same fashion as paired fish but held individually) trout. Elevated liver AMPK activity in subordinate fish likely reflected a significantly higher ratio of phosphorylated AMPK (phospho-AMPK) to total AMPK protein, which was accompanied by significantly higher AMPKα 1 relative mRNA abundance. Liver ATP and creatine phosphate concentrations in subordinate fish also were elevated, perhaps as a result of AMPK activity. Sham fish that were fasted for 3 days exhibited effects parallel to those of subordinate fish, suggesting that low food intake was an important trigger of elevated AMPK activity in subordinate fish. Effects on white muscle appeared to be influenced by the physical activity associated with social interaction. Overall, muscle AMPK activity was significantly higher in dominant and subordinate than sham fish. The ratio of phospho-AMPK to total AMPK protein in muscle was highest in subordinate fish, while muscle AMPKα 1 relative mRNA abundance was elevated by social dominance. Muscle ATP and creatine phosphate concentrations were high in dominant and subordinate fish at 6 h of interaction and decreased significantly thereafter. Collectively, the findings of the present study support a role for AMPK in mediating liver and white muscle metabolic adjustments associated with social hierarchy formation in rainbow trout. Copyright © 2017 the American Physiological Society.

  9. Beyond muscles: The untapped potential of creatine.

    Science.gov (United States)

    Riesberg, Lisa A; Weed, Stephanie A; McDonald, Thomas L; Eckerson, Joan M; Drescher, Kristen M

    2016-08-01

    Creatine is widely used by both elite and recreational athletes as an ergogenic aid to enhance anaerobic exercise performance. Older individuals also use creatine to prevent sarcopenia and, accordingly, may have therapeutic benefits for muscle wasting diseases. Although the effect of creatine on the musculoskeletal system has been extensively studied, less attention has been paid to its potential effects on other physiological systems. Because there is a significant pool of creatine in the brain, the utility of creatine supplementation has been examined in vitro as well as in vivo in both animal models of neurological disorders and in humans. While the data are preliminary, there is evidence to suggest that individuals with certain neurological conditions may benefit from exogenous creatine supplementation if treatment protocols can be optimized. A small number of studies that have examined the impact of creatine on the immune system have shown an alteration in soluble mediator production and the expression of molecules involved in recognizing infections, specifically toll-like receptors. Future investigations evaluating the total impact of creatine supplementation are required to better understand the benefits and risks of creatine use, particularly since there is increasing evidence that creatine may have a regulatory impact on the immune system. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. p21-activated Kinase1(PAK1) can promote ERK activation in a kinase independent manner

    DEFF Research Database (Denmark)

    Wang, Zhipeng; Fu, Meng; Wang, Lifeng

    2013-01-01

    204) although phosphorylation of b-Raf (Ser445) and c-Raf (Ser 338) remained unchanged. Furthermore, increased activation of the PAK1 activator Rac1 induced the formation of a triple complex of Rac1, PAK1 and Mek1, independent of the kinase activity of PAK1. These data suggest that PAK1 can stimulate...... MEK activity in a kinase independent manner, probably by serving as a scaffold to facilitate interaction of c-Raf....

  11. Mitogen-activated protein kinases interacting kinases are autoinhibited by a reprogrammed activation segment.

    Science.gov (United States)

    Jauch, Ralf; Cho, Min-Kyu; Jäkel, Stefan; Netter, Catharina; Schreiter, Kay; Aicher, Babette; Zweckstetter, Markus; Jäckle, Herbert; Wahl, Markus C

    2006-09-06

    Autoinhibition is a recurring mode of protein kinase regulation and can be based on diverse molecular mechanisms. Here, we show by crystal structure analysis, nuclear magnetic resonance (NMR)-based nucleotide affinity studies and rational mutagenesis that nonphosphorylated mitogen-activated protein (MAP) kinases interacting kinase (Mnk) 1 is autoinhibited by conversion of the activation segment into an autoinhibitory module. In a Mnk1 crystal structure, the activation segment is repositioned via a Mnk-specific sequence insertion at the N-terminal lobe with the following consequences: (i) the peptide substrate binding site is deconstructed, (ii) the interlobal cleft is narrowed, (iii) an essential Lys-Glu pair is disrupted and (iv) the magnesium-binding loop is locked into an ATP-competitive conformation. Consistently, deletion of the Mnk-specific insertion or removal of a conserved phenylalanine side chain, which induces a blockade of the ATP pocket, increase the ATP affinity of Mnk1. Structural rearrangements required for the activation of Mnks are apparent from the cocrystal structure of a Mnk2 D228G -staurosporine complex and can be modeled on the basis of crystal packing interactions. Our data suggest a novel regulatory mechanism specific for the Mnk subfamily.

  12. The cellular and compartmental profile of mouse retinal glycolysis, tricarboxylic acid cycle, oxidative phosphorylation, and ~P transferring kinases.

    Science.gov (United States)

    Rueda, Elda M; Johnson, Jerry E; Giddabasappa, Anand; Swaroop, Anand; Brooks, Matthew J; Sigel, Irena; Chaney, Shawnta Y; Fox, Donald A

    2016-01-01

    The homeostatic regulation of cellular ATP is achieved by the coordinated activity of ATP utilization, synthesis, and buffering. Glucose is the major substrate for ATP synthesis through glycolysis and oxidative phosphorylation (OXPHOS), whereas intermediary metabolism through the tricarboxylic acid (TCA) cycle utilizes non-glucose-derived monocarboxylates, amino acids, and alpha ketoacids to support mitochondrial ATP and GTP synthesis. Cellular ATP is buffered by specialized equilibrium-driven high-energy phosphate (~P) transferring kinases. Our goals were twofold: 1) to characterize the gene expression, protein expression, and activity of key synthesizing and regulating enzymes of energy metabolism in the whole mouse retina, retinal compartments, and/or cells and 2) to provide an integrative analysis of the results related to function. mRNA expression data of energy-related genes were extracted from our whole retinal Affymetrix microarray data. Fixed-frozen retinas from adult C57BL/6N mice were used for immunohistochemistry, laser scanning confocal microscopy, and enzymatic histochemistry. The immunoreactivity levels of well-characterized antibodies, for all major retinal cells and their compartments, were obtained using our established semiquantitative confocal and imaging techniques. Quantitative cytochrome oxidase (COX) and lactate dehydrogenase (LDH) activity was determined histochemically. The Affymetrix data revealed varied gene expression patterns of the ATP synthesizing and regulating enzymes found in the muscle, liver, and brain. Confocal studies showed differential cellular and compartmental distribution of isozymes involved in glucose, glutamate, glutamine, lactate, and creatine metabolism. The pattern and intensity of the antibodies and of the COX and LDH activity showed the high capacity of photoreceptors for aerobic glycolysis and OXPHOS. Competition assays with pyruvate revealed that LDH-5 was localized in the photoreceptor inner segments. The

  13. 21 CFR 862.1210 - Creatine test system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Creatine test system. 862.1210 Section 862.1210....1210 Creatine test system. (a) Identification. A creatine test system is a device intended to measure creatine (a substance synthesized in the liver and pancreas and found in biological fluids) in plasma...

  14. Treatment of Creatine Transporter (SLC6A8) Deficiency With Oral S-Adenosyl Methionine as Adjunct to L-arginine, Glycine, and Creatine Supplements.

    Science.gov (United States)

    Jaggumantri, Sravan; Dunbar, Mary; Edgar, Vanessa; Mignone, Cristina; Newlove, Theresa; Elango, Rajavel; Collet, Jean Paul; Sargent, Michael; Stockler-Ipsiroglu, Sylvia; van Karnebeek, Clara D M

    2015-10-01

    Creatine transporter (SLC6A8) deficiency is an X-linked inborn error of metabolism characterized by cerebral creatine deficiency, behavioral problems, seizures, hypotonia, and intellectual developmental disability. A third of patients are amenable to treatment with high-dose oral creatine, glycine, and L-arginine supplementation. Given the limited treatment response, we initiated an open-label observational study to evaluate the effect of adjunct S-adenosyl methionine to further enhance intracerebral creatine synthesis. Significant and reproducible issues with sleep and behavior were noted in both male patients on a dose of 50/mg/kg. One of the two patients stopped S-adenosyl methionine and did not come for any follow-up. A safe and tolerable dose (17 mg/kg/day) was identified in the other patient. On magnetic resonance spectroscopy, this 8-year-old male did not show an increase in intracerebral creatine. However, significant improvement in speech/language skills, muscle mass were observed as well as in personal outcomes as defined by the family in activities related to communication and decision making. Further research is needed to assess the potential of S-adenosyl methionine as an adjunctive therapy for creatine transporter deficiency patients and to define the optimal dose. Our study also illustrates the importance of pathophysiology-based treatment, individualized outcome assessment, and patient/family participation in rare diseases research. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. The effects of creatine pyruvate and creatine citrate on performance during high intensity exercise

    Directory of Open Access Journals (Sweden)

    Purpura Martin

    2008-02-01

    Full Text Available Abstract Background A double-blind, placebo-controlled, randomized study was performed to evaluate the effect of oral creatine pyruvate (Cr-Pyr and creatine citrate (Cr-Cit supplementation on exercise performance in healthy young athletes. Methods Performance during intermittent handgrip exercise of maximal intensity was evaluated before (pretest and after (posttest 28 days of Cr-Pyr (5 g/d, n = 16, Cr-Cit (5 g/d, n = 16 or placebo (pla, 5 g/d, n = 17 intake. Subjects performed ten 15-sec exercise intervals, each followed by 45 sec rest periods. Results Cr-Pyr (p Conclusion It is concluded that four weeks of Cr-Pyr and Cr-Cit intake significantly improves performance during intermittent handgrip exercise of maximal intensity and that Cr-Pyr might benefit endurance, due to enhanced activity of the aerobic metabolism.

  16. Creatine synthesis and exchanges between brain cells: What can be learned from human creatine deficiencies and various experimental models?

    Science.gov (United States)

    Hanna-El-Daher, Layane; Braissant, Olivier

    2016-08-01

    While it has long been thought that most of cerebral creatine is of peripheral origin, the last 20 years has provided evidence that the creatine synthetic pathway (AGAT and GAMT enzymes) is expressed in the brain together with the creatine transporter (SLC6A8). It has also been shown that SLC6A8 is expressed by microcapillary endothelial cells at the blood-brain barrier, but is absent from surrounding astrocytes, raising the concept that the blood-brain barrier has a limited permeability for peripheral creatine. The first creatine deficiency syndrome in humans was also discovered 20 years ago (GAMT deficiency), followed later by AGAT and SLC6A8 deficiencies, all three diseases being characterized by creatine deficiency in the CNS and essentially affecting the brain. By reviewing the numerous and latest experimental studies addressing creatine transport and synthesis in the CNS, as well as the clinical and biochemical characteristics of creatine-deficient patients, our aim was to delineate a clearer view of the roles of the blood-brain and blood-cerebrospinal fluid barriers in the transport of creatine and guanidinoacetate between periphery and CNS, and on the intracerebral synthesis and transport of creatine. This review also addresses the question of guanidinoacetate toxicity for brain cells, as probably found under GAMT deficiency.

  17. Creatine affords protection against glutamate-induced nitrosative and oxidative stress.

    Science.gov (United States)

    Cunha, Mauricio P; Lieberknecht, Vicente; Ramos-Hryb, Ana Belén; Olescowicz, Gislaine; Ludka, Fabiana K; Tasca, Carla I; Gabilan, Nelson H; Rodrigues, Ana Lúcia S

    2016-05-01

    Creatine has been reported to exert beneficial effects in several neurodegenerative diseases in which glutamatergic excitotoxicity and oxidative stress play an etiological role. The purpose of this study was to investigate the protective effects of creatine, as compared to the N-Methyl-d-Aspartate (NMDA) receptor antagonist dizocilpine (MK-801), against glutamate or hydrogen peroxide (H2O2)-induced injury in human neuroblastoma SH-SY5Y cells. Exposure of cells to glutamate (60-80 mM) or H2O2 (200-300 μM) for 24 h decreased cellular viability and increased dichlorofluorescein (DCF) fluorescence (indicative of increased reactive oxygen species, ROS) and nitric oxide (NO) production (assessed by mono-nitrogen oxides, NOx, levels). Creatine (1-10 mM) or MK-801 (0.1-10 μM) reduced glutamate- and H2O2-induced toxicity. The protective effect of creatine against glutamate-induced toxicity involves its antioxidant effect, since creatine, similar to MK-801, prevented the increase on DCF fluorescence induced by glutamate or H2O2. Furthermore, creatine or MK-801 blocked glutamate- and H2O2-induced increases in NOx levels. In another set of experiments, the repeated, but not acute, administration of creatine (300 mg/kg, po) in mice prevented the decreases on cellular viability and mitochondrial membrane potential (assessed by tetramethylrhodamine ethyl ester, TMRE, probe) of hippocampal slices incubated with glutamate (10 mM). Creatine concentration-dependent decreased the amount of nitrite formed in the reaction of oxygen with NO produced from sodium nitroprusside solution, suggesting that its protective effect against glutamate or H2O2-induced toxicity might be due to its scavenger activity. Overall, the results suggest that creatine may be useful as adjuvant therapy for neurodegenerative disease treatments. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Hierarchical modeling of activation mechanisms in the ABL and EGFR kinase domains: thermodynamic and mechanistic catalysts of kinase activation by cancer mutations.

    Directory of Open Access Journals (Sweden)

    Anshuman Dixit

    2009-08-01

    Full Text Available Structural and functional studies of the ABL and EGFR kinase domains have recently suggested a common mechanism of activation by cancer-causing mutations. However, dynamics and mechanistic aspects of kinase activation by cancer mutations that stimulate conformational transitions and thermodynamic stabilization of the constitutively active kinase form remain elusive. We present a large-scale computational investigation of activation mechanisms in the ABL and EGFR kinase domains by a panel of clinically important cancer mutants ABL-T315I, ABL-L387M, EGFR-T790M, and EGFR-L858R. We have also simulated the activating effect of the gatekeeper mutation on conformational dynamics and allosteric interactions in functional states of the ABL-SH2-SH3 regulatory complexes. A comprehensive analysis was conducted using a hierarchy of computational approaches that included homology modeling, molecular dynamics simulations, protein stability analysis, targeted molecular dynamics, and molecular docking. Collectively, the results of this study have revealed thermodynamic and mechanistic catalysts of kinase activation by major cancer-causing mutations in the ABL and EGFR kinase domains. By using multiple crystallographic states of ABL and EGFR, computer simulations have allowed one to map dynamics of conformational fluctuations and transitions in the normal (wild-type and oncogenic kinase forms. A proposed multi-stage mechanistic model of activation involves a series of cooperative transitions between different conformational states, including assembly of the hydrophobic spine, the formation of the Src-like intermediate structure, and a cooperative breakage and formation of characteristic salt bridges, which signify transition to the active kinase form. We suggest that molecular mechanisms of activation by cancer mutations could mimic the activation process of the normal kinase, yet exploiting conserved structural catalysts to accelerate a conformational transition

  19. CIKS, a connection to Ikappa B kinase and stress-activated protein kinase.

    Science.gov (United States)

    Leonardi, A; Chariot, A; Claudio, E; Cunningham, K; Siebenlist, U

    2000-09-12

    Pathogens, inflammatory signals, and stress cause acute transcriptional responses in cells. The induced expression of genes in response to these signals invariably involves transcription factors of the NF-kappaB and AP-1/ATF families. Activation of NF-kappaB factors is thought to be mediated primarily via IkappaB kinases (IKK), whereas that of AP-1/ATF can be mediated by stress-activated protein kinases (SAPKs; also named Jun kinases or JNKs). IKKalpha and IKKbeta are two catalytic subunits of a core IKK complex that also contains the regulatory subunit NEMO (NF-kappaB essential modulator)/IKKgamma. The latter protein is essential for activation of the IKKs, but its mechanism of action is not known. Here we describe the molecular cloning of CIKS (connection to IKK and SAPK/JNK), a previously unknown protein that directly interacts with NEMO/IKKgamma in cells. When ectopically expressed, CIKS stimulates IKK and SAPK/JNK kinases and it transactivates an NF-kappaB-dependent reporter. Activation of NF-kappaB is prevented in the presence of kinase-deficient, interfering mutants of the IKKs. CIKS may help to connect upstream signaling events to IKK and SAPK/JNK modules. CIKS could coordinate the activation of two stress-induced signaling pathways, functions reminiscent of those noted for tumor necrosis factor receptor-associated factor adaptor proteins.

  20. CIKS, a connection to IκB kinase and stress-activated protein kinase

    Science.gov (United States)

    Leonardi, Antonio; Chariot, Alain; Claudio, Estefania; Cunningham, Kirk; Siebenlist, Ulrich

    2000-01-01

    Pathogens, inflammatory signals, and stress cause acute transcriptional responses in cells. The induced expression of genes in response to these signals invariably involves transcription factors of the NF-κB and AP-1/ATF families. Activation of NF-κB factors is thought to be mediated primarily via IκB kinases (IKK), whereas that of AP-1/ATF can be mediated by stress-activated protein kinases (SAPKs; also named Jun kinases or JNKs). IKKα and IKKβ are two catalytic subunits of a core IKK complex that also contains the regulatory subunit NEMO (NF-κB essential modulator)/IKKγ. The latter protein is essential for activation of the IKKs, but its mechanism of action is not known. Here we describe the molecular cloning of CIKS (connection to IKK and SAPK/JNK), a previously unknown protein that directly interacts with NEMO/IKKγ in cells. When ectopically expressed, CIKS stimulates IKK and SAPK/JNK kinases and it transactivates an NF-κB-dependent reporter. Activation of NF-κB is prevented in the presence of kinase-deficient, interfering mutants of the IKKs. CIKS may help to connect upstream signaling events to IKK and SAPK/JNK modules. CIKS could coordinate the activation of two stress-induced signaling pathways, functions reminiscent of those noted for tumor necrosis factor receptor-associated factor adaptor proteins. PMID:10962033

  1. Genetics Home Reference: X-linked creatine deficiency

    Science.gov (United States)

    ... Health Conditions X-linked creatine deficiency X-linked creatine deficiency Printable PDF Open All Close All Enable ... view the expand/collapse boxes. Description X-linked creatine deficiency is an inherited disorder that primarily affects ...

  2. SH2 domains: modulators of nonreceptor tyrosine kinase activity.

    Science.gov (United States)

    Filippakopoulos, Panagis; Müller, Susanne; Knapp, Stefan

    2009-12-01

    The Src homology 2 (SH2) domain is a sequence-specific phosphotyrosine-binding module present in many signaling molecules. In cytoplasmic tyrosine kinases, the SH2 domain is located N-terminally to the catalytic kinase domain (SH1) where it mediates cellular localization, substrate recruitment, and regulation of kinase activity. Initially, structural studies established a role of the SH2 domain stabilizing the inactive state of Src family members. However, biochemical characterization showed that the presence of the SH2 domain is frequently required for catalytic activity, suggesting a crucial function stabilizing the active state of many nonreceptor tyrosine kinases. Recently, the structure of the SH2-kinase domain of Fes revealed that the SH2 domain stabilizes the active kinase conformation by direct interactions with the regulatory helix alphaC. Stabilizing interactions between the SH2 and the kinase domains have also been observed in the structures of active Csk and Abl. Interestingly, mutations in the SH2 domain found in human disease can be explained by SH2 domain destabilization or incorrect positioning of the SH2. Here we summarize our understanding of mechanisms that lead to tyrosine kinase activation by direct interactions mediated by the SH2 domain and discuss how mutations in the SH2 domain trigger kinase inactivation.

  3. X-linked creatine transporter deficiency: clinical aspects and pathophysiology

    NARCIS (Netherlands)

    van de Kamp, J.M.; Mancini, G.M.; Salomons, G.S.

    2014-01-01

    Creatine transporter deficiency was discovered in 2001 as an X-linked cause of intellectual disability characterized by cerebral creatine deficiency. This review describes the current knowledge regarding creatine metabolism, the creatine transporter and the clinical aspects of creatine transporter

  4. URINARY CREATINE AT REST AND AFTER REPEATED SPRINTS IN ATHLETES: A PILOT STUDY

    Science.gov (United States)

    Nasrallah, F.; Feki, M.; Chamari, K.; Omar, S.; Alouane-Trabelsi, L.; Ben Mansour, A.; Kaabachi, N.

    2014-01-01

    Creatine plays a key role in muscle function and its evaluation is important in athletes. In this study, urinary creatine concentration was measured in order to highlight its possible significance in monitoring sprinters. The study included 51 sprinters and 25 age- and sex-matched untrained subjects as a control group. Body composition was measured and dietary intake estimated. Urine samples were collected before and after standardized physical exercise. Creatine was assessed by gas chromatography mass spectrometry. Basal urinary creatine (UC) was significantly lower in sprinters than controls (34±30 vs. 74±3 µmol/mmol creatinine, p creatine significantly decreased in both athletes and controls. UC is low in sprinters at rest and further decreases after exercise, most likely due to a high uptake and use of creatine by muscles, as muscle mass and physical activity are supposed to be greater in athletes than untrained subjects. Further studies are needed to test the value of urinary creatine as a non-invasive marker of physical condition and as a parameter for managing Cr supplementation in athletes. PMID:24917689

  5. ROS and CDPK-like kinase-mediated activation of MAP kinase in rice roots exposed to lead.

    Science.gov (United States)

    Huang, Tsai-Lien; Huang, Hao-Jen

    2008-04-01

    Lead (Pb2+) is a cytotoxic metal ion in plants, the mechanism of which is not yet established. The aim of this study is to investigate the signalling pathways that are activated by elevated concentrations of Pb2+ in rice roots. Root growth was stunted and cell death was accelerated when exposed to different dosages of Pb2+ during extended time periods. Using ROS-sensitive dye and Ca2+ indicator, we demonstrated that Pb2+ induced ROS production and Ca2+ accumulation, respectively. In addition, Pb2+ elicited a remarkable increase in myelin basic protein (MBP) kinase activities. By immunoblot and immunoprecipitation analysis, 40- and 42-kDa MBP kinases that were activated by Pb2+ were identified to be mitogen-activated protein (MAP) kinases. Pre-treatment of rice roots with an antioxidant and a NADPH oxidase inhibitor, glutathione (GSH) and diphenylene iodonium (DPI), effectively reduced Pb2+-induced cell death and MAP kinase activation. Moreover, calcium-dependent protein kinase (CDPK) antagonist, W7, attenuated Pb2+-induced cell death and MAP kinase activation. These results suggested that the ROS and CDPK may function in the Pb2+-triggered cell death and MAP kinase signalling pathway in rice roots.

  6. The cataract and glucosuria associated monocarboxylate transporter MCT12 is a new creatine transporter

    Science.gov (United States)

    Abplanalp, Jeannette; Laczko, Endre; Philp, Nancy J.; Neidhardt, John; Zuercher, Jurian; Braun, Philipp; Schorderet, Daniel F.; Munier, Francis L.; Verrey, François; Berger, Wolfgang; Camargo, Simone M.R.; Kloeckener-Gruissem, Barbara

    2013-01-01

    Creatine transport has been assigned to creatine transporter 1 (CRT1), encoded by mental retardation associated SLC6A8. Here, we identified a second creatine transporter (CRT2) known as monocarboxylate transporter 12 (MCT12), encoded by the cataract and glucosuria associated gene SLC16A12. A non-synonymous alteration in MCT12 (p.G407S) found in a patient with age-related cataract (ARC) leads to a significant reduction of creatine transport. Furthermore, Slc16a12 knockout (KO) rats have elevated creatine levels in urine. Transport activity and expression characteristics of the two creatine transporters are distinct. CRT2 (MCT12)-mediated uptake of creatine was not sensitive to sodium and chloride ions or creatine biosynthesis precursors, breakdown product creatinine or creatine phosphate. Increasing pH correlated with increased creatine uptake. Michaelis–Menten kinetics yielded a Vmax of 838.8 pmol/h/oocyte and a Km of 567.4 µm. Relative expression in various human tissues supports the distinct mutation-associated phenotypes of the two transporters. SLC6A8 was predominantly found in brain, heart and muscle, while SLC16A12 was more abundant in kidney and retina. In the lens, the two transcripts were found at comparable levels. We discuss the distinct, but possibly synergistic functions of the two creatine transporters. Our findings infer potential preventive power of creatine supplementation against the most prominent age-related vision impaired condition. PMID:23578822

  7. Beyond Muscles: The Untapped Potential of Creatine

    OpenAIRE

    Riesberg, Lisa A.; Weed, Stephanie A.; McDonald, Thomas L.; Eckerson, Joan M.; Drescher, Kristen M.

    2016-01-01

    Creatine is widely used by both elite and recreational athletes as an ergogenic aid to enhance anaerobic exercise performance. Older individuals also use creatine to prevent sarcopenia and, accordingly, may have therapeutic benefits for muscle wasting diseases. Although the effect of creatine on the musculoskeletal system has been extensively studied, less attention has been paid to its potential effects on other physiological systems. Because there is a significant pool of creatine in the br...

  8. Creatine-induced activation of antioxidative defence in myotube cultures revealed by explorative NMR-based metabonomics and proteomics

    DEFF Research Database (Denmark)

    Young, Jette Feveile; Larsen, Lotte Bach; Malmendal, Anders

    2010-01-01

    Creatine is a key intermediate in energy metabolism and supplementation of creatine has been used for increasing muscle mass, strength and endurance. Creatine supplementation has also been reported to trigger the skeletal muscle expression of insulin like growth factor I, to increase the fat......-free mass and improve cognition in elderly, and more explorative approaches like transcriptomics has revealed additional information. The aim of the present study was to reveal additional insight into the biochemical effects of creatine supplementation at the protein and metabolite level by integrating...... the explorative techniques, proteomics and NMR metabonomics, in a systems biology approach. METHODS: Differentiated mouse myotube cultures (C2C12) were exposed to 5 mM creatine monohydrate (CMH) for 24 hours. For proteomics studies, lysed myotubes were analyzed in single 2-DGE gels where the first dimension...

  9. Effects of phorbol ester on mitogen-activated protein kinase kinase activity in wild-type and phorbol ester-resistant EL4 thymoma cells.

    Science.gov (United States)

    Gause, K C; Homma, M K; Licciardi, K A; Seger, R; Ahn, N G; Peterson, M J; Krebs, E G; Meier, K E

    1993-08-05

    Phorbol ester-sensitive and -resistant EL4 thymoma cell lines differ in their ability to activate mitogen-activated protein kinase (MAPK) in response to phorbol ester. Treatment of wild-type EL4 cells with phorbol ester results in the rapid activations of MAPK and pp90rsk kinase, a substrate for MAPK, while neither kinase is activated in response to phorbol ester in variant EL4 cells. This study examines the activation of MAPK kinase (MAPKK), an activator of MAPK, in wild-type and variant EL4 cells. Phosphorylation of a 40-kDa substrate, identified as MAPK, was observed following in vitro phosphorylation reactions using cytosolic extracts or Mono Q column fractions prepared from phorbol ester-treated wild-type EL4 cells. MAPKK activity coeluted with a portion of the inactive MAPK upon Mono Q anion-exchange chromatography, permitting detection of the MAPKK activity in fractions containing both kinases. This MAPKK activity was present in phorbol ester-treated wild-type cells, but not in phorbol ester-treated variant cells or in untreated wild-type or variant cells. The MAPKK from wild-type cells was able to activate MAPK prepared from either wild-type or variant cells. MAPKK activity could be stimulated in both wildtype and variant EL4 cells in response to treatment of cells with okadaic acid. These results indicate that the failure of variant EL4 cells to activate MAP kinase in response to phorbol ester is due to a failure to activate MAPKK. Therefore, the step that confers phorbol ester resistance to variant EL4 cells lies between the activation of protein kinase C and the activation of MAPKK.

  10. The use of creatine supplements in the military.

    Science.gov (United States)

    Havenetidis, Konstantinos

    2016-08-01

    Creatine is considered an effective nutritional ergogenic aid to enhance exercise performance. In spite of the publication of several reviews in the last decade on the topic of exercise performance/sports and creatine there is a need for an update related to the military given the lack of information in this area. The aim of this study was to critically assess original research addressing the use of creatine supplements in the military. A search of the electronic databases PubMed and SPORTDiscus, for the following key words: military personnel, trainees, recruit, soldier, physical fitness, physical conditioning, creatine supplementation, creatine ingestion, nutritional supplements to identify surveys and randomised clinical trials from journal articles and technical reports investigating the effect of creatine supplementation on military populations. Thirty-three out of 90 articles examined the use of creatine as a dietary supplement in military personnel. Twenty-one studies were finally selected on the basis of stated inclusion criteria for military surveys and randomised clinical trials. Most of the surveys (15/17) in the military indicate a high popularity of creatine (average 27%) among supplement users. In contrast, in most of the exercise protocols used (6/9) during randomised clinical trials creatine has produced a non-significant performance-enhancing effect. Creatine is one of the most widely used supplemental compounds in the military. It is not considered a doping infraction or related to any adverse health effects but its long-term usage needs further investigation. Experimental research suggests that creatine supplementation does not enhance physical performance in the military. However, limitations in creatine dosage, military fitness testing and sample group selection might have underestimated the ergogenic properties of creatine. Recent studies also indicate positive effects on various aspects of total force fitness such as cognitive

  11. A buffered form of creatine does not promote greater changes in muscle creatine content, body composition, or training adaptations than creatine monohydrate

    Directory of Open Access Journals (Sweden)

    Jagim Andrew R

    2012-09-01

    Full Text Available Abstract Background Creatine monohydrate (CrM has been consistently reported to increase muscle creatine content and improve high-intensity exercise capacity. However, a number of different forms of creatine have been purported to be more efficacious than CrM. The purpose of this study was to determine if a buffered creatine monohydrate (KA that has been purported to promote greater creatine retention and training adaptations with fewer side effects at lower doses is more efficacious than CrM supplementation in resistance-trained individuals. Methods In a double-blind manner, 36 resistance-trained participants (20.2 ± 2 years, 181 ± 7 cm, 82.1 ± 12 kg, and 14.7 ± 5% body fat were randomly assigned to supplement their diet with CrM (Creapure® AlzChem AG, Trostberg, Germany at normal loading (4 x 5 g/d for 7-days and maintenance (5 g/d for 21-days doses; KA (Kre-Alkalyn®, All American Pharmaceutical, Billings, MT, USA at manufacturer’s recommended doses (KA-L, 1.5 g/d for 28-days; or, KA with equivalent loading (4 x 5 g/d for 7-days and maintenance (5 g/d doses of CrM (KA-H. Participants were asked to maintain their current training programs and record all workouts. Muscle biopsies from the vastus lateralis, fasting blood samples, body weight, DEXA determined body composition, and Wingate Anaerobic Capacity (WAC tests were performed at 0, 7, and 28-days while 1RM strength tests were performed at 0 and 28-days. Data were analyzed by a repeated measures multivariate analysis of variance (MANOVA and are presented as mean ± SD changes from baseline after 7 and 28-days, respectively. Results Muscle free creatine content obtained in a subgroup of 25 participants increased in all groups over time (1.4 ± 20.7 and 11.9 ± 24.0 mmol/kg DW, p = 0.03 after 7 and 28-days, respectively, with no significant differences among groups (KA-L −7.9 ± 22.3, 4.7 ± 27.0; KA-H 1.0 ± 12.8, 9.1

  12. A buffered form of creatine does not promote greater changes in muscle creatine content, body composition, or training adaptations than creatine monohydrate.

    Science.gov (United States)

    Jagim, Andrew R; Oliver, Jonathan M; Sanchez, Adam; Galvan, Elfego; Fluckey, James; Riechman, Steven; Greenwood, Michael; Kelly, Katherine; Meininger, Cynthia; Rasmussen, Christopher; Kreider, Richard B

    2012-09-13

    Creatine monohydrate (CrM) has been consistently reported to increase muscle creatine content and improve high-intensity exercise capacity. However, a number of different forms of creatine have been purported to be more efficacious than CrM. The purpose of this study was to determine if a buffered creatine monohydrate (KA) that has been purported to promote greater creatine retention and training adaptations with fewer side effects at lower doses is more efficacious than CrM supplementation in resistance-trained individuals. In a double-blind manner, 36 resistance-trained participants (20.2 ± 2 years, 181 ± 7 cm, 82.1 ± 12 kg, and 14.7 ± 5% body fat) were randomly assigned to supplement their diet with CrM (Creapure® AlzChem AG, Trostberg, Germany) at normal loading (4 x 5 g/d for 7-days) and maintenance (5 g/d for 21-days) doses; KA (Kre-Alkalyn®, All American Pharmaceutical, Billings, MT, USA) at manufacturer's recommended doses (KA-L, 1.5 g/d for 28-days); or, KA with equivalent loading (4 x 5 g/d for 7-days) and maintenance (5 g/d) doses of CrM (KA-H). Participants were asked to maintain their current training programs and record all workouts. Muscle biopsies from the vastus lateralis, fasting blood samples, body weight, DEXA determined body composition, and Wingate Anaerobic Capacity (WAC) tests were performed at 0, 7, and 28-days while 1RM strength tests were performed at 0 and 28-days. Data were analyzed by a repeated measures multivariate analysis of variance (MANOVA) and are presented as mean ± SD changes from baseline after 7 and 28-days, respectively. Muscle free creatine content obtained in a subgroup of 25 participants increased in all groups over time (1.4 ± 20.7 and 11.9 ± 24.0 mmol/kg DW, p = 0.03) after 7 and 28-days, respectively, with no significant differences among groups (KA-L -7.9 ± 22.3, 4.7 ± 27.0; KA-H 1.0 ± 12.8, 9.1 ± 23.2; CrM 11.3 ± 23.9, 22.3 ± 21

  13. Interaction of human biliverdin reductase with Akt/protein kinase B and phosphatidylinositol-dependent kinase 1 regulates glycogen synthase kinase 3 activity: a novel mechanism of Akt activation.

    Science.gov (United States)

    Miralem, Tihomir; Lerner-Marmarosh, Nicole; Gibbs, Peter E M; Jenkins, Jermaine L; Heimiller, Chelsea; Maines, Mahin D

    2016-08-01

    Biliverdin reductase A (BVR) and Akt isozymes have overlapping pleiotropic functions in the insulin/PI3K/MAPK pathway. Human BVR (hBVR) also reduces the hemeoxygenase activity product biliverdin to bilirubin and is directly activated by insulin receptor kinase (IRK). Akt isoenzymes (Akt1-3) are downstream of IRK and are activated by phosphatidylinositol-dependent kinase 1 (PDK1) phosphorylating T(308) before S(473) autophosphorylation. Akt (RxRxxSF) and PDK1 (RFxFPxFS) binding motifs are present in hBVR. Phosphorylation of glycogen synthase kinase 3 (GSK3) isoforms α/β by Akts inhibits their activity; nonphosphorylated GSK3β inhibits activation of various genes. We examined the role of hBVR in PDK1/Akt1/GSK3 signaling and Akt1 in hBVR phosphorylation. hBVR activates phosphorylation of Akt1 at S(473) independent of hBVR's kinase competency. hBVR and Akt1 coimmunoprecipitated, and in-cell Förster resonance energy transfer (FRET) and glutathione S-transferase pulldown analyses identified Akt1 pleckstrin homology domain as the interactive domain. hBVR activates phosphorylation of Akt1 at S(473) independent of hBVR's kinase competency. Site-directed mutagenesis, mass spectrometry, and kinetic analyses identified S(230) in hBVR (225)RNRYLSF sequence as the Akt1 target. Underlined amino acids are the essential residues of the signaling motifs. In cells, hBVR-activated Akt1 increased both GSK3α/β and forkhead box of the O class transcription class 3 (FoxO3) phosphorylation and inhibited total GSK3 activity; depletion of hBVR released inhibition and stimulated glucose uptake. Immunoprecipitation analysis showed that PDK1 and hBVR interact through hBVR's PDK1 binding (161)RFGFPAFS motif and formation of the PDK1/hBVR/Akt1 complex. sihBVR blocked complex formation. Findings identify hBVR as a previously unknown coactivator of Akt1 and as a key mediator of Akt1/GSK3 pathway, as well as define a key role for hBVR in Akt1 activation by PDK1.-Miralem, T., Lerner

  14. Early detection of response in small cell bronchogenic carcinoma by changes in serum concentrations of creatine kinase, neuron specific enolase, calcitonin, ACTH, serotonin and gastrin releasing peptide

    DEFF Research Database (Denmark)

    Bork, E; Hansen, M; Urdal, P

    1988-01-01

    Creatine kinase (CK-BB), neuron specific enolase (NSE), ACTH, calcitonin, serotonin and gastrin releasing peptide (GRP) were measured in serum or plasma before and immediately after initiation of treatment in patients with small cell lung cancer (SCC). Pretherapeutic elevated concentrations of CK...... stage patients and 71% in limited stage patients. Frequent initial monitoring of the substances showed an increase in the concentrations of pretherapeutic elevated CK-BB and NSE on day 1 or 2 followed by a sharp decrease within 1 week. These changes were correlated to objective clinical response...... determined within 4-8 weeks. The results indicate that serum CK-BB and NSE are potential markers for SCC at the time of diagnosis and that changes in the concentrations during the first course of cytostatic therapy are promising as biochemical tests for early detection of response to chemotherapy....

  15. THE EFFECT OF CREATINE SUPPLEMENTATION ON ATHLETE ISOKINETIC PERFORMANCE

    OpenAIRE

    Erkan Faruk ŞİRİN; Suzan YALÇIN

    2009-01-01

    The purpose of this study is to find the effects of Creatin Monohydrate (CrH2O) on athlete performance (isokinetic power measured as a total workout) used as an ergojenic aid in long-term (6 weeks) supplementation. There are 38 participants willing to join to the study. Their ages are between 20 and 27. All of them are choosed from active athletes. From the findings of this study; all the participants’ in the creatin group have increased the total workout production in all cycles of isokineti...

  16. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    Directory of Open Access Journals (Sweden)

    Oyku Gulmez

    2008-12-01

    Full Text Available Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Aslı Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs during percutaneous coronary intervention (PCI has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB levels in patients undergoing elective PCI.Methods: A total of 570 patients who underwent PCI were evaluated for CK-MB elevation. Patients who were on CCB therapy when admitted to the hospital constituted the CCB group. No CCBs were given to the rest of the patients during the periprocedural period and these patients served as the control group. Blood samples for CK-MB were obtained before and at 6 h, 24 h, and 36 h after the procedure.Results: 217 patients were in the CCB group (mean age 60.2 ± 9.3 years, 162 males, and 353 were in the control group (mean age 60.0 ± 10.1 years, 262 males. CK-MB levels increased above the normal values in 41 patients (18.9% of the CCBs group and in 97 patients (27.5% of the control group (p = 0.02. Median CK-MB levels were significantly higher in the control group for all studied hours (for all p < 0.05.Conclusions: Prior oral CCB therapy may have favorable effects in preventing myocyte necrosis after elective PCI.Keywords: calcium channel blockers, myonecrosis, percutaneous coronary interventions

  17. Quantitative and Dynamic Imaging of ATM Kinase Activity.

    Science.gov (United States)

    Nyati, Shyam; Young, Grant; Ross, Brian Dale; Rehemtulla, Alnawaz

    2017-01-01

    Ataxia telangiectasia mutated (ATM) is a serine/threonine kinase critical to the cellular DNA-damage response, including DNA double-strand breaks (DSBs). ATM activation results in the initiation of a complex cascade of events facilitating DNA damage repair, cell cycle checkpoint control, and survival. Traditionally, protein kinases have been analyzed in vitro using biochemical methods (kinase assays using purified proteins or immunological assays) requiring a large number of cells and cell lysis. Genetically encoded biosensors based on optical molecular imaging such as fluorescence or bioluminescence have been developed to enable interrogation of kinase activities in live cells with a high signal to background. We have genetically engineered a hybrid protein whose bioluminescent activity is dependent on the ATM-mediated phosphorylation of a substrate. The engineered protein consists of the split luciferase-based protein complementation pair with a CHK2 (a substrate for ATM kinase activity) target sequence and a phospho-serine/threonine-binding domain, FHA2, derived from yeast Rad53. Phosphorylation of the serine residue within the target sequence by ATM would lead to its interaction with the phospho-serine-binding domain, thereby preventing complementation of the split luciferase pair and loss of reporter activity. Bioluminescence imaging of reporter expressing cells in cultured plates or as mouse xenografts provides a quantitative surrogate for ATM kinase activity and therefore the cellular DNA damage response in a noninvasive, dynamic fashion.

  18. Measuring Kinase Activity-A Global Challenge.

    Science.gov (United States)

    Cann, Marissa L; McDonald, Ian M; East, Michael P; Johnson, Gary L; Graves, Lee M

    2017-11-01

    The kinase enzymes within a cell, known collectively as the kinome, play crucial roles in many signaling pathways, including survival, motility, differentiation, stress response, and many more. Aberrant signaling through kinase pathways is often linked to cancer, among other diseases. A major area of scientific research involves understanding the relationships between kinases, their targets, and how the kinome adapts to perturbations of the cellular system. This review will discuss many of the current and developing methods for studying kinase activity, and evaluate their applications, advantages, and disadvantages. J. Cell. Biochem. 118: 3595-3606, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  19. Use of oral creatine as an ergogenic aid for increased sports performance: perceptions of adolescent athletes.

    Science.gov (United States)

    Ray, T R; Eck, J C; Covington, L A; Murphy, R B; Williams, R; Knudtson, J

    2001-06-01

    Competitive athletes, including adolescents, seek ways to gain advantage over competitors. One ergogenic aid is creatine, a naturally occurring nitrogen compound found primarily in skeletal muscle. Increasing creatine levels may prolong skeletal muscle activity, enhancing work output. A questionnaire assessing awareness and use of creatine supplementation was completed by 674 athletes from 11 high schools. Data were statistically analyzed to determine variation among groups. Of those surveyed, 75% had knowledge of creatine supplements, and 16% used creatine to enhance athletic performance. Percentage of use increased with age and grade level. Awareness and use were greater among boys than girls. Adverse effects were reported by 26%. Most athletes consumed creatine using a method inconsistent with scientific recommendations. Use of creatine by adolescent athletes is significant and inconsistent with optimal dosing. Physicians, athletic trainers, and coaches should disseminate proper information and advise these adolescent athletes.

  20. Effects of whole-body x irradiation on the biogenesis of creatine in the rat

    International Nuclear Information System (INIS)

    Thyagarajan, P.; Vakil, U.K.; Sreenivasan, A.

    1977-01-01

    Influences of whole-body x irradiation on various aspects of creatine metabolism have been studied. Exposures to sublethal or lethal doses of x radiation results in excessive urinary excretion as well as higher accumulation of creatine in the skeletal muscle of x-irradiated rats. A sudden fall in CPK activity in muscle with a concomitant rise in serum suggests that changes in serum and tissue CPK activity are of an adaptive nature in rats exposed to sublethal doses of x radiation. In vitro studies on creatine synthesis shows that transaminidase and methyl transferase activities in kidneys and liver, respectively, are decreased on the 5th day in the x-irradiated, are decreased on the 5th day in the x-irradiated rat. However, on the 8th day, the enzyme activities are restored to normal

  1. Protein phosphatases active on acetyl-CoA carboxylase phosphorylated by casein kinase I, casein kinase II and the cAMP-dependent protein kinase

    International Nuclear Information System (INIS)

    Witters, L.A.; Bacon, G.W.

    1985-01-01

    The protein phosphatases in rat liver cytosol, active on rat liver acetyl-CoA carboxylase (ACC) phosphorylated by casein kinase I, casein kinase II and the cAMP-dependent protein kinase, have been partially purified by anion-exchange and gel filtration chromatography. The major phosphatase activities against all three substrates copurify through fractionation and appear to be identical to protein phosphatases 2A1 and 2A2. No unique protein phosphatase active on 32 P-ACC phosphorylated by the casein kinases was identified

  2. Preventive effects of physical exercise on the inhibition of creatine kinase in the cerebral cortex of mice exposed to cigarette smoke. DOI: 10.5007/1980-0037.2011v13n2p106

    Directory of Open Access Journals (Sweden)

    Daiane Bittencourt Fraga

    2011-03-01

    Full Text Available Recent studies have shown the health benefits of physical exercise, increasing the oxidative response of muscle. However, the effects of exercise on the brain are poorly understood and contradictory. The inhibition of creatine kinase (CK activity has been associated with the pathogenesis of a large number of diseases, especially in the brain. The objective of this study was to determine the preventive effects of physical exercise in the hippocampus and cerebral cortex of mice after chronic cigarette smoke exposure. Eight to 10-week-old male mice (C57BL-6 were divided into four groups and submitted to an exercise program (swimming, 5 times a week, for 8 weeks. After this period, the animals were passively exposed to cigarette smoke for 60 consecutive days, 3 times a day (4 Marlboro red cigarettes per session, for a total of 12 cigarettes. CK activity was measured in cerebral cortex and hippocampal homogenates. Enzyme activity was inhibited in the cerebral cortex of animals submitted to the inhalation of cigarette smoke. However, exercise prevented this inhibition. In contrast, CK activity remained unchanged in the hippocampus. This inhibition of CK by inhalation of cigarette smoke might be related to the process of cell death. Physical exercise played a preventive role in the inhibition of CK activity caused by exposure to cigarette smoke.

  3. Preventive effects of physical exercise on the inhibition of creatine kinase in the cerebral cortex of mice exposed to cigarette smoke. DOI: 10.5007/1980-0037.2011v13n2p106

    Directory of Open Access Journals (Sweden)

    Daiane Bittencourt Fraga

    2011-02-01

    Full Text Available Recent studies have shown the health benefits of physical exercise, increasing the oxidative response of muscle. However, the effects of exercise on the brain are poorly understood and contradictory. The inhibition of creatine kinase (CK activity has been associated with the pathogenesis of a large number of diseases, especially in the brain. The objective of this study was to determine the preventive effects of physical exercise in the hippocampus and cerebral cortex of mice after chronic cigarette smoke exposure. Eight to 10-week-old male mice (C57BL-6 were divided into four groups and submitted to an exercise program (swimming, 5 times a week, for 8 weeks. After this period, the animals were passively exposed to cigarette smoke for 60 consecutive days, 3 times a day (4 Marlboro red cigarettes per session, for a total of 12 cigarettes. CK activity was measured in cerebral cortex and hippocampal homogenates. Enzyme activity was inhibited in the cerebral cortex of animals submitted to the inhalation of cigarette smoke. However, exercise prevented this inhibition. In contrast, CK activity remained unchanged in the hippocampus. This inhibition of CK by inhalation of cigarette smoke might be related to the process of cell death. Physical exercise played a preventive role in the inhibition of CK activity caused by exposure to cigarette smoke.

  4. The kinase activity of the Ser/Thr kinase BUB1 promotes TGF-β signaling.

    Science.gov (United States)

    Nyati, Shyam; Schinske-Sebolt, Katrina; Pitchiaya, Sethuramasundaram; Chekhovskiy, Katerina; Chator, Areeb; Chaudhry, Nauman; Dosch, Joseph; Van Dort, Marcian E; Varambally, Sooryanarayana; Kumar-Sinha, Chandan; Nyati, Mukesh Kumar; Ray, Dipankar; Walter, Nils G; Yu, Hongtao; Ross, Brian Dale; Rehemtulla, Alnawaz

    2015-01-06

    Transforming growth factor-β (TGF-β) signaling regulates cell proliferation and differentiation, which contributes to development and disease. Upon binding TGF-β, the type I receptor (TGFBRI) binds TGFBRII, leading to the activation of the transcription factors SMAD2 and SMAD3. Using an RNA interference screen of the human kinome and a live-cell reporter for TGFBR activity, we identified the kinase BUB1 (budding uninhibited by benzimidazoles-1) as a key mediator of TGF-β signaling. BUB1 interacted with TGFBRI in the presence of TGF-β and promoted the heterodimerization of TGFBRI and TGFBRII. Additionally, BUB1 interacted with TGFBRII, suggesting the formation of a ternary complex. Knocking down BUB1 prevented the recruitment of SMAD3 to the receptor complex, the phosphorylation of SMAD2 and SMAD3 and their interaction with SMAD4, SMAD-dependent transcription, and TGF-β-mediated changes in cellular phenotype including epithelial-mesenchymal transition (EMT), migration, and invasion. Knockdown of BUB1 also impaired noncanonical TGF-β signaling mediated by the kinases AKT and p38 MAPK (mitogen-activated protein kinase). The ability of BUB1 to promote TGF-β signaling depended on the kinase activity of BUB1. A small-molecule inhibitor of the kinase activity of BUB1 (2OH-BNPP1) and a kinase-deficient mutant of BUB1 suppressed TGF-β signaling and formation of the ternary complex in various normal and cancer cell lines. 2OH-BNPP1 administration to mice bearing lung carcinoma xenografts reduced the amount of phosphorylated SMAD2 in tumor tissue. These findings indicated that BUB1 functions as a kinase in the TGF-β pathway in a role beyond its established function in cell cycle regulation and chromosome cohesion. Copyright © 2015, American Association for the Advancement of Science.

  5. Does maternal-fetal transfer of creatine occur in pregnant sheep?

    Science.gov (United States)

    Baharom, Syed; De Matteo, Robert; Ellery, Stacey; Della Gatta, Paul; Bruce, Clinton R; Kowalski, Greg M; Hale, Nadia; Dickinson, Hayley; Harding, Richard; Walker, David; Snow, Rodney J

    2017-07-01

    Our aim was to determine the disposition of creatine in ovine pregnancy and whether creatine is transferred across the placenta from mother to fetus. Pregnant ewes received either 1 ) a continuous intravenous infusion of creatine monohydrate or saline from 122 to 131 days gestation, with maternal and fetal arterial blood and amniotic fluid samples collected daily for creatine analysis and fetal tissues collected at necropsy at 133 days for analysis of creatine content, or 2 ) a single systemic bolus injection of [ 13 C]creatine monohydrate at 130 days of gestation, with maternal and fetal arterial blood, uterine vein blood, and amniotic fluid samples collected before and for 4 h after injection and analyzed for creatine, creatine isotopic enrichment, and guanidinoacetic acid (GAA; precursor of creatine) concentrations. Presence of the creatine transporter-1 (SLC6A8) and l-arginine:glycine amidinotransferase (AGAT; the enzyme synthesizing GAA) proteins were determined by Western blots of placental cotyledons. The 10-day creatine infusion increased maternal plasma creatine concentration three- to fourfold ( P creatine content. Maternal arterial 13 C enrichment was increased ( P creatine injection without change of fetal arterial 13 C enrichment. SLC6A8 and AGAT proteins were identified in placental cotyledons, and GAA concentration was significantly higher in uterine vein than maternal artery plasma. Despite the presence of SLC6A8 protein in cotyledons, these results suggest that creatine is not transferred from mother to fetus in near-term sheep and that the ovine utero-placental unit releases GAA into the maternal circulation. Copyright © 2017 the American Physiological Society.

  6. Creatine metabolism: detection of creatine and guanidinoacetate in saliva of healthy subjects.

    Science.gov (United States)

    Martínez, Lidia D; Bezard, Miriam; Brunotto, Mabel; Dodelson de Kremer, Raquel

    2016-04-01

    Creatine (Cr) plays an important role in storage and transmission of phosphate-bound energy. Cerebral creatine deficiency syndromes comprise three inherited defects in Cr biosynthesis and transport. The aim of this study was to investigate whether Cr and Guanidinoacetate (GAA) can be detected in saliva of healthy subjects and to establish the relationship between salivary and plasma levels of these molecules. An adapted gas chromatography (GC) method is described for the quantification of Cr and GAA biomarkers in saliva. Reference values were established for GAA and Cr in saliva. These values were age dependent (p= 0.001). No difference between genders was observed. We detected a difference between GAA and Cr concentrations in saliva and in plasma. The GC method for simultaneous determination of GAA and Cr in human saliva is fast, reliable, sensitive, non-invasive and precise to use as a biochemical approach in early detection of cerebral creatine deficiency syndromes. Sociedad Argentina de Investigación Odontológica.

  7. Stress-induced activation of protein kinase CK2 by direct interaction with p38 mitogen-activated protein kinase

    DEFF Research Database (Denmark)

    Sayed, M; Kim, S O; Salh, B S

    2000-01-01

    Protein kinase CK2 has been implicated in the regulation of a wide range of proteins that are important in cell proliferation and differentiation. Here we demonstrate that the stress signaling agents anisomycin, arsenite, and tumor necrosis factor-alpha stimulate the specific enzyme activity of CK2...... in the human cervical carcinoma HeLa cells by up to 8-fold, and this could be blocked by the p38 MAP kinase inhibitor SB203580. We show that p38alpha MAP kinase, in a phosphorylation-dependent manner, can directly interact with the alpha and beta subunits of CK2 to activate the holoenzyme through what appears...

  8. Differences in muscle pain and plasma creatine kinase activity after ...

    African Journals Online (AJOL)

    encephalopathy,18 and the decrement in muscle power associated with muscle damage.6 ... A high degree of intra-individual variability in plasma. CK activity was ..... 21. Komi PV. Stretch-shortening cycle exercise: a powerful model to study.

  9. Profiling bacterial kinase activity using a genetic circuit

    DEFF Research Database (Denmark)

    van der Helm, Eric; Bech, Rasmus; Lehning, Christina Eva

    Phosphorylation is a post-translational modification that regulates the activity of several key proteins in bacteria and eukaryotes. Accordingly, a variety of tools has been developed to measure kinase activity. To couple phosphorylation to an in vivo fluorescent readout we used the Bacillus...... subtilis kinase PtkA, transmembrane activator TkmA and the repressor FatR to construct a genetic circuit in E. coli. By tuning the repressor and kinase expression level at the same time, we were able to show a 4.2-fold increase in signal upon kinase induction. We furthermore validated that the previously...... reported FatR Y45E mutation1 attenuates operator repression. This genetic circuit provides a starting point for computational protein design and a metagenomic library-screening tool....

  10. X-Linked Creatine Transporter Deficiency Presenting as a Mitochondrial Disorder

    NARCIS (Netherlands)

    Hathaway, S.C.; Friez, M.; Limbo, K.; Parker, C.; Salomons, G.S.; Vockley, J.; Wood, T.; Abdul-Rahman, O.A.

    2010-01-01

    X-linked creatine transporter defect is caused by mutations in SLC6A8 at Xq28, which encodes the sodium-dependent creatine transporter. Reduction in creatine uptake results in elevated urine creatine and CSF creatine deficiency, which can be detected on magnetic resonance spectroscopy. We report a

  11. Creatine kinase rate constant in the human heart measured with 3D‐localization at 7 tesla

    Science.gov (United States)

    Robson, Matthew D.; Neubauer, Stefan; Rodgers, Christopher T.

    2016-01-01

    Purpose We present a new Bloch‐Siegert four Angle Saturation Transfer (BOAST) method for measuring the creatine kinase (CK) first‐order effective rate constant kf in human myocardium at 7 tesla (T). BOAST combines a variant of the four‐angle saturation transfer (FAST) method using amplitude‐modulated radiofrequency pulses, phosphorus Bloch‐Siegert B1+‐mapping to determine the per‐voxel flip angles, and nonlinear fitting to Bloch simulations for postprocessing. Methods Optimal flip angles and repetition time parameters were determined from Monte Carlo simulations. BOAST was validated in the calf muscle of two volunteers at 3T and 7T. The myocardial CK forward rate constant was then measured in 10 volunteers at 7T in 82 min (after 1H localization). Results BOAST kfCK values were 0.281 ± 0.002 s−1 in the calf and 0.35 ± 0.05 s−1 in myocardium. These are consistent with literature values from lower fields. Using a literature values for adenosine triphosphate concentration, we computed CK flux values of 4.55 ± 1.52 mmol kg−1 s−1. The sensitive volume for BOAST depends on the B1 inhomogeneity of the transmit coil. Conclusion BOAST enables measurement of the CK rate constant in the human heart at 7T, with spatial localization in three dimensions to 5.6 mL voxels, using a 10‐cm loop coil. Magn Reson Med 78:20–32, 2017. © 2016 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. PMID:27579566

  12. Effects of creatine supplementation on exercise performance.

    Science.gov (United States)

    Demant, T W; Rhodes, E C

    1999-07-01

    While creatine has been known to man since 1835, when a French scientist reported finding this constitutent of meat, its presence in athletics as a performance enhancer is relatively new. Amid claims of increased power and strength, decreased performance time and increased muscle mass, creatine is being hailed as a true ergogenic aid. Creatinine is synthesised from the amino acids glycine, arginine and methionine in the kidneys, liver and pancreas, and is predominantly found in skeletal muscle, where it exists in 2 forms. Approximately 40% is in the free creatine form (Crfree), while the remaining 60% is in the phosphorylated form, creatine phosphate (CP). The daily turnover rate of approximately 2 g per day is equally met via exogenous intake and endogenous synthesis. Although creatine concentration (Cr) is greater in fast twitch muscle fibres, slow twitch fibres have a greater resynthesis capability due to their increased aerobic capacity. There appears to be no significant difference between males and females in Cr, and training does not appear to effect Cr. The 4 roles in which creatine is involved during performance are temporal energy buffering, spatial energy buffering, proton buffering and glycolysis regulation. Creatine supplementation of 20 g per day for at least 3 days has resulted in significant increases in total Cr for some individuals but not others, suggesting that there are 'responders' and 'nonresponders'. These increases in total concentration among responders is greatest in individuals who have the lowest initial total Cr, such as vegetarians. Increased concentrations of both Crfree and CP are believed to aid performance by providing more short term energy, as well as increase the rate of resynthesis during rest intervals. Creatine supplementation does not appear to aid endurance and incremental type exercises, and may even be detrimental. Studies investigating the effects of creatine supplementation on short term, high intensity exercises have

  13. Genetic Depletion of Adipocyte Creatine Metabolism Inhibits Diet-Induced Thermogenesis and Drives Obesity.

    Science.gov (United States)

    Kazak, Lawrence; Chouchani, Edward T; Lu, Gina Z; Jedrychowski, Mark P; Bare, Curtis J; Mina, Amir I; Kumari, Manju; Zhang, Song; Vuckovic, Ivan; Laznik-Bogoslavski, Dina; Dzeja, Petras; Banks, Alexander S; Rosen, Evan D; Spiegelman, Bruce M

    2017-10-03

    Diet-induced thermogenesis is an important homeostatic mechanism that limits weight gain in response to caloric excess and contributes to the relative stability of body weight in most individuals. We previously demonstrated that creatine enhances energy expenditure through stimulation of mitochondrial ATP turnover, but the physiological role and importance of creatine energetics in adipose tissue have not been explored. Here, we have inactivated the first and rate-limiting enzyme of creatine biosynthesis, glycine amidinotransferase (GATM), selectively in fat (Adipo-Gatm KO). Adipo-Gatm KO mice are prone to diet-induced obesity due to the suppression of elevated energy expenditure that occurs in response to high-calorie feeding. This is paralleled by a blunted capacity for β3-adrenergic activation of metabolic rate, which is rescued by dietary creatine supplementation. These results provide strong in vivo genetic support for a role of GATM and creatine metabolism in energy expenditure, diet-induced thermogenesis, and defense against diet-induced obesity. Published by Elsevier Inc.

  14. Effects of Butyltins (BTs) on Mitogen-Activated-Protein Kinase Kinase Kinase (MAP3K) and Ras Activity in Human Natural Killer Cells

    Science.gov (United States)

    Celada, Lindsay J.; Whalen, Margaret M.

    2013-01-01

    Butyltins (BTs) contaminate the environment and are found in human blood. BTs, tributyltin (TBT) and dibutyltin (DBT), diminish the cytotoxic function and levels of key proteins of human natural killer (NK) cells. NK cells are an initial immune defense against tumors, virally-infected cells and antibody-coated cells and thus critical to human health. The signaling pathways that regulate NK cell functions include mitogen-activated protein kinases (MAPKs). Studies have shown that exposure to BTs leads to the activation of specific MAPKs and MAPK kinases (MAP2Ks) in human NK cells. MAP2K kinases (MAP3Ks) are upstream activators of MAP2Ks, which then activate MAPKs. The current study examined if BT-induced activation of MAP3Ks was responsible for MAP2K and thus, MAPK activation. This study examines the effects of TBT and DBT on the total levels of two MAP3Ks, c-Raf and ASK1, as well as activating and inhibitory phosphorylation sites on these MAP3Ks. In addition, the immediate upstream activator of c-Raf, Ras, was examined for BT-induced alterations. Our results show significant activation of the MAP3K, c-Raf, in human NK cells within 10 minutes of TBT exposure and the MAP3K, ASK1, after one hour exposures to TBT. In addition, our results suggest that both TBT and DBT are impacting the regulation of c-Raf. PMID:24038145

  15. Constitutive Activity in an Ancestral Form of Abl Tyrosine Kinase.

    Directory of Open Access Journals (Sweden)

    Saadat U Aleem

    Full Text Available The c-abl proto-oncogene encodes a nonreceptor tyrosine kinase that is found in all metazoans, and is ubiquitously expressed in mammalian tissues. The Abl tyrosine kinase plays important roles in the regulation of mammalian cell physiology. Abl-like kinases have been identified in the genomes of unicellular choanoflagellates, the closest relatives to the Metazoa, and in related unicellular organisms. Here, we have carried out the first characterization of a premetazoan Abl kinase, MbAbl2, from the choanoflagellate Monosiga brevicollis. The enzyme possesses SH3, SH2, and kinase domains in a similar arrangement to its mammalian counterparts, and is an active tyrosine kinase. MbAbl2 lacks the N-terminal myristoylation and cap sequences that are critical regulators of mammalian Abl kinase activity, and we show that MbAbl2 is constitutively active. When expressed in mammalian cells, MbAbl2 strongly phosphorylates cellular proteins on tyrosine, and transforms cells much more potently than mammalian Abl kinase. Thus, MbAbl2 appears to lack the autoinhibitory mechanism that tightly constrains the activity of mammalian Abl kinases, suggesting that this regulatory apparatus arose more recently in metazoan evolution.

  16. Creatine monohydrate supplementation does not increase muscle strength, lean body mass, or muscle phosphocreatine in patients with myotonic dystrophy type 1.

    Science.gov (United States)

    Tarnopolsky, Mark; Mahoney, Douglas; Thompson, Terry; Naylor, Heather; Doherty, Timothy J

    2004-01-01

    Creatine monohydrate (CrM) supplementation may increase strength in some types of muscular dystrophy. A recent study in myotonic muscular dystrophy type 1 (DM1) did not find a significant treatment effect, but measurements of muscle phosphocreatine (PCr) were not performed. We completed a randomized, double-blind, cross-over trial using 34 genetically confirmed adult DM1 patients without significant cognitive impairment. Participants received CrM (5 g, approximately 0.074 g/kg daily) and a placebo for each 4-month phase with a 6-week wash-out. Spirometry, manual muscle testing, quantitative isometric strength testing of handgrip, foot dorsiflexion, and knee extension, handgrip and foot dorsiflexion endurance, functional tasks, activity of daily living scales, body composition (total, bone, and fat-free mass), serum creatine kinase activity, serum creatinine concentration and clearance, and liver function tests were completed before and after each intervention, and muscle PCr/beta-adenosine triphosphate (ATP) ratios of the forearm flexor muscles were completed at the end of each phase. CrM supplementation did not increase any of the outcome measurements except for plasma creatinine concentration (but not creatinine clearance). Thus, CrM supplementation at 5 g daily does not have any effects on muscle strength, body composition, or activities of daily living in patients with DM1, perhaps because of a failure of the supplementation to increase muscle PCr/beta-ATP content.

  17. Creatine and the Liver: Metabolism and Possible Interactions.

    Science.gov (United States)

    Barcelos, R P; Stefanello, S T; Mauriz, J L; Gonzalez-Gallego, J; Soares, F A A

    2016-01-01

    The process of creatine synthesis occurs in two steps, catalyzed by L-arginine:glycine amidinotransferase (AGAT) and guanidinoacetate N-methyltransferase (GAMT), which take place mainly in kidney and liver, respectively. This molecule plays an important energy/pH buffer function in tissues, and to guarantee the maintenance of its total body pool, the lost creatine must be replaced from diet or de novo synthesis. Creatine administration is known to decrease the consumption of Sadenosyl methionine and also reduce the homocysteine production in liver, diminishing fat accumulation and resulting in beneficial effects in fatty liver and non-alcoholic liver disease. Different studies have shown that creatine supplementation could supply brain energy, presenting neuroprotective effects against the encephalopathy induced by hyperammonemia in acute liver failure. Creatine is also taken by many athletes for its ergogenic properties. However, little is known about the adverse effects of creatine supplementation, which are barely described in the literature, with reports of mainly hypothetical effects arising from a small number of scientific publications. Antioxidant effects have been found in several studies, although one of the theories regarding the potential for toxicity from creatine supplementation is that it can increase oxidative stress and potentially form carcinogenic compounds.

  18. LmxMPK4, an essential mitogen-activated protein kinase of Leishmania mexicana is phosphorylated and activated by the STE7-like protein kinase LmxMKK5

    DEFF Research Database (Denmark)

    John von Freyend, Simona; Rosenqvist, Heidi; Fink, Annette

    2010-01-01

    The essential mitogen-activated protein kinase (MAP kinase), LmxMPK4, of Leishmania mexicana is minimally active when purified following recombinant expression in Escherichia coli and was therefore unsuitable for drug screening until now. Using an E. coli protein co-expression system we identified...... LmxMKK5, a STE7-like protein kinase from L. mexicana, which phosphorylates and activates recombinant LmxMPK4 in vitro. LmxMKK5 is comprised of 525 amino acids and has a calculated molecular mass of 55.9kDa. The co-expressed, purified LmxMPK4 showed strong phosphotransferase activity in radiometric...... kinase assays and was confirmed by immunoblot and tandem mass spectrometry analyses to be phosphorylated on threonine 190 and tyrosine 192 of the typical TXY MAP kinase activation motif. The universal protein kinase inhibitor staurosporine reduced the phosphotransferase activity of co...

  19. Chronic high-dose creatine has opposing effects on depression-related gene expression and behavior in intact and sex hormone-treated gonadectomized male and female rats.

    Science.gov (United States)

    Allen, Patricia J; DeBold, Joseph F; Rios, Maribel; Kanarek, Robin B

    2015-03-01

    Creatine is an antioxidant, neuromodulator and key regulator of energy metabolism shown to improve depressive symptoms in humans and animals, especially in females. To better understand the pharmacological effects of creatine, we examined its influence on depression-related hippocampal gene expression and behaviors in the presence and absence of sex steroids. Sham-operated and gonadectomized male and female rats were fed chow alone or chow blended with either 2% or 4% w/w creatine monohydrate for five weeks before forced swim, open field, and wire suspension tests, or seven weeks total. Before supplementation, males were chronically implanted with an empty or a testosterone-filled (T) capsule (10-mm surface release), and females were administered progesterone (P, 250 μg), estradiol benzoate (EB, 2.5 μg), EB+P, or sesame oil vehicle weekly. Relative to non-supplemented shams, all hippocampal plasticity-related mRNAs measured, including brain-derived neurotrophic factor (BDNF), tyrosine kinase B, doublecortin, calretinin, and calbindin, were downregulated in sham males given 4% creatine, and BDNF, doublecortin, and calbindin mRNAs were downregulated in sham females given 4% creatine. In contrast, combined 4% creatine+T in castrates prevented downregulation of BDNF, doublecortin, and calretinin mRNAs. Similarly, combined 4% creatine+EB+P in ovariectomized females attenuated downregulation of BDNF and calbindin mRNA levels. Moderate antidepressant and anxiolytic-like behaviors were observed in EB+P-treated ovariectomized females fed creatine, with similar trends in T-treated castrates fed creatine. Altogether, these data show that chronic, high-dose creatine has opposing effects on neuroplasticity-related genes and depressive behavior in intact and gonadectomized male and female rats. The dose and schedule of creatine used negatively impacted hippocampal neuronal integrity in otherwise healthy brains, possibly through negative compensatory changes in energy

  20. Scientific basis and practical aspects of creatine supplementation for athletes.

    Science.gov (United States)

    Volek, Jeff S; Rawson, Eric S

    2004-01-01

    A large number of studies have been published on creatine supplementation over the last decade. Many studies show that creatine supplementation in conjunction with resistance training augments gains in muscle strength and size. The underlying physiological mechanism(s) to explain this ergogenic effect remain unclear. Increases in muscle fiber hypertrophy and myosin heavy chain expression have been observed with creatine supplementation. Creatine supplementation increases acute weightlifting performance and training volume, which may allow for greater overload and adaptations to training. Creatine supplementation may also induce a cellular swelling in muscle cells, which in turn may affect carbohydrate and protein metabolism. Several studies point to the conclusion that elevated intramuscular creatine can enhance glycogen levels but an effect on protein synthesis/degradation has not been consistently detected. As expected there is a distribution of responses to creatine supplementation that can be largely explained by the degree of creatine uptake into muscle. Thus, there is wide interest in methods to maximize muscle creatine levels. A carbohydrate or carbohydrate/protein-induced insulin response appears to benefit creatine uptake. In summary, the predominance of research indicates that creatine supplementation represents a safe, effective, and legal method to enhance muscle size and strength responses to resistance training.

  1. URINARY CREATINE AT REST AND AFTER REPEATED SPRINTS IN ATHLETES: A PILOT STUDY

    Directory of Open Access Journals (Sweden)

    I. Bezrati-Benayed

    2014-07-01

    Full Text Available Creatine plays a key role in muscle function and its evaluation is important in athletes. In this study, urinary creatine concentration was measured in order to highlight its possible significance in monitoring sprinters. The study included 51 sprinters and 25 age- and sex-matched untrained subjects as a control group. Body composition was measured and dietary intake estimated. Urine samples were collected before and after standardized physical exercise. Creatine was assessed by gas chromatography mass spectrometry. Basal urinary creatine (UC was significantly lower in sprinters than controls (34±30 vs. 74±3 μmol/mmol creatinine, p<0.05. UC was inversely correlated with body mass (r=-0.34, p<0.01 and lean mass (r=- 0.30, p<0.05, and positively correlated with fat mass (r=0.32, p<0.05. After acute exercise, urinary creatine significantly decreased in both athletes and controls. UC is low in sprinters at rest and further decreases after exercise, most likely due to a high uptake and use of creatine by muscles, as muscle mass and physical activity are supposed to be greater in athletes than untrained subjects. Further studies are needed to test the value of urinary creatine as a non-invasive marker of physical condition and as a parameter for managing Cr supplementation in athletes.

  2. Synapses of Amphids Defective (SAD-A) Kinase Promotes Glucose-stimulated Insulin Secretion through Activation of p21-activated Kinase (PAK1) in Pancreatic β-Cells*

    Science.gov (United States)

    Nie, Jia; Sun, Chao; Faruque, Omar; Ye, Guangming; Li, Jia; Liang, Qiangrong; Chang, Zhijie; Yang, Wannian; Han, Xiao; Shi, Yuguang

    2012-01-01

    The p21-activated kinase-1 (PAK1) is implicated in regulation of insulin exocytosis as an effector of Rho GTPases. PAK1 is activated by the onset of glucose-stimulated insulin secretion (GSIS) through phosphorylation of Thr-423, a major activation site by Cdc42 and Rac1. However, the kinase(s) that phosphorylates PAK1 at Thr-423 in islet β-cells remains elusive. The present studies identified SAD-A (synapses of amphids defective), a member of AMP-activated protein kinase-related kinases exclusively expressed in brain and pancreas, as a key regulator of GSIS through activation of PAK1. We show that SAD-A directly binds to PAK1 through its kinase domain. The interaction is mediated by the p21-binding domain (PBD) of PAK1 and requires both kinases in an active conformation. The binding leads to direct phosphorylation of PAK1 at Thr-423 by SAD-A, triggering the onset of GSIS from islet β-cells. Consequently, ablation of PAK1 kinase activity or depletion of PAK1 expression completely abolishes the potentiating effect of SAD-A on GSIS. Consistent with its role in regulating GSIS, overexpression of SAD-A in MIN6 islet β-cells significantly stimulated cytoskeletal remodeling, which is required for insulin exocytosis. Together, the present studies identified a critical role of SAD-A in the activation of PAK1 during the onset of insulin exocytosis. PMID:22669945

  3. Synapses of amphids defective (SAD-A) kinase promotes glucose-stimulated insulin secretion through activation of p21-activated kinase (PAK1) in pancreatic β-Cells.

    Science.gov (United States)

    Nie, Jia; Sun, Chao; Faruque, Omar; Ye, Guangming; Li, Jia; Liang, Qiangrong; Chang, Zhijie; Yang, Wannian; Han, Xiao; Shi, Yuguang

    2012-07-27

    The p21-activated kinase-1 (PAK1) is implicated in regulation of insulin exocytosis as an effector of Rho GTPases. PAK1 is activated by the onset of glucose-stimulated insulin secretion (GSIS) through phosphorylation of Thr-423, a major activation site by Cdc42 and Rac1. However, the kinase(s) that phosphorylates PAK1 at Thr-423 in islet β-cells remains elusive. The present studies identified SAD-A (synapses of amphids defective), a member of AMP-activated protein kinase-related kinases exclusively expressed in brain and pancreas, as a key regulator of GSIS through activation of PAK1. We show that SAD-A directly binds to PAK1 through its kinase domain. The interaction is mediated by the p21-binding domain (PBD) of PAK1 and requires both kinases in an active conformation. The binding leads to direct phosphorylation of PAK1 at Thr-423 by SAD-A, triggering the onset of GSIS from islet β-cells. Consequently, ablation of PAK1 kinase activity or depletion of PAK1 expression completely abolishes the potentiating effect of SAD-A on GSIS. Consistent with its role in regulating GSIS, overexpression of SAD-A in MIN6 islet β-cells significantly stimulated cytoskeletal remodeling, which is required for insulin exocytosis. Together, the present studies identified a critical role of SAD-A in the activation of PAK1 during the onset of insulin exocytosis.

  4. Effects of in ovo feeding of creatine pyruvate on the hatchability, growth performance and energy status in embryos and broiler chickens.

    Science.gov (United States)

    Zhao, M M; Gao, T; Zhang, L; Li, J L; Lv, P A; Yu, L L; Gao, F; Zhou, G H

    2017-10-01

    The effects of in ovo feeding (IOF) of creatine pyruvate (CrPyr) on the hatchability, growth performance and energy status of embryos and broilers (Arbor Acres) were investigated. Five treatments were arranged as non-injected treatment (Control), 0.6 ml physiological saline (0.75%) injected treatment (Saline), and IOF treatments injected with 0.6 ml physiological saline (0.75%) containing 3, 6 or 12 mg CrPyr (CrPyr3, CrPyr6 or CrPyr12) into the amnion per fertile egg on day 17.5 of incubation. After hatching, 80 male chicks from each treatment with similar weight close to the average BW of their pooled group were selected and randomly assigned into eight replicates of 10 chicks each. The results showed that the hatchability was not affected among groups, whereas the hatching weight of broilers in CrPyr12 was significantly higher than the control and saline groups (P0.05). Irrespective of dosage, the concentrations of creatine and phosphocreatine, and activities of creatine kinase in embryos were enhanced in CrPyr treatments at 19 E when compared with the control and saline groups (P<0.05). The activities of glucose-6-phosphatase in liver in CrPyr6 and CrPyr12 treatments were higher than the control and saline groups at 19 E (P<0.05). In conclusion, these results indicated that IOF of CrPyr, especially at the level of 12 mg/egg, could improve energy status of embryos and hatchlings, which was useful for enhancing hatching weight, BW and pectoral muscle weight until the end of the experiments at 21 days post-hatch in broilers.

  5. Creatine and Caffeine: Considerations for Concurrent Supplementation.

    Science.gov (United States)

    Trexler, Eric T; Smith-Ryan, Abbie E

    2015-12-01

    Nutritional supplementation is a common practice among athletes, with creatine and caffeine among the most commonly used ergogenic aids. Hundreds of studies have investigated the ergogenic potential of creatine supplementation, with consistent improvements in strength and power reported for exercise bouts of short duration (≤ 30 s) and high intensity. Caffeine has been shown to improve endurance exercise performance, but results are mixed in the context of strength and sprint performance. Further, there is conflicting evidence from studies comparing the ergogenic effects of coffee and caffeine anhydrous supplementation. Previous research has identified independent mechanisms by which creatine and caffeine may improve strength and sprint performance, leading to the formulation of multi-ingredient supplements containing both ingredients. Although scarce, research has suggested that caffeine ingestion may blunt the ergogenic effect of creatine. While a pharmacokinetic interaction is unlikely, authors have suggested that this effect may be explained by opposing effects on muscle relaxation time or gastrointestinal side effects from simultaneous consumption. The current review aims to evaluate the ergogenic potential of creatine and caffeine in the context of high-intensity exercise. Research directly comparing coffee and caffeine anhydrous is discussed, along with previous studies evaluating the concurrent supplementation of creatine and caffeine.

  6. Protein Kinase Mitogen-activated Protein Kinase Kinase Kinase Kinase 4 (MAP4K4) Promotes Obesity-induced Hyperinsulinemia.

    Science.gov (United States)

    Roth Flach, Rachel J; Danai, Laura V; DiStefano, Marina T; Kelly, Mark; Menendez, Lorena Garcia; Jurczyk, Agata; Sharma, Rohit B; Jung, Dae Young; Kim, Jong Hun; Kim, Jason K; Bortell, Rita; Alonso, Laura C; Czech, Michael P

    2016-07-29

    Previous studies revealed a paradox whereby mitogen-activated protein kinase kinase kinase kinase 4 (Map4k4) acted as a negative regulator of insulin sensitivity in chronically obese mice, yet systemic deletion of Map4k4 did not improve glucose tolerance. Here, we report markedly reduced glucose-responsive plasma insulin and C-peptide levels in whole body Map4k4-depleted mice (M4K4 iKO) as well as an impaired first phase of insulin secretion from islets derived from M4K4 iKO mice ex vivo After long-term high fat diet (HFD), M4K4 iKO mice pancreata also displayed reduced β cell mass, fewer proliferating β cells and reduced islet-specific gene mRNA expression compared with controls, although insulin content was normal. Interestingly, the reduced plasma insulin in M4K4 iKO mice exposed to chronic (16 weeks) HFD was not observed in response to acute HFD challenge or short term treatment with the insulin receptor antagonist S961. Furthermore, the improved insulin sensitivity in obese M4K4 iKO mice was abrogated by high exogenous insulin over the course of a euglycemic clamp study, indicating that hypoinsulinemia promotes insulin sensitivity in chronically obese M4K4 iKO mice. These results demonstrate that protein kinase Map4k4 drives obesity-induced hyperinsulinemia and insulin resistance in part by promoting insulin secretion from β cells in mice. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Side effects of creatine supplementation in athletes.

    OpenAIRE

    Francaux, Marc; Poortmans, Jacques R

    2006-01-01

    Context: Allegations about side effects of creatine supplementation by athletes have been published in the popular media and scientific publications. Purpose: To examine the experimental evidence relating to the physiological effects of creatine supplementation. Results: One of the purported effects of oral creatine supplementation is increased muscle mass. A review of the literature reveals a 1.0% to 2.3% increase in body mass, which is attributed to fat-free mass and, more specifically, to ...

  8. Is there a rationale for the use of creatine either as nutritional supplementation or drug administration in humans participating in a sport?

    Science.gov (United States)

    Benzi, G

    2000-03-01

    Even though no unambiguous proof for enhanced performance during high-intensity exercise has yet been reported, the creatine administration is charged to improve physical performance and has become a popular practice among subjects participating in different sports. Appropriate creatine dosage may be also used as a medicinal product since, in accordance with the Council Directive 65/65/CEE, any substance which may be administered with a view to restoring, correcting or modifying physiological functions in human beings is considered a medicinal product. Thus, quality, efficacy and safety must characterize the substance. In biochemical terms, creatine administration enhances both creatine and phosphocreatine concentrations, allowing for an increased total creatine pool in skeletal muscle. In thermodynamics terms, creatine interferes with the creatine-creatine kinase-phosphocreatine circuit, which is related to the mitochondrial function as a highly organized system for the energy control of the subcellular adenylate pool. In pharmacokinetics terms, creatine entry into skeletal muscle is initially dependent on the extracellular concentration, but the creatine transport is subsequently down-regulated. In pharmacodynamics terms, the creatine enhances the possibility to maintain power output during brief periods of high-intensity exercises. In spite of uncontrolled daily dosage and long-term administration, no research on creatine safety in humans has been set up by specific standard protocol of clinical pharmacology and toxicology, as currently occurs in phase I for the products for human use. More or less documented side effects induced by creatine are weight gain; influence on insulin production; feedback inhibition of endogenous creatine synthesis; long-term damages on renal function. A major point that related to the quality of creatine products is the amount of creatine ingested in relation to the amount of contaminants present. During the production of creatine

  9. A comparison of mutagen production in fried ground chicken and beef: effect of supplemental creatine.

    Science.gov (United States)

    Knize, M G; Shen, N H; Felton, J S

    1988-11-01

    Ground chicken breast and ground beef with either endogenous or a 10-fold increase in the concentration of creatine were fried at 220 degrees C for 10 min per side. One patty (100 g) of chicken meat yielded 120,000 Salmonella (TA1538) revertants following metabolic activation. The pan residues had 39% of the total activity. Added creatine (10-fold the endogenous level) increased mutagen yields an average of 2-fold. Beef cooked under identical conditions yielded 150,000 revertants/100 g for the meat patties and pan residues combined. Added creatine to beef prior to cooking increased mutagen yields 3-fold. The mutagenic profiles following initial HPLC separation showed that chicken samples with endogenous or added creatine were remarkably similar. Chicken and beef HPLC mutagenicity profiles were also similar to each other, but not identical. This suggests that the general mutagen-forming reactions with the two different types of muscle are qualitatively similar with only minor quantitative differences. The pan residues from both meat types with and without added creatine showed some significant differences in the mutagen peak profile. This work suggests that the types of mutagens formed in chicken are similar to those formed in beef and that creatine appears to be involved in the formation of all the mutagenic compounds produced from fried muscle tissue.

  10. Intramolecular Crosstalk between Catalytic Activities of Receptor Kinases

    KAUST Repository

    Kwezi, Lusisizwe

    2018-01-22

    Signal modulation is important for the growth and development of plants and this process is mediated by a number of factors including physiological growth regulators and their associated signal transduction pathways. Protein kinases play a central role in signaling, including those involving pathogen response mechanisms. We previously demonstrated an active guanylate cyclase (GC) catalytic center in the brassinosteroid insensitive receptor (AtBRI1) within an active intracellular kinase domain resulting in dual enzymatic activity. Here we propose a novel type of receptor architecture that is characterized by a functional GC catalytic center nested in the cytosolic kinase domain enabling intramolecular crosstalk. This may be through a cGMP-AtBRI1 complex forming that may induce a negative feedback mechanism leading to desensitisation of the receptor, regulated through the cGMP production pathway. We further argue that the comparatively low but highly localized cGMP generated by the GC in response to a ligand is sufficient to modulate the kinase activity. This type of receptor therefore provides a molecular switch that directly and/or indirectly affects ligand dependent phosphorylation of downstream signaling cascades and suggests that subsequent signal transduction and modulation works in conjunction with the kinase in downstream signaling.

  11. Intramolecular Crosstalk between Catalytic Activities of Receptor Kinases

    KAUST Repository

    Kwezi, Lusisizwe; Wheeler, Janet I; Marondedze, Claudius; Gehring, Christoph A; Irving, Helen R

    2018-01-01

    Signal modulation is important for the growth and development of plants and this process is mediated by a number of factors including physiological growth regulators and their associated signal transduction pathways. Protein kinases play a central role in signaling, including those involving pathogen response mechanisms. We previously demonstrated an active guanylate cyclase (GC) catalytic center in the brassinosteroid insensitive receptor (AtBRI1) within an active intracellular kinase domain resulting in dual enzymatic activity. Here we propose a novel type of receptor architecture that is characterized by a functional GC catalytic center nested in the cytosolic kinase domain enabling intramolecular crosstalk. This may be through a cGMP-AtBRI1 complex forming that may induce a negative feedback mechanism leading to desensitisation of the receptor, regulated through the cGMP production pathway. We further argue that the comparatively low but highly localized cGMP generated by the GC in response to a ligand is sufficient to modulate the kinase activity. This type of receptor therefore provides a molecular switch that directly and/or indirectly affects ligand dependent phosphorylation of downstream signaling cascades and suggests that subsequent signal transduction and modulation works in conjunction with the kinase in downstream signaling.

  12. [Laboratory diagnostics in transient loss of consciousness : Serum lactate compared to serum creatine kinase as diagnostic indicator for generalized tonic-clonic seizures].

    Science.gov (United States)

    Dafotakis, M; Heckelmann, J; Zechbauer, S; Litmathe, J; Brokmann, J; Willmes, K; Surges, R; Matz, O

    2018-03-21

    Laboratory parameters can help in the differential diagnostics of acute episodes of transient loss of consciousness. Especially serum lactate and serum creatine kinase (CK) levels may provide valuable hints to distinguish generalized tonic-clonic seizures (GTCS) from syncope. Serum lactate levels at admission and CK levels 10-48 h after the episodes that led to admission were compared between patients with GTCS (n = 30) and those with syncope (n = 15). In addition, sensitivity and specificity of lactate and CK as diagnostic markers for syncope and GTCS were determined. The serum lactate and serum CK levels were significantly increased in patients with GTCS as compared to syncope patients (serum lactate: p lactate as an indicator for GTCS was 0.94 (95% confidence interval [CI] 0.88-1.0). For CK the receiver operating characteristics (ROC) analysis produced an AUC of only 0.77 (95% CI: 0.63-0.9). The determination of the lactate value as point-of-care diagnostics appears to be highly relevant in the rapid clarification of unclear episodes with transient loss of consciousness. The CK level at follow-up is also suitable for distinguishing GTCS from syncope but is inferior to the serum lactate value.

  13. Interaction of human biliverdin reductase with Akt/protein kinase B and phosphatidylinositol-dependent kinase 1 regulates glycogen synthase kinase 3 activity: a novel mechanism of Akt activation

    OpenAIRE

    Miralem, Tihomir; Lerner-Marmarosh, Nicole; Gibbs, Peter E. M.; Jenkins, Jermaine L.; Heimiller, Chelsea; Maines, Mahin D.

    2016-01-01

    Biliverdin reductase A (BVR) and Akt isozymes have overlapping pleiotropic functions in the insulin/PI3K/MAPK pathway. Human BVR (hBVR) also reduces the hemeoxygenase activity product biliverdin to bilirubin and is directly activated by insulin receptor kinase (IRK). Akt isoenzymes (Akt1–3) are downstream of IRK and are activated by phosphatidylinositol-dependent kinase 1 (PDK1) phosphorylating T308 before S473 autophosphorylation. Akt (RxRxxSF) and PDK1 (RFxFPxFS) binding motifs are present ...

  14. Molecular Imaging of the ATM Kinase Activity

    Energy Technology Data Exchange (ETDEWEB)

    Williams, Terence M. [Department of Radiation Oncology, Ohio State University, Columbus, Ohio (United States); Nyati, Shyam [Department of Radiation Oncology, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Center for Molecular Imaging, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Ross, Brian D. [Department of Radiation Oncology, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Department of Radiology, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Rehemtulla, Alnawaz, E-mail: alnawaz@umich.edu [Department of Radiation Oncology, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Center for Molecular Imaging, University of Michigan Medical Center, Ann Arbor, Michigan (United States); Department of Radiology, University of Michigan Medical Center, Ann Arbor, Michigan (United States)

    2013-08-01

    Purpose: Ataxia telangiectasia mutated (ATM) is a serine/threonine kinase critical to the cellular DNA-damage response, including from DNA double-strand breaks. ATM activation results in the initiation of a complex cascade of events including DNA damage repair, cell cycle checkpoint control, and survival. We sought to create a bioluminescent reporter that dynamically and noninvasively measures ATM kinase activity in living cells and subjects. Methods and Materials: Using the split luciferase technology, we constructed a hybrid cDNA, ATM-reporter (ATMR), coding for a protein that quantitatively reports on changes in ATM kinase activity through changes in bioluminescence. Results: Treatment of ATMR-expressing cells with ATM inhibitors resulted in a dose-dependent increase in bioluminescence activity. In contrast, induction of ATM kinase activity upon irradiation resulted in a decrease in reporter activity that correlated with ATM and Chk2 activation by immunoblotting in a time-dependent fashion. Nuclear targeting improved ATMR sensitivity to both ATM inhibitors and radiation, whereas a mutant ATMR (lacking the target phosphorylation site) displayed a muted response. Treatment with ATM inhibitors and small interfering (si)RNA-targeted knockdown of ATM confirm the specificity of the reporter. Using reporter expressing xenografted tumors demonstrated the ability of ATMR to report in ATM activity in mouse models that correlated in a time-dependent fashion with changes in Chk2 activity. Conclusions: We describe the development and validation of a novel, specific, noninvasive bioluminescent reporter that enables monitoring of ATM activity in real time, in vitro and in vivo. Potential applications of this reporter include the identification and development of novel ATM inhibitors or ATM-interacting partners through high-throughput screens and in vivo pharmacokinetic/pharmacodynamic studies of ATM inhibitors in preclinical models.

  15. Creatine and guanidinoacetate reference values in a French population

    NARCIS (Netherlands)

    Joncquel-Cheval Curt, M.; Cheillan, D.; Briand, G.; Salomons, G.S.; Mention-Mulliez, K.; Dobbelaere, D.; Cuisset, J.M.; Lion-Francois, L.; Portes, V.D.; Chabli, A.; Valayannopoulos, V.; Benoist, J.F.; Pinard, J.M.; Simard, G.; Douay, O.; Deiva, K.; Tardieu, M.; Afenjar, A.; Heron, D.; Rivier, F.; Chabrol, B.; Prieur, F.; Cartault, F.; Pitelet, G.; Goldenberg, A.; Bekri, S.; Gerard, M.; Delorme, R.; Porchet, N.; Vianey-Saban, C.; Vamecq, J.

    2013-01-01

    Creatine and guanidinoacetate are biomarkers of creatine metabolism. Their assays in body fluids may be used for detecting patients with primary creatine deficiency disorders (PCDD), a class of inherited diseases. Their laboratory values in blood and urine may vary with age, requiring that reference

  16. Clinical usefulness of creatine Kinase BB determination by a Ria method in serum of patients with cerebrovascular accidents

    Energy Technology Data Exchange (ETDEWEB)

    Nuti, A; Giraldi, C; Piccini, P; Bonucelli, U; Clerico, A; Del Chicca, M G

    1988-01-01

    The measurement of creatine kinase BB isoenzyme (CK-BB) using RIA methods could have diagnostic utility as a biological marker of cerebral damage. The aim of the present study is to evaluate whether frequent sampling (4 samples/day for 3 days) permits a better correlation between serum CK/sub B/B values and the clinical outcomes of patients with cerebrovascular accidents. 16 in-patients (12 men and 4 women) with stroke (15 of an ischemic nature and 1 hemorrhagic) have benn studied. The presence of stroke was confirmed by clinical symptoms and by CAT results. Blood samples were drawn at 6 and 12 a.m. and at 6 and 12 p.m. over 3 consecutive days after hospitalization. Values of serum CK-BB above the normal range (>7 ng/ml) were found in the 5 of the 16 (31.3%) patients studied. The mean CK-BB value observed in the patients' group was significantly higher than that found in a group of 112 control subjects (controls, mean+-SD=2.1+-1.7 ng/ml vs patients 3.3+-3.4 ng/ml, unpaired t-test p<0.025). We observed a very wide range of serum CKBB levels in most of the patients studied. Some prominent peaks of CK-BB concetrations were found in patients' outcame (Spearman correlation coefficient r-s=0.618, p<0,01). Although our results indicate that the measurement of CK-BB concentrations cannot be considered a sensitive marker of stroke, the significant correlation between serum CK-BB values and outcome suggests that high CK-BB levels could be a sign of worse prognosis in patients with cerebrovascular accidents. 24 refs.

  17. Clinical usefulness of creatine Kinase BB determination by a Ria method in serum of patients with cerebrovascular accidents

    International Nuclear Information System (INIS)

    Nuti, A.; Giraldi, C.; Piccini, P.; Bonucelli, U.; Clerico, A.; Del Chicca, M. G.

    1988-01-01

    The measurement of creatine kinase BB isoenzyme (CK-BB) using RIA methods could have diagnostic utility as a biological marker of cerebral damage. The aim of the present study is to evaluate whether frequent sampling (4 samples/day for 3 days) permits a better correlation between serum CK B B values and the clinical outcomes of patients with cerebrovascular accidents. 16 in-patients (12 men and 4 women) with stroke (15 of an ischemic nature and 1 hemorrhagic) have benn studied. The presence of stroke was confirmed by clinical symptoms and by CAT results. Blood samples were drawn at 6 and 12 a.m. and at 6 and 12 p.m. over 3 consecutive days after hospitalization. Values of serum CK-BB above the normal range (>7 ng/ml) were found in the 5 of the 16 (31.3%) patients studied. The mean CK-BB value observed in the patients' group was significantly higher than that found in a group of 112 control subjects (controls, mean+-SD=2.1+-1.7 ng/ml vs patients 3.3+-3.4 ng/ml, unpaired t-test p<0.025). We observed a very wide range of serum CKBB levels in most of the patients studied. Some prominent peaks of CK-BB concetrations were found in patients' outcame (Spearman correlation coefficient r-s=0.618, p<0,01). Although our results indicate that the measurement of CK-BB concentrations cannot be considered a sensitive marker of stroke, the significant correlation between serum CK-BB values and outcome suggests that high CK-BB levels could be a sign of worse prognosis in patients with cerebrovascular accidents

  18. [Effect of Low-Intensity 900 MHz Frequency Electromagnetic Radiation on Rat Brain Enzyme Activities Linked to Energy Metabolism].

    Science.gov (United States)

    Petrosyan, M S; Nersesova, L S; Gazaryants, M G; Meliksetyan, G O; Malakyan, M G; Bajinyan, S A; Akopian, J I

    2015-01-01

    The research deals with the effect of low-intensity 900 MHz frequency electromagnetic radiation (EMR), power density 25 μW/cm2, on the following rat brain and blood serum enzyme activities: creatine kinase (CK), playing a central role in the process of storing and distributing the cell energy, as well as alanine aminotransferase (ALT) and aspartate aminotransferase (AST) that play a key role in providing the conjunction of carbohydrate and amino acid metabolism. The comparative analysis of the changes in the enzyme activity studied at different times following the two-hour single, as well as fractional, radiation equivalent of the total time showed that the most radiosensitive enzyme is the brain creatine kinase, which may then be recommended as a marker of the radio frequency radiation impact. According to the analysis of the changing dynamics of the CK, ALT and AST activity level, with time these changes acquire the adaptive character and are directed to compensate the damaged cell energy metabolism.

  19. Elevation of serum creatine kinase during methimazole treatment of Graves disease in a 13-year-old girl and a literature review of similar cases

    Directory of Open Access Journals (Sweden)

    Hyeseon Kim

    2015-06-01

    Full Text Available We report a 13-year-old girl with Graves disease, who showed an increased level of serum creatine kinase (CK accompanied by myalgia after methimazole (MMI treatment. This patient developed muscular pain two weeks after MMI administration, along with increased CK levels. The level of thyroid hormone was within the normal range when she showed increased CK levels. After the MMI dose was decreased and levo-thyroxine was added, serum CK levels decreased to normal and the myalgia improved. The pathophysiologic mechanism of this effect has not yet been elucidated. An acute relatively hypothyroid state occurs secondary to antithyroid drug (ATD administration in chronic hyperthyroidism, which may cause changes in the CK levels. In this report, we present a rare pediatric case, along with a literature review of similar cases. In the initial state of MMI treatment, myalgia should be detected and when it occurs, CK levels should be measured. The clinical strategy of monitoring CK levels with the aim of normalizing thyroid hormones is helpful in case of the development of adverse reactions, such as myalgia, during ATD treatment for Graves disease in children.

  20. Visual Snapshots of Intracellular Kinase Activity At The Onset of Mitosis

    Science.gov (United States)

    Dai, Zhaohua; Dulyaninova, Natalya G.; Kumar, Sanjai; Bresnick, Anne R.; Lawrence, David S.

    2007-01-01

    Summary Visual snapshots of intracellular kinase activity can be acquired with exquisite temporal control using a light-activatable (caged) sensor, thereby providing a means to interrogate enzymatic activity at any point during the cell division cycle. Robust protein kinase activity transpires just prior to, but not immediately following, nuclear envelope breakdown (NEB). Furthermore, kinase activity is required for progression from prophase into metaphase. Finally, the application of selective protein kinase C (PKC) inhibitors, in combination with the caged sensor, correlates the action of the PKC β isoform with subsequent NEB. PMID:18022564

  1. A Requirement for ZAK Kinase Activity in Canonical TGF-β Signaling

    Directory of Open Access Journals (Sweden)

    Shyam Nyati

    2016-12-01

    Full Text Available The sterile alpha motif and leucine zipper containing kinase ZAK (AZK, MLT, MLK7, is a MAPK-kinase kinase (MKKK. Like most MAPKKKs which are known to activate the c-Jun. amino-terminal kinase (JNK pathway, ZAK has been shown to participate in the transduction of Transforming growth factor-β (TGF-β-mediated non-canonical signaling. A role for ZAK in SMAD-dependent, canonical TGF-β signaling has not been previously appreciated. Using a combination of functional genomics and biochemical techniques, we demonstrate that ZAK regulates canonical TGFβRI/II signaling in lung and breast cancer cell lines and may serve as a key node in the regulation of TGFBR kinase activity. Remarkably, we demonstrate that siRNA mediated depletion of ZAK strongly inhibited TGF-β dependent SMAD2/3 activation and subsequent promoter activation (SMAD binding element driven luciferase expression; SBE4-Luc. A ZAK specific inhibitor (DHP-2, dose-dependently activated the bioluminescent TGFBR-kinase activity reporter (BTR, blocked TGF-β induced SMAD2/3 phosphorylation and SBE4-Luc activation and cancer cell-invasion. In aggregate, these findings identify a novel role for the ZAK kinase in canonical TGF-β signaling and an invasive cancer cell phenotype thus providing a novel target for TGF-β inhibition.

  2. Creatine supplementation and glycemic control: a systematic review.

    Science.gov (United States)

    Pinto, Camila Lemos; Botelho, Patrícia Borges; Pimentel, Gustavo Duarte; Campos-Ferraz, Patrícia Lopes; Mota, João Felipe

    2016-09-01

    The focus of this review is the effects of creatine supplementation with or without exercise on glucose metabolism. A comprehensive examination of the past 16 years of study within the field provided a distillation of key data. Both in animal and human studies, creatine supplementation together with exercise training demonstrated greater beneficial effects on glucose metabolism; creatine supplementation itself demonstrated positive results in only a few of the studies. In the animal studies, the effects of creatine supplementation on glucose metabolism were even more distinct, and caution is needed in extrapolating these data to different species, especially to humans. Regarding human studies, considering the samples characteristics, the findings cannot be extrapolated to patients who have poorer glycemic control, are older, are on a different pharmacological treatment (e.g., exogenous insulin therapy) or are physically inactive. Thus, creatine supplementation is a possible nutritional therapy adjuvant with hypoglycemic effects, particularly when used in conjunction with exercise.

  3. Mitogen-activated protein kinases mediate Mycobacterium ...

    Indian Academy of Sciences (India)

    2012-01-19

    Jan 19, 2012 ... CD44, an adhesion molecule, has been reported to be a binding site for ... receptors in mediating mitogen-activated protein kinase activation. ... surface expression and tumour necrosis factor-alpha levels, ... Abbreviations used: Abs, antibodies; ANOVA, analysis of variance; AP-1, activator protein -1; BCG, ...

  4. Can creatine supplementation form carcinogenic heterocyclic amines in humans?

    Science.gov (United States)

    Pereira, Renato Tavares dos Santos; Dörr, Felipe Augusto; Pinto, Ernani; Solis, Marina Yazigi; Artioli, Guilherme Giannini; Fernandes, Alan Lins; Murai, Igor Hisashi; Dantas, Wagner Silva; Seguro, Antônio Carlos; Santinho, Mirela Aparecida Rodrigues; Roschel, Hamilton; Carpentier, Alain; Poortmans, Jacques Remi; Gualano, Bruno

    2015-01-01

    Abstract Creatine supplementation has been associated with increased cancer risk. In fact, there is evidence indicating that creatine and/or creatinine are important precursors of carcinogenic heterocyclic amines (HCAs). The present study aimed to investigate the acute and chronic effects of low- and high-dose creatine supplementation on the production of HCAs in healthy humans (i.e. 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (8-MeIQx),  2-amino-(1,6-dimethylfuro[3,2-e]imidazo[4,5-b])pyridine (IFP) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx)). This was a non-counterbalanced single-blind crossover study divided into two phases, in which low- and high-dose creatine protocols were tested. After acute (1 day) and chronic supplementation (30 days), the HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx were assessed through a newly developed HPLC–MS/MS method. Dietary HCA intake and blood and urinary creatinine were also evaluated. Out of 576 assessments performed (from 149 urine samples), only nine (3 from creatine and 6 from placebo) showed quantifiable levels of HCAs (8-MeIQx: n = 3; 4,8-DiMeIQx: n = 2; PhIP: n = 4). Individual analyses revealed that diet rather than creatine supplementation was the main responsible factor for HCA formation in these cases. This study provides compelling evidence that both low and high doses of creatine supplementation, given either acutely or chronically, did not cause increases in the carcinogenic HCAs PhIP, 8-MeIQx, IFP and 4,8-DiMeIQx in healthy subjects. These findings challenge the long-existing notion that creatine supplementation could potentially increase the risk of cancer by stimulating the formation of these mutagens. Key points There is a long-standing concern that creatine supplementation could be associated with cancer, possibly by facilitating the formation of carcinogenic heterocyclic amines (HCAs). This study provides compelling evidence

  5. S -Nitrosylation inhibits the kinase activity of tomato phosphoinositide-dependent kinase 1 (PDK1)

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jian-Zhong; Duan, Jicheng; Ni, Min; Liu, Zhen; Qiu, Wen-Li; Whitham, Steven A.; Qian, Wei-Jun

    2017-09-29

    It is well known that the reactive oxygen species, nitric oxide (NO), can trigger cell death in plants, but the underlying molecular mechanisms are not well understood. Here, we provide evidence that NO may trigger cell death in tomato (Solanum lycopersicon) through inhibiting the phosphoinositide-dependent kinase 1 (PDK1) kinase activity via S-nitrosylation. Biotin-switch assays and LC-MS/MS analyses demonstrated that SlPDK1 was a target of S-nitrosylation modification, which primarily occurred on the cysteine residue at position 128 (Cys128). Accordingly, the kinase activity of SlPDK1 was inhibited by S-nitrosoglutathione (GSNO) both in vitro and in vivo in a concentration-dependent manner, indicating that SlPDK1 activity is regulated by S-nitrosylation. The inhibition of SlPDK1 kinase activity by GSNO was reversible in the presence of a reducing agent but synergistically enhanced by hydrogen peroxide (H2O2). Mutation of Cys128 to serine completely abolished SlPDK1 kinase activity, suggesting that S-nitrosylation of Cys128 is responsible for the inhibition of the kinase activity of SlPDK1. In sum, our results established a potential link between NO-triggered cell death and inhibition of the kinase activity of tomato PDK1, a conserved negative regulator of cell death in yeasts, mammals and plants. Nitric oxide (NO) potentiates the induction of hypersensitive cell death in soybean cells by reactive oxygen species (ROS) (1). However, the molecular mechanism of the NO-induced cell death remains an enigma. One potential mechanism is that the activity of proteins that control cell death may be altered by a post-translational modification, S-nitrosylation. S-nitrosylation is the addition of the NO moiety to thiol groups, including cysteine (Cys) residues in proteins, to form S-nitrosothiols (SNOs). S-nitrosylation is an enzyme-independent post-translational and labile modification that can function as an on/off switch of protein activity (2- 4). Thousands of diverse

  6. Treatment by oral creatine, L-arginine and L-glycine in six severely affected patients with creatine transporter defect

    NARCIS (Netherlands)

    Valayannopoulos, V.; Boddaert, N.; Chabli, A.; Barbier, V.; Desguerre, I.; Philippe, A.; Afenjar, A.; Mazzuca, M.; Cheillan, D.; Munnich, A.; de Keyzer, Y.; Jakobs, C.A.J.M.; Salomons, G.S.; de Lonlay, P.

    2012-01-01

    Background X-linked cerebral creatine deficiency is caused by the deficiency of the creatine transporter (CTP) encoded by the SLC6A8 gene. Patients and Methods We report here a series of six patients with severe CTP deficiency, four males and two females; clinical presentations include mild to

  7. Creatine kinase rate constant in the human heart measured with 3D-localization at 7 tesla.

    Science.gov (United States)

    Clarke, William T; Robson, Matthew D; Neubauer, Stefan; Rodgers, Christopher T

    2017-07-01

    We present a new Bloch-Siegert four Angle Saturation Transfer (BOAST) method for measuring the creatine kinase (CK) first-order effective rate constant k f in human myocardium at 7 tesla (T). BOAST combines a variant of the four-angle saturation transfer (FAST) method using amplitude-modulated radiofrequency pulses, phosphorus Bloch-Siegert B1+-mapping to determine the per-voxel flip angles, and nonlinear fitting to Bloch simulations for postprocessing. Optimal flip angles and repetition time parameters were determined from Monte Carlo simulations. BOAST was validated in the calf muscle of two volunteers at 3T and 7T. The myocardial CK forward rate constant was then measured in 10 volunteers at 7T in 82 min (after 1 H localization). BOAST kfCK values were 0.281 ± 0.002 s -1 in the calf and 0.35 ± 0.05 s -1 in myocardium. These are consistent with literature values from lower fields. Using a literature values for adenosine triphosphate concentration, we computed CK flux values of 4.55 ± 1.52 mmol kg -1 s -1 . The sensitive volume for BOAST depends on the B 1 inhomogeneity of the transmit coil. BOAST enables measurement of the CK rate constant in the human heart at 7T, with spatial localization in three dimensions to 5.6 mL voxels, using a 10-cm loop coil. Magn Reson Med 78:20-32, 2017. © 2016 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2016 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine.

  8. Purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae

    OpenAIRE

    Elbing, Karin; McCartney, Rhonda R.; Schmidt, Martin C.

    2006-01-01

    Members of the Snf1/AMPK family of protein kinases are activated by distinct upstream kinases that phosphorylate a conserved threonine residue in the Snf1/AMPK activation loop. Recently, the identities of the Snf1- and AMPK-activating kinases have been determined. Here we describe the purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae. The identities of proteins associated with the Snf1-activating kinases were determined by peptide mass fingerpr...

  9. The acute effect of beta-guanidinopropionic acid versus creatine or placebo in healthy men (ABC-Trial): A randomized controlled first-in-human trial.

    Science.gov (United States)

    Karamat, Fares A; Horjus, Deborah L; Haan, Yentl C; van der Woude, Lisa; Schaap, Marianne C; Oudman, Inge; van Montfrans, Gert A; Nieuwland, Rienk; Salomons, Gajja S; Clark, Joseph F; Brewster, Lizzy M

    2017-12-01

    Increasing evidence indicates that the ATP-generating enzyme creatine kinase (CK) is involved in hypertension. CK rapidly regenerates ATP from creatine phosphate and ADP. Recently, it has been shown that beta-guanidinopropionic acid (GPA), a kidney-synthesized creatine analogue and competitive CK inhibitor, reduced blood pressure in spontaneously hypertensive rats. To further develop the substance as a potential blood pressure-lowering agent, we assessed the tolerability of a sub-therapeutic GPA dose in healthy men. In this active and placebo-controlled, triple-blind, single-centre trial, we recruited 24 healthy men (18-50 years old, BMI 18.5-29.9 kg m -2 ) in the Netherlands. Participants were randomized (1:1:1) to one week daily oral administration of GPA 100 mg, creatine 5 g, or matching placebo. The primary outcome was the tolerability of GPA, in an intent-to-treat analysis. Twenty-four randomized participants received the allocated intervention and 23 completed the study. One participant in the placebo arm dropped out for personal reasons. GPA was well tolerated, without serious or severe adverse events. No abnormalities were reported with GPA use in clinical safety parameters, including physical examination, laboratory studies, or 12-Lead ECG. At day 8, mean plasma GPA was 213.88 (SE 0.07) in the GPA arm vs. 32.75 (0.00) nmol l -1 in the placebo arm, a mean difference of 181.13 (95% CI 26.53-335.72). In this first-in-human trial, low-dose GPA was safe and well-tolerated when used during 1 week in healthy men. Subsequent studies should focus on human pharmacokinetic and pharmacodynamic assessments with different doses. © 2017 The Authors. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.

  10. Activation of Extracellular Signal-Regulated Kinase but Not of p38 Mitogen-Activated Protein Kinase Pathways in Lymphocytes Requires Allosteric Activation of SOS

    Science.gov (United States)

    Jun, Jesse E.; Yang, Ming; Chen, Hang; Chakraborty, Arup K.

    2013-01-01

    Thymocytes convert graded T cell receptor (TCR) signals into positive selection or deletion, and activation of extracellular signal-related kinase (ERK), p38, and Jun N-terminal protein kinase (JNK) mitogen-activated protein kinases (MAPKs) has been postulated to play a discriminatory role. Two families of Ras guanine nucleotide exchange factors (RasGEFs), SOS and RasGRP, activate Ras and the downstream RAF-MEK-ERK pathway. The pathways leading to lymphocyte p38 and JNK activation are less well defined. We previously described how RasGRP alone induces analog Ras-ERK activation while SOS and RasGRP cooperate to establish bimodal ERK activation. Here we employed computational modeling and biochemical experiments with model cell lines and thymocytes to show that TCR-induced ERK activation grows exponentially in thymocytes and that a W729E allosteric pocket mutant, SOS1, can only reconstitute analog ERK signaling. In agreement with RasGRP allosterically priming SOS, exponential ERK activation is severely decreased by pharmacological or genetic perturbation of the phospholipase Cγ (PLCγ)-diacylglycerol-RasGRP1 pathway. In contrast, p38 activation is not sharply thresholded and requires high-level TCR signal input. Rac and p38 activation depends on SOS1 expression but not allosteric activation. Based on computational predictions and experiments exploring whether SOS functions as a RacGEF or adaptor in Rac-p38 activation, we established that the presence of SOS1, but not its enzymatic activity, is critical for p38 activation. PMID:23589333

  11. Neural cell adhesion molecule-stimulated neurite outgrowth depends on activation of protein kinase C and the Ras-mitogen-activated protein kinase pathway

    DEFF Research Database (Denmark)

    Kolkova, K; Novitskaya, V; Pedersen, N

    2000-01-01

    , inhibitors of the nonreceptor tyrosine kinase p59(fyn), PLC, PKC and MEK and an activator of PKC, phorbol-12-myristate-13-acetate (PMA). MEK2 transfection rescued cells treated with all inhibitors. The same was found for PMA treatment, except when cells concomitantly were treated with the MEK inhibitor....... Arachidonic acid rescued cells treated with antibodies to the FGF receptor or the PLC inhibitor, but not cells in which the activity of PKC, p59(fyn), FAK, Ras, or MEK was inhibited. Interaction of NCAM with a synthetic NCAM peptide ligand, known to induce neurite outgrowth, was shown to stimulate...... phosphorylation of the MAP kinases extracellular signal-regulated kinases ERK1 and ERK2. The MAP kinase activation was sustained, because ERK1 and ERK2 were phosphorylated in PC12-E2 cells and primary hippocampal neurons even after 24 hr of cultivation on NCAM-expressing fibroblasts. Based on these results, we...

  12. Involvement of protein kinase B and mitogen-activated protein kinases in experimental normothermic liver ischaemia-reperfusion injury.

    Science.gov (United States)

    Cursio, R; Filippa, N; Miele, C; Van Obberghen, E; Gugenheim, J

    2006-06-01

    This study evaluated the role of protein kinase B (PKB), phosphatidylinositol 3-kinase (PI3-K), Bcl-2-associated death protein (BAD) and mitogen-activated protein kinases (MAPKs) in normothermic ischaemia-reperfusion (IR)-induced apoptosis in rat liver. Rats were divided into two groups that received either phosphate-buffered saline (control) or the caspase inhibitor Z-Asp-2,6-dichorobenzoyloxymethylketone (Z-Asp-cmk), injected intravenously 2 min before the induction of 120 min of normothermic liver ischaemia. Liver apoptosis was assessed by the terminal deoxyribonucleotidyltransferase-mediated dUTP nick end labelling (TUNEL) method. PI3-K, PKB, BAD and MAPK activities were measured in ischaemic and non-ischaemic lobes at various times after reperfusion. The number of TUNEL-positive cells was significantly decreased after pretreatment with Z-Asp-cmk. In controls, PI3-K and PKB activities and BAD phosphorylation were inhibited in ischaemic liver lobes. The MAPKs (extracellular signal-regulated kinases, c-Jun N-terminal kinase and p38) showed different patterns of activation during IR. PKB activity was not modified by pretreatment with Z-Asp-cmk. Induction of apoptosis during IR liver injury might be triggered by inactivation of the antiapoptotic PI3-K-PKB pathway and activation of the proapoptotic MAPKs. Copyright (c) 2006 British Journal of Surgery Society Ltd. Published by John Wiley & Sons, Ltd.

  13. Creatine pretreatment protects cortical axons from energy depletion in vitro

    Science.gov (United States)

    Shen, Hua; Goldberg, Mark P.

    2012-01-01

    Creatine is a natural nitrogenous guanidino compound involved in bioenergy metabolism. Although creatine has been shown to protect neurons of the central nervous system (CNS) from experimental hypoxia/ischemia, it remains unclear if creatine may also protect CNS axons, and if the potential axonal protection depends on glial cells. To evaluate the direct impact of creatine on CNS axons, cortical axons were cultured in a separate compartment from their somas and proximal neurites using a modified two-compartment culture device. Axons in the axon compartment were subjected to acute energy depletion, an in vitro model of white matter ischemia, by exposure to 6 mM sodium azide for 30 min in the absence of glucose and pyruvate. Energy depletion reduced axonal ATP by 65%, depolarized axonal resting potential, and damaged 75% of axons. Application of creatine (10 mM) to both compartments of the culture at 24 h prior to energy depletion significantly reduced axonal damage by 50%. In line with the role of creatine in the bioenergy metabolism, this application also alleviated the axonal ATP loss and depolarization. Inhibition of axonal depolarization by blocking sodium influx with tetrodotoxin also effectively reduced the axonal damage caused by energy depletion. Further study revealed that the creatine effect was independent of glial cells, as axonal protection was sustained even when creatine was applied only to the axon compartment (free from somas and glial cells) for as little as 2 h. In contrast, application of creatine after energy depletion did not protect axons. The data provide the first evidence that creatine pretreatment may directly protect CNS axons from energy deficiency. PMID:22521466

  14. Creatine Enhances Mitochondrial-Mediated Oligodendrocyte Survival After Demyelinating Injury.

    Science.gov (United States)

    Chamberlain, Kelly A; Chapey, Kristen S; Nanescu, Sonia E; Huang, Jeffrey K

    2017-02-08

    Chronic oligodendrocyte loss, which occurs in the demyelinating disorder multiple sclerosis (MS), contributes to axonal dysfunction and neurodegeneration. Current therapies are able to reduce MS severity, but do not prevent transition into the progressive phase of the disease, which is characterized by chronic neurodegeneration. Therefore, pharmacological compounds that promote oligodendrocyte survival could be beneficial for neuroprotection in MS. Here, we investigated the role of creatine, an organic acid involved in adenosine triphosphate (ATP) buffering, in oligodendrocyte function. We found that creatine increased mitochondrial ATP production directly in oligodendrocyte lineage cell cultures and exerted robust protection on oligodendrocytes by preventing cell death in both naive and lipopolysaccharide-treated mixed glia. Moreover, lysolecithin-mediated demyelination in mice deficient in the creatine-synthesizing enzyme guanidinoacetate-methyltransferase ( Gamt ) did not affect oligodendrocyte precursor cell recruitment, but resulted in exacerbated apoptosis of regenerated oligodendrocytes in central nervous system (CNS) lesions. Remarkably, creatine administration into Gamt -deficient and wild-type mice with demyelinating injury reduced oligodendrocyte apoptosis, thereby increasing oligodendrocyte density and myelin basic protein staining in CNS lesions. We found that creatine did not affect the recruitment of macrophages/microglia into lesions, suggesting that creatine affects oligodendrocyte survival independently of inflammation. Together, our results demonstrate a novel function for creatine in promoting oligodendrocyte viability during CNS remyelination. SIGNIFICANCE STATEMENT We report that creatine enhances oligodendrocyte mitochondrial function and protects against caspase-dependent oligodendrocyte apoptosis during CNS remyelination. This work has important implications for the development of therapeutic targets for diseases characterized by

  15. Umbrella sampling of proton transfer in a creatine-water system

    Science.gov (United States)

    Ivchenko, Olga; Bachert, Peter; Imhof, Petra

    2014-04-01

    Proton transfer reactions are among the most common processes in chemistry and biology. Proton transfer between creatine and surrounding solvent water is underlying the chemical exchange saturation transfer used as a contrast in magnetic resonance imaging. The free energy barrier, determined by first-principles umbrella sampling simulations (EaDFT 3 kcal/mol) is in the same order of magnitude as the experimentally obtained activation energy. The underlying mechanism is a first proton transfer from the guanidinium group to the water pool, followed by a second transition where a proton is "transferred back" from the nearest water molecule to the deprotonated nitrogen atom of creatine.

  16. Puerarin activates endothelial nitric oxide synthase through estrogen receptor-dependent PI3-kinase and calcium-dependent AMP-activated protein kinase

    International Nuclear Information System (INIS)

    Hwang, Yong Pil; Kim, Hyung Gyun; Hien, Tran Thi; Jeong, Myung Ho; Jeong, Tae Cheon; Jeong, Hye Gwang

    2011-01-01

    The cardioprotective properties of puerarin, a natural product, have been attributed to the endothelial nitric oxide synthase (eNOS)-mediated production of nitric oxide (NO) in EA.hy926 endothelial cells. However, the mechanism by which puerarin activates eNOS remains unclear. In this study, we sought to identify the intracellular pathways underlying eNOS activation by puerarin. Puerarin induced the activating phosphorylation of eNOS on Ser1177 and the production of NO in EA.hy926 cells. Puerarin-induced eNOS phosphorylation required estrogen receptor (ER)-mediated phosphatidylinositol 3-kinase (PI3K)/Akt signaling and was reversed by AMP-activated protein kinase (AMPK) and calcium/calmodulin-dependent kinase II (CaMKII) inhibition. Importantly, puerarin inhibited the adhesion of tumor necrosis factor (TNF)-α-stimulated monocytes to endothelial cells and suppressed the TNF-α induced expression of intercellular cell adhesion molecule-1. Puerarin also inhibited the TNF-α-induced nuclear factor-κB activation, which was attenuated by pretreatment with N G -nitro-L-arginine methyl ester, a NOS inhibitor. These results indicate that puerarin stimulates eNOS phosphorylation and NO production via activation of an estrogen receptor-mediated PI3K/Akt- and CaMKII/AMPK-dependent pathway. Puerarin may be useful for the treatment or prevention of endothelial dysfunction associated with diabetes and cardiovascular disease. -- Highlights: ► Puerarin induced the phosphorylation of eNOS and the production of NO. ► Puerarin activated eNOS through ER-dependent PI3-kinase and Ca 2+ -dependent AMPK. ► Puerarin-induced NO was involved in the inhibition of NF-kB activation. ► Puerarin may help for prevention of vascular dysfunction and diabetes.

  17. Recruitment of focal adhesion kinase and paxillin to β1 integrin promotes cancer cell migration via mitogen activated protein kinase activation

    International Nuclear Information System (INIS)

    Crowe, David L; Ohannessian, Arthur

    2004-01-01

    Integrin-extracellular matrix interactions activate signaling cascades such as mitogen activated protein kinases (MAPK). Integrin binding to extracellular matrix increases tyrosine phosphorylation of focal adhesion kinase (FAK). Inhibition of FAK activity by expression of its carboxyl terminus decreases cell motility, and cells from FAK deficient mice also show reduced migration. Paxillin is a focal adhesion protein which is also phosphorylated on tyrosine. FAK recruitment of paxillin to the cell membrane correlates with Shc phosphorylation and activation of MAPK. Decreased FAK expression inhibits papilloma formation in a mouse skin carcinogenesis model. We previously demonstrated that MAPK activation was required for growth factor induced in vitro migration and invasion by human squamous cell carcinoma (SCC) lines. Adapter protein recruitment to integrin subunits was examined by co-immunoprecipitation in SCC cells attached to type IV collagen or plastic. Stable clones overexpressing FAK or paxillin were created using the lipofection technique. Modified Boyden chambers were used for invasion assays. In the present study, we showed that FAK and paxillin but not Shc are recruited to the β1 integrin cytoplasmic domain following attachment of SCC cells to type IV collagen. Overexpression of either FAK or paxillin stimulated cancer cell migration on type IV collagen and invasion through reconstituted basement membrane which was dependent on MAPK activity. We concluded that recruitment of focal adhesion kinase and paxillin to β1 integrin promoted cancer cell migration via the mitogen activated protein kinase pathway

  18. Recruitment of focal adhesion kinase and paxillin to β1 integrin promotes cancer cell migration via mitogen activated protein kinase activation

    Directory of Open Access Journals (Sweden)

    Ohannessian Arthur

    2004-05-01

    Full Text Available Abstract Background Integrin-extracellular matrix interactions activate signaling cascades such as mitogen activated protein kinases (MAPK. Integrin binding to extracellular matrix increases tyrosine phosphorylation of focal adhesion kinase (FAK. Inhibition of FAK activity by expression of its carboxyl terminus decreases cell motility, and cells from FAK deficient mice also show reduced migration. Paxillin is a focal adhesion protein which is also phosphorylated on tyrosine. FAK recruitment of paxillin to the cell membrane correlates with Shc phosphorylation and activation of MAPK. Decreased FAK expression inhibits papilloma formation in a mouse skin carcinogenesis model. We previously demonstrated that MAPK activation was required for growth factor induced in vitro migration and invasion by human squamous cell carcinoma (SCC lines. Methods Adapter protein recruitment to integrin subunits was examined by co-immunoprecipitation in SCC cells attached to type IV collagen or plastic. Stable clones overexpressing FAK or paxillin were created using the lipofection technique. Modified Boyden chambers were used for invasion assays. Results In the present study, we showed that FAK and paxillin but not Shc are recruited to the β1 integrin cytoplasmic domain following attachment of SCC cells to type IV collagen. Overexpression of either FAK or paxillin stimulated cancer cell migration on type IV collagen and invasion through reconstituted basement membrane which was dependent on MAPK activity. Conclusions We concluded that recruitment of focal adhesion kinase and paxillin to β1 integrin promoted cancer cell migration via the mitogen activated protein kinase pathway.

  19. Purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae.

    Science.gov (United States)

    Elbing, Karin; McCartney, Rhonda R; Schmidt, Martin C

    2006-02-01

    Members of the Snf1/AMPK family of protein kinases are activated by distinct upstream kinases that phosphorylate a conserved threonine residue in the Snf1/AMPK activation loop. Recently, the identities of the Snf1- and AMPK-activating kinases have been determined. Here we describe the purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae. The identities of proteins associated with the Snf1-activating kinases were determined by peptide mass fingerprinting. These kinases, Sak1, Tos3 and Elm2 do not appear to require the presence of additional subunits for activity. Sak1 and Snf1 co-purify and co-elute in size exclusion chromatography, demonstrating that these two proteins form a stable complex. The Snf1-activating kinases phosphorylate the activation loop threonine of Snf1 in vitro with great specificity and are able to do so in the absence of beta and gamma subunits of the Snf1 heterotrimer. Finally, we showed that the Snf1 kinase domain isolated from bacteria as a GST fusion protein can be activated in vitro and shows substrate specificity in the absence of its beta and gamma subunits.

  20. Damage-induced DNA replication stalling relies on MAPK-activated protein kinase 2 activity

    DEFF Research Database (Denmark)

    Köpper, Frederik; Bierwirth, Cathrin; Schön, Margarete

    2013-01-01

    knockdown of the MAP kinase-activated protein kinase 2 (MK2), a kinase currently implicated in p38 stress signaling and G2 arrest. Depletion or inhibition of MK2 also protected cells from DNA damage-induced cell death, and mice deficient for MK2 displayed decreased apoptosis in the skin upon UV irradiation...

  1. Estimation of infarct size by myocardial emission computed tomography with thallium-201 and its relation to creatine kinase-MB release after myocardial infarction in man

    International Nuclear Information System (INIS)

    Tamaki, S.; Nakajima, H.; Murakami, T.

    1982-01-01

    Emission computed tomography (ECT) for thallium-201 ( 201 Tl) myocardial imaging was evaluated in estimating infarct size (IS). In 18 patients in whom IS was estimated enzymatically at the time of the acute episode, planar 201 Tl perfusion scintigraphy and ECT with a rotating gamma camera were performed 4 weeks after the first myocardial infarction. From the size of 201 Tl perfusion defects, the infarct area in planar images and the infarct volume in reconsturcted ECT images were measured by computerized planimetry. When scintigraphic IS was compared with the accumulated creatine kinase-MB isoenzyme release (CK-MBr), infarct volume determined from ECT correlated closely with CK-MBr (r=0.89), whereas infarct area measured from planar images correlated less satisfactorily with the enzymatic IS (for an average infarct area from three views, r=0.69; for the largest infarct area, r=0.73). Although conventional scintigraphic evaluation is useful for detecting and localizing infarction, quantification of ischemic injury with this two-dimensional technique has a significant inherent limitation. The ECT approach can provide a more accurate three-dimensional quantitative estimate of infarction, and can corroborate the enzymatic estimate of IS

  2. Estimation of infarct size by myocardial emission computed tomography with 201Tl and its relation to creatine kinase-MB release after myocardial infarction in man

    International Nuclear Information System (INIS)

    Tamaki, S.; Nakajima, H.; Murakami, T.

    1982-01-01

    We evaluated emission computed tomography (ECT) 201 Tl myocardial imaging in estimating infarct size (IS). In 18 patients in whom IS was estimated enzymatically at the time of the acute episode, planar 201 Tl perfusion scintigraphy and ECT with a rotating gamma camera were performed 4 weeks after the first myocardial infarction. From the size of 201 Tl perfusion defects, the infarct area in planar images and the infarct volume in reconstructed ECT images were measured by computerized planimetry. When scintigraphic IS was compared with the accumulated creatine kinase-MB isoenzyme release (CK-MBr), infarct volume determined from ECT correlated closely with CK-MBr (r . 0.89), whereas infarct area measured from planar images correlated less satisfactorily with the enzymatic IS (for an average infarct area from three views, r . 0.69; for the largest infarct area, r . 0.73). Although conventional scintigraphic evaluation is useful for detecting and localizing infarction, quantification of ischemic injury with this two-dimensional technique has a significant inherent limitation. The ECT approach can provide a more accurate three-dimensional quantitative estimate of infarction, and can corroborate the enzymatic estimate of IS

  3. Angiotensin II regulation of neuromodulation: downstream signaling mechanism from activation of mitogen-activated protein kinase.

    Science.gov (United States)

    Lu, D; Yang, H; Raizada, M K

    1996-12-01

    Angiotensin II (Ang II) stimulates expression of tyrosine hydroxylase and norepinephrine transporter genes in brain neurons; however, the signal-transduction mechanism is not clearly defined. This study was conducted to determine the involvement of the mitogen-activated protein (MAP) kinase signaling pathway in Ang II stimulation of these genes. MAP kinase was localized in the perinuclear region of the neuronal soma. Ang II caused activation of MAP kinase and its subsequent translocation from the cytoplasmic to nuclear compartment, both effects being mediated by AT1 receptor subtype. Ang II also stimulated SRE- and AP1-binding activities and fos gene expression and its translocation in a MAP kinase-dependent process. These observations are the first demonstration of a downstream signaling pathway involving MAP kinase in Ang II-mediated neuromodulation in noradrenergic neurons.

  4. Protective features of resveratrol on human spermatozoa cryopreservation may be mediated through 5' AMP-activated protein kinase activation.

    Science.gov (United States)

    Shabani Nashtaei, M; Amidi, F; Sedighi Gilani, M A; Aleyasin, A; Bakhshalizadeh, Sh; Naji, M; Nekoonam, S

    2017-03-01

    Biochemical and physical modifications during the freeze-thaw process adversely influence the restoration of energy-dependent sperm functions required for fertilization. Resveratrol, a phytoalexin, has been introduced to activate 5' AMP-activated protein kinase which is a cell energy sensor and a cell metabolism regulator. The cryoprotection of resveratrol on sperm cryoinjury via activation of AMP-activated protein kinase also remains to be elucidated. Our aim, thus, was to investigate: (i) the presence and intracellular localization of AMP-activated protein kinase protein; (ii) whether resveratrol may exert a protective effect on certain functional properties of fresh and post-thaw human spermatozoa through modulation of AMP-activated protein kinase. Spermatozoa from normozoospermic men were incubated with or without different concentrations of Compound C as an AMP-activated protein kinase inhibitor or resveratrol as an AMP-activated protein kinase activator for different lengths of time and were then cryopreserved. AMP-activated protein kinase is expressed essentially in the entire flagellum and the post-equatorial region. Viability of fresh spermatozoa was not significantly affected by the presence of Compound C or resveratrol. However, although Compound C caused a potent inhibition of spermatozoa motility parameters, resveratrol did not induce negative effect, except a significant reduction in motility at 25 μm for 1 h. Furthermore, resveratrol significantly increased AMP-activated protein kinase phosphorylation and mitochondrial membrane potential and decreased reactive oxygen species and apoptosis-like changes in frozen-thawed spermatozoa. Nevertheless, it was not able to compensate decreased sperm viability and motility parameters following cryopreservation. In contrast, Compound C showed opposite effects to resveratrol on AMP-activated protein kinase phosphorylation, reactive oxygen species, apoptosis-like changes, mitochondrial membrane potential, and

  5. Creatine Supplementation and Skeletal Muscle Metabolism for Building Muscle Mass- Review of the Potential Mechanisms of Action.

    Science.gov (United States)

    Farshidfar, Farnaz; Pinder, Mark A; Myrie, Semone B

    2017-01-01

    Creatine, a very popular supplement among athletic populations, is of growing interest for clinical applications. Since over 90% of creatine is stored in skeletal muscle, the effect of creatine supplementation on muscle metabolism is a widely studied area. While numerous studies over the past few decades have shown that creatine supplementation has many favorable effects on skeletal muscle physiology and metabolism, including enhancing muscle mass (growth/hypertrophy); the underlying mechanisms are poorly understood. This report reviews studies addressing the mechanisms of action of creatine supplementation on skeletal muscle growth/hypertrophy. Early research proposed that the osmotic effect of creatine supplementation serves as a cellular stressor (osmosensing) that acts as an anabolic stimulus for protein synthesis signal pathways. Other reports indicated that creatine directly affects muscle protein synthesis via modulations of components in the mammalian target of rapamycin (mTOR) pathway. Creatine may also directly affect the myogenic process (formation of muscle tissue), by altering secretions of myokines, such as myostatin and insulin-like growth factor-1, and expressions of myogenic regulatory factors, resulting in enhanced satellite cells mitotic activities and differentiation into myofiber. Overall, there is still no clear understanding of the mechanisms of action regarding how creatine affects muscle mass/growth, but current evidence suggests it may exert its effects through multiple approaches, with converging impacts on protein synthesis and myogenesis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  6. Nuclear localization of Lyn tyrosine kinase mediated by inhibition of its kinase activity

    International Nuclear Information System (INIS)

    Ikeda, Kikuko; Nakayama, Yuji; Togashi, Yuuki; Obata, Yuuki; Kuga, Takahisa; Kasahara, Kousuke; Fukumoto, Yasunori; Yamaguchi, Naoto

    2008-01-01

    Src-family kinases, cytoplasmic enzymes that participate in various signaling events, are found at not only the plasma membrane but also subcellular compartments, such as the nucleus, the Golgi apparatus and late endosomes/lysosomes. Lyn, a member of the Src-family kinases, is known to play a role in DNA damage response and cell cycle control in the nucleus. However, it is still unclear how the localization of Lyn to the nucleus is regulated. Here, we investigated the mechanism of the distribution of Lyn between the cytoplasm and the nucleus in epitheloid HeLa cells and hematopoietic THP-1 cells. Lyn was definitely detected in purified nuclei by immunofluorescence and immunoblotting analyses. Nuclear accumulation of Lyn was enhanced upon treatment of cells with leptomycin B (LMB), an inhibitor of Crm1-mediated nuclear export. Moreover, Lyn mutants lacking the sites for lipid modification were highly accumulated in the nucleus upon LMB treatment. Intriguingly, inhibition of the kinase activity of Lyn by SU6656, Csk overexpression, or point mutation in the ATP-binding site induced an increase in nuclear Lyn levels. These results suggest that Lyn being imported into and rapidly exported from the nucleus preferentially accumulates in the nucleus by inhibition of the kinase activity and lipid modification

  7. Moonlighting kinases with guanylate cyclase activity can tune regulatory signal networks

    KAUST Repository

    Irving, Helen R.; Kwezi, Lusisizwe; Wheeler, Janet I.; Gehring, Christoph A

    2012-01-01

    Guanylate cyclase (GC) catalyzes the formation of cGMP and it is only recently that such enzymes have been characterized in plants. One family of plant GCs contains the GC catalytic center encapsulated within the intracellular kinase domain of leucine rich repeat receptor like kinases such as the phytosulfokine and brassinosteroid receptors. In vitro studies show that both the kinase and GC domain have catalytic activity indicating that these kinase-GCs are examples of moonlighting proteins with dual catalytic function. The natural ligands for both receptors increase intracellular cGMP levels in isolated mesophyll protoplast assays suggesting that the GC activity is functionally relevant. cGMP production may have an autoregulatory role on receptor kinase activity and/or contribute to downstream cell expansion responses. We postulate that the receptors are members of a novel class of receptor kinases that contain functional moonlighting GC domains essential for complex signaling roles.

  8. Moonlighting kinases with guanylate cyclase activity can tune regulatory signal networks

    KAUST Repository

    Irving, Helen R.

    2012-02-01

    Guanylate cyclase (GC) catalyzes the formation of cGMP and it is only recently that such enzymes have been characterized in plants. One family of plant GCs contains the GC catalytic center encapsulated within the intracellular kinase domain of leucine rich repeat receptor like kinases such as the phytosulfokine and brassinosteroid receptors. In vitro studies show that both the kinase and GC domain have catalytic activity indicating that these kinase-GCs are examples of moonlighting proteins with dual catalytic function. The natural ligands for both receptors increase intracellular cGMP levels in isolated mesophyll protoplast assays suggesting that the GC activity is functionally relevant. cGMP production may have an autoregulatory role on receptor kinase activity and/or contribute to downstream cell expansion responses. We postulate that the receptors are members of a novel class of receptor kinases that contain functional moonlighting GC domains essential for complex signaling roles.

  9. Activation of G-proteins by receptor-stimulated nucleoside diphosphate kinase in Dictyostelium.

    Science.gov (United States)

    Bominaar, A A; Molijn, A C; Pestel, M; Veron, M; Van Haastert, P J

    1993-01-01

    Recently, interest in the enzyme nucleoside diphosphate kinase (EC2.7.4.6) has increased as a result of its possible involvement in cell proliferation and development. Since NDP kinase is one of the major sources of GTP in cells, it has been suggested that the effects of an altered NDP kinase activity on cellular processes might be the result of altered transmembrane signal transduction via guanine nucleotide-binding proteins (G-proteins). In the cellular slime mould Dictyostelium discoideum, extracellular cAMP induces an increase of phospholipase C activity via a surface cAMP receptor and G-proteins. In this paper it is demonstrated that part of the cellular NDP kinase is associated with the membrane and stimulated by cell surface cAMP receptors. The GTP produced by the action of NDP kinase is capable of activating G-proteins as monitored by altered G-protein-receptor interaction and the activation of the effector enzyme phospholipase C. Furthermore, specific monoclonal antibodies inhibit the effect of NDP kinase on G-protein activation. These results suggest that receptor-stimulated NDP kinase contributes to the mediation of hormone action by producing GTP for the activation of GTP-binding proteins. Images PMID:8389692

  10. Calcium-Oxidant Signaling Network Regulates AMP-activated Protein Kinase (AMPK) Activation upon Matrix Deprivation*

    Science.gov (United States)

    Sundararaman, Ananthalakshmy; Amirtham, Usha; Rangarajan, Annapoorni

    2016-01-01

    The AMP-activated protein kinase (AMPK) has recently been implicated in anoikis resistance. However, the molecular mechanisms that activate AMPK upon matrix detachment remain unexplored. In this study, we show that AMPK activation is a rapid and sustained phenomenon upon matrix deprivation, whereas re-attachment to the matrix leads to its dephosphorylation and inactivation. Because matrix detachment leads to loss of integrin signaling, we investigated whether integrin signaling negatively regulates AMPK activation. However, modulation of focal adhesion kinase or Src, the major downstream components of integrin signaling, failed to cause a corresponding change in AMPK signaling. Further investigations revealed that the upstream AMPK kinases liver kinase B1 (LKB1) and Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ) contribute to AMPK activation upon detachment. In LKB1-deficient cells, we found AMPK activation to be predominantly dependent on CaMKKβ. We observed no change in ATP levels under detached conditions at early time points suggesting that rapid AMPK activation upon detachment was not triggered by energy stress. We demonstrate that matrix deprivation leads to a spike in intracellular calcium as well as oxidant signaling, and both these intracellular messengers contribute to rapid AMPK activation upon detachment. We further show that endoplasmic reticulum calcium release-induced store-operated calcium entry contributes to intracellular calcium increase, leading to reactive oxygen species production, and AMPK activation. We additionally show that the LKB1/CaMKK-AMPK axis and intracellular calcium levels play a critical role in anchorage-independent cancer sphere formation. Thus, the Ca2+/reactive oxygen species-triggered LKB1/CaMKK-AMPK signaling cascade may provide a quick, adaptable switch to promote survival of metastasizing cancer cells. PMID:27226623

  11. Puerarin activates endothelial nitric oxide synthase through estrogen receptor-dependent PI3-kinase and calcium-dependent AMP-activated protein kinase

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Yong Pil; Kim, Hyung Gyun [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon (Korea, Republic of); Hien, Tran Thi [College of Pharmacy, Chosun University, Gwangju (Korea, Republic of); Jeong, Myung Ho [Heart Research Center, Chonnam National University Hospital, Gwangju (Korea, Republic of); Jeong, Tae Cheon, E-mail: taecheon@ynu.ac.kr [College of Pharmacy, Yeungnam University, Gyungsan (Korea, Republic of); Jeong, Hye Gwang, E-mail: hgjeong@cnu.ac.kr [Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon (Korea, Republic of)

    2011-11-15

    The cardioprotective properties of puerarin, a natural product, have been attributed to the endothelial nitric oxide synthase (eNOS)-mediated production of nitric oxide (NO) in EA.hy926 endothelial cells. However, the mechanism by which puerarin activates eNOS remains unclear. In this study, we sought to identify the intracellular pathways underlying eNOS activation by puerarin. Puerarin induced the activating phosphorylation of eNOS on Ser1177 and the production of NO in EA.hy926 cells. Puerarin-induced eNOS phosphorylation required estrogen receptor (ER)-mediated phosphatidylinositol 3-kinase (PI3K)/Akt signaling and was reversed by AMP-activated protein kinase (AMPK) and calcium/calmodulin-dependent kinase II (CaMKII) inhibition. Importantly, puerarin inhibited the adhesion of tumor necrosis factor (TNF)-{alpha}-stimulated monocytes to endothelial cells and suppressed the TNF-{alpha} induced expression of intercellular cell adhesion molecule-1. Puerarin also inhibited the TNF-{alpha}-induced nuclear factor-{kappa}B activation, which was attenuated by pretreatment with N{sup G}-nitro-L-arginine methyl ester, a NOS inhibitor. These results indicate that puerarin stimulates eNOS phosphorylation and NO production via activation of an estrogen receptor-mediated PI3K/Akt- and CaMKII/AMPK-dependent pathway. Puerarin may be useful for the treatment or prevention of endothelial dysfunction associated with diabetes and cardiovascular disease. -- Highlights: Black-Right-Pointing-Pointer Puerarin induced the phosphorylation of eNOS and the production of NO. Black-Right-Pointing-Pointer Puerarin activated eNOS through ER-dependent PI3-kinase and Ca{sup 2+}-dependent AMPK. Black-Right-Pointing-Pointer Puerarin-induced NO was involved in the inhibition of NF-kB activation. Black-Right-Pointing-Pointer Puerarin may help for prevention of vascular dysfunction and diabetes.

  12. The phosphatidylinositol 3-kinase inhibitor, wortmannin, inhibits insulin-induced activation of phosphatidylcholine hydrolysis and associated protein kinase C translocation in rat adipocytes.

    OpenAIRE

    Standaert, M L; Avignon, A; Yamada, K; Bandyopadhyay, G; Farese, R V

    1996-01-01

    We questioned whether phosphatidylinositol 3-kinase (PI 3-kinase) and protein kinase C (PKC) function as interrelated signalling mechanisms during insulin action in rat adipocytes. Insulin rapidly activated a phospholipase D that hydrolyses phosphatidylcholine (PC), and this activation was accompanied by increases in diacylglycerol and translocative activation of PKC-alpha and PKC-beta in the plasma membrane. Wortmannin, an apparently specific PI 3-kinase inhibitor, inhibited insulin-stimulat...

  13. Creatine protects against mitochondrial dysfunction associated with HIV-1 Tat-induced neuronal injury

    Science.gov (United States)

    Stevens, Patrick R.; Gawryluk, Jeremy W.; Hui, Liang; Chen, Xuesong; Geiger, Jonathan D.

    2015-01-01

    HIV-1 infected individuals are living longer but experiencing a prevalence rate of over 50% for HIV-1 associated neurocognitive disorders (HAND) for which no effective treatment is available. Viral and cellular factors secreted by HIV-1 infected cells leads to neuronal injury and HIV-1 Tat continues to be implicated in the pathogenesis of HAND. Here we tested the hypothesis that creatine protected against HIV-1 Tat-induced neuronal injury by preventing mitochondrial bioenergetic crisis and/or redox catastrophe. Creatine blocked HIV-1 Tat1-72-induced increases in neuron cell death and synaptic area loss. Creatine protected against HIV-1 Tat-induced decreases in ATP. Creatine and creatine plus HIV-1 Tat increased cellular levels of creatine, and creatine plus HIV-1 Tat further decreased ratios of phosphocreatine to creatine observed with creatine or HIV-1 Tat treatments alone. Additionally, creatine protected against HIV-1 Tat-induced mitochondrial hypopolarization and HIV-1 Tat-induced mitochondrial permeability transition pore opening. Thus, creatine may be a useful adjunctive therapy against HAND. PMID:25613139

  14. Effect of transportation stress on heat shock protein 70 concentration and mRNA expression in heart and kidney tissues and serum enzyme activities and hormone concentrations of pigs.

    Science.gov (United States)

    Yu, Hong; Bao, En-Dong; Zhao, Ru-Qian; Lv, Qiong-Xia

    2007-11-01

    To determine the enzymatic and hormonal responses, heat shock protein 70 (Hsp70) production, and Hsp70 mRNA expression in heart and kidney tissues of transport-stressed pigs. 24 pigs (mean weight, 20 +/- 1 kg). Pigs were randomly placed into groups of 12 each. One group was transported for 2 hours. The other group was kept under normal conditions and used as control pigs. Sera were used to detect triiodothyronine, thyroxine, and cortisol concentrations and alanine aminotransferase, aspartate aminotransferase, and creatine kinase activities. The heart and kidneys of anesthetized pigs were harvested and frozen in liquid nitrogen for quantification of Hsp70 and Hsp70 mRNA. No significant differences were detected in serum alanine aminotransferase activity and triiodothyronine and cortisol concentrations between groups; however, the serum creatine kinase and aspartate aminotransferase activities and thyroxine concentrations were higher in transported pigs. Densitometric readings of western blots revealed that the amount of Hsp70 in heart and kidney tissues was significantly higher in transported pigs, compared with control pigs. Results of fluorescence quantitative real-time PCR assay revealed that the Hsp70 mRNA transcription in heart tissue, but not kidney tissue, was significantly higher in transported pigs, compared with control pigs. Transportation imposed a severe stress on pigs that was manifested as increased serum activities of aspartate aminotransferase and creatine kinase and increased amounts of Hsp70 and Hsp70 mRNA expression in heart and kidney tissues. Changes in serum enzyme activities were related to the tissue damage of transport-stressed pigs.

  15. EXERCISE PERFORMANCE AND MUSCLE CONTRACTILE PROPERTIES AFTER CREATINE MONOHYDRATE SUPPLEMENTATION IN AEROBIC-ANAEROBIC TRAINING RATS

    Directory of Open Access Journals (Sweden)

    Nickolay Boyadjiev

    2007-12-01

    Full Text Available The purpose of this study was to investigate the effects of creatine monohydrate supplementation on exercise performance and contractile variables in aerobic-anaerobic training rats. Twenty 90-day-old male Sprague Dawley rats were divided into two groups - creatine (Cr and controls (K. The creatine group received creatine monohydrate as a nutritional supplement, whereas the control group was given placebo. Both groups were trained 5 days a week on a treadmill for 20 days in a mixed (aerobic-anaerobic metabolic working regimen (27 m·min-1, 15% elevation for 40 min. The exercise performance (sprint-test, contractile properties (m. tibialis anterior, oxidative enzyme activity (SDH, LDH, NADH2 in m. soleus and blood hematological and chemical variables were assessed in the groups at the end of the experiment. It was found out that creatine supplementation improved the exercise performance after 20 days of administration in a dose of 60 mg per day on the background of a mixed (aerobic-anaerobic exercise training. At the end of the trial the Cr-group demonstrated better values for the variables which characterize the contractile properties of m. tibialis anterior containing predominantly types IIA and IIB muscle fibers. On the other hand, a higher oxidative capacity was found out in m. soleus (type I muscle fibers as a result of 20-day creatine supplementation. No side effects of creatine monohydrate supplementation were assessed by the hematological and blood biochemical indices measured in this study

  16. On the importance of exchangeable NH protons in creatine for the magnetic coupling of creatine methyl protons in skeletal muscle

    NARCIS (Netherlands)

    Kruiskamp, M.J.; Nicolaij, K.

    2001-01-01

    The methyl protons of creatine in skeletal muscle exhibit a strong off-resonance magnetization transfer effect. The mechanism of this process is unknown. We previously hypothesized that the exchangeable amide/amino protons of creatine might be involved. To test this the characteristics of the

  17. The potent, indirect adenosine monophosphate-activated protein kinase activator R419 attenuates mitogen-activated protein kinase signaling, inhibits nociceptor excitability, and reduces pain hypersensitivity in mice

    Directory of Open Access Journals (Sweden)

    Galo L. Mejia

    2016-07-01

    Full Text Available Abstract. There is a great need for new therapeutics for the treatment of pain. A possible avenue to development of such therapeutics is to interfere with signaling pathways engaged in peripheral nociceptors that cause these neurons to become hyperexcitable. There is strong evidence that mitogen-activated protein kinases and phosphoinositide 3-kinase (PI3K/mechanistic target of rapamycin signaling pathways are key modulators of nociceptor excitability in vitro and in vivo. Activation of adenosine monophosphate-activated protein kinase (AMPK can inhibit signaling in both of these pathways, and AMPK activators have been shown to inhibit nociceptor excitability and pain hypersensitivity in rodents. R419 is one of, if not the most potent AMPK activator described to date. We tested whether R419 activates AMPK in dorsal root ganglion (DRG neurons and if this leads to decreased pain hypersensitivity in mice. We find that R419 activates AMPK in DRG neurons resulting in decreased mitogen-activated protein kinase signaling, decreased nascent protein synthesis, and enhanced P body formation. R419 attenuates nerve growth factor (NGF-induced changes in excitability in DRG neurons and blocks NGF-induced mechanical pain amplification in vivo. Moreover, locally applied R419 attenuates pain hypersensitivity in a model of postsurgical pain and blocks the development of hyperalgesic priming in response to both NGF and incision. We conclude that R419 is a promising lead candidate compound for the development of potent and specific AMPK activation to inhibit pain hypersensitivity as a result of injury.

  18. Effects of acute creatine supplementation on iron homeostasis and uric acid-based antioxidant capacity of plasma after wingate test

    Directory of Open Access Journals (Sweden)

    Barros Marcelo P

    2012-06-01

    Full Text Available Abstract Background Dietary creatine has been largely used as an ergogenic aid to improve strength and athletic performance, especially in short-term and high energy-demanding anaerobic exercise. Recent findings have also suggested a possible antioxidant role for creatine in muscle tissues during exercise. Here we evaluate the effects of a 1-week regimen of 20 g/day creatine supplementation on the plasma antioxidant capacity, free and heme iron content, and uric acid and lipid peroxidation levels of young subjects (23.1 ± 5.8 years old immediately before and 5 and 60 min after the exhaustive Wingate test. Results Maximum anaerobic power was improved by acute creatine supplementation (10.5 %, but it was accompanied by a 2.4-fold increase in pro-oxidant free iron ions in the plasma. However, potential iron-driven oxidative insult was adequately counterbalanced by proportional increases in antioxidant ferric-reducing activity in plasma (FRAP, leading to unaltered lipid peroxidation levels. Interestingly, the FRAP index, found to be highly dependent on uric acid levels in the placebo group, also had an additional contribution from other circulating metabolites in creatine-fed subjects. Conclusions Our data suggest that acute creatine supplementation improved the anaerobic performance of athletes and limited short-term oxidative insults, since creatine-induced iron overload was efficiently circumvented by acquired FRAP capacity attributed to: overproduction of uric acid in energy-depleted muscles (as an end-product of purine metabolism and a powerful iron chelating agent and inherent antioxidant activity of creatine.

  19. Prevalence of Creatine Deficiency Syndromes in Children With Nonsyndromic Autism.

    Science.gov (United States)

    Schulze, Andreas; Bauman, Margaret; Tsai, Anne Chun-Hui; Reynolds, Ann; Roberts, Wendy; Anagnostou, Evdokia; Cameron, Jessie; Nozzolillo, Alixandra A; Chen, Shiyi; Kyriakopoulou, Lianna; Scherer, Stephen W; Loh, Alvin

    2016-01-01

    Creatine deficiency may play a role in the neurobiology of autism and may represent a treatable cause of autism. The goal of the study was to ascertain the prevalence of creatine deficiency syndromes (CDSs) in children with autism spectrum disorder (ASD). In a prospective multicenter study, 443 children were investigated after a confirmed diagnosis of ASD. Random spot urine screening for creatine metabolites (creatine, guanidinoacetate, creatinine, and arginine) with liquid chromatography-tandem mass spectrometry and second-tier testing with high-performance liquid chromatography methodology was followed by recall testing in 24-hour urines and confirmatory testing by Sanger-based DNA sequencing of GAMT, GATM, and SLC6A8 genes. Additional diagnostic tests included plasma creatine metabolites and in vivo brain proton magnetic resonance spectroscopy. The creatine metabolites in spot urine in the autism group were compared with 128 healthy controls controlled for age. In 443 subjects with ASD investigated for CDS, we had 0 events (event: 0, 95% confidence interval 0-0.0068), therefore with 95% confidence the prevalence of CDS is creatine metabolites (P > .0125) in urine. Our study revealed a very low prevalence of CDS in children with nonsyndromic ASD and no obvious association between creatine metabolites and autism. Unlike our study population, we expect more frequent CDS among children with severe developmental delay, speech impairment, seizures, and movement disorders in addition to impairments in social communication, restricted interests, and repetitive behaviors. Copyright © 2016 by the American Academy of Pediatrics.

  20. Protein kinase activity associated with the corticosteroid binder IB

    International Nuclear Information System (INIS)

    Vujicic, M.; Djordjevic-Markovic, R.; Radic, O.; Krstic, M.; Kanazir, D.

    1997-01-01

    The physiological effects elicited by glucocorticoids are mediated via glucocorticoid receptors (GR). Analysis of specific glucocorticoid binding to radioactively labelled [ 3 H] triamcinolone acetonide in rat liver cytosol and analysis by ion exchange chromatography have revealed the presence of two distinct molecular species. The major form, designated as binder II appears to correspond to the well characterized glucocorticoid receptor by virtue of its size, charge, steroid binding characteristics and ability to bind to DNA.The second form, designated as corticosteroid binder IB, is a minor binding component in the liver. The binder IB differs from the binder II receptor by virtue of its lower molecular weight and its elution in the pre gradient of DEAE-Sephadex A-50 column which retains the un activated binder II receptor complexes. We examined the kinase activity of partially purified corticosteroid binder IB. Using (γ 3 2 P) ATP we detected kinase activity associated with the IB fraction from the rat liver. This kinase phosphorylate mixed histones and and dose not phosphorylate IB protein in vitro. The kinase activity is completely inhibited by the addition of Mg 2 + ions and is partially inhibited by the addition of Ca 2 +ions. (author)

  1. AMP-activated protein kinase induces actin cytoskeleton reorganization in epithelial cells

    International Nuclear Information System (INIS)

    Miranda, Lisa; Carpentier, Sarah; Platek, Anna; Hussain, Nusrat; Gueuning, Marie-Agnes; Vertommen, Didier; Ozkan, Yurda; Sid, Brice; Hue, Louis; Courtoy, Pierre J.; Rider, Mark H.; Horman, Sandrine

    2010-01-01

    AMP-activated protein kinase (AMPK), a known regulator of cellular and systemic energy balance, is now recognized to control cell division, cell polarity and cell migration, all of which depend on the actin cytoskeleton. Here we report the effects of A769662, a pharmacological activator of AMPK, on cytoskeletal organization and signalling in epithelial Madin-Darby canine kidney (MDCK) cells. We show that AMPK activation induced shortening or radiation of stress fibers, uncoupling from paxillin and predominance of cortical F-actin. In parallel, Rho-kinase downstream targets, namely myosin regulatory light chain and cofilin, were phosphorylated. These effects resembled the morphological changes in MDCK cells exposed to hyperosmotic shock, which led to Ca 2+ -dependent AMPK activation via calmodulin-dependent protein kinase kinase-β(CaMKKβ), a known upstream kinase of AMPK. Indeed, hypertonicity-induced AMPK activation was markedly reduced by the STO-609 CaMKKβ inhibitor, as was the increase in MLC and cofilin phosphorylation. We suggest that AMPK links osmotic stress to the reorganization of the actin cytoskeleton.

  2. Partial purification and characterization of a wortmannin-sensitive and insulin-stimulated protein kinase that activates heart 6-phosphofructo-2-kinase.

    OpenAIRE

    Deprez, J; Bertrand, L; Alessi, D R; Krause, U; Hue, L; Rider, M H

    2000-01-01

    A wortmannin-sensitive and insulin-stimulated protein kinase (WISK), which phosphorylates and activates cardiac 6-phosphofructo-2-kinase (PFK-2), was partially purified from perfused rat hearts. Immunoblotting showed that WISK was devoid of protein kinase B (PKB), serum- and glucocorticoid-regulated protein kinase and protein kinase Czeta (PKCzeta). Comparison of the inhibition of WISK, PKCalpha and PKCzeta by different protein kinase inhibitors suggested that WISK was not a member of the PKC...

  3. Src-family-tyrosine kinase Lyn is critical for TLR2-mediated NF-κB activation through the PI 3-kinase signaling pathway.

    Science.gov (United States)

    Toubiana, Julie; Rossi, Anne-Lise; Belaidouni, Nadia; Grimaldi, David; Pene, Frederic; Chafey, Philippe; Comba, Béatrice; Camoin, Luc; Bismuth, Georges; Claessens, Yann-Erick; Mira, Jean-Paul; Chiche, Jean-Daniel

    2015-10-01

    TLR2 has a prominent role in host defense against a wide variety of pathogens. Stimulation of TLR2 triggers MyD88-dependent signaling to induce NF-κB translocation, and activates a Rac1-PI 3-kinase dependent pathway that leads to transactivation of NF-κB through phosphorylation of the P65 NF-κB subunit. This transactivation pathway involves tyrosine phosphorylations. The role of the tyrosine kinases in TLR signaling is controversial, with discrepancies between studies using only chemical inhibitors and knockout mice. Here, we show the involvement of the tyrosine-kinase Lyn in TLR2-dependent activation of NF-κB in human cellular models, by using complementary inhibition strategies. Stimulation of TLR2 induces the formation of an activation cluster involving TLR2, CD14, PI 3-kinase and Lyn, and leads to the activation of AKT. Lyn-dependent phosphorylation of the p110 catalytic subunit of PI 3-kinase is essential to the control of PI 3-kinase biological activity upstream of AKT and thereby to the transactivation of NF-κB. Thus, Lyn kinase activity is crucial in TLR2-mediated activation of the innate immune response in human mononuclear cells. © The Author(s) 2015.

  4. Fluvoxamine alters the activity of energy metabolism enzymes in the brain

    Directory of Open Access Journals (Sweden)

    Gabriela K. Ferreira

    2014-09-01

    Full Text Available Objective: Several studies support the hypothesis that metabolism impairment is involved in the pathophysiology of depression and that some antidepressants act by modulating brain energy metabolism. Thus, we evaluated the activity of Krebs cycle enzymes, the mitochondrial respiratory chain, and creatine kinase in the brain of rats subjected to prolonged administration of fluvoxamine. Methods: Wistar rats received daily administration of fluvoxamine in saline (10, 30, and 60 mg/kg for 14 days. Twelve hours after the last administration, rats were killed by decapitation and the prefrontal cortex, cerebral cortex, hippocampus, striatum, and cerebellum were rapidly isolated. Results: The activities of citrate synthase, malate dehydrogenase, and complexes I, II-III, and IV were decreased after prolonged administration of fluvoxamine in rats. However, the activities of complex II, succinate dehydrogenase, and creatine kinase were increased. Conclusions: Alterations in activity of energy metabolism enzymes were observed in most brain areas analyzed. Thus, we suggest that the decrease in citrate synthase, malate dehydrogenase, and complexes I, II-III, and IV can be related to adverse effects of pharmacotherapy, but long-term molecular adaptations cannot be ruled out. In addition, we demonstrated that these changes varied according to brain structure or biochemical analysis and were not dose-dependent.

  5. Protective Effects of Kaempferol against Myocardial Ischemia/Reperfusion Injury in Isolated Rat Heart via Antioxidant Activity and Inhibition of Glycogen Synthase Kinase-3β

    Science.gov (United States)

    Zhou, Mingjie; Ren, Huanhuan; Wang, Wenjuan; Zheng, Qiusheng; Wang, Dong

    2015-01-01

    Objective. This study aimed to evaluate the protective effect of kaempferol against myocardial ischemia/reperfusion (I/R) injury in rats. Method. Left ventricular developed pressure (LVDP) and its maximum up/down rate (±dp/dt max) were recorded as myocardial function. Infarct size was detected with 2,3,5-triphenyltetrazolium chloride staining. Cardiomyocyte apoptosis was determined using terminal deoxynucleotidyl nick-end labeling (TUNEL). The levels of creatine kinase (CK), lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione/glutathione disulfide (GSH/GSSG) ratio, and tumor necrosis factor-alpha (TNF-α) were determined using enzyme linked immunosorbent assay (ELISA). Moreover, total glycogen synthase kinase-3β (GSK-3β), phospho-GSK-3β (P-GSK-3β), precaspase-3, cleaved caspase-3, and cytoplasm cytochrome C were assayed using Western blot analysis. Results. Pretreatment with kaempferol significantly improved the recovery of LVDP and ±dp/dt max, as well as increased the levels of SOD and P-GSK-3β and GSH/GSSG ratio. However, the pretreatment reduced myocardial infarct size and TUNEL-positive cell rate, as well as decreased the levels of cleaved caspase-3, cytoplasm cytochrome C, CK, LDH, MDA, and TNF-α. Conclusion. These results suggested that kaempferol provides cardioprotection via antioxidant activity and inhibition of GSK-3β activity in rats with I/R. PMID:26265983

  6. Creatine Protects against Excitoxicity in an In Vitro Model of Neurodegeneration

    Science.gov (United States)

    Genius, Just; Geiger, Johanna; Bender, Andreas; Möller, Hans-Jürgen; Klopstock, Thomas; Rujescu, Dan

    2012-01-01

    Creatine has been shown to be neuroprotective in aging, neurodegenerative conditions and brain injury. As a common molecular background, oxidative stress and disturbed cellular energy homeostasis are key aspects in these conditions. Moreover, in a recent report we could demonstrate a life-enhancing and health-promoting potential of creatine in rodents, mainly due to its neuroprotective action. In order to investigate the underlying pharmacology mediating these mainly neuroprotective properties of creatine, cultured primary embryonal hippocampal and cortical cells were challenged with glutamate or H2O2. In good agreement with our in vivo data, creatine mediated a direct effect on the bioenergetic balance, leading to an enhanced cellular energy charge, thereby acting as a neuroprotectant. Moreover, creatine effectively antagonized the H2O2-induced ATP depletion and the excitotoxic response towards glutamate, while not directly acting as an antioxidant. Additionally, creatine mediated a direct inhibitory action on the NMDA receptor-mediated calcium response, which initiates the excitotoxic cascade. Even excessive concentrations of creatine had no neurotoxic effects, so that high-dose creatine supplementation as a health-promoting agent in specific pathological situations or as a primary prophylactic compound in risk populations seems feasible. In conclusion, we were able to demonstrate that the protective potential of creatine was primarily mediated by its impact on cellular energy metabolism and NMDA receptor function, along with reduced glutamate spillover, oxidative stress and subsequent excitotoxicity. PMID:22347384

  7. Creatine-induced activation of antioxidative defence in myotube cultures revealed by explorative NMR-based metabonomics and proteomics

    Directory of Open Access Journals (Sweden)

    Nielsen Niels

    2010-02-01

    Full Text Available Abstract Background Creatine is a key intermediate in energy metabolism and supplementation of creatine has been used for increasing muscle mass, strength and endurance. Creatine supplementation has also been reported to trigger the skeletal muscle expression of insulin like growth factor I, to increase the fat-free mass and improve cognition in elderly, and more explorative approaches like transcriptomics has revealed additional information. The aim of the present study was to reveal additional insight into the biochemical effects of creatine supplementation at the protein and metabolite level by integrating the explorative techniques, proteomics and NMR metabonomics, in a systems biology approach. Methods Differentiated mouse myotube cultures (C2C12 were exposed to 5 mM creatine monohydrate (CMH for 24 hours. For proteomics studies, lysed myotubes were analyzed in single 2-DGE gels where the first dimension of protein separation was pI 5-8 and second dimension was a 12.5% Criterion gel. Differentially expressed protein spots of significance were excised from the gel, desalted and identified by peptide mass fingerprinting using MALDI-TOF MS. For NMR metabonomic studies, chloroform/methanol extractions of the myotubes were subjected to one-dimensional 1H NMR spectroscopy and the intracellular oxidative status of myotubes was assessed by intracellular DCFH2 oxidation after 24 h pre-incubation with CMH. Results The identified differentially expressed proteins included vimentin, malate dehydrogenase, peroxiredoxin, thioredoxin dependent peroxide reductase, and 75 kDa and 78 kDa glucose regulated protein precursors. After CMH exposure, up-regulated proteomic spots correlated positively with the NMR signals from creatine, while down-regulated proteomic spots were negatively correlated with these NMR signals. The identified differentially regulated proteins were related to energy metabolism, glucose regulated stress, cellular structure and the

  8. The phosphatidylinositol 3-kinase inhibitor, wortmannin, inhibits insulin-induced activation of phosphatidylcholine hydrolysis and associated protein kinase C translocation in rat adipocytes.

    Science.gov (United States)

    Standaert, M L; Avignon, A; Yamada, K; Bandyopadhyay, G; Farese, R V

    1996-02-01

    We questioned whether phosphatidylinositol 3-kinase (PI 3-kinase) and protein kinase C (PKC) function as interrelated signalling mechanisms during insulin action in rat adipocytes. Insulin rapidly activated a phospholipase D that hydrolyses phosphatidylcholine (PC), and this activation was accompanied by increases in diacylglycerol and translocative activation of PKC-alpha and PKC-beta in the plasma membrane. Wortmannin, an apparently specific PI 3-kinase inhibitor, inhibited insulin-stimulated, phospholipase D-dependent PC hydrolysis and subsequent translocation of PKC-alpha and PKC-beta to the plasma membrane. Wortmannin did not inhibit PKC directly in vitro, or the PKC-dependent effects of phorbol esters on glucose transport in intact adipocytes. The PKC inhibitor RO 31-8220 did not inhibit PI 3-kinase directly or its activation in situ by insulin, but inhibited both insulin-stimulated and phorbol ester-stimulated glucose transport. Our findings suggest that insulin acts through PI 3-kinase to activate a PC-specific phospholipase D and causes the translocative activation of PKC-alpha and PKC-beta in plasma membranes of rat adipocytes.

  9. Thymidine uptake, thymidine incorporation, and thymidine kinase activity in marine bacterium isolates

    International Nuclear Information System (INIS)

    Jeffrey, W.H.; Paul, J.H.

    1990-01-01

    One assumption made in bacterial production estimates from [ 3 H]thymidine incorporation is that all heterotrophic bacteria can incorporate exogenous thymidine into DNA. Heterotrophic marine bacterium isolates from Tampa Bay, Fla., Chesapeake Bay, Md., and a coral surface microlayer were examined for thymidine uptake (transport), thymidine incorporation, the presence of thymidine kinase genes, and thymidine kinase enzyme activity. Of the 41 isolates tested, 37 were capable of thymidine incorporation into DNA. The four organisms that could not incorporate thymidine also transported the thymidine poorly and lacked thymidine kinase activity. Attempts to detect thymidine kinase genes in the marine isolates by molecular probing with gene probes made from Escherichia coli and herpes simplex virus thymidine kinase genes proved unsuccessful. To determine if the inability to incorporate thymidine was due to the lack of thymidine kinase, one organism, Vibro sp. strain DI9, was transformed with a plasmid (pGQ3) that contained an E. coli thymidine kinase gene. Although enzyme assays indicated high levels of thymidine kinase activity in transformants, these cells still failed to incorporate exogenous thymidine into DNA or to transport thymidine into cells. These results indicate that the inability of certain marine bacteria to incorporate thymidine may not be solely due to the lack of thymidine kinase activity but may also be due to the absence of thymidine transport systems

  10. Creatine co-ingestion with carbohydrate or cinnamon extract provides no added benefit to anaerobic performance.

    Science.gov (United States)

    Islam, Hashim; Yorgason, Nick J; Hazell, Tom J

    2016-09-01

    The insulin response following carbohydrate ingestion enhances creatine transport into muscle. Cinnamon extract is promoted to have insulin-like effects, therefore this study examined if creatine co-ingestion with carbohydrates or cinnamon extract improved anaerobic capacity, muscular strength, and muscular endurance. Active young males (n = 25; 23.7 ± 2.5 y) were stratified into 3 groups: (1) creatine only (CRE); (2) creatine+ 70 g carbohydrate (CHO); or (3) creatine+ 500 mg cinnamon extract (CIN), based on anaerobic capacity (peak power·kg(-1)) and muscular strength at baseline. Three weeks of supplementation consisted of a 5 d loading phase (20 g/d) and a 16 d maintenance phase (5 g/d). Pre- and post-supplementation measures included a 30-s Wingate and a 30-s maximal running test (on a self-propelled treadmill) for anaerobic capacity. Muscular strength was measured as the one-repetition maximum 1-RM for chest, back, quadriceps, hamstrings, and leg press. Additional sets of the number of repetitions performed at 60% 1-RM until fatigue measured muscular endurance. All three groups significantly improved Wingate relative peak power (CRE: 15.4% P = .004; CHO: 14.6% P = .004; CIN: 15.7%, P = .003), and muscular strength for chest (CRE: 6.6% P creatine ingestion lead to similar changes in anaerobic power, strength, and endurance.

  11. Role of nongenomic activation of phosphatidylinositol 3-kinase/Akt and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase/extracellular signal-regulated kinase 1/2 pathways in 1,25D3-mediated apoptosis in squamous cell carcinoma cells.

    Science.gov (United States)

    Ma, Yingyu; Yu, Wei-Dong; Kong, Rui-Xian; Trump, Donald L; Johnson, Candace S

    2006-08-15

    Vitamin D is a steroid hormone that regulates calcium homeostasis and bone metabolism. The active form of vitamin D [1 alpha,25-dihydroxyvitamin D(3) (1,25D3)] acts through both genomic and nongenomic pathways. 1,25D3 has antitumor effects in a variety of cancers, including colorectal, prostate, breast, ovarian, and skin cancers. 1,25D3 exerts growth-inhibitory effects in cancer cells through the induction of apoptosis, cell cycle arrest, and differentiation. The mechanisms regulating 1,25D3-induced apoptosis remain unclear. We investigated the role of nongenomic signaling in 1,25D3-mediated apoptosis in squamous cell carcinoma (SCC) cells. 1,25D3 induced rapid and sustained activation of phosphatidylinositol 3-kinase/Akt and mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) 1/2 pathways in SCC cells. These effects were nongenomic: they occurred rapidly and were not inhibited by cycloheximide or actinomycin D. To examine whether the nongenomic activation of Akt and ERK1/2 plays a role in 1,25D3-mediated apoptosis, the expression of Akt or ERK1/2 was reduced by small interfering RNA (siRNA). siRNA-Akt significantly enhanced 1,25D3-induced apoptosis as indicated by increased levels of Annexin V-positive cells and increased sub-G(1) population and DNA fragmentation. In contrast, siRNA-ERK1/2 had no effects on 1,25D3-induced apoptosis. In addition, siRNA-Akt transfection followed by 1,25D3 treatment induced apoptosis much sooner than 1,25D3 alone. siRNA-Akt and 1,25D3 induced caspase-10 activation, suppressed the expression of c-IAP1 and XIAP, and promoted 1,25D3-induced caspase-3 activation. These results support a link between 1,25D3-induced nongenomic signaling and apoptosis. 1,25D3 induces the activation of phosphatidylinositol 3-kinase/Akt, which suppresses 1,25D3-mediated apoptosis and prolongs the survival of SCC cells.

  12. Ghrelin augments murine T-cell proliferation by activation of the phosphatidylinositol-3-kinase, extracellular signal-regulated kinase and protein kinase C signaling pathways

    Science.gov (United States)

    Lee, Jun Ho; Patel, Kalpesh; Tae, Hyun Jin; Lustig, Ana; Kim, Jie Wan; Mattson, Mark P.; Taub, Dennis D.

    2014-01-01

    Thymic atrophy occurs during normal aging, and is accelerated by exposure to chronic stressors that elevate glucocorticoid levelsand impair the naïve T cell output. The orexigenic hormone ghrelin was recently shown to attenuate age-associated thymic atrophy. Here, we report that ghrelin enhances the proliferation of murine CD4+ primary T cells and a CD4+ T-cell line. Ghrelin induced activation of the ERK1/2 and Akt signaling pathways, via upstream activation of phosphatidylinositol-3-kinase and protein kinase C, to enhance T-cell proliferation. Moreover, ghrelin induced expression of the cell cycle proteins cyclin D1, cyclin E, cyclin-dependent kinase 2 (CDK2) and retinoblastoma phosphorylation. Finally, ghrelin activated the above-mentioned signaling pathways and stimulated thymocyte proliferation in young and older mice in vivo. PMID:25447526

  13. Creatine Transporter Deficiency in Two Brothers with Autism Spectrum Disorder.

    Science.gov (United States)

    Aydin, Halil Ibrahim

    2018-01-15

    Creatine transporter deficiency (CTD) is a treatable, X-linked, inborn error of metabolism. Two brothers with autism spectrum disorder were diagnosed with CTD at the ages of 17 and 12 years. Both were found to have a previously reported hemizygous p.408delF (c.1216_1218delTTC) deletion mutation. Both patients were given creatine monohydrate, L-arginine, L-glycine and S-adenosylmethionine, which partially improved the behavioral problems. Serum creatinine levels, creatine peak at brain MR spectroscopy or creatine/creatinine ratio in urine should be evaluated to identify CTD in children with autistic behavior and language disorders.

  14. Cell-type-specific activation of mitogen-activated protein kinases in PAN-induced progressive renal disease in rats

    International Nuclear Information System (INIS)

    Park, Sang-Joon; Jeong, Kyu-Shik

    2004-01-01

    We examined the time-course activation and the cell-type specific role of MAP kinases in puromycin aminonucleoside (PAN)-induced renal disease. The maximal activation of c-Jun-NH 2 -terminal kinase (JNK), extracellular signal regulated kinase (ERK), and p38 MAP kinase was detected on Days 52, 38, and 38 after PAN-treatment, respectively. p-JNK was localized in mesangial and proximal tubular cells at the early renal injury. It was expressed, therefore, in the inflammatory cells of tubulointerstitial lesions. While, p-ERK was markedly increased in the glomerular regions and macrophages p-p38 was observed in glomerular endothelial cells, tubular cells, and some inflammatory cells. The results show that the activation of MAP kinases in the early renal injury by PAN-treatment involves cellular changes such as cell proliferation or apoptosis in renal native cells. The activation of MAP kinases in infiltrated inflammatory cells and fibrotic cells plays an important role in destructive events such as glomerulosclerosis and tubulointerstitial fibrosis

  15. SH2 domains: modulators of nonreceptor tyrosine kinase activity

    OpenAIRE

    Filippakopoulos, Panagis; Müller, Susanne; Knapp, Stefan

    2009-01-01

    The Src homology 2 (SH2) domain is a sequence-specific phosphotyrosine-binding module present in many signaling molecules. In cytoplasmic tyrosine kinases, the SH2 domain is located N-terminally to the catalytic kinase domain (SH1) where it mediates cellular localization, substrate recruitment, and regulation of kinase activity. Initially, structural studies established a role of the SH2 domain stabilizing the inactive state of Src family members. However, biochemical characterization showed ...

  16. Subunits of the Snf1 kinase heterotrimer show interdependence for association and activity.

    Science.gov (United States)

    Elbing, Karin; Rubenstein, Eric M; McCartney, Rhonda R; Schmidt, Martin C

    2006-09-08

    The Snf1 kinase and its mammalian orthologue, the AMP-activated protein kinase (AMPK), function as heterotrimers composed of a catalytic alpha-subunit and two non-catalytic subunits, beta and gamma. The beta-subunit is thought to hold the complex together and control subcellular localization whereas the gamma-subunit plays a regulatory role by binding to and blocking the function of an auto-inhibitory domain (AID) present in the alpha-subunit. In addition, catalytic activity requires phosphorylation by a distinct upstream kinase. In yeast, any one of three Snf1-activating kinases, Sak1, Tos3, or Elm1, can fulfill this role. We have previously shown that Sak1 is the only Snf1-activating kinase that forms a stable complex with Snf1. Here we show that the formation of the Sak1.Snf1 complex requires the beta- and gamma-subunits in vivo. However, formation of the Sak1.Snf1 complex is not necessary for glucose-regulated phosphorylation of the Snf1 activation loop. Snf1 kinase purified from cells lacking the beta-subunits do not contain any gamma-subunit, indicating that the Snf1 kinase does not form a stable alphagamma dimer in vivo. In vitro kinase assays using purified full-length and truncated Snf1 proteins demonstrate that the kinase domain, which lacks the AID, is significantly more active than the full-length Snf1 protein. Addition of purified beta- and gamma-subunits could stimulate the kinase activity of the full-length alpha-subunit but only when all three subunits were present, suggesting an interdependence of all three subunits for assembly of a functional complex.

  17. Role of Creatine Supplementation on Exercise-Induced Cardiovascular Function and Oxidative Stress

    Directory of Open Access Journals (Sweden)

    Michael I. C. Kingsley

    2009-01-01

    Full Text Available Many degenerative diseases are associated with increased oxidative stress. Creatine has the potential to act as an indirect and direct antioxidant; however, limited data exist to evaluate the antioxidant capabilities of creatine supplementation within in vivo human systems. This study aimed to investigate the effects of oral creatine supplementation on markers of oxidative stress and antioxidant defenses following exhaustive cycling exercise. Following preliminary testing and two additional familiarization sessions, 18 active males repeated two exhaustive incremental cycling trials (T1 and T2 separated by exactly 7 days. The subjects were assigned, in a double-blind manner, to receive either 20 g of creatine (Cr or a placebo (P for the 5 days preceding T2. Breath-by-breath respiratory data and heart rate were continually recorded throughout the exercise protocol and blood samples were obtained at rest (preexercise, at the end of exercise (postexercise, and the day following exercise (post24 h. Serum hypdroperoxide concentrations were elevated at postexercise by 17 ± 5% above preexercise values (p = 0.030. However, supplementation did not influence lipid peroxidation (serum hypdroperoxide concentrations, resistance of low density lipoprotein to oxidative stress (t1/2max LDL oxidation and plasma concentrations of non-enzymatic antioxidants (retinol, α-carotene, β-carotene, α-tocopherol, γ-tocopherol, lycopene and vitamin C. Heart rate and oxygen uptake responses to exercise were not affected by supplementation. These findings suggest that short-term creatine supplementation does not enhance non-enzymatic antioxidant defence or protect against lipid peroxidation induced by exhaustive cycling in healthy males.

  18. Autoinhibition of Bruton's tyrosine kinase (Btk) and activation by soluble inositol hexakisphosphate

    Science.gov (United States)

    Wang, Qi; Vogan, Erik M; Nocka, Laura M; Rosen, Connor E; Zorn, Julie A; Harrison, Stephen C; Kuriyan, John

    2015-01-01

    Bruton's tyrosine kinase (Btk), a Tec-family tyrosine kinase, is essential for B-cell function. We present crystallographic and biochemical analyses of Btk, which together reveal molecular details of its autoinhibition and activation. Autoinhibited Btk adopts a compact conformation like that of inactive c-Src and c-Abl. A lipid-binding PH-TH module, unique to Tec kinases, acts in conjunction with the SH2 and SH3 domains to stabilize the inactive conformation. In addition to the expected activation of Btk by membranes containing phosphatidylinositol triphosphate (PIP3), we found that inositol hexakisphosphate (IP6), a soluble signaling molecule found in both animal and plant cells, also activates Btk. This activation is a consequence of a transient PH-TH dimerization induced by IP6, which promotes transphosphorylation of the kinase domains. Sequence comparisons with other Tec-family kinases suggest that activation by IP6 is unique to Btk. DOI: http://dx.doi.org/10.7554/eLife.06074.001 PMID:25699547

  19. Fenproporex increases locomotor activity and alters energy metabolism, and mood stabilizers reverse these changes: a proposal for a new animal model of mania.

    Science.gov (United States)

    Rezin, Gislaine T; Furlanetto, Camila B; Scaini, Giselli; Valvassori, Samira S; Gonçalves, Cinara L; Ferreira, Gabriela K; Jeremias, Isabela C; Resende, Wilson R; Cardoso, Mariane R; Varela, Roger B; Quevedo, João; Streck, Emilio L

    2014-04-01

    Fenproporex (Fen) is converted in vivo into amphetamine, which is used to induce mania-like behaviors in animals. In the present study, we intend to present a new animal model of mania. In order to prove through face, construct, and predictive validities, we evaluated behavioral parameters (locomotor activity, stereotypy activity, and fecal boli amount) and brain energy metabolism (enzymes citrate synthase; malate dehydrogenase; succinate dehydrogenase; complexes I, II, II-III, and IV of the mitochondrial respiratory chain; and creatine kinase) in rats submitted to acute and chronic administration of fenproporex, treated with lithium (Li) and valproate (VPA). The administration of Fen increased locomotor activity and decreased the activity of Krebs cycle enzymes, mitochondrial respiratory chain complexes, and creatine kinase, in most brain structures evaluated. In addition, treatment with mood stabilizers prevented and reversed this effect. Our results are consistent with the literature that demonstrates behavioral changes and mitochondrial dysfunction caused by psychostimulants. These findings suggest that chronic administration of Fen may be a potential animal model of mania.

  20. Interactions of Aging, Overload, and Creatine Supplementation in Rat Plantaris Muscle

    Directory of Open Access Journals (Sweden)

    Mark D. Schuenke

    2011-01-01

    Full Text Available Attenuation of age-related sarcopenia by creatine supplementation has been equivocal. In this study, plantaris muscles of young (Y; 5m and aging (A; 24m Fisher 344 rats underwent four weeks of either control (C, creatine supplementation (Cr, surgical overload (O, or overload plus creatine (OCr. Creatine alone had no effect on muscle fiber cross-sectional area (CSA or heat shock protein (HSP70 and increased myonuclear domain (MND only in young rats. Overload increased CSA and HSP70 content in I and IIA fibers, regardless of age, and MND in IIA fibers of YO rats. CSA and MND increased in all fast fibers of YOCr, and CSA increased in I and IIA fibers of AOCr. OCR did not alter HSP70, regardless of age. MND did not change in aging rats, regardless of treatment. These data indicate creatine alone had no significant effect. Creatine with overload produced no additional hypertrophy relative to overload alone and attenuated overload-induced HSP70 expression.

  1. Disruption of the LOV-Jalpha helix interaction activates phototropin kinase activity.

    Science.gov (United States)

    Harper, Shannon M; Christie, John M; Gardner, Kevin H

    2004-12-28

    Light plays a crucial role in activating phototropins, a class of plant photoreceptors that are sensitive to blue and UV-A wavelengths. Previous studies indicated that phototropin uses a bound flavin mononucleotide (FMN) within its light-oxygen-voltage (LOV) domain to generate a protein-flavin covalent bond under illumination. In the C-terminal LOV2 domain of Avena sativa phototropin 1, formation of this bond triggers a conformational change that results in unfolding of a helix external to this domain called Jalpha [Harper, S. M., et al. (2003) Science 301, 1541-1545]. Though the structural effects of illumination were characterized, it was unknown how these changes are coupled to kinase activation. To examine this, we made a series of point mutations along the Jalpha helix to disrupt its interaction with the LOV domain in a manner analogous to light activation. Using NMR spectroscopy and limited proteolysis, we demonstrate that several of these mutations displace the Jalpha helix from the LOV domain independently of illumination. When placed into the full-length phototropin protein, these point mutations display constitutive kinase activation, without illumination of the sample. These results indicate that unfolding of the Jalpha helix is the critical event in regulation of kinase signaling for the phototropin proteins.

  2. Regulatory crosstalk by protein kinases on CFTR trafficking and activity

    Science.gov (United States)

    Farinha, Carlos Miguel; Swiatecka-Urban, Agnieszka; Brautigan, David; Jordan, Peter

    2016-01-01

    Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a member of the ATP binding cassette (ABC) transporter superfamily that functions as a cAMP-activated chloride ion channel in fluid-transporting epithelia. There is abundant evidence that CFTR activity (i.e. channel opening and closing) is regulated by protein kinases and phosphatases via phosphorylation and dephosphorylation. Here, we review recent evidence for the role of protein kinases in regulation of CFTR delivery to and retention in the plasma membrane. We review this information in a broader context of regulation of other transporters by protein kinases because the overall functional output of transporters involves the integrated control of both their number at the plasma membrane and their specific activity. While many details of the regulation of intracellular distribution of CFTR and other transporters remain to be elucidated, we hope that this review will motivate research providing new insights into how protein kinases control membrane transport to impact health and disease.

  3. Expression, purification and kinase activity analysis of maize ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-07-06

    Jul 6, 2009 ... Kinase activity is essential for a protein kinase to perform its biological function. In previous study we have cloned a novel plant SnRK2 subfamily gene from maize and named it as ZmSPK1. In this study the. cDNA of ZmSPK1 with dHA-His6 tag was amplified by PCR and was subcloned into the yeast.

  4. Human adenylate kinases – classification, structure, physiological and pathological importance

    Directory of Open Access Journals (Sweden)

    Magdalena Wujak

    2015-01-01

    Full Text Available Adenylate kinase (AK, EC 2.7.4.3 is a ubiquitous phosphotransferase which catalyzes the reversible transfer of high-energy β – and γ-phosphate groups between nucleotides. All classified AKs show a similar structure: they contain a large central CORE region, nucleoside monophosphate and triphosphate binding domains (NMPbd and NTPbd and the LID domain. Analysis of amino acid sequence similarity revealed the presence of as many as nine human AK isoenzymes, which demonstrate different organ-tissue and intercellular localization. Among these kinases, only two, AK1 and AK2, fulfill the structural and functional criterion by the highest affinity for adenine nucleotides and the utilization of only AMP or dAMP as phosphate acceptors. Human AK isoenzymes are involved in nucleotide homeostasis and monitor disturbances of cell energy charge. Participating in large regulatory protein complexes, AK supplies high energy substrates for controlling the functions of channels and transporters as well as ligands for extracellular P2 nucleotide receptors. In pathological conditions AK can take over the function of other kinases, such as creatine kinase in oxygen-depleted myocardium. Directed mutagenesis and genetic studies of diseases (such as aleukocytosis, hemolytic anemia, primary ciliary dyskinesia (PCD link the presence and activity of AK with etiology of these disturbances. Moreover, AK participates in regulation of differentiation and maturation of cells as well as in apoptosis and oncogenesis. Involvement of AK in a wide range of processes and the correlation between AK and etiology of diseases support the medical potential for the use of adenylate kinases in the diagnosis and treatment of certain diseases. This paper summarizes the current knowledge on the structure, properties and functions of human adenylate kinase.

  5. Src kinase conformational activation: thermodynamics, pathways, and mechanisms.

    Directory of Open Access Journals (Sweden)

    Sichun Yang

    2008-03-01

    Full Text Available Tyrosine kinases of the Src-family are large allosteric enzymes that play a key role in cellular signaling. Conversion of the kinase from an inactive to an active state is accompanied by substantial structural changes. Here, we construct a coarse-grained model of the catalytic domain incorporating experimental structures for the two stable states, and simulate the dynamics of conformational transitions in kinase activation. We explore the transition energy landscapes by constructing a structural network among clusters of conformations from the simulations. From the structural network, two major ensembles of pathways for the activation are identified. In the first transition pathway, we find a coordinated switching mechanism of interactions among the alphaC helix, the activation-loop, and the beta strands in the N-lobe of the catalytic domain. In a second pathway, the conformational change is coupled to a partial unfolding of the N-lobe region of the catalytic domain. We also characterize the switching mechanism for the alphaC helix and the activation-loop in detail. Finally, we test the performance of a Markov model and its ability to account for the structural kinetics in the context of Src conformational changes. Taken together, these results provide a broad framework for understanding the main features of the conformational transition taking place upon Src activation.

  6. High quality, small molecule-activity datasets for kinase research [version 3; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Rajan Sharma

    2016-10-01

    Full Text Available Kinases regulate cell growth, movement, and death. Deregulated kinase activity is a frequent cause of disease. The therapeutic potential of kinase inhibitors has led to large amounts of published structure activity relationship (SAR data. Bioactivity databases such as the Kinase Knowledgebase (KKB, WOMBAT, GOSTAR, and ChEMBL provide researchers with quantitative data characterizing the activity of compounds across many biological assays. The KKB, for example, contains over 1.8M kinase structure-activity data points reported in peer-reviewed journals and patents. In the spirit of fostering methods development and validation worldwide, we have extracted and have made available from the KKB 258K structure activity data points and 76K associated unique chemical structures across eight kinase targets. These data are freely available for download within this data note.

  7. Cytogenetical Effect of Creatine Monohydrate in Vicia faba Root Tips

    International Nuclear Information System (INIS)

    Ali, A.A.M.; El-zahrani, N.H.; El-shamrani, S.M.

    2010-01-01

    The present study has been conducted to evaluate the creatine effect on the cellular behavior at mitosis of Vicia faba using four concentrations (1.50, 2, 2.50 and 3 g/ 100 ml) with three exposure times (6, 12, 24 hour). Marked reduction of mitotic index was recorded at all creatine treatments and this trait was affected by creatine concentration and exposure time. Unbalanced mitotic stages percentages were observed after all treatments whereas, prophase % was decreased in all treatments but the opposite was true for metaphase %. While, (ana-telo) phases % were either increased or decreased after creatine treatments. Alteration of DNA or RNA contents, were obtained at different treatments. On the other hand, abnormalities were shown at all treatments with an increase percentage by increasing creatine concentration and exposure time. The most common of these abnormalities were: stickiness, disturbed and C metaphase. In addition, laggards, multipolor, and bridges were observed in some treatments but with low percentage

  8. AcEST: BP912187 [AcEST

    Lifescience Database Archive (English)

    Full Text Available 9 Definition sp|P70079|KCRU_CHICK Creatine kinase, ubiquitous mitochondrial OS=Ga...gnments: (bits) Value sp|P70079|KCRU_CHICK Creatine kinase, ubiquitous mitochondr...ial O... 34 0.33 sp|Q9TTK8|KCRU_BOVIN Creatine kinase, ubiquitous mitochondrial O... 33 0.74 sp|P46430|GSTT1...NH_SHESA Ribonuclease H OS=Shewanella sp. (strain ANA... 32 1.7 sp|P12532|KCRU_HUMAN Creatine kinase, ubiqui...tous mitochondrial O... 31 2.2 sp|P25809|KCRU_RAT Creatine kinase, ubiquitous mitochondrial OS=... 31 2.8 sp|P30275|KCRU_MOUSE Creati

  9. Cellular reprogramming through mitogen-activated protein kinases

    Directory of Open Access Journals (Sweden)

    Justin eLee

    2015-10-01

    Full Text Available Mitogen-activated protein kinase (MAPK cascades are conserved eukaryote signaling modules where MAPKs, as the final kinases in the cascade, phosphorylate protein substrates to regulate cellular processes. While some progress in the identification of MAPK substrates has been made in plants, the knowledge on the spectrum of substrates and their mechanistic action is still fragmentary. In this focused review, we discuss the biological implications of the data in our original paper (Sustained mitogen-activated protein kinase activation reprograms defense metabolism and phosphoprotein profile in Arabidopsis thaliana; Frontiers in Plant Science 5: 554 in the context of related research. In our work, we mimicked in vivo activation of two stress-activated MAPKs, MPK3 and MPK6, through transgenic manipulation of Arabidopsis thaliana and used phosphoproteomics analysis to identify potential novel MAPK substrates. Here, we plotted the identified putative MAPK substrates (and downstream phosphoproteins as a global protein clustering network. Based on a highly stringent selection confidence level, the core networks highlighted a MAPK-induced cellular reprogramming at multiple levels of gene and protein expression – including transcriptional, post-transcriptional, translational, post-translational (such as protein modification, folding and degradation steps, and also protein re-compartmentalization. Additionally, the increase in putative substrates/phosphoproteins of energy metabolism and various secondary metabolite biosynthesis pathways coincides with the observed accumulation of defense antimicrobial substances as detected by metabolome analysis. Furthermore, detection of protein networks in phospholipid or redox elements suggests activation of downstream signaling events. Taken in context with other studies, MAPKs are key regulators that reprogram cellular events to orchestrate defense signaling in eukaryotes.

  10. Kinase activity determination of specific AMPK complexes/heterotrimers in the skeletal muscle

    DEFF Research Database (Denmark)

    Birk, Jesper Bratz; Wojtaszewski, Jørgen

    2018-01-01

    Measuring the kinase activity of the 5'-AMP-activated protein kinase (AMPK) is an essential part of understanding the regulation of this metabolic master switch. The AMPK heterotrimer can exist in 12 different constellations with potentially diverse activation patterns. It is therefore important ...

  11. Severe falciparum malaria with dengue coinfection complicated by rhabdomyolysis and acute kidney injury: an unusual case with myoglobinemia, myoglobinuria but normal serum creatine kinase

    Directory of Open Access Journals (Sweden)

    Yong Kok Pin

    2012-12-01

    Full Text Available Abstract Background Acute kidney injury (AKI is a complication of severe malaria, and rhabdomyolysis with myoglobinuria is an uncommon cause. We report an unusual case of severe falciparum malaria with dengue coinfection complicated by AKI due to myoglobinemia and myoglobinuria while maintaining a normal creatine kinase (CK. Case presentation A 49-year old Indonesian man presented with fever, chills, and rigors with generalized myalgia and was diagnosed with falciparum malaria based on a positive blood smear. This was complicated by rhabdomyolysis with raised serum and urine myoglobin but normal CK. Despite rapid clearance of the parasitemia with intravenous artesunate and aggressive hydration maintaining good urine output, his myoglobinuria and acidosis worsened, progressing to uremia requiring renal replacement therapy. High-flux hemodiafiltration effectively cleared his serum and urine myoglobin with recovery of renal function. Further evaluation revealed evidence of dengue coinfection and past infection with murine typhus. Conclusion In patients with severe falciparum malaria, the absence of raised CK alone does not exclude a diagnosis of rhabdomyolysis. Raised serum and urine myoglobin levels could lead to AKI and should be monitored. In the event of myoglobin-induced AKI requiring dialysis, clinicians may consider using high-flux hemodiafiltration instead of conventional hemodialysis for more effective myoglobin removal. In Southeast Asia, potential endemic coinfections that can also cause or worsen rhabdomyolysis, such as dengue, rickettsiosis and leptospirosis, should be considered.

  12. Regulation of AMP-activated protein kinase by LKB1 and CaMKK in adipocytes

    DEFF Research Database (Denmark)

    Gormand, Amélie; Henriksson, Emma; Ström, Kristoffer

    2011-01-01

    AMP-activated protein kinase (AMPK) is a serine/threonine kinase that regulates cellular and whole body energy homeostasis. In adipose tissue, activation of AMPK has been demonstrated in response to a variety of extracellular stimuli. However, the upstream kinase that activates AMPK in adipocytes...

  13. IKAP: A heuristic framework for inference of kinase activities from Phosphoproteomics data.

    Science.gov (United States)

    Mischnik, Marcel; Sacco, Francesca; Cox, Jürgen; Schneider, Hans-Christoph; Schäfer, Matthias; Hendlich, Manfred; Crowther, Daniel; Mann, Matthias; Klabunde, Thomas

    2016-02-01

    Phosphoproteomics measurements are widely applied in cellular biology to detect changes in signalling dynamics. However, due to the inherent complexity of phosphorylation patterns and the lack of knowledge on how phosphorylations are related to functions, it is often not possible to directly deduce protein activities from those measurements. Here, we present a heuristic machine learning algorithm that infers the activities of kinases from Phosphoproteomics data using kinase-target information from the PhosphoSitePlus database. By comparing the estimated kinase activity profiles to the measured phosphosite profiles, it is furthermore possible to derive the kinases that are most likely to phosphorylate the respective phosphosite. We apply our approach to published datasets of the human cell cycle generated from HeLaS3 cells, and insulin signalling dynamics in mouse hepatocytes. In the first case, we estimate the activities of 118 at six cell cycle stages and derive 94 new kinase-phosphosite links that can be validated through either database or motif information. In the second case, the activities of 143 kinases at eight time points are estimated and 49 new kinase-target links are derived. The algorithm is implemented in Matlab and be downloaded from github. It makes use of the Optimization and Statistics toolboxes. https://github.com/marcel-mischnik/IKAP.git. marcel.mischnik@gmail.com Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  14. Activation of protein kinase C inhibits synthesis and release of decidual prolactin

    International Nuclear Information System (INIS)

    Harman, I.; Costello, A.; Ganong, B.; Bell, R.M.; Handwerger, S.

    1986-01-01

    Activation of calcium-activated, phospholipid-dependent protein kinase C by diacylglycerol and phorbol esters has been shown to mediate release of hormones in many systems. To determine whether protein kinase C activation is also involved in the regulation of prolactin release from human decidual, the authors have examined the effects of various acylglycerols and phorbol esters on the synthesis and release of prolactin from cultured human decidual cells. sn-1,2-Dioctanolyglycerol (diC 8 ), which is known to stimulate protein kinase C in other systems, inhibited prolactin release in a dose-dependent manner with maximal inhibition of 53.1% at 100 μM. Diolein (100 μM), which also stimulates protein kinase C activity in some systems, inhibited prolactin release by 21.3%. Phorbol 12-myristate 13-acetate (PMA), phorbol 12,13-didecanoate, and 4β-phorbol 12,13-dibutyrate, which activate protein kinase C in other systems, also inhibited the release of prolactin, which the protein kinase C inactivate 4α-phorbol-12,13-didecanoate was without effect. The inhibition of prolactin release was secondary to a decrease in prolactin synthesis. Although diC 8 and PMA inhibited the synthesis and release of prolactin, these agents had no effect on the synthesis or release of trichloroacetic acid-precipitable [ 35 S]methionine-labeled decidual proteins and did not cause the release of the cytosolic enzymes lactic dehydrogenase and alkaline phosphatase. DiC 8 and PMA stimulates the specific activity of protein kinase C in decidual tissue by 14.6 and 14.0-fold, respectively. The inhibition of the synthesis and release of prolactin by diC 8 and phorbol esters strongly implicates protein kinase C in the regulation of the production and release of prolactin from the decidua

  15. Effect of age, diet, and tissue type on PCr response to creatine supplementation.

    Science.gov (United States)

    Solis, Marina Yazigi; Artioli, Guilherme Giannini; Otaduy, Maria Concepción García; Leite, Claudia da Costa; Arruda, Walquiria; Veiga, Raquel Ramos; Gualano, Bruno

    2017-08-01

    Creatine/phosphorylcreatine (PCr) responses to creatine supplementation may be modulated by age, diet, and tissue, but studies assessing this possibility are lacking. Therefore we aimed to determine whether PCr responses vary as a function of age, diet, and tissue. Fifteen children, 17 omnivorous and 14 vegetarian adults, and 18 elderly individuals ("elderly") participated in this study. Participants were given placebo and subsequently creatine (0.3 g·kg -1 ·day -1 ) for 7 days in a single-blind fashion. PCr was measured through phosphorus magnetic resonance spectroscopy ( 31 P-MRS) in muscle and brain. Creatine supplementation increased muscle PCr in children ( P creatine supplementation in any group, and delta changes in brain PCr (-0.7 to +3.9%) were inferior to those in muscle PCr content (+10.3 to +27.6%; P creatine protocol (0.3 g·kg -1 ·day -1 for 7 days) may be affected by age, diet, and tissue. Whereas creatine supplementation was able to increase muscle PCr in all groups, although to different extents, brain PCr was shown to be unresponsive overall. These findings demonstrate the need to tailor creatine protocols to optimize creatine/PCr accumulation both in muscle and in brain, enabling a better appreciation of the pleiotropic properties of creatine. NEW & NOTEWORTHY A standardized creatine supplementation protocol (0.3 g·kg -1 ·day -1 for 7 days) effectively increased muscle, but not brain, phosphorylcreatine. Older participants responded better than younger participants whereas vegetarians responded better than omnivores. Responses to supplementation are thus dependent on age, tissue, and diet. This suggests that a single "universal" protocol, originally designed for increasing muscle creatine in young individuals, may lead to heterogeneous muscle responses in different populations or even no responses in tissues other than skeletal muscle. Copyright © 2017 the American Physiological Society.

  16. Timeless links replication termination to mitotic kinase activation.

    Directory of Open Access Journals (Sweden)

    Jayaraju Dheekollu

    2011-05-01

    Full Text Available The mechanisms that coordinate the termination of DNA replication with progression through mitosis are not completely understood. The human Timeless protein (Tim associates with S phase replication checkpoint proteins Claspin and Tipin, and plays an important role in maintaining replication fork stability at physical barriers, like centromeres, telomeres and ribosomal DNA repeats, as well as at termination sites. We show here that human Tim can be isolated in a complex with mitotic entry kinases CDK1, Auroras A and B, and Polo-like kinase (Plk1. Plk1 bound Tim directly and colocalized with Tim at a subset of mitotic structures in M phase. Tim depletion caused multiple mitotic defects, including the loss of sister-chromatid cohesion, loss of mitotic spindle architecture, and a failure to exit mitosis. Tim depletion caused a delay in mitotic kinase activity in vivo and in vitro, as well as a reduction in global histone H3 S10 phosphorylation during G2/M phase. Tim was also required for the recruitment of Plk1 to centromeric DNA and formation of catenated DNA structures at human centromere alpha satellite repeats. Taken together, these findings suggest that Tim coordinates mitotic kinase activation with termination of DNA replication.

  17. Selective anticancer activity of a hexapeptide with sequence homology to a non-kinase domain of Cyclin Dependent Kinase 4

    Directory of Open Access Journals (Sweden)

    Agarwala Usha

    2011-06-01

    Full Text Available Abstract Background Cyclin-dependent kinases 2, 4 and 6 (Cdk2, Cdk4, Cdk6 are closely structurally homologous proteins which are classically understood to control the transition from the G1 to the S-phases of the cell cycle by combining with their appropriate cyclin D or cyclin E partners to form kinase-active holoenzymes. Deregulation of Cdk4 is widespread in human cancer, CDK4 gene knockout is highly protective against chemical and oncogene-mediated epithelial carcinogenesis, despite the continued presence of CDK2 and CDK6; and overexpresssion of Cdk4 promotes skin carcinogenesis. Surprisingly, however, Cdk4 kinase inhibitors have not yet fulfilled their expectation as 'blockbuster' anticancer agents. Resistance to inhibition of Cdk4 kinase in some cases could potentially be due to a non-kinase activity, as recently reported with epidermal growth factor receptor. Results A search for a potential functional site of non-kinase activity present in Cdk4 but not Cdk2 or Cdk6 revealed a previously-unidentified loop on the outside of the C'-terminal non-kinase domain of Cdk4, containing a central amino-acid sequence, Pro-Arg-Gly-Pro-Arg-Pro (PRGPRP. An isolated hexapeptide with this sequence and its cyclic amphiphilic congeners are selectively lethal at high doses to a wide range of human cancer cell lines whilst sparing normal diploid keratinocytes and fibroblasts. Treated cancer cells do not exhibit the wide variability of dose response typically seen with other anticancer agents. Cancer cell killing by PRGPRP, in a cyclic amphiphilic cassette, requires cells to be in cycle but does not perturb cell cycle distribution and is accompanied by altered relative Cdk4/Cdk1 expression and selective decrease in ATP levels. Morphological features of apoptosis are absent and cancer cell death does not appear to involve autophagy. Conclusion These findings suggest a potential new paradigm for the development of broad-spectrum cancer specific therapeutics with

  18. Structural Basis for Selective Small Molecule Kinase Inhibition of Activated c-Met

    Energy Technology Data Exchange (ETDEWEB)

    Rickert, Keith W.; Patel, Sangita B.; Allison, Timothy J.; Byrne, Noel J.; Darke, Paul L.; Ford, Rachael E.; Guerin, David J.; Hall, Dawn L.; Kornienko, Maria; Lu, Jun; Munshi, Sanjeev K.; Reid, John C.; Shipman, Jennifer M.; Stanton, Elizabeth F.; Wilson, Kevin J.; Young, Jonathon R.; Soisson, Stephen M.; Lumb, Kevin J. (Merck)

    2012-03-15

    The receptor tyrosine kinase c-Met is implicated in oncogenesis and is the target for several small molecule and biologic agents in clinical trials for the treatment of cancer. Binding of the hepatocyte growth factor to the cell surface receptor of c-Met induces activation via autophosphorylation of the kinase domain. Here we describe the structural basis of c-Met activation upon autophosphorylation and the selective small molecule inhibiton of autophosphorylated c-Met. MK-2461 is a potent c-Met inhibitor that is selective for the phosphorylated state of the enzyme. Compound 1 is an MK-2461 analog with a 20-fold enthalpy-driven preference for the autophosphorylated over unphosphorylated c-Met kinase domain. The crystal structure of the unbound kinase domain phosphorylated at Tyr-1234 and Tyr-1235 shows that activation loop phosphorylation leads to the ejection and disorder of the activation loop and rearrangement of helix {alpha}C and the G loop to generate a viable active site. Helix {alpha}C adopts a orientation different from that seen in activation loop mutants. The crystal structure of the complex formed by the autophosphorylated c-Met kinase domain and compound 1 reveals a significant induced fit conformational change of the G loop and ordering of the activation loop, explaining the selectivity of compound 1 for the autophosphorylated state. The results highlight the role of structural plasticity within the kinase domain in imparting the specificity of ligand binding and provide the framework for structure-guided design of activated c-Met inhibitors.

  19. Muscle enzyme activities in a deep-sea squaloid shark, Centroscyllium fabricii, compared with its shallow-living relative, Squalus acanthias.

    Science.gov (United States)

    Treberg, Jason R; Martin, R Aidan; Driedzic, William R

    2003-12-01

    The activities of several enzymes of energy metabolism were measured in the heart, red muscle, and white muscle of a deep and a shallow living squaloid shark, Centroscyllium fabricii and Squalus acanthias, respectively. The phylogenetic closeness of these species, combined with their active predatory nature, similar body form, and size makes them well matched for comparison. This is the first time such a comparison has been made involving a deep-sea elasmobranch. Enzyme activities were similar in the heart, but generally lower in the red muscle of C. fabricii. Paralleling the trend seen in deep-sea teleosts, the white muscle of C. fabricii had substantially lower activities of key glycolytic enzymes, pyruvate kinase and lactate dehydrogenase, relative to S. acanthias or other shallow living elasmobranchs. Unexpectedly, between the squaloid sharks examined, creatine phosphokinase activity was higher in all tissues of the deep living C. fabricii. Low white muscle glycolytic enzyme activities in the deep-sea species coupled with high creatine phosphokinase activity suggests that the capacity for short burst swimming is likely limited once creatine phosphate supplies have been exhausted. Copyright 2003 Wiley-Liss, Inc.

  20. Arctigenin protects against steatosis in WRL68 hepatocytes through activation of phosphoinositide 3-kinase/protein kinase B and AMP-activated protein kinase pathways.

    Science.gov (United States)

    Chen, Kung-Yen; Lin, Jui-An; Yao, Han-Yun; Hsu, An-Chih; Tai, Yu-Ting; Chen, Jui-Tai; Hsieh, Mao-Chih; Shen, Tang-Long; Hsu, Ren-Yi; Wu, Hong-Tan; Wang, Guey Horng; Ho, Bing-Ying; Chen, Yu-Pei

    2018-04-01

    Arctigenin (ATG), a lignin extracted from Arctium lappa (L.), exerts antioxidant and anti-inflammatory effects. We hypothesized that ATG exerts a protective effect on hepatocytes by preventing nonalcoholic fatty liver disease (NAFLD) progression associated with lipid oxidation-associated lipotoxicity and inflammation. We established an in vitro NAFLD cell model by using normal WRL68 hepatocytes to investigate oleic acid (OA) accumulation and the potential bioactive role of ATG. The results revealed that ATG inhibited OA-induced lipid accumulation, lipid peroxidation, and inflammation in WRL68 hepatocytes, as determined using Oil Red O staining, thiobarbituric acid reactive substance assay, and inflammation antibody array assays. Quantitative RT-PCR analysis demonstrated that ATG significantly mitigated the expression of acetylcoenzyme A carboxylase 1 and sterol regulatory element-binding protein-1 and significantly increased the expression of carnitine palmitoyltransferase 1 and peroxisome proliferator-activated receptor alpha. The 40 targets of the Human Inflammation Antibody Array indicated that ATG significantly inhibited the elevation of the U937 lymphocyte chemoattractant, ICAM-1, IL-1β, IL-6, IL-6sR, IL-7, and IL-8. ATG could activate the phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and AMP-activated protein kinase (AMPK) pathways and could increase the phosphorylation levels of Akt and AMPK to mediate cell survival, lipid metabolism, oxidation stress, and inflammation. Thus, we demonstrated that ATG could inhibit NAFLD progression associated with lipid oxidation-associated lipotoxicity and inflammation, and we provided insights into the underlying mechanisms and revealed potential targets to enable a thorough understanding of NAFLD progression. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. Concentrações de creatino quinase, aspartato aminotransferase e desidrogenase lática em potros do nascimento até os seis meses de idade Concentration of creatine kinase, aspartate aminotransferase and lactate dehydrogenase in foals from birth up to sixth month

    Directory of Open Access Journals (Sweden)

    Elisiane Lourdes Da Cás

    2001-12-01

    Full Text Available Dez potros da raça Puro Sangue de Corrida (PSC, de ambos os sexos, foram avaliados quanto à concentração das enzimas séricas creatino quinase (CK, aspartato aminotransferase (AST e deshidrogenase lática (DHL. Foram colhidas amostras sangüíneas diariamente do 1º ao 7ºdia de vida e depois aos 15, 30, 60, 90, 120, 150 e 180 dias de idade. A concentração da CK mostrou um decréscimo significativo (pTen Thoroughbred foals, male and female, had the seric concentration of creatine kinase (CK, aspartate aminotransferase (AST and lactate dehydrogenase (LDH determined. Blood samples were collected every day from days 1 to 7 and on days 15, 30, 60, 90, 120, 150 and 180 of age. CK activity decreased significantly (p< 0.0003 in the first week and showed significant variation between day 15 and 6 months of age. AST showed a significant (p< 0.0001 increase in its values until 102 days of age, decreasing subsequently until 6 months of age. LDH values decreased significantly (p< 0.0002 between days 15 and 120, increasing subsequently until 6 months of age. At 6 months of age CK, AST and LDH activities were close to those of adult horses.

  2. Acute effect of a fight of Mixed Martial Arts (MMA on the serum concentrations of testosterone, cortisol, creatine kinase, lactate, and glucose

    Directory of Open Access Journals (Sweden)

    Rodrigo Poderoso de Souza

    2017-06-01

    Full Text Available The aim of this study was to analyse the serum concentrations of testosterone (T, cortisol (C, lactate (LAC, creatine kinase (CK and glucose (GLU on mixed martial arts (MMA athletes, before and after a fight. We divided 20 MMA athletes into two groups of 10 fighters each, according to the result of a fight, and were then evaluated four times: 24 hours before (-24h, one hour before (-1h, immediately after (0h and 24 hours after the fight (+24h. It was observed: a significant decrease in T and T/C between moment -24h and 0h and a subsequent increase between the moment 0h and +24h and a reverse behaviour in variables C, LAC and GLU (p<0.0001; a decrease in CK between moment -24h and -1h and an increase between moment -1h and +24h (p<0.0001; and differences between winners and losers T levels, in moments -24h, -1h, 0h and +24h (p = 0.009 e p < 0.001, p = 0.005 e p = 0,001, T and C, respectively, in T/C in the moments -24h and 0h (p=0.006 and p=0.001, respectively and in GLU levels (p<0.0001 in the moment 0h. Therefore, it seems that an MMA fight leads to metabolic stress and muscle damage, regardless of the result of the fight. The coaches have now more biochemical and hormonal references and indicators in response to an MMA fight.

  3. Stimulation of casein kinase II by epidermal growth factor: Relationship between the physiological activity of the kinase and the phosphorylation state of its beta subunit

    International Nuclear Information System (INIS)

    Ackerman, P.; Osheroff, N.; Glover, C.V.C.

    1990-01-01

    To determine relationships between the hormonal activation of casein kinase II and its phosphorylation state, epidermal growth factor (EGF)-treated and EGF-naive human A-431 carcinoma cells were cultured in the presence of [ 32 P]orthophosphate. Immunoprecipitation experiments indicated that casein kinase II in the cytosol of EGF-treated cells contained approximately 3-fold more incorporated [ 32 P]phosphate than did its counterpart in untreated cells. Levels of kinase phosphorylation paralleled levels of kinase activity over a wide range of EGF concentrations as well as over a time course of hormone action. Approximately 97% of the incorporated [ 32 P]phosphate was found in the β subunit of casein kinase II. Both activated and hormone-naive kinase contained radioactive phosphoserine and phosphothreonine but no phosphotyronsine. On the basis of proteolytic mapping experiments, EGF treatment of A-431 cells led to an increase in the average [ 32 P]phosphate content (i.e., hyperphosphorylation) of casein kinase II β subunit peptides which were modified prior to hormone treatment. Finally, the effect of alkaline phosphatase on the reaction kinetics of activated casein kinase II indicated that hormonal stimulation of the kinase resulted from the increase in its phosphorylation state

  4. The Effect of One Session Continuous and Intermittent Aerobic Exercise on Blood Responses of HSP72 , Cortisol and Creatine Kinase

    Directory of Open Access Journals (Sweden)

    M. Amani

    2013-10-01

    Full Text Available Introduction & Objective: Heat shock proteins help the cells’ ability to keep their structures against different stresses. The purpose of this study was to investigate the effect of one ses-sion continuous and intermittent aerobic exercise on blood responses of HSP72, cortisol and creatine kinase (CK. Materials & Methods: This study is semi-experimental in which 21 male student athletes were divided in continuous group (n=7, intermittent group (n=7 and control group (n=7. Exer-cise protocol of continuous group included 1 hour running with 80% maximum heart rate in-tensity and that of intermittent group was 3 stages of 20 minute running with the same inten-sity as of continuous group . Blood sampling of basal, pre exercise, immediately after exer-cise and 90 minutes after exercise were gathered and the amounts of HSP72, cortisol and CK, were measured by ELISA, RIA and Enzymatic methods respectively. The data was analyzed with one way ANOVA and repeated measure analysis of variance at P?0.05 significance level. Results: HSP72 levels in the continuous group and intermittent group despite an increase in the average did not show a statistically significant difference. Changes between the groups were significant in immediately after exercise and 90 minutes after exercise (P.values respectively 0.017 and 0.002. CK changes in continuous group were significant but cortisol changes in different stages hadn’t significant difference Conclusion: Exercise with its role associated with cortisol and CK will stimulate HSP72 and continuous exercise will make further increase in HSP72 and CK increasing leads to a greater HSP72 response. (Sci J Hamadan Univ Med Sci 2013; 20 (3:223-231

  5. Muscle pain and serum creatine kinase are not associated with low serum 25(OH) vitamin D levels in patients receiving statins.

    Science.gov (United States)

    Kurnik, Daniel; Hochman, Israel; Vesterman-Landes, Janet; Kenig, Tali; Katzir, Itzhak; Lomnicky, Yosef; Halkin, Hillel; Loebstein, Ronen

    2012-07-01

    Vitamin D deficiency has been associated in some studies with nonspecific musculoskeletal pain and, more specifically, with statin-induced myalgia, which was ameliorated by high-dose vitamin D supplements. Our objective was to explore the association between vitamin D status and statin-induced myalgia and elevation of serum creatine kinase (CK). Retrospective cohort study, based on the electronic database of a health maintenance organization. Six thousand eight hundred and eight patients (71·5% women) to whom statins were dispensed during 2008 and who had ≥1 CK and 25-hydroxy vitamin D (25OHD) levels measured during statin exposure. Of these, 376 patients (5·5%) had switched from a first-line statin to atorvastatin because of muscle pain (n = 220) or other reasons (n = 156). Measurements; In the entire cohort, we compared serum CK levels among serum 25OHD quartiles. In addition, we compared CK and 25OHD levels among patients with myalgia, other switchers and nonswitchers. The median 25OHD level in the entire cohort was 21·8 ng/ml [interquartile range (IQR), 16·3-27·4]. CK levels were marginally lower in patients in the lowest 25OHD quartile [median CK (IQR) in 25OHD quartiles 1-4, 87 (61-130), 90 (65-131), 91 (65-132) and 91 (67-131) IU/ml, respectively; P = 0·007]. 25OHD levels in statin switchers were similar to those in nonswitchers; moreover, there were no differences in 25OHD among switchers with muscle pain and other switchers. Our findings do not support an association between low 25OHD levels and statin-induced myalgia or CK elevation. © 2011 Blackwell Publishing Ltd.

  6. Novel autophosphorylation sites of Src family kinases regulate kinase activity and SH2 domain-binding capacity.

    Science.gov (United States)

    Weir, Marion E; Mann, Jacqueline E; Corwin, Thomas; Fulton, Zachary W; Hao, Jennifer M; Maniscalco, Jeanine F; Kenney, Marie C; Roman Roque, Kristal M; Chapdelaine, Elizabeth F; Stelzl, Ulrich; Deming, Paula B; Ballif, Bryan A; Hinkle, Karen L

    2016-04-01

    Src family tyrosine kinases (SFKs) are critical players in normal and aberrant biological processes. While phosphorylation importantly regulates SFKs at two known tyrosines, large-scale phosphoproteomics have revealed four additional tyrosines commonly phosphorylated in SFKs. We found these novel tyrosines to be autophosphorylation sites. Mimicking phosphorylation at the C-terminal site to the activation loop decreased Fyn activity. Phosphomimetics and direct phosphorylation at the three SH2 domain sites increased Fyn activity while reducing phosphotyrosine-dependent interactions. While 68% of human SH2 domains exhibit conservation of at least one of these tyrosines, few have been found phosphorylated except when found in cis to a kinase domain. © 2016 Federation of European Biochemical Societies.

  7. Effect of creatine supplementation and drop-set resistance training in untrained aging adults.

    Science.gov (United States)

    Johannsmeyer, Sarah; Candow, Darren G; Brahms, C Markus; Michel, Deborah; Zello, Gordon A

    2016-10-01

    To investigate the effects of creatine supplementation and drop-set resistance training in untrained aging adults. Participants were randomized to one of two groups: Creatine (CR: n=14, 7 females, 7 males; 58.0±3.0yrs, 0.1g/kg/day of creatine+0.1g/kg/day of maltodextrin) or Placebo (PLA: n=17, 7 females, 10 males; age: 57.6±5.0yrs, 0.2g/kg/day of maltodextrin) during 12weeks of drop-set resistance training (3days/week; 2 sets of leg press, chest press, hack squat and lat pull-down exercises performed to muscle fatigue at 80% baseline 1-repetition maximum [1-RM] immediately followed by repetitions to muscle fatigue at 30% baseline 1-RM). Prior to and following training and supplementation, assessments were made for body composition, muscle strength, muscle endurance, tasks of functionality, muscle protein catabolism and diet. Drop-set resistance training improved muscle mass, muscle strength, muscle endurance and tasks of functionality (pcreatine to drop-set resistance training significantly increased body mass (p=0.002) and muscle mass (p=0.007) compared to placebo. Males on creatine increased muscle strength (lat pull-down only) to a greater extent than females on creatine (p=0.005). Creatine enabled males to resistance train at a greater capacity over time compared to males on placebo (p=0.049) and females on creatine (p=0.012). Males on creatine (p=0.019) and females on placebo (p=0.014) decreased 3-MH compared to females on creatine. The addition of creatine to drop-set resistance training augments the gains in muscle mass from resistance training alone. Creatine is more effective in untrained aging males compared to untrained aging females. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. New Developments in Creatine Supplementation Research: Mechanisms of Athletic Performance Enhancement

    OpenAIRE

    DerHovanessian, Ariss

    2002-01-01

    In the last decade creatine supplementation has become the most popular ergogenic aid among athletes, with particular performance enhancements found in high-power output, anaerobic exercises. Physiologically, creatine and phosphocreatine provide an energy reservoir in skeletal muscle. Recent studies have also shown that the ergogenic effects of creatine are caused by muscle protein metabolism (or reduced catabolism), satellite cell proliferation, protective oxidant scavenging, and membrane st...

  9. 31 P magnetic resonance fingerprinting for rapid quantification of creatine kinase reaction rate in vivo.

    Science.gov (United States)

    Wang, Charlie Y; Liu, Yuchi; Huang, Shuying; Griswold, Mark A; Seiberlich, Nicole; Yu, Xin

    2017-12-01

    The purpose of this work was to develop a 31 P spectroscopic magnetic resonance fingerprinting (MRF) method for fast quantification of the chemical exchange rate between phosphocreatine (PCr) and adenosine triphosphate (ATP) via creatine kinase (CK). A 31 P MRF sequence (CK-MRF) was developed to quantify the forward rate constant of ATP synthesis via CK ( kfCK), the T 1 relaxation time of PCr ( T1PCr), and the PCr-to-ATP concentration ratio ( MRPCr). The CK-MRF sequence used a balanced steady-state free precession (bSSFP)-type excitation with ramped flip angles and a unique saturation scheme sensitive to the exchange between PCr and γATP. Parameter estimation was accomplished by matching the acquired signals to a dictionary generated using the Bloch-McConnell equation. Simulation studies were performed to examine the susceptibility of the CK-MRF method to several potential error sources. The accuracy of nonlocalized CK-MRF measurements before and after an ischemia-reperfusion (IR) protocol was compared with the magnetization transfer (MT-MRS) method in rat hindlimb at 9.4 T (n = 14). The reproducibility of CK-MRF was also assessed by comparing CK-MRF measurements with both MT-MRS (n = 17) and four angle saturation transfer (FAST) (n = 7). Simulation results showed that CK-MRF quantification of kfCK was robust, with less than 5% error in the presence of model inaccuracies including dictionary resolution, metabolite T 2 values, inorganic phosphate metabolism, and B 1 miscalibration. Estimation of kfCK by CK-MRF (0.38 ± 0.02 s -1 at baseline and 0.42 ± 0.03 s -1 post-IR) showed strong agreement with MT-MRS (0.39 ± 0.03 s -1 at baseline and 0.44 ± 0.04 s -1 post-IR). kfCK estimation was also similar between CK-MRF and FAST (0.38 ± 0.02 s -1 for CK-MRF and 0.38 ± 0.11 s -1 for FAST). The coefficient of variation from 20 s CK-MRF quantification of kfCK was 42% of that by 150 s MT-MRS acquisition and was 12% of that by 20 s FAST

  10. Co-administration of creatine and guanidinoacetic acid for augmented tissue bioenergetics: A novel approach?

    Science.gov (United States)

    Ostojic, Sergej M

    2017-07-01

    A confined absorption of exogenous creatine through creatine transporter (CRT1) seems to hamper its optimal uptake in bioenergetical deficits. Co-administration of guanidinoacetic acid (GAA) along with creatine could target other transport channels besides CRT1, and supremely improve cellular levels of creatine. This innovative approach might tackle tissues difficult to reach with conventional creatine interventions, providing a potentially more effective and safe mixture in clinical pharmacology and therapeutics. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  11. Creatine Use and Exercise Heat Tolerance in Dehydrated Men

    OpenAIRE

    Watson, Greig; Casa, Douglas J; Fiala, Kelly A; Hile, Amy; Roti, Melissa W; Healey, Julie C; Armstrong, Lawrence E; Maresh, Carl M

    2006-01-01

    Context: Creatine monohydrate (CrM) use is highly prevalent in team sports (eg, football, lacrosse, ice hockey) and by athletes at the high school, college, professional, and recreational levels. Concerns have been raised about whether creatine use is associated with increased cramping, muscle injury, heat intolerance, and risk of dehydration.

  12. RhoA/Rho Kinase Mediates Neuronal Death Through Regulating cPLA2 Activation.

    Science.gov (United States)

    Wu, Xiangbing; Walker, Chandler L; Lu, Qingbo; Wu, Wei; Eddelman, Daniel B; Parish, Jonathan M; Xu, Xiao-Ming

    2017-11-01

    Activation of RhoA/Rho kinase leads to growth cone collapse and neurite retraction. Although RhoA/Rho kinase inhibition has been shown to improve axon regeneration, remyelination and functional recovery, its role in neuronal cell death remains unclear. To determine whether RhoA/Rho kinase played a role in neuronal death after injury, we investigated the relationship between RhoA/Rho kinase and cytosolic phospholipase A 2 (cPLA 2 ), a lipase that mediates inflammation and cell death, using an in vitro neuronal death model and an in vivo contusive spinal cord injury model performed at the 10th thoracic (T10) vertebral level. We found that co-administration of TNF-α and glutamate induced spinal neuron death, and activation of RhoA, Rho kinase and cPLA 2 . Inhibition of RhoA, Rho kinase and cPLA 2 significantly reduced TNF-α/glutamate-induced cell death by 33, 52 and 43 %, respectively (p < 0.001). Inhibition of RhoA and Rho kinase also significantly downregulated cPLA 2 activation by 66 and 60 %, respectively (p < 0.01). Furthermore, inhibition of RhoA and Rho kinase reduced the release of arachidonic acid, a downstream substrate of cPLA 2 . The immunofluorescence staining showed that ROCK 1 or ROCK 2 , two isoforms of Rho kinase, was co-localized with cPLA 2 in neuronal cytoplasm. Interestingly, co-immunoprecipitation (Co-IP) assay showed that ROCK 1 or ROCK 2 bonded directly with cPLA 2 and phospho-cPLA 2 . When the Rho kinase inhibitor Y27632 was applied in mice with T10 contusion injury, it significantly decreased cPLA 2 activation and expression and reduced injury-induced apoptosis at and close to the lesion site. Taken together, our results reveal a novel mechanism of RhoA/Rho kinase-mediated neuronal death through regulating cPLA 2 activation.

  13. Detection of protein kinase activity by renaturation in sodium dodecyl sulfate-polyacrylamide gels

    International Nuclear Information System (INIS)

    Anostario, M. Jr.; Harrison, M.L.; Geahlen, R.L.

    1986-01-01

    The authors have developed a procedure for identifying protein kinase activity in protein samples following electrophoresis on SDS-polyacrylamide gels. Proteins are allowed to renature directly in the gel by removal of detergent. The gel is then incubated with [γ- 32 P]ATP to allow renatured protein kinases to autophosphorylate or to phosphorylate various substrates which can be incorporated into the gel. The positions of the radiolabeled proteins can then be detected by autoradiography. With this technique, using purified catalytic subunit of cAMP-dependent protein kinase, enzyme concentrations as low as 0.01 μg can be detected on gels containing 1.0 mg/ml casein. The procedure is also applicable for the determination of active subunits of multisubunit protein kinases. For example, when the two subunits of casein kinase II are separated by SDS-polyacrylamide gel electrophoresis and allowed to renature, only the larger α subunit shows activity. This procedure can also be used to detect and distinguish kinases present in heterogeneous mixtures. Starting with a particulate fraction from LSTRA, a murine T cell lymphoma, several distinct enzymes were detected, including a 30,000 Dalton protein with protein-tyrosine kinase activity. This same enzyme has also been detected in T lymphocytes and other T lymphoid cell lines

  14. Lindersin B from Lindernia crustacea induces neuritogenesis by activation of tyrosine kinase A/phosphatidylinositol 3 kinase/extracellular signal-regulated kinase signaling pathway.

    Science.gov (United States)

    Cheng, Lihong; Ye, Ying; Xiang, Lan; Osada, Hiroyuki; Qi, Jianhua

    2017-01-15

    Neurotrophic factors such as nerve growth factor (NGF) play important roles in nervous system. NGF is a potential therapeutic drug for treatment of neurodegenerative diseases. However, because of physicochemical property, NGF cannot pass through the blood-brain barrier (BBB). Hence, small molecules which exhibit NGF-mimic activity and can pass through the BBB are considered to be promising drug candidates for treatment of such diseases. The present study was designed to isolate NGF-mimic substance from extract of natural products, determine their structures and investigate mechanism of action of the active substance. Extract of Lindernia crustacean was partitioned between water and ethyl acetate to obtain water layer and ethyl acetate layer samples, respectively, and then evaluated their neuritogenic activity in PC12 cells. The active sample was separated by open columns, followed by HPLC purification to obtain active compound. Then, specific inhibitors were used to investigate signaling pathway of neurite outgrowth induced by the active compound. Finally, western blot analysis was performed to confirm the pathway proposed by inhibitor experiments. The ethyl acetate layer sample of extract of Lindernia crustacea exhibited significant neuritogenic activity. Two new compounds, named as linderside A and lindersin B, were isolated; their structures were elucidated by spectroscopic and chemical derivatization methods. Linderside A is a cucurbitane glycoside, whereas lindersin B is a cucurbitane triterpenoid. Each compound has an unusual isopentene unit, namely, a double bond bound to an unmodified isopropyl group at the end of cucurbitane triterpenoid side chain. Among them, lindersin B induced significant neurite outgrowth in PC12 cells, while linderside A was inactive against PC12 cells. Western blotting analysis results showed that lindersin B-induced neuritogenic activity depended on the activation of the mitogen-activated protein kinase (MAPK)/extracellular signal

  15. N-Acetylcysteine Supplementation Controls Total Antioxidant Capacity, Creatine Kinase, Lactate, and Tumor Necrotic Factor-Alpha against Oxidative Stress Induced by Graded Exercise in Sedentary Men

    Directory of Open Access Journals (Sweden)

    Donrawee Leelarungrayub

    2011-01-01

    Full Text Available Aim of this study was to evaluate the effects of short-term (7 days N-acetylcysteine (NAC at 1,200 mg daily supplementation on muscle fatigue, maximal oxygen uptake (VO2max, total antioxidant capacity (TAC, lactate, creatine kinase (CK, and tumor necrotic factor-alpha (TNF-α. Twenty-nine sedentary men (13 controls; 16 in the supplement group from a randomized control were included. At before and after supplementation, fatigue index (FI was evaluated in the quadriceps muscle, and performed a graded exercise treadmill test to induce oxidative stress, and as a measure of VO2max. Blood samples were taken before exercise and 20 minutes after it at before and after supplementation, to determine TAC, CK, lactate, and TNF-α levels. Results showed that FI and VO2max increased significantly in the supplement group. After exercise decreased the levels of TAC and increased lactate, CK, and TNF-α of both groups at before supplementation. After supplementation, lactate, CK, and TNF-α levels significantly increased and TAC decreased after exercise in the control group. Whereas the TAC and lactate levels did not change significantly, but CK and TNF-α increased significantly in the supplement group. Therefore, this results showed that NAC improved the muscle fatigue, VO2max, maintained TAC, controlled lactate production, but had no influence on CK and TNF-α.

  16. AcEST: BP916012 [AcEST

    Lifescience Database Archive (English)

    Full Text Available ne... 29 6.4 sp|P07335|KCRB_RAT Creatine kinase B-type OS=Rattus norvegicus G... 29 6.4 sp|Q04447|KCRB_MOUSE Creatine... kinase B-type OS=Mus musculus GN=C... 29 6.4 sp|P05122|KCRB_CHICK Creatine kinase B-type OS=Gallu...N=ARSH... 29 6.4 sp|O13879|YE1F_SCHPO Uncharacterized transporter C1B3.15C OS=Sch... 29 8.3 sp|P24722|KCRT_ONCMY Creatine...6 LESGFDAQSRTKLSDLS 36 +ES +D +++ D+S Sbjct: 119 IESKYDVHTKSVTVDVS 135 >sp|P07335|KCRB_RAT Creatine...DGDLSGRYYALKSMTEAEQQQLIDDHFLFDKPV 198 >sp|Q04447|KCRB_MOUSE Creatine kinase B-type OS=Mus musculus GN=Ckb PE

  17. Caffeine and Creatine Content of Dietary Supplements Consumed by Brazilian Soccer Players.

    Science.gov (United States)

    Inácio, Suelen Galante; de Oliveira, Gustavo Vieira; Alvares, Thiago Silveira

    2016-08-01

    Caffeine and creatine are ingredients in the most popular dietary supplements consumed by soccer players. However, some products may not contain the disclosed amounts of the ingredients listed on the label, compromising the safe usage and the effectiveness of these supplements. Therefore, the aim of this study was to evaluate the content of caffeine and creatine in dietary supplements consumed by Brazilian soccer players. The results obtained were compared with the caffeine content listed on the product label. Two batches of the supplement brands consumed by ≥ 50% of the players were considered for analysis. The quantification of caffeine and creatine in the supplements was determined by a high-performance liquid chromatography system with UV detector. Nine supplements of caffeine and 7 supplements of creatine met the inclusion criteria for analysis. Eight brands of caffeine and five brands of creatine showed significantly different values (p soccer players present inaccurate values listed on the label, although most presented no difference among batches. To ensure consumer safety and product efficacy, accurate information on caffeine and creatine content should be provided on all dietary supplement labels.

  18. Rho kinase inhibitor fasudil mitigates high-cholesterol diet-induced hypercholesterolemia and vascular damage.

    Science.gov (United States)

    Abdali, Nibrass Taher; Yaseen, Awny H; Said, Eman; Ibrahim, Tarek M

    2017-04-01

    The current study was designed to investigate the potential beneficial therapeutic outcome of Rho kinase inhibitor (fasudil) against hypercholesterolemia-induced myocardial and vascular injury in rabbits together with diet modification. Sixteen male rabbits were randomly divided into four groups: normal control group which received standard rabbit chow, hypercholesterolemic control group, and treated groups which received cholesterol-rich rabbit chow (1.5% cholesterol) for 8 weeks. Treated groups received either fasudil (100 mg/kg/day) or rosuvastatin (2.5 mg/kg/day) starting from the ninth week for further 4 weeks with interruption of the cholesterol-rich chow. Biochemical assessment of serum cholesterol, triglyceride, high-density lipoprotein (HDL), low-density lipoprotein (LDL), and myocardial oxidative/antioxidant biomarkers malondialdehyde (MDA), superoxide dismutase (SOD), and reduced glutathione (GSH), besides biochemical assessment of serum nitric oxide (NO), creatine kinase (CK), and lactate dehydrogenase (LDH) activities and serum total antioxidant capacity (TAC), was conducted. Serum vascular cell adhesion molecule 1 (VCAM-1) and serum Rho-associated protein kinase 1 (ROCK-1) were also evaluated along with histopathological examination of aorta specimens. Fasudil administration significantly decreased serum cholesterol, triglyceride (TG), and LDL and significantly increased serum HDL, with concomitant decrease in serum CK and LDH activities, NO, and restoration of serum TAC. Myocardial MDA significantly declined; SOD activity and GSH contents were restored. Serum ROCK-1 and VCAM-1 levels significantly declined as well. Vascular improvement was confirmed with histopathological examination, which revealed normal aortic intema with the absence of atheromas. Fasudil has promising anti-atherogenic activity mediated primarily via alleviation of hypercholesterolemia-induced oxidative stress and modulation of inflammatory response.

  19. Meta-Analysis of Creatine for Neuroprotection Against Parkinson's Disease.

    Science.gov (United States)

    Attia; Ahmed, Hussien; Gadelkarim, Mohamed; Morsi, Mahmoud; Awad, Kamal; Elnenny, Mohamed; Ghanem, Esraa; El-Jaafary, Shaimaa; Negida, Ahmed

    2017-01-01

    Creatine is an antioxidant agent that showed neuroprotective effects in animal models of Parkinson's disease (PD). Creatine was selected by the National Institute of Neurological Disorders and Stroke as a possible disease modifying agent for Parkinson's disease. Therefore, many clinical trials evaluated the efficacy of creatine for patients with PD. The aim of this systematic review and meta-analysis is to synthesize evidence from published randomized controlled trials (RCTs) about the efficacy of Creatine for patients with PD. We followed PRISMA statement guidelines during the preparation of this systematic review and meta-analysis. A computer literature search for PubMed, EBSCO, web of science and Ovid Midline was carried out. We included RCTs comparing creatine with placebo in terms of motor functions and quality of life. Outcomes of total Unified Parkinson's Disease Rating Scale (UPDRS), UPDRS I, UPDRS II, and UPDRS III were pooled as mean difference (MD) between two groups from baseline to the endpoint. Statistical heterogeneity was assessed by visual inspection of the forest plot and measured by chi-square and I square tests. Three RCTs (n=1935) were included in this study. The overall effect did not favor either of the two groups in terms of: UPDRS total score (MD 1.07, 95% CI [3.38 to 1.25], UPDRS III (MD 0.62, 95% CI [2.27 to 1.02]), UPDRS II (MD 0.03, 95% CI [0.81 to 0.86], or UPDRS I (MD 0.03, 95% CI [0.33 to 0.28]). Current evidence does not support the use of creatine for neuroprotection against PD. Future well-designed, randomized controlled trials are needed. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  20. A chimeric cyclic interferon-α2b peptide induces apoptosis by sequential activation of phosphatidylinositol 3-kinase, protein kinase Cδ and p38 MAP kinase.

    Science.gov (United States)

    Blank, V C; Bertucci, L; Furmento, V A; Peña, C; Marino, V J; Roguin, L P

    2013-06-10

    We have previously demonstrated that tyrosine phosphorylation of STAT1/3 and p38 mitogen-activated protein kinase (p38 MAPK) activation are involved in the apoptotic response triggered by a chimeric cyclic peptide of the interferon-α2b (IFN-α2b) in WISH cells. Since the peptide also induced serine phosphorylation of STAT proteins, in the present study we examined the kinase involved in serine STAT1 phosphorylation and the signaling effectors acting upstream such activation. We first found that p38 MAPK is involved in serine STAT1 phosphorylation, since a reduction of phophoserine-STAT1 levels was evident after incubating WISH cells with cyclic peptide in the presence of a p38 pharmacological inhibitor or a dominant-negative p38 mutant. Next, we demonstrated that the peptide induced activation of protein kinase Cδ (PKCδ). Based on this finding, the role of this kinase was then evaluated. After incubating WISH cells with a PKCδ inhibitor or after decreasing PKCδ expression levels by RNA interference, both peptide-induced serine STAT1 and p38 phosphorylation levels were significantly decreased, indicating that PKCδ functions as an upstream regulator of p38. We also showed that PKCδ and p38 activation stimulated by the peptide was inhibited by a specific pharmacological inhibitor of phosphatidylinositol 3-kinase (PI3K) or by a dominant-negative p85 PI3K-regulatory subunit, suggesting that PI3K is upstream in the signaling cascade. In addition, the role of PI3K and PKCδ in cyclic peptide-induced apoptosis was examined. Both signaling effectors were found to regulate the antiproliferative activity and the apoptotic response triggered by the cyclic peptide in WISH cells. In conclusion, we herein demonstrated that STAT1 serine phosphorylation is mediated by the sequential activation of PI3K, PKCδ and p38 MAPK. This signaling cascade contributes to the antitumor effect induced by the chimeric IFN-α2b cyclic peptide in WISH cells. Copyright © 2013 Elsevier Inc

  1. [Tricholoma equestre--animal toxicity study].

    Science.gov (United States)

    Chodorowski, Zygmunt; Sznitowska, Małgorzata; Wiśniewski, Marek; Sein Anand, Jacek; Waldman, Wojciech; Ronikier, Anna

    2004-01-01

    Animal toxicity study of Tricholoma equestre mushrooms stored for 12 months at (-)20 degrees C was performed using 30 male BALB/c mice. Three groups of 5 mice each were given suspension of T. equestre powder in water, boiled aqueous extract and chloroform-methanol extract dissolved in Miglyol 812 by gavage for three consecutive days. Mice in control groups were given water, Miglyol 812 and p-phenylenediamine (CAS 106-50-3). Creatine kinase activity was determined in serum collected 72 hours after the final dose. Mean activity of serum creatine kinase in mice treated with T. equestre powder, aqueous extract, chloroform-methanol extract and Miglyol 812 were 157 +/- 93, 129 +/- 30, 96 +/- 38, 111 +/- 66 U/L respectively and did not differ significantly from mean activity in mice which were given water (107 +/- 38 U/L). Mean serum creatine kinase activity in p-phenylenediamine group (265 +/- 63 U/L) was significantly higher than in group treated with water (p<0.01). Extracts of Tricholoma equestre mushrooms stored for 12 months at (-)20 degrees C did not cause rhabdomyolysis in male BALB/c mice.

  2. Cloning and characterization of a G protein-activated human phosphoinositide-3 kinase.

    Science.gov (United States)

    Stoyanov, B; Volinia, S; Hanck, T; Rubio, I; Loubtchenkov, M; Malek, D; Stoyanova, S; Vanhaesebroeck, B; Dhand, R; Nürnberg, B

    1995-08-04

    Phosphoinositide-3 kinase activity is implicated in diverse cellular responses triggered by mammalian cell surface receptors and in the regulation of protein sorting in yeast. Receptors with intrinsic and associated tyrosine kinase activity recruit heterodimeric phosphoinositide-3 kinases that consist of p110 catalytic subunits and p85 adaptor molecules containing Src homology 2 (SH2) domains. A phosphoinositide-3 kinase isotype, p110 gamma, was cloned and characterized. The p110 gamma enzyme was activated in vitro by both the alpha and beta gamma subunits of heterotrimeric guanosine triphosphate (GTP)-binding proteins (G proteins) and did not interact with p85. A potential pleckstrin homology domain is located near its amino terminus. The p110 gamma isotype may link signaling through G protein-coupled receptors to the generation of phosphoinositide second messengers phosphorylated in the D-3 position.

  3. Dietary guanidinoacetic acid increases brain creatine levels in healthy men

    DEFF Research Database (Denmark)

    Ostojic, Sergej M; Ostojic, Jelena; Drid, Patrik

    2017-01-01

    OBJECTIVE: Guanidinoacetic acid (GAA) is an experimental dietary additive that might act as a creatine source in tissues with high-energy requirements. In this case study, we evaluated brain levels of creatine in white matter, gray matter, cerebellum, and thalamus during 8 wk oral GAA......, and 8 wk, the participants underwent brain magnetic resonance spectroscopy, clinical chemistry studies, and open-ended questionnaire for side-effect prevalence and severity. RESULTS: Brain creatine levels increased in similar fashion in cerebellum, and white and gray matter after GAA supplementation......, with an initial increase of 10.7% reported after 4 wk, and additional upsurge (7.7%) from the weeks 4 to 8 follow-up (P creatine levels decreased after 4 wk for 6.5% (P = 0.02), and increased nonsignificantly after 8 wk for 8% (P = 0.09). GAA induced an increase in N-acetylaspartate levels at 8...

  4. Caffeine, creatine, GRIN2A and Parkinson's disease progression.

    Science.gov (United States)

    Simon, David K; Wu, Cai; Tilley, Barbara C; Lohmann, Katja; Klein, Christine; Payami, Haydeh; Wills, Anne-Marie; Aminoff, Michael J; Bainbridge, Jacquelyn; Dewey, Richard; Hauser, Robert A; Schaake, Susen; Schneider, Jay S; Sharma, Saloni; Singer, Carlos; Tanner, Caroline M; Truong, Daniel; Wei, Peng; Wong, Pei Shieen; Yang, Tianzhong

    2017-04-15

    Caffeine is neuroprotective in animal models of Parkinson's disease (PD) and caffeine intake is inversely associated with the risk of PD. This association may be influenced by the genotype of GRIN2A, which encodes an NMDA-glutamate-receptor subunit. In two placebo-controlled studies, we detected no association of caffeine intake with the rate of clinical progression of PD, except among subjects taking creatine, for whom higher caffeine intake was associated with more rapid progression. We now have analyzed data from 420 subjects for whom DNA samples and caffeine intake data were available from a placebo-controlled study of creatine in PD. The GRIN2A genotype was not associated with the rate of clinical progression of PD in the placebo group. However, there was a 4-way interaction between GRIN2A genotype, caffeine, creatine and the time since baseline. Among subjects in the creatine group with high levels of caffeine intake, but not among those with low caffeine intake, the GRIN2A T allele was associated with more rapid progression (p=0.03). These data indicate that the deleterious interaction between caffeine and creatine with respect to rate of progression of PD is influenced by GRIN2A genotype. This example of a genetic factor interacting with environmental factors illustrates the complexity of gene-environment interactions in the progression of PD. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. The Structural Basis for Activation and Inhibition of ZAP-70 Kinase Domain.

    Science.gov (United States)

    Huber, Roland G; Fan, Hao; Bond, Peter J

    2015-10-01

    ZAP-70 (Zeta-chain-associated protein kinase 70) is a tyrosine kinase that interacts directly with the activated T-cell receptor to transduce downstream signals, and is hence a major player in the regulation of the adaptive immune response. Dysfunction of ZAP-70 causes selective T cell deficiency that in turn results in persistent infections. ZAP-70 is activated by a variety of signals including phosphorylation of the kinase domain (KD), and binding of its regulatory tandem Src homology 2 (SH2) domains to the T cell receptor. The present study investigates molecular mechanisms of activation and inhibition of ZAP-70 via atomically detailed molecular dynamics simulation approaches. We report microsecond timescale simulations of five distinct states of the ZAP-70 KD, comprising apo, inhibited and three phosphorylated variants. Extensive analysis of local flexibility and correlated motions reveal crucial transitions between the states, thus elucidating crucial steps in the activation mechanism of the ZAP-70 KD. Furthermore, we rationalize previously observed staurosporine-bound crystal structures, suggesting that whilst the KD superficially resembles an "active-like" conformation, the inhibitor modulates the underlying protein dynamics and restricts it in a compact, rigid state inaccessible to ligands or cofactors. Finally, our analysis reveals a novel, potentially druggable pocket in close proximity to the activation loop of the kinase, and we subsequently use its structure in fragment-based virtual screening to develop a pharmacophore model. The pocket is distinct from classical type I or type II kinase pockets, and its discovery offers promise in future design of specific kinase inhibitors, whilst mutations in residues associated with this pocket are implicated in immunodeficiency in humans.

  6. The Structural Basis for Activation and Inhibition of ZAP-70 Kinase Domain.

    Directory of Open Access Journals (Sweden)

    Roland G Huber

    2015-10-01

    Full Text Available ZAP-70 (Zeta-chain-associated protein kinase 70 is a tyrosine kinase that interacts directly with the activated T-cell receptor to transduce downstream signals, and is hence a major player in the regulation of the adaptive immune response. Dysfunction of ZAP-70 causes selective T cell deficiency that in turn results in persistent infections. ZAP-70 is activated by a variety of signals including phosphorylation of the kinase domain (KD, and binding of its regulatory tandem Src homology 2 (SH2 domains to the T cell receptor. The present study investigates molecular mechanisms of activation and inhibition of ZAP-70 via atomically detailed molecular dynamics simulation approaches. We report microsecond timescale simulations of five distinct states of the ZAP-70 KD, comprising apo, inhibited and three phosphorylated variants. Extensive analysis of local flexibility and correlated motions reveal crucial transitions between the states, thus elucidating crucial steps in the activation mechanism of the ZAP-70 KD. Furthermore, we rationalize previously observed staurosporine-bound crystal structures, suggesting that whilst the KD superficially resembles an "active-like" conformation, the inhibitor modulates the underlying protein dynamics and restricts it in a compact, rigid state inaccessible to ligands or cofactors. Finally, our analysis reveals a novel, potentially druggable pocket in close proximity to the activation loop of the kinase, and we subsequently use its structure in fragment-based virtual screening to develop a pharmacophore model. The pocket is distinct from classical type I or type II kinase pockets, and its discovery offers promise in future design of specific kinase inhibitors, whilst mutations in residues associated with this pocket are implicated in immunodeficiency in humans.

  7. Adenosine monophosphate-activated protein kinase modulates the activated phenotype of hepatic stellate cells.

    Science.gov (United States)

    Caligiuri, Alessandra; Bertolani, Cristiana; Guerra, Cristina Tosti; Aleffi, Sara; Galastri, Sara; Trappoliere, Marco; Vizzutti, Francesco; Gelmini, Stefania; Laffi, Giacomo; Pinzani, Massimo; Marra, Fabio

    2008-02-01

    Adiponectin limits the development of liver fibrosis and activates adenosine monophosphate-activated protein kinase (AMPK). AMPK is a sensor of the cellular energy status, but its possible modulation of the fibrogenic properties of hepatic stellate cells (HSCs) has not been established. In this study, we investigated the role of AMPK activation in the biology of activated human HSCs. A time-dependent activation of AMPK was observed in response to a number of stimuli, including globular adiponectin, 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR), or metformin. All these compounds significantly inhibited platelet-derived growth factor (PDGF)-stimulated proliferation and migration of human HSCs and reduced the secretion of monocyte chemoattractant protein-1. In addition, AICAR limited the secretion of type I procollagen. Knockdown of AMPK by gene silencing increased the mitogenic effects of PDGF, confirming the negative modulation exerted by this pathway on HSCs. AMPK activation did not reduce PDGF-dependent activation of extracellular signal-regulated kinase (ERK) or Akt at early time points, whereas a marked inhibition was observed 24 hours after addition of PDGF, reflecting a block in cell cycle progression. In contrast, AICAR blocked short-term phosphorylation of ribosomal S6 kinase (p70(S6K)) and 4E binding protein-1 (4EBP1), 2 downstream effectors of the mammalian target of rapamycin (mTOR) pathway, by PDGF. The ability of interleukin-a (IL-1) to activate nuclear factor kappa B (NF-kappaB) was also reduced by AICAR. Activation of AMPK negatively modulates the activated phenotype of HSCs.

  8. Pervanadate induces Mammalian Ste20 Kinase 3 (MST3) tyrosine phosphorylation but not activation.

    Science.gov (United States)

    Kan, Wei-Chih; Lu, Te-Ling; Ling, Pin; Lee, Te-Hsiu; Cho, Chien-Yu; Huang, Chi-Ying F; Jeng, Wen-Yih; Weng, Yui-Ping; Chiang, Chun-Yen; Wu, Jin Bin; Lu, Te-Jung

    2016-07-01

    The yeast Ste20 (sterile) protein kinase, which is a serine/threonine kinase, responds to the stimulation of the G proteincoupled receptor (GPCR) pheromone receptor. Ste20 protein kinase serves as the critical component that links signaling from the GPCR/G proteins to the mitogen-activated protein kinase (MAPK) cascade in yeast. The yeast Ste20p functions as a MAP kinase kinase kinase kinase (MAP4K) in the pheromone response. Ste20-like kinases are structurally conserved from yeast to mammals. The mechanism by which MAP4K links GPCR to the MAPK pathway is less clearly defined in vertebrates. In addition to MAP4K, the tyrosine kinase cascade bridges G proteins and the MAPK pathway in vertebrate cells. Mammalian Ste20 Kinase 3 (MST3) has been categorized into the Ste20 family and has been reported to function in the regulation of cell polarity and migration. However, whether MST3 tyrosine phosphorylation regulates diverse signaling pathways is unknown. In this study, the tyrosine phosphatase inhibitor pervanadate was found to induce MST3 tyrosine phosphorylation in intact cells, and the activity of tyrosine-phosphorylated MST3 was measured. This tyrosine-directed phosphorylation was independent of MST3 activity. Parameters including protein conformation, Triton concentration and ionic concentration influenced the sensitivity of MST3 activity. Taken together, our data suggests that the serine/threonine kinase MST3 undergoes tyrosinedirected phosphorylation. The tyrosine-phosphorylated MST3 may create a docking site for the structurally conserved SH2/SH3 (Src Homology 2 and 3) domains within the Src oncoprotein. The unusual tyrosinephosphorylated MST3 may recruit MST3 to various signaling components. Copyright © 2016. Published by Elsevier Inc.

  9. Creatine kinase kinetics in professional soccer players during a competitive season. DOI: 10.5007/1980-0037.2011v13n3p189

    Directory of Open Access Journals (Sweden)

    Daniel Barbosa Coelho

    2011-04-01

    Full Text Available Serum creatine kinase (CK concentration has been widely used as an indicator of skeletal muscle damage in sports. However, there are no longitunal studies on post-game CK kinetics in Soccer during a competitive season. The aim of this study was to evaluate serum CK kinetics in professional Soccer players at different post-game times during a competitive season without training interruption. Seventeen professional soccer players (age: 22.2±3.1 years, height: 179±6.0 cm, body fat percentage: 9.5±1.1, and 67.0±3.5 mL O2/kg/min were evaluated over a period of 3 months of the national championship. Serum CK concentration was measured before the beginning of the season (baseline and at four different times after a soccer game (post-1: 12-20 h, post-2: 36-48 h, post-3: 60-65 h, and post-4: 90-110 h. Plasma CK concentrations were higher at all times when compared to baseline (p<0.05. Post-2 CK concentration was lower than post-1 and higher than post-3 and -4 (p<0.05, with no significant differences between post-3 and post-4. In conclusion, serum CK kinetics was influenced by the training routine of the soccer players, with a peak between 12 and 20 h after the game, returning to normal within 60-65 h. This procedure can be used to monitor the recovery state of athletes and game and training intensities.

  10. Maternal creatine supplementation affects the morpho-functional development of hippocampal neurons in rat offspring.

    Science.gov (United States)

    Sartini, S; Lattanzi, D; Ambrogini, P; Di Palma, M; Galati, C; Savelli, D; Polidori, E; Calcabrini, C; Rocchi, M B L; Sestili, P; Cuppini, R

    2016-01-15

    Creatine supplementation has been shown to protect neurons from oxidative damage due to its antioxidant and ergogenic functions. These features have led to the hypothesis of creatine supplementation use during pregnancy as prophylactic treatment to prevent CNS damage, such as hypoxic-ischemic encephalopathy. Unfortunately, very little is known on the effects of creatine supplementation during neuron differentiation, while in vitro studies revealed an influence on neuron excitability, leaving the possibility of creatine supplementation during the CNS development an open question. Using a multiple approach, we studied the hippocampal neuron morphological and functional development in neonatal rats born by dams supplemented with 1% creatine in drinking water during pregnancy. CA1 pyramidal neurons of supplemented newborn rats showed enhanced dendritic tree development, increased LTP maintenance, larger evoked-synaptic responses, and higher intrinsic excitability in comparison to controls. Moreover, a faster repolarizing phase of action potential with the appearance of a hyperpolarization were recorded in neurons of the creatine-treated group. Consistently, CA1 neurons of creatine exposed pups exhibited a higher maximum firing frequency than controls. In summary, we found that creatine supplementation during pregnancy positively affects morphological and electrophysiological development of CA1 neurons in offspring rats, increasing neuronal excitability. Altogether, these findings emphasize the need to evaluate the benefits and the safety of maternal intake of creatine in humans. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

  11. Auxin efflux by PIN-FORMED proteins is activated by two different protein kinases, D6 PROTEIN KINASE and PINOID

    KAUST Repository

    Zourelidou, Melina; Absmanner, Birgit; Weller, Benjamin; Barbosa, Inê s CR; Willige, Bjö rn C; Fastner, Astrid; Streit, Verena; Port, Sarah A; Colcombet, Jean; de la Fuente van Bentem, Sergio; Hirt, Heribert; Kuster, Bernhard; Schulze, Waltraud X; Hammes, Ulrich Z; Schwechheimer, Claus

    2014-01-01

    The development and morphology of vascular plants is critically determined by synthesis and proper distribution of the phytohormone auxin. The directed cell-to-cell distribution of auxin is achieved through a system of auxin influx and efflux transporters. PIN-FORMED (PIN) proteins are proposed auxin efflux transporters, and auxin fluxes can seemingly be predicted based on the-in many cells-asymmetric plasma membrane distribution of PINs. Here, we show in a heterologous Xenopus oocyte system as well as in Arabidopsis thaliana inflorescence stems that PIN-mediated auxin transport is directly activated by D6 PROTEIN KINASE (D6PK) and PINOID (PID)/WAG kinases of the Arabidopsis AGCVIII kinase family. At the same time, we reveal that D6PKs and PID have differential phosphosite preferences. Our study suggests that PIN activation by protein kinases is a crucial component of auxin transport control that must be taken into account to understand auxin distribution within the plant.

  12. Auxin efflux by PIN-FORMED proteins is activated by two different protein kinases, D6 PROTEIN KINASE and PINOID

    KAUST Repository

    Zourelidou, Melina

    2014-06-19

    The development and morphology of vascular plants is critically determined by synthesis and proper distribution of the phytohormone auxin. The directed cell-to-cell distribution of auxin is achieved through a system of auxin influx and efflux transporters. PIN-FORMED (PIN) proteins are proposed auxin efflux transporters, and auxin fluxes can seemingly be predicted based on the-in many cells-asymmetric plasma membrane distribution of PINs. Here, we show in a heterologous Xenopus oocyte system as well as in Arabidopsis thaliana inflorescence stems that PIN-mediated auxin transport is directly activated by D6 PROTEIN KINASE (D6PK) and PINOID (PID)/WAG kinases of the Arabidopsis AGCVIII kinase family. At the same time, we reveal that D6PKs and PID have differential phosphosite preferences. Our study suggests that PIN activation by protein kinases is a crucial component of auxin transport control that must be taken into account to understand auxin distribution within the plant.

  13. Erythrocyte creatine as a marker of intravascular hemolysis due to left ventricular outflow tract obstruction in hypertrophic cardiomyopathy.

    Science.gov (United States)

    Kubo, Toru; Okumiya, Toshika; Baba, Yuichi; Hirota, Takayoshi; Tanioka, Katsutoshi; Yamasaki, Naohito; Sugiura, Tetsuro; Doi, Yoshinori L; Kitaoka, Hiroaki

    2016-03-01

    Erythrocyte creatine, a marker of erythrocyte age that increases with shortening of erythrocyte survival, has been reported to be a quantitative and reliable marker for intravascular hemolysis. We hypothesized that hemolysis could also occur due to intraventricular obstruction in patients with hypertrophic cardiomyopathy (HCM). The purpose of this study was to examine the presence of subclinical hemolysis and the relation between intravascular hemolysis and intraventricular pressure gradient (IVPG). We measured erythrocyte creatine in 92 HCM patients. Twelve patients had left ventricular outflow tract obstruction (LVOTO), 4 had midventricular obstruction (MVO), and the remaining 76 were non-obstructive. Erythrocyte creatine levels ranged from 0.92 to 4.36μmol/g hemoglobin. Higher levels of erythrocyte creatine were associated with higher IVPG (r=0.437, pcreatine levels are high (≥1.8μmol/g hemoglobin), subclinical hemolysis is considered to be present. Half of LVOTO patients and no MVO patients showed high erythrocyte creatine levels. Although non-obstructive patients did not show significant intraventricular obstruction at rest, some showed high erythrocyte creatine levels. When LVOT-PG was measured during the strain phase of the Valsalva maneuver in 20 non-obstructive patients, 7 of those 20 patients showed LVOTO. In the 20 patients, there was no relation between erythrocyte creatine levels and LVOT-PG before the Valsalva maneuver (r=0.125, p=0.600), whereas there was a significant correlation between erythrocyte creatine and LVOT-PG provoked by the Valsalva maneuver (r=0.695, p=0.001). There is biochemical evidence of subclinical hemolysis in patients with HCM, and this hemolysis seems to be associated with LVOTO provoked by daily physical activities. Copyright © 2015 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.

  14. AMP-activated protein kinase downregulates Kv7.1 cell surface expression

    DEFF Research Database (Denmark)

    Andersen, Martin N; Krzystanek, Katarzyna; Jespersen, Thomas

    2012-01-01

    in response to polarization of the epithelial Madin-Darby canine kidney (MDCK) cell line and that this was mediated by activation of protein kinase C (PKC). In this study, the pathway downstream of PKC, which leads to internalization of Kv7.1 upon cell polarization, is elucidated. We show by confocal...... microscopy that Kv7.1 is endocytosed upon initiation of the polarization process and sent for degradation by the lysosomal pathway. The internalization could be mimicked by pharmacological activation of the AMP-activated protein kinase (AMPK) using three different AMPK activators. We demonstrate...

  15. Activation of c-Raf-1 kinase signal transduction pathway in alpha(7) integrin-deficient mice.

    Science.gov (United States)

    Saher, G; Hildt, E

    1999-09-24

    Integrin alpha(7)-deficient mice develop a novel form of muscular dystrophy. Here we report that deficiency of alpha(7) integrin causes an activation of the c-Raf-1/mitogen-activated protein (MAP) 2 kinase signal transduction pathway in muscle cells. The observed activation of c-Raf-1/MAP2 kinases is a specific effect, because the alpha(7) integrin deficiency does not cause unspecific stress as determined by measurement of the Hsp72/73 level and activity of the JNK2 kinase. Because an increased level of activated FAK was found in muscle of alpha(7) integrin-deficient mice, the activation of c-Raf-1 kinase is triggered most likely by an integrin-dependent pathway. In accordance with this, in the integrin alpha(7)-deficient mice, part of the integrin beta(1D) variant in muscle is replaced by the beta(1A) variant, which permits the FAK activation. A recent report describes that integrin activity can be down-modulated by the c-Raf-1/MAP2 kinase pathway. Specific activation of the c-Raf-1/MAP2 kinases by cell-permeable peptides in skeletal muscle of rabbits causes degeneration of muscle fibers. Therefore, we conclude that in alpha(7) integrin-deficient mice, the continuous activation of c-Raf-1 kinase causes a permanent reduction of integrin activity diminishing integrin-dependent cell-matrix interactions and thereby contributing to the development of the dystrophic phenotype.

  16. Characterization of breakpoint cluster region kinase and SH2-binding activities.

    Science.gov (United States)

    Afar, D E; Witte, O N

    1995-01-01

    BCR is an interesting signaling protein, whose cellular function is currently unknown. Its biochemical properties include serine kinase activity, SH2-binding activity, and a GTPase-activating activity. The SH2-binding activity is particularly interesting because it may link BCR to signaling pathways involving SH2-containing molecules. Since tyrosine phosphorylation of BCR has been detected in CML-derived cell lines and since tyrosine-phosphorylated BCR shows increased affinity toward certain SH2 domains, it seems particularly important to further characterize this activity. This chapter described a simple purification scheme for partial purification of BCR, which can be used to assess in vitro kinase and SH2-binding activities.

  17. Mitogen-Activated Protein Kinase Kinase 3 Regulates Seed Dormancy in Barley.

    Science.gov (United States)

    Nakamura, Shingo; Pourkheirandish, Mohammad; Morishige, Hiromi; Kubo, Yuta; Nakamura, Masako; Ichimura, Kazuya; Seo, Shigemi; Kanamori, Hiroyuki; Wu, Jianzhong; Ando, Tsuyu; Hensel, Goetz; Sameri, Mohammad; Stein, Nils; Sato, Kazuhiro; Matsumoto, Takashi; Yano, Masahiro; Komatsuda, Takao

    2016-03-21

    Seed dormancy has fundamental importance in plant survival and crop production; however, the mechanisms regulating dormancy remain unclear [1-3]. Seed dormancy levels generally decrease during domestication to ensure that crops successfully germinate in the field. However, reduction of seed dormancy can cause devastating losses in cereals like wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) due to pre-harvest sprouting, the germination of mature seed (grain) on the mother plant when rain occurs before harvest. Understanding the mechanisms of dormancy can facilitate breeding of crop varieties with the appropriate levels of seed dormancy [4-8]. Barley is a model crop [9, 10] and has two major seed dormancy quantitative trait loci (QTLs), SD1 and SD2, on chromosome 5H [11-19]. We detected a QTL designated Qsd2-AK at SD2 as the single major determinant explaining the difference in seed dormancy between the dormant cultivar "Azumamugi" (Az) and the non-dormant cultivar "Kanto Nakate Gold" (KNG). Using map-based cloning, we identified the causal gene for Qsd2-AK as Mitogen-activated Protein Kinase Kinase 3 (MKK3). The dormant Az allele of MKK3 is recessive; the N260T substitution in this allele decreases MKK3 kinase activity and appears to be causal for Qsd2-AK. The N260T substitution occurred in the immediate ancestor allele of the dormant allele, and the established dormant allele became prevalent in barley cultivars grown in East Asia, where the rainy season and harvest season often overlap. Our findings show fine-tuning of seed dormancy during domestication and provide key information for improving pre-harvest sprouting tolerance in barley and wheat. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. CSK negatively regulates nerve growth factor induced neural differentiation and augments AKT kinase activity

    International Nuclear Information System (INIS)

    Dey, Nandini; Howell, Brian W.; De, Pradip K.; Durden, Donald L.

    2005-01-01

    Src family kinases are involved in transducing growth factor signals for cellular differentiation and proliferation in a variety of cell types. The activity of all Src family kinases (SFKs) is controlled by phosphorylation at their C-terminal 527-tyrosine residue by C-terminal SRC kinase, CSK. There is a paucity of information regarding the role of CSK and/or specific Src family kinases in neuronal differentiation. Pretreatment of PC12 cells with the Src family kinase inhibitor, PP1, blocked NGF-induced activation of SFKs and obliterated neurite outgrowth. To confirm a role for CSK and specific isoforms of SFKs in neuronal differentiation, we overexpressed active and catalytically dead CSK in the rat pheochromocytoma cell line, PC12. CSK overexpression caused a profound inhibition of NGF-induced activation of FYN, YES, RAS, and ERK and inhibited neurite outgrowth, NGF-stimulated integrin-directed migration and blocked the NGF-induced conversion of GDP-RAC to its GTP-bound active state. CSK overexpression markedly augmented the activation state of AKT following NGF stimulation. In contrast, kinase-dead CSK augmented the activation of FYN, RAS, and ERK and increased neurite outgrowth. These data suggest a distinct requirement for CSK in the regulation of NGF/TrkA activation of RAS, RAC, ERK, and AKT via the differential control of SFKs in the orchestration of neuronal differentiation

  19. Urinary excretion of creatine and creatinine in gamma irradiated rats

    Energy Technology Data Exchange (ETDEWEB)

    Basu, S K; Srinivasan, M N; Chuttani, K; Bhatnagar, A; Ghose, A

    1985-06-01

    Dose response relationships of creatine, creatinine excretions and their ratio in 24 hr urine samples have been studied on each individual day upto 4 days after 1-7 Gy whole body gamma irradiation to rats. Creatine excretion reaches the peak on the 2nd day while creatinine excretion reaches the peak on the first day and a plateau is maintained up to the 4th day in each case. Good dose response correlationship is maintained for creatine or creatinine levels up to the 4th day and for creatine creatinine ratio up to the 3rd day. Seperate dose response curves are needed on each individual day for using these parameters for biological dosimetry purpose. Administration of the radioprotectors viz., combination of 5-hydroxytryptophan (HT) and 2-amino-ethylisothiuronium bromide hydrobromide (AET), HT alone and optimum radioprotecting dose of AET before 5 Gy whole body ..gamma..-irradiation have not been of help for reducing creatinineurea. (author).

  20. Urinary excretion of creatine and creatinine in gamma irradiated rats

    International Nuclear Information System (INIS)

    Basu, S.K.; Srinivasan, M.N.; Chuttani, K.; Bhatnagar, A.; Ghose, A.

    1985-01-01

    Dose response relationships of creatine, creatinie excretions and their ratio in 24 hr urine samples have been studied on each individual day upto 4 days after 1-7 Gy whole body gamma irradiation to rats. Creatine excretion reaches the peak on the 2nd day while creatinine excretion reaches the peak on the first day and a plateau is maintained upto the 4th day in each case. Good dose response correlationship is maintained for creatine or creatinine levels upto the 4th day and for creatine creatinine ratio upto the 3rd day. Seperate dose response curves are needed on each individual day for using these parameters for biological dosimetry purpose. Administration of the radioprotectors viz., combination of 5-hydroxytryptophan (HT) and 2-amino-ethylisothiuronium bromide hydrobromide (AET), HT alone and optimum radioprotecting dose of AET before 5 Gy whole body γ-irradiation have not been of help for reducing creatinineurea. (author)

  1. Theoretical Insights Reveal Novel Motions in Csk's SH3 Domain That Control Kinase Activation.

    Directory of Open Access Journals (Sweden)

    Sulyman Barkho

    Full Text Available The Src family of tyrosine kinases (SFKs regulate numerous aspects of cell growth and differentiation and are under the principal control of the C-terminal Src Kinase (Csk. Although Csk and SFKs share conserved kinase, SH2 and SH3 domains, they differ considerably in three-dimensional structure, regulatory mechanism, and the intrinsic kinase activities. Although the SH2 and SH3 domains are known to up- or down-regulate tyrosine kinase function, little is known about the global motions in the full-length kinase that govern these catalytic variations. We use a combination of accelerated Molecular Dynamics (aMD simulations and experimental methods to provide a new view of functional motions in the Csk scaffold. These computational studies suggest that high frequency vibrations in the SH2 domain are coupled through the N-terminal lobe of the kinase domain to motions in the SH3 domain. The effects of these reflexive movements on the kinase domain can be viewed using both Deuterium Exchange Mass Spectrometry (DXMS and steady-state kinetic methods. Removal of several contacts, including a crystallographically unobserved N-terminal segment, between the SH3 and kinase domains short-circuit these coupled motions leading to reduced catalytic efficiency and stability of N-lobe motifs within the kinase domain. The data expands the model of Csk's activation whereby separate domains productively interact with two diametrically opposed surfaces of the kinase domain. Such reversible transitions may organize the active structure of the tyrosine kinase domain of Csk.

  2. Insulin-induced decrease in protein phosphorylation in rat adipocytes not explained by decreased A-kinase activity

    International Nuclear Information System (INIS)

    Egan, J.J.; Greenberg, A.S.; Chang, M.K.; Londos, C.

    1987-01-01

    In isolated rat adipocytes, insulin inhibits lipolysis to a greater extent than would be predicted by the decrease in (-/+)cAMP activity ratio of cAMP-dependent protein kinase [A-kinase], from which it was speculated that insulin promotes the dephosphorylation of hormone-sensitive lipase. They have examined the phosphorylation state of cellular proteins under conditions of varying A-kinase activities in the presence and absence of insulin. Protein phosphorylation was determined by SDS-PAGE electrophoresis of extracts from 32 P-loaded cells; glycerol and A-kinase activity ratios were measured in the cytosolic extracts from control, non-radioactive cells. Increased protein phosphorylation in general occurred over the same range of A-kinase activity ratios, 0.1-0.3, associated with increased glycerol release. The insulin-induced decrease in lipolysis was associated with a decrease in the 32 P content of several proteins, an effect not explained by the modest reduction in A-kinase activity by insulin. This effect of insulin on protein phosphorylation was lost as the A-kinase activity ratios exceeded 0.5. The results suggest that insulin promotes the dephosphorylation of those adipocyte proteins which are subject to phosphorylation by A-kinase

  3. Eotaxin induces degranulation and chemotaxis of eosinophils through the activation of ERK2 and p38 mitogen-activated protein kinases

    DEFF Research Database (Denmark)

    Kampen, G T; Stafford, S; Adachi, T

    2000-01-01

    Eotaxin and other CC chemokines acting via CC chemokine receptor-3 (CCR3) are believed to play an integral role in the development of eosinophilic inflammation in asthma and allergic inflammatory diseases. However, little is known about the intracellular events following agonist binding to CCR3...... and the relationship of these events to the functional response of the cell. The objectives of this study were to investigate CCR3-mediated activation of the mitogen-activated protein (MAP) kinases extracellular signal-regulated kinase-2 (ERK2), p38, and c-jun N-terminal kinase (JNK) in eosinophils and to assess...... the requirement for MAP kinases in eotaxin-induced eosinophil cationic protein (ECP) release and chemotaxis. MAP kinase activation was studied in eotaxin-stimulated eosinophils (more than 97% purity) by Western blotting and immune-complex kinase assays. ECP release was measured by radioimmunoassay. Chemotaxis...

  4. Structures of the inactive and active states of RIP2 kinase inform on the mechanism of activation.

    Directory of Open Access Journals (Sweden)

    Erika Pellegrini

    Full Text Available Innate immune receptors NOD1 and NOD2 are activated by bacterial peptidoglycans leading to recruitment of adaptor kinase RIP2, which, upon phosphorylation and ubiquitination, becomes a scaffold for downstream effectors. The kinase domain (RIP2K is a pharmaceutical target for inflammatory diseases caused by aberrant NOD2-RIP2 signalling. Although structures of active RIP2K in complex with inhibitors have been reported, the mechanism of RIP2K activation remains to be elucidated. Here we analyse RIP2K activation by combining crystal structures of the active and inactive states with mass spectrometric characterization of their phosphorylation profiles. The active state has Helix αC inwardly displaced and the phosphorylated Activation Segment (AS disordered, whilst in the inactive state Helix αC is outwardly displaced and packed against the helical, non-phosphorylated AS. Biophysical measurements show that the active state is a stable dimer whilst the inactive kinase is in a monomer-dimer equilibrium, consistent with the observed structural differences at the dimer interface. We conclude that RIP2 kinase auto-phosphorylation is intimately coupled to dimerization, similar to the case of BRAF. Our results will help drug design efforts targeting RIP2 as a potential treatment for NOD2-RIP2 related inflammatory diseases.

  5. Structures of apicomplexan calcium-dependent protein kinases reveal mechanism of activation by calcium

    Energy Technology Data Exchange (ETDEWEB)

    Wernimont, Amy K; Artz, Jennifer D.; Jr, Patrick Finerty; Lin, Yu-Hui; Amani, Mehrnaz; Allali-Hassani, Abdellah; Senisterra, Guillermo; Vedadi, Masoud; Tempel, Wolfram; Mackenzie, Farrell; Chau, Irene; Lourido, Sebastian; Sibley, L. David; Hui, Raymond (Toronto); (WU-MED)

    2010-09-21

    Calcium-dependent protein kinases (CDPKs) have pivotal roles in the calcium-signaling pathway in plants, ciliates and apicomplexan parasites and comprise a calmodulin-dependent kinase (CaMK)-like kinase domain regulated by a calcium-binding domain in the C terminus. To understand this intramolecular mechanism of activation, we solved the structures of the autoinhibited (apo) and activated (calcium-bound) conformations of CDPKs from the apicomplexan parasites Toxoplasma gondii and Cryptosporidium parvum. In the apo form, the C-terminal CDPK activation domain (CAD) resembles a calmodulin protein with an unexpected long helix in the N terminus that inhibits the kinase domain in the same manner as CaMKII. Calcium binding triggers the reorganization of the CAD into a highly intricate fold, leading to its relocation around the base of the kinase domain to a site remote from the substrate binding site. This large conformational change constitutes a distinct mechanism in calcium signal-transduction pathways.

  6. Creatine maintains intestinal homeostasis and protects against colitis.

    Science.gov (United States)

    Turer, Emre; McAlpine, William; Wang, Kuan-Wen; Lu, Tianshi; Li, Xiaohong; Tang, Miao; Zhan, Xiaoming; Wang, Tao; Zhan, Xiaowei; Bu, Chun-Hui; Murray, Anne R; Beutler, Bruce

    2017-02-14

    Creatine, a nitrogenous organic acid, replenishes cytoplasmic ATP at the expense of mitochondrial ATP via the phosphocreatine shuttle. Creatine levels are maintained by diet and endogenous synthesis from arginine and glycine. Glycine amidinotransferase (GATM) catalyzes the rate-limiting step of creatine biosynthesis: the transfer of an amidino group from arginine to glycine to form ornithine and guanidinoacetate. We screened 36,530 third-generation germline mutant mice derived from N -ethyl- N -nitrosourea-mutagenized grandsires for intestinal homeostasis abnormalities after oral administration of dextran sodium sulfate (DSS). Among 27 colitis susceptibility phenotypes identified and mapped, one was strongly correlated with a missense mutation in Gatm in a recessive model of inheritance, and causation was confirmed by CRISPR/Cas9 gene targeting. Supplementation of homozygous Gatm mutants with exogenous creatine ameliorated the colitis phenotype. CRISPR/Cas9-targeted ( Gatm c/c ) mice displayed a normal peripheral immune response and immune cell homeostasis. However, the intestinal epithelium of the Gatm c/c mice displayed increased cell death and decreased proliferation during DSS treatment. In addition, Gatm c/c colonocytes showed increased metabolic stress in response to DSS with higher levels of phospho-AMPK and lower levels of phosphorylation of mammalian target of rapamycin (phospho-mTOR). These findings establish an in vivo requirement for rapid replenishment of cytoplasmic ATP within colonic epithelial cells in the maintenance of the mucosal barrier after injury.

  7. Creatine supplementation in the aging population: effects on skeletal muscle, bone and brain.

    Science.gov (United States)

    Gualano, Bruno; Rawson, Eric S; Candow, Darren G; Chilibeck, Philip D

    2016-08-01

    This narrative review aims to summarize the recent findings on the adjuvant application of creatine supplementation in the management of age-related deficits in skeletal muscle, bone and brain metabolism in older individuals. Most studies suggest that creatine supplementation can improve lean mass and muscle function in older populations. Importantly, creatine in conjunction with resistance training can result in greater adaptations in skeletal muscle than training alone. The beneficial effect of creatine upon lean mass and muscle function appears to be applicable to older individuals regardless of sex, fitness or health status, although studies with very old (>90 years old) and severely frail individuals remain scarce. Furthermore, there is evidence that creatine may affect the bone remodeling process; however, the effects of creatine on bone accretion are inconsistent. Additional human clinical trials are needed using larger sample sizes, longer durations of resistance training (>52 weeks), and further evaluation of bone mineral, bone geometry and microarchitecture properties. Finally, a number of studies suggest that creatine supplementation improves cognitive processing under resting and various stressed conditions. However, few data are available on older adults, and the findings are discordant. Future studies should focus on older adults and possibly frail elders or those who have already experienced an age-associated cognitive decline.

  8. Effects of overexpression of IL-1 receptor-associated kinase on NFkappaB activation, IL-2 production and stress-activated protein kinases in the murine T cell line EL4.

    Science.gov (United States)

    Knop, J; Wesche, H; Lang, D; Martin, M U

    1998-10-01

    The association and activation of the IL-1 receptor-associated protein kinase (IRAK) to the IL-1 receptor complex is one of the earliest events detectable in IL-1 signal transduction. We generated permanent clones of the murine T cell line EL4 6.1 overexpressing human (h)IRAK to evaluate the role of this kinase in IL-1 signaling. Overexpression of hIRAK enhanced IL-1-stimulated activation of the transcription factor NFkappaB, whereas a truncated form (N-IRAK) specifically inhibited IL-1-dependent NFkappaB activity. In clones stably overexpressing hIRAK a weak constitutive activation of NFkappaB correlated with a low basal IL-2 production which was enhanced in an IL-1-dependent manner. Compared to the parental cell line the dose-response curve of IL-1-induced IL-2 production was shifted in both potency and efficacy. These results demonstrate that IRAK directly triggers NFkappaB-mediated gene expression in EL4 cells. Qualitatively different effects were observed for the IL-1-induced activation of stress-activated protein (SAP) kinases: permanent overexpression of IRAK did not affect the dose dependence but prolonged the kinetics of IL-1-induced activation of SAP kinases, suggesting that this signaling branch may be regulated by distinct mechanisms.

  9. Activated platelet-derived growth factor β receptor and Ras-mitogen-activated protein kinase pathway in natural bovine urinary bladder carcinomas.

    Science.gov (United States)

    Corteggio, Annunziata; Di Geronimo, Ornella; Roperto, Sante; Roperto, Franco; Borzacchiello, Giuseppe

    2012-03-01

    Bovine papillomavirus types 1 or 2 (BPV-1/2) are involved in the aetiopathogenesis of bovine urinary bladder cancer. BPV-1/2 E5 activates the platelet-derived growth factor β receptor (PDGFβR). The aim of this study was to analyse the Ras/mitogen-activated protein kinase (MAPK) pathway in relation to activation of PDGFβR in natural bovine urinary bladder carcinomas. Co-immunoprecipitation and Western blot analysis demonstrated that recruitment of growth factor receptor bound protein 2 (GRB-2) and Sos-1 to the activated PDGFβR was increased in carcinomas compared to normal tissues. Higher grade bovine urinary bladder carcinomas were associated with activation of Ras, but not with activation of downstream mitogen-activated protein kinase/extracellular signal-regulated kinase (Mek 1/2) or extracellular signal-regulated kinase (Erk 1/2). Copyright © 2011 Elsevier Ltd. All rights reserved.

  10. MARK/Par1 Kinase Is Activated Downstream of NMDA Receptors through a PKA-Dependent Mechanism.

    Directory of Open Access Journals (Sweden)

    Laura P Bernard

    Full Text Available The Par1 kinases, also known as microtubule affinity-regulating kinases (MARKs, are important for the establishment of cell polarity from worms to mammals. Dysregulation of these kinases has been implicated in autism, Alzheimer's disease and cancer. Despite their important function in health and disease, it has been unclear how the activity of MARK/Par1 is regulated by signals from cell surface receptors. Here we show that MARK/Par1 is activated downstream of NMDA receptors in primary hippocampal neurons. Further, we show that this activation is dependent on protein kinase A (PKA, through the phosphorylation of Ser431 of Par4/LKB1, the major upstream kinase of MARK/Par1. Together, our data reveal a novel mechanism by which MARK/Par1 is activated at the neuronal synapse.

  11. Her4 and Her2/neu tyrosine kinase domains dimerize and activate in a reconstituted in vitro system.

    Science.gov (United States)

    Monsey, John; Shen, Wei; Schlesinger, Paul; Bose, Ron

    2010-03-05

    Her4 (ErbB-4) and Her2/neu (ErbB-2) are receptor-tyrosine kinases belonging to the epidermal growth factor receptor (EGFR) family. Crystal structures of EGFR and Her4 kinase domains demonstrate kinase dimerization and activation through an allosteric mechanism. The kinase domains form an asymmetric dimer, where the C-lobe surface of one monomer contacts the N-lobe of the other monomer. EGFR kinase dimerization and activation in vitro was previously reported using a nickel-chelating lipid-liposome system, and we now apply this system to all other members of the EGFR family. Polyhistidine-tagged Her4, Her2/neu, and Her3 kinase domains are bound to these nickel-liposomes and are brought to high local concentration, mimicking what happens to full-length receptors in vivo following ligand binding. Addition of nickel-liposomes to Her4 kinase domain results in 40-fold activation in kinase activity and marked enhancement of C-terminal tail autophosphorylation. Activation of Her4 shows a sigmoidal dependence on kinase concentration, consistent with a cooperative process requiring kinase dimerization. Her2/neu kinase activity is also activated by nickel-liposomes, and is increased further by heterodimerization with Her3 or Her4. The ability of Her3 and Her4 to heterodimerize and activate other family members is studied in vitro. Her3 kinase domain readily activates Her2/neu but is a poor activator of Her4, which differs from the prediction made by the asymmetric dimer model. Mutation of Her3 residues (952)ENI(954) to the corresponding sequence in Her4 enhanced the ability of Her3 to activate Her4, demonstrating that sequence differences on the C-lobe surface influence the heterodimerization and activation of ErbB kinase domains.

  12. Activation of AMP-activated protein kinase by tributyltin induces neuronal cell death

    International Nuclear Information System (INIS)

    Nakatsu, Yusuke; Kotake, Yaichiro; Hino, Atsuko; Ohta, Shigeru

    2008-01-01

    AMP-activated protein kinase (AMPK), a member of the metabolite-sensing protein kinase family, is activated by energy deficiency and is abundantly expressed in neurons. The environmental pollutant, tributyltin chloride (TBT), is a neurotoxin, and has been reported to decrease cellular ATP in some types of cells. Therefore, we investigated whether TBT activates AMPK, and whether its activation contributes to neuronal cell death, using primary cultures of cortical neurons. Cellular ATP levels were decreased 0.5 h after exposure to 500 nM TBT, and the reduction was time-dependent. It was confirmed that most neurons in our culture system express AMPK, and that TBT induced phosphorylation of AMPK. Compound C, an AMPK inhibitor, reduced the neurotoxicity of TBT, suggesting that AMPK is involved in TBT-induced cell death. Next, the downstream target of AMPK activation was investigated. Nitric oxide synthase, p38 phosphorylation and Akt dephosphorylation were not downstream of TBT-induced AMPK activation because these factors were not affected by compound C, but glutamate release was suggested to be controlled by AMPK. Our results suggest that activation of AMPK by TBT causes neuronal death through mediating glutamate release

  13. Sex-specific antidepressant effects of dietary creatine with and without sub-acute fluoxetine in rats

    Science.gov (United States)

    Allen, Patricia J.; D'Anci, Kristen E.; Kanarek, Robin B.; Renshaw, Perry F.

    2013-01-01

    The potential role of metabolic impairments in the pathophysiology of depression is motivating researchers to evaluate the treatment efficacy of creatine, a naturally occurring energetic and neuroprotective compound found in brain and muscle tissues. Growing evidence is demonstrating the benefit of oral creatine supplements for reducing depressive symptoms in humans and animals. A novel question is whether dietary creatine, when combined with antidepressant drug therapy, would be more effective than either compound alone. To answer this question, four studies were conducted to investigate the behavioral effects of combined creatine and low-dose fluoxetine treatment using the forced swim test in male and female rats. Sprague-Dawley rats were fed powdered rodent chow supplemented with 0%, 2% or 4% w/w creatine monohydrate for 5 weeks. Rats were injected with fluoxetine (5.0 or 10.0 mg/kg) or saline according to a sub-acute dosing schedule. Female rats maintained on a 4% creatine diet displayed antidepressant-like effects compared to non-supplemented females prior to fluoxetine treatment. In contrast, creatine did not alter behavior reliably in males. Following drug treatment and a second forced swim trial, the antidepressant-like profile of creatine remained significant only in females co-administered 5.0 mg/kg fluoxetine. Moreover, in females only, supplementation with 4% creatine produced a more robust antidepressant-like behavioral profile compared to either dose of fluoxetine alone. Estrous cycle data indicated that ovarian hormones influenced the antidepressant-like effects of creatine. Addressing the issue of sex differences in response to treatment may affect our understanding of creatine, its relationship with depressive behavior, and may lead to sex-specific therapeutic strategies. PMID:22429992

  14. A role for Pyk2 and Src in linking G-protein-coupled receptors with MAP kinase activation.

    Science.gov (United States)

    Dikic, I; Tokiwa, G; Lev, S; Courtneidge, S A; Schlessinger, J

    1996-10-10

    The mechanisms by which mitogenic G-protein-coupled receptors activate the MAP kinase signalling pathway are poorly understood. Candidate protein tyrosine kinases that link G-protein-coupled receptors with MAP kinase include Src family kinases, the epidermal growth factor receptor, Lyn and Syk. Here we show that lysophosphatidic acid (LPA) and bradykinin induce tyrosine phosphorylation of Pyk2 and complex formation between Pyk2 and activated Src. Moreover, tyrosine phosphorylation of Pyk2 leads to binding of the SH2 domain of Src to tyrosine 402 of Pyk2 and activation of Src. Transient overexpression of a dominant interfering mutant of Pyk2 or the protein tyrosine kinase Csk reduces LPA- or bradykinin-induced activation of MAP kinase. LPA- or bradykinin-induced MAP kinase activation was also inhibited by overexpression of dominant interfering mutants of Grb2 and Sos. We propose that Pyk2 acts with Src to link Gi- and Gq-coupled receptors with Grb2 and Sos to activate the MAP kinase signalling pathway in PC12 cells.

  15. Creatine supplementation with methylglyoxal: a potent therapy for cancer in experimental models.

    Science.gov (United States)

    Pal, Aparajita; Roy, Anirban; Ray, Manju

    2016-08-01

    The anti-cancer effect of methylglyoxal (MG) is now well established in the literature. The main aim of this study was to investigate the effect of creatine as a supplement in combination with MG both in vitro and in vivo. In case of the in vitro studies, two different cell lines, namely MCF-7 (human breast cancer cell line) and C2C12 (mouse myoblast cell line) were chosen. MG in combination with creatine showed enhanced apoptosis as well as higher cytotoxicity in the breast cancer MCF-7 cell line, compared to MG alone. Pre-treatment of well-differentiated C2C12 myotubes with cancerogenic 3-methylcholanthrene (3MC) induced a dedifferentiation of these myotubes towards cancerous cells (that mimic the effect of 3MC observed in solid fibro-sarcoma animal models) and subsequent exposure of these induced cancer cells with MG proved to be cytotoxic. Thus, creatine plus ascorbic acid enhanced the anti-cancer effects of MG. In contrast, when normal C2C12 muscle cells or myotubes (mouse normal myoblast cell line) were treated with MG or MG plus creatine and ascorbic acid, no detrimental effects were seen. This indicated that cytotoxic effects of MG are specifically limited towards cancer cells and are further enhanced when MG is used in combination with creatine and ascorbic acid. For the in vivo studies, tumors were induced by injecting Sarcoma-180 cells (2 × 10(6) cells/mouse) in the left hind leg. After 7 days of tumor inoculation, treatments were started with MG (20 mg/kg body wt/day, via the intravenous route), with or without creatine (150 mg/kg body wt/day, fed orally) and ascorbic acid (50 mg/kg body wt/day, fed orally) and continued for 10 consecutive days. Significant regression of tumor size was observed when Sarcoma-180 tumor-bearing mice were treated with MG and even more so with the aforesaid combination. The creatine-supplemented group demonstrated better overall survival in comparison with tumor-bearing mice without creatine. In conclusion, it may be

  16. Loss of ATM kinase activity leads to embryonic lethality in mice.

    Science.gov (United States)

    Daniel, Jeremy A; Pellegrini, Manuela; Lee, Baeck-Seung; Guo, Zhi; Filsuf, Darius; Belkina, Natalya V; You, Zhongsheng; Paull, Tanya T; Sleckman, Barry P; Feigenbaum, Lionel; Nussenzweig, André

    2012-08-06

    Ataxia telangiectasia (A-T) mutated (ATM) is a key deoxyribonucleic acid (DNA) damage signaling kinase that regulates DNA repair, cell cycle checkpoints, and apoptosis. The majority of patients with A-T, a cancer-prone neurodegenerative disease, present with null mutations in Atm. To determine whether the functions of ATM are mediated solely by its kinase activity, we generated two mouse models containing single, catalytically inactivating point mutations in Atm. In this paper, we show that, in contrast to Atm-null mice, both D2899A and Q2740P mutations cause early embryonic lethality in mice, without displaying dominant-negative interfering activity. Using conditional deletion, we find that the D2899A mutation in adult mice behaves largely similar to Atm-null cells but shows greater deficiency in homologous recombination (HR) as measured by hypersensitivity to poly (adenosine diphosphate-ribose) polymerase inhibition and increased genomic instability. These results may explain why missense mutations with no detectable kinase activity are rarely found in patients with classical A-T. We propose that ATM kinase-inactive missense mutations, unless otherwise compensated for, interfere with HR during embryogenesis.

  17. Involvement of PI3K/Akt Signaling Pathway and Its Downstream Intracellular Targets in the Antidepressant-Like Effect of Creatine.

    Science.gov (United States)

    Cunha, Mauricio P; Budni, Josiane; Ludka, Fabiana K; Pazini, Francis L; Rosa, Julia Macedo; Oliveira, Ágatha; Lopes, Mark W; Tasca, Carla I; Leal, Rodrigo B; Rodrigues, Ana Lúcia S

    2016-07-01

    Creatine has been proposed to exert beneficial effects in the management of depression, but the cell signaling pathways implicated in its antidepressant effects are not well established. This study investigated the involvement of PI3K/Akt signaling pathway and its downstream intracellular targets in the antidepressant-like effect of creatine. The acute treatment of mice with creatine (1 mg/kg, po) increased the Akt and P70S6K phosphorylation, and HO-1, GPx and PSD95 immunocontents. The pretreatment of mice with LY294002 (10 nmol/mouse, icv, PI3K inhibitor), wortmannin (0.1 μg/mouse, icv, PI3K inhibitor), ZnPP (10 μg/mouse, icv, HO-1 inhibitor), or rapamycin (0.2 nmol/mouse, icv, mTOR inhibitor) prevented the antidepressant-like effect of creatine (1 mg/kg, po) in the TST. In addition, the administration of subeffective dose of either the selective GSK3 inhibitor AR-A014418 (0.01 μg/mouse, icv), the nonselective GSK3 inhibitor lithium chloride (10 mg/kg, po), or the HO-1 inductor CoPP (0.01 μg/mouse, icv), in combination with a subeffective dose of creatine (0.01 mg/kg, po) reduced the immobility time in the TST as compared with either drug alone. No treatment caused significant changes in the locomotor activity of mice. These results indicate that the antidepressant-like effect of creatine in the TST depends on the activation of Akt, Nrf2/HO-1, GPx, and mTOR, and GSK3 inhibition.

  18. Protein tyrosine kinase and mitogen-activated protein kinase signalling pathways contribute to differences in heterophil-mediated innate immune responsiveness between two lines of broilers

    Science.gov (United States)

    Protein tyrosine phosphorylation mediates signal transduction of cellular processes, with protein tyrosine kinases (PTKs) regulating virtually all signaling events. The mitogen-activated protein kinase (MAPK) super-family consists of three conserved pathways that convert receptor activation into ce...

  19. Polarization of migrating monocytic cells is independent of PI 3-kinase activity.

    Directory of Open Access Journals (Sweden)

    Silvia Volpe

    Full Text Available BACKGROUND: Migration of mammalian cells is a complex cell type and environment specific process. Migrating hematopoietic cells assume a rapid amoeboid like movement when exposed to gradients of chemoattractants. The underlying signaling mechanisms remain controversial with respect to localization and distribution of chemotactic receptors within the plasma membrane and the role of PI 3-kinase activity in cell polarization. METHODOLOGY/PRINCIPAL FINDINGS: We present a novel model for the investigation of human leukocyte migration. Monocytic THP-1 cells transfected with the alpha(2A-adrenoceptor (alpha(2AAR display comparable signal transduction responses, such as calcium mobilization, MAP-kinase activation and chemotaxis, to the noradrenaline homologue UK 14'304 as when stimulated with CCL2, which binds to the endogenous chemokine receptor CCR2. Time-lapse video microscopy reveals that chemotactic receptors remain evenly distributed over the plasma membrane and that their internalization is not required for migration. Measurements of intramolecular fluorescence resonance energy transfer (FRET of alpha(2AAR-YFP/CFP suggest a uniform activation of the receptors over the entire plasma membrane. Nevertheless, PI 3-kinase activation is confined to the leading edge. When reverting the gradient of chemoattractant by moving the dispensing micropipette, polarized monocytes--in contrast to neutrophils--rapidly flip their polarization axis by developing a new leading edge at the previous posterior side. Flipping of the polarization axis is accompanied by re-localization of PI-3-kinase activity to the new leading edge. However, reversal of the polarization axis occurs in the absence of PI 3-kinase activation. CONCLUSIONS/SIGNIFICANCE: Accumulation and internalization of chemotactic receptors at the leading edge is dispensable for cell migration. Furthermore, uniformly distributed receptors allow the cells to rapidly reorient and adapt to changes in the

  20. Magnolol Alleviates Inflammatory Responses and Lipid Accumulation by AMP-Activated Protein Kinase-Dependent Peroxisome Proliferator-Activated Receptor α Activation

    Directory of Open Access Journals (Sweden)

    Ye Tian

    2018-02-01

    Full Text Available Magnolol (MG is a kind of lignin isolated from Magnolia officinalis, which serves several different biological functions, such as antifungal, anticancer, antioxidant, and hepatoprotective functions. This study aimed to evaluate the protective effect of MG against oleic acid (OA-induced hepatic steatosis and inflammatory damage in HepG2 cells and in a tyloxapol (Ty-induced hyperlipidemia mouse model. Our findings indicated that MG can effectively inhibit OA-stimulated tumor necrosis factor α (TNF-α secretion, reactive oxygen species generation, and triglyceride (TG accumulation. Further study manifested that MG significantly suppressed OA-activated mitogen-activated protein kinase (MAPK and nuclear factor-kappa B (NF-κB signaling pathways and that these inflammatory responses can be negated by pretreatment with inhibitors of extracellular regulated protein kinase and c-Jun N-terminal kinase (U0126 and SP600125, respectively. In addition, MG dramatically upregulated peroxisome proliferator-activated receptor α (PPARα translocation and reduced sterol regulatory element-binding protein 1c (SREBP-1c protein synthesis and excretion, both of which are dependent upon the phosphorylation of adenosine monophosphate (AMP-activated protein kinase (AMPK, acetyl-CoA carboxylase, and AKT kinase (AKT. However, MG suspended the activation of PPARα expression and was thus blocked by pretreatment with LY294002 and compound c (specific inhibitors of AKT and AMPK. Furthermore, MG clearly alleviated serum TG and total cholesterol release; upregulated AKT, AMPK, and PPARα expression; suppressed SREBP-1c generation; and alleviated hepatic steatosis and dyslipidemia in Ty-induced hyperlipidemia mice. Taken together, these results suggest that MG exerts protective effects against steatosis, hyperlipidemia, and the underlying mechanism, which may be closely associated with AKT/AMPK/PPARα activation and MAPK/NF-κB/SREBP-1c inhibition.

  1. Differential sensitivity of Src-family kinases to activation by SH3 domain displacement.

    Directory of Open Access Journals (Sweden)

    Jamie A Moroco

    Full Text Available Src-family kinases (SFKs are non-receptor protein-tyrosine kinases involved in a variety of signaling pathways in virtually every cell type. The SFKs share a common negative regulatory mechanism that involves intramolecular interactions of the SH3 domain with the PPII helix formed by the SH2-kinase linker as well as the SH2 domain with a conserved phosphotyrosine residue in the C-terminal tail. Growing evidence suggests that individual SFKs may exhibit distinct activation mechanisms dictated by the relative strengths of these intramolecular interactions. To elucidate the role of the SH3:linker interaction in the regulation of individual SFKs, we used a synthetic SH3 domain-binding peptide (VSL12 to probe the sensitivity of downregulated c-Src, Hck, Lyn and Fyn to SH3-based activation in a kinetic kinase assay. All four SFKs responded to VSL12 binding with enhanced kinase activity, demonstrating a conserved role for SH3:linker interaction in the control of catalytic function. However, the sensitivity and extent of SH3-based activation varied over a wide range. In addition, autophosphorylation of the activation loops of c-Src and Hck did not override regulatory control by SH3:linker displacement, demonstrating that these modes of activation are independent. Our results show that despite the similarity of their downregulated conformations, individual Src-family members show diverse responses to activation by domain displacement which may reflect their adaptation to specific signaling environments in vivo.

  2. Preparation of sup 125 I-creatine phosphokinase-MM

    Energy Technology Data Exchange (ETDEWEB)

    Jingxian, Su; Jingmin, Ma [Academia Sinica, Beijing, BJ (China). Inst. of Atomic Energy

    1988-09-01

    {sup 125}I-creatine phosphokinase-MM ({sup 125}I-CPK-MM) was prepared by {sup 125}I-labelled Bolton-Hunter reagent (HPNS). Iodinating conditions of HPNS and its conjugation to protein were studied. {sup 125}I-CPK-MM with immune activity was obtained and used to establish the {sup 125}I-CPK-MM radioimmunoassay method by the General Hospital of PLA. {sup 125}I-CPK-MM in PBS-G solution containing 0.015 mol/l ethyl mercaptan at 4-10 deg C can be used for one month.

  3. Crystal Structure of Ripk4 Reveals Dimerization-Dependent Kinase Activity.

    Science.gov (United States)

    Huang, Christine S; Oberbeck, Nina; Hsiao, Yi-Chun; Liu, Peter; Johnson, Adam R; Dixit, Vishva M; Hymowitz, Sarah G

    2018-05-01

    Receptor-interacting protein kinase 4 (RIPK4) is a highly conserved regulator of epidermal differentiation. Members of the RIPK family possess a common kinase domain as well as unique accessory domains that likely dictate subcellular localization and substrate preferences. Mutations in human RIPK4 manifest as Bartsocas-Papas syndrome (BPS), a genetic disorder characterized by severe craniofacial and limb abnormalities. We describe the structure of the murine Ripk4 (MmRipk4) kinase domain, in ATP- and inhibitor-bound forms. The crystallographic dimer of MmRipk4 is similar to those of RIPK2 and BRAF, and we show that the intact dimeric entity is required for MmRipk4 catalytic activity through a series of engineered mutations and cell-based assays. We also assess the impact of BPS mutations on protein structure and activity to elucidate the molecular origins of the disease. Copyright © 2018 Elsevier Ltd. All rights reserved.

  4. Selective blockade of protein kinase B protects the rat and human myocardium against ischaemic injury

    Science.gov (United States)

    Linares-Palomino, José; Husainy, Muhammad A; Lai, Vien K; Dickenson, John M; Galiñanes, Manuel

    2010-01-01

    Protein kinase B (PKB/Akt) plays a critical role in cell survival but the investigation of its involvement has been limited by the lack of specific pharmacological agents. In this study, using novel PKB inhibitors (VIII and XI), we investigated the role of PKB in cardioprotection of the rat and human myocardium, the location of PKB in relation to mitoKATP channels and p38 mitogen-activated protein kinase (p38 MAPK), and whether the manipulation of PKB can overcome the unresponsiveness to protection of the diabetic myocardium. Myocardial slices from rat left ventricle and from the right atrial appendage of patients undergoing elective cardiac surgery were subjected to 90 min ischaemia/120 min reoxygenation at 37°C. Tissue injury was assessed by creatine kinase (CK) released and determination of cell necrosis and apoptosis. The results showed that blockade of PKB activity caused significant reduction of CK release and cell death, a benefit that was as potent as ischaemic preconditioning and could be reproduced by blockade of phosphatidylinositol 3-kinase (PI-3K) with wortmannin and LY 294002. The protection was time dependent with maximal benefit seen when PKB and PI-3K were inhibited before ischaemia or during both ischaemia and reoxygenation. In addition, it was revealed that PKB is located downstream of mitoKATP channels but upstream of p38 MAPK. PKB inhibition induced a similar degree of protection in the human and rat myocardium and, importantly, it reversed the unresponsiveness to protection of the diabetic myocardium. In conclusion, inhibition of PKB plays a critical role in protection of the mammalian myocardium and may represent a clinical target for the reduction of ischaemic injury. PMID:20403980

  5. Expression and Purification of PI3 Kinase {alpha} and Development of an ATP Depletion and an AlphaScreen PI3 Kinase Activity Assay

    DEFF Research Database (Denmark)

    Boldyreff, Brigitte; Rasmussen, Tine L; Jensen, Hans H

    2008-01-01

    Phosphoinositide-3-kinases are important targets for drug development because many proteins in the PI3 kinase signaling pathway are mutated, hyperactivated, or overexpressed in human cancers. Here, the authors coexpressed the human class Ia PI3 kinase p110alpha catalytic domain with an N-terminal....... In parallel, a second assay format using the AlphaScreen technology was optimized to measure PI3 kinase activity. Both assay formats used should be suitable for high-throughput screening for the identification of PI3 kinase inhibitors. (Journal of Biomolecular Screening XXXX:xx-xx)....

  6. Low dose creatine supplementation enhances sprint phase of 400 meters swimming performance.

    Science.gov (United States)

    Anomasiri, Wilai; Sanguanrungsirikul, Sompol; Saichandee, Pisut

    2004-09-01

    This study demonstrated the effect of low dose creatine supplement (10 g. per day) on the sprinting time in the last 50 meters of 400 meters swimming competition, as well as the effect on exertion. Nineteen swimmers in the experimental group received creatine monohydrate 5 g with orange solution 15 g, twice per day for 7 days and nineteen swimmers in the control group received the same quantity of orange solution. The results showed that the swimmers who received creatine supplement lessened the sprinting time in the last 50 meters of 400 meters swimming competition than the control group. (p<0.05). The results of Wingate test (anaerobic power, anaerobic capacity and fatigue index) compared between pre and post supplementation. There was significant difference at p<0.05 in the control group from training effect whereas there was significant difference at p<0.000 from training effect and creatine supplement in the experiment group. Therefore, the creatine supplement in amateur swimmers in the present study enhanced the physical performance up to the maximum capacity.

  7. Kinase Activity Studied in Living Cells Using an Immunoassay

    Science.gov (United States)

    Bavec, Aljos?a

    2014-01-01

    This laboratory exercise demonstrates the use of an immunoassay for studying kinase enzyme activity in living cells. The advantage over the classical method, in which students have to isolate the enzyme from cell material and measure its activity in vitro, is that enzyme activity is modulated and measured in living cells, providing a more…

  8. The CREST-E study of creatine for Huntington disease: A randomized controlled trial.

    Science.gov (United States)

    Hersch, Steven M; Schifitto, Giovanni; Oakes, David; Bredlau, Amy-Lee; Meyers, Catherine M; Nahin, Richard; Rosas, Herminia Diana

    2017-08-08

    To investigate whether creatine administration could slow progressive functional decline in adults with early symptoms of Huntington disease. We conducted a multicenter, randomized, double-blind, placebo-controlled study of up to 40 g daily of creatine monohydrate in participants with stage I and II HD treated for up to 48 months. The primary outcome measure was the rate of change in total functional capacity (TFC) between baseline and end of follow-up. Secondary outcome measures included changes in additional clinical scores, tolerability, and quality of life. Safety was assessed by adverse events and laboratory studies. At 46 sites in North America, Australia, and New Zealand, 553 participants were randomized to creatine (275) or placebo (278). The trial was designed to enroll 650 patients, but was halted for futility after the first interim analysis. The estimated rates of decline in the primary outcome measure (TFC) were 0.82 points per year for participants on creatine, 0.70 points per year for participants on placebo, favoring placebo (nominal 95% confidence limits -0.11 to 0.35). Adverse events, mainly gastrointestinal, were significantly more common in participants on creatine. Serious adverse events, including deaths, were more frequent in the placebo group. Subgroup analysis suggested that men and women may respond differently to creatine treatment. Our data do not support the use of creatine treatment for delaying functional decline in early manifest HD. NCT00712426. This study provides Class II evidence that for patients with early symptomatic HD, creatine monohydrate is not beneficial for slowing functional decline. Copyright © 2017 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.

  9. Effects of creatine supplementation along with resistance training on urinary formaldehyde and serum enzymes in wrestlers.

    Science.gov (United States)

    Nasseri, Azadeh; Jafari, Afshar

    2016-04-01

    Formaldehyde is a cytotoxic agent produced from creatine through a metabolic pathway, and in this regard, it has been claimed that creatine supplementation could be cytotoxic. Even though the cytotoxic effects of creatine supplementation have been widely studied, yet little is known about how resistance training can alter these toxic effects. This study aimed to determine the effects of short-term creatine supplementation plus resistance training on the level of urinary formaldehyde and concentrations of serum enzymes in young male wrestlers. In a double-blind design twenty-one subjects were randomized into creatine supplementation (Cr), creatine supplementation plus resistance training (Cr + T) and placebo plus resistance training (Pl + T) groups. Participants ingested creatine (0.3 g/kg/day) or placebo for 7 days. The training protocol consisted of 3 sessions in one week, each session including three sets of 6-9 repetitions at 80-85% of one-repetition maximum for whole-body exercise. Urine and blood samples were collected at baseline and at the end of the supplementation. Creatine supplementation significantly increased the excretion rate of urinary formaldehyde in the Cr and Cr + T groups by 63.4% and 30.4%, respectively (P0.05). These findings indicate that resistance training may lower the increase of urinary formaldehyde excretion induced by creatine supplementation, suggesting that creatine consumption could be relatively less toxic when combined with resistance training.

  10. Oral creatine supplementation attenuates muscle loss caused by limb immobilization: a systematic review

    Directory of Open Access Journals (Sweden)

    Camila Souza Padilha

    Full Text Available Abstract Introduction: Recent studies have pointing creatine supplementation as a promising therapeutic alterna- tive in several diseases, especially myopathies and neurodegenerative disorder. Objective: elucidate the role of creatine supplementation on deleterious effect caused by limb immobilization in humans and rats. Methods: Analyzed articles were searched by three online databases, PubMed, SportDicus e Scielo. After a review and analysis, the studies were included in this review articles on effect of creatine supplementation on skeletal muscle in humans and rat, before, during and after a period of limb immobilization. Results: Studies analyzed demonstrated positive points in use of creatine supplementation as a therapeutic tool to mitigating the deleterious effects of limb immobilization, in humans and rat. Conclusion: The dataset of this literature review allows us to conclude that creatine supplementation may reduce muscle loss and/or assist in the recovery of muscle atrophy caused by immobilization and disuse in rats and humans. Also, we note that further research with better methodological rigor is needed to clarify the mechanisms by which creatine support the recovery of muscle atrophy. Moreover, these effects are positive and promising in the field of muscle rehabilitation, especially after member’s immobilization.

  11. Total body skeletal muscle mass: estimation by creatine (methyl-d3) dilution in humans

    Science.gov (United States)

    Walker, Ann C.; O'Connor-Semmes, Robin L.; Leonard, Michael S.; Miller, Ram R.; Stimpson, Stephen A.; Turner, Scott M.; Ravussin, Eric; Cefalu, William T.; Hellerstein, Marc K.; Evans, William J.

    2014-01-01

    Current methods for clinical estimation of total body skeletal muscle mass have significant limitations. We tested the hypothesis that creatine (methyl-d3) dilution (D3-creatine) measured by enrichment of urine D3-creatinine reveals total body creatine pool size, providing an accurate estimate of total body skeletal muscle mass. Healthy subjects with different muscle masses [n = 35: 20 men (19–30 yr, 70–84 yr), 15 postmenopausal women (51–62 yr, 70–84 yr)] were housed for 5 days. Optimal tracer dose was explored with single oral doses of 30, 60, or 100 mg D3-creatine given on day 1. Serial plasma samples were collected for D3-creatine pharmacokinetics. All urine was collected through day 5. Creatine and creatinine (deuterated and unlabeled) were measured by liquid chromatography mass spectrometry. Total body creatine pool size and muscle mass were calculated from D3-creatinine enrichment in urine. Muscle mass was also measured by magnetic resonance imaging (MRI), dual-energy x-ray absorptiometry (DXA), and traditional 24-h urine creatinine. D3-creatine was rapidly absorbed and cleared with variable urinary excretion. Isotopic steady-state of D3-creatinine enrichment in the urine was achieved by 30.7 ± 11.2 h. Mean steady-state enrichment in urine provided muscle mass estimates that correlated well with MRI estimates for all subjects (r = 0.868, P creatine dose determined by urine D3-creatinine enrichment provides an estimate of total body muscle mass strongly correlated with estimates from serial MRI with less bias than total lean body mass assessment by DXA. PMID:24764133

  12. CARDIOPROTECTIVE EFFECT OF ESCULETIN ON CARDIAC MARKER ENZYMES AND MEMRANE BOUND ENZYMES IN ISOPROTERENOL-INDUCED MYOCARDIAL INFARCTION IN WISTAR RATS

    OpenAIRE

    Palanivel Karthika; Murugan Rajadurai; Palanisamy Ganapathy; Ganesan Kanchana

    2011-01-01

    This study evaluates the cardioprotective effect of esculetin on isoproterenol (ISO)-induced myocardial infarction (MI) in rats. Rats were pretreated with esculetin (10 and 20 mg/kg) orally for a period of 21 days. After the treatment period ISO (85 mg/kg) was administered subcutaneously to rats at an interval of 24 h for 2 days. ISO-induced rats showed a significant increase in the activities of marker enzymes such as creatine kinase (CK), creatine kinase-MB (CK-MB), aspartate transaminase (...

  13. Effects of Sesame (Sesamum indicum L.) Supplementation on Creatine Kinase, Lactate Dehydrogenase, Oxidative Stress Markers, and Aerobic Capacity in Semi-Professional Soccer Players.

    Science.gov (United States)

    Barbosa, Carlos V da Silva; Silva, Alexandre S; de Oliveira, Caio V C; Massa, Nayara M L; de Sousa, Yasmim R F; da Costa, Whyara K A; Silva, Ayice C; Delatorre, Plínio; Carvalho, Rhayane; Braga, Valdir de Andrade; Magnani, Marciane

    2017-01-01

    Nutritional intervention with antioxidants rich foods has been considered a strategy to minimize the effects of overtraining in athletes. This experimental, randomized, and placebo-controlled study evaluated the effects of consumption of sesame ( Sesamum indicum L.) on muscle damage markers, oxidative stress, systemic inflammation, and aerobic performance in male semi-professional soccer players. Twenty athletes were randomly assigned to groups that received 40 g (two tablespoons) per day of sesame or a placebo during 28 days of regular training (exposed to routine training that includes loads of heavy training in the final half of the season). Before and after intervention, creatine kinase (CK), lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD), C-reactive protein (hs-CRP), and aerobic capacity were evaluated. Before intervention, a physiologic imbalance was noted in both groups related to CK and LDH levels. Sesame intake caused a reduction of CK (19%, p < 0.05), LDH (37%, p < 0.05), MDA (55%, p < 0.05) and hs-CRP (53%, p < 0.05) and increased SOD (14%, p < 0.05), vitamin A (25%, p < 0.05), and vitamin E (65%, p < 0.05) in the experimental group. These phenomena were accompanied by increased aerobic capacity (17%, p < 0.05). The placebo group showed an increase in CK (5%, p < 0.05) and no significant change in LDH, SOD or vitamin A. MDA levels decreased (21%, p < 0.05) and vitamin E increased (14%, p < 0.05) in the placebo group, but to a much lesser extent than in the experimental group. These results show that sesame consumption may reduce muscle damage and oxidative stress while improving the aerobic capacity in soccer players.

  14. Calcium is the switch in the moonlighting dual function of the ligand-activated receptor kinase phytosulfokine receptor 1

    KAUST Repository

    Muleya, Victor

    2014-09-23

    Background: A number of receptor kinases contain guanylate cyclase (GC) catalytic centres encapsulated in the cytosolic kinase domain. A prototypical example is the phytosulfokine receptor 1 (PSKR1) that is involved in regulating growth responses in plants. PSKR1 contains both kinase and GC activities however the underlying mechanisms regulating the dual functions have remained elusive. Findings: Here, we confirm the dual activity of the cytoplasmic domain of the PSKR1 receptor. We show that mutations within the guanylate cyclase centre modulate the GC activity while not affecting the kinase catalytic activity. Using physiologically relevant Ca2+ levels, we demonstrate that its GC activity is enhanced over two-fold by Ca2+ in a concentration-dependent manner. Conversely, increasing Ca2+ levels inhibits kinase activity up to 500-fold at 100 nM Ca2+. Conclusions: Changes in calcium at physiological levels can regulate the kinase and GC activities of PSKR1. We therefore propose a functional model of how calcium acts as a bimodal switch between kinase and GC activity in PSKR1 that could be relevant to other members of this novel class of ligand-activated receptor kinases.

  15. A review of creatine supplementation in age-related diseases: more than a supplement for athletes

    Science.gov (United States)

    Smith, Rachel N.; Agharkar, Amruta S.; Gonzales, Eric B.

    2014-01-01

    Creatine is an endogenous compound synthesized from arginine, glycine and methionine. This dietary supplement can be acquired from food sources such as meat and fish, along with athlete supplement powders. Since the majority of creatine is stored in skeletal muscle, dietary creatine supplementation has traditionally been important for athletes and bodybuilders to increase the power, strength, and mass of the skeletal muscle. However, new uses for creatine have emerged suggesting that it may be important in preventing or delaying the onset of neurodegenerative diseases associated with aging. On average, 30% of muscle mass is lost by age 80, while muscular weakness remains a vital cause for loss of independence in the elderly population. In light of these new roles of creatine, the dietary supplement’s usage has been studied to determine its efficacy in treating congestive heart failure, gyrate atrophy, insulin insensitivity, cancer, and high cholesterol. In relation to the brain, creatine has been shown to have antioxidant properties, reduce mental fatigue, protect the brain from neurotoxicity, and improve facets/components of neurological disorders like depression and bipolar disorder. The combination of these benefits has made creatine a leading candidate in the fight against age-related diseases, such as Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, long-term memory impairments associated with the progression of Alzheimer’s disease, and stroke. In this review, we explore the normal mechanisms by which creatine is produced and its necessary physiology, while paying special attention to the importance of creatine supplementation in improving diseases and disorders associated with brain aging and outlining the clinical trials involving creatine to treat these diseases. PMID:25664170

  16. Navigating the conformational landscape of G protein-coupled receptor kinases during allosteric activation.

    Science.gov (United States)

    Yao, Xin-Qiu; Cato, M Claire; Labudde, Emily; Beyett, Tyler S; Tesmer, John J G; Grant, Barry J

    2017-09-29

    G protein-coupled receptors (GPCRs) are essential for transferring extracellular signals into carefully choreographed intracellular responses controlling diverse aspects of cell physiology. The duration of GPCR-mediated signaling is primarily regulated via GPCR kinase (GRK)-mediated phosphorylation of activated receptors. Although many GRK structures have been reported, the mechanisms underlying GRK activation are not well-understood, in part because it is unknown how these structures map to the conformational landscape available to this enzyme family. Unlike most other AGC kinases, GRKs rely on their interaction with GPCRs for activation and not phosphorylation. Here, we used principal component analysis of available GRK and protein kinase A crystal structures to identify their dominant domain motions and to provide a framework that helps evaluate how close each GRK structure is to being a catalytically competent state. Our results indicated that disruption of an interface formed between the large lobe of the kinase domain and the regulator of G protein signaling homology domain (RHD) is highly correlated with establishment of the active conformation. By introducing point mutations in the GRK5 RHD-kinase domain interface, we show with both in silico and in vitro experiments that perturbation of this interface leads to higher phosphorylation activity. Navigation of the conformational landscape defined by this bioinformatics-based study is likely common to all GPCR-activated GRKs. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Creatine Enhances Transdifferentiation of Bone Marrow Stromal Cell-Derived Neural Stem Cell Into GABAergic Neuron-Like Cells Characterized With Differential Gene Expression.

    Science.gov (United States)

    Darabi, Shahram; Tiraihi, Taki; Delshad, AliReza; Sadeghizadeh, Majid; Taheri, Taher; Hassoun, Hayder K

    2017-04-01

    Creatine was reported to induce bone marrow stromal cells (BMSC) into GABAergic neuron-like cells (GNLC). In a previous study, creatine was used as a single inducer for BMSC into GNLC with low yield. In this study, BMSC-derived neurospheres (NS) have been used in generating GABAergic phenotype. The BMSC were isolated from adult rats and used in generating neurospheres and used for producing neural stem cells (NSC). A combination of all-trans-retinoic acid (RA), the ciliary neurotrophic factor (CNTF), and creatine was used in order to improve the yield of GNLC. We also used other protocols for the transdifferentiation including RA alone; RA and creatine; RA and CNTF; and RA, CNTF, and creatine. The BMSC, NSC, and GNLC were characterized by specific markers. The activity of the GNLC was evaluated using FM1-43. The isolated BMSC expressed Oct4, fibronectin, and CD44. The NS were immunoreactive to nestin and SOX2, the NSC were immunoreactive to nestin, NF68 and NF160, while the GNLC were immunoreactive to GAD1/2, VGAT, GABA, and synaptophysin. Oct4 and c-MYC, pluripotency genes, were expressed in the BMSC, while SOX2 and c-MYC were expressed in the NSC. The activity of GNLC indicates that the synaptic vesicles were released upon stimulation. The conclusion is that the combination of RA, CNTF, and creatine induced differentiation of neurosphere-derived NSC into GNLC within 1 week. This protocol gives higher yield than the other protocols used in this study. The mechanism of induction was clearly associated with several differential pluripotent genes.

  18. The Effects of Creatine Supplementation on Explosive Performance and Optimal Individual Postactivation Potentiation Time

    Directory of Open Access Journals (Sweden)

    Chia-Chi Wang

    2016-03-01

    Full Text Available Creatine plays an important role in muscle energy metabolism. Postactivation potentiation (PAP is a phenomenon that can acutely increase muscle power, but it is an individualized process that is influenced by muscle fatigue. This study examined the effects of creatine supplementation on explosive performance and the optimal individual PAP time during a set of complex training bouts. Thirty explosive athletes performed tests of back squat for one repetition maximum (1RM strength and complex training bouts for determining the individual optimal timing of PAP, height and peak power of a counter movement jump before and after the supplementation. Subjects were assigned to a creatine or placebo group and then consumed 20 g of creatine or carboxymethyl cellulose per day for six days. After the supplementation, the 1RM strength in the creatine group significantly increased (p < 0.05. The optimal individual PAP time in the creatine group was also significant earlier than the pre-supplementation and post-supplementation of the placebo group (p < 0.05. There was no significant difference in jump performance between the groups. This study demonstrates that creatine supplementation improves maximal muscle strength and the optimal individual PAP time of complex training but has no effect on explosive performance.

  19. The Axl kinase domain in complex with a macrocyclic inhibitor offers first structural insights into an active TAM receptor kinase.

    Science.gov (United States)

    Gajiwala, Ketan S; Grodsky, Neil; Bolaños, Ben; Feng, Junli; Ferre, RoseAnn; Timofeevski, Sergei; Xu, Meirong; Murray, Brion W; Johnson, Ted W; Stewart, Al

    2017-09-22

    The receptor tyrosine kinase family consisting of Tyro3, Axl, and Mer (TAM) is one of the most recently identified receptor tyrosine kinase families. TAM receptors are up-regulated postnatally and maintained at high levels in adults. They all play an important role in immunity, but Axl has also been implicated in cancer and therefore is a target in the discovery and development of novel therapeutics. However, of the three members of the TAM family, the Axl kinase domain is the only one that has so far eluded structure determination. To this end, using differential scanning fluorimetry and hydrogen-deuterium exchange mass spectrometry, we show here that a lower stability and greater dynamic nature of the Axl kinase domain may account for its poor crystallizability. We present the first structural characterization of the Axl kinase domain in complex with a small-molecule macrocyclic inhibitor. The Axl crystal structure revealed two distinct conformational states of the enzyme, providing a first glimpse of what an active TAM receptor kinase may look like and suggesting a potential role for the juxtamembrane region in enzyme activity. We noted that the ATP/inhibitor-binding sites of the TAM members closely resemble each other, posing a challenge for the design of a selective inhibitor. We propose that the differences in the conformational dynamics among the TAM family members could potentially be exploited to achieve inhibitor selectivity for targeted receptors. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Ca2+/Calmodulin-Dependent Protein Kinase Kinases (CaMKKs) Effects on AMP-Activated Protein Kinase (AMPK) Regulation of Chicken Sperm Functions.

    Science.gov (United States)

    Nguyen, Thi Mong Diep; Combarnous, Yves; Praud, Christophe; Duittoz, Anne; Blesbois, Elisabeth

    2016-01-01

    Sperm require high levels of energy to ensure motility and acrosome reaction (AR) accomplishment. The AMP-activated protein kinase (AMPK) has been demonstrated to be strongly involved in the control of these properties. We address here the question of the potential role of calcium mobilization on AMPK activation and function in chicken sperm through the Ca(2+)/calmodulin-dependent protein kinase kinases (CaMKKs) mediated pathway. The presence of CaMKKs and their substrates CaMKI and CaMKIV was evaluated by western-blotting and indirect immunofluorescence. Sperm were incubated in presence or absence of extracellular Ca(2+), or of CaMKKs inhibitor (STO-609). Phosphorylations of AMPK, CaMKI, and CaMKIV, as well as sperm functions were evaluated. We demonstrate the presence of both CaMKKs (α and β), CaMKI and CaMKIV in chicken sperm. CaMKKα and CaMKI were localized in the acrosome, the midpiece, and at much lower fluorescence in the flagellum, whereas CaMKKβ was mostly localized in the flagellum and much less in the midpiece and the acrosome. CaMKIV was only present in the flagellum. The presence of extracellular calcium induced an increase in kinases phosphorylation and sperm activity. STO-609 reduced AMPK phosphorylation in the presence of extracellular Ca(2+) but not in its absence. STO-609 did not affect CaMKIV phosphorylation but decreased CaMKI phosphorylation and this inhibition was quicker in the presence of extracellular Ca(2+) than in its absence. STO-609 efficiently inhibited sperm motility and AR, both in the presence and absence of extracellular Ca(2+). Our results show for the first time the presence of CaMKKs (α and β) and one of its substrate, CaMKI in different subcellular compartments in germ cells, as well as the changes in the AMPK regulation pathway, sperm motility and AR related to Ca(2+) entry in sperm through the Ca(2+)/CaM/CaMKKs/CaMKI pathway. The Ca(2+)/CaMKKs/AMPK pathway is activated only under conditions of extracellular Ca(2+) entry

  1. Ca2+/Calmodulin-Dependent Protein Kinase Kinases (CaMKKs Effects on AMP-Activated Protein Kinase (AMPK Regulation of Chicken Sperm Functions.

    Directory of Open Access Journals (Sweden)

    Thi Mong Diep Nguyen

    Full Text Available Sperm require high levels of energy to ensure motility and acrosome reaction (AR accomplishment. The AMP-activated protein kinase (AMPK has been demonstrated to be strongly involved in the control of these properties. We address here the question of the potential role of calcium mobilization on AMPK activation and function in chicken sperm through the Ca(2+/calmodulin-dependent protein kinase kinases (CaMKKs mediated pathway. The presence of CaMKKs and their substrates CaMKI and CaMKIV was evaluated by western-blotting and indirect immunofluorescence. Sperm were incubated in presence or absence of extracellular Ca(2+, or of CaMKKs inhibitor (STO-609. Phosphorylations of AMPK, CaMKI, and CaMKIV, as well as sperm functions were evaluated. We demonstrate the presence of both CaMKKs (α and β, CaMKI and CaMKIV in chicken sperm. CaMKKα and CaMKI were localized in the acrosome, the midpiece, and at much lower fluorescence in the flagellum, whereas CaMKKβ was mostly localized in the flagellum and much less in the midpiece and the acrosome. CaMKIV was only present in the flagellum. The presence of extracellular calcium induced an increase in kinases phosphorylation and sperm activity. STO-609 reduced AMPK phosphorylation in the presence of extracellular Ca(2+ but not in its absence. STO-609 did not affect CaMKIV phosphorylation but decreased CaMKI phosphorylation and this inhibition was quicker in the presence of extracellular Ca(2+ than in its absence. STO-609 efficiently inhibited sperm motility and AR, both in the presence and absence of extracellular Ca(2+. Our results show for the first time the presence of CaMKKs (α and β and one of its substrate, CaMKI in different subcellular compartments in germ cells, as well as the changes in the AMPK regulation pathway, sperm motility and AR related to Ca(2+ entry in sperm through the Ca(2+/CaM/CaMKKs/CaMKI pathway. The Ca(2+/CaMKKs/AMPK pathway is activated only under conditions of extracellular Ca(2

  2. Independence of protein kinase C-delta activity from activation loop phosphorylation: structural basis and altered functions in cells.

    Science.gov (United States)

    Liu, Yin; Belkina, Natalya V; Graham, Caroline; Shaw, Stephen

    2006-04-28

    Activation loop phosphorylation plays critical regulatory roles for many kinases. Unlike other protein kinase Cs (PKC), PKC-delta does not require phosphorylation of its activation loop (Thr-507) for in vitro activity. We investigated the structural basis for this unusual capacity and its relevance to PKC-delta function in intact cells. Mutational analysis demonstrated that activity without Thr-507 phosphorylation depends on 20 residues N-terminal to the kinase domain and a pair of phenylalanines (Phe-500/Phe-527) unique to PKC-delta in/near the activation loop. Molecular modeling demonstrated that these elements stabilize the activation loop by forming a hydrophobic chain of interactions from the C-lobe to activation loop to N-terminal (helical) extension. In cells PKC-delta mediates both apoptosis and transcription regulation. We found that the T507A mutant of the PKC-delta kinase domain resembled the corresponding wild type in mediating apoptosis in transfected HEK293T cells. But the T507A mutant was completely defective in AP-1 and NF-kappaB reporter assays. A novel assay in which the kinase domain of PKC-delta and its substrate (a fusion protein of PKC substrate peptide with green fluorescent protein) were co-targeted to lipid rafts revealed a major substrate-selective defect of the T507A mutant in phosphorylating the substrate in cells. In vitro analysis showed strong product inhibition on the T507A mutant with particular substrates whose characteristics suggest it contributes to the substrate selective defect of the PKC-delta T507A mutant in cells. Thus, activation loop phosphorylation of PKC-delta may regulate its function in cells in a novel way.

  3. SAM domain-dependent activity of PfTKL3, an essential tyrosine kinase-like kinase of the human malaria parasite Plasmodium falciparum.

    Science.gov (United States)

    Abdi, Abdirahman; Eschenlauer, Sylvain; Reininger, Luc; Doerig, Christian

    2010-10-01

    Over the last decade, several protein kinases inhibitors have reached the market for cancer chemotherapy. The kinomes of pathogens represent potentially attractive targets in infectious diseases. The functions of the majority of protein kinases of Plasmodium falciparum, the parasitic protist responsible for the most virulent form of human malaria, remain unknown. Here we present a thorough characterisation of PfTKL3 (PF13_0258), an enzyme that belongs to the tyrosine kinase-like kinase (TKL) group. We demonstrate by reverse genetics that PfTKL3 is essential for asexual parasite proliferation in human erythrocytes. PfTKL3 is expressed in both asexual and gametocytes stages, and in the latter the protein co-localises with cytoskeleton microtubules. Recombinant PfTKL3 displays in vitro autophosphorylation activity and is able to phosphorylate exogenous substrates, and both activities are dramatically dependent on the presence of an N-terminal "sterile alpha-motif" domain. This study identifies PfTKL3 as a validated drug target amenable to high-throughput screening.

  4. DNA Damage-Induced Acetylation of Lysine 3016 of ATM Activates ATM Kinase Activity▿ †

    OpenAIRE

    Sun, Yingli; Xu, Ye; Roy, Kanaklata; Price, Brendan D.

    2007-01-01

    The ATM protein kinase is essential for cells to repair and survive genotoxic events. The activation of ATM's kinase activity involves acetylation of ATM by the Tip60 histone acetyltransferase. In this study, systematic mutagenesis of lysine residues was used to identify regulatory ATM acetylation sites. The results identify a single acetylation site at lysine 3016, which is located in the highly conserved C-terminal FATC domain adjacent to the kinase domain. Antibodies specific for acetyl-ly...

  5. Concentrations in beef and lamb of taurine, carnosine, coenzyme Q(10), and creatine.

    Science.gov (United States)

    Purchas, R W; Rutherfurd, S M; Pearce, P D; Vather, R; Wilkinson, B H P

    2004-03-01

    Levels of taurine, carnosine, coenzyme Q(10), and creatine were measured in beef liver and several muscles of beef and lamb and in cooked and uncooked meat. The amino acid taurine has numerous biological functions, the dipeptide carnosine is a buffer as well as an antioxidant, coenzyme Q(10) is also an antioxidant present within mitochondria, and creatine along with creatine phosphate is involved with energy metabolism in muscle. Large differences were shown for all compounds between beef cheek muscle (predominantly red fibres) and beef semitendinosus muscle (mainly white fibres), with cheek muscle containing 9.9 times as much taurine, and 3.2 times as much coenzyme Q(10), but only 65% as much creatine and 9% as much carnosine. Levels in lamb relative to beef semitendinosus muscles were higher for taurine but slightly lower for carnosine, coenzyme Q(10) and creatine. Values for all the compounds varied significantly between eight lamb muscles, possibly due in part to differences in the proportion of muscle fibre types. Slow cooking (90 min at 70 °C) of lamb longissimus and semimembranosus muscles led to significant reductions in the content of taurine, carnosine, and creatine (Plamb, but that these levels vary between muscles, between animals, and with cooking.

  6. Effects of protein kinase C activators on phorbol ester-sensitive and -resistant EL4 thymoma cells.

    Science.gov (United States)

    Sansbury, H M; Wisehart-Johnson, A E; Qi, C; Fulwood, S; Meier, K E

    1997-09-01

    Phorbol ester-sensitive EL4 murine thymoma cells respond to phorbol 12-myristate 13-acetate with activation of ERK mitogen-activated protein kinases, synthesis of interleukin-2, and death, whereas phorbol ester-resistant variants of this cell line do not exhibit these responses. Additional aspects of the resistant phenotype were examined, using a newly-established resistant cell line. Phorbol ester induced morphological changes, ERK ac