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Sample records for dose increases leucine

  1. Increasing leucine concentration stimulates mechanistic target of rapamycin signaling and cell growth in C2C12 skeletal muscle cells.

    PubMed

    Areta, José L; Hawley, John A; Ye, Ji-Ming; Chan, M H Stanley; Coffey, Vernon G

    2014-11-01

    Leucine is a key amino acid for initiating translation in muscle cells, but the dose-dependent effects of leucine on intracellular signaling are poorly characterized. This study examined the effect that increasing doses of leucine would have on changes in mechanistic target of rapamycin (mTOR)-mediated signaling, rates of protein synthesis, and cell size in C2C12 cells. We hypothesized that a leucine "threshold" exists, which represents the minimum stimulus required to initiate mTOR signaling in muscle cells. Acute exposure to 1.5, 3.2, 5.0, and 16.1 mM leucine increased phosphorylation of mTOR(Ser2448) (~1.4-fold; P < .04), 4E-BP1 (Thr37/46) (~1.9-fold; P < .001), and rpS6(Ser235/6) (~2.3-fold; P < .001). However, only p70S6k(Thr389) exhibited a dose-dependent response to leucine with all treatments higher than control (~4-fold; P < .001) and at least 5 mM higher than the 1.5-mM concentration (1.2-fold; P < .02). Rates of protein synthesis were not altered by any treatment. Seven days of exposure to 0.5, 1.5, 5.0, and 16.5 mM leucine resulted in an increase in cell size in at least 5 mM treatments (~1.6-fold, P < .001 vs control). Our findings indicate that even at low leucine concentrations, phosphorylation of proteins regulating translation initiation signaling is enhanced. The phosphorylation of p70S6k(Thr389) follows a leucine dose-response relationship, although this was not reflected by the acute protein synthetic response. Nevertheless, under the conditions of the present study, it appears that leucine concentrations of at least 5 mM are necessary to enhance cell growth.

  2. Dose and Latency Effects of Leucine Supplementation in Modulating Glucose Homeostasis: Opposite Effects in Healthy and Glucocorticoid-Induced Insulin-Resistance States

    PubMed Central

    Zanchi, Nelo Eidy; Guimarães-Ferreira, Lucas; de Siqueira-Filho, Mário Alves; Felitti, Vitor; Nicastro, Humberto; Bueno, Carlos; Lira, Fábio Santos; Naimo, Marshall Alan; Campos-Ferraz, Patrícia; Nunes, Maria Tereza; Seelaender, Marília; de Oliveira Carvalho, Carla Roberta; Blachier, François; Lancha, Antonio Herbert

    2012-01-01

    Dexamethasone (DEXA) is a potent immunosupressant and anti-inflammatory agent whose main side effects are muscle atrophy and insulin resistance in skeletal muscles. In this context, leucine supplementation may represent a way to limit the DEXA side effects. In this study, we have investigated the effects of a low and a high dose of leucine supplementation (via a bolus) on glucose homeostasis, muscle mass and muscle strength in energy-restricted and DEXA-treated rats. Since the leucine response may also be linked to the administration of this amino acid, we performed a second set of experiments with leucine given in bolus (via gavage) versus leucine given via drinking water. Leucine supplementation was found to produce positive effects (e.g., reduced insulin levels) only when administrated in low dosage, both via the bolus or via drinking water. However, under DEXA treatment, leucine administration was found to significantly influence this response, since leucine supplementation via drinking water clearly induced a diabetic state, whereas the same effect was not observed when supplied via the gavage. PMID:23363994

  3. Increased IGFBP-1 phosphorylation in response to leucine deprivation is mediated by CK2 and PKC.

    PubMed

    Malkani, Niyati; Biggar, Kyle; Shehab, Majida Abu; Li, Shawn Shun-Cheng; Jansson, Thomas; Gupta, Madhulika B

    2016-04-15

    Insulin-like growth factor binding protein-1 (IGFBP-1), secreted by fetal liver, is a key regulator of IGF-I bioavailability and fetal growth. IGFBP-1 phosphorylation decreases IGF-I bioavailability and diminishes its growth-promoting effects. Growth-restricted fetuses have decreased levels of circulating essential amino acids. We recently showed that IGFBP-1 hyperphosphorylation (pSer101/119/169) in response to leucine deprivation is regulated via activation of the amino acid response (AAR) in HepG2 cells. Here we investigated nutrient-sensitive protein kinases CK2/PKC/PKA in mediating IGFBP-1 phosphorylation in leucine deprivation. We demonstrated that leucine deprivation stimulated CK2 activity (enzymatic assay) and induced IGFBP-1 phosphorylation (immunoblotting/MRM-MS). Inhibition (pharmacological/siRNA) of CK2/PKC, but not PKA, prevented IGFBP-1 hyperphosphorylation in leucine deprivation. PKC inhibition also prevented leucine deprivation-stimulated CK2 activity. Functionally, leucine deprivation decreased IGF-I-induced-IGF-1R autophosphorylation when CK2/PKC were not inhibited. Our data strongly support that PKC promotes leucine deprivation-induced IGFBP-1 hyperphosphorylation via CK2 activation, mechanistically linking decreased amino acid availability and reduced fetal growth.

  4. Comparison of the transcriptional responses of skeletal muscle and bone to a flooding dose of leucine in the gilthead sea bream (Sparus aurata).

    PubMed

    Garcia de la Serrana, Daniel; Mareco, Edson A; LA Vieira, Vera; Power, Deborah M; Johnston, Ian A

    2016-09-01

    Skeletal muscle, cartilage and bone must function in a co-ordinated fashion during locomotion and growth. In the present study on the gilthead sea bream (Sparus aurata) we tested the hypothesis that muscle and bone differ in their responsiveness to stimuli eliciting fast growth, providing a potential mechanism for generating the skeletal deformities observed in aquaculture. To investigate transcription regulation in skeletal muscle and bone we stimulated protein synthesis using a flooding dose of the branched chain amino acid leucine and compared the results with saline-injected controls. To increase the amount of available sequence information for gene expression analysis a de novo transcriptome was assembled using publicly available Next Generation Sequencing libraries from embryo, fast skeletal muscle, bone and cartilage. The resulting 5 million reads were assembled into 125,646 isotigs representing around 16,000 unique genes, including most components of the Pi3k/Akt/mTor signalling pathway. Principal components analysis was able to distinguish the transcriptional responses between leucine and saline injected controls in skeletal muscle, but not in the bone. General Linear Modelling revealed significant temporal changes in gene expression following leucine injection including the tissue-specific markers sparc, bglap (bone), mlc2 and myod2 (muscle) and gene transcripts associated with Pi3k/Akt/mTor signalling, p70sk6, akt2, ampka and mtor. Skeletal muscle showed more pronounced and rapid changes in transcript abundance than the bone to the same pro-growth signal. The observed differences in transcriptional response are consistent with the idea that fast growth results in a miss-match between muscle and bone development and may contribute to a higher incidence of skeletal deformities.

  5. Leucine supplementation of a low-protein meal increases skeletal muscle and visceral tissue protein synthesis in neonatal pigs by stimulating mTOR-dependent translation initiation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Protein synthesis and eukaryotic initiation factor (eIF) activation are increased in skeletal muscle of neonatal pigs parenterally infused with amino acids. Leucine appears to be the most effective single amino acid to trigger these effects. To examine the response to enteral leucine supplementation...

  6. Estrogen supplementation reduces whole body leucine and carbohydrate oxidation and increases lipid oxidation in men during endurance exercise.

    PubMed

    Hamadeh, Mazen J; Devries, Michaela C; Tarnopolsky, Mark A

    2005-06-01

    Healthy active men exhibit higher rates of carbohydrate (CHO) and leucine oxidation and lower rates of lipid oxidation compared with their female counterparts both at rest and during moderate intensity endurance exercise. We postulated that this reduced dependence on amino acids as a fuel source in women was due to the female sex hormone estrogen. In a randomized, double-blind, placebo-controlled, cross-over design, we investigated the effect of supplementing 12 recreationally active men with estrogen on whole body substrate oxidation and leucine kinetics at rest and during moderate intensity endurance exercise. Subjects cycled for 90 min at an intensity of 65% maximum O(2) consumption after 8 d of either estrogen supplementation (2 mg 17beta-estradiol/d) or placebo (polycose). After a 2-wk washout period, they repeated the test after 8 d of the alternate treatment. On the test day, after a primed continuous infusion of l-[(13)C]leucine, O(2) consumption, CO(2) production, steady-state breath (13)CO(2), and plasma alpha-[(13)C]ketoisocaproate enrichments were measured at rest and at 60, 75, and 90 min during exercise in the postabsorptive state. Exercise increased energy expenditure more than 5-fold, CHO oxidation more than 6-fold, lipid oxidation more than 4-fold, and leucine oxidation 2.2-fold (all P < 0.0001), whereas it decreased the ratio of lipid to CHO oxidation by 50-70% (P = 0.003) compared with values at rest. Estrogen supplementation decreased respiratory exchange ratio during exercise (P = 0.03). Estrogen supplementation significantly decreased CHO oxidation by 5-16% (P = 0.04) and leucine oxidation by 16% (P = 0.01), whereas it significantly increased lipid oxidation by 22-44% (P = 0.024) at rest and during exercise. We conclude that estrogen influences fuel source selection at rest and during endurance exercise in recreationally active men, characterized by a reduced dependence on amino acids and CHO and an increased reliance on lipids as a fuel

  7. Dietary l-leucine supplementation of lactating rats results in a tendency to increase lean/fat ratio associated to lower orexigenic neuropeptide expression in hypothalamus.

    PubMed

    López, N; Sánchez, J; Picó, C; Palou, A; Serra, F

    2010-07-01

    The aim of this study was to assess the effects of dietary leucine supplementation in lactating dams, particularly on energy homeostasis through signaling mechanisms in the central nervous system. Dams were fed ad libitum with standard diet during pregnancy (control dams) or supplemented with 2% leucine (leucine-supplemented dams) from delivery onwards. Food intake, body weight and composition were periodically recorded. Hypothalamus was collected at the end of lactation, and the expression of neuropeptide Y (NPY), agouti-related protein (AgRP) pro-opiomelanocortin (POMC), cocaine and amphetamine regulated transcript (CART), insulin receptor (InsR), ghrelin receptor (GSHR), melanocortin receptor (MCR4), leptin receptor (Ob-Rb) and suppressor of cytokine signaling 3 (SOCS3) were analyzed. Dietary leucine supplementation to lactating rats increased plasma leucine by 56%, modulated body composition and contributed to a tendency of higher ratio of lean/fat mass content of dams during lactation, without affecting food intake, thermogenesis capacity or body or tissue/organs weights. No differences in body weight of offspring from control and leucine-supplemented dams were found. The expression of orexigenic peptides (NPY and AgRP) decreased in leucine-dams, whereas the expression of anorexigenic peptides (POMC and CART), the hypothalamic receptors of insulin, ghrelin, melanocortin and leptin and SOCS3 did not change by leucine supplementation. In conclusion, increased leucine intake during lactation may contribute to a healthier profile of body composition in dams, without compromising the growth and development of the progeny by a mechanism associated with lower expression of orexigenic neuropeptides in hypothalamus.

  8. Leucine stimulation of skeletal muscle protein synthesis

    SciTech Connect

    Layman, D.K.; Grogan, C.K.

    1986-03-01

    Previous work in this laboratory has demonstrated a stimulatory effect of leucine on skeletal muscle protein synthesis measured in vitro during catabolic conditions. Studies in other laboratories have consistently found this effect in diaphragm muscle, however, studies examining effects on nitrogen balance or with in vivo protein synthesis in skeletal muscle are equivocal. This experiment was designed to determine the potential of leucine to stimulate skeletal muscle protein synthesis in vivo. Male Sprague-Dawley rats weighing 200 g were fasted for 12 hrs, anesthetized, a jugular cannula inserted, and protein synthesis measured using a primed continuous infusion of /sup 14/C-tyrosine. A plateau in specific activity was reached after 30 to 60 min and maintained for 3 hrs. The leucine dose consisted of a 240 umole priming dose followed by a continuous infusion of 160 umoles/hr. Leucine infusion stimulated protein synthesis in the soleus muscle (28%) and in the red (28%) and white portions (12%) of the gastrocnemius muscle compared with controls infused with only tyrosine. The increased rates of protein synthesis were due to increased incorporation of tyrosine into protein and to decreased specific activity of the free tyrosine pool. These data indicate that infusion of leucine has the potential to stimulate in vivo protein synthesis in skeletal muscles.

  9. Differential effects of leucine on translation initiation factor activation and protein synthesis in skeletal muscle, renal and adipose tissues of neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In adult rats, protein synthesis in skeletal muscle and adipose tissue increases in response to pharmacological doses of leucine (Leu) administered orally. In neonatal pigs, a physiological increase in plasma leucine stimulates protein synthesis in skeletal muscle without increasing hepatic protein...

  10. Anthocyanin excretion increases linearly with increasing strawberry dose.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A clinical study was conducted to investigate the dose response and metabolism of strawberry anthocyanins. In a crossover study design, twelve healthy adults consumed each of three strawberry treatments. The treatments were 100 g, 200 g, and 400 g of pureed strawberries, delivering 15 micromol, 30 m...

  11. Leucine markedly regulates pancreatic exocrine secretion in goats.

    PubMed

    Yu, Z P; Xu, M; Liu, K; Yao, J H; Yu, H X; Wang, F

    2014-02-01

    Four goats (30.1 ± 1.3 kg) with common bile duct re-entrant catheter and duodenal catheter were used to evaluate the effects of duodenal leucine infusion on pancreatic exocrine secretion and plasma parameters with two 4 × 4 Latin square design experiments. In the long-term infusion experiment, goats were fed twice daily [700 g/day, dry matter (DM) basis] at 8:00 and 18:00 hours and were duodenally infused with 0, 3, 6, 9 g/day leucine for 14 days. Pancreatic juice and jugular blood samples were collected over 1-h intervals for 6 h daily from d 11 to 14 days to encompass a 24-h day. In the short-term experiment, goats were infused leucine for 10 h continuously at the same infusion rate with Experiment 1 after feed deprivation for 24 h repeated every 10 days. Pancreatic juice and blood samples were collected at 0, 1, 2, 4, 6, 8 and 10 h of infusion. The results showed that the long-term leucine infusion did not affect pancreatic juice secretion, protein output, trypsin and lipase secretion and plasma insulin concentration, but linearly increased α-amylase secretion. No changes in pancreatic protein and lipase secretion were observed in the short-term infusion. Pancreatic juice and α-amylase secretion responded quadratically, with the greatest values observed in the 3 and 6 g/day leucine respectively. Trypsin secretion linearly decreased, while plasma insulin concentration increased linearly with increased leucine infusion. The results demonstrated that duodenal leucine infusion dose and time dependently regulated pancreatic enzyme secretion not associated with the change in plasma insulin concentration.

  12. Effect of very high dose D-leucine6-gonadotropin-releasing hormone proethylamide on the hypothalamic-pituitary testicular axis in patients with prostatic cancer.

    PubMed Central

    Warner, B; Worgul, T J; Drago, J; Demers, L; Dufau, M; Max, D; Santen, R J

    1983-01-01

    Potent synthetic analogs of gonadotropin-releasing hormone produce parodoxical antireproductive effects when administered chronically. These compounds are minimally toxic and may exhibit no plateau of the dose-response curve even at very high doses. These considerations served as the basis for our systematic evaluation of [D-leucine6-desarginine-glycine-NH2(10)]gonadotropin-releasing hormone (GnRH-A) proethylamide in the very high dose range (i.e., 10-fold larger amounts than previously used). In rats given the analog for 12 wk, prostate, testis, and seminal vesicle weights were suppressed to a greater extent with 200 micrograms q.d. than with 40 micrograms q.d. (P less than 0.01 prostate, less than 0.01 testis, less than 0.01 seminal vesicles), indicating dose-response effects in the very high dose range. 200 micrograms of [D-Leu6-des-Gly-NH2(10]-GnRH-A consistently suppressed leutinizing hormone (LH) values at 6 and 12 wk (basal 71 +/- 9.5; 6 wk 34 +/- 3.8; 12 wk 28 +/- 5 ng/ml) whereas 40 micrograms suppressed LH variably (basal 33 +/- 3.8; 6 wk 17 +/- 3.9; 12 wk 32 +/- 5.2). Testosterone fell to 15 +/- 2.4 and 19 +/- 2.0 ng/100 ml in response to 200 micrograms q.d. and to 27 +/- 6.4 and 22 +/- 7.4 ng/100 ml with the 40-micrograms dose. These findings in the rodent prompted treatment of stage D prostate cancer patients with similarly high doses of [D-Leu6-des-Gly-NH2(10)]-GnRH-A. After treatment for 11 wk with 1,000 or 10,000 micrograms/d of the analog, testosterone and dihydrotestosterone levels transiently rose and then fell into the surgically castrate range (testosterone 19 +/- 4.4 ng/100 ml [D-Leu6-des-Gly-NH2(10)]-GnRH-A vs. surgically castrate 11 +/- 0.9 ng/100 ml, P = NS; dihydrotestosterone 15 +/- 1.7 ng/100 ml GnRH-A vs. surgically castrate 15 +/- 4.1 ng/100 ml. P = NS). However, unlike the chronic stimulatory effect on the pituitary at lower doses, very high dose therapy resulted in profound suppression of plasma and urine LH. Plasma levels fell to

  13. Chronic leucine supplementation of a low protein diet increases protein synthesis in skeletal muscle and visceral tissues of neonatal pigs through mTOR signaling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leucine acutely stimulates protein synthesis by activating the mammalian target of rapamycin (mTOR) signaling pathway. We hypothesized that leucine supplementation of a low protein diet will enhance protein synthesis and mTOR signaling in the neonate for prolonged periods. Fasted 5-d-old pigs (n=6–8...

  14. Multilateral analysis of increasing collective dose and new ALARA programme.

    PubMed

    Oumi, Tadashi; Morii, Yasuki; Imai, Toshirou

    2011-07-01

    JAPC (The Japan Atomic Power Company) is the only electric power company that operates different types of nuclear reactors in Japan; it operates two BWRs (boiling water reactors), one pressurised water reactor and one gas cooled reactor. JAPC has been conducting various activities aimed at reducing radiation dose received by workers for over 45 y. Recently, the collective dose resulting from periodic maintenance has increased at each plant because of the replacement of large equipment and the unexpected extension of the outage period. In particular, the collective dose at Tokai-2 is one of the highest among Japanese BWR plants((1)), owing to the replacement and strengthening of equipment to meet earthquake-proof requirements. In this study, the authors performed a multilateral analysis of unacceptably a large collective dose and devised a new ALARA programme that includes a 3D dose prediction map and the development of machines to assist workers.

  15. Does administering iodine in radiological procedures increase patient doses?

    SciTech Connect

    He, Wenjun; Yao, Hai; Huda, Walter; Mah, Eugene

    2014-11-01

    Purpose: The authors investigated the changes in the pattern of energy deposition in tissue equivalent phantoms following the introduction of iodinated contrast media. Methods: The phantom consisted of a small “contrast sphere,” filled with water or iodinated contrast, located at the center of a 28 cm diameter water sphere. Monte Carlo simulations were performed using MCNP5 codes, validated by simulating irradiations with analytical solutions. Monoenergetic x-rays ranging from 35 to 150 keV were used to simulate exposures to spheres containing contrast agent with iodine concentrations ranging from 1 to 100 mg/ml. Relative values of energy imparted to the contrast sphere, as well as to the whole phantom, were calculated. Changes in patterns of energy deposition around the contrast sphere were also investigated. Results: Small contrast spheres can increase local absorbed dose by a factor of 13, but the corresponding increase in total energy absorbed was negligible (<1%). The highest localized dose increases were found to occur at incident photon energies of about 60 keV. For a concentration of about 10 mg/ml, typical of clinical practice, localized absorbed doses were generally increased by about a factor of two. At this concentration of 10 mg/ml, the maximum increase in total energy deposition in the phantom was only 6%. These simulations demonstrated that increases in contrast sphere doses were offset by corresponding dose reductions at distal and posterior locations. Conclusions: Adding iodine can result in values of localized absorbed dose increasing by more than an order of magnitude, but the total energy deposition is generally very modest (i.e., <10%). Their data show that adding iodine primarily changes the pattern of energy deposition in the irradiated region, rather than increasing patient doses per se.

  16. N-acetyl-L-leucine accelerates vestibular compensation after unilateral labyrinthectomy by action in the cerebellum and thalamus.

    PubMed

    Günther, Lisa; Beck, Roswitha; Xiong, Guoming; Potschka, Heidrun; Jahn, Klaus; Bartenstein, Peter; Brandt, Thomas; Dutia, Mayank; Dieterich, Marianne; Strupp, Michael; la Fougère, Christian; Zwergal, Andreas

    2015-01-01

    An acute unilateral vestibular lesion leads to a vestibular tone imbalance with nystagmus, head roll tilt and postural imbalance. These deficits gradually decrease over days to weeks due to central vestibular compensation (VC). This study investigated the effects of i.v. N-acetyl-DL-leucine, N-acetyl-L-leucine and N-acetyl-D-leucine on VC using behavioural testing and serial [18F]-Fluoro-desoxyglucose ([18F]-FDG)-μPET in a rat model of unilateral chemical labyrinthectomy (UL). Vestibular behavioural testing included measurements of nystagmus, head roll tilt and postural imbalance as well as sequential whole-brain [18F]-FDG-μPET was done before and on days 1,3,7 and 15 after UL. A significant reduction of postural imbalance scores was identified on day 7 in the N-acetyl-DL-leucine (p < 0.03) and the N-acetyl-L-leucine groups (p < 0.01), compared to the sham treatment group, but not in the N-acetyl-D-leucine group (comparison for applied dose of 24 mg i.v. per rat, equivalent to 60 mg/kg body weight, in each group). The course of postural compensation in the DL- and L-group was accelerated by about 6 days relative to controls. The effect of N-acetyl-L-leucine on postural compensation depended on the dose: in contrast to 60 mg/kg, doses of 15 mg/kg and 3.75 mg/kg had no significant effect. N-acetyl-L-leucine did not change the compensation of nystagmus or head roll tilt at any dose. Measurements of the regional cerebral glucose metabolism (rCGM) by means of μPET revealed that only N-acetyl-L-leucine but not N-acetyl-D-leucine caused a significant increase of rCGM in the vestibulocerebellum and a decrease in the posterolateral thalamus and subthalamic region on days 3 and 7. A similar pattern was found when comparing the effect of N-acetyl-L-leucine on rCGM in an UL-group and a sham UL-group without vestibular damage. In conclusion, N-acetyl-L-leucine improves compensation of postural symptoms after UL in a dose-dependent and specific manner, most likely by

  17. N-Acetyl-L-Leucine Accelerates Vestibular Compensation after Unilateral Labyrinthectomy by Action in the Cerebellum and Thalamus

    PubMed Central

    Xiong, Guoming; Potschka, Heidrun; Jahn, Klaus; Bartenstein, Peter; Brandt, Thomas; Dutia, Mayank; Dieterich, Marianne; Strupp, Michael; la Fougère, Christian; Zwergal, Andreas

    2015-01-01

    An acute unilateral vestibular lesion leads to a vestibular tone imbalance with nystagmus, head roll tilt and postural imbalance. These deficits gradually decrease over days to weeks due to central vestibular compensation (VC). This study investigated the effects of i.v. N-acetyl-DL-leucine, N-acetyl-L-leucine and N-acetyl-D-leucine on VC using behavioural testing and serial [18F]-Fluoro-desoxyglucose ([18F]-FDG)-μPET in a rat model of unilateral chemical labyrinthectomy (UL). Vestibular behavioural testing included measurements of nystagmus, head roll tilt and postural imbalance as well as sequential whole-brain [18F]-FDG-μPET was done before and on days 1,3,7 and 15 after UL. A significant reduction of postural imbalance scores was identified on day 7 in the N-acetyl-DL-leucine (p < 0.03) and the N-acetyl-L-leucine groups (p < 0.01), compared to the sham treatment group, but not in the N-acetyl-D-leucine group (comparison for applied dose of 24 mg i.v. per rat, equivalent to 60 mg/kg body weight, in each group). The course of postural compensation in the DL- and L-group was accelerated by about 6 days relative to controls. The effect of N-acetyl-L-leucine on postural compensation depended on the dose: in contrast to 60 mg/kg, doses of 15 mg/kg and 3.75 mg/kg had no significant effect. N-acetyl-L-leucine did not change the compensation of nystagmus or head roll tilt at any dose. Measurements of the regional cerebral glucose metabolism (rCGM) by means of μPET revealed that only N-acetyl-L-leucine but not N-acetyl-D-leucine caused a significant increase of rCGM in the vestibulocerebellum and a decrease in the posterolateral thalamus and subthalamic region on days 3 and 7. A similar pattern was found when comparing the effect of N-acetyl-L-leucine on rCGM in an UL-group and a sham UL-group without vestibular damage. In conclusion, N-acetyl-L-leucine improves compensation of postural symptoms after UL in a dose-dependent and specific manner, most likely by

  18. Overexpression of the leucine-rich receptor-like kinase gene LRK2 increases drought tolerance and tiller number in rice.

    PubMed

    Kang, Junfang; Li, Jianmin; Gao, Shuang; Tian, Chao; Zha, Xiaojun

    2017-02-09

    Drought represents a key limiting factor of global crop distribution. Receptor-like kinases play major roles in plant development and defense responses against stresses such as drought. In this study, LRK2, which encodes a leucine-rich receptor-like kinase, was cloned and characterized and found to be localized on the plasma membrane in rice. Promoter-GUS analysis revealed strong expression in tiller buds, roots, nodes and anthers. Transgenic plants overexpressing LRK2 exhibited enhanced tolerance to drought stress due to an increased number of lateral roots compared to the wild-type at the vegetative stage. Moreover, ectopic expression of LRK2 seedlings resulted in increased tiller development. Yeast two-hybrid screening and bimolecular fluorescence complementation (BiFC) indicated a possible interaction between LRK2 and elongation factor 1 alpha (OsEF1A) in vitro. These results suggest that LRK2 functions as a positive regulator of the drought stress response and tiller development via increased branch development in rice. These findings will aid our understanding of branch regulation in other grasses and support improvements in rice genetics. This article is protected by copyright. All rights reserved.

  19. Increasing Stem Cell Dose Promotes Posttransplant Immune Reconstitution.

    PubMed

    Xu, Ning; Shen, Sylvie; Dolnikov, Alla

    2017-01-16

    Umbilical cord blood (UCB) transplantation can provide a successful therapeutic option for patients that have no suitable related donor. UCB transplantation is often limited by the relatively small hematopoietic stem cell (HSC) numbers in UCB especially for adult recipients. Early neutrophil and platelet engraftment correlates with the stem cell numbers in UCB transplant. Compared to other HSC sources, immune reconstitution following UCB transplant is slower and complicated by increased frequency of opportunistic infections. The effect of HSC numbers in UCB transplant on immune reconstitution was not thoroughly examined. Using immunocompromised mice transplanted with purified UCB CD34+ stem cells, we have demonstrated that increasing the numbers of CD34+ cells in the transplant promotes hematopoietic and immune reconstitution. At early stages posttransplant, high stem cell dose generated relatively more B cells, while lower dose generated more myeloid and T cells. Thus, the size of the stem cell graft appears to modulate the differentiation potential of infused stem cells. In addition, increasing stem cell dose in the transplant improved CD8+ T cell development and delayed late memory T cell skewing in expense of naive T cells highlighting the importance of HSC dose to maintain the pool of naive T cells able to develop strong immune responses. Transplantation of ex vivo expanded CD34+ cells did not promote, but rather delayed immune reconstitution suggesting the loss of primitive lymphoid precursor cells during ex vivo expansion.

  20. Leucine metabolism in patients with Hepatic Encephalopathy

    SciTech Connect

    McGhee, A.S.; Kassouny, M.E.; Matthews, D.E.; Millikan, W.

    1986-03-01

    A primed continuous infusion of (/sup 15/N, 1-/sup 13/C)leucine was used to determine whether increased oxidation and/or protein synthesis of leucine occurs in patients with cirrhosis. Five controls and patients were equilibrated on a metabolic balance diet (0.6 g protein per kg ideal body weight (IBW)). An additional four patients were equilibrated in the same manner with the same type of diet with a protein level of 0.75 g per kg IBW. Plasma leucine and breath CO/sub 2/ enrichments were measured by mass spectrometry. Protein synthesis and leucine metabolism were identical in controls and patients when both were fed a diet with 0.6 g protein/kg IBW. Results indicate that systemic derangements of leucine metabolism are not the cause of Hepatic Encephalopathy.

  1. Overexpression of Glucocorticoid-induced Leucine Zipper (GILZ) increases susceptibility to Imiquimod-induced psoriasis and involves cutaneous activation of TGF-β1

    PubMed Central

    Carceller, Elena; Ballegeer, Marlies; Deckers, Julie; Riccardi, Carlo; Bruscoli, Stefano; Hochepied, Tino; Libert, Claude; Pérez, Paloma

    2016-01-01

    Psoriasis vulgaris is a chronic inflammatory skin disease affecting millions of people. Its pathophysiology is complex and involves a skin compartment with epidermal and immune cells which produce cytokines, e.g. belonging to the IL-23–Th17-cell axis. Glucocorticoids (GCs) are the most common therapeutics used in cutaneous inflammatory disorders and GC-induced leucine zipper (GILZ) has emerged as a mediator of GCs due to its anti-inflammatory actions, theoretically lacking GC side-effects. We evaluated whether GILZ may provide a better therapeutic index in comparison to GCs during the onset and progression of psoriasis by generating and characterizing a mouse model with generalized overexpression of this protein (GILZ-Tg mice) and the imiquimod (IMQ) psoriasis model. Unexpectedly, in GILZ-Tg mice, the severity of IMQ-induced psoriasis-like skin lesions as well as induction of cytokines commonly up-regulated in human psoriasis (Il-17, Il-22, Il-23, Il-6, S100a8/a9, and Stat3) was significantly more pronounced relative to GILZ-Wt mice. The increased susceptibility to IMQ-induced psoriasis of GILZ-Tg mice was significantly associated with skin-specific over-activation of TGF-β1-mediated signaling via SMAD2/3. Our findings demonstrate that GILZ may behave as pro-inflammatory protein in certain tissues and that, similar to prolonged GC therapy, GILZ as an alternative treatment for psoriasis may also have adverse effects. PMID:27934944

  2. Atazanavir increases the plasma concentrations of 1200 mg raltegravir dose.

    PubMed

    Krishna, Rajesh; East, Lilly; Larson, Patrick; Valiathan, Chandni; Deschamps, Kathleen; Luk, Julie Ann; Bethel-Brown, Crystal; Manthos, Helen; Brejda, John; Gartner, Michael

    2016-12-01

    Raltegravir is a human immunodeficiency virus (HIV)-1 integrase strand transfer inhibitor currently marketed at a dose of 400 mg twice-daily (b.i.d.). Raltegravir 1200 mg once-daily (q.d.) (investigational q.d. formulation of 2 × 600 mg tablets; q.d. RAL) was found to be generally well tolerated and non-inferior to the marketed 400 mg b.i.d. dose at 48 weeks in a phase 3 trial. Since raltegravir is eliminated mainly by metabolism via a uridine diphosphate glucuronosyltransferase (UGT) 1A1-mediated glucuronidation pathway, co-administration of UGT1A1 inhibitors may increase the plasma levels of q.d. RAL. To assess this potential, the drug interaction of 1200 mg raltegravir using atazanavir, a known UGT1A1 inhibitor, was studied. An open-label, randomized, 2-period, fixed-sequence phase 1 study was performed in adult healthy male and female (non-childbearing potential) subjects ≥ 19 and ≤ 55 years of age, with a body mass index (BMI) ≥ 18.5 and ≤ 32.0 kg/m(2) . Subjects (n = 14) received a single oral dose of 1200 mg raltegravir in period 1. After a washout period of at least 7 days, the subjects received oral doses of 400 mg atazanavir q.d. for 9 consecutive days, with a single oral dose of 1200 mg raltegravir co-administered on day 7 of period 2. Serial blood samples were collected for 72 h following raltegravir dosing and analysed using a validated bioanalytical method to quantify raltegravir plasma concentrations. Co-administration with atazanavir yielded GMRs (90% CIs) for raltegravir AUC0-∞ , Cmax and C24 of 1.67 (1.34, 2.10), 1.16 (1.01, 1.33) and 1.26 (1.08, 1.46), respectively. There was no effect of raltegravir on serum total bilirubin. In contrast, atazanavir increased the mean bilirubin by up to 200%, an effect that was preserved in the atazanavir/raltegravir treatment group. Administration of single q.d. RAL alone and co-administered with multiple oral doses of atazanavir were generally well tolerated in healthy subjects. The results show that

  3. Global Metabolic Profiling of Plasma Shows that Three-Year Mild-Caloric Restriction Lessens an Age-Related Increase in Sphingomyelin and Reduces L-leucine and L-phenylalanine in Overweight and Obese Subjects.

    PubMed

    Kim, Minjoo; Lee, Sang-Hyun; Lee, Jong Ho

    2016-12-01

    The effect of weight loss from long-term, mild-calorie diets (MCD) on plasma metabolites is unknown. This study was to examine whether MCD-induced weight reduction caused changes in the extended plasma metabolites. Overweight and obese subjects aged 40-59 years consumed a MCD (approximately 100 kcal/day deficit, n=47) or a weight-maintenance diet (control, n=47) in a randomized, controlled design with a three-year clinical intervention period and plasma samples were analyzed by using UPLC-LTQ-Orbitrap mass spectrometry. The three-year MCD intervention resulted in weight loss (-8.87%) and significant decreases in HOMA-IR and TG. The three-year follow-up of the MCD group showed reductions in the following 13 metabolites: L-leucine; L-phenylalanine; 9 lysoPCs; PC (18:0/20:4); and SM (d18:0/16:1). The three-year MCD group follow-up identified increases in palmitic amide, oleamide, and PC (18:2/18:2). Considering the age-related alterations in the identified metabolites, the MCD group showed a greater decrease in L-leucine, L-phenylalanine, and SM (d18:0/16:1) compared with those of the control group. Overall, the change (Δ) in BMI positively correlated with the ΔTG, ΔHOMA-IR, ΔL-leucine, and ΔSM (d18:0/16:1). The ΔHOMA-IR positively correlated with ΔTG, ΔL-leucine, ΔL-phenylalanine, and ΔSM (d18:0/16:1). The weight loss resulting from three-year mild-caloric restriction lessens the age-related increase in SM and reduces L-leucine and L-phenylalanine in overweight and obese subjects. These changes were coupled with improved insulin resistance (ClinicalTrials.gov: NCT02081898).

  4. Global Metabolic Profiling of Plasma Shows that Three-Year Mild-Caloric Restriction Lessens an Age-Related Increase in Sphingomyelin and Reduces L-leucine and L-phenylalanine in Overweight and Obese Subjects

    PubMed Central

    Kim, Minjoo; Lee, Sang-Hyun; Lee, Jong Ho

    2016-01-01

    The effect of weight loss from long-term, mild-calorie diets (MCD) on plasma metabolites is unknown. This study was to examine whether MCD-induced weight reduction caused changes in the extended plasma metabolites. Overweight and obese subjects aged 40-59 years consumed a MCD (approximately 100 kcal/day deficit, n=47) or a weight-maintenance diet (control, n=47) in a randomized, controlled design with a three-year clinical intervention period and plasma samples were analyzed by using UPLC-LTQ-Orbitrap mass spectrometry. The three-year MCD intervention resulted in weight loss (-8.87%) and significant decreases in HOMA-IR and TG. The three-year follow-up of the MCD group showed reductions in the following 13 metabolites: L-leucine; L-phenylalanine; 9 lysoPCs; PC (18:0/20:4); and SM (d18:0/16:1). The three-year MCD group follow-up identified increases in palmitic amide, oleamide, and PC (18:2/18:2). Considering the age-related alterations in the identified metabolites, the MCD group showed a greater decrease in L-leucine, L-phenylalanine, and SM (d18:0/16:1) compared with those of the control group. Overall, the change (Δ) in BMI positively correlated with the ΔTG, ΔHOMA-IR, ΔL-leucine, and ΔSM (d18:0/16:1). The ΔHOMA-IR positively correlated with ΔTG, ΔL-leucine, ΔL-phenylalanine, and ΔSM (d18:0/16:1). The weight loss resulting from three-year mild-caloric restriction lessens the age-related increase in SM and reduces L-leucine and L-phenylalanine in overweight and obese subjects. These changes were coupled with improved insulin resistance (ClinicalTrials.gov: NCT02081898). PMID:28053823

  5. Enteral leucine supplementation increases protein synthesis in skeletal and cardiac muscles and visceral tissues of neonatal pigs through mTORC1-dependent pathways

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leucine activates mammalian target of rapamycin (mTOR) to upregulate protein synthesis (PS). To examine enteral Leu effects on PS and signaling activation, 5-d-old piglets were fed for 24 h diets containing: (i) LP, (ii) LP+L, or (iii) HP. PS in skeletal muscles, heart, liver, pancreas, and jejunum...

  6. Leucine supplementation of an infant formula increases skeletal muscle and visceral tissue protein synthesis in a neonatal animal model by stimulating mTOR-dependent translation initiation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Low birth weight infants are often discharged weighing less than the tenth percentile for age and some remain small to adulthood with adverse long-term outcomes related to inadequate nutrition. Infant formula fortifiers had been used to improve the nutritional value of milk formulas. Leucine (Leu) a...

  7. Pathology effects at radiation doses below those causing increased mortality

    NASA Technical Reports Server (NTRS)

    Carnes, Bruce A.; Gavrilova, Natalia; Grahn, Douglas

    2002-01-01

    Mortality data from experiments conducted at the Argonne National Laboratory (ANL) on the long-term effects of external whole-body irradiation on B6CF(1) mice were used to investigate radiation-induced effects at intermediate doses of (60)Co gamma rays or fission-spectrum neutrons either delivered as a single exposure or protracted over 60 once-weekly exposures. Kaplan-Meier analyses were used to identify the lowest dose in the ANL data (within radiation quality, pattern of exposure, and sex) at which radiation-induced mortality caused by primary tumors could be detected (approximately 1-2 Gy for gamma rays and 10-15 cGy for neutrons). Doses at and below these levels were then examined for radiation-induced shifts in the spectrum of pathology detected at death. To do this, specific pathology events were pooled into larger assemblages based on whether they were cancer, cardiovascular disease or non-neoplastic diseases detected within the lungs and pleura, liver and biliary tract, reproductive organs, or urinary tract. Cancer and cardiovascular disease were further subdivided into categories based on whether they caused death, contributed to death, or were simply observed at death. Counts of how often events falling within each of these combined pathology categories occurred within a mouse were then used as predictor variables in logistic regression to determine whether irradiated mice could be distinguished from control mice. Increased pathology burdens were detected in irradiated mice at doses lower than those causing detectable shifts in mortality-22 cGy for gamma rays and 2 cGy for neutrons. These findings suggest that (1) models based on mortality data alone may underestimate radiation effects, (2) radiation may have adverse health consequences (i.e. elevated health risks) even when mortality risks are not detected, and (3) radiation-induced pathologies other than cancer do occur, and they involve multiple organ systems.

  8. Leucine aminopeptidase - urine

    MedlinePlus

    ... GO About MedlinePlus Site Map FAQs Customer Support Health Topics Drugs & Supplements Videos & Tools Español You Are Here: Home → Medical Encyclopedia → Leucine aminopeptidase - urine URL of this page: //medlineplus.gov/ ...

  9. Increased radiation dose at mammography due to prolonged exposure, delayed processing, and increased film darkening

    SciTech Connect

    Kimme-Smith, C.; Bassett, L.W.; Gold, R.H.; Chow, S. )

    1991-02-01

    Four single-emulsion films introduced over the past 2 years--Du Pont Microvision, Fuji MiMa, Konica CM, and Eastman Kodak OM--were compared with Eastman Kodak OM SO-177 (Min-RE) film to evaluate their varying effects on mean glandular dose of reciprocity law failure due to prolonged exposure, delayed processing, and increased film darkening as a result of increased radiation exposure to improve penetration of glandular tissue. Exposures over 1.3 seconds led to increased radiation doses of 20%-30%. Delays in processing of 6 hours decreased processing speed by 11%-32% for all films except Du Pont Microvision. Optical density increases of 0.40 required 20%-30% more skin exposure for all five films. Optimal viewing densities were also evaluated and found to be different for each of the five films. Mammographers need to be aware of these differences in mammographic films to achieve maximum contrast at mammography.

  10. Increased radiation dose at mammography due to prolonged exposure, delayed processing, and increased film darkening.

    PubMed

    Kimme-Smith, C; Bassett, L W; Gold, R H; Chow, S

    1991-02-01

    Four single-emulsion films introduced over the past 2 years--Du Pont Microvision, Fuji MiMa, Konica CM, and Eastman Kodak OM--were compared with Eastman Kodak OM SO-177 (Min-RE) film to evaluate their varying effects on mean glandular dose of reciprocity law failure due to prolonged exposure, delayed processing, and increased film darkening as a result of increased radiation exposure to improve penetration of glandular tissue. Exposures over 1.3 seconds led to increased radiation doses of 20%-30%. Delays in processing of 6 hours decreased processing speed by 11%-32% for all films except Du Pont Microvision. Optical density increases of 0.40 required 20%-30% more skin exposure for all five films. Optimal viewing densities were also evaluated and found to be different for each of the five films. Mammographers need to be aware of these differences in mammographic films to achieve maximum contrast at mammography.

  11. Potent anti-seizure effects of D-leucine.

    PubMed

    Hartman, Adam L; Santos, Polan; O'Riordan, Kenneth J; Stafstrom, Carl E; Hardwick, J Marie

    2015-10-01

    There are no effective treatments for millions of patients with intractable epilepsy. High-fat ketogenic diets may provide significant clinical benefit but are challenging to implement. Low carbohydrate levels appear to be essential for the ketogenic diet to work, but the active ingredients in dietary interventions remain elusive, and a role for ketogenesis has been challenged. A potential antiseizure role of dietary protein or of individual amino acids in the ketogenic diet is understudied. We investigated the two exclusively ketogenic amino acids, L-leucine and L-lysine, and found that only L-leucine potently protects mice when administered prior to the onset of seizures induced by kainic acid injection, but not by inducing ketosis. Unexpectedly, the D-enantiomer of leucine, which is found in trace amounts in the brain, worked as well or better than L-leucine against both kainic acid and 6Hz electroshock-induced seizures. However, unlike L-leucine, D-leucine potently terminated seizures even after the onset of seizure activity. Furthermore, D-leucine, but not L-leucine, reduced long-term potentiation but had no effect on basal synaptic transmission in vitro. In a screen of candidate neuronal receptors, D-leucine failed to compete for binding by cognate ligands, potentially suggesting a novel target. Even at low doses, D-leucine suppressed ongoing seizures at least as effectively as diazepam but without sedative effects. These studies raise the possibility that D-leucine may represent a new class of anti-seizure agents, and that D-leucine may have a previously unknown function in eukaryotes.

  12. Leucine aminopeptidase urine test (image)

    MedlinePlus

    Leucine aminopeptidase (LAP) is a proteolytic enzyme that breaks chemical bonds in proteins at specific sites next to leucine amino acids. Serum (blood) LAP is measured to diagnose liver (hepatic) dysfunction.

  13. Increased aggressive responding in male volunteers following the administration of gradually increasing doses of testosterone cypionate.

    PubMed

    Kouri, E M; Lukas, S E; Pope, H G; Oliva, P S

    1995-11-01

    The present study assessed the effects of supraphysiologic doses of testosterone on aggressive responding in a controlled laboratory setting. Eight male subjects received gradually increasing doses of testosterone cypionate (150 mg/week for two weeks, 300 mg/week for two weeks, and 600 mg/week for two weeks) or placebo using a double-blind, randomized, cross-over design. Subjects were tested both before and after the series of injections. During the experimental session subjects could press a button to accumulate points exchangeable for money (non-aggressive response) or press another button to subtract points from a fictitious opponent (aggressive response). Aggressive responding was instigated by subtracting points from the subject which was attributable to the fictitious opponent. Testosterone administration resulted in a significantly higher number of aggressive responding compared to placebo.

  14. Leucine metabolism in human newborns

    SciTech Connect

    Denne, S.C.; Kalhan, S.C. )

    1987-12-01

    The present study was designed to (1) determine whether a relationship exists between newborn birth weight and leucine metabolism and (2) compare leucine and energy metabolism in a period of rapid growth and development (i.e., newborn) with a constant nongrowth period (i.e., adult). Leucine kinetics and energy expenditure were measured in the postabsorptive state in 12 normal full-term newborns in early neonatal life and in 11 normal adults using a primed constant L-(1-{sup 13}C)leucine infusion combined with respiratory calorimetry. A significant positive correlation between newborn birth weight and leucine flux was observed. These data suggest the following. (1) A relationship exists between newborn birth weight and protein metabolism, as reflected by the correlation between leucine flux when expressed as micromoles per kilogram per hour and birth weight. (2) The high rate of leucine flux measured in newborns probably reflects the rapid remodeling of protein that occurs in this period of development, even during fasting. (3) The similar values in newborns and adults of leucine kinetics and energy expenditure when normalized to metabolic body weight and the nearly equivalent allometric exponents relating body weight to leucine flux and energy expenditure support a close relationship between leucine and energy metabolism, at least at the extremes of human growth.

  15. Effect of insulin and plasma amino acid concentrations on leucine metabolism in man. Role of substrate availability on estimates of whole body protein synthesis.

    PubMed Central

    Castellino, P; Luzi, L; Simonson, D C; Haymond, M; DeFronzo, R A

    1987-01-01

    We examined the effect of insulin and plasma amino acid concentrations on leucine kinetics in 15 healthy volunteers (age 22 +/- 2 yr) using the euglycemic insulin clamp technique and an infusion of [1-14C]leucine. Four different experimental conditions were examined: (a) study one, high insulin with reduced plasma amino acid concentrations; (b) study two, high insulin with maintenance of basal plasma amino acid concentrations; (c) study three, high insulin with elevated plasma amino acid concentrations; and (d) study four, basal insulin with elevated plasma amino acid concentrations. Data were analyzed using both the plasma leucine and alpha-ketoisocaproate (the alpha-ketoacid of leucine) specific activities. In study one total leucine flux, leucine oxidation, and nonoxidative leucine disposal (an index of whole body protein synthesis) all decreased (P less than 0.01) regardless of the isotope model utilized. In study two leucine flux did not change, while leucine oxidation increased (P less than 0.01) and nonoxidative leucine disposal was maintained at the basal rate; endogenous leucine flux (an index of whole body protein degradation) decreased (P less than 0.01). In study three total leucine flux, leucine oxidation, and nonoxidative leucine disposal all increased significantly (P less than 0.01). In study four total leucine flux, leucine oxidation, and nonoxidative leucine disposal all increased (P less than 0.001), while endogenous leucine flux decreased (P less than 0.001). We conclude that: (a) hyperinsulinemia alone decreases plasma leucine concentration and inhibits endogenous leucine flux (protein breakdown), leucine oxidation, and nonoxidative leucine disposal (protein synthesis); (b) hyperaminoacidemia, whether in combination with hyperinsulinemia or with maintained basal insulin levels decreases endogenous leucine flux and stimulates both leucine oxidation and nonoxidative leucine disposal. PMID:3316280

  16. Mitochondrial leucine tRNA level and PTCD1 are regulated in response to leucine starvation.

    PubMed

    Schild, Christof; Hahn, Dagmar; Schaller, André; Jackson, Christopher Benjamin; Rothen-Rutishauser, Barbara; Mirkovitch, Jelena; Nuoffer, Jean-Marc

    2014-07-01

    Pentatricopeptide repeat domain protein 1 (PTCD1) is a novel human protein that was recently shown to decrease the levels of mitochondrial leucine tRNAs. The physiological role of this regulation, however, remains unclear. Here we show that amino acid starvation by leucine deprivation significantly increased the mRNA steady-state levels of PTCD1 in human hepatocarcinoma (HepG2) cells. Amino acid starvation also increased the mitochondrially encoded leucine tRNA (tRNA(Leu(CUN))) and the mRNA for the mitochondrial leucyl-tRNA synthetase (LARS2). Despite increased PTCD1 mRNA steady-state levels, amino acid starvation decreased PTCD1 on the protein level. Decreasing PTCD1 protein concentration increases the stability of the mitochondrial leucine tRNAs, tRNA(Leu(CUN)) and tRNA(Leu(UUR)) as could be shown by RNAi experiments against PTCD1. Therefore, it is likely that decreased PTCD1 protein contributes to the increased tRNA(Leu(CUN)) levels in amino acid-starved cells. The stabilisation of the mitochondrial leucine tRNAs and the upregulation of the mitochondrial leucyl-tRNA synthetase LARS2 might play a role in adaptation of mitochondria to amino acid starvation.

  17. Increasing the Dose of Autologous Chondrocytes Improves Articular Cartilage Repair

    PubMed Central

    Guillén-García, Pedro; Rodríguez-Iñigo, Elena; Guillén-Vicente, Isabel; Caballero-Santos, Rosa; Guillén-Vicente, Marta; Abelow, Stephen; Giménez-Gallego, Guillermo

    2014-01-01

    Background: We hypothesized that implanting cells in a chondral defect at a density more similar to that of the intact cartilage could induce them to synthesize matrix with the features more similar to that of the uninjured one. Methods: We compared the implantation of different doses of chondrocytes: 1 million (n = 5), 5 million (n = 5), or 5 million mesenchymal cells (n = 5) in the femoral condyle of 15 sheep. Tissue generated by microfracture at the trochlea, and normal cartilage from a nearby region, processed as the tissues resulting from the implantation, were used as references. Histological and molecular (expression of type I and II collagens and aggrecan) studies were performed. Results: The features of the cartilage generated by implantation of mesenchymal cells and elicited by microfractures were similar and typical of a poor repair of the articular cartilage (presence of fibrocartilage, high expression of type I collagen and a low mRNA levels of type II collagen and aggrecan). Nevertheless, in the samples obtained from tissues generated by implantation of chondrocytes, hyaline-like cartilage, cell organization, low expression rates of type I collagen and high levels of mRNA corresponding to type II collagen and aggrecan were observed. These histological features, show less variability and are more similar to those of the normal cartilage used as control in the case of 5 million cells implantation than when 1 million cells were used. Conclusions: The implantation of autologous chondrocytes in type I/III collagen membranes at high density could be a promising tool to repair articular cartilage. PMID:26069691

  18. Prolonged oral treatment with an essential amino acid L-leucine does not affect female reproductive function and embryo-fetal development in rats.

    PubMed

    Mawatari, Kazunori; Katsumata, Toyohisa; Uematsu, Masanobu; Katsumata, Tomoyoshi; Yoshida, Junichi; Smriga, Miro; Kimura, Takeshi

    2004-09-01

    L-leucine, an essential amino acid, is one of the most popular ingredients in dietary supplements. To investigate a possibility of its embryo-fetal toxicity in rats, 11- to 12-week old dams were orally administered an aqueous solution of L-leucine at doses of 300 or 1000 mg/kg body weight on gestational days 7-17. Body weight and feed intake was evaluated throughout the whole course of pregnancy (days 0-20). L-Leucine did not influence body weight, but at a dose of 1000 mg/kg, slightly enhanced feed intake on days 14 and 18 of pregnancy. Caesarean section (day 20) revealed no influences on the litter size and weight of live-born fetuses, the number of corpora lutea, implantation index or the quality of placenta, and the minor increase in feed intake was considered irrelevant to the pregnancy outcomes. Fetuses were evaluated in a battery of external, visceral and skeletal examinations. No effects of L-leucine on gender ratio and external abnormalities, and no significant treatment-related variations in visceral and skeletal pathologies were observed. These results suggested that L-leucine, administered orally during organogenesis at doses up to 1000 mg/kg body weight, did not affect the outcome of pregnancy and did not cause fetotoxicity in rats.

  19. Leucine as a treatment for muscle wasting: a critical review.

    PubMed

    Ham, Daniel J; Caldow, Marissa K; Lynch, Gordon S; Koopman, René

    2014-12-01

    Amino acids are potent modulators of protein turnover and skeletal muscle cells are highly sensitive to changes in amino acid availability. During amino acid abundance increased activity of mTORC1 drives protein synthesis and growth. In skeletal muscle, it has been clearly demonstrated that of all the amino acids, leucine is the most potent stimulator of mTORC1 and protein synthesis in vitro and in vivo. As such, leucine has received considerable attention as a potential pharmaconutrient for the treatment of numerous muscle wasting conditions. However, despite a multitude of studies showing enhanced acute protein synthesis with leucine or leucine-rich supplements in healthy individuals, additional leucine intake does not necessarily enhance protein synthesis during muscle wasting conditions. In addition, long-term, placebo controlled, iso-caloric studies in humans consistently show no beneficial effect of leucine supplementation on skeletal muscle mass or function. This review, critically evaluates the therapeutic potential of leucine to attenuate the skeletal muscle wasting associated with ageing, cancer and immobilization/bed rest. It also highlights the impact of inflammation on amino acid sensing, mTORC1 activation and stimulation of protein synthesis and challenges the underlying hypothesis that the acute activation of mTORC1 and stimulation of protein synthesis by leucine increases in muscle mass over time. We conclude that leucine, as a standalone nutritional intervention, is not effective in the prevention of muscle wasting. Future work should focus on identifying and utilizing other nutrients or treatments that sensitize skeletal muscle to leucine, thereby enhancing its therapeutic potential for muscle wasting conditions.

  20. Leucine transport is affected by Bacillus thuringiensis Cry1 toxins in brush border membrane vesicles from Ostrinia nubilalis Hb (Lepidoptera: Pyralidae) and Sesamia nonagrioides Lefebvre (Lepidoptera: Noctuidae) midgut.

    PubMed

    Leonardi, M Giovanna; Caccia, Silvia; González-Cabrera, Joel; Ferré, Juan; Giordana, Barbara

    2006-01-01

    The pore-forming activity of Cry1Ab, Cry1Fa and Cry1Ca toxins and their interaction with leucine transport mediated by the K(+)/leucine cotransporter were studied in brush border membrane vesicles (BBMVs) isolated from the midgut of Ostrinia nubilalis and Sesamia nonagrioides. In both species, as in other Lepidoptera, leucine uptake by BBMVs can take place in the absence of cations, but it can also be driven by a K(+) gradient. Experiments with the voltage-sensitive fluorescent dye 3,3'-diethylthiacarbocyanine iodide proved that Cry1Ab, a Bacillus thuringiensis toxin active in vivo, enhanced the membrane permeability to potassium in O. nubilalis BBMVs. This result is in agreement with similar effects observed in S. nonagrioides BBMV incubated with various Cry1 toxins active in vivo. The effect of the above toxins was tested on the initial rate of 0.1 mM: leucine influx. Instead of an increase in leucine influx, a reduction was observed with the Cry1 toxins active in vivo. Cry1Ab and Cry1Fa, but not the inactive toxin Cry1Da, inhibited in a dose-dependent manner leucine uptake both in the absence and in the presence of a K(+) gradient, a clear indication that their effect is independent of the channel formed by the toxins and that this effect is exerted directly on the amino acid transport system.

  1. Relation between plasma and tissue parameters of leucine metabolism in fed and starved rats

    SciTech Connect

    Vazquez, J.A.; Paul, H.S.; Adibi, S.A.

    1986-06-01

    By use of a primed continuous infusion of (1-/sup 14/C)leucine, the authors investigated parameters of leucine metabolism in plasma, expired air, and tissues of fed and 48-h starved rats. The ratios of muscle to plasma specific activity of ..cap alpha..-ketoisocaproate (KIC) in fed and starved rats were not significantly different from 1. The ratio of muscle to plasma specific activity of leucine was also not significantly different from 1 in fed rats, but was significantly lower than 1 in starved rats. The rate of leucine oxidation was 28-34% higher when calculation was based on plasma KIC rather than leucine specific activity. However, starvation significantly increased the rate of leucine oxidation with either specific activity. The rates of leucine incorporation into whole-body protein, calculated as the difference between plasma leucine turnover and oxidation, were unaffected by starvation, but the incorporations into total protein measured directly were significantly decreased in liver and muscle. They conclude that a) leucine or KIC specific activity in muscle is better predicted by plasma KIC than leucine specific activity, and b) the difference between rates of plasma leucine turnover and oxidation does not appear to be a valid measurement of leucine incorporation into whole-body protein.

  2. Relationship between surface concentration of L-leucine and bulk powder properties in spray dried formulations.

    PubMed

    Mangal, Sharad; Meiser, Felix; Tan, Geoffrey; Gengenbach, Thomas; Denman, John; Rowles, Matthew R; Larson, Ian; Morton, David A V

    2015-08-01

    The amino acid L-leucine has been demonstrated to act as a lubricant and improve the dispersibility of otherwise cohesive fine particles. It was hypothesized that optimum surface L-leucine concentration is necessary to achieve optimal surface and bulk powder properties. Polyvinylpyrrolidone was spray dried with different concentration of L-leucine and the change in surface composition of the formulations was determined using X-ray photoelectron spectroscopy (XPS) and time of flight-secondary ion mass spectrometry (ToF-SIMS). The formulations were also subjected to powder X-ray diffraction analysis in order to understand the relationship between surface concentration and solid-state properties of L-leucine. In addition, the morphology, surface energy and bulk cohesion of spray dried formulations were also assessed to understand the relation between surface L-leucine concentration and surface and bulk properties. The surface concentration of L-leucine increased with higher feed concentrations and plateaued at about 10% L-leucine. Higher surface L-leucine concentration also resulted in the formation of larger L-leucine crystals and not much change in crystal size was noted above 10% L-leucine. A change in surface morphology of particles from spherical to increasingly corrugated was also observed with increasing surface l-leucine concentration. Specific collapsed/folded over particles were only seen in formulations with 10% or higher l-leucine feed concentration suggesting a change in particle surface formation process. In addition, bulk cohesion also reduced and approached a minimum with 10% L-leucine concentration. Thus, the surface concentration of L-leucine governs particle formation and optimum surface L-leucine concentration results in optimum surface and bulk powder properties.

  3. Effectiveness of an Increased Dose of Bovamine Compared to a Lower Dose to Reduce Salmonella in Fed Cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Category: Pre-harvest pathogen reduction Published: unpublished to date Objective: To examine the effect of increasing the probiotic dose from Bovamine® to Bovamine® Defend™ on the prevalence of Salmonella in pen environments, fecal samples and subiliac lymph nodes of fed cattle. Experimental ...

  4. Quantitative role of splanchnic region in leucine metabolism: L-(1-13C,15N)leucine and substrate balance studies

    SciTech Connect

    Yu, Y.M.; Wagner, D.A.; Tredget, E.E.; Walaszewski, J.A.; Burke, J.F.; Young, V.R. )

    1990-07-01

    The role of the splanchnic region (Sp) in whole body leucine metabolism was assessed in six chronically catheterized fasting mongrel dogs and in eight dogs during constant enteral feeding of a complete amino acid solution (0.24 g.kg-1.h-1). We used primed continuous intravenous infusions of L-(1-13C,15N)leucine and L-(1-14C)leucine and measurements of arteriovenous isotope and leucine balance across the gut, liver, and Sp. In the fasted condition, 3.5% of arterial leucine supply was oxidized in the Sp, accounting for 13% of total body leucine oxidation, with 10% by liver. With amino acid feeding (1) leucine carbon and nitrogen fluxes and oxidation were increased (P less than 0.01) at the whole body level; (2) the percent of whole body leucine oxidation occurring in the Sp and liver increased (P less than 0.01) to 41 and 27%, respectively; (3) fractional metabolic utilization of leucine delivered to the Sp was reduced (P less than 0.01) from 47 to 35%; (4) the deamination rate of leucine in the gut was increased (P less than 0.05), along with an increased reamination rate of alpha-ketoisocaproic acid in the Sp (P less than 0.05). These findings reveal that the Sp accounts for a small fraction of whole body leucine oxidation during the fasting condition, but it plays a quantitatively important role in total body leucine oxidation during amino acid feeding; the gut and liver play cooperative roles in controlling leucine supply to peripheral tissues.

  5. Postprandial leucine and insulin responses and toxicological effects of a novel whey protein hydrolysate-based supplement in rats

    PubMed Central

    2012-01-01

    The purpose of this study was: aim 1) compare insulin and leucine serum responses after feeding a novel hydrolyzed whey protein (WPH)-based supplement versus a whey protein isolate (WPI) in rats during the post-absorptive state, and aim 2) to perform a thorough toxicological analysis on rats that consume different doses of the novel WPH-based supplement over a 30-day period. In male Wistar rats (~250 g, n = 40), serum insulin and leucine concentrations were quantified up to 120 min after one human equivalent dose of a WPI or the WPH-based supplement. In a second cohort of rats (~250 g, n = 20), we examined serum/blood and liver/kidney histopathological markers after 30 days of feeding low (1human equivalent dose), medium (3 doses) and high (6 doses) amounts of the WPH-based supplement. In aim 1, higher leucine levels existed at 15 min after WPH vs. WPI ingestion (p = 0.04) followed by higher insulin concentrations at 60 min (p = 0.002). In aim 2, liver and kidney histopathology/toxicology markers were not different 30 days after feeding with low, medium, high dose WPH-based supplementation or water only. There were no between-condition differences in body fat or lean mass or circulating clinical chemistry markers following the 30-day feeding intervention in aim 2. In comparison to WPI, acute ingestion of a novel WPH-based supplement resulted in a higher transient leucine response with a sequential increase in insulin. Furthermore, chronic ingestion of the tested whey protein hydrolysate supplement appears safe. PMID:22672725

  6. Increasing the Therapeutic Ratio of Stereotactic Ablative Radiotherapy by Individualized Isotoxic Dose Prescription.

    PubMed

    Zindler, Jaap D; Thomas, Charles R; Hahn, Stephen M; Hoffmann, Aswin L; Troost, Esther G C; Lambin, Philippe

    2016-02-01

    To obtain a favorable tradeoff between treatment benefits and morbidity ("therapeutic ratio"), radiotherapy (RT) dose is prescribed according to the tumor volume, with the goal of controlling the disease while respecting normal tissue tolerance levels. We propose a new paradigm for tumor dose prescription in stereotactic ablative radiotherapy (SABR) based on organ-at-risk (OAR) tolerance levels called isotoxic dose prescription (IDP), which is derived from experiences and limitations of conventionally fractionated radiotherapy. With IDP, the radiation dose is prescribed based on the predefined level of normal tissue complication probability of a nearby dose-limiting OAR at a prespecified dose-volume constraint. Simultaneously, the prescribed total tumor dose (TTD) is maximized to the technically highest achievable level in order to increase the local tumor control probability (TCP). IDP is especially relevant for tumors located at eloquent locations or for large tumors in which severe toxicity has been described. IDP will result in a lower RT dose or a treatment scheduled with more fractions if the OAR tolerance level is exceeded, and potential dose escalation occurs when the OAR tolerance level allows it and when it is expected to be beneficial (if TCP < 90%). For patients with small tumors at noneloquent sites, the current SABR dose prescription already results in high rates of local control at low toxicity rates. In this review, the concept of IDP is described in the context of SABR.

  7. Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway.

    PubMed

    Saxton, Robert A; Knockenhauer, Kevin E; Wolfson, Rachel L; Chantranupong, Lynne; Pacold, Michael E; Wang, Tim; Schwartz, Thomas U; Sabatini, David M

    2016-01-01

    Eukaryotic cells coordinate growth with the availability of nutrients through the mechanistic target of rapamycin complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag guanosine triphosphatases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. Here we present the 2.7 angstrom crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucine leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. These results provide a structural mechanism of amino acid sensing by the mTORC1 pathway.

  8. Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway

    SciTech Connect

    Saxton, Robert A.; Knockenhauer, Kevin E.; Wolfson, Rachel L.; Chantranupong, Lynne; Pacold, Michael E.; Wang, Tim; Schwartz, Thomas U.; Sabatini, David M.

    2015-11-19

    Eukaryotic cells coordinate growth with the availability of nutrients through mTOR complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag GTPases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. We present the 2.7-Å crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucine leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. Lastly, these results provide a structural mechanism of amino acid sensing by the mTORC1 pathway.

  9. Inverse relationship of leucine flux and oxidation to free fatty acid availability in vivo.

    PubMed Central

    Tessari, P; Nissen, S L; Miles, J M; Haymond, M W

    1986-01-01

    To determine the effect of fatty acid availability on leucine metabolism, 14-h fasted dogs were infused with either glycerol or triglyceride plus heparin, and 46-h fasted dogs were infused with either nicotinic acid or nicotinic acid plus triglyceride and heparin. Leucine metabolism was assessed using a simultaneous infusion of L-[4,5-3H]leucine and alpha-[1-14C]ketoisocaproate. Leucine, alpha-ketoisocaproate (KIC), and totalleucine carbon (leucine plus KIC) flux and oxidation rates were calculated at steady state. In 14-h fasted animals, infusion of triglyceride and heparin increased plasma free fatty acids (FFA) by 0.7 mM (P less than 0.01) and decreased leucine (P less than 0.01), total leucine carbon flux (P less than 0.02), and oxidation (P less than 0.05). The estimated rate of leucine utilization not accounted for by oxidation and KIC flux decreased, but the changes were not significant. During glycerol infusion, leucine and KIC flux and oxidation did not change. In 46-h fasted dogs, nicotinic acid decreased FFA by 1.0 mM (P less than 0.01) and increased (P less than 0.05) the rate of leucine and total leucine carbon flux, but did not affect KIC flux. Leucine oxidation increased (P less than 0.01) by nearly threefold, whereas nonoxidized leucine utilization decreased. Infusion of triglyceride plus heparin together with nicotinic acid blunted some of the responses observed with nicotinic acid alone. In that changes in oxidation under steady state condition reflect changes in net leucine balance, these data suggest that FFA availability may positively affect the sparing of at least one essential amino acid and may influence whole body protein metabolism. PMID:3080479

  10. Increasing Use of Dose-Escalated External Beam Radiation Therapy for Men With Nonmetastatic Prostate Cancer

    SciTech Connect

    Swisher-McClure, Samuel; Mitra, Nandita; Woo, Kaitlin; Smaldone, Marc; Uzzo, Robert; Bekelman, Justin E.

    2014-05-01

    Purpose: To examine recent practice patterns, using a large national cancer registry, to understand the extent to which dose-escalated external beam radiation therapy (EBRT) has been incorporated into routine clinical practice for men with prostate cancer. Methods and Materials: We conducted a retrospective observational cohort study using the National Cancer Data Base, a nationwide oncology outcomes database in the United States. We identified 98,755 men diagnosed with nonmetastatic prostate cancer between 2006 and 2011 who received definitive EBRT and classified patients into National Comprehensive Cancer Network (NCCN) risk groups. We defined dose-escalated EBRT as total prescribed dose of ≥75.6 Gy. Using multivariable logistic regression, we examined the association of patient, clinical, and demographic characteristics with the use of dose-escalated EBRT. Results: Overall, 81.6% of men received dose-escalated EBRT during the study period. The use of dose-escalated EBRT did not vary substantially by NCCN risk group. Use of dose-escalated EBRT increased from 70.7% of patients receiving treatment in 2006 to 89.8% of patients receiving treatment in 2011. On multivariable analysis, year of diagnosis and use of intensity modulated radiation therapy were significantly associated with receipt of dose-escalated EBRT. Conclusions: Our study results indicate that dose-escalated EBRT has been widely adopted by radiation oncologists treating prostate cancer in the United States. The proportion of patients receiving dose-escalated EBRT increased nearly 20% between 2006 and 2011. We observed high utilization rates of dose-escalated EBRT within all disease risk groups. Adoption of intensity modulated radiation therapy was strongly associated with use of dose-escalated treatment.

  11. Leucine supplementation improves regeneration of skeletal muscles from old rats.

    PubMed

    Pereira, Marcelo G; Silva, Meiricris T; da Cunha, Fernanda M; Moriscot, Anselmo S; Aoki, Marcelo S; Miyabara, Elen H

    2015-12-01

    The decreased regenerative capacity of old skeletal muscles involves disrupted turnover of proteins. This study investigated whether leucine supplementation in old rats could improve muscle regenerative capacity. Young and old male Wistar rats were supplemented with leucine; then, the muscles were cryolesioned and examined after 3 and 10 days. Leucine supplementation attenuated the decrease in the expression of eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1) and eukaryotic translation initiation factor 4E (eIF4E) in young and old muscles on day 3 post-injury and promoted an increase in the cross-sectional area of regenerating myofibers from both young and old soleus muscles on day 10 post-injury. This supplementation decreased the levels of ubiquitinated proteins and increased the proteasome activity in young regenerating muscles, but the opposite effect was observed in old regenerating muscles. Moreover, leucine decreased the inflammation area and induced an increase in the number of proliferating satellite cells in both young and old muscles. Our results suggest that leucine supplementation improves the regeneration of skeletal muscles from old rats, through the preservation of certain biological responses upon leucine supplementation. Such responses comprise the decrease in the inflammation area, increase in the number of proliferating satellite cells and size of regenerating myofibers, combined with the modulation of components of the phosphoinositide 3-kinase/Akt-protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway and ubiquitin-proteasome system.

  12. Increased marihuana-induced fetotoxicity by a low dose of concomitant alcohol administration.

    PubMed

    Abel, E L; Dintcheff, B A

    1986-09-01

    Many pregnant women use both alcohol and marihuana. To evaluate the effects of this combination on fetotoxicity, pregnant mice in the experimental group were injected with a relatively low dose of alcohol (1 g/kg) and with one of two doses of marihuana extract (equivalent of 50 or 100 g/kg delta 9-THC). Control mice received marihuana extract or alcohol alone. The combination of alcohol and the high dose of marihuana produced a greater effect on fetotoxicity (83%) than either marihuana or alcohol alone or that due to the additive effects of either of these substances (63%). The combination of alcohol and the lower dose of marihuana extract did not increase fetotoxicity significantly. Doses of alcohol that are otherwise without effect on pregnancy may thus have the potential for greatly increasing the effects of drugs on pregnancy outcome.

  13. Dietary L-leucine supplementation enhances intestinal development in suckling piglets.

    PubMed

    Sun, Yuli; Wu, Zhenlong; Li, Wei; Zhang, Chen; Sun, Kaiji; Ji, Yun; Wang, Bin; Jiao, Ning; He, Beibei; Wang, Weiwei; Dai, Zhaolai; Wu, Guoyao

    2015-08-01

    L-Leucine is a signaling amino acid in animal metabolism. It is unknown whether supplementing L-leucine to breast-fed neonates may enhance their small-intestinal development. This hypothesis was tested with a piglet model. Seven-day-old sow-reared pigs with an average birth weight of 1.45 kg were assigned randomly to the control or leucine group (n = 30/group). Piglets in the leucine group were orally administrated with 1.4 g L-leucine/kg body weight, whereas piglets in the control group received isonitrogenous L-alanine, twice daily for 14 days. The supplemental L-leucine amounted to 200 % of L-leucine intake from sow's milk by 7-day-old pigs. At the end of the 2-week experiment, tissue samples were collected for determining intestinal morphology, expression of genes for intestinal leucine transporters (real-time RT-PCR and western blot analysis), and plasma metabolites and hormones. L-leucine administration increased (P < 0.05) villus height in the duodenum, an elevated ratio of villus height to crypt depth in the duodenum and ileum, plasma concentrations of leucine, glutamine and asparagine, as well as body-weight gains. mRNA levels for L-leucine transporters (SLC6A14, SLC6A19 and SLC7A9) and the abundance of the ATB(0,+) protein were increased (P < 0.05) but those for SLC7A7 mRNA and the LAT2 protein were decreased (P < 0.05) in the jejunum of leucine-supplemented piglets, compared with the control. Plasma concentrations of ammonia, urea, triglycerides, and growth-related hormones did not differ between the control and leucine groups. Collectively, these results indicate that L-leucine supplementation improves intestinal development and whole-body growth in suckling piglets with a normal birth weight.

  14. Synergistic Effects of Polyphenols and Methylxanthines with Leucine on AMPK/Sirtuin-Mediated Metabolism in Muscle Cells and Adipocytes

    PubMed Central

    Bruckbauer, Antje; Zemel, Michael B.

    2014-01-01

    The AMPK-Sirt1 pathway is an important regulator of energy metabolism and therefore a potential target for prevention and therapy of metabolic diseases. We recently demonstrated leucine and its metabolite β-hydroxy-β-methylbutyrate (HMB) to synergize with low-dose resveratrol (200 nM) to activate sirtuin signaling and stimulate energy metabolism. Here we show that leucine exerts a direct effect on Sirt1 kinetics, reducing its Km for NAD+ by >50% and enabling low doses of resveratrol to further activate the enzyme (p = 0.012). To test which structure elements of resveratrol are necessary for synergy, we assessed potential synergy of structurally similar and dissimilar polyphenols as well as other compounds converging on the same pathways with leucine using fatty acid oxidation (FAO) as screening tool. Dose-response curves for FAO were constructed and the highest non-effective dose (typically 1–10 nM) was used with either leucine (0.5 mM) or HMB (5 µM) to treat adipocytes and myotubes for 24 h. Significant synergy was detected for stilbenes with FAO increase in adipocytes by 60–70% (p<0.05) and in myotubes >2000% (p<0.01). Sirt1 and AMPK activities were stimulated by ∼65% (p<0.001) and ∼50% (p<0.03), respectively. Similarly, hydroxycinnamic acids and derivatives (chlorogenic, cinnamic, and ferulic acids) combined with leucine/HMB increased FAO (300–1300%, p<0.01), AMPK activity (50–150%, p<0.01), and Sirt1 activity (∼70%, p<0.001). In contrast, more complex polyphenol structures, such as ellagic acid and epigallocatechin gallate required higher concentrations (>1 µM) and exhibited little or no synergy. Thus, the six-carbon ring structure bound to a carboxylic group seems to be a necessary element for leucine/HMB synergy with other stilbenes and hydroxycinnamic acids to stimulate AMPK/Sirt1 dependent FAO; these effects occur at concentrations that produce no independent effects and are readily achievable via oral administration. PMID:24551237

  15. Stimulation of muscle protein synthesis by prolonged parenteral infusion of leucine is dependent on amino acid availability in neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The postprandial rise in amino acids, particularly leucine, stimulates muscle protein synthesis in neonates. Previously, we showed that a 1-h infusion of leucine increased protein synthesis, but this response was not sustained for 2 h unless the leucine-induced decrease in amino acids was prevented....

  16. Stimulation of skeletal muscle protein synthesis in neonatal pigs by long-term infusion of leucine is amino acid dependent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infusing leucine for 1 hr increases skeletal muscle protein synthesis in neonatal pigs, but this is not sustained for 2 h unless the leucine-induced fall in amino acids is prevented. We aimed to determine whether continuous leucine infusion can stimulate protein synthesis for a prolonged period whe...

  17. Is Increased Low-dose somatic Radiosensitivity Associated with Increased Transgenerational Germline Mutation

    SciTech Connect

    Brenner, David J.

    2008-10-02

    Using single-molecule polymerase chain reaction, the frequency of spontaneous and radiation-induced mutation at an expanded simple tandem repeat (ESTR) locus was studied in DNA samples extracted from sperm and bone marrow of Atm knockout (Atm+/–) heterozygous male mice. The frequency of spontaneous mutation in sperm and bone marrow in Atm+/– males did not significantly differ from that in wild-type BALB/c mice. Acute gamma-ray exposure did not affect ESTR mutation frequency in bone marrow and resulted in similar increases in sperm samples taken from Atm+/– and BALB/c males. Taken together, these results suggest that the Atm haploinsufficiency analyzed in our study does not affect spontaneous and radiation-induced ESTR mutation frequency in mice.

  18. Liver functional metabolomics discloses an action of L-leucine against Streptococcus iniae infection in tilapias.

    PubMed

    Ma, Yan-Mei; Yang, Man-Jun; Wang, Sanying; Li, Hui; Peng, Xuan-Xian

    2015-08-01

    Streptococcus iniae seriously affects the intensive farming of tilapias. Much work has been conducted on prevention and control of S. iniae infection, but little published information on the metabolic response is available in tilapias against the bacterial infection, and no metabolic modulation way may be adopted to control this disease. The present study used GC/MS based metabolomics to characterize the metabolic profiling of tilapias infected by a lethal dose (LD50) of S. iniae and determined two characteristic metabolomes separately responsible for the survival and dying fishes. A reversal changed metabolite, decreased and increased l-leucine in the dying and survival groups, respectively, was identified as a biomarker which featured the difference between the two metabolomes. More importantly, exogenous l-leucine could be used as a metabolic modulator to elevate survival ability of tilapias infected by S. iniae. These results indicate that tilapias mount metabolic strategies to deal with bacterial infection, which can be regulated by exogenous metabolites such as l-leucine. The present study establishes an alternative way, metabolic modulation, to cope with bacterial infections.

  19. A method to evaluate the dose increase in CT with iodinated contrast medium

    SciTech Connect

    Amato, Ernesto; Lizio, Domenico; Settineri, Nicola; Di Pasquale, Andrea; Salamone, Ignazio; Pandolfo, Ignazio

    2010-08-15

    Purpose: The objective of this study is to develop a method to calculate the relative dose increase when a computerized tomography scan (CT) is carried out after administration of iodinated contrast medium, with respect to the same CT scan in absence of contrast medium. Methods: A Monte Carlo simulation in GEANT4 of anthropomorphic neck and abdomen phantoms exposed to a simplified model of CT scanner was set up in order to calculate the increase of dose to thyroid, liver, spleen, kidneys, and pancreas as a function of the quantity of iodine accumulated; a series of experimental measurements of Hounsfield unit (HU) increment for known concentrations of iodinated contrast medium was carried out on a Siemens Sensation 16 CT scanner in order to obtain a relationship between the increment in HU and the relative dose increase in the organs studied. The authors applied such a method to calculate the average dose increase in three patients who underwent standard CT protocols consisting of one native scan in absence of contrast, followed by a contrast-enhanced scan in venous phase. Results: The authors validated their GEANT4 Monte Carlo simulation by comparing the resulting dose increases for iodine solutions in water with the ones presented in literature and with their experimental data obtained through a Roentgen therapy unit. The relative dose increases as a function of the iodine mass fraction accumulated and as a function of the Hounsfield unit increment between the contrast-enhanced scan and the native scan are presented. The data shown for the three patients exhibit an average relative dose increase between 22% for liver and 74% for kidneys; also, spleen (34%), pancreas (28%), and thyroid (48%) show a remarkable average increase. Conclusions: The method developed allows a simple evaluation of the dose increase when iodinated contrast medium is used in CT scans, basing on the increment in Hounsfield units observed on the patients' organs. Since many clinical protocols

  20. Parkinson's disease: low-dose haloperidol increases dopamine receptor sensitivity and clinical response.

    PubMed

    Hudson, Craig J; Seeman, Philip; Seeman, Mary V

    2014-01-01

    Background. It is known that ultra-low doses of haloperidol can cause dopamine supersensitivity of dopamine D2 receptors and related behaviour in animals. Objective. The objective was to determine whether a daily ultra-low dose of 40 micrograms of haloperidol could enhance the clinical action of levodopa in Parkinson's disease patients. Method. While continuing their daily treatment with levodopa, 16 patients with Parkinson's disease were followed weekly for six weeks. They received an add-on daily dose of 40 micrograms of haloperidol for the first two weeks only. The SPES/SCOPA scale (short scale for assessment of motor impairments and disabilities in Parkinson's disease) was administered before treatment and weekly throughout the trial. Results. The results showed a mean decrease in SPES/SCOPA scores after one week of the add-on treatment. Conclusion. SCOPA scores decreased after the addition of low-dose haloperidol to the standard daily levodopa dose. This finding is consistent with an increase in sensitivity of dopamine D2 receptors induced by haloperidol. Such treatment for Parkinson's disease may possibly permit the levodopa dose to be reduced and, thus, delay the onset of levodopa side effects.

  1. Effect of hyperammonemia on leucine and protein metabolism in rats.

    PubMed

    Holecek, M; Sprongl, L; Tichý, M

    2000-10-01

    The cause of muscle wasting and decreased plasma levels of branched chain amino acids (BCAA), valine, leucine, and isoleucine in liver cirrhosis is obscure. Here we have evaluated the effect of hyperammonemia. Rats were infused with either an ammonium acetate/bicarbonate mixture, a sodium acetate/bicarbonate mixture, or saline for 320 minutes. The parameters of leucine and protein metabolism were evaluated in the whole body and in several tissues using a primed constant intravenous infusion of L-[1-14C]leucine. Ammonium infusion caused an increase in ammonia and glutamine levels in plasma, a decrease in BCAA and alanine in plasma and skeletal muscle, a significant decrease in whole-body proteolysis and protein synthesis, and an increase in leucine oxidized fraction. A significant decrease in protein synthesis after ammonium infusion was observed in skeletal muscle while a nonsignificant effect was observed in liver, gut, heart, spleen, and kidneys. We conclude that the decrease in plasma BCAA after ammonia infusion is associated with decreased proteolysis and increased leucine oxidized fraction.

  2. L-Leucine and NO-mediated cardiovascular function.

    PubMed

    Yang, Ying; Wu, Zhenlong; Meininger, Cynthia J; Wu, Guoyao

    2015-03-01

    Reduced availability of nitric oxide (NO) in the vasculature is a major factor contributing to the impaired action of insulin on blood flow and, therefore, insulin resistance in obese and diabetic subjects. Available evidence shows that vascular insulin resistance plays an important role in the pathogenesis of cardiovascular disease, the leading cause of death in developed nations. Interestingly, increased concentrations of L-leucine in the plasma occur in obese humans and other animals with vascular dysfunction. Among branched-chain amino acids, L-leucine is unique in inhibiting NO synthesis from L-arginine in endothelial cells and may modulate cardiovascular homeostasis in insulin resistance. Results of recent studies indicate that L-leucine is an activator of glutamine:fructose-6-phosphate aminotransferase (GFAT), which is the first and a rate-controlling enzyme in the synthesis of glucosamine (an inhibitor of endothelial NO synthesis). Through stimulating the mammalian target of rapamycin signaling pathway and thus protein synthesis, L-leucine may enhance GFAT protein expression, thereby inhibiting NO synthesis in endothelial cells. We propose that reducing circulating levels of L-leucine or endothelial GFAT activity may provide a potentially novel strategy for preventing and/or treating cardiovascular disease in obese and diabetic subjects. Such means may include dietary supplementation with either α-ketoglutarate to enhance the catabolism of L-leucine in the small intestine and other tissues or with N-ethyl-L-glutamine to inhibit GFAT activity in endothelial cells. Preventing leucine-induced activation of GFAT by nutritional supplements or pharmaceutical drugs may contribute to improved cardiovascular function by enhancing vascular NO synthesis.

  3. Triennial growth symposium: Leucine acts as a nutrient signal to stimulate protein synthesis in neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The postprandial increases in AA and insulin independently stimulate protein synthesis in skeletal muscle of piglets. Leucine is an important mediator of the response to AA. We have shown that the postprandial increase in leucine, but not isoleucine or valine, acutely stimulates muscle protein synth...

  4. Stimulation of muscle protein synthesis by leucine is dependent on plasma amino acid availability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have reported that a physiological increase in plasma leucine increased translation initiation factor activity during 60- and 120-min leucine infusion. Muscle protein synthesis was stimulated at 60 min but not at 120 min, perhaps due to the decrease (-50%) in plasma essential amino acids (AA). ...

  5. Increased Skin Dose With the Use of a Custom Mattress for Prone Breast Radiotherapy

    SciTech Connect

    Becker, Stewart J. Patel, Rakesh R.; Mackie, Thomas R.

    2007-10-01

    The purpose of this study was to measure and compare the loss of buildup to the skin of the breast in the prone position due to 2 different positioning systems during tangential external beam irradiation. Two experiments were performed; one with a standard nylon-covered foam support and another with a novel helium-filled Mylar bag support. The choice of helium-filled Mylar was to reduce the contamination to as low as possible. The experiments were designed to allow a surface dose measurement and a depth dose profile with the pads placed in the path of the beam in front of the detector. All measurements were taken using a Capintec PS-033 thin-window parallel plate ionization chamber. The standard nylon-covered foam pad caused the surface dose to rise as it got closer to the skin. When the pad was directly touching the surface, the surface dose increased by 300% compared to the result when no pad was present. This loss of buildup to the surface was similar to that of a custom bolus material. The opposite effect occurred with the use of the helium-filled Mylar bag, namely the surface dose gradually decreased as the pad got closer to the phantom. When the Mylar pad was directly touching the phantom, the surface dose was decreased by 7% compared to when no pad was present. The use of a foam pad could potentially result in a significant higher dose to the skin, resulting in an enhanced acute skin reaction. Therefore, special care should be taken in this clinical scenario and further investigation of an air- or helium-based mylar support pad should be investigated in the context of definitive breast radiation treatment.

  6. High Dose Astaxanthin Lowers Blood Pressure and Increases Insulin Sensitivity in Rats: Are These Effects Interdependent?

    PubMed Central

    Preuss, Harry G.; Echard, Bobby; Yamashita, Eiji; Perricone, Nicholas V.

    2011-01-01

    The present investigation in Sprague-Dawley rats (SD) was designed to examine effects of astaxanthin (Asta) at different doses on elevated blood pressure (BP) and glucose-insulin perturbations produced by heavy sucrose ingestion. We also examined effects of Asta on BP during restraint stress. SD were divided into six groups each containing eight rats. All SD ate a basic diet of ground regular rat chow with sucrose added at 30% w/w. The Control group received only the basic diet containing added sucrose, while the other five groups each received the same diet with added test material: captopril, (30 mg/Kg), pioglitazone (15.0 mg/Kg), low Asta (25 mg/Kg), medium Asta (50 mg/kg) or high Asta (100 mg/Kg). Many tests were carried out to examine the mechanisms behind the effects of Asta on BP (serum ACE activity, losartan challenge, and LNAME challenge) and the glucose-insulin system (glucose tolerance, HOMA measurement, and insulin challenge). In SD, a relatively low dose of Asta decreased SBP, but produced no major changes in the glucose-insulin system simulating results from a previous study using Zucker Fatty Rats. Increasing the dose of Asta resulted in both a lowering of elevated systolic BP and enhanced insulin sensitivity determined by many different estimations. BP lowering was consistent with changes in the renin-angiotensin (RAS) and nitric oxide (NO) systems. At the examined doses of each, captopril lowered BP in SD without influencing glucose-insulin metabolism, whereas pioglitazone favorably affected glucose-insulin metabolism while showing essentially no effects on BP. Accordingly, Asta beneficially affects both sucrose-induced elevations of BP and insulin resistance at relatively high doses in SD. Also, Asta at higher doses lessens restraint stress, whereas, captopril and pioglitazone did not at the doses examined, even though they influenced the BP and glucose-insulin systems respectively. PMID:21326955

  7. Low-dose gamma irradiation of food protein increases its allergenicity in a chronic oral challenge.

    PubMed

    Vaz, A F M; Souza, M P; Medeiros, P L; Melo, A M M A; Silva-Lucca, R A; Santana, L A; Oliva, M L V; Perez, K R; Cuccovia, I M; Correia, M T S

    2013-01-01

    Few chronic food protein models have described the relationship between allergenicity and the molecular structure of food protein after physical processing. The effect of γ-radiation on the structure of food protein was measured by fluorescence, circular dichroism and microcalorimetry. BALB/c mice were intraperitoneally sensitized and then given non-irradiated and irradiated Con-A by daily gavage for 28days. The tendency to form insoluble amorphous aggregates and partially unfolded species was observed after irradiation. The administration of non-irradiated and irradiated samples at low-dose significantly increased weight loss as well as plasma levels of eotaxin in animals repeatedly exposed to Con-A. Significant lymphocytic infiltrate filling completely the stroma of microvilli and tubular glands was observed in the small intestinal of the group given Con-A irradiated at a low dose. This phenotype was not observed in animals treated with Con-A irradiated at a high dose.

  8. Increased dose near the skin due to electromagnetic surface beacon transponder.

    PubMed

    Ahn, Kang-Hyun; Manger, Ryan; Halpern, Howard J; Aydogan, Bulent

    2015-05-08

    The purpose of this study was to evaluate the increased dose near the skin from an electromagnetic surface beacon transponder, which is used for localization and tracking organ motion. The bolus effect due to the copper coil surface beacon was evaluated with radiographic film measurements and Monte Carlo simulations. Various beam incidence angles were evaluated for both 6 MV and 18 MV experimentally. We performed simulations using a general-purpose Monte Carlo code MCNPX (Monte Carlo N-Particle) to supplement the experimental data. We modeled the surface beacon geometry using the actual mass of the glass vial and copper coil placed in its L-shaped polyethylene terephthalate tubing casing. Film dosimetry measured factors of 2.2 and 3.0 enhancement in the surface dose for normally incident 6 MV and 18 MV beams, respectively. Although surface dose further increased with incidence angle, the relative contribution from the bolus effect was reduced at the oblique incidence. The enhancement factors were 1.5 and 1.8 for 6 MV and 18 MV, respectively, at an incidence angle of 60°. Monte Carlo simulation confirmed the experimental results and indicated that the epidermal skin dose can reach approximately 50% of the dose at dmax at normal incidence. The overall effect could be acceptable considering the skin dose enhancement is confined to a small area (~ 1 cm2), and can be further reduced by using an opposite beam technique. Further clinical studies are justified in order to study the dosimetric benefit versus possible cosmetic effects of the surface beacon. One such clinical situation would be intact breast radiation therapy, especially large-breasted women.

  9. Activation of mammalian target of rapamycin signaling in skeletal muscle of neonatal chicks: effects of dietary leucine and age.

    PubMed

    Deng, Huiling; Zheng, Aijuan; Liu, Guohua; Chang, Wenhuan; Zhang, Shu; Cai, Huiyi

    2014-01-01

    The mammalian target of rapamycin (mTOR) signaling pathway is necessary for cellular protein synthesis regulation. Leucine was reported to stimulate muscle protein synthesis in mammalian embryos and neonates, but in higher animals (chickens) the effect of dietary leucine on mTOR signaling is unknown. Thus, we investigated the effects of dietary leucine and age on mRNA expression and phosphorylation of mTOR as well as its downstream targets, ribosomal protein S6 kinase (S6K1) and eukaryotic initiation factor 4E binding protein 1 (4E-BP1) in chick pectoral muscles. One hundred eighty newly hatched male chicks were randomly assigned to 1 of 3 dietary leucine treatment groups (1.43, 1.73, and 2.03% leucine) for 14 d, respectively. Each treatment group consisted of 6 cages with 10 chicks each. On d 3, 7, and 14, plasma insulin and leucine were measured and target gene expression and phosphorylation was assessed. Dietary leucine influenced plasma leucine but not insulin, and plasma leucine and insulin declined with chick age. The mTOR, S6K1, and 4E-BP1 mRNA expression and phosphorylation within chick pectoral muscles were upregulated with increased dietary leucine but downregulated with increased chick age. Thus, high dietary leucine activates target of rapamycin signaling pathways in skeletal muscle of neonatal chicks to stimulate muscle protein synthesis, and this pathway is attenuated with aging.

  10. Comparison of the effects of recombinant human insulin-like growth factor-I and insulin on glucose and leucine kinetics in humans.

    PubMed Central

    Laager, R; Ninnis, R; Keller, U

    1993-01-01

    To compare the metabolic effects of elevated plasma concentrations of IGF-I and insulin, overnight-fasted normal subjects were studied twice, once receiving IGF-I and once insulin at doses that resulted in identical increases in glucose uptake during 8-h euglycemic clamping. Recombinant human IGF-I or insulin were infused in one group at high doses (30 micrograms/kg per h IGF-I or 0.23 nmol/kg per h insulin) and in another group at low doses (5 micrograms/kg per h IGF-I or 0.04 nmol/kg per h insulin). Glucose rate of disappearance (measured by [6,6-D2]-glucose infusions) increased from baseline by 239 +/- 16% during high dose IGF-I vs 197 +/- 18% during insulin (P = 0.021 vs IGF-I). Hepatic glucose production decreased by 37 +/- 6% during high dose IGF-I vs 89 +/- 13% during insulin (P = 0.0028 vs IGF-I). IGF-I suppressed whole body leucine flux ([1-13C]-leucine infusion technique) more than insulin (42 +/- 4 vs 32 +/- 3% during high doses, P = 0.0082). Leucine oxidation rate decreased during high dose IGF-I more than during insulin (55 +/- 4 vs 32 +/- 6%, P = 0.0001). The decreases of plasma concentrations of free fatty acids, acetoacetate, and beta-hydroxybutyrate after 8 h of IGF-I and insulin administration were similar. Plasma C-peptide levels decreased by 57 +/- 4% during high doses of IGF-I vs 36 +/- 6% during insulin (P = 0.005 vs IGF-I). The present data demonstrate that, compared to insulin, an acute increase in plasma IGF-I levels results in preferential enhancement of peripheral glucose utilization, diminished suppression of hepatic glucose production, augmented decrease of whole body protein breakdown (leucine flux), and of irreversible leucine catabolism but in similar antilipolytic effects. The data suggest that insulin-like effects of IGF-I in humans are mediated in part via IGF-I receptors and in part via insulin receptors. PMID:8408642

  11. Effect of Increasing Doses of Saw Palmetto on Lower Urinary Tract Symptoms: A Randomized Trial

    PubMed Central

    Barry, Michael J.; Meleth, Sreelatha; Lee, Jeannette Y.; Kreder, Karl J.; Avins, Andrew L.; Nickel, J. Curtis; Roehrborn, Claus G.; Crawford, E. David; Foster, Harris E.; Kaplan, Steven A.; McCullough, Andrew; Andriole, Gerald L.; Naslund, Michael J.; Williams, O. Dale; Kusek, John W.; Meyers, Catherine M.; Betz, Joseph M.; Cantor, Alan; McVary, Kevin T.

    2012-01-01

    Context Saw palmetto fruit extracts are widely used for treating lower urinary tract symptoms attributed to benign prostatic hyperplasia. However, recent clinical trials have questioned their efficacy, at least at standard doses (320 mg daily). Objective To determine the effect of a saw palmetto extract at up to three times the standard dose on lower urinary tract symptoms attributed to benign prostatic hyperplasia. Design Multicenter placebo-controlled randomized trial conducted from June, 2008 through October, 2010. Setting Eleven North American clinical sites. Participants Were men at least 45 years old, with a peak urinary flow rate ≥ 4 ml/sec, an AUA Symptom Index (AUASI) score ≥ 8 and ≤ 24, and no exclusions. Interventions One, two, and then three 320 mg daily doses of saw palmetto extract or placebo, with dose increases at 24 and 48 weeks. Main Outcome Measures Primary outcome was the difference in AUASI score from baseline to 72 weeks. Secondary outcomes were measures of urinary bother; nocturia; uroflow; postvoid residual; prostate-specific antigen; participants’ global assessments; and indices of sexual function, continence, sleep quality, and prostatitis symptoms. Results From baseline to 72 weeks, mean AUASI scores decreased from 14.4 to 12.2 points with saw palmetto and from 14.7 to 11.7 points with placebo. The group mean difference in AUASI score change from baseline to 72 weeks between the saw palmetto and placebo groups was 0.79 points favoring placebo (bound of the 95% confidence interval most favorable to saw palmetto was 1.77 points, one-sided P=0.91). Saw palmetto was no more effective than placebo for any secondary outcome. No attributable side effects were identified. Conclusions Increasing doses of a saw palmetto fruit extract did not reduce lower urinary tract symptoms more than placebo. (CAMUS study number NCT00603304 http://www.ClinicalTrials.gov) PMID:21954478

  12. Long-term leucine induced stimulation of muscle protein synthesis is amino acid dependent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infusing leucine for 1 h increases skeletal muscle protein synthesis in the neonate, but this is not sustained for 2 h unless the corresponding fall in amino acids is prevented. This study aimed to determine whether a continuous leucine infusion can stimulate protein synthesis for a prolonged period...

  13. Protein-leucine ingestion activates a regenerative inflammo-myogenic transcriptome in skeletal muscle following intense endurance exercise.

    PubMed

    Rowlands, David S; Nelson, Andre R; Raymond, Frederic; Metairon, Sylviane; Mansourian, Robert; Clarke, Jim; Stellingwerff, Trent; Phillips, Stuart M

    2016-01-01

    Protein-leucine supplement ingestion following strenuous endurance exercise accentuates skeletal-muscle protein synthesis and adaptive molecular responses, but the underlying transcriptome is uncharacterized. In a randomized single-blind triple-crossover design, 12 trained men completed 100 min of high-intensity cycling then ingested 70/15/180/30 g protein-leucine-carbohydrate-fat (15LEU), 23/5/180/30 g (5LEU), or 0/0/274/30 g (CON) beverages during the first 90 min of a 240 min recovery period. Vastus lateralis muscle samples (30 and 240 min postexercise) underwent transcriptome analysis by microarray followed by bioinformatic analysis. Gene expression was regulated by protein-leucine in a dose-dependent manner affecting the inflammatory response and muscle growth and development. At 30 min, 15LEU and 5LEU vs. CON activated transcriptome networks with gene-set functions involving cell-cycle arrest (Z-score 2.0-2.7, P < 0.01), leukocyte maturation (1.7, P = 0.007), cell viability (2.4, P = 0.005), promyogenic networks encompassing myocyte differentiation and myogenin (MYOD1, MYOG), and a proteinaceous extracellular matrix, adhesion, and development program correlated with plasma lysine, arginine, tyrosine, taurine, glutamic acid, and asparagine concentrations. High protein-leucine dose (15LEU-5LEU) activated an IL-1I-centered proinflammatory network and leukocyte migration, differentiation, and survival functions (2.0-2.6, <0.001). By 240 min, the protein-leucine transcriptome was anti-inflammatory and promyogenic (IL-6, NF- β, SMAD, STAT3 network inhibition), with overrepresented functions including decreased leukocyte migration and connective tissue development (-1.8-2.4, P < 0.01), increased apoptosis of myeloid and muscle cells (2.2-3.0, P < 0.002), and cell metabolism (2.0-2.4, P < 0.01). The analysis suggests protein-leucine ingestion modulates inflammatory-myogenic regenerative processes during skeletal muscle recovery from endurance exercise. Further

  14. Dietary leucine requirement of juvenile Japanese seabass ( Lateolabrax japonicus)

    NASA Astrophysics Data System (ADS)

    Li, Yan; Cheng, Zhenyan; Mai, Kangsen; Ai, Qinghui

    2015-02-01

    A 56-day feeding trial was conducted to examine the dietary leucine requirement of juvenile Japanese seabass in seawater floating net cages (1.5 m × 1.5 m × 2.0 m). Six isonitrogenous (crude protein 40%) and isoenergetic (gross energy 20 kJ g-1) diets were formulated to contain different concentrations of leucine (0.9%, 1.49%, 2.07%, 2.70%, 3.30% and 3.88% of dry matter). Crystalline L-amino acids were supplemented to simulate the whole body amino acid pattern of Japanese seabass except for leucine. Three groups (30 fish individuals each, 8.0 g ± 0.20 g in initial weight) were fed to apparent satiation at 5:00 and 17:30 every day. During the experimental period, the water temperature ranged from 26 to 32δC and salinity from 26 to 30, and the dissolved oxygen was maintained at 7 mg L-1. The results showed that weight gain ( WG), nitrogen retention ( NR), feed efficiency ( FE) and protein efficiency ratio ( PER) were significantly increased when dietary leucine was increased from 0.90% to 2.70% of dry matter, and then declined. WG was the highest when fish were fed D4 containing 2.70% of leucine. No significant differences were observed in body composition among dietary treatments ( P > 0.05). Considering the change of WG, the optimum dietary leucine requirement of juvenile Japanese seabass was either 2.39% of dry matter or 5.68% of dietary protein.

  15. Leucine improves protein nutritional status and regulates hepatic lipid metabolism in calorie-restricted rats.

    PubMed

    Pedroso, João Alfredo B; Nishimura, Luciana Sigueta; de Matos-Neto, Emídio Marques; Donato, Jose; Tirapegui, Julio

    2014-06-01

    Several studies have highlighted the potential of leucine supplementation for the treatment of metabolic diseases including type 2 diabetes and obesity. Caloric restriction is a common approach to improve the health in diabetic and obese subjects. However, very few studies assessed the effects of leucine supplementation in calorie-restricted animals. Rats were subjected to a 30% calorie-restricted diet for 6 weeks to study the effects of leucine supplementation on protein status markers and lipid metabolism. Caloric restriction reduced the body weight. However, increased leucine intake preserved body lean mass and protein mass and improved protein anabolism as indicated by the increased circulating levels of albumin and insulin-like growth factor-1 (IGF-1), and the liver expression of albumin and IGF-1 messenger RNA. Leucine supplementation also increased the circulating levels of interleukin-6 and leptin but did not affect the tumour necrosis factor-α and monocyte chemotactic protein-1 concentrations. Ketone bodies were increased in rats consuming a leucine-rich diet, but we observed no changes in cholesterol or triglycerides concentrations. Caloric restriction reduced the liver expression of peroxisome proliferator activated receptor-α and glucose-6-phosphatase, whereas leucine supplementation increased the liver expression of 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA) reductase and sterol regulatory element-binding transcription factor 1. A leucine-rich diet during caloric restriction preserved whole body protein mass and improved markers of protein anabolism. In addition, leucine modulated the hepatic lipid metabolism. These results indicate that increased leucine intake may be useful in preventing excessive protein waste in conditions of large weight loss.

  16. Acute Low-Dose Caffeine Supplementation Increases Electromyographic Fatigue Threshold in Healthy Men.

    PubMed

    Morse, Jacob J; Pallaska, Gramos; Pierce, Patrick R; Fields, Travis M; Galen, Sujay S; Malek, Moh H

    2016-11-01

    Morse, JJ, Pallaska, G, Pierce, PR, Fields, TM, Galen, SS, and Malek, MH. Acute low-dose caffeine supplementation increases electromyographic fatigue threshold in healthy men. J Strength Cond Res 30(11): 3236-3241, 2016-The purpose of this study is to determine whether consumption of a single low-dose caffeine drink will delay the onset of the electromyographic fatigue threshold (EMGFT) in the superficial quadriceps femoris muscles. We hypothesize that the EMGFT values for the caffeine condition will be significantly higher than the EMGFT values for the placebo condition. On separate occasions, 10 physically active men performed incremental single-leg knee-extensor ergometry 1 hour after caffeine (200 mg) or placebo consumption. The EMGFT was determined for each participant for both conditions. The results indicated a significant increase for maximal power output (16%; p = 0.004) and EMGFT (45%; p = 0.004) in the caffeine condition compared with placebo. These findings suggest that acute low-dose caffeine supplementation delays neuromuscular fatigue in the quadriceps femoris muscles.

  17. Dietary L-leucine improves the anemia in a mouse model for Diamond-Blackfan anemia.

    PubMed

    Jaako, Pekka; Debnath, Shubhranshu; Olsson, Karin; Bryder, David; Flygare, Johan; Karlsson, Stefan

    2012-09-13

    Diamond-Blackfan anemia (DBA) is a congenital erythroid hypoplasia caused by a functional haploinsufficiency of genes encoding for ribosomal proteins. Recently, a case study reported a patient who became transfusion-independent in response to treatment with the amino acid L-leucine. Therefore, we have validated the therapeutic effect of L-leucine using our recently generated mouse model for RPS19-deficient DBA. Administration of L-leucine significantly improved the anemia in Rps19-deficient mice (19% improvement in hemoglobin concentration; 18% increase in the number of erythrocytes), increased the bone marrow cellularity, and alleviated stress hematopoiesis. Furthermore, the therapeutic response to L-leucine appeared specific for Rps19-deficient hematopoiesis and was associated with down-regulation of p53 activity. Our study supports the rationale for clinical trials of L-leucine as a therapeutic agent for DBA.

  18. Leucine supplementation via drinking water reduces atherosclerotic lesions in apoE null mice

    PubMed Central

    Zhao, Yang; Dai, Xiao-yan; Zhou, Zhou; Zhao, Ge-xin; Wang, Xian; Xu, Ming-jiang

    2016-01-01

    Aim: Recent evidence suggests that the essential amino acid leucine may be involved in systemic cholesterol metabolism. In this study, we investigated the effects of leucine supplementation on the development of atherosclerosis in apoE null mice. Methods: ApoE null mice were fed with chow supplemented with leucine (1.5% w/v) in drinking water for 8 week. Aortic atherosclerotic lesions were examined using Oil Red O staining. Plasma lipoprotein-cholesterol levels were measured with fast protein liquid chromatography. Hepatic gene expression was detected using real-time PCR and Western blot analyses. Results: Leucine supplementation resulted in 57.6% reduction of aortic atherosclerotic lesion area in apoE null mice, accompanied by 41.2% decrease of serum LDL-C levels and 40.2% increase of serum HDL-C levels. The body weight, food intake and blood glucose level were not affected by leucine supplementation. Furthermore, leucine supplementation increased the expression of Abcg5 and Abcg8 (that were involved in hepatic cholesterol efflux) by 1.28- and 0.86-fold, respectively, and significantly increased their protein levels. Leucine supplementation also increased the expression of Srebf1, Scd1 and Pgc1b (that were involved in hepatic triglyceride metabolism) by 3.73-, 1.35- and 1.71-fold, respectively. Consequently, leucine supplementation resulted in 51.77% reduction of liver cholesterol content and 2.2-fold increase of liver triglyceride content. Additionally, leucine supplementation did not affect the serum levels of IL-6, IFN-γ, TNF-α, IL-10 and IL-12, but markedly decreased the serum level of MCP-1. Conclusion: Leucine supplementation effectively attenuates atherosclerosis in apoE null mice by improving the plasma lipid profile and reducing systemic inflammation. PMID:26687933

  19. Leucine metabolism in cirrhotic patients with hepatic encephalopathy

    SciTech Connect

    McGhee, A.S.

    1985-01-01

    The purpose of this study was to determine whether increased oxidation of or protein synthesis requiring leucine occurs in cirrhotic patients. Five control subjects and four subjects with cirrhosis were equilibrated on a baseline diet (0.6 g protein per kg ideal body weight (IBW)) with sufficient nonprotein calories to preclude negative nitrogen balance. An additional four patients were equilibrated on the same type of diet with a higher protein level (0.75 g per kg IBW). Control subjects and the patients were then studied during continuous infusion of L-(/sup 15/N, 1-/sup 13/C) leucine in the fasted state and, in the fed state, with a Propac diet which had the same distribution of energy nutrients as the baseline diets. Plasma levels of L-(/sup 15/N, 1-/sup 13/C), L-(1-/sup 13/C) and L-(/sup 15/N) leucine were measured during isotopic steady state by gas chromatography-mass spectrometry and fractional excretion of /sup 13/CO/sup 2/ in breath samples were analyzed by isotopic ratio mass spectrometry. During the fasted and fed states leucine metabolism was measured to quantitate rates of nitrogen flux (Q/sub N/), carbon flux (Q/sub c/) and oxidation to carbon dioxide and water (C). From these measured values, proteins breakdown (B), protein synthesis (S), deamination (X/sup 0/) and reamination (X/sub N/) were calculated. The results showed that protein synthesis and leucine metabolism were identical in controls and patients when both were fed a diet with 0.6 g protein/kg IBW and maintenance level of nonprotein calories. The data also showed that leucine metabolism can be quantitatively and reproducibly measured in subjects with cirrhosis.

  20. Leucine Modulates Mitochondrial Biogenesis and SIRT1-AMPK Signaling in C2C12 Myotubes

    PubMed Central

    Liang, Chunzi; Curry, Benjamin J.; Brown, Patricia L.; Zemel, Michael B.

    2014-01-01

    Previous studies from this laboratory demonstrate that dietary leucine protects against high fat diet-induced mitochondrial impairments and stimulates mitochondrial biogenesis and energy partitioning from adipocytes to muscle cells through SIRT1-mediated mechanisms. Moreover, β-hydroxy-β-methyl butyrate (HMB), a metabolite of leucine, has been reported to activate AMPK synergistically with resveratrol in C2C12 myotubes. Therefore, we hypothesize that leucine-induced activation of SIRT1 and AMPK is the central event that links the upregulated mitochondrial biogenesis and fatty acid oxidation in skeletal muscle. Thus, C2C12 myotubes were treated with leucine (0.5 mM), alanine (0.5 mM), valine (0.5 mM), EX527 (SIRT1 inhibitor, 25 μM), and Compound C (AMPK inhibitor, 25 μM) alone or in combination to determine the roles of AMPK and SIRT1 in leucine-modulation of energy metabolism. Leucine significantly increased mitochondrial content, mitochondrial biogenesis-related genes expression, fatty acid oxidation, SIRT1 activity and gene expression, and AMPK phosphorylation in C2C12 myotubes compared to the controls, while EX527 and Compound C markedly attenuated these effects. Furthermore, leucine treatment for 24 hours resulted in time-dependent increases in cellular NAD+, SIRT1 activity, and p-AMPK level, with SIRT1 activation preceding that of AMPK, indicating that leucine activation of SIRT1, rather than AMPK, is the primary event. PMID:25400942

  1. Down Regulation of Asparagine Synthetase and 3-Phosphoglycerate Dehydrogenase, and the Up-Regulation of Serine Dehydratase in Rat Liver from Intake of Excess Amount of Leucine Are Not Related to Leucine-Caused Amino Acid Imbalance.

    PubMed

    Yoshimura, Ryoji; Takai, Marie; Namaki, Hiroya; Minami, Kimiko; Imamura, Wataru; Kato, Hisanori; Kamei, Yasutomi; Kanamoto, Ryuhei

    2015-01-01

    Asparagine synthetase (ASNS), 3-phosphoglycerate dehydrogenase (PHGDH) and serine dehydratase (SDS) in rat liver are expressed in response to protein and amino acid intake. In the present study, we examined the expression of these enzymes in relation to amino acid imbalance caused by leucine. Rats were subjected to leucine administration in the diet or orally between meals. Consumption of more than 2% leucine in a 6% casein diet suppressed food intake and caused growth retardation in a dose-dependent manner, but this was not seen in a 12% or 40% casein diet. ASNS and PHGDH expression in the liver was significantly induced by the 6% casein diet and was suppressed by leucine in a dose-dependent manner, whereas the SDS expression was induced. These effects were leucine specific and not seen with ingestion of isoleucine or valine. However, leucine orally administered between meals did not change the food intake or growth of rats fed a 6% casein die, though it similarly affected the expression of ASNS, PHGDH and SDS in the liver. These results suggest that the growth retardation caused by leucine imbalance was mainly because of the suppression of food intake, and demonstrated that there are no causal relationships between ASNS, PHGDH and SDS expression and amino acid imbalance caused by leucine.

  2. Genetic studies of leucine transport in mammalian cells.

    PubMed

    Shotwell, M A; Lobatón, C D; Collarini, E J; Moreno, A; Giles, R E; Oxender, D L

    1984-05-15

    We have taken two approaches to the study of the genetics of leucine transport in mammalian cells. First, from a mutant Chinese hamster ovary cell line that has a temperature-sensitive leucyl-tRNA synthetase, we isolated temperature-resistant revertants with increased leucine transport activity. This transport elevation is reflected by increased Vmax values of leucine uptake and unchanged Km values of uptake. The temperature resistance in each revertant appears to result from the increased transport and not from any change in the leucyl-tRNA synthetase. We conclude that in each revertant there is a stable derepression of amino acid transport system L. In a second approach, we started with a Chinese hamster-human hybrid strain formed by the fusion of a temperature-sensitive leucyl-tRNA synthetase mutant hamster cell line and normal human leukocytes. From this temperature-sensitive hybrid strain we selected temperature-resistant hybrids, one class of which we found to have greatly elevated leucine transport activity. We have allowed human chromosomes to segregate from these high-transport hybrids, promoted by the presence of low concentrations of colcemid. The loss of the high-transport phenotype coincides with the loss of a single small human chromosome, which we are attempting to identify by using G-11 and G-banding staining techniques.

  3. Measurement of L-(1-/sup 14/C)leucine kinetics in splanchnic and leg tissues in humans. Effect of amino acid infusion

    SciTech Connect

    Gelfand, R.A.; Glickman, M.G.; Castellino, P.; Louard, R.J.; DeFronzo, R.A.

    1988-10-01

    Although whole-body leucine flux is widely measured to study body protein turnover in humans, the contribution of specific tissues to the total-body measurement remains unknown. By combining the organ-balance technique with the systemic infusion of L-(1-14C)leucine, we quantitated leucine production and disposal by splanchnic and leg tissues and by the whole body, simultaneously, in six normal men before and during amino acid infusion. At steady state, disposal of arterial leucine by splanchnic and leg tissues was calculated from the percent extraction (E) of L-(1-14C)leucine counts: uptake = E x (Leu)a x flow. Tissue release of cold leucine (from protein turnover) into vein was calculated as the difference between leucine uptake and the net tissue leucine balance. In the postabsorptive state, despite substantial (P less than .01) extraction of L-(1-14C)leucine by splanchnic (23 +/- 1%) and leg (18 +/- 2%) tissues, net leucine balance across both tissue beds was small, indicating active simultaneous disposal and production of leucine at nearly equivalent rates. Splanchnic tissues accounted for approximately 50% of the measured total-body leucine flux. During amino acid infusion, the net leucine balance across splanchnic and leg tissues became positive, reflecting not only an increase in leucine uptake but also a marked suppression (by approximately 50%, P less than .02) of cold leucine release. This reduction in splanchnic and leg leucine release was indicated by a sharp decline in whole-body endogenous leucine flux.

  4. A (14)C-leucine absorption, distribution, metabolism and excretion (ADME) study in adult Sprague-Dawley rat reveals β-hydroxy-β-methylbutyrate as a metabolite.

    PubMed

    Lee, Anthony J; Beno, David W A; Zhang, Xiaolin; Shapiro, Robin; Mason, Mark; Mason-Bright, Tanita; Surber, Bruce; Edens, Neilé K

    2015-05-01

    Leucine is an essential branched-chain amino acid that acts as a substrate for protein synthesis and as a signaling molecule. Leucine not incorporated into muscle protein is ultimately oxidized through intermediates such as β-hydroxy-β-methylbutyrate (HMB) which itself is reported to enhance muscle mass and function in rats and humans. HMB has been reported in the plasma following oral leucine administration in sheep and pigs but not in Sprague-Dawley rats, the standard preclinical model. Therefore, we conducted radiolabeled absorption, distribution, metabolism and excretion (ADME) studies in rats using a low (3 mg/kg) or high dose (1,000 mg/kg) of (14)C-leucine. Blood, tissue, and urine samples were analyzed for (14)C-leucine and its metabolites by HPLC-MS. Our results show for the first time that (14)C-HMB appears in plasma and urine of rats following an oral dose of (14)C-leucine. (14)C-leucine appears in plasma as (14)C-α-ketoisocaproic acid (KIC) with a slower time course than (14)C-HMB, a putative product of KIC. Further, two novel metabolites of leucine were detected in urine, N-acetyl leucine and glycyl leucine. Mass balance studies demonstrate that excretory routes accounted for no more than 0.9 % of the radiolabel and approximately 61 % of the dose was recovered in the carcass. Approximately 65 % of the dose was recovered in total, suggesting that approximately one-third of the leucine dose is oxidized to CO2. In conclusion, this study demonstrates endogenous production of HMB from leucine in adult rats, a standard preclinical model used to guide design of clinical trials in nutrition.

  5. Low doses of isosorbide mononitrate attenuate the postprandial increase in portal pressure in patients with cirrhosis.

    PubMed

    Bellis, Lia; Berzigotti, Annalisa; Abraldes, Juan G; Moitinho, Eduardo; García-Pagán, Juan C; Bosch, Jaime; Rodés, Juan

    2003-02-01

    Postprandial hyperemia is associated with a significant increase in portal pressure in cirrhosis, which may contribute to progressive dilation and rupture of gastroesophageal varices. In cirrhosis, an insufficient hepatic production of nitric oxide (NO) may impair the expected hepatic vasodilatory response to increased blood flow, further exaggerating the postprandial increase in portal pressure. This study was aimed at investigating whether low doses of an oral NO donor might counteract the postprandial peak in portal pressure. Twenty-three portal hypertensive cirrhotics, 8 of them under propranolol therapy, were randomized to receive orally 5-isosorbide mononitrate (ISMN; 10 mg; n = 11) or placebo (n = 12) and a standard liquid meal 15 minutes later. Hepatic venous pressure gradient (HVPG), mean arterial pressure (MAP), and hepatic blood flow (HBF) were measured at baseline and 15, 30, and 45 minutes after a meal. ISMN significantly attenuated the postprandial increase in portal pressure as compared with placebo (peak HVPG increase: 2.4 +/- 1.4 mm Hg vs. 5.2 +/- 2.1 mm Hg, P =.002). Percentual increases in HBF were similar in both groups. MAP decreased slightly in ISMN group (-7.5% +/-.5%; P <.01 vs. baseline). These effects were also observed in patients on chronic propranolol therapy. In conclusion, hepatic NO supplementation by low doses of ISMN effectively reduces the postprandial increase of portal pressure in cirrhosis, with only a mild effect on arterial pressure. The same was observed in patients receiving propranolol. Our results suggest that therapeutic strategies based on selective hepatic NO delivery may improve the treatment of portal hypertension.

  6. Calculations of increased solar UV fluxes and DUV doses due to stratospheric-ozone depletions

    SciTech Connect

    Zardecki, A.; Gerstl, S.A.W.

    1982-02-01

    Accurate radiative transfer calculations are performed in the middle ultraviolet spectral region for aerosol-loaded atmospheres with the goal of determining the solar irradiance at the ground and quantifying the irradiance perturbations due to the presence of aerosols and various ozone depletions. The extent of the increase of UV-B radiation as a function of wave-length and solar zenith angle is calculated for five model atmospheres. In addition, the damaging ultraviolet dose rates and radiation amplification factors are evaluated at different latitudes and seasons for erythemal and DNA action spectra.

  7. Bi-Directional Effect of Increasing Doses of Baclofen on Reinforcement Learning

    PubMed Central

    Terrier, Jean; Ort, Andres; Yvon, Cédric; Saj, Arnaud; Vuilleumier, Patrik; Lüscher, Christian

    2011-01-01

    In rodents as well as in humans, efficient reinforcement learning depends on dopamine (DA) released from ventral tegmental area (VTA) neurons. It has been shown that in brain slices of mice, GABAB-receptor agonists at low concentrations increase the firing frequency of VTA–DA neurons, while high concentrations reduce the firing frequency. It remains however elusive whether baclofen can modulate reinforcement learning in humans. Here, in a double-blind study in 34 healthy human volunteers, we tested the effects of a low and a high concentration of oral baclofen, a high affinity GABAB-receptor agonist, in a gambling task associated with monetary reward. A low (20 mg) dose of baclofen increased the efficiency of reward-associated learning but had no effect on the avoidance of monetary loss. A high (50 mg) dose of baclofen on the other hand did not affect the learning curve. At the end of the task, subjects who received 20 mg baclofen p.o. were more accurate in choosing the symbol linked to the highest probability of earning money compared to the control group (89.55 ± 1.39 vs. 81.07 ± 1.55%, p = 0.002). Our results support a model where baclofen, at low concentrations, causes a disinhibition of DA neurons, increases DA levels and thus facilitates reinforcement learning. PMID:21811448

  8. Bi-directional effect of increasing doses of baclofen on reinforcement learning.

    PubMed

    Terrier, Jean; Ort, Andres; Yvon, Cédric; Saj, Arnaud; Vuilleumier, Patrik; Lüscher, Christian

    2011-01-01

    In rodents as well as in humans, efficient reinforcement learning depends on dopamine (DA) released from ventral tegmental area (VTA) neurons. It has been shown that in brain slices of mice, GABA(B)-receptor agonists at low concentrations increase the firing frequency of VTA-DA neurons, while high concentrations reduce the firing frequency. It remains however elusive whether baclofen can modulate reinforcement learning in humans. Here, in a double-blind study in 34 healthy human volunteers, we tested the effects of a low and a high concentration of oral baclofen, a high affinity GABA(B)-receptor agonist, in a gambling task associated with monetary reward. A low (20 mg) dose of baclofen increased the efficiency of reward-associated learning but had no effect on the avoidance of monetary loss. A high (50 mg) dose of baclofen on the other hand did not affect the learning curve. At the end of the task, subjects who received 20 mg baclofen p.o. were more accurate in choosing the symbol linked to the highest probability of earning money compared to the control group (89.55 ± 1.39 vs. 81.07 ± 1.55%, p = 0.002). Our results support a model where baclofen, at low concentrations, causes a disinhibition of DA neurons, increases DA levels and thus facilitates reinforcement learning.

  9. A Randomized Controlled Trial of Increased Dose and Frequency of Albendazole with Standard Dose DEC for Treatment of Wuchereria bancrofti Microfilaremics in Odisha, India

    PubMed Central

    Kerketa, Anna Salomi; Maharana, Antaryami; Panda, Sudanshu S; Mohanty, Prafulla Chandra; Horton, John; Ramachandran, Cherubala P

    2015-01-01

    Although current programmes to eliminate lymphatic filariasis have made significant progress it may be necessary to use different approaches to achieve the global goal, especially where compliance has been poor and ‘hot spots’ of continued infection exist. In the absence of alternative drugs, the use of higher or more frequent dosing with the existing drugs needs to be explored. We examined the effect of higher and/or more frequent dosing with albendazole with a fixed 300mg dose of diethylcarbamazine in a Wuchereria bancrofti endemic area in Odisha, India. Following screening, 104 consenting adults were randomly assigned to treatment with the standard regimen annually for 24 months (S1), or annually with increased dose (800mg albendazole)(H1) or with increased frequency (6 monthly) with either standard (S2) or increased (H2) dose. Pre-treatment microfilaria counts (GM) ranged from 348 to 459 mf/ml. Subjects were followed using microfilaria counts, OG4C3 antigen levels and ultrasound scanning for adult worm nests. Microfilarial counts tended to decrease more rapidly with higher or more frequent dosing at all time points. At 12 months, Mf clearance was marginally greater with the high dose regimens, while by 24 months, there was a trend to higher Mf clearance in the arm with increased frequency and 800mg of albendazole (76.9%) compared to other arms, (S1:64%, S2:69.2% & H1:73.1%). Although higher and/or more frequent dosing showed a trend towards a greater decline in antigenemia and clearance of “nests”, all regimens demonstrated the potential macrofilaricidal effect of the combination. The higher doses of albendazole did not result in a greater number or more severe side effects. The alternative regimens could be useful in the later stages of existing elimination programmes or achieving elimination more rapidly in areas where programmes have yet to start. PMID:25781977

  10. A single dose of EGLN1 siRNA yields increased erythropoiesis in nonhuman primates.

    PubMed

    Abrams, Marc T; Koser, Martin; Burchard, Julja; Strapps, Walter; Mehmet, Huseyin; Gindy, Marian; Zaller, Dennis; Sepp-Lorenzino, Laura; Stickens, Dominique

    2014-12-01

    Decreased production of erythropoietin (EPO) causes anemia in patients with chronic kidney disease, and recombinant human EPO is used to treat renal failure associated anemia. The liver, the main EPO-producing organ in utero, maintains the capacity to produce EPO in the adult but in insufficient quantities to restore hemoglobin levels to normal in patients with impaired renal function. Inhibition of prolyl-4-hydroxylase domain (PHD) proteins is known to cause an increase in EPO production through its effects on hypoxia inducible factor. Here, we utilized small interfering RNA (siRNA) targeting EGLN1, the gene encoding the PHD2 protein, to investigate the phenotypic consequences in nonhuman primates. A single, well-tolerated intravenous dose of an optimized EGLN1 siRNA encapsulated in a lipid nanoparticle formulation caused robust mRNA silencing in the liver, leading to increases in serum EPO and hemoglobin. The siRNA-induced erythropoiesis was dose-dependent and was sustained for at least 2 months. These data point to the potential for an RNA interference-based, liver-targeted therapeutic approach for the treatment of anemia.

  11. Study on increasing production of natural silk by using low dose irradiation

    SciTech Connect

    Ruiying, Z.; Yinfen, Z.; Dingzhu, C.; Jinxian, R.

    1985-01-01

    Radiation effect on silkworm irradiated by low dose fast neutron and ..gamma..-ray emitted from Ra-Be neutron source are reported. It is shown that increasing production of natural silk can only be obtained by irradiation under specified conditions. It was found that an appropriate fluence employed could lead to increase hatching rate of silkworm eggs, make silkworms' bodies strong, grow fast, possess high disease resistance and reduce the whole stadium by 1/2 to 2 1/2 days. In addition, the irradiated silkworm can be expected to spin bigger cocoons with thick layers and the quality of cocoon silk are remarkable improved. The application of irradiation technique has now been extended to the suburbs of Beijing and welcomed by sericulturist.

  12. A Heterospecific Leucine Zipper Tetramer

    SciTech Connect

    Deng, Y.; Liu, J; Zheng, Q; Li, Q; Kallenbach, N; Lu, M

    2008-01-01

    Protein-protein interactions dictate the assembly of the macromolecular complexes essential for functional networks and cellular behavior. Elucidating principles of molecular recognition governing important interfaces such as coiled coils is a challenging goal for structural and systems biology. We report here that two valine-containing mutants of the GCN4 leucine zipper that fold individually as four-stranded coiled coils associate preferentially in mixtures to form an antiparallel, heterotetrameric structure. X-ray crystallographic analysis reveals that the coinciding hydrophobic interfaces of the hetero- and homotetramers differ in detail, explaining their partnering and structural specificity. Equilibrium disulfide exchange and thermal denaturation experiments show that the 50-fold preference for heterospecificity results from a combination of preferential packing and hydrophobicity. The extent of preference is sensitive to the side chains comprising the interface. Thus, heterotypic versus homotypic interaction specificity in coiled coils reflects a delicate balance in complementarity of shape and chemistry of the participating side chains.

  13. Extracting the normal lung dose-response curve from clinical DVH data: a possible role for low dose hyper-radiosensitivity, increased radioresistance

    NASA Astrophysics Data System (ADS)

    Gordon, J. J.; Snyder, K.; Zhong, H.; Barton, K.; Sun, Z.; Chetty, I. J.; Matuszak, M.; Ten Haken, R. K.

    2015-09-01

    In conventionally fractionated radiation therapy for lung cancer, radiation pneumonitis’ (RP) dependence on the normal lung dose-volume histogram (DVH) is not well understood. Complication models alternatively make RP a function of a summary statistic, such as mean lung dose (MLD). This work searches over damage profiles, which quantify sub-volume damage as a function of dose. Profiles that achieve best RP predictive accuracy on a clinical dataset are hypothesized to approximate DVH dependence. Step function damage rate profiles R(D) are generated, having discrete steps at several dose points. A range of profiles is sampled by varying the step heights and dose point locations. Normal lung damage is the integral of R(D) with the cumulative DVH. Each profile is used in conjunction with a damage cutoff to predict grade 2 plus (G2+) RP for DVHs from a University of Michigan clinical trial dataset consisting of 89 CFRT patients, of which 17 were diagnosed with G2+ RP. Optimal profiles achieve a modest increase in predictive accuracy—erroneous RP predictions are reduced from 11 (using MLD) to 8. A novel result is that optimal profiles have a similar distinctive shape: enhanced damage contribution from low doses (<20 Gy), a flat contribution from doses in the range ~20-40 Gy, then a further enhanced contribution from doses above 40 Gy. These features resemble the hyper-radiosensitivity / increased radioresistance (HRS/IRR) observed in some cell survival curves, which can be modeled using Joiner’s induced repair model. A novel search strategy is employed, which has the potential to estimate RP dependence on the normal lung DVH. When applied to a clinical dataset, identified profiles share a characteristic shape, which resembles HRS/IRR. This suggests that normal lung may have enhanced sensitivity to low doses, and that this sensitivity can affect RP risk.

  14. Extracting the normal lung dose-response curve from clinical DVH data: a possible role for low dose hyper-radiosensitivity, increased radioresistance.

    PubMed

    Gordon, J J; Snyder, K; Zhong, H; Barton, K; Sun, Z; Chetty, I J; Matuszak, M; Ten Haken, R K

    2015-09-07

    In conventionally fractionated radiation therapy for lung cancer, radiation pneumonitis' (RP) dependence on the normal lung dose-volume histogram (DVH) is not well understood. Complication models alternatively make RP a function of a summary statistic, such as mean lung dose (MLD). This work searches over damage profiles, which quantify sub-volume damage as a function of dose. Profiles that achieve best RP predictive accuracy on a clinical dataset are hypothesized to approximate DVH dependence.Step function damage rate profiles R(D) are generated, having discrete steps at several dose points. A range of profiles is sampled by varying the step heights and dose point locations. Normal lung damage is the integral of R(D) with the cumulative DVH. Each profile is used in conjunction with a damage cutoff to predict grade 2 plus (G2+) RP for DVHs from a University of Michigan clinical trial dataset consisting of 89 CFRT patients, of which 17 were diagnosed with G2+ RP.Optimal profiles achieve a modest increase in predictive accuracy--erroneous RP predictions are reduced from 11 (using MLD) to 8. A novel result is that optimal profiles have a similar distinctive shape: enhanced damage contribution from low doses (<20 Gy), a flat contribution from doses in the range ~20-40 Gy, then a further enhanced contribution from doses above 40 Gy. These features resemble the hyper-radiosensitivity / increased radioresistance (HRS/IRR) observed in some cell survival curves, which can be modeled using Joiner's induced repair model.A novel search strategy is employed, which has the potential to estimate RP dependence on the normal lung DVH. When applied to a clinical dataset, identified profiles share a characteristic shape, which resembles HRS/IRR. This suggests that normal lung may have enhanced sensitivity to low doses, and that this sensitivity can affect RP risk.

  15. Phosphorylation of 4EBP by oral leucine administration was suppressed in the skeletal muscle of PGC-1α knockout mice.

    PubMed

    Yoshimura, Ryoji; Minami, Kimiko; Matsuda, Junichiro; Sawada, Naoki; Miura, Shinji; Kamei, Yasutomi

    2016-01-01

    Leucine is known to increase mTOR-mediated phosphorylation of 4EBP. In this study, leucine was administered to skeletal muscle-PGC-1α knockout mice. We observed attenuated 4EBP phosphorylation in the skeletal muscle, but not in the liver, of the PGC-1α knockout mice. These data suggest that skeletal muscle-PGC-1α is important for leucine-mediated mTOR activation and protein biosynthesis.

  16. High-dose thalidomide increases the risk of peripheral neuropathy in the treatment of ankylosing spondylitis.

    PubMed

    Xue, Hong-Xia; Fu, Wen-Yi; Cui, Hua-Dong; Yang, Li-Li; Zhang, Ning; Zhao, Li-Juan

    2015-05-01

    Thalidomide is an effective drug for the treatment of ankylosing spondylitis but might induce peripheral neuropathy. This major adverse reaction has attracted much concern. The current study aimed to observe the incidence of thalidomide-induced peripheral neuropathy among ankylosing spondylitis patients for 1 year after treatment. In this study, 207 ankylosing spondylitis cases received thalidomide treatment, while 116 ankylosing spondylitis cases received other treatments. Results showed that the incidence of thalidomide-induced peripheral neuropathy in the thalidomide group was higher than that in the non-thalidomide group. There was no significant difference in the incidence of neuropathy between the < 6 months medication and ≥ 6 months medication groups. There were no differences in the mean age, gender, or daily dose between the two groups. The incidence of peripheral neuropathy among patients receiving 25, 50, 75, or 100 mg thalidomide per day was 4.6%, 8.5%, 17.1%, 21.7%, respectively. The incidence was significantly different between the groups receiving 25 mg and 100 mg thalidomide. In conclusion, thalidomide can induce peripheral neuropathy within 1 year after treatment of ankylosing spondylitis; however, age and gender have no obvious impact on the incidence of peripheral neuropathy. The incidence of peripheral neuropathy is associated with increasing daily doses of thalidomide.

  17. Low dose nicotine treatment during early adolescence increases subsequent cocaine reward.

    PubMed

    McQuown, Susan C; Belluzzi, James D; Leslie, Frances M

    2007-01-01

    Adolescence is a critical period for the initiation of drug use, starting with tobacco and alcohol and progressing to marijuana and other illicit drugs. These findings have led to the suggestion that tobacco and alcohol are 'gateway' drugs that sensitize maturing reward pathways to the effects of illicit substances such as cocaine. To test this hypothesis, we have examined whether low-dose nicotine pretreatment alters acquisition of cocaine self-administration in adolescents more than in adults. Male and female Sprague-Dawley rats, aged postnatal day (P) 28 or P86, were given two daily intravenous injections of nicotine (0.03 mg/kg/0.1 ml) or saline for 4 days. At P32 and P90, rats were placed in self-administration chambers and tested for acquisition of cocaine (0.2 or 0.5 mg/kg/inj) for 5 days. Data were collapsed across cocaine dose and sex since there was no significant effect of these variables. Adolescent rats pretreated with nicotine exhibited significantly greater cocaine-reinforced responding as compared to saline controls or adults (p<0.01). This drug pretreatment effect did not generalize to all rewards, since nicotine did not increase responding for sucrose pellets in adolescents. These findings provide evidence that the adolescent brain is uniquely vulnerable to the effects of nicotine on subsequent drug reward.

  18. Leucine supplementation accelerates connective tissue repair of injured tibialis anterior muscle.

    PubMed

    Pereira, Marcelo G; Silva, Meiricris T; Carlassara, Eduardo O C; Gonçalves, Dawit A; Abrahamsohn, Paulo A; Kettelhut, Isis C; Moriscot, Anselmo S; Aoki, Marcelo S; Miyabara, Elen H

    2014-09-29

    This study investigated the effect of leucine supplementation on the skeletal muscle regenerative process, focusing on the remodeling of connective tissue of the fast twitch muscle tibialis anterior (TA). Young male Wistar rats were supplemented with leucine (1.35 g/kg per day); then, TA muscles from the left hind limb were cryolesioned and examined after 10 days. Although leucine supplementation induced increased protein synthesis, it was not sufficient to promote an increase in the cross-sectional area (CSA) of regenerating myofibers (p > 0.05) from TA muscles. However, leucine supplementation reduced the amount of collagen and the activation of phosphorylated transforming growth factor-β receptor type I (TβR-I) and Smad2/3 in regenerating muscles (p < 0.05). Leucine also reduced neonatal myosin heavy chain (MyHC-n) (p < 0.05), increased adult MyHC-II expression (p < 0.05) and prevented the decrease in maximum tetanic strength in regenerating TA muscles (p < 0.05). Our results suggest that leucine supplementation accelerates connective tissue repair and consequent function of regenerating TA through the attenuation of TβR-I and Smad2/3 activation. Therefore, future studies are warranted to investigate leucine supplementation as a nutritional strategy to prevent or attenuate muscle fibrosis in patients with several muscle diseases.

  19. Repeated exposure to moderate doses of ethanol augments hippocampal glutamate neurotransmission by increasing release

    PubMed Central

    Chefer, Vladimir; Meis, Jennifer; Wang, Grace; Kuzmin, Alexander; Bakalkin, Georgy; Shippenberg, Toni

    2013-01-01

    The present study used conventional and quantitative microdialysis to assess glutamatergic and GABAergic neurotransmission in the hippocampal CA3 area of the rat following a moderate-dose ethanol treatment regimen. Male Wistar rats received 3.4 g/kg of ethanol or water for 6 days via gastric gavage. Microdialysis experiments commenced 2 days later. Basal and depolarization-induced glutamate overflow were significantly elevated in ethanol-treated animals. Basal and depolarization-induced gamma-aminobutyric acid (GABA) overflow were unaltered. Quantitative no-net-flux microdialysis was used to determine if changes in dialysate glutamate levels following ethanol administration are due to an increase in release or a decrease in uptake.To confirm the validity of this method for quantifying basal glutamate dynamics, extracellular concentrations of glutamate and the extraction fraction, which reflects changes in analyte clearance, were quantified in response to retro-dialysis of the glutamate uptake blocker trans-pyrrolidine-2,4-dicarboxylic acid (tPDC). tPDC significantly decreased the extraction fraction for glutamate, resulting in augmented extracellular glutamate concentrations. Repeated ethanol administration did not alter the glutamate extraction fraction. However, extracellular glutamate concentrations were significantly elevated, indicating that glutamate release is increased as a consequence of repeated ethanol administration. These data demonstrate that repeated bouts of moderate ethanol consumption alter basal glutamate dynamics in the CA3 region of the dorsal hippocampus. Basal glutamate release is augmented, whereas glutamate uptake is unchanged. Furthermore, they suggest that dysregulation of glutamate transmission in this region may contribute to the previously documented deficits in cognitive function associated with moderate dose ethanol use. PMID:21182572

  20. Medium doses of daily vitamin D decrease falls and higher doses of daily vitamin D3 increase falls: A randomized clinical trial.

    PubMed

    Smith, Lynette M; Gallagher, J Christopher; Suiter, Corinna

    2017-03-18

    Falls are a serious health problem in the aging population. Because low levels of vitamin D have been associated with increased fall rates, many trials have been performed with vitamin D; two meta-analyses showed either a small effect or no effect of vitamin D on falls. We conducted a study of the effect of vitamin D on serum 25 hydroxyvitamin D (25OHD) and data on falls was collected as a secondary outcome. In a 12-month double blind randomized placebo trial, elderly women, mean age 66 years, were randomized to one of seven daily oral doses of vitamin D or placebo. The main inclusion criterion for study was a baseline serum 25OHD<20ng/ml (50nmol/L). A history of falls was collected at baseline and fall events were collected every 3 months. Results showed that the effect of vitamin D on falls followed a U-shaped curve whether analyzed by dose or serum 25OHD levels. There was no decrease in falls on low vitamin D doses 400, 800 IU, a significant decrease on medium doses 1600, 2400,3200 IU (p=0.020) and no decrease on high doses 4000, 4800 IU compared to placebo (p=0.55). When compared to 12-month serum 25OHD quintiles, the faller rate was 60% in the lowest quintile <25ng/ml (<50nmol/L), 21% in the low middle quintile 32-38ng/ml (80-95nmo/L), 72% in the high middle quintile 38-46ng/ml (95-115nmo/L) and 45% in the highest quintile 46-66ng/ml (115-165nmol/L). In the subgroup with a fall history, fall rates were 68% on low dose, 27% on medium doses and 100% on higher doses. Fall rates on high doses were increased compared to medium doses (Odds Ratio 5.6.95% CI: 2.1-14.8). In summary, the maximum decrease in falls corresponds to a 12- month serum 25OHD of 32-38ng/ml (80-95nmol/L) and faller rates increase as serum 25OHD exceed 40-45ng/ml (100-112.5nmol/L). The Tolerable upper limit (TUL) recently increased in 2010 from 2000 to 4000 IU/day may need to be reduced in elderly women especially in those with a fall history.

  1. A Leucine Zipper Motif Essential for Gating of Hyperpolarization-activated Channels*

    PubMed Central

    Wemhöner, Konstantin; Silbernagel, Nicole; Marzian, Stefanie; Netter, Michael F.; Rinné, Susanne; Stansfeld, Phillip J.; Decher, Niels

    2012-01-01

    Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are pacemakers in cardiac myocytes and neurons. Although their membrane topology closely resembles that of voltage-gated K+ channels, the mechanism of their unique gating behavior in response to hyperpolarization is still poorly understood. We have identified a highly conserved leucine zipper motif in the S5 segment of HCN family members. In order to study the role of this motif for channel function, the leucine residues of the zipper were individually mutated to alanine, arginine, or glutamine residues. Leucine zipper mutants traffic to the plasma membrane, but the channels lose their sensitivity to open upon hyperpolarization. Thus, our data indicate that the leucine zipper is an important molecular determinant for hyperpolarization-activated channel gating. Residues of the leucine zipper interact with the adjacent S6 segment of the channel. This interaction is essential for voltage-dependent gating of the channel. The lower part of the leucine zipper, at the intracellular mouth of the channel, is important for stabilizing the closed state. Mutations at these sites increase current amplitudes or result in channels with deficient closing and increased min-Po. Our data are further supported by homology models of the open and closed state of the HCN2 channel pore. Thus, we conclude that the leucine zipper of HCN channels is a major determinant for hyperpolarization-activated channel gating. PMID:23048023

  2. Food deprivation increases the low-dose locomotor stimulant response to ethanol in Drosophila melanogaster.

    PubMed

    Kliethermes, Christopher L

    2013-10-01

    Acute and chronic states of food deprivation result in increased sensitivity to a variety of natural reinforcers as well as to drugs of abuse. Food deprived animals show increased locomotor activity during periods of food deprivation, as well as increased locomotor stimulant responses to drugs of abuse, including cocaine, amphetamine, morphine, and ethanol, implying that drugs of abuse act in part on neural systems that underlie responses towards food. To determine whether this effect extends to an invertebrate, highly genetically tractable animal, the locomotor stimulant effects of low dose ethanol were assessed under a variety of feeding conditions in the fruit fly, Drosophila melanogaster. Food deprivation resulted in strain specific increases in ethanol-stimulated locomotor activity in most strains tested, although elevated baseline activity confounded interpretation in some strains. Experiments conducted with Canton S flies found that the effects of food deprivation on the locomotor stimulant response to ethanol increased with the duration of deprivation, and could be blocked by refeeding the flies with standard food or sucrose, but not yeast, immediately prior to the ethanol exposure. Life-span extending dietary depletion procedures or previous periods of food deprivation did not affect the response to ethanol, indicating that only animals in an acutely food deprived state are more sensitive to the stimulant effects of ethanol. These results suggest that increased sensitivity to the stimulant effects of some drugs of abuse might reflect an evolutionarily conserved neural mechanism that underlies behavioral responses to natural reinforcers and drugs of abuse. The identification of this mechanism, and the genes that underlie its development and function, will constitute a novel approach towards the study of alcohol abuse and dependence.

  3. Diuretics enhance effects of increased dose of candesartan on ambulatory blood pressure reduction in Japanese patients with uncontrolled hypertension treated with medium-dose angiotensin II receptor blockers.

    PubMed

    Sakima, Atsushi; Kita, Toshihiro; Nakada, Seigo; Yokota, Naoto; Tamaki, Noboru; Etoh, Takuma; Shimokubo, Toru; Kitamura, Kazuo; Takishita, Shuichi; Ohya, Yusuke

    2014-01-01

    Abstract Although blockade of the renin-angiotensin system by increasing the dose of angiotensin II receptor blockers (ARBs) is recommended to achieve clinical benefits in terms of blood pressure (BP) control and cardiovascular and renal outcomes, the effect of this increased dose on ambulatory BP monitoring has not been evaluated completely in Japanese patients with uncontrolled hypertension undergoing medium-dose ARB therapy. The primary objective of this study was to examine the effect of the relatively high dose of the ARB candesartan (12 mg/day) on 24-h systolic BP and the attainment of target BP levels in uncontrolled hypertension treated with a medium dose of ARBs. A total of 146 hypertensive patients (age: 69.9 ± 9.3 years; females: 65.8%) completed the study. After switching to candesartan at 12 mg/day, all these BP measurements decreased significantly (p<0.001). Attainment of the target office BP (p=0.0014) and 24-h BP levels (p=0.0296) also improved significantly. Subgroup analysis indicated that the reduction of 24-h systolic BP was larger in patients treated with diuretics than those without (p=0.0206). Multivariate analysis revealed a significant correlation between the combined ARB and diuretic therapy, and the change in 24-h systolic BP irrespective of preceding ARBs. In conclusion, the switching therapy to increased dose of candesartan caused significant reductions in office and ambulatory BP levels, and improved the attainment of target BP levels in patients with uncontrolled hypertension treated with a medium dose of ARBs. Combination with diuretics enhanced this effect.

  4. Increase of Escherichia coli inoculum doses induces faster innate immune response in primiparous cows.

    PubMed

    Vangroenweghe, F; Rainard, P; Paape, M; Duchateau, L; Burvenich, C

    2004-12-01

    The objective of the current study was to evaluate the dynamics of infection and the immunological response to varying numbers of Escherichia coli injected into the mammary glands of primiparous cows during the periparturient period. Primiparous cows have been shown to be more resistant to intramammary E. coli challenge, and an increase of the inoculum dose by 2 log10 units induced a more rapid clinical response and clearance of the organisms. Recognition of lipopolysaccharide (LPS) is a key event in the innate immunity response to gram-negative infection and is mediated by the accessory molecules CD14 and LPS-binding protein (LBP). Primiparous cows were inoculated with 1 x 10(4) (Group A; n=8) or 1 x 10(6) (Group B; n=8) cfu E. coli P4:O32 in their 2 left quarters during the periparturient period. Clinical examination and analysis of blood and milk parameters, including IL-8, complement fragment 5a (C5a), LBP, and soluble CD14 (sCD14), were performed from d -4 to d +3 relative to infection. Primiparous cows in Group B initiated a more rapid clinical response following intramammary infection (IMI), resulting in typical clinical signs and changes in blood and milk parameters approximately 3 h earlier compared with primiparous cows in Group A. Based on average milk production in the noninfected quarters on d +2 postinoculation, all heifers reacted as moderate responders. Distinct differences in the kinetic patterns of rectal temperature, somatic cell count (SCC), IL-8, C5a, LBP, and sCD14 were observed between both groups during the early phase of inflammation. Both C5a and IL-8 increased before cellular influx into the infected glands, followed by increases in sCD14 and LBP. In conclusion, primiparous cows are able to clear an intramammary E. coli infection efficiently. Moreover, increasing the inoculum dose induces a more rapid inflammatory reaction, mainly because of early activation of the innate host immune response.

  5. L-leucine supplementation worsens the adiposity of already obese rats by promoting a hypothalamic pattern of gene expression that favors fat accumulation.

    PubMed

    Zampieri, Thais T; Torres-Leal, Francisco L; Campaña, Amanda B; Lima, Fabio B; Donato, Jose

    2014-04-02

    Several studies showed that l-leucine supplementation reduces adiposity when provided before the onset of obesity. We studied rats that were exposed to a high-fat diet (HFD) for 10 weeks before they started to receive l-leucine supplementation. Fat mass was increased in l-leucine-supplemented rats consuming the HFD. Accordingly, l-leucine produced a hypothalamic pattern of gene expression that favors fat accumulation. In conclusion, l-leucine supplementation worsened the adiposity of rats previously exposed to HFD possibly by central mechanisms.

  6. Ingestion of a moderately high caffeine dose before exercise increases postexercise energy expenditure.

    PubMed

    Fernández-Elías, Valentín E; Del Coso, Juan; Hamouti, Nassim; Ortega, Juan F; Muñoz, Gloria; Muñoz-Guerra, Jesus; Mora-Rodríguez, Ricardo

    2015-02-01

    Caffeine is an ergogenic aid widely used before and during prolonged exercise. Due to its prolonged biological half-life caffeine effects could remain after exercise. We aimed to investigate the metabolic, respiratory, and cardiovascular postexercise responses to preexercise graded caffeine ingestion. Twelve aerobically trained subjects (mean VO₂max = 54 ± 7 ml · min⁻¹ · kg⁻¹) cycled for 60-min at 75% VO₂max after ingesting placebo (0 mg of caffeine per kg of body weight) or 0.5, 1.5, 3.0 and 4.5 mg · kg⁻¹ on five occasions. During the 3 hr postexercise, heart rate, blood pressure, glucose, lactate, and fatty acids were analyzed. None of these variables were statistically affected by preexercise caffeine ingestion between 0.5 and 4.5 mg · kg⁻¹. However, ingestion of 4.5 mg · kg⁻¹ of caffeine raised postexercise energy expenditure 15% above placebo (233 ± 58 vs. 202 ± 49 kcal/3 hr; p < .05). Ventilation and tidal volume were elevated after the 4.5 mg · kg⁻¹ caffeine dose above placebo (9.2 ± 2.5 L · min⁻¹ and 0.67 ± 0.29 L · breath⁻¹ vs. 7.8 ± 1.5 L · min⁻¹ and 0.56 ± 0.20 L · breath⁻¹, respectively; p < .05). Ventilation correlated with tidal volume (r = .45; p < .05) and energy expenditure (r = .72; p < .05). In summary, preexercise ingestion of ergogenic caffeine doses do not alter postexercise cardiovascular responses. However, ingestion of 4.5 mg · kg⁻¹ of caffeine raises 3-hr postexercise energy expenditure (i.e., 31 kcal) likely through increased energy cost of ventilation.

  7. Stunted growth, increased mortality, and liver tumors in offspring of polybrominated biphenyl (PBB) dosed sherman rats.

    PubMed

    Groce, D F; Kimbrough, R D

    1984-01-01

    Firemaster FF-1, a polybrominated biphenyl (PBB) mixture, was dissolved in corn oil and given as a dose of 200 mg/kg body weight to Sherman rats on d 7 and 14 of pregnancy. Control rats received equivalent doses of corn oil alone. Selected pups and all dams were killed 1 mo after pups were weaned. A total of 50 male and 50 female offspring per group were followed until they were 2 yr old. The livers of offspring killed at the ages of 2 mo and 2 yr had PBB levels of 2,4 (SD 1.2) and 0.8 (SD 0.65) mg/kg for females and 3.0 (SD 1.6) and 0.6 (SD 0.37) mg/kg for males, respectively. The incidence of hepatocellular carcinomas was 3/51 (5.9%) and 4/41 (9.6%) after 2 yr in females and males, respectively. Hepatocellular carcinomas were not observed among the controls. Neoplastic (hyperplastic) nodules of the liver were present in 9/51 (17.6%) and 2/41 (4.9%) of exposed females and males, respectively, whereas only 2/48 (4.2%) of control females and no control males had neoplastic (hyperplastic) nodules. Body weights were lower in PBB-exposed rats at ages 1, 6, 12, and 24 mo. Survival rates from birth to weaning were lower in PBB-exposed pups (89%) than in controls (98%). Mortality was two times higher in PBB-exposed males (64%) than in control males (32%) after 2 yr. Transplacental PBB exposure and exposure through milk resulted in PBB body burdens in the offspring still measurable at the end of their lifespan. These offspring had increased mortality rates and lower body weights than controls, and they developed hepatocellular carcinomas.

  8. Increased interleukin-1β levels following low dose MDMA induces tolerance against the 5-HT neurotoxicity produced by challenge MDMA

    PubMed Central

    2011-01-01

    Background Preconditioning is a phenomenon by which tolerance develops to injury by previous exposure to a stressor of mild severity. Previous studies have shown that single or repeated low dose MDMA can attenuate 5-HT transporter loss produced by a subsequent neurotoxic dose of the drug. We have explored the mechanism of delayed preconditioning by low dose MDMA. Methods Male Dark Agouti rats were given low dose MDMA (3 mg/kg, i.p.) 96 h before receiving neurotoxic MDMA (12.5 mg/kg, i.p.). IL-1β and IL1ra levels and 5-HT transporter density in frontal cortex were quantified at 1 h, 3 h or 7 days. IL-1β, IL-1ra and IL-1RI were determined between 3 h and 96 h after low dose MDMA. sIL-1RI combined with low dose MDMA or IL-1β were given 96 h before neurotoxic MDMA and toxicity assessed 7 days later. Results Pretreatment with low dose MDMA attenuated both the 5-HT transporter loss and elevated IL-1β levels induced by neurotoxic MDMA while producing an increase in IL-1ra levels. Low dose MDMA produced an increase in IL-1β at 3 h and in IL-1ra at 96 h. sIL-1RI expression was also increased after low dose MDMA. Coadministration of sIL-1RI (3 μg, i.c.v.) prevented the protection against neurotoxic MDMA provided by low dose MDMA. Furthermore, IL-1β (2.5 pg, intracortical) given 96 h before neurotoxic MDMA protected against the 5-HT neurotoxicity produced by the drug, thus mimicking preconditioning. Conclusions These results suggest that IL-1β plays an important role in the development of delayed preconditioning by low dose MDMA. PMID:22114930

  9. Leucine induced dephosphorylation of Sestrin2 promotes mTORC1 activation.

    PubMed

    Kimball, Scot R; Gordon, Bradley S; Moyer, Jenna E; Dennis, Michael D; Jefferson, Leonard S

    2016-08-01

    The studies described herein were designed to explore the role of Sestrin2 in mediating the selective action of leucine to activate mTORC1. The results demonstrate that Sestrin2 is a phosphoprotein and that its phosphorylation state is responsive to the availability of leucine, but not other essential amino acids. Moreover, leucine availability-induced alterations in Sestrin2 phosphorylation correlated temporally and dose dependently with the activation state of mTORC1, there being a reciprocal relationship between the degree of phosphorylation of Sestrin2 and the extent of repression of mTORC1. With leucine deprivation, Sestrin2 became more highly phosphorylated and interacted more strongly with proteins of the GATOR2 complex. Notably, in cells lacking the protein kinase ULK1, the activation state of mTORC1 was elevated in leucine-deficient medium, such that the effect of re-addition of the amino acid was blunted. In contrast, overexpression of ULK1 led to hyperphosphorylation of Sestrin2 and enhanced its interaction with GATOR2. Neither rapamycin nor Torin2 had any effect on Sestrin2 phosphorylation, suggesting that leucine deprivation-induced repression of mTORC1 was not responsible for the action of ULK1 on Sestrin2. Mass spectrometry analysis of Sestrin2 revealed three phosphorylation sites that are conserved across mammalian species. Mutation of the three sites to phospho-mimetic amino acids in exogenously expressed Sestrin2 promoted its interaction with GATOR2 and dramatically repressed mTORC1 even in the presence of leucine. Overall, the results support a model in which leucine selectively promotes dephosphorylation of Sestrin2, causing it to dissociate from and thereby activate GATOR2, leading to activation of mTORC1.

  10. Purification and identification of a novel leucine aminopeptidase from Bacillus thuringiensis israelensis.

    PubMed

    Cahan, Rivka; Hetzroni, Efrat; Nisnevitch, Marina; Nitzan, Yeshayahu

    2007-11-01

    A novel leucine aminopeptidase was purified from a Bacillus thuringiensis israelensis (Bti) culture. The purification stages included heating the concentrated supernatant to 65 degrees C for 90 min, anion-exchange chromatography by DEAE cellulose, and hydrophobic chromatography by phenyl Sepharose. The specific activity of leucine aminopeptidase after the hydrophobic chromatography increased by 215.5-fold and the yield was 16%. The molecular weight of the active enzyme was 59 kDa. Mass spectrometry analysis of the 59-kDa leucine aminopeptidase revealed that this protein has at least 41% homology with the cytosol leucine aminopeptidase produced by Bacillus cereus. Maximal leucine aminopeptidase activity occurred at 65 degrees C, pH 10 toward leucine as the amino acid terminus. The enzyme was strongly inhibited by bestatin, dithiothreitol, and 1,10-phenanthroline, indicating that the enzyme might be considered as a metallo-aminopeptidase that has disulfide bonds at the catalytic site or at a region that influences its configuration. Examination of the purified leucine aminopeptidase's effect on the activation of the protoxin Cyt1Aa from Bti revealed that when it acts synergistically with Bti endogenous proteases, it has only a minor role in the processing of Cyt1Aa into an active toxin.

  11. Increased thermal pain sensitivity in animals exposed to chronic high dose Vicodin but not pure hydrocodone.

    PubMed

    O'Connell, Thomas F; Carpenter, Patrick S; Caballero, Nadia; Putnam, Andrew J; Steere, Joshua T; Matz, Gregory J; Foecking, Eileen M

    2014-01-01

    Vicodin, the combination drug of acetaminophen and the opioid hydrocodone, is one of the most prescribed drugs on the market today. Opioids have demonstrated the ability to paradoxically cause increased pain sensitivity to users in a phenomena called opioid-induced hyperalgesia (OIH). While selected opioids have been shown to produce OIH symptoms in an animal model, hydrocodone and the combination drug Vicodin have yet to be studied. The purpose of this study was to explore the effect of exposure to chronic high dose Vicodin or its components on the sensitivity to both thermal and mechanical pain. Animals were randomly divided into 4 groups, Vicodin, acetaminophen, hydrocodone, or vehicle control, and administered the drug daily for 120 days. Rats were subsequently tested for thermal and mechanical sensitivity. The rats in the Vicodin group displayed a significant decrease in withdrawal time to thermal pain. The rats receiving acetaminophen, hydrocodone, and vehicle showed no statistically significant hypersensitivity in thermal testing. None of the groups demonstrated statistically significant hypersensitivity to mechanical testing. The data suggests Vicodin produces signs of OIH in a rodent model. However, increased pain sensitivity was only noted in the thermal pathway and the hypersensitivity was only seen with the opioid combination drug, not the opioid alone. The results of this study both support the results of previous rodent opioid studies while generating further questions about the specific properties of Vicodin that contribute to pain hypersensitivity. The growing use of Vicodin to treat chronic pain necessitates further research looking into this paradoxical pain response.

  12. Beta-carotene conversion to vitamin A decreases as the dietary dose increases in humans

    Technology Transfer Automated Retrieval System (TEKTRAN)

    It has been suggested that high doses of B-carotene limit its conversion to vitamin A, yet this effect has not been well established in humans. A feeding study was conducted in which volunteers consumed two doses of deuterium labeled B-carotene on two occasions, with B-carotene and vitamin A respon...

  13. Do poor-responder patients benefit from increasing the daily gonadotropin dose during controlled ovarian hyperstimulation for IVF?

    PubMed

    Haas, Jigal; Zilberberg, Eran; Machtinger, Ronit; Kedem, Alon; Hourvitz, Ariel; Orvieto, Raoul

    2015-01-01

    We aim to assess the in vitro fertilization-embryo transfer (IVF-ET) outcome in patients receiving an extremely high 450 daily dose (IU) of gonadotropins during controlled ovarian hyperstimulation (COH) for IVF. Moreover, in those who failed to conceive while using 450 daily dose (IU) of gonadotropins, we aim to evaluate whether increasing the daily dose gonadotropins to 600 IU will improve IVF outcome. All consecutive women, admitted to our IVF unit and underwent COH consisting of daily gonadotropin dose of 450 IU were included. Ovarian stimulation characteristics, number of oocytes retrieved, number of embryo transferred and pregnancy rate were assessed. Nine-hundred one consecutive IVF cycles were evaluated. While there was no between-group difference in the duration of COH, patients who conceived were significantly younger, yielded higher number of oocytes retrieved and embryos transferred and had significantly lower cancellations. In a sub-analysis, including only those patients who failed to conceive while using 450 daily dose (IU) of gonadotropins, and who underwent a subsequent IVF cycle attempt with the used of 600 IU daily dose of gonadotropins, no improvements in COH characteristics or cancellation rates were observed with increasing the daily gonadotropin dose to 600 IU. To conclude, in poor responders undergoing COH with an extremely high daily gonadotropin dose (450 IU), the most important factors that predict IVF success are female age and the number of oocytes retrieved. Moreover, patients who failed to conceive on a daily gonadotropin dose of 450 IU will not benefit from increasing the dose to 600 IU and should therefore consider the options of egg donation or adoption.

  14. Enhanced Incorporation of 3-Hydroxy-4-Methylvalerate Unit into Biosynthetic Polyhydroxyalkanoate Using Leucine as a Precursor

    PubMed Central

    2011-01-01

    Ralstonia eutropha PHB-4 expressing Pseudomonas sp. 61-3 polyhydroxyalkanoate (PHA) synthase 1 (PhaC1Ps) synthesizes PHA copolymer containing 3-hydroxybutyrate (3HB) and a small amount (0.5 mol%) of 3-hydroxy-4-methylvalerate (3H4MV) from fructose as a carbon source. In this study, enhanced incorporation of 3H4MV into PHA was investigated using branched amino acid leucine as a precursor of 3H4MV. Leucine has the same carbon backbone as 3H4MV and is expected to be a natural and self-producible precursor. We found that the incorporation of 3H4MV was enhanced by the supplementation of excess amount (10 g/L) of leucine in the culture medium. This finding indicates that 3H4MV can be derived from leucine. To increase metabolic flux to leucine biosynthesis in the host strain by eliminating the feedback inhibition, the cells were subjected to N-methyl-N'-nitro-N-nitrosoguanidine (NTG) mutagenesis and leucine analog resistant mutants were generated. The mutants showed statistically higher 3H4MV fraction than the parent strain without supplementing leucine. Additionally, by supplying excess amount of leucine, the mutants synthesized 3HB-based PHA copolymer containing 3.1 mol% 3H4MV and 1.2 mol% 3-hydroxyvalerate (3HV) as minor constituents, which significantly affected the thermal properties of the copolymer. This study demonstrates that it is possible to enhance the monomer supply of 3H4MV into PHA by manipulating leucine metabolism. PMID:21906338

  15. Regulation of Leucine Catabolism in Pseudomonas putida

    PubMed Central

    Massey, Linda K.; Conrad, Robert S.; Sokatch, John R.

    1974-01-01

    The generation time of Pseudomonas putida with l-leucine was 20 h in synthetic media but only 3 h with d-leucine. Slow growth in the presence of l-leucine was partially overcome by addition of 0.1 mM amounts of either d-valine, l-valine, or 2-ketoisovalerate. The activities of five enzymes which take part in the oxidation of leucine by P. putida were measured under various conditions of growth. Four enzymes were induced by growth with dl-leucine as sole source of carbon: d-amino acid dehydrogenase, branched-chain keto acid dehydrogenase, 3-methylcrotonyl-coenzyme A carboxylase, and 3-hydroxy-3-methylglutaryl-coenzyme A lyase. The segment of the pathway required for oxidation of 3-methylcrotonate was induced by growth on isovalerate or 3-methylcrotonate without formation of the preceding enzymes. The synthesis of carboxylase and lyase appeared to have been repressed by the addition of l-glutamate or glucose to cells growing on dl-leucine as the sole carbon source. Mutants unable to grow at the expense of isovalerate had reduced levels of carboxylase and lyase, whereas the levels of three enzymes common to the catabolism of all three branched-chain amino acids and those of two isoleucine catabolic enzymes were normal. PMID:4150714

  16. Influence of increased target dose inhomogeneity on margins for breathing motion compensation in conformal stereotactic body radiotherapy

    PubMed Central

    Richter, Anne; Baier, Kurt; Meyer, Juergen; Wilbert, Juergen; Krieger, Thomas; Flentje, Michael; Guckenberger, Matthias

    2008-01-01

    Background Breathing motion should be considered for stereotactic body radiotherapy (SBRT) of lung tumors. Four-dimensional computer tomography (4D-CT) offers detailed information of tumor motion. The aim of this work is to evaluate the influence of inhomogeneous dose distributions in the presence of breathing induced target motion and to calculate margins for motion compensation. Methods Based on 4D-CT examinations, the probability density function of pulmonary tumors was generated for ten patients. The time-accumulated dose to the tumor was calculated using one-dimensional (1D) convolution simulations of a 'static' dose distribution and target probability density function (PDF). In analogy to stereotactic body radiotherapy (SBRT), different degrees of dose inhomogeneity were allowed in the target volume: minimum doses of 100% were prescribed to the edge of the target and maximum doses varied between 102% (P102) and 150% (P150). The dose loss due to breathing motion was quantified and margins were added until this loss was completely compensated. Results With the time-weighted mean tumor position as the isocentre, a close correlation with a quadratic relationship between the standard deviation of the PDF and the margin size was observed. Increased dose inhomogeneity in the target volume required smaller margins for motion compensation: margins of 2.5 mm, 2.4 mm and 1.3 mm were sufficient for compensation of 11.5 mm motion range and standard deviation of 3.9 mm in P105, P125 and P150, respectively. This effect of smaller margins for increased dose inhomogeneity was observed for all patients. Optimal sparing of the organ-at-risk surrounding the target was achieved for dose prescriptions P105 to P118. The internal target volume concept over-compensated breathing motion with higher than planned doses to the target and increased doses to the surrounding normal tissue. Conclusion Treatment planning with inhomogeneous dose distributions in the target volume required

  17. Fitness costs of increased cataract frequency and cumulative radiation dose in natural mammalian populations from Chernobyl

    PubMed Central

    Lehmann, Philipp; Boratyński, Zbyszek; Mappes, Tapio; Mousseau, Timothy A.; Møller, Anders P.

    2016-01-01

    A cataract is a clouding of the lens that reduces light transmission to the retina, and it decreases the visual acuity of the bearer. The prevalence of cataracts in natural populations of mammals, and their potential ecological significance, is poorly known. Cataracts have been reported to arise from high levels of oxidative stress and a major cause of oxidative stress is ionizing radiation. We investigated whether elevated frequencies of cataracts are found in eyes of bank voles Myodes glareolus collected from natural populations in areas with varying levels of background radiation in Chernobyl. We found high frequencies of cataracts in voles collected from different areas in Chernobyl. The frequency of cataracts was positively correlated with age, and in females also with the accumulated radiation dose. Furthermore, the number of offspring in female voles was negatively correlated with cataract severity. The results suggest that cataracts primarily develop as a function of ionizing background radiation, most likely as a plastic response to high levels of oxidative stress. It is therefore possible that the elevated levels of background radiation in Chernobyl affect the ecology and fitness of local mammals both directly through, for instance, reduced fertility and indirectly, through increased cataractogenesis. PMID:26814168

  18. Increased iron deposition in rat liver fibrosis induced by a high-dose injection of dimethylnitrosamine.

    PubMed

    Guo, Limei; Enzan, Hideaki; Hayashi, Yoshihiro; Miyazaki, Eriko; Jin, Yulan; Toi, Makoto; Kuroda, Naoto; Hiroi, Makoto

    2006-12-01

    Using a developed rat model of hepatic necrosis and subsequent fibrosis induced by a high-dose intraperitoneal injection of dimethylnitrosamine (DMN), we studied iron deposition and expression of transforming growth factor-beta(1) (TGF-beta(1)) during the development of persistent liver fibrosis. Rats were sacrificed at several timepoints from 6 h to 10 months post-injection and the livers were examined for iron content and distribution, and for expression of alpha-smooth muscle actin, ED-1, TGF-beta(1), and collagen (alpha(2))I. Morphologic evidence of acute submassive hemorrhagic necrosis peaked at 36 h; on day 3 the residual parenchyma contained activated hepatic stellate cells (HSCs) and necrotic areas contained numerous macrophages; and on day 5, necrotic tissues and erythrocytes had been phagocytosed and macrophages contained abundant iron deposits. From days 7 to 10, iron-laden macrophages and activated HSCs (myofibroblasts) populated the fibrous septa in parallel. From week 2 to month 10, closely arranged macrophages and myofibroblasts were found in central-to-central bridging fibrotic tissue. TGF-beta(1) was strongly detected in both macrophages and HSCs during development of liver fibrosis. Our data suggest that increased iron deposition may be involved in the initiation and perpetuation of rat liver fibrosis. Iron-laden macrophages may influence HSCs through the action of TGF-beta(1) in DMN-induced liver fibrosis.

  19. Fitness costs of increased cataract frequency and cumulative radiation dose in natural mammalian populations from Chernobyl.

    PubMed

    Lehmann, Philipp; Boratyński, Zbyszek; Mappes, Tapio; Mousseau, Timothy A; Møller, Anders P

    2016-01-27

    A cataract is a clouding of the lens that reduces light transmission to the retina, and it decreases the visual acuity of the bearer. The prevalence of cataracts in natural populations of mammals, and their potential ecological significance, is poorly known. Cataracts have been reported to arise from high levels of oxidative stress and a major cause of oxidative stress is ionizing radiation. We investigated whether elevated frequencies of cataracts are found in eyes of bank voles Myodes glareolus collected from natural populations in areas with varying levels of background radiation in Chernobyl. We found high frequencies of cataracts in voles collected from different areas in Chernobyl. The frequency of cataracts was positively correlated with age, and in females also with the accumulated radiation dose. Furthermore, the number of offspring in female voles was negatively correlated with cataract severity. The results suggest that cataracts primarily develop as a function of ionizing background radiation, most likely as a plastic response to high levels of oxidative stress. It is therefore possible that the elevated levels of background radiation in Chernobyl affect the ecology and fitness of local mammals both directly through, for instance, reduced fertility and indirectly, through increased cataractogenesis.

  20. Habitual Chocolate Consumption May Increase Body Weight in a Dose-Response Manner

    PubMed Central

    Greenberg, James A.; Buijsse, Brian

    2013-01-01

    Objective Habitual chocolate intake was recently found to be associated with lower body weight in three cross-sectional epidemiological studies. Our objective was to assess whether these cross-sectional results hold up in a more rigorous prospective analysis. Methods We used data from the Atherosclerosis Risk in Communities cohort. Usual dietary intake was assessed by questionnaire at baseline (1987–98), and after six years. Participants reported usual chocolate intake as the frequency of eating a 1-oz (∼28 g) serving. Body weight and height were measured at the two visits. Missing data were replaced by multiple imputation. Linear mixed-effects models were used to evaluate cross-sectional and prospective associations between chocolate intake and adiposity. Results Data were from 15,732 and 12,830 participants at the first and second visit, respectively. More frequent chocolate consumption was associated with a significantly greater prospective weight gain over time, in a dose-response manner. For instance, compared to participants who ate a chocolate serving less often than monthly, those who ate it 1–4 times a month and at least weekly experienced an increase in Body Mass Index (kg/m2) of 0.26 (95% CI 0.08, 0.44) and 0.39 (0.23, 0.55), respectively, during the six-year study period. In cross-sectional analyses the frequency of chocolate consumption was inversely associated with body weight. This inverse association was attenuated after excluding participants with preexisting obesity-related illness. Compared to participants without such illness, those with it had higher BMI and reported less frequent chocolate intake, lower caloric intake, and diets richer in fruits and vegetables. They tended to make these dietary changes after becoming ill. Conclusions Our prospective analysis found that a chocolate habit was associated with long-term weight gain, in a dose-response manner. Our cross-sectional finding that chocolate intake was associated with lower body

  1. Bisphosphonate treatment of postmenopausal osteoporosis is associated with a dose dependent increase in serum sclerostin.

    PubMed

    Gatti, Davide; Viapiana, Ombretta; Adami, Silvano; Idolazzi, Luca; Fracassi, Elena; Rossini, Maurizio

    2012-03-01

    The benefits coming from long-term treatment of postmenopausal osteoporosis with bisphophonates are limited by a coupled decrease in bone formation. The objective of this study is to determine whether this decrease in bone formation is associated with changes in serum levels of the WNT signaling antagonist sclerostin or Dickkopf-1 (DKK1). This is an ancillary observation from patients participating in a 12 months, phase 2, randomized clinical trial. We analyzed 107 patients given either monthly intramuscular neridronate (12.5, 25 or 50 mg) or placebo. Serum C-terminal telopeptide of type I collagen (sCTX, a bone-resorption marker) decreased by 61%, 75% and 73% in the 12.5, 25 and 50 mg dose groups, respectively. Mean changes in bone alkaline phosphatase (bAP) at 12 months were -47%, -60.0% and -52.6% in the groups receiving 12.5, 25 or 50 mg neridronate, respectively. Serum DKK1 remained unchanged at all time points in the 3 groups. Serum sclerostin increased versus placebo group gradually and significantly only in patients treated with 25 or 50 mg neridronate monthly, reaching 138-148% of baseline values (P<0.001). Changes in serum sclerostin at 12 months were negatively correlated with changes in bAP (P<0.001) even when data were adjusted for sCTX changes and only treated patients were included. In conclusions, decreased bone formation after several months of bisphosphonate therapy is associated with increased serum levels of sclerostin. This might suggest that Wnt signaling may play a role in the coupling between resorption and formation.

  2. Increased Mitochondrial DNA Copy Number in Occupations Associated with Low-Dose Benzene Exposure

    PubMed Central

    Pesatori, Angela Cecilia; Dioni, Laura; Hoxha, Mirjam; Bollati, Valentina; Albetti, Benedetta; Byun, Hyang-Min; Bonzini, Matteo; Fustinoni, Silvia; Cocco, Pierluigi; Satta, Giannina; Zucca, Mariagrazia; Merlo, Domenico Franco; Cipolla, Massimo; Bertazzi, Pier Alberto; Baccarelli, Andrea

    2011-01-01

    Background: Benzene is an established leukemogen at high exposure levels. Although low-level benzene exposure is widespread and may induce oxidative damage, no mechanistic biomarkers are available to detect biological dysfunction at low doses. Objectives: Our goals were to determine in a large multicenter cross-sectional study whether low-level benzene is associated with increased blood mitochondrial DNA copy number (mtDNAcn, a biological oxidative response to mitochondrial DNA damage and dysfunction) and to explore potential links between mtDNAcn and leukemia-related epigenetic markers. Methods: We measured blood relative mtDNAcn by real-time polymerase chain reaction in 341 individuals selected from various occupational groups with low-level benzene exposures (> 100 times lower than the Occupational Safety and Health Administration/European Union standards) and 178 referents from three Italian cities (Genoa, Milan, Cagliari). Results: In each city, benzene-exposed participants showed higher mtDNAcn than referents: mtDNAcn was 0.90 relative units in Genoa bus drivers and 0.75 in referents (p = 0.019); 0.90 in Milan gas station attendants, 1.10 in police officers, and 0.75 in referents (p-trend = 0.008); 1.63 in Cagliari petrochemical plant workers, 1.25 in referents close to the plant, and 0.90 in referents farther from the plant (p-trend = 0.046). Using covariate-adjusted regression models, we estimated that an interquartile range increase in personal airborne benzene was associated with percent increases in mtDNAcn equal to 10.5% in Genoa (p = 0.014), 8.2% (p = 0.008) in Milan, 7.5% in Cagliari (p = 0.22), and 10.3% in all cities combined (p < 0.001). Using methylation data available for the Milan participants, we found that mtDNAcn was associated with LINE-1 hypomethylation (–2.41%; p = 0.007) and p15 hypermethylation (+15.95%, p = 0.008). Conclusions: Blood MtDNAcn was increased in persons exposed to low benzene levels, potentially reflecting mitochondrial

  3. Effect of increased CRM₁₉₇ carrier protein dose on meningococcal C bactericidal antibody response.

    PubMed

    Lee, Lucia H; Blake, Milan S

    2012-04-01

    New multivalent CRM(197)-based conjugate vaccines are available for childhood immunization. Clinical studies were reviewed to assess meningococcal group C (MenC) antibody responses following MenC-CRM(197) coadministration with CRM(197)-based pneumococcal or Haemophilus influenzae type b conjugate vaccines. Infants receiving a total CRM(197) carrier protein dose of ∼50 μg and concomitant diphtheria-tetanus-acellular pertussis (DTaP)-containing vaccine tended to have lower MenC geometric mean antibody titers and continued to have low titers after the toddler dose. Nevertheless, at least 95% of children in the reported studies achieved a MenC serum bactericidal antibody (SBA) titer of ≥ 1:8 after the last infant or toddler dose. SBA was measured using an assay with a baby rabbit or human complement source. Additional studies are needed to assess long-term antibody persistence and MenC CRM(197) conjugate vaccine immunogenicity using alternative dosing schedules.

  4. Low-dose ionizing radiation induces mitochondrial fusion and increases expression of mitochondrial complexes I and III in hippocampal neurons

    PubMed Central

    Chang, Chuang-Rung; Kao, Mou-Chieh; Chen, Kuan-Wei; Chiu, Shih-Che; Hsu, Ming-Ling; Hsiang, I-Chou; Chen, Yu-Jen; Chen, Linyi

    2015-01-01

    High energy ionizing radiation can cause DNA damage and cell death. During clinical radiation therapy, the radiation dose could range from 15 to 60 Gy depending on targets. While 2 Gy radiation has been shown to cause cancer cell death, studies also suggest a protective potential by low dose radiation. In this study, we examined the effect of 0.2-2 Gy radiation on hippocampal neurons. Low dose 0.2 Gy radiation treatment increased the levels of MTT. Since hippocampal neurons are post-mitotic, this result reveals a possibility that 0.2 Gy irradiation may increase mitochondrial activity to cope with stimuli. Maintaining neural plasticity is an energy-demanding process that requires high efficient mitochondrial function. We thus hypothesized that low dose radiation may regulate mitochondrial dynamics and function to ensure survival of neurons. Our results showed that five days after 0.2 Gy irradiation, no obvious changes on neuronal survival, neuronal synapses, membrane potential of mitochondria, reactive oxygen species levels, and mitochondrial DNA copy numbers. Interestingly, 0.2 Gy irradiation promoted the mitochondria fusion, resulting in part from the increased level of a mitochondrial fusion protein, Mfn2, and inhibition of Drp1 fission protein trafficking to the mitochondria. Accompanying with the increased mitochondrial fusion, the expressions of complexes I and III of the electron transport chain were also increased. These findings suggest that, hippocampal neurons undergo increased mitochondrial fusion to modulate cellular activity as an adaptive mechanism in response to low dose radiation. PMID:26415228

  5. Low-dose ionizing radiation induces mitochondrial fusion and increases expression of mitochondrial complexes I and III in hippocampal neurons.

    PubMed

    Chien, Ling; Chen, Wun-Ke; Liu, Szu-Ting; Chang, Chuang-Rung; Kao, Mou-Chieh; Chen, Kuan-Wei; Chiu, Shih-Che; Hsu, Ming-Ling; Hsiang, I-Chou; Chen, Yu-Jen; Chen, Linyi

    2015-10-13

    High energy ionizing radiation can cause DNA damage and cell death. During clinical radiation therapy, the radiation dose could range from 15 to 60 Gy depending on targets. While 2 Gy radiation has been shown to cause cancer cell death, studies also suggest a protective potential by low dose radiation. In this study, we examined the effect of 0.2-2 Gy radiation on hippocampal neurons. Low dose 0.2 Gy radiation treatment increased the levels of MTT. Since hippocampal neurons are post-mitotic, this result reveals a possibility that 0.2 Gy irradiation may increase mitochondrial activity to cope with stimuli. Maintaining neural plasticity is an energy-demanding process that requires high efficient mitochondrial function. We thus hypothesized that low dose radiation may regulate mitochondrial dynamics and function to ensure survival of neurons. Our results showed that five days after 0.2 Gy irradiation, no obvious changes on neuronal survival, neuronal synapses, membrane potential of mitochondria, reactive oxygen species levels, and mitochondrial DNA copy numbers. Interestingly, 0.2 Gy irradiation promoted the mitochondria fusion, resulting in part from the increased level of a mitochondrial fusion protein, Mfn2, and inhibition of Drp1 fission protein trafficking to the mitochondria. Accompanying with the increased mitochondrial fusion, the expressions of complexes I and III of the electron transport chain were also increased. These findings suggest that, hippocampal neurons undergo increased mitochondrial fusion to modulate cellular activity as an adaptive mechanism in response to low dose radiation.

  6. Increasing the dose of television advertising in a national antismoking media campaign: results from a randomised field trial

    PubMed Central

    McAfee, Tim; Davis, Kevin C; Shafer, Paul; Patel, Deesha; Alexander, Robert; Bunnell, Rebecca

    2017-01-01

    Background While antismoking media campaigns have demonstrated effectiveness, less is known about the country-level effects of increased media dosing. The 2012 US Tips From Former Smokers (Tips) campaign generated approximately 1.6 million quit attempts overall; however, the specific dose–response from the campaign was only assessed by self-report. Objective Assess the impact of higher ad exposure during the 2013 Tips campaign on quit-related behaviours and intentions, campaign awareness, communication about campaign, and disease knowledge. Methods A 3-month national media buy was supplemented within 67 (of 190) randomly selected local media markets. Higher-dose markets received media buys 3 times that of standard-dose markets. We compared outcomes of interest using data collected via web-based surveys from nationally representative, address-based probability samples of 5733 cigarette smokers and 2843 non-smokers. Results In higher-dose markets, 87.2% of smokers and 83.9% of non-smokers recalled television campaign exposure versus 75.0% of smokers and 73.9% of non-smokers in standard-dose markets. Among smokers overall, the relative quit attempt rate was 11% higher in higher-dose markets (38.8% vs 34.9%; p<0.04). The higher-dose increase was larger in African-Americans (50.9% vs 31.8%; p<0.01). Smokers in higher-dose markets without a mental health condition, with a chronic health condition, or with only some college education made quit attempts at a higher rate than those in standard-dose markets. Non-smokers in higher-dose markets were more likely to talk with family or friends about smoking dangers (43.1% vs 35.7%; p<0.01) and had greater knowledge of smoking-related diseases. Conclusions The US 2013 Tips antismoking media campaign compared standard and higher doses by randomisation of local media markets. Results demonstrate the effectiveness of a higher dose for engaging non-smokers and further increasing quit attempts among smokers, especially African

  7. Dose- and time-dependent increase of lysosomal enzymes in embryonic cartilage in vitro after ionizing radiation

    SciTech Connect

    Cornelissen, M.; de Ridder, L. )

    1990-09-01

    Radiation doses of 20, 50 or 100 Gy caused the same time related decrease for RNA and proteoglycan (PG) synthesis in embryonic cartilage in vitro (4 days culture). In this paper, participation of lysosomes in this radiation response is investigated. Therefore, we employ a cytochemical method using beta-glycerophosphate as substrate for acid phosphatase (AP) detection. Increase of AP was found 2 days after irradiation and increased during the whole culture period. The increase was more pronounced with a higher radiation dose. Stimulation of AP activity explains the observed radiation response of RNA and PG synthesis.

  8. Platelet response to increased aspirin dose in patients with persistent platelet aggregation while treated with aspirin 81 mg.

    PubMed

    Gengo, Fran; Westphal, Erica S; Rainka, Michelle M; Janda, Maria; Robson, Matthew J; Hourihane, J Maurice; Bates, Vernice

    2016-04-01

    This study demonstrates that patients who are taking 81 mg of aspirin and are nonresponsive benefit from a dose of 162 mg or greater vs a different antiplatelet therapy. We identified 100 patients who were nonresponsive to aspirin 81 mg via whole blood aggregometry and observed how many patients became responsive at a dose of 162 mg or greater. Platelet nonresponsiveness was defined as >10 Ω of resistance to collagen 1 µg/mL and/or an ohms ratio of collagen 1 µg/mL to collagen 5 µg/mL >0.5 and/or >6 Ω to arachidonate. Borderline response was defined as an improvement in 1 but not both of the above criteria. Of the initial 100 patients who were nonresponsive to an aspirin dose of 81 mg, 79% became responsive at a dose of 162 mg or >162 mg. Only 6% did not respond to any increase in dose. We believe that patients treated with low-dose aspirin who have significant risk for secondary vascular events should be individually assessed to determine their antiplatelet response. Those found to have persistent platelet aggregation despite treatment with 81 mg of aspirin have a higher likelihood of obtaining an adequate antiplatelet response at a higher aspirin dose.

  9. Responses of astrocytes in culture after low dose laser irradiation

    SciTech Connect

    Yew, D.T.; Zheng, D.R.; Au, C.; Li, W.W. )

    1990-03-01

    The effect of Helium-Neon low dose laser on astrocytes was investigated in cultures of isolated astrocytes from albino neonatal rats. The laser appeared to inhibit the growth of astrocytes as exemplified by the smaller sizes of the cells and the decreased leucine uptake in each cell after treatment. Temporary decrease in the number of mitoses was also observed, but this trend was reversed soon after. Electron microscopic studies revealed an increase in buddings from cell bodies and processes (branches) after irradiation.

  10. Relationships between leucine and the pancreatic exocrine function for improving starch digestibility in ruminants.

    PubMed

    Liu, K; Liu, Y; Liu, S M; Xu, M; Yu, Z P; Wang, X; Cao, Y C; Yao, J H

    2015-04-01

    Four Holstein heifers (215 ± 7 kg; means ± SD), fitted with one pancreatic pouch, duodenal re-entrant cannulas, and duodenal infusion catheters, were used in this experiment. In phase 1, the 24-h profile of pancreatic fluid was determined. Pancreatic fluid flow peaked 1h after feeding, but peaks of similar magnitude also occurred before the morning feed, necessitating 24-h collection of pancreatic fluid to estimate daily excretion. In phase 2, the effects of duodenal infusions of 0, 10, 20, or 30 g of leucine on pancreatic fluid flow were determined in a 4 × 4 Latin square design. The leucine was infused for 12h in 2,500 mL of the infusate, and samples of pancreatic fluid and jugular blood were collected in 1-h intervals from the beginning of the infusion for 36 h. The results showed that the secretion rate of pancreatic fluid (mL/h) was significantly higher in 10-g leucine group than the other groups (mL/h). Protein concentration (mg/mL) in pancreatic fluid was elevated proportional to the amount of leucine infused. Leucine infusions increased both the concentration (U/mL) and secretion rate (U/h) of α-amylase. Infusion of 10 g of leucine also increased the secretion rates (U/h) of trypsin, chymotrypsin, and lipase, but did not change their concentrations. No significant effects of leucine infusions on plasma glucose and insulin concentrations were found. The results indicate that leucine could act as a nutrient signal to stimulate α-amylase production and pancreatic exocrine function in dairy heifers.

  11. Leucine Promotes Proliferation and Differentiation of Primary Preterm Rat Satellite Cells in Part through mTORC1 Signaling Pathway.

    PubMed

    Dai, Jie-Min; Yu, Mu-Xue; Shen, Zhen-Yu; Guo, Chu-Yi; Zhuang, Si-Qi; Qiu, Xiao-Shan

    2015-05-08

    Signaling through the mammalian target of rapamycin (mTOR) in response to leucine modulates many cellular and developmental processes. However, in the context of satellite cell proliferation and differentiation, the role of leucine and mTORC1 is less known. This study investigates the role of leucine in the process of proliferation and differentiation of primary preterm rat satellite cells, and the relationship with mammalian target of rapamycin complex 1 (mTORC1) activation. Dissociation of primary satellite cells occurred with type I collagenase and trypsin, and purification, via different speed adherence methods. Satellite cells with positive expression of Desmin were treated with leucine and rapamycin. We observed that leucine promoted proliferation and differentiation of primary satellite cells and increased the phosphorylation of mTOR. Rapamycin inhibited proliferation and differentiation, as well as decreased the phosphorylation level of mTOR. Furthermore, leucine increased the expression of MyoD and myogenin while the protein level of MyoD decreased due to rapamycin. However, myogenin expressed no affect by rapamycin. In conclusion, leucine may up-regulate the activation of mTORC1 to promote proliferation and differentiation of primary preterm rat satellite cells. We have shown that leucine promoted the differentiation of myotubes in part through the mTORC1-MyoD signal pathway.

  12. Oral Leucine Supplementation Is Sensed by the Brain but neither Reduces Food Intake nor Induces an Anorectic Pattern of Gene Expression in the Hypothalamus

    PubMed Central

    Zampieri, Thais T.; Pedroso, João A. B.; Furigo, Isadora C.; Tirapegui, Julio; Donato, Jose

    2013-01-01

    Leucine activates the intracellular mammalian target of the rapamycin (mTOR) pathway, and hypothalamic mTOR signaling regulates food intake. Although central infusion of leucine reduces food intake, it is still uncertain whether oral leucine supplementation is able to affect the hypothalamic circuits that control energy balance. We observed increased phosphorylation of p70s6k in the mouse hypothalamus after an acute oral gavage of leucine. We then assessed whether acute oral gavage of leucine induces the activation of neurons in several hypothalamic nuclei and in the brainstem. Leucine did not induce the expression of Fos in hypothalamic nuclei, but it increased the number of Fos-immunoreactive neurons in the area postrema. In addition, oral gavage of leucine acutely increased the 24 h food intake of mice. Nonetheless, chronic leucine supplementation in the drinking water did not change the food intake and the weight gain of ob/ob mice and of wild-type mice consuming a low- or a high-fat diet. We assessed the hypothalamic gene expression and observed that leucine supplementation increased the expression of enzymes (BCAT1, BCAT2 and BCKDK) that metabolize branched-chain amino acids. Despite these effects, leucine supplementation did not induce an anorectic pattern of gene expression in the hypothalamus. In conclusion, our data show that the brain is able to sense oral leucine intake. However, the food intake is not modified by chronic oral leucine supplementation. These results question the possible efficacy of leucine supplementation as an appetite suppressant to treat obesity. PMID:24349566

  13. Controlling Mechanical Properties of Bis-leucine Oxalyl Amide Gels

    NASA Astrophysics Data System (ADS)

    Chang, William; Carvajal, Daniel; Shull, Kenneth

    2011-03-01

    is-leucine oxalyl amide is a low molecular weight gelator capable of gelling polar and organic solvents. A fundamental understanding of self-assembled systems can lead to new methods in drug delivery and the design of new soft material systems. An important feature of self-assembled systems are the intermolecular forces between solvent and gelator molecule; by changing the environment the gel is in, the mechanical properties also change. In this project two variables were considered: the degree of neutralization present for the gelator molecule from neutral to completely ionized, and the concentration of the gelator molecule, from 1 weight percent to 8 weight percent in 1-butanol. Mechanical properties were studied using displacement controlled indentation techniques and temperature sweep rheometry. It has been found that properties such as the storage modulus, gelation temperature and maximum stress allowed increase with bis-leucine oxalyl amide concentration. The results from this study establish a 3-d contour map between the gelator concentration, the gelator degree of ionization and mechanical properties such as storage modulus and maximum stress allowed. The intermolecular forces between the bis-leucine low molecular weight gelator and 1-butanol govern the mechanical properties of the gel system, and understanding these interactions will be key to rationally designed self-assembled systems.

  14. Low-dose recombinant human growth hormone increases body weight and lean body mass in patients with short bowel syndrome.

    PubMed Central

    Ellegård, L; Bosaeus, I; Nordgren, S; Bengtsson, B A

    1997-01-01

    OBJECTIVE: The authors investigate the effects of low dose recombinant human growth hormone (rhGH) on body composition and absorptive capacity in patients with short bowel syndrome from Crohn's disease. SUMMARY BACKGROUND DATA: Patients with short bowel syndrome usually are malnourished because of malabsorption. The anabolic effects of high doses of rhGH have been tested in different clinical catabolic conditions, recently including patients with short bowel syndrome. The authors have investigated the effects of low-dose rhGH in short bowel syndrome in a placebo-controlled crossover clinical trial. METHODS: Ten patients were treated with daily subcutaneous doses of rhGH/placebo (0.5 international units/kg-1 per week-1 = 0.024 mg/kg-1 per day-1) for 8 weeks in a randomized, double-blind, placebo-controlled crossover clinical trial with a minimum of 12 weeks wash-out. Absorptive capacity and biochemical parameters were investigated in a metabolic ward before treatment and during first and last week of treatment. Body composition was determined by DEXA-Scan (Lunar DPX, Scanexport Medical, Helsingborg, Sweden), impedance analysis, and whole body potassium counting. RESULTS: Low-dose rhGH doubled serum levels of insulin-like growth factor-1 (IGF-1) and increased body weight, lean body mass, and total body potassium by 5% (p < 0.05). Fat-free mass and total body water increased by 6% (p = 0.008). Increases in IGF-1 levels correlated with increases in fat-free mass (r = 0.77, p < 0.02). No significant changes in absorptive capacity of water, energy, or protein were detected. CONCLUSION: Eight weeks of low-dose rhGH treatment leads to increases in body weight, lean body mass, and fat-free mass in patients with short bowel syndrome, correlated to increases in IGF-1 levels. PMID:8998124

  15. The time variation of dose rate artificially increased by the Fukushima nuclear crisis

    PubMed Central

    Hosoda, Masahiro; Tokonami, Shinji; Sorimachi, Atsuyuki; Monzen, Satoru; Osanai, Minoru; Yamada, Masatoshi; Kashiwakura, Ikuo; Akiba, Suminori

    2011-01-01

    A car-borne survey for dose rate in air was carried out in March and April 2011 along an expressway passing northwest of the Fukushima Dai-ichi Nuclear Power Station which released radionuclides starting after the Great East Japan Earthquake on March 11, 2011, and in an area closer to the Fukushima NPS which is known to have been strongly affected. Dose rates along the expressway, i.e. relatively far from the power station were higher after than before March 11, in some places by several orders of magnitude, implying that there were some additional releases from Fukushima NPS. The maximum dose rate in air within the high level contamination area was 36 μGy h−1, and the estimated maximum cumulative external dose for evacuees who came from Namie Town to evacuation sites (e.g. Fukushima, Koriyama and Nihonmatsu Cities) was 68 mSv. The evacuation is justified from the viewpoint of radiation protection. PMID:22355606

  16. Effect of Increased CRM197 Carrier Protein Dose on Meningococcal C Bactericidal Antibody Response

    PubMed Central

    Blake, Milan S.

    2012-01-01

    New multivalent CRM197-based conjugate vaccines are available for childhood immunization. Clinical studies were reviewed to assess meningococcal group C (MenC) antibody responses following MenC-CRM197 coadministration with CRM197-based pneumococcal or Haemophilus influenzae type b conjugate vaccines. Infants receiving a total CRM197 carrier protein dose of ∼50 μg and concomitant diphtheria-tetanus-acellular pertussis (DTaP)-containing vaccine tended to have lower MenC geometric mean antibody titers and continued to have low titers after the toddler dose. Nevertheless, at least 95% of children in the reported studies achieved a MenC serum bactericidal antibody (SBA) titer of ≥1:8 after the last infant or toddler dose. SBA was measured using an assay with a baby rabbit or human complement source. Additional studies are needed to assess long-term antibody persistence and MenC CRM197 conjugate vaccine immunogenicity using alternative dosing schedules. PMID:22336285

  17. Increased Subventricular Zone Radiation Dose Correlates With Survival in Glioblastoma Patients After Gross Total Resection

    SciTech Connect

    Chen, Linda; Guerrero-Cazares, Hugo; Ye, Xiaobu; Ford, Eric; McNutt, Todd; Kleinberg, Lawrence; Lim, Michael; Chaichana, Kaisorn; Quinones-Hinojosa, Alfredo; Redmond, Kristin

    2013-07-15

    Purpose: Neural progenitor cells in the subventricular zone (SVZ) have a controversial role in glioblastoma multiforme (GBM) as potential tumor-initiating cells. The purpose of this study was to examine the relationship between radiation dose to the SVZ and survival in GBM patients. Methods and Materials: The study included 116 patients with primary GBM treated at the Johns Hopkins Hospital between 2006 and 2009. All patients underwent surgical resection followed by adjuvant radiation therapy with intensity modulated radiation therapy (60 Gy/30 fractions) and concomitant temozolomide. Ipsilateral, contralateral, and bilateral SVZs were contoured on treatment plans by use of coregistered magnetic resonance imaging and computed tomography. Multivariate Cox regression was used to examine the relationship between mean SVZ dose and progression-free survival (PFS), as well as overall survival (OS). Age, Karnofsky Performance Status score, and extent of resection were used as covariates. The median age was 58 years (range, 29-80 years). Results: Of the patients, 12% underwent biopsy, 53% had subtotal resection (STR), and 35% had gross total resection (GTR). The Karnofsky Performance Status score was less than 90 in 54 patients and was 90 or greater in 62 patients. The median ipsilateral, contralateral, and bilateral mean SVZ doses were 48.7 Gy, 34.4 Gy, and 41.5 Gy, respectively. Among patients who underwent GTR, a mean ipsilateral SVZ dose of 40 Gy or greater was associated with a significantly improved PFS compared with patients who received less than 40 Gy (15.1 months vs 10.3 months; P=.028; hazard ratio, 0.385 [95% confidence interval, 0.165-0.901]) but not in patients undergoing STR or biopsy. The subgroup of GTR patients who received an ipsilateral dose of 40 Gy or greater also had a significantly improved OS (17.5 months vs 15.6 months; P=.027; hazard ratio, 0.385 [95% confidence interval, 0.165-0.895]). No association was found between SVZ radiation dose and PFS

  18. Isolation and characterization of awamori yeast mutants with L-leucine accumulation that overproduce isoamyl alcohol.

    PubMed

    Takagi, Hiroshi; Hashida, Keisuke; Watanabe, Daisuke; Nasuno, Ryo; Ohashi, Masataka; Iha, Tomoya; Nezuo, Maiko; Tsukahara, Masatoshi

    2015-02-01

    Awamori shochu is a traditional distilled alcoholic beverage made from steamed rice in Okinawa, Japan. Although it has a unique aroma that is distinguishable from that of other types of shochu, no studies have been reported on the breeding of awamori yeasts. In yeast, isoamyl alcohol (i-AmOH), known as the key flavor of bread, is mainly produced from α-ketoisocaproate in the pathway of L-leucine biosynthesis, which is regulated by end-product inhibition of α-isopropylmalate synthase (IPMS). Here, we isolated mutants resistant to the L-leucine analog 5,5,5-trifluoro-DL-leucine (TFL) derived from diploid awamori yeast of Saccharomyces cerevisiae. Some of the mutants accumulated a greater amount of intracellular L-leucine, and among them, one mutant overproduced i-AmOH in awamori brewing. This mutant carried an allele of the LEU4 gene encoding the Ser542Phe/Ala551Val variant IPMS, which is less sensitive to feedback inhibition by L-leucine. Interestingly, we found that either of the constituent mutations (LEU4(S542F) and LEU4(A551V)) resulted in the TFL tolerance of yeast cells and desensitization to L-leucine feedback inhibition of IPMS, leading to intracellular L-leucine accumulation. Homology modeling also suggested that L-leucine binding was drastically inhibited in the Ser542Phe, Ala551Val, and Ser542Phe/Ala551Val variants due to steric hindrance in the cavity of IPMS. As we expected, awamori yeast cells expressing LEU4(S542F), LEU4(A551V), and LEU4(S542F/A551V) showed a prominent increase in extracellular i-AmOH production, compared with that of cells carrying the vector only. The approach described here could be a practical method for the breeding of novel awamori yeasts to expand the diversity of awamori taste and flavor.

  19. Leucine alleviates dexamethasone-induced suppression of muscle protein synthesis via synergy involvement of mTOR and AMPK pathways

    PubMed Central

    Wang, Xiao J.; Yang, Xin; Wang, Ru X.; Jiao, Hong C.; Zhao, Jing P.; Song, Zhi G.; Lin, Hai

    2016-01-01

    Glucocorticoids (GCs) are negative muscle protein regulators that contribute to the whole-body catabolic state during stress. Mammalian target of rapamycin (mTOR)-signalling pathway, which acts as a central regulator of protein metabolism, can be activated by branched-chain amino acids (BCAA). In the present study, the effect of leucine on the suppression of protein synthesis induced by GCs and the pathway involved were investigated. In vitro experiments were conducted using cultured C2C12 myoblasts to study the effect of GCs on protein synthesis, and the involvement of mTOR pathway was investigated as well. After exposure to dexamethasone (DEX, 100 μmol/l) for 24 h, protein synthesis in muscle cells was significantly suppressed (P<0.05), the phosphorylations of mTOR, ribosomal protein S6 protein kinase 1 (p70s6k1) and eukaryotic initiation factor 4E binding protein 1 (4EBP1) were significantly reduced (P<0.05). Leucine supplementation (5 mmol/l, 10 mmol/l and 15 mmol/l) for 1 h alleviated the suppression of protein synthesis induced by DEX (P<0.05) and was accompanied with the increased phosphorylation of mTOR and decreased phosphorylation of AMPK (P<0.05). Branched-chain amino transferase 2 (BCAT2) mRNA level was not influenced by DEX (P>0.05) but was increased by leucine supplementation at a dose of 5 mmol/l (P<0.05). PMID:27129299

  20. Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway

    DOE PAGES

    Saxton, Robert A.; Knockenhauer, Kevin E.; Wolfson, Rachel L.; ...

    2015-11-19

    Eukaryotic cells coordinate growth with the availability of nutrients through mTOR complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag GTPases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. We present the 2.7-Å crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucinemore » leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. Lastly, these results provide a structural mechanism of amino acid sensing by the mTORC1 pathway.« less

  1. Directed evolution of leucine dehydrogenase for improved efficiency of L-tert-leucine synthesis.

    PubMed

    Zhu, Lin; Wu, Zhe; Jin, Jian-Ming; Tang, Shuang-Yan

    2016-07-01

    L-tert-Leucine and its derivatives are used as synthetic building blocks for pharmaceutical active ingredients, chiral auxiliaries, and ligands. Leucine dehydrogenase (LeuDH) is frequently used to prepare L-tert-leucine from the α-keto acid precursor trimethylpyruvate (TMP). In this study, a high-throughput screening method for the L-tert-leucine synthesis reaction based on a spectrophotometric approach was developed. Directed evolution strategy was applied to engineer LeuDH from Lysinibacillus sphaericus for improved efficiency of L-tert-leucine synthesis. After two rounds of random mutagenesis, the specific activity of LeuDH on the substrate TMP was enhanced by more than two-fold, compared with that of the wild-type enzyme, while the activity towards its natural substrate, leucine, decreased. The catalytic efficiencies (k cat/K m) of the best mutant enzyme, H6, on substrates TMP and NADH were all enhanced by more than five-fold as compared with that of the wild-type enzyme. The efficiency of L-tert-leucine synthesis by mutant H6 was significantly improved. A productivity of 1170 g/l/day was achieved for the mutant enzyme H6, compared with 666 g/l/day for the wild-type enzyme.

  2. The behaviour of leucine aminopeptidase towards thionopeptides.

    PubMed Central

    Beattie, R E; Elmore, D T; Williams, C H; Guthrie, D J

    1987-01-01

    Thionoleucine S-anilide (Leut-anilide), Leut-Gly-OEt and Leut-Phe-OMe were synthesized and shown to be competitive inhibitors of leucine aminopeptidase from pig kidney. The kinetics of inhibition were determined in the presence of leucine 4-methylcoumarin-7-amide as substrate. Although the compounds showed only moderate inhibitory potency, it was found that all were resistant to hydrolysis by the enzyme, in contrast with the reported behaviour of some thionopeptide analogues of substrates for other Zn2+-peptidases such as carboxypeptidase A and angiotensin-converting enzyme. PMID:3663153

  3. Effect of exercise training on leucine oxidation

    SciTech Connect

    Hendrix, M.K.; Layman, D.K.

    1986-03-01

    Oxidation of the BCAA leucine is increased during a bout of exhaustive exercise. The purpose of this study was to determine the effects of exercise training on leu oxidation during aerobic exercise. Female Sprague-Dawley rats were fed a commercial diet ad lib and divided into sedentary and two trained groups. Animals were trained to run on a treadmill with a 10/sup 0/ incline at 28 m/min for 5 wks for either 50 or 120 min/day. There were no differences in food intake or body weight. After a 12 hr fast, animals were run for 50 or 120 min and changes in leu catabolism determined by measurement of in vivo leu oxidation and activity of branched chain keto acid dehydrogenase (BCKAD). For measurement of leu oxidation, rats were injected IP with 4 ..mu..Ci 1-/sup 14/C-leu during the last 15 min of exercise, placed in glass metabolic chambers, and /sup 14/CO/sub 2/ collected in 1 N NaOH for 30 min periods. Leu oxidation was increased by 40% after 50 min of exercise and by 79% after 120 min of exercise. Five weeks of training reduced the rate of leu oxidation during an exercise bout. The activity of the BCKAD was not increased in the trained animals after either 50 or 120 min of exercise. These data indicate that the rate of leu oxidation during exercises is dependent on the duration of the exercise and that training will reduce the magnitude of this effect.

  4. Increasing dose gradient and uniformity in small fields using modulation: Theory and prototypes for cone-based stereotactic radiosurgery

    SciTech Connect

    Bender, Edward T.

    2014-05-15

    Purpose: To investigate the theoretical limits to the tradeoff between dose gradient and uniformity when modulation is used in the context of cone based SRS, and to design a prototype collimation device that allows for steeper dose gradients and/or higher target uniformity as compared to a standard circular collimator. Methods: An inverse planning optimization is performed in the context of idealized phantom geometry to determine the ideal fluence pattern that best approximates a “rect function” dose distribution. Ideal fluence patterns were approximated in a prototype device and radiochromic film dosimetry was utilized to compare the prototype device to a standard circular collimator. Results: For choices of prescription isodose lines above approximately 50%, utilizing modulation allows for an improved tradeoff between dose gradient index and dose heterogeneity index. Compensators placed within the circular collimator can achieve the necessary modulation. Conclusions: Using modulation with features on a submillimeter distance scale, it is possible to increase the dose gradient and/or uniformity in small fields.

  5. Double-blind, Randomized Trial of Alternative Letrozole Dosing Regimens in Postmenopausal Women with Increased Breast Cancer Risk

    PubMed Central

    López, Ana Maria; Pruthi, Sandhya; Boughey, Judy C.; Perloff, Marjorie; Hsu, Chiu-Hsieh; Lang, Julie E.; Ley, Michele; Frank, Denise; Taverna, Josephine A.; Chow, H-H Sherry

    2015-01-01

    Aromatase inhibitors (AIs) profoundly suppress estrogen levels in postmenopausal women and are effective in breast cancer prevention among high-risk postmenopausal women. Unfortunately, AI treatment is associated with undesirable side effects that limit patient acceptance for primary prevention of breast cancer. A double-blind, randomized trial was conducted to determine whether low and intermittent doses of letrozole can achieve effective estrogen suppression with a more favorable side effect profile. Overall, 112 postmenopausal women at increased risk for breast cancer were randomized to receive letrozole at 2.5 mg once daily (QD, standard dose arm), 2.5 mg every Monday, Wednesday, and Friday (Q-MWF), 1.0 mg Q-MWF or 0.25 mg Q-MWF for 24 weeks. Primary endpoint was suppression in serum estradiol levels at the end of letrozole intervention. Secondary endpoints included changes in serum estrone, testosterone, C-telopeptide (marker of bone resorption), lipid profile and quality of life measures (QoL) following treatment. Significant estrogen suppression was observed in all dose arms with an average of 75 – 78% and 86 – 93% reduction in serum estradiol and estrone levels, respectively. There were no differences among dose arms with respect to changes in C-telopeptide levels, lipid profile, adverse events (AEs) or QoL measures. We conclude that low and intermittent doses of letrozole are not inferior to standard dose in estrogen suppression and resulted in a similar side effect profile compared to standard dose. Further studies are needed to determine the feasibility of selecting an effective AI dosing schedule with better tolerability. PMID:26667449

  6. Dose escalation in brachytherapy for cervical cancer: impact on (or increased need for) MRI-guided plan optimisation

    PubMed Central

    Paton, A M; Chalmers, K E; Coomber, H; Cameron, A L

    2012-01-01

    Objective The aim of this study was to assess the impact of dose escalation on the proportion of patients requiring MR image-guided optimisation rather than standard Manchester-based CT-guided planning, and the level of escalation achievable. Methods 30 patients with cervical cancer treated with external beam radiotherapy and image-guided brachytherapy (IGBT) had MR images acquired at the first fraction of IGBT. Gross tumour volume and high-risk clinical target volume (HR CTV) were contoured and treatment plans retrospectively produced for a range of total 2-Gy equivalent (EQD2) prescription doses from 66 Gyα/β=10 to 90 Gyα/β=10 (HR CTV D90). Standard Manchester system-style plans were produced, prescribed to point A and then optimised where necessary with the aim of delivering at least the prescription dose to the HR CTV D90 while respecting organ-at-risk (OAR) tolerances. Results Increasing the total EQD2 from 66 Gyα/β=10 to 90 Gyα/β=10 increased the number of plans requiring optimisation from 13.3% to 90%. After optimisation, the number of plans achieving the prescription dose ranged from 93.3% (66 Gyα/β=10) to 63.3% (90 Gyα/β=10) with the mean±standard deviation for HR CTV D90 EQD2 from 78.4±12.4 Gyα/β=10 (66 Gyα/β=10) to 94.1±19.9 Gyα/β=10 (90 Gyα/β=10). Conclusion As doses are escalated, the need for non-standard optimised planning increases, while benefits in terms of increased target doses actually achieved diminish. The maximum achievable target dose is ultimately limited by proximity of OARs. Advances in knowledge This work represents a guide for other centres in determining the highest practicable prescription doses while considering patient throughput and maintaining acceptable OAR doses. PMID:23175490

  7. Carcinogenically relevant split dose repair increased with age in rat skin model.

    NASA Astrophysics Data System (ADS)

    Burns, Fredric; Tang, Moon-Shong Eric; Wu, Feng; Uddin, Ahmed

    2012-07-01

    These experiments utilize cancer induction to evaluate cancer-relevant repair during the interval between dose fractions. Low LET electron radiation(LET ~ 0.34 keV/u) were utilized in experiments that involved exposing rat dorsal skin to 2 equal 8 Gy dose fractions separated at various intervals from 0.25 h to 24 h. Cancer onset was established for 80 weeks after the exposures and only histologically verified cancers were included in the analysis. This experiment involved a total of 540 rats and 880 induced cancers. In the youngest rats (irradiated at 28 days of age) the cancer yield declined with a halftime of approximately 3.5 hrs. In 113 day old rats the cancer yield halftime was shortened to 1.3 hrs. In the oldest rats (182 days of age), the halftime could not be established quantitatively, because it was less than the shortest interval (15 min) utilized in the protocol (best estimate ~5 min). In the oldest rats the cancer yields for all fractionated exposures dropped essentially to the expected level of 2 single fractions, below which theoretically no further reduction is possible. The follow-up times for obtaining cancer yields were the same for all exposure groups in spite of the differing ages at exposure. These results indicate that repair of carcinogenically-relevant damage accelerates with age of the rat. No information is available on the possible mechanistic basis for this finding, although the model might be useful for delineating which of the many postulated split dose repair pathways is the correct one. The finding indicates that older rats should be less susceptible to the carcinogenic action of single doses of low LET radiation in comparison to younger rats, which has been verified in separate studies.

  8. Effect of increasing oral doses of loperamide on gallbladder motility in man.

    PubMed Central

    Hopman, W P; Rosenbusch, G; Jansen, J B; Lamers, C B

    1990-01-01

    1. Loperamide, a peripherally acting opiate receptor agonist with antidiarrhoeal action, inhibits ileal and colonic motor function. To determine the effect of loperamide on gallbladder motility, we have pretreated five healthy volunteers with 2 mg oral loperamide 24 h, 20, 12 and 2.5 h before; six healthy volunteers with 16 mg oral loperamide 2.5 h before; and eight healthy volunteers with 16 mg oral loperamide 12 and 2.5 h before intravenous infusion of a 'physiological dose' of 12.5 pmol kg-1 cholecystokinin (CCK) for 1 h to stimulate gallbladder contraction. All subjects served as their own controls. Gallbladder volume was measured by ultrasonography and plasma CCK by radioimmunoassay until 90 min after start of the CCK infusion. 2. Infusion of CCK resulted in plasma CCK concentrations similar to those after intraduodenal fat. Integrated gallbladder contraction after 4 X 2 mg loperamide (4600 +/- 891% min) was similar to that without pretreatment (5270 +/- 1037% min; NS). Integrated gallbladder contraction after 1 X 16 mg loperamide diminished from 5458 +/- 412% min without to 2632 +/- 816% min with loperamide (P less than 0.05), and was completely abolished to -596 +/- 762% min (P less than 0.0005 vs without loperamide) after 2 X 16 mg loperamide. 3. It is concluded that loperamide inhibits gallbladder contraction in response to a physiological dose of cholecystokinin in a dose-dependent manner. PMID:2297461

  9. Life-Span Exposure to Low Doses of Aspartame Beginning during Prenatal Life Increases Cancer Effects in Rats

    PubMed Central

    Soffritti, Morando; Belpoggi, Fiorella; Tibaldi, Eva; Esposti, Davide Degli; Lauriola, Michelina

    2007-01-01

    Background In a previous study conducted at the Cesare Maltoni Cancer Research Center of the European Ramazzini Foundation (CMCRC/ERF), we demonstrated for the first time that aspartame (APM) is a multipotent carcinogenic agent when various doses are administered with feed to Sprague-Dawley rats from 8 weeks of age throughout the life span. Objective The aim of this second study is to better quantify the carcinogenic risk of APM, beginning treatment during fetal life. Methods We studied groups of 70–95 male and female Sprague-Dawley rats administered APM (2,000, 400, or 0 ppm) with feed from the 12th day of fetal life until natural death. Results Our results show a) a significant dose-related increase of malignant tumor–bearing animals in males (p < 0.01), particularly in the group treated with 2,000 ppm APM (p < 0.01); b) a significant increase in incidence of lymphomas/leukemias in males treated with 2,000 ppm (p < 0.05) and a significant dose-related increase in incidence of lymphomas/leukemias in females (p < 0.01), particularly in the 2,000-ppm group (p < 0.01); and c) a significant dose-related increase in incidence of mammary cancer in females (p < 0.05), particularly in the 2,000-ppm group (p < 0.05). Conclusions The results of this carcinogenicity bioassay confirm and reinforce the first experimental demonstration of APM’s multipotential carcinogenicity at a dose level close to the acceptable daily intake for humans. Furthermore, the study demonstrates that when life-span exposure to APM begins during fetal life, its carcinogenic effects are increased. PMID:17805418

  10. 21 CFR 582.5406 - Leucine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Leucine. 582.5406 Section 582.5406 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  11. 21 CFR 582.5406 - Leucine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Leucine. 582.5406 Section 582.5406 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  12. 21 CFR 582.5406 - Leucine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Leucine. 582.5406 Section 582.5406 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  13. 21 CFR 582.5406 - Leucine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Leucine. 582.5406 Section 582.5406 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  14. 21 CFR 582.5406 - Leucine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Leucine. 582.5406 Section 582.5406 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  15. Amino acid availability and age affect the leucine stimulation of protein synthesis and eIF4F formation in muscle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have previously shown that a physiological increase in plasma leucine for 60 and 120 min increases translation initiation factor activation in muscle of neonatal pigs. Although muscle protein synthesis is increased by leucine at 60 min, it is not maintained at 120 min, perhaps because of the decr...

  16. Doses and risks from uranium are not increased significantly by interactions with natural background photon radiation.

    PubMed

    Tanner, R J; Eakins, J S; Jansen, J T M; Harrison, J D

    2012-08-01

    The impact of depleted uranium (DU) on human health has been the subject of much conjecture. Both the chemical and radiological aspects of its behaviour in the human body have previously been investigated in detail, with the radiological impact being assumed to be linked to the alpha decay of uranium. More recently, it has been proposed that the accumulation in tissue of high-Z materials, such as DU, may give rise to enhanced local energy deposition in the presence of natural background photon radiation due to the high photoelectric interaction cross sections of high-Z atoms. It is speculated that, in addition to producing short-range photoelectrons, these events will be followed by intense Auger and Coster-Kronig electron emission, thereby causing levels of cell damage that are unaccounted for in conventional models of radiological risk. In this study, the physical and biological bases of these claims are investigated. The potential magnitudes of any effect are evaluated and discussed, and compared with the risks from other radiological or chemical hazards. Monte Carlo calculations are performed to estimate likely energy depositions due to the presence of uranium in human tissues in photon fields: whole body doses, organ doses in anthropomorphic phantoms and nano-/micro-dosimetric scenarios are each considered. The proposal is shown generally to be based on sound physics, but overall the impact on human health is expected to be negligible.

  17. Rapamycin-mediated lifespan increase in mice is dose and sex dependent and metabolically distinct from dietary restriction

    PubMed Central

    Miller, Richard A; Harrison, David E; Astle, Clinton M; Fernandez, Elizabeth; Flurkey, Kevin; Han, Melissa; Javors, Martin A; Li, Xinna; Nadon, Nancy L; Nelson, James F; Pletcher, Scott; Salmon, Adam B; Sharp, Zelton Dave; Van Roekel, Sabrina; Winkleman, Lynn; Strong, Randy

    2014-01-01

    Rapamycin, an inhibitor of mTOR kinase, increased median lifespan of genetically heterogeneous mice by 23% (males) to 26% (females) when tested at a dose threefold higher than that used in our previous studies; maximal longevity was also increased in both sexes. Rapamycin increased lifespan more in females than in males at each dose evaluated, perhaps reflecting sexual dimorphism in blood levels of this drug. Some of the endocrine and metabolic changes seen in diet-restricted mice are not seen in mice exposed to rapamycin, and the pattern of expression of hepatic genes involved in xenobiotic metabolism is also quite distinct in rapamycin-treated and diet-restricted mice, suggesting that these two interventions for extending mouse lifespan differ in many respects. PMID:24341993

  18. Gestational and lactational exposure to low-dose bisphenol A increases Th17 cells in mice offspring.

    PubMed

    Luo, Shimeng; Li, Yun; Li, Yingpei; Zhu, Qixing; Jiang, Jianhua; Wu, Changhao; Shen, Tong

    2016-10-01

    Increasing evidence demonstrates that perinatal exposure to Bisphenol A (BPA) can cause immune disorders throughout the life span. However, the biological basis for these immune disorders is poorly understood and the effects of exposure to BPA on Th17 development are unknown. The present study sought to characterize alterations of Th17 cells in childhood and adulthood following gestational and lactational exposure to environmentally relevant low-dose of BPA and the underlying mechanisms. Pregnant dams were exposed to BPA (10, 100 or 1000nM) via drinking water from gestational day (GD) 0 to postnatal day (PND) 21. At PNDs 21 and 42, offspring mice were anesthetized, blood was obtained for cytokine assay and spleens were collected for Th17 cell frequency and RORγt mRNA expression analysis. Perinatal exposure to low-dose BPA resulted in a dose-dependent and gender-specific persistent rise in Th17 cells accompanied by an increase of RORγt mRNA expression in the offsprings. The contents of major Th17 cell-derived cytokines (IL-17 and IL-21) and those essential for Th17 cell differentiation (IL-6 and IL-23) were also increased compared to those in controls. These changes were more pronounced in female than in male offsprings. However, perinatal exposure to low-dose BPA had little effect on serum TGF-β, another key regulator for Th17 cell development. Our results suggest that gestational and lactational exposure to a low-dose of BPA can affect Th17 cell development via an action on its transcription factor and the regulatory cytokines. These findings provide novel insight into sustained immune disorders by BPA exposure during development.

  19. METABOLIC RESPONSES TO DIETARY LEUCINE RESTRICTION INVOLVE REMODELING OF ADIPOSE TISSUE AND ENHANCED HEPATIC INSULIN SIGNALING

    PubMed Central

    Wanders, Desiree; Stone, Kirsten P.; Dille, Kelly; Simon, Jacob; Pierse, Alicia; Gettys, Thomas W.

    2015-01-01

    Dietary leucine was incrementally restricted to test whether limiting this essential amino acid (EAA) would fully reproduce the beneficial responses produced by dietary methionine restriction. Restricting leucine by 85% increased energy intake and expenditure within five to seven days of its introduction and reduced overall accumulation of adipose tissue. Leucine restriction (LR) also improved glucose tolerance, increased hepatic release of FGF21 into the blood stream, and enhanced insulin-dependent activation of Akt in liver. However, LR had no effect on hepatic lipid levels and failed to lower lipogenic gene expression in the liver. LR did affect remodeling of white and brown adipose tissue, increasing expression of both thermogenic and lipogenic genes. These findings illustrate that dietary LR reproduces many but not all of the physiological responses of methionine restriction. The primary differences occur in the liver, where methionine and leucine restriction cause opposite effects on tissue lipid levels and expression of lipogenic genes. Together these findings suggest that the sensing systems which detect and respond to dietary restriction of EAAs act through mechanisms that both leucine and methionine are able to engage, and in the case of hepatic lipid metabolism, may be unique to specific EAAs such as methionine. PMID:26643647

  20. Low-dose exposure to di-(2-ethylhexyl) phthalate (DEHP) increases susceptibility to testicular autoimmunity in mice.

    PubMed

    Hirai, Shuichi; Naito, Munekazu; Kuramasu, Miyuki; Ogawa, Yuki; Terayama, Hayato; Qu, Ning; Hatayama, Naoyuki; Hayashi, Shogo; Itoh, Masahiro

    2015-09-01

    Exposure to di-(2-ethylhexyl) phthalate (DEHP) induces spermatogenic disturbance (SD) through oxidative stress, and affects the immune system by acting as an adjuvant. Recently, we reported that in mice, a low dose of DEHP, which did not affect spermatogenesis, was able to alter the testicular immune microenvironment. Experimental autoimmune orchitis (EAO) can be induced by repeated immunization with testicular antigens, and its pathology is characterized by production of autoantibodies and SD. In the present study, we investigated the effect of a low-dose DEHP on the susceptibility of mice to EAO. The exposure to DEHP-containing feed (0.01%) caused a modest functional damage to the blood-testis barrier (BTB) with an increase in testicular number of interferon gamma (IFN-γ)-positive cells and resulted in the production of autoantibodies targeting haploid cells, but did not affect spermatogenesis. While only single immunization with testicular antigens caused very mild EAO, the concurrent DEHP exposure induced severe EAO with significant increases in number of interferon gamma-positive cells and macrophages, as well as lymphocytic infiltration and serum autoantibody titer accompanied by severe SD. To summarize, the exposure of mice to the low-dose DEHP does not induce significant SD, but it may cause an increase in IFN-γ positive cells and modest functional damage to the BTB in the testis. These changes lead to an autoimmune response against haploid cell autoantigens, resulting in increased susceptibility to EAO.

  1. Morphological templating of metastable calcium carbonates by the amino acid leucine

    NASA Astrophysics Data System (ADS)

    Thompson, S. P.; Parker, J. E.; Street, S. R.; Tang, C. C.

    2011-03-01

    The in vitro precipitation of the metastable CaCO3 phases aragonite and vaterite in the presence of leucine is investigated. Under normal conditions, the production of CaCO3 via the hydrolysis of urea method favours the formation of regular needle-like aragonite crystals, with very minor quantities of vaterite and calcite. However in the presence of leucine, aragonite forms highly branched structures and the vaterite yield is increased, forming flower-like clusters composed of nano-thin sheets. Both the degree of aragonite branching and the occurrence, regularity of shape and number of vaterite "petals" increases with leucine concentration. The two phases exhibit different variations in their crystallographic parameters with increasing concentration, while the molecular structure appears unaffected.

  2. Protein Ingestion Induces Muscle Insulin Resistance Independent of Leucine-Mediated mTOR Activation.

    PubMed

    Smith, Gordon I; Yoshino, Jun; Stromsdorfer, Kelly L; Klein, Seth J; Magkos, Faidon; Reeds, Dominic N; Klein, Samuel; Mittendorfer, Bettina

    2015-05-01

    Increased plasma branched-chain amino acid concentrations are associated with insulin resistance, and intravenous amino acid infusion blunts insulin-mediated glucose disposal. We tested the hypothesis that protein ingestion impairs insulin-mediated glucose disposal by leucine-mediated mTOR signaling, which can inhibit AKT. We measured glucose disposal and muscle p-mTOR(Ser2448), p-AKT(Ser473), and p-AKT(Thr308) in 22 women during a hyperinsulinemic-euglycemic clamp procedure with and without concomitant ingestion of whey protein (0.6 g/kg fat-free mass; n = 11) or leucine that matched the amount given with whey protein (n = 11). Both whey protein and leucine ingestion raised plasma leucine concentration by approximately twofold and muscle p-mTOR(Ser2448) by ∼30% above the values observed in the control (no amino acid ingestion) studies; p-AKT(Ser473) and p-AKT(Thr308) were not affected by whey protein or leucine ingestion. Whey protein ingestion decreased insulin-mediated glucose disposal (median 38.8 [quartiles 30.8, 61.8] vs. 51.9 [41.0, 77.3] µmol glucose/µU insulin · mL(-1) · min(-1); P < 0.01), whereas ingestion of leucine did not (52.3 [43.3, 65.4] vs. 52.3 [43.9, 73.2]). These results indicate that 1) protein ingestion causes insulin resistance and could be an important regulator of postprandial glucose homeostasis and 2) the insulin-desensitizing effect of protein ingestion is not due to inhibition of AKT by leucine-mediated mTOR signaling.

  3. Protein Ingestion Induces Muscle Insulin Resistance Independent of Leucine-Mediated mTOR Activation

    PubMed Central

    Smith, Gordon I.; Yoshino, Jun; Stromsdorfer, Kelly L.; Klein, Seth J.; Magkos, Faidon; Reeds, Dominic N.; Klein, Samuel

    2015-01-01

    Increased plasma branched-chain amino acid concentrations are associated with insulin resistance, and intravenous amino acid infusion blunts insulin-mediated glucose disposal. We tested the hypothesis that protein ingestion impairs insulin-mediated glucose disposal by leucine-mediated mTOR signaling, which can inhibit AKT. We measured glucose disposal and muscle p-mTORSer2448, p-AKTSer473, and p-AKTThr308 in 22 women during a hyperinsulinemic-euglycemic clamp procedure with and without concomitant ingestion of whey protein (0.6 g/kg fat-free mass; n = 11) or leucine that matched the amount given with whey protein (n = 11). Both whey protein and leucine ingestion raised plasma leucine concentration by approximately twofold and muscle p-mTORSer2448 by ∼30% above the values observed in the control (no amino acid ingestion) studies; p-AKTSer473 and p-AKTThr308 were not affected by whey protein or leucine ingestion. Whey protein ingestion decreased insulin-mediated glucose disposal (median 38.8 [quartiles 30.8, 61.8] vs. 51.9 [41.0, 77.3] µmol glucose/µU insulin · mL−1 · min−1; P < 0.01), whereas ingestion of leucine did not (52.3 [43.3, 65.4] vs. 52.3 [43.9, 73.2]). These results indicate that 1) protein ingestion causes insulin resistance and could be an important regulator of postprandial glucose homeostasis and 2) the insulin-desensitizing effect of protein ingestion is not due to inhibition of AKT by leucine-mediated mTOR signaling. PMID:25475435

  4. Fractionated low doses of abdominal irradiation alters jejunal uptake of nutrients

    SciTech Connect

    Thomson, A.B.; Keelan, M.; Cheeseman, C.I.; Walker, K.

    1986-06-01

    Abdominal radiation is associated with changes in intestinal uptake of nutrients that begins within three days and persist for over 33 weeks. Clinically, fractionated doses of radiation (FDR) are used in an attempt to minimize the complications of this therapy, but the effects of fractionated doses of radiation on intestinal transport have not been defined. An in vitro technique was used to assess the jejunal and ileal uptake of varying concentrations of glucose and leucine, as well as the uptake of single concentrations of fatty acids and decanol in rats exposed 3, 7, and 14 days previously to a course of 200 cGy given on each of five consecutive days. FDR was associated with an increase in the uptake of decanol, and therefore a decrease in the effective resistance of the unstirred water layer. FDR had a variable effect on the uptake of glucose and leucine, with a decline in the value of the Michaelis constant (Km) and the passive permeability coefficient for glucose (Pd), whereas the Km for leucine was unchanged and the Pd for leucine was variably affected by FDR. The maximal transport rate (Jdm) for leucine progressively rose following FDR, whereas the Jdm for glucose initially rose, then fell. The uptake of galactose and medium chain-length fatty acids was unchanged by FDR, whereas the jejunal uptake of myristic acid rose, and the uptake of cholic acid declined, then returned to normal. FDR was associated with greater body weight gain and jejunal and ileal weight. The changes in nutrient uptake following FDR differed from the absorption changes occurring after a single dose of radiation. Thus, fractionated doses of abdominal radiation produce complex changes in the intestinal uptake of actively and passively transported nutrients, and these variable changes are influenced by the time following radiation exposure and by the solute studied.

  5. Experimental studies on the impact of an increased dose of anionic salts on the metabolism of dairy cows.

    PubMed

    Gelfert, C C; Passfeld, M; Löptien, A; Montag, N; Baumgartner, W; Staufenbiel, R

    2006-12-01

    This study was initiated to investigate the influence of a daily dose of anionic salts (AS) above the valid upper limit at present on metabolism of dairy cows. Eleven non-pregnant and non-lactating Holstein-Friesian crossbred cows with a permanent rumen cannula were used in a study with a controlled feeding design. The initial daily dose was 2500 meq/day, which resulted in a Dietary Cation Anion Difference (DCAD) of -211 meq/kg dry matter. Every seven days,the daily dose was raised by 500 meq. If a cow stopped eating, the application of AS was stopped and these cows were monitored over the next seven days. On day 30 another batch of hay, having the same DCAD but higher concentrations of minerals and energy, was fed. Blood and urine samples were taken to monitor acid-base balance and calcium concentrations. Acid-base balance was strongly influenced by AS. Blood pH dropped steadily and reached values around 7.23. Urine pH dropped quickly below 6 and remained at that level regardless of the increased dosage of AS. Net acid base excretion (NABE) fell continuously with the increase of the dosage of AS and reached values below -200 mmol/l. Calcium concentrations in the serum were nearly stable, but those in urine increased sharply and remained on an elevated level with increasing doses of AS. A few days before the individual cow's refusal of feed intake, calcium excretion in urine decreased. The majority of cows stopped eating while consuming a diet containing 3500 to 4000 meq AS except two animals who consumed up to 6000 meq/day AS but they received the better hay in the second half of the treatment period. In this time pH in blood increased slowly. NABE remained stable on a low level at -100 mmol/l. The results showed that with an increasing amount of AS fed the risk of clinical acidosis increased. The decreasing urine concentrations of calcium indicate a breakdown of the compensation capability of the single cow. Besides the dose of AS fed, the quality of the feed

  6. Protective effects of leucine against lipopolysaccharide-induced inflammatory response in Labeo rohita fingerlings.

    PubMed

    Giri, Sib Sankar; Sen, Shib Sankar; Jun, Jin Woo; Sukumaran, Venkatachalam; Park, Se Chang

    2016-05-01

    The present study investigated the protective effects of leucine against lipopolysaccharide (LPS)-induced inflammatory responses in Labeo rohita (rohu) in vivo and in vitro. Primary hepatocytes, isolated from the hepatopancreas, were exposed to different concentrations of LPS for 24 h to induce an inflammatory response, and the protective effects of leucine against LPS-induced inflammation were studied. Finally, we investigated the efficiency of dietary leucine supplementation in attenuating an immune challenge induced by LPS in vivo. Exposure of cells to 10-25 μg mL(-1) of LPS for 24 h resulted in a significant production of nitric oxide and release of lactate dehydrogenase to the medium, whereas cell viability and protein content were reduced (p < 0.05). LPS exposure (10 μg mL(-1)) increased mRNA levels of the pro-inflammatory cytokines TNF-α, IL-1β and IL-8 in vitro (p < 0.05). However, pretreatment with leucine prevented the LPS-induced upregulation of TNF-α, IL-1β and IL-8 mRNAs by downregulating TLR4, MyD88, NF-κBp65, and MAPKp38 mRNA expression. Interestingly, mRNA expression of the anti-inflammatory cytokine, IL-10, which was increased by LPS treatment, was further enhanced (p < 0.05) by leucine pretreatment. The enhanced expression of IL-10 might inhibit the production of other pro-inflammatory cytokines. It was found that leucine pretreatment attenuated the excessive activation of LPS-induced TLR4-MyD88 signaling as manifested by lower level of TLR4, MyD88, MAPKp38, NF-κBp65 and increased level of IκB-α protein in leucine pre-treatment group. In vivo experiments demonstrated that leucine pre-supplementation could protect fish against LPS-induced inflammation through an attenuation of TLR4-MyD88 signaling pathway. Taken together, we propose that leucine pre-supplementation decreases LPS-induced immune damage in rohu by enhancing the expression of IL-10 and by regulating the TLR4-MyD88 signaling pathways.

  7. Chronic leucine exposure results in reduced but reversible glucose-stimulated insulin secretion in INS-1 cells.

    PubMed

    Zhang, Xiujuan; Han, Wenxia; Jiang, Xiuyun; Li, Min; Gao, Ling; Zhao, Jia Jun

    2014-06-01

    Previous studies have demonstrated that sustained high leucine exposure decreases glucose-stimulated insulin secretion (GSIS). However, whether this effect is recoverable following the removal of leucine is unclear. Pancreatic/duodenal homeobox-1 (PDX-1) and its downstream target, glucose transporter 2 (GLUT2), are reported to be positively associated with insulin secretion. However, it also remains unclear whether the effect of leucine on GSIS is accompanied by alterations in PDX-1 and GLUT2. In the present study, insulin secretion, insulin content, PDX-1 and GLUT2 protein expression in INS-1 (rat insulinoma cell line) cells were assessed following a 24-h incubation in 40 mmol/l leucine. Half of the cells were incubated in leucine-free media for a further 24 h to observe the abovementioned effects. In contrast to the control, 40 mmol/l leucine for 24 or 48 h diminished GSIS at high glucose concentrations by 11% (P=0.026) or 22% (P=0.003), insulin content by 14% (P=0.008) or 20% (P=0.002), as well as decreasing PDX-1 and GLUT2 expression. When leucine was removed from the media for a further 24-h incubation, in comparison with those cells that were maintained in leucine treatment for 24 and 48 h, the high GSIS increased by 13% (P=0.032) and 27% (P=0.002), insulin content was augmented by 10% (P=0.014) and 20% (P=0.003), and the protein expression of PDX-1 and GLUT2 also increased. The present study demonstrates that sustained high concentrations of leucine induce a reversible impairment of GSIS and alter insulin content, which is mediated by PDX-1 and GLUT2, in INS-1 cells.

  8. Leptin and leucine synergistically regulate protein metabolism in C2C12 myotubes and mouse skeletal muscles.

    PubMed

    Mao, Xiangbing; Zeng, Xiangfang; Huang, Zhimin; Wang, Junjun; Qiao, Shiyan

    2013-07-28

    Leucine and leptin play important roles in regulating protein synthesis and degradation in skeletal muscles in vitro and in vivo. However, the objective of the present study was to determine whether leptin and leucine function synergistically in regulating protein metabolism of skeletal muscles. In the in vitro experiment, C2C12 myotubes were cultured for 2 h in the presence of 5 mm-leucine and/or 50 ng/ml of leptin. In the in vivo experiment, C57BL/6 and ob/ob mice were randomly assigned to be fed a non-purified diet supplemented with 3 % L-leucine or 2·04 % L-alanine (isonitrogenous control) for 14 d. Ob/ob mice were injected intraperitoneally with sterile PBS or recombinant mouse leptin (0·1 μg/g body weight) for 14 d. In C57BL/6 mice, dietary leucine supplementation increased (P< 0·05) plasma leptin, leptin receptor expression and protein synthesis in skeletal muscles, but reduced (P< 0·05) plasma urea and protein degradation in skeletal muscles. Dietary leucine supplementation and leptin injection increased the relative weight of the gastrocnemius and soleus muscles in ob/ob mice. Moreover, leucine and leptin treatments stimulated (P< 0·05) protein synthesis and inhibited (P< 0·05) protein degradation in C2C12 myotubes and skeletal muscles of ob/ob mice. There were interactions (P< 0·05) between the leucine and leptin treatments with regard to protein metabolism in C2C12 myotubes and soleus muscles of ob/ob mice but not in the gastrocnemius muscles of ob/ob mice. Collectively, these results suggest that leptin and leucine synergistically regulate protein metabolism in skeletal muscles both in vitro and in vivo.

  9. Leucine stimulates PPARβ/δ-dependent mitochondrial biogenesis and oxidative metabolism with enhanced GLUT4 content and glucose uptake in myotubes.

    PubMed

    Schnuck, Jamie K; Sunderland, Kyle L; Gannon, Nicholas P; Kuennen, Matthew R; Vaughan, Roger A

    2016-01-01

    Leucine stimulates anabolic and catabolic processes in skeletal muscle, however little is known about the effects of leucine on peroxisome proliferator-activated receptor (PPAR) activity. This work characterized the effects of 24-h leucine treatment on metabolic parameters and protein expression in cultured myotubes. Leucine significantly increased PPARβ/δ expression as well as markers of mitochondrial biogenesis, leading to significantly increased mitochondrial content and oxidative metabolism in a PPARβ/δ-dependent manner. However, leucine-treated cells did not display significant alterations in uncoupling protein expression or oxygen consumed per relative mitochondrial content suggesting leucine-mediated increases in oxidative metabolism are a function of increased mitochondrial content and not altered mitochondrial efficiency. Leucine treatment also increased GLUT4 content and glucose uptake as well as PPARγ and FAS expression leading to increased total lipid content. Leucine appears to activate PPAR activity leading to increased mitochondrial biogenesis and elevated substrate oxidation, while simultaneously promoting substrate/lipid storage and protein synthesis.

  10. Novel dosing strategies increase exposures of the potent antituberculosis drug rifapentine but are poorly tolerated in healthy volunteers.

    PubMed

    Dooley, Kelly E; Savic, Radojka M; Park, Jeong-Gun; Cramer, Yoninah; Hafner, Richard; Hogg, Evelyn; Janik, Jennifer; Marzinke, Mark A; Patterson, Kristine; Benson, Constance A; Hovind, Laura; Dorman, Susan E; Haas, David W

    2015-01-01

    Rifapentine is a potent antituberculosis drug currently in phase III trials. Bioavailability decreases with increasing dose, yet high daily exposures are likely needed to improve efficacy and shorten the tuberculosis treatment duration. Further, the limits of tolerability are poorly defined. The phase I multicenter trial in healthy adults described here investigated two strategies to increase rifapentine exposures: dividing the dose or giving the drug with a high-fat meal. In arm 1, rifapentine was administered at 10 mg/kg of body weight twice daily and 20 mg/kg once daily, each for 14 days, separated by a 28-day washout; the dosing sequence was randomized. In arm 2, 15 mg/kg rifapentine once daily was given with a high-fat versus a low-fat breakfast. Sampling for pharmacokinetic analysis was performed on days 1 and 14. Population pharmacokinetic analyses were performed. This trial was stopped early for poor tolerability and because of safety concerns. Of 44 subjects, 20 discontinued prematurely; 11 of these discontinued for protocol-defined toxicity (a grade 3 or higher adverse event or grade 2 or higher rifamycin hypersensitivity). Taking rifapentine with a high-fat meal increased the median steady-state area under the concentration-time curve from time zero to 24 h (AUC0-24ss) by 31% (relative standard error, 6%) compared to that obtained when the drug was taken with a low-fat breakfast. Dividing the dose increased exposures substantially (e.g., 38% with 1,500 mg/day). AUC0-24ss was uniformly higher in our study than in recent tuberculosis treatment trials, in which toxicity was rare. In conclusion, two strategies to increase rifapentine exposures, dividing the dose or giving it with a high-fat breakfast, successfully increased exposures, but toxicity was common in healthy adults. The limits of tolerability in patients with tuberculosis remain to be defined. (AIDS Clinical Trials Group study A5311 has been registered at ClinicalTrials.gov under registration no

  11. Novel Dosing Strategies Increase Exposures of the Potent Antituberculosis Drug Rifapentine but Are Poorly Tolerated in Healthy Volunteers

    PubMed Central

    Savic, Radojka M.; Park, Jeong-Gun; Cramer, Yoninah; Hafner, Richard; Hogg, Evelyn; Janik, Jennifer; Marzinke, Mark A.; Patterson, Kristine; Benson, Constance A.; Hovind, Laura; Dorman, Susan E.; Haas, David W.

    2015-01-01

    Rifapentine is a potent antituberculosis drug currently in phase III trials. Bioavailability decreases with increasing dose, yet high daily exposures are likely needed to improve efficacy and shorten the tuberculosis treatment duration. Further, the limits of tolerability are poorly defined. The phase I multicenter trial in healthy adults described here investigated two strategies to increase rifapentine exposures: dividing the dose or giving the drug with a high-fat meal. In arm 1, rifapentine was administered at 10 mg/kg of body weight twice daily and 20 mg/kg once daily, each for 14 days, separated by a 28-day washout; the dosing sequence was randomized. In arm 2, 15 mg/kg rifapentine once daily was given with a high-fat versus a low-fat breakfast. Sampling for pharmacokinetic analysis was performed on days 1 and 14. Population pharmacokinetic analyses were performed. This trial was stopped early for poor tolerability and because of safety concerns. Of 44 subjects, 20 discontinued prematurely; 11 of these discontinued for protocol-defined toxicity (a grade 3 or higher adverse event or grade 2 or higher rifamycin hypersensitivity). Taking rifapentine with a high-fat meal increased the median steady-state area under the concentration-time curve from time zero to 24 h (AUC0–24ss) by 31% (relative standard error, 6%) compared to that obtained when the drug was taken with a low-fat breakfast. Dividing the dose increased exposures substantially (e.g., 38% with 1,500 mg/day). AUC0–24ss was uniformly higher in our study than in recent tuberculosis treatment trials, in which toxicity was rare. In conclusion, two strategies to increase rifapentine exposures, dividing the dose or giving it with a high-fat breakfast, successfully increased exposures, but toxicity was common in healthy adults. The limits of tolerability in patients with tuberculosis remain to be defined. (AIDS Clinical Trials Group study A5311 has been registered at ClinicalTrials.gov under registration

  12. Low Z target switching to increase tumor endothelial cell dose enhancement during gold nanoparticle-aided radiation therapy

    SciTech Connect

    Berbeco, Ross I. Detappe, Alexandre; Tsiamas, Panogiotis; Parsons, David; Yewondwossen, Mammo; Robar, James

    2016-01-15

    Purpose: Previous studies have introduced gold nanoparticles as vascular-disrupting agents during radiation therapy. Crucial to this concept is the low energy photon content of the therapy radiation beam. The authors introduce a new mode of delivery including a linear accelerator target that can toggle between low Z and high Z targets during beam delivery. In this study, the authors examine the potential increase in tumor blood vessel endothelial cell radiation dose enhancement with the low Z target. Methods: The authors use Monte Carlo methods to simulate delivery of three different clinical photon beams: (1) a 6 MV standard (Cu/W) beam, (2) a 6 MV flattening filter free (Cu/W), and (3) a 6 MV (carbon) beam. The photon energy spectra for each scenario are generated for depths in tissue-equivalent material: 2, 10, and 20 cm. The endothelial dose enhancement for each target and depth is calculated using a previously published analytic method. Results: It is found that the carbon target increases the proportion of low energy (<150 keV) photons at 10 cm depth to 28% from 8% for the 6 MV standard (Cu/W) beam. This nearly quadrupling of the low energy photon content incident on a gold nanoparticle results in 7.7 times the endothelial dose enhancement as a 6 MV standard (Cu/W) beam at this depth. Increased surface dose from the low Z target can be mitigated by well-spaced beam arrangements. Conclusions: By using the fast-switching target, one can modulate the photon beam during delivery, producing a customized photon energy spectrum for each specific situation.

  13. Low Z target switching to increase tumor endothelial cell dose enhancement during gold nanoparticle-aided radiation therapy

    PubMed Central

    Berbeco, Ross I.; Detappe, Alexandre; Tsiamas, Panogiotis; Parsons, David; Yewondwossen, Mammo; Robar, James

    2016-01-01

    Purpose: Previous studies have introduced gold nanoparticles as vascular-disrupting agents during radiation therapy. Crucial to this concept is the low energy photon content of the therapy radiation beam. The authors introduce a new mode of delivery including a linear accelerator target that can toggle between low Z and high Z targets during beam delivery. In this study, the authors examine the potential increase in tumor blood vessel endothelial cell radiation dose enhancement with the low Z target. Methods: The authors use Monte Carlo methods to simulate delivery of three different clinical photon beams: (1) a 6 MV standard (Cu/W) beam, (2) a 6 MV flattening filter free (Cu/W), and (3) a 6 MV (carbon) beam. The photon energy spectra for each scenario are generated for depths in tissue-equivalent material: 2, 10, and 20 cm. The endothelial dose enhancement for each target and depth is calculated using a previously published analytic method. Results: It is found that the carbon target increases the proportion of low energy (<150 keV) photons at 10 cm depth to 28% from 8% for the 6 MV standard (Cu/W) beam. This nearly quadrupling of the low energy photon content incident on a gold nanoparticle results in 7.7 times the endothelial dose enhancement as a 6 MV standard (Cu/W) beam at this depth. Increased surface dose from the low Z target can be mitigated by well-spaced beam arrangements. Conclusions: By using the fast-switching target, one can modulate the photon beam during delivery, producing a customized photon energy spectrum for each specific situation. PMID:26745936

  14. Dose Relations between Goal Setting, Theory-Based Correlates of Goal Setting and Increases in Physical Activity during a Workplace Trial

    ERIC Educational Resources Information Center

    Dishman, Rod K.; Vandenberg, Robert J.; Motl, Robert W.; Wilson, Mark G.; DeJoy, David M.

    2010-01-01

    The effectiveness of an intervention depends on its dose and on moderators of dose, which usually are not studied. The purpose of the study is to determine whether goal setting and theory-based moderators of goal setting had dose relations with increases in goal-related physical activity during a successful workplace intervention. A…

  15. Age Attenuates Leucine Oxidation after Eccentric Exercise

    PubMed Central

    Kullman, E. L.; Campbell, W. W.; Krishnan, R. K.; Yarasheski, K. E.; Evans, W. J.; Kirwan, J. P.

    2013-01-01

    Aging may alter protein metabolism during periods of metabolic and physiologic challenge. The purpose of this study was to assess the effects of age on whole-body amino acid turnover in response to eccentric exercise and hyperglycemia-induced hyperinsulinemia. 16 healthy men were divided into young (N = 8) and older (N = 8) groups. Protein metabolism was assessed using a [1-13C]-leucine isotopic tracer approach. Measures were obtained under fasted basal conditions and during 3-h hyperglycemic clamps that were performed without (control) and 48 h after eccentric exercise. Exercise reduced leucine oxidation in the younger men (P < 0.05), but not in older men. Insulin sensitivity was inversely correlated with leucine oxidation (P < 0.05), and was lower in older men (P < 0.05). Healthy aging is associated with an impaired capacity to adjust protein oxidation in response to eccentric exercise. The decreased efficiency of protein utilization in older men may contribute to impaired maintenance, growth, and repair of body tissues with advancing age. PMID:23325713

  16. High-Dose Testosterone Treatment Increases Serotonin Transporter Binding in Transgender People

    PubMed Central

    Kranz, Georg S.; Wadsak, Wolfgang; Kaufmann, Ulrike; Savli, Markus; Baldinger, Pia; Gryglewski, Gregor; Haeusler, Daniela; Spies, Marie; Mitterhauser, Markus; Kasper, Siegfried; Lanzenberger, Rupert

    2015-01-01

    Background Women are two times more likely to be diagnosed with depression than men. Sex hormones modulating serotonergic transmission are proposed to partly underlie these epidemiologic findings. Here, we used the cross-sex steroid hormone treatment of transsexuals seeking sex reassignment as a model to investigate acute and chronic effects of testosterone and estradiol on serotonin reuptake transporter (SERT) binding in female-to-male and male-to-female transsexuals. Methods Thirty-three transsexuals underwent [11C]DASB positron emission tomography before start of treatment, a subset of which underwent a second scan 4 weeks and a third scan 4 months after treatment start. SERT nondisplaceable binding potential was quantified in 12 regions of interest. Treatment effects were analyzed using linear mixed models. Changes of hormone plasma levels were correlated with changes in regional SERT nondisplaceable binding potential. Results One and 4 months of androgen treatment in female-to-male transsexuals increased SERT binding in amygdala, caudate, putamen, and median raphe nucleus. SERT binding increases correlated with treatment-induced increases in testosterone levels, suggesting that testosterone increases SERT expression on the cell surface. Conversely, 4 months of antiandrogen and estrogen treatment in male-to-female transsexuals led to decreases in SERT binding in insula, anterior, and mid-cingulate cortex. Increases in estradiol levels correlated negatively with decreases in regional SERT binding, indicating a protective effect of estradiol against SERT loss. Conclusions Given the central role of the SERT in the treatment of depression and anxiety disorders, these findings may lead to new treatment modalities and expand our understanding of the mechanism of action of antidepressant treatment properties. PMID:25497691

  17. Functional profiling discovers the dieldrin organochlorinated pesticide affects leucine availability in yeast.

    PubMed

    Gaytán, Brandon D; Loguinov, Alex V; Lantz, Stephen R; Lerot, Jan-Michael; Denslow, Nancy D; Vulpe, Chris D

    2013-04-01

    Exposure to organochlorinated pesticides such as dieldrin has been linked to Parkinson's and Alzheimer's diseases, endocrine disruption, and cancer, but the cellular and molecular mechanisms of toxicity behind these effects remain largely unknown. Here we demonstrate, using a functional genomics approach in the model eukaryote Saccharomyces cerevisiae, that dieldrin alters leucine availability. This model is supported by multiple lines of congruent evidence: (1) mutants defective in amino acid signaling or transport are sensitive to dieldrin, which is reversed by the addition of exogenous leucine; (2) dieldrin sensitivity of wild-type or mutant strains is dependent upon leucine concentration in the media; (3) overexpression of proteins that increase intracellular leucine confer resistance to dieldrin; (4) leucine uptake is inhibited in the presence of dieldrin; and (5) dieldrin induces the amino acid starvation response. Additionally, we demonstrate that appropriate negative regulation of the Ras/protein kinase A pathway, along with an intact pyruvate dehydrogenase complex, is required for dieldrin tolerance. Many yeast genes described in this study have human orthologs that may modulate dieldrin toxicity in humans.

  18. Hypertrophy-Promoting Effects of Leucine Supplementation and Moderate Intensity Aerobic Exercise in Pre-Senescent Mice.

    PubMed

    Xia, Zhi; Cholewa, Jason; Zhao, Yan; Yang, Yue-Qin; Shang, Hua-Yu; Guimarães-Ferreira, Lucas; Naimo, Marshall Alan; Su, Quan-Sheng; Zanchi, Nelo Eidy

    2016-05-02

    Several studies have indicated a positive influence of leucine supplementation and aerobic training on the aging skeletal muscle signaling pathways that control muscle protein balance and muscle remodeling. However, the effect of a combined intervention requires further clarification. Thirteen month old CD-1(®) mice were subjected to moderate aerobic exercise (45 min swimming per day with 3% body weight workload) and fed a chow diet with 5% leucine or 3.4% alanine for 8 weeks. Serum and plasma were prepared for glucose, urea nitrogen, insulin and amino acid profile analysis. The white gastrocnemius muscles were used for determination of muscle size and signaling proteins involved in protein synthesis and degradation. The results show that both 8 weeks of leucine supplementation and aerobic training elevated the activity of mTOR (mammalian target of rapamycin) and its downstream target p70S6K and 4E-BP1, inhibited the ubiquitin-proteasome system, and increased fiber cross-sectional area (CSA) in white gastrocnemius muscle. Moreover, leucine supplementation in combination with exercise demonstrated more significant effects, such as greater CSA, protein content and altered phosphorylation (suggestive of increased activity) of protein synthesis signaling proteins, in addition to lower expression of proteins involved in protein degradation compared to leucine or exercise alone. The current study shows moderate aerobic training combined with 5% leucine supplementation has the potential to increase muscle size in fast-twitch skeletal muscle during aging, potentially through increased protein synthesis and decreased protein breakdown.

  19. Hypertrophy-Promoting Effects of Leucine Supplementation and Moderate Intensity Aerobic Exercise in Pre-Senescent Mice

    PubMed Central

    Xia, Zhi; Cholewa, Jason; Zhao, Yan; Yang, Yue-Qin; Shang, Hua-Yu; Guimarães-Ferreira, Lucas; Naimo, Marshall Alan; Su, Quan-Sheng; Zanchi, Nelo Eidy

    2016-01-01

    Several studies have indicated a positive influence of leucine supplementation and aerobic training on the aging skeletal muscle signaling pathways that control muscle protein balance and muscle remodeling. However, the effect of a combined intervention requires further clarification. Thirteen month old CD-1® mice were subjected to moderate aerobic exercise (45 min swimming per day with 3% body weight workload) and fed a chow diet with 5% leucine or 3.4% alanine for 8 weeks. Serum and plasma were prepared for glucose, urea nitrogen, insulin and amino acid profile analysis. The white gastrocnemius muscles were used for determination of muscle size and signaling proteins involved in protein synthesis and degradation. The results show that both 8 weeks of leucine supplementation and aerobic training elevated the activity of mTOR (mammalian target of rapamycin) and its downstream target p70S6K and 4E-BP1, inhibited the ubiquitin-proteasome system, and increased fiber cross-sectional area (CSA) in white gastrocnemius muscle. Moreover, leucine supplementation in combination with exercise demonstrated more significant effects, such as greater CSA, protein content and altered phosphorylation (suggestive of increased activity) of protein synthesis signaling proteins, in addition to lower expression of proteins involved in protein degradation compared to leucine or exercise alone. The current study shows moderate aerobic training combined with 5% leucine supplementation has the potential to increase muscle size in fast-twitch skeletal muscle during aging, potentially through increased protein synthesis and decreased protein breakdown. PMID:27144582

  20. Incorporation of fucose and leucine into PNS myelin proteins in nerves undergoing early Wallerian degeneration

    SciTech Connect

    Peterson, R.G.; Baughman, S.; Scheidler, D.M.

    1981-02-01

    The simultaneous incorporation of (/sup 3/H)fucose and (1-/sup 14/C)leucine into normal rat sciatic nerve was examined using an in vitro incubation model. A linear rate of protein precursor uptake was found in purified myelin protein over 1/2-6 hr of incubation utilizing a supplemented medium containing amino acids. This model was then used to examine myelin protein synthesis in nerves undergoing degeneration at 1-4 days following a crush injury. Data showed a statistically significant decrease in the ratio of fucose to leucine at 2, 3, and 4 days of degeneration, which was the consequence of a significant increase in leucine uptake. These results, plus substantial protein recovery in axotomized nerves, are indicative of active synthesis of proteins that purify with myelin during early Wallerian degeneration.

  1. Exposure to Sublethal Doses of Fipronil and Thiacloprid Highly Increases Mortality of Honeybees Previously Infected by Nosema ceranae

    PubMed Central

    Vidau, Cyril; Diogon, Marie; Aufauvre, Julie; Fontbonne, Régis; Viguès, Bernard; Brunet, Jean-Luc; Texier, Catherine; Biron, David G.; Blot, Nicolas; El Alaoui, Hicham; Belzunces, Luc P.; Delbac, Frédéric

    2011-01-01

    Background The honeybee, Apis mellifera, is undergoing a worldwide decline whose origin is still in debate. Studies performed for twenty years suggest that this decline may involve both infectious diseases and exposure to pesticides. Joint action of pathogens and chemicals are known to threaten several organisms but the combined effects of these stressors were poorly investigated in honeybees. Our study was designed to explore the effect of Nosema ceranae infection on honeybee sensitivity to sublethal doses of the insecticides fipronil and thiacloprid. Methodology/Finding Five days after their emergence, honeybees were divided in 6 experimental groups: (i) uninfected controls, (ii) infected with N. ceranae, (iii) uninfected and exposed to fipronil, (iv) uninfected and exposed to thiacloprid, (v) infected with N. ceranae and exposed 10 days post-infection (p.i.) to fipronil, and (vi) infected with N. ceranae and exposed 10 days p.i. to thiacloprid. Honeybee mortality and insecticide consumption were analyzed daily and the intestinal spore content was evaluated 20 days after infection. A significant increase in honeybee mortality was observed when N. ceranae-infected honeybees were exposed to sublethal doses of insecticides. Surprisingly, exposures to fipronil and thiacloprid had opposite effects on microsporidian spore production. Analysis of the honeybee detoxification system 10 days p.i. showed that N. ceranae infection induced an increase in glutathione-S-transferase activity in midgut and fat body but not in 7-ethoxycoumarin-O-deethylase activity. Conclusions/Significance After exposure to sublethal doses of fipronil or thiacloprid a higher mortality was observed in N. ceranae-infected honeybees than in uninfected ones. The synergistic effect of N. ceranae and insecticide on honeybee mortality, however, did not appear strongly linked to a decrease of the insect detoxification system. These data support the hypothesis that the combination of the increasing

  2. Monocytes from patients with rheumatoid arthritis and type 2 diabetes mellitus display an increased production of interleukin (IL)-1β via the nucleotide-binding domain and leucine-rich repeat containing family pyrin 3(NLRP3)-inflammasome activation: a possible implication for therapeutic decision in these patients.

    PubMed

    Ruscitti, P; Cipriani, P; Di Benedetto, P; Liakouli, V; Berardicurti, O; Carubbi, F; Ciccia, F; Alvaro, S; Triolo, G; Giacomelli, R

    2015-10-01

    A better understanding about the mechanisms involved in the pathogenesis of type 2 diabetes mellitus (T2D) showed that inflammatory cytokines such as tumour necrosis factor (TNF) and interleukin (IL)-1β play a pivotal role, mirroring data largely reported in rheumatoid arthritis (RA). IL-1β is produced mainly by monocytes (MO), and hyperglycaemia may be able to modulate, in the cytoplasm of these cells, the assembly of a nucleotide-binding domain and leucine-rich repeat containing family pyrin (NLRP3)-inflammosome, a cytosolic multi-protein platform where the inactive pro-IL-1β is cleaved into active form, via caspase-1 activity. In this paper, we evaluated the production of IL-1 β and TNF, in peripheral blood MO of patients affected by RA or T2D or both diseases, in order to understand if an alteration of the glucose metabolism may influence their proinflammatory status. Our data showed, after 24 h of incubation with different glucose concentrations, a significantly increased production of IL-1β and TNF in all evaluated groups when compared with healthy controls. However, a significant increase of IL-1β secretion by T2D/RA was observed when compared with other groups. The analysis of relative mRNA expression confirmed these data. After 24 h of incubation with different concentrations of glucose, our results showed a significant increase in NLRP3 expression. In this work, an increased production of IL-1β by MO obtained from patients affected by both RA and T2D via NLRP3-inflammasome activation may suggest a potential IL-1β targeted therapy in these patients.

  3. Image gently, step lightly: increasing radiation dose awareness in pediatric interventions through an international social marketing campaign.

    PubMed

    Sidhu, Manrita K; Goske, Marilyn J; Coley, Brian J; Connolly, Bairbre; Racadio, John; Yoshizumi, Terry T; Utley, Tara; Strauss, Keith J

    2009-09-01

    In the past several decades, advances in imaging and interventional techniques have been accompanied by an increase in medical radiation dose to the public. Radiation exposure is even more important in children, who are more sensitive to radiation and have a longer lifespan during which effects may manifest. To address radiation safety in pediatric computed tomography, in 2008 the Alliance for Radiation Safety in Pediatric Imaging launched an international social marketing campaign entitled Image Gently. This article describes the next phase of the Image Gently campaign, entitled Step Lightly, which focuses on radiation safety in pediatric interventional radiology.

  4. Increased disease activity in a patient with sarcoidosis after high dose interleukin 2 treatment for metastatic renal cancer.

    PubMed

    Logan, T F; Bensadoun, E S

    2005-07-01

    Sarcoidosis is a disease of unknown aetiology in which cytokines such as interleukin 2 (IL-2) are thought to play an important role. We present the case history of a 48 year old man with sarcoidosis who received treatment with high dose IL-2 for metastatic renal cell cancer, following which he developed hypercalcaemia characterised by a raised level of 1,25-dihydroxyvitamin D (1,25-(OH)2-D3), a finding consistent with sarcoidosis associated hypercalcaemia. The increased activity in his sarcoidosis following IL-2 treatment provides direct supportive evidence for the role of IL-2 in the pathogenesis of sarcoidosis.

  5. Reduced cerebral glucose metabolism and increased brain capillary permeability following high-dose methotrexate chemotherapy: a positron emission tomographic study

    SciTech Connect

    Phillips, P.C.; Dhawan, V.; Strother, S.C.; Sidtis, J.J.; Evans, A.C.; Allen, J.C.; Rottenberg, D.A.

    1987-01-01

    Regional glucose metabolic rate constants and blood-to-brain transport of rubidium were estimated using positron emission tomography in an adolescent patient with a brain tumor, before and after chemotherapy with intravenous high-dose methotrexate. Widespread depression of cerebral glucose metabolism was apparent 24 hours after drug administration, which may reflect reduced glucose phosphorylation, and the influx rate constant for /sup 82/Rb was increased, indicating a drug-induced alteration in blood-brain barrier function. Associated changes in neuropsychological performance, electroencephalogram, and plasma amino acid concentration were identified in the absence of evidence of systemic methotrexate toxicity, suggesting primary methotrexate neurotoxicity.

  6. Evidence That Lifelong Low Dose Rates of Ionizing Radiation Increase Lifespan in Long- and Short-Lived Dogs

    PubMed Central

    Feinendegen, Ludwig E.; Socol, Yehoshua

    2017-01-01

    After the 1956 radiation scare to stop weapons testing, studies focused on cancer induction by low-level radiation. Concern has shifted to protecting “radiation-sensitive individuals.” Since longevity is a measure of health impact, this analysis reexamined data to compare the effect of dose rate on the lifespans of short-lived (5% and 10% mortality) dogs and on the lifespans of dogs at 50% mortality. The data came from 2 large-scale studies. One exposed 10 groups to different γ dose rates; the other exposed 8 groups to different lung burdens of plutonium. Reexamination indicated that normalized lifespans increased more for short-lived dogs than for average dogs, when radiation was moderately above background. This was apparent by interpolating between the lifespans of nonirradiated dogs and exposed dogs. The optimum lifespan increase appeared at 50 mGy/y. The threshold for harm (decreased lifespan) was 700 mGy/y for 50% mortality dogs and 1100 mGy/y for short-lived dogs. For inhaled α-emitting particulates, longevity was remarkably increased for short-lived dogs below the threshold for harm. Short-lived dogs seem more radiosensitive than average dogs and they benefit more from low radiation. If dogs model humans, this evidence would support a change to radiation protection policy. Maintaining exposures “as low as reasonably achievable” (ALARA) appears questionable. PMID:28321175

  7. Effect of a single dose of salmeterol on the increase in airway eosinophils induced by allergen challenge in asthmatic subjects

    PubMed Central

    Dente, F.; Bancalari, L.; Bacci, E.; Bartoli, M.; Carnevali, S.; Cianchetti, S.; Di, F; Giannini, D.; Vagaggini, B.; Testi, R.; Paggiaro, P.

    1999-01-01

    BACKGROUND—The long acting β2 agonist salmeterol is very effective in preventing asthmatic responses to specific stimuli, and this effect could theoretically be due to some anti-inflammatory property in addition to bronchodilator property.
METHODS—The protective effect of a single dose of salmeterol (50 µg) on allergen induced early and late responses and on the associated airway inflammation was investigated in a double blind, placebo controlled, crossover study in 11 atopic asthmatic subjects. Eosinophil percentages and concentrations of eosinophil cationic protein (ECP) in peripheral blood and in hypertonic saline induced sputum were measured 24 hours after allergen inhalation.
RESULTS—Salmeterol effectively inhibited both early and late asthmatic responses in comparison with placebo. Salmeterol also inhibited the increase in the percentage of eosinophils in the sputum 24hours after allergen inhalation (median (range) baseline 6% (1-36), after placebo 31% (5-75), after salmeterol 12% (1-63)). However, the increase in both sputum and serum ECP concentrations 24 hours after allergen challenge was not affected by pretreatment with salmeterol.
CONCLUSIONS—A single dose of salmeterol inhibits the allergen induced airway responses and the increase in sputum eosinophils after allergen challenge.

 PMID:10377209

  8. Evidence That Lifelong Low Dose Rates of Ionizing Radiation Increase Lifespan in Long- and Short-Lived Dogs.

    PubMed

    Cuttler, Jerry M; Feinendegen, Ludwig E; Socol, Yehoshua

    2017-01-01

    After the 1956 radiation scare to stop weapons testing, studies focused on cancer induction by low-level radiation. Concern has shifted to protecting "radiation-sensitive individuals." Since longevity is a measure of health impact, this analysis reexamined data to compare the effect of dose rate on the lifespans of short-lived (5% and 10% mortality) dogs and on the lifespans of dogs at 50% mortality. The data came from 2 large-scale studies. One exposed 10 groups to different γ dose rates; the other exposed 8 groups to different lung burdens of plutonium. Reexamination indicated that normalized lifespans increased more for short-lived dogs than for average dogs, when radiation was moderately above background. This was apparent by interpolating between the lifespans of nonirradiated dogs and exposed dogs. The optimum lifespan increase appeared at 50 mGy/y. The threshold for harm (decreased lifespan) was 700 mGy/y for 50% mortality dogs and 1100 mGy/y for short-lived dogs. For inhaled α-emitting particulates, longevity was remarkably increased for short-lived dogs below the threshold for harm. Short-lived dogs seem more radiosensitive than average dogs and they benefit more from low radiation. If dogs model humans, this evidence would support a change to radiation protection policy. Maintaining exposures "as low as reasonably achievable" (ALARA) appears questionable.

  9. SdAb heterodimer formation using leucine zippers

    NASA Astrophysics Data System (ADS)

    Goldman, Ellen R.; Anderson, George P.; Brozozog-Lee, P. Audrey; Zabetakis, Dan

    2013-05-01

    Single domain antibodies (sdAb) are variable domains cloned from camel, llama, or shark heavy chain only antibodies, and are among the smallest known naturally derived antigen binding fragments. SdAb derived from immunized llamas are able to bind antigens with high affinity, and most are capable of refolding after heat or chemical denaturation to bind antigen again. We hypothesized that the ability to produce heterodimeric sdAb would enable reagents with the robust characteristics of component sdAb, but with dramatically improved overall affinity through increased avidity. Previously we had constructed multimeric sdAb by genetically linking sdAb that bind non-overlapping epitopes on the toxin, ricin. In this work we explored a more flexible approach; the construction of multivalent binding reagents using multimerization domains. We expressed anti-ricin sdAb that recognize different epitopes on the toxin as fusions with differently charged leucine zippers. When the initially produced homodimers are mixed the leucine zipper domains will pair to produce heterodimers. We used fluorescence resonance energy transfer to confirm heterodimer formation. Surface plasmon resonance, circular dichroism, enzyme linked immunosorbent assays, and fluid array assays were used to characterize the multimer constructs, and evaluate their utility in toxin detection.

  10. A Combination of Leucine, Metformin, and Sildenafil Treats Nonalcoholic Fatty Liver Disease and Steatohepatitis in Mice

    PubMed Central

    Fu, Lizhi; Li, Fenfen; Cao, Qiang; Cui, Xin; Wu, Rui; Shi, Hang

    2016-01-01

    Sirt1, AMPK, and eNOS modulate hepatic energy metabolism and inflammation and are key players in the development of NASH. L-leucine, an allosteric Sirt1 activator, synergizes with low doses of metformin or sildenafil on the AMPK-eNOS-Sirt1 pathway to reverse mild NAFLD in preclinical mouse models. Here we tested a possible multicomponent synergy to yield greater therapeutic efficacy in NAFLD/NASH. Liver cells and macrophages or an atherogenic diet induced NASH mouse model was treated with two-way and three-way combinations. The three-way combination Sild-Met-Leu increased hepatic fatty acid oxidation and reduced lipogenic gene expression and inflammatory marker in vitro. In mice, Sild-Met-Leu reduced the diet induced increases of ALT, TGFβ, PAI-1, IL1β, and TNFα, hepatic collagen expression, and nearly completely reversed hepatocyte ballooning and triglyceride accumulation, while all two-way combinations had only modest effects. Therefore, these data provide preclinical evidence for therapeutic efficacy of Sild-Met-Leu in the treatment of NAFLD and NASH. PMID:28042486

  11. Regulation of amino acid transport system L by amino acid availability in CHO-K1 cells. A special role for leucine.

    PubMed

    Moreno, A; Lobatón, C D; Oxender, D L

    1985-10-10

    Starvation of CHO-K1 cells for leucine leads to a 3-4-fold increase in transport system L activity, without modification of transport through systems A and ASC. The concentration of leucine must be below 10 microM before the enhancement of transport can be clearly seen. To achieve low concentrations of leucine such as 10 microM, extensive dialysis of fetal calf serum was required. The enhancement of transport was completed after 12-24 h of starvation and was fully reversed within 1 h of re-feeding with leucine. Starvation for isoleucine, valine or phenylalanine also produced an increase in system L transport activity, but the effect was only one half of that seen following leucine starvation.

  12. 21 CFR 862.1460 - Leucine aminopeptidase test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ..., plasma, and urine. Leucine aminopeptidase measurements are used in the diagnosis and treatment of liver diseases such as viral hepatitis and obstructive jaundice. (b) Classification. Class I (general...

  13. 21 CFR 862.1460 - Leucine aminopeptidase test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ..., plasma, and urine. Leucine aminopeptidase measurements are used in the diagnosis and treatment of liver diseases such as viral hepatitis and obstructive jaundice. (b) Classification. Class I (general...

  14. 21 CFR 862.1460 - Leucine aminopeptidase test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ..., plasma, and urine. Leucine aminopeptidase measurements are used in the diagnosis and treatment of liver diseases such as viral hepatitis and obstructive jaundice. (b) Classification. Class I (general...

  15. 21 CFR 862.1460 - Leucine aminopeptidase test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ..., plasma, and urine. Leucine aminopeptidase measurements are used in the diagnosis and treatment of liver diseases such as viral hepatitis and obstructive jaundice. (b) Classification. Class I (general...

  16. Impact of leucine supplementation on exercise training induced anti-cardiac remodeling effect in heart failure mice.

    PubMed

    de Moraes, Wilson Max Almeida Monteiro; Melara, Thaís Plasti; de Souza, Pamella Ramona Moraes; Guimarães, Fabiana de Salvi; Bozi, Luiz Henrique Marchesi; Brum, Patricia Chakur; Medeiros, Alessandra

    2015-05-15

    Leucine supplementation potentiates the effects of aerobic exercise training (AET) on skeletal muscle; however, its potential effects associated with AET on cardiac muscle have not been clarified yet. We tested whether leucine supplementation would potentiate the anti-cardiac remodeling effect of AET in a genetic model of sympathetic hyperactivity-induced heart failure in mice (α2A/α2CARKO). Mice were assigned to five groups: wild type mice treated with placebo and sedentary (WT, n = 11), α2A/α2CARKO treated with placebo and sedentary (KO, n = 9), α2A/α2CARKO treated with leucine and sedentary (KOL, n = 11), α2A/α2CARKO treated with placebo and AET (KOT, n = 12) or α2A/α2CARKO treated with leucine and AET (KOLT, n = 12). AET consisted of four weeks on a treadmill with 60 min sessions (six days/week, 60% of maximal speed) and administration by gavage of leucine (1.35 g/kg/day) or placebo (distilled water). The AET significantly improved exercise capacity, fractional shortening and re-established cardiomyocytes' diameter and collagen fraction in KOT. Additionally, AET significantly prevented the proteasome hyperactivity, increased misfolded proteins and HSP27 expression. Isolated leucine supplementation displayed no effect on cardiac function and structure (KOL), however, when associated with AET (KOLT), it increased exercise tolerance to a higher degree than isolated AET (KOT) despite no additional effects on AET induced anti-cardiac remodeling. Our results provide evidence for the modest impact of leucine supplementation on cardiac structure and function in exercised heart failure mice. Leucine supplementation potentiated AET effects on exercise tolerance, which might be related to its recognized impact on skeletal muscle.

  17. Ethanol Dose- and Time-dependently Increases α and β Subunits of Mitochondrial ATP Synthase of Cultured Neonatal Rat Cardiomyocytes.

    PubMed

    Mashimo, Keiko; Arthur, Peter G; Ohno, Youkichi

    2015-01-01

    Mitochondria are target subcellular organelles of ethanol. In this study, the effects of ethanol on protein composition was examined with 2-dimensional electrophoresis of protein extracts from cultured neonatal rat cardiomyocytes exposed to 100 mM ethanol for 24 hours. A putative β subunit of mitochondrial ATP synthase was increased, which was confirmed by Western blot. The cellular protein abundances in the α and β subunits of ATP synthase increased in dose (0, 10, 50, and 100 mM) - and time (0.5 hour and 24 hours) -dependent manners. The DNA microarray analysis of total RNA extract demonstrated that gene expression of the corresponding messenger RNAs of these subunit proteins did not significantly alter due to 24-hour ethanol exposure. Therefore, protein expression of these nuclear-encoded mitochondrial proteins may be regulated at the translational, rather than the transcriptional, level. Alternatively, degradation of these subunit proteins might be decreased. Additionally, cellular ATP content of cardiomyocytes scarcely decreased following 24-hour exposure to any examined concentrations of ethanol. Previous studies, together with this study, have demonstrated that protein abundance of the α subunit or β subunit or both subunits of ATP synthase after ethanol exposure or dysfunctional conditions might differ according to tissue: significant increases in heart but decreases in liver and brain. Thus, it is suggested that the abundance of subunit proteins of mitochondrial ATP synthase in the ethanol-exposed heart, being different from that in the liver and brain, should increase dose-dependently through either translational upregulation or decreased degradation or both to maintain ATP production, as the heart requires much more energy than other tissues for continuing sustained contractions.

  18. Perineural Invasion Predicts Increased Recurrence, Metastasis, and Death From Prostate Cancer Following Treatment With Dose-Escalated Radiation Therapy

    SciTech Connect

    Feng, Felix Y.; Qian Yushen; Stenmark, Matthew H.; Halverson, Schuyler; Blas, Kevin; Vance, Sean; Sandler, Howard M.; Hamstra, Daniel A.

    2011-11-15

    Purpose: To assess the prognostic value of perineural invasion (PNI) for patients treated with dose-escalated external-beam radiation therapy for prostate cancer. Methods and Materials: Outcomes were analyzed for 651 men treated for prostate cancer with EBRT to a minimum dose {>=}75 Gy. We assessed the impact of PNI as well as pretreatment and treatment-related factors on freedom from biochemical failure (FFBF), freedom from metastasis (FFM), cause-specific survival (CSS), and overall survival. Results: PNI was present in 34% of specimens at biopsy and was significantly associated with higher Gleason score (GS), T stage, and prostate-specific antigen level. On univariate and multivariate analysis, the presence of PNI was associated with worse FFBF (hazard ratio = 1.7, p <0.006), FFM (hazard ratio = 1.8, p <0.03), and CSS (HR = 1.4, p <0.05) compared with absence of PNI; there was no difference in overall survival. Seven-year rates of FFBF, FFM, and CCS were 64% vs. 80%, 84% vs. 92%, and 91% vs. 95% for those patients with and without PNI, respectively. On recursive partitioning analysis, PNI predicted for worse FFM and CSS in patients with GS 8-10, with FFM of 67% vs. 89% (p <0.02), and CSS of 69% vs. 91%, (p <0.04) at 7 years for those with and without PNI, respectively. Conclusions: The presence of PNI in the prostate biopsy predicts worse clinical outcome for patients treated with dose-escalated external-beam radiation therapy. Particularly in patients with GS 8-10 disease, the presence of PNI suggests an increased risk of metastasis and prostate cancer death.

  19. Metabolic responses to dietary leucine restriction involve remodeling of adipose tissue and enhanced hepatic insulin signaling.

    PubMed

    Wanders, Desiree; Stone, Kirsten P; Dille, Kelly; Simon, Jacob; Pierse, Alicia; Gettys, Thomas W

    2015-01-01

    Dietary leucine was incrementally restricted to test whether limiting this essential amino acid (EAA) would fully reproduce the beneficial responses produced by dietary methionine restriction. Restricting leucine by 85% increased energy intake and expenditure within 5 to 7 days of its introduction and reduced overall accumulation of adipose tissue. Leucine restriction (LR) also improved glucose tolerance, increased hepatic release of fibroblast growth factor 21 into the blood stream, and enhanced insulin-dependent activation of Akt in liver. However, LR had no effect on hepatic lipid levels and failed to lower lipogenic gene expression in the liver. LR did affect remodeling of white and brown adipose tissues, increasing expression of both thermogenic and lipogenic genes. These findings illustrate that dietary LR reproduces many but not all of the physiological responses of methionine restriction. The primary differences occur in the liver, where methionine and LR cause opposite effects on tissue lipid levels and expression of lipogenic genes. Altogether, these findings suggest that the sensing systems which detect and respond to dietary restriction of EAAs act through mechanisms that both leucine and methionine are able to engage, and in the case of hepatic lipid metabolism, may be unique to specific EAAs such as methionine.

  20. Low-Dose Inorganic Mercury Increases Severity and Frequency of Chronic Coxsackievirus-Induced Autoimmune Myocarditis in Mice

    PubMed Central

    Nyland, Jennifer F.; Fairweather, DeLisa; Shirley, Devon L.; Davis, Sarah E.; Rose, Noel R.; Silbergeld, Ellen K.

    2012-01-01

    Mercury is a widespread environmental contaminant with neurotoxic impacts that have been observed over a range of exposures. In addition, there is increasing evidence that inorganic mercury (iHg) and organic mercury (including methyl mercury) have a range of immunotoxic effects, including immune suppression and induction of autoimmunity. In this study, we investigated the effect of iHg on a model of autoimmune heart disease in mice induced by infection with coxsackievirus B3 (CVB3). We examined the role of timing of iHg exposure on disease; in some experiments, mice were pretreated with iHg (200 μg/kg, every other day for 15 days) before disease induction with virus inoculation, and in others, they were treated with iHg after the acute (viral) phase of disease but before the development of dilated cardiomyopathy (DCM). iHg alone had no effect on heart pathology. Pretreatment with iHg before CVB3 infection significantly increased the severity of chronic myocarditis and DCM compared with control animals receiving vehicle alone. In contrast, treatment with iHg after acute myocarditis did not affect the severity of chronic disease. The increased chronic myocarditis, fibrosis, and DCM induced by iHg pretreatment were not due to increased viral replication in the heart, which was unaltered by iHg treatment. iHg pretreatment induced a macrophage infiltrate and mixed cytokine response in the heart during acute myocarditis, including significantly increased interleukin (IL)-12, IL-17, interferon-γ, and tumor necrosis factor-α levels. IL-17 levels were also significantly increased in the spleen during chronic disease. Thus, we show for the first time that low-dose Hg exposure increases chronic myocarditis and DCM in a murine model. PMID:21984480

  1. Pushing product formation to its limit: metabolic engineering of Corynebacterium glutamicum for L-leucine overproduction.

    PubMed

    Vogt, Michael; Haas, Sabine; Klaffl, Simon; Polen, Tino; Eggeling, Lothar; van Ooyen, Jan; Bott, Michael

    2014-03-01

    Using metabolic engineering, an efficient L-leucine production strain of Corynebacterium glutamicum was developed. In the wild type of C. glutamicum, the leuA-encoded 2-isopropylmalate synthase (IPMS) is inhibited by low L-leucine concentrations with a K(i) of 0.4 mM. We identified a feedback-resistant IMPS variant, which carries two amino acid exchanges (R529H, G532D). The corresponding leuA(fbr) gene devoid of the attenuator region and under control of a strong promoter was integrated in one, two or three copies into the genome and combined with additional genomic modifications aimed at increasing L-leucine production. These modifications involved (i) deletion of the gene encoding the repressor LtbR to increase expression of leuBCD, (ii) deletion of the gene encoding the transcriptional regulator IolR to increase glucose uptake, (iii) reduction of citrate synthase activity to increase precursor supply, and (iv) introduction of a gene encoding a feedback-resistant acetohydroxyacid synthase. The production performance of the resulting strains was characterized in bioreactor cultivations. Under fed-batch conditions, the best producer strain accumulated L-leucine to levels exceeding the solubility limit of about 24 g/l. The molar product yield was 0.30 mol L-leucine per mol glucose and the volumetric productivity was 4.3 mmol l⁻¹ h⁻¹. These values were obtained in a defined minimal medium with a prototrophic and plasmid-free strain, making this process highly interesting for industrial application.

  2. Leucine pulses enhance skeletal muscle protein synthesis during continuous feeding in neonatal pigs

    PubMed Central

    Boutry, Claire; El-Kadi, Samer W.; Suryawan, Agus; Wheatley, Scott M.; Orellana, Renán A.; Kimball, Scot R.; Nguyen, Hanh V.

    2013-01-01

    Infants unable to maintain oral feeding can be nourished by orogastric tube. We have shown that orogastric continuous feeding restricts muscle protein synthesis compared with intermittent bolus feeding in neonatal pigs. To determine whether leucine infusion can be used to enhance protein synthesis during continuous feeding, neonatal piglets received the same amount of formula enterally by orogastric tube for 25.25 h continuously (CON) with or without LEU or intermittently by bolus every 4 h (BOL). For the CON+LEU group, leucine pulses were administered parenterally (800 μmol·kg−1·h−1) every 4 h. Insulin and glucose concentrations increased after the BOL meal and were unchanged in groups fed continuously. LEU infusion during CON feeding increased plasma leucine after the leucine pulse and decreased essential amino acids compared with CON feeding. Protein synthesis in longissimus dorsi (LD), gastrocnemius, and soleus muscles, but not liver or heart, were greater in CON+LEU and BOL than in the CON group. BOL feeding increased protein synthesis in the small intestine. Muscle S6K1 and 4E-BP1 phosphorylation and active eIF4E·eIF4G complex formation were higher in CON+LEU and BOL than in CON but AMPKα, eIF2α, and eEF2 phosphorylation were unchanged. LC3-II-to-total LC3 ratio was lower in CON+LEU and BOL than in CON, but there were no differences in atrogin-1 and MuRF-1 abundance and FoxO3 phosphorylation. In conclusion, administration of leucine pulses during continuous orogastric feeding in neonates increases muscle protein synthesis by stimulating translation initiation and may reduce protein degradation via the autophagy-lysosome, but not the ubiquitin-proteasome pathway. PMID:23839523

  3. Hypoalbuminemia is significantly associated with increased clearance time of high dose methotrexate in patients being treated for lymphoma or leukemia.

    PubMed

    Reiss, Samantha N; Buie, Larry W; Adel, Nelly; Goldman, Debra A; Devlin, Sean M; Douer, Dan

    2016-12-01

    As a weak acid, methotrexate (MTX) is bound to serum albumin and has variable protein binding. The purpose of this study was to assess serum albumin's relationship with MTX pharmacokinetics by comparing MTX clearance and toxicities between patients with normal serum albumin to those with hypoalbuminemia. This single-center retrospective study included adult patients with leukemia or lymphoma who received their first MTX at a dose ≥1 g/m(2). Hypoalbuminemia was defined as serum albumin ≤3.4 g/dL. MTX clearance was defined as the first documented time the MTX level ≤0.05 μM. Fisher's exact tests and Wilcoxon rank sum tests were used to examine differences in toxicities, and Cox proportional hazard regression was used to assess relationship with time to clearance. Of 523 patients identified, 167 patients were evaluable. One hundred thirty-five patients had normal serum albumin and 32 had hypoalbuminemia. Hypoalbuminemia was associated with a higher proportion of patients experiencing edema, ascites or pleural effusions (34 vs. 12 %, p = 0.006), and the concomitant use of nephrotoxic agents (41 vs. 20 %, p = 0.021). Hypoalbuminemia was associated with a significantly longer time to MTX clearance (median 96 vs. 72 h, p = 0.004). In addition, patients with hypoalbuminemia had a higher proportion of hyperbilirubinemia and significantly longer hospitalization (median 14 vs. 5 days, p < 0.001). In conclusion, hypoalbuminemia was associated with increased time to MTX clearance and increased length of hospitalization. High dose MTX is safe to administer in patients with low albumin levels, with appropriate leucovorin rescue, and good supportive care.

  4. Leucine acts as a nutrient signal to stimulate protein synthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The postprandial rise in amino acids and insulin independently stimulates protein synthesis in skeletal muscle of piglets. Leucine is an important mediator of the response to amino acids. We have shown that the postprandial rise in leucine, but not isoleucine or valine, acutely stimulates muscle pro...

  5. Dose-dependent increases in flow-mediated dilation following acute cocoa ingestion in healthy older adults

    PubMed Central

    Feehan, Robert P.; Kunselman, Allen R.; Preston, Amy G.; Miller, Debra L.; Lott, Mary E. J.

    2011-01-01

    An inverse relation exists between intake of flavonoid-rich foods, such as cocoa, and cardiovascular-related mortality. Favorable effects of flavonoids on the endothelium may underlie these associations. We performed a randomized, double-blind, placebo-controlled study to test the hypothesis that acute cocoa ingestion dose dependently increases endothelium-dependent vasodilation, as measured by an increase in brachial artery flow-mediated dilation (FMD), in healthy older adults. Measurements were obtained before (preingestion) and after (1- and 2-h postingestion) ingestion of 0 (placebo), 2, 5, 13, and 26 g of cocoa in 23 adults (63 ± 2 yr old, mean ± SE). Changes in brachial artery FMD 1- and 2-h postingestion compared with preingestion were used to determine the effects of cocoa. FMD was unchanged 1 (Δ−0.3 ± 0.2%)- and 2-h (Δ0.1 ± 0.1%) after placebo (0 g cocoa). In contrast, FMD increased both 1-h postingestion (2 g cocoa Δ0.0 ± 0.2%, 5 g cocoa Δ0.8 ± 0.3%, 13 g cocoa Δ1.0 ± 0.3%, and 26 g cocoa Δ1.6 ± 0.3%: P < 0.05 compared with placebo for 5, 13, and 26 g cocoa) and 2-h postingestion (2 g cocoa Δ0.5 ± 0.3%, 5 g cocoa Δ1.0 ± 0.3%, 13 g cocoa Δ1.4 ± 0.2%, and 26 g cocoa Δ2.5 ± 0.4%: P < 0.05 compared with placebo for 5, 13, and 26 g cocoa) on the other study days. A serum marker of cocoa ingestion (total epicatechin) correlated with increased FMD 1- and 2-h postingestion (r = 0.44–0.48; both P < 0.05). Collectively, these results indicate that acute cocoa ingestion dose dependently increases brachial artery FMD in healthy older humans. These responses may help to explain associations between flavonoid intake and cardiovascular-related mortality in humans. PMID:21903881

  6. [An evaluation of the biological action of increased doses of EMI in the UV range on the functional state and productivity of sheep].

    PubMed

    Ivanov, V L; Ipatova, A G; Zeĭnalov, A A; Kozlov, V A; Sarukhanov, V Ia

    2000-01-01

    The effect of increased UV-radiation doses modeling 25 and 50% of ozone layer depletion on sheep's organisms was studied in the field experiment. The character of changes in animal organisms was found to depend on irradiation doses, sensitivity of individual system of living organism to electromagnetic radiation and physiological peculiarities of protection.

  7. Single-Dose Local Simvastatin Injection Improves Implant Fixation via Increased Angiogenesis and Bone Formation in an Ovariectomized Rat Model

    PubMed Central

    Tan, Jie; Yang, Ning; Fu, Xin; Cui, Yueyi; Guo, Qi; Ma, Teng; Yin, Xiaoxue; Leng, Huijie; Song, Chunli

    2015-01-01

    Background Statins have been reported to promote bone formation. However, taken orally, their bioavailability is low to the bones. Implant therapies require a local repair response, topical application of osteoinductive agents, or biomaterials that promote implant fixation. Material/Methods The present study evaluated the effect of a single local injection of simvastatin on screw fixation in an ovariectomized rat model of osteoporosis. Results Dual-energy X-ray absorptiometry, micro-computed tomography, histology, and biomechanical tests revealed that 5 and 10 mg simvastatin significantly improved bone mineral density by 18.2% and 22.4%, respectively (P<0.05); increased bone volume fraction by 51.0% and 57.9%, trabecular thickness by 16.4% and 18.9%, trabeculae number by 112.0% and 107.1%, and percentage of osseointegration by 115.7% and 126.3%; and decreased trabeculae separation by 34.1% and 36.6%, respectively (all P<0.01). Bone mineral apposition rate was significantly increased (P<0.01). Furthermore, implant fixation was significantly increased (P<0.05), and bone morphogenetic protein 2 (BMP2) expression was markedly increased. Local injection of a single dose of simvastatin also promoted angiogenesis. Vessel number, volume, thickness, surface area, and vascular volume per tissue volume were significantly increased (all P<0.01). Vascular endothelial growth factor (VEGF), VEGF receptor-2, von Willebrand factor, and platelet endothelial cell adhesion molecule-1 expression were enhanced. Conclusions A single local injection of simvastatin significantly increased bone formation, promoted osseointegration, and enhanced implant fixation in ovariectomized rats. The underlying mechanism appears to involve enhanced BMP2 expression and angiogenesis in the target bone. PMID:25982481

  8. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    PubMed Central

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise. PMID:27367725

  9. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running.

    PubMed

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-06-28

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d₃-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise.

  10. Dose-dependent increase and decrease in active glucose uptake in jejunal epithelium of broilers after acute exposure to ethanol.

    PubMed

    Yunus, Agha Waqar; Awad, Wageha A; Kröger, Susan; Zentek, Jürgen; Böhm, Josef

    2011-06-01

    Little is known about the effects of ethanol on gastrointestinal tract of chicken. In this study, we investigated the effects of low levels of ethanol on electrophysiological variables of jejunal epithelium of commercial broilers. Jejunal tissues from 35- to 39-day-old broilers were exposed to either 0 or 0.1% ethanol in Ussing chambers, and electrophysiological variables were monitored for 40 min. After 40 and 60 min of incubation, glucose (20 mM) and carbamoylcholine (200 μM), respectively, were introduced into the chambers. The absolute and percent increase in short-circuit current (Isc) and potential difference (Vt) induced by glucose were increased significantly with 0.1% ethanol. There was no significant effect of 0.1% ethanol on carbamoylcholine-induced electrophysiological variables. To investigate if higher levels of ethanol have similar effects, we tested the effects of 0, 0.33, and 0.66% ethanol under similar experimental conditions until the glucose-addition step. Contrary to 0.1% ethanol, both the 0.33 and 0.66% ethanol levels significantly decreased the basal and glucose-induced Isc and Vt. Tissue conductivity remained unaffected in all cases. These results indicate that intestinal epithelia of chicken may be more sensitive to the effects of ethanol as compared with other species. This is the first report indicating dose-dependent increase and decrease in active glucose absorption in intestinal epithelia in the presence of ethanol.

  11. Leukemia patient-derived lymphoblastoid cell lines exhibit increased induction of leukemia-associated transcripts following high-dose irradiation.

    PubMed

    Spencer, A; Granter, N

    1999-09-01

    Improvement in diagnostic cytogenetic techniques has led to the recognition of an increasing number of leukemia-associated chromosomal translocations and inversions. These genetic lesions frequently are associated with the disruption of putative transcription factors and the production of hybrid transcripts that are implicated in leukemogenesis. Epidemiologic evidence suggests that some, but not all, individuals with a history of gamma-irradiation exposure are at increased risk of developing chronic myeloid leukemia (CML). CML is characterized by the Philadelphia chromosome and transcription of the resulting hybrid BCR-ABL gene. Utilizing the leukemia-associated BCR-ABL p210 transcript as a marker, we sought differences in the induction of illegitimate genetic recombination following high-dose gamma-irradiation of karyotypically normal lymphoblastoid cell lines (LCL) derived from individuals with and without a history of myeloid leukemias. Six LCL [4 leukemia patient derived [2 acute myeloid leukemia and 2 CML] and 2 from normal individuals were analyzed with reverse transcriptase polymerase chain reaction for BCR-ABL under stringent conditions following exposure to 0, 50, or 100 Gy of LET gamma-irradiation delivered via a Varian linear accelerator at 4 MV. Transcripts identical to disease-associated b2a2 and b3a2 transcripts were detected both spontaneously (background illegitimate genetic recombination) and following gamma-irradiation. Background BCR-ABL positivity was demonstrable in 4 of the 6 LCL, with no significant difference in detection between leukemic- and nonleukemic-derived LCL. Overall, increasing gamma-irradiation dose resulted in an increased frequency of BCR-ABL transcript detection (0 Gy vs 50 Gy vs 100 Gy,p = 0.0023, Chi-square test). Within the leukemic- but not the nonleukemic-derived LCL there was significantly greater BCR-ABL positivity after gamma-irradiation compared to unirradiated equivalents. Furthermore, the BCR-ABL positivity of both

  12. Leucine supplementation has an anabolic effect on proteins in rabbit skin wound and muscle.

    PubMed

    Zhang, Xiao-Jun; Chinkes, David L; Wolfe, Robert R

    2004-12-01

    We investigated the effect of leucine supplementation on protein metabolism in skin wounds and muscle in anesthetized rabbits. l-[ring-(13)C(6)]phenylalanine was infused on d 7 after the ear was scalded, and the scalded ear and uninjured hindlimb were used as arteriovenous units to reflect protein kinetics in skin wounds and muscle. In comparison with a commercially available amino acid solution (10% Travasol), isonitrogenous [1638 micromol/(kg . h)] infusion of the amino acid solution with supplemental leucine to account for 35% of total nitrogen increased the net phenylalanine balance (P < 0.05) in the skin wound and muscle from -6.7 +/- 6.1 to 0.9 +/- 1.4 and from -4.4 +/- 2.4 to -1.0 +/- 0.4 micromol/(100 g . h), respectively. Infusion of leucine alone did not significantly improve the net phenylalanine balance in either skin wounds [-4.0 +/- 4.6 micromol/(100 g . h)] or muscle [-2.7 +/- 0.7 micromol/(100 g . h)]. We conclude that leucine supplementation had an anabolic effect on proteins in skin wounds and muscle, provided that adequate additional amino acids were also available.

  13. Transient increase in intrahepatic pressure mediates successful treatment of the Gunn rat with reduced doses of lentiviral vector.

    PubMed

    Schmitt, Françoise; Flageul, Maude; Dariel, Anne; Pichard, Virginie; Pontes, Cecilia Abarrategui; Boni, Sébastien; Podevin, Guillaume; Myara, Anne; Ferry, Nicolas; Nguyen, Tuan Huy

    2010-10-01

    Lentiviral vectors can stably transduce hepatocytes and are promising tools for gene therapy of hepatic diseases. Although hepatocytes are accessible to blood-borne viral vectors through fenestrations of the hepatic endothelium, improved liver transduction after delivery of vectors to the blood stream is needed. As the normal endothelial fenestration and lentiviral vectors are similar in size (150 nm), we hypothesized that a transient increase in hepatic blood pressure may enhance in vivo gene transfer to hepatocytes. We designed a simple surgical procedure, by which the liver is temporarily excluded from blood flow. Lentiviral vectors were injected in a large volume to increase intrahepatic pressure. We demonstrated that in the Gunn rat, a model of Crigler-Najjar disease, the administration of low vector doses (corresponding to a multiplicity of infection of 0.2) by this procedure resulted in therapeutic correction of hyperbilirubinemia, without toxicity. The correction was sustained for 10 months (end of study). The same vector amounts yielded only partial correction after intraportal delivery. We believe that this new and clinically applicable strategy may broaden the range of genetic liver diseases accessible to gene therapy.

  14. Atrazine Does Not Induce Pica Behavior at Doses that Increase Hypothalamic-Pituitary-Adrenal Axis Activation and Cause Conditioned Taste Avoidance.

    EPA Science Inventory

    Previous work has shown that a single oral administration of atrazine (ATR), a chlorotriazine herbicide, induces dose-dependent increases in plasma adrenocorticotropic hormone (ACTH), serum corticosterone (CORT) and progesterone. The mechanism for these effects is unknown. To tes...

  15. Severe, Protracted Spasm of Urinary Bladder and Autonomic Dysreflexia Caused by Changing the Suprapubic Catheter in a Cervical Spinal Cord Injury Patient: Treatment by a Bolus Dose and Increased Total Daily Dose of Intrathecal Baclofen

    PubMed Central

    Vaidyanathan, Subramanian; Oo, Tun; Soni, Bakul M.; Hughes, Peter L.; Singh, Gurpreet

    2016-01-01

    BACKGROUND Intrathecal administration of baclofen by implanted pump reduces rigidity and muscle spasms. Its use specifically to control bladder spasms has not been reported. CASE REPORT A tetraplegic patient developed severe, protracted, bladder spasms, abdominal muscles spasms, and high blood pressure after change of suprapubic catheter; nifedipine, diazepam, and paracetamol did not control spasms; bolus dose of baclofen intrathecally produced prompt relief via baclofen pump. CONCLUSION Severe, protracted bladder spasms, abdominal muscles spasms, and autonomic dysreflexia, induced by change of suprapubic catheter in a spinal cord injury patient, were treated successfully by a bolus dose and increased total daily dose of intrathecal baclofen. PMID:28008298

  16. The combination of ursolic acid and leucine potentiates the differentiation of C2C12 murine myoblasts through the mTOR signaling pathway.

    PubMed

    Kim, Minjung; Sung, Bokyung; Kang, Yong Jung; Kim, Dong Hwan; Lee, Yujin; Hwang, Seong Yeon; Yoon, Jeong-Hyun; Yoo, Mi-Ae; Kim, Cheol Min; Chung, Hae Young; Kim, Nam Deuk

    2015-03-01

    Aging causes phenotypic changes in skeletal muscle progenitor cells that lead to the progressive loss of myogenic differentiation and thus a decrease in muscle mass. The naturally occurring triterpene, ursolic acid, has been reported to be an effective agent for the prevention of muscle loss by suppressing degenerative muscular dystrophy. Leucine, a branched-chain amino acid, and its metabolite, β-hydroxy-β-methylbutyric acid, have been reported to enhance protein synthesis in skeletal muscle. Therefore, the aim of the present study was to investigate whether the combination of ursolic acid and leucine promotes greater myogenic differentiation compared to either agent alone in C2C12 murine myoblasts. Morphological changes were observed and creatine kinase (CK) activity analysis was performed to determine the conditions through which the combination of ursolic acid and leucine would exert the most prominent effects on muscle cell differentiation. The effect of the combination of ursolic acid and leucine on the expression of myogenic differentiation marker genes was examined by RT-PCR and western blot analysis. The combination of ursolic acid (0.5 µM) and leucine (10 µM) proved to be the most effective in promoting myogenic differentiation. The combination of ursolic acid and leucine significantly increased CK activity than treatment with either agent alone. The level of myosin heavy chain, a myogenic differentiation marker protein, was also enhanced by the combination of ursolic acid and leucine. The combination of ursolic acid and leucine significantly induced the expression of myogenic differentiation marker genes, such as myogenic differentiation 1 (MyoD) and myogenin, at both the mRNA and protein level. In addition, the number of myotubes and the fusion index were increased. These findings indicate that the combination of ursolic acid and leucine promotes muscle cell differentiation, thus suggesting that this combination of agents may prove to be beneficial

  17. Leucine supplementation at the onset of high-fat feeding does not prevent weight gain or improve glycemic regulation in male Sprague-Dawley rats.

    PubMed

    Baum, Jamie I; Washington, Tyrone A; Shouse, Stephanie A; Bottje, Walter; Dridi, Sami; Davis, Gina; Smith, Dameon

    2016-12-01

    Obesity is a major public health concern and it is essential to identify effective treatments and preventative strategies to stop continued increases in obesity rates. The potential functional roles of the branched chain amino acid leucine make this amino acid an attractive candidate for the treatment and/or prevention of obesity. The objective of this study was to determine if long-term leucine supplementation could prevent the development of obesity and reduce the risk factors for chronic disease in rats fed a high-fat (60 % fat) diet. Male Sprague-Dawley rats (n = 30 per dietary treatment) were meal-fed (3 meals/day) either a control, low-fat diet (LF), control + leucine (LFL), high-fat (HF), or high-fat + leucine (HFL) for 42 days. On day 42, rats were sacrificed at 0, 30, or 90 min postprandial. Animals fed the HF and HFL diets had higher (P < 0.05) final body weights and weight gain compared to animals fed the LF and LFL diets. Leucine supplementation increased epididymal fat mass (P < 0.05) and decreased muscle mass (P < 0.05). There was no effect of leucine supplementation on postprandial glucose or insulin response. However, there was a significant effect (P < 0.05) of diet and time on free fatty acid concentrations. There was no effect of leucine on muscle markers of protein synthesis (4E-BP1, p70S6K) or energy metabolism (Akt, AMPK). Leucine supplementation decreased (P < 0.05) PGC1α expression and increased (P < 0.05) PPARγ expression in skeletal muscle. In conclusion, long-term leucine supplementation does not prevent weight gain, improve body composition, or improve glycemic control in rats fed a high-fat diet.

  18. Oral challenge with increasing doses of LPS modulated the patterns of plasma metabolites and minerals in periparturient dairy cows.

    PubMed

    Zebeli, Qendrim; Mansmann, Dominik; Sivaraman, Shanti; Dunn, Suzanna M; Ametaj, Burim N

    2013-06-01

    We showed recently that repeated oral exposure to LPS stimulated humoral immune responses in periparturient dairy cows. Here, metabolic and mineral responses to repeated oral administration of LPS were investigated. Sixteen clinically healthy, pregnant Holstein cows were orally administered 3 ml of saline solution (control) or 3 ml of saline solution containing 3 increasing doses of LPS, at 07:00 h, as follows: (i) 0.01 µg/kg body mass (BM) on d -14 and -10, (ii) 0.05 µg/kg BM on d -7 and -3, and (iii) 0.1 µg/kg BM on d 3 and 7 relative to parturition. Blood samples were measured shortly before, and at 8 different time-points after (up to 6 h), the first challenge of each LPS dosage to evaluate the post-challenge plasma profile, as well as weekly up to 4 wk postpartum. Results showed that oral administration of LPS lowered concentrations of non-esterified fatty acids (P < 0.01) and β-hydroxy-butyrate (P < 0.01) in the plasma, particularly after the third LPS challenge. Also, after the third oral LPS challenge, treatment tended to increase plasma glucose. Plasma calcium did not change, but concentrations of insulin (P < 0.01) and zinc (P < 0.01) were greater, while that of copper was lower (P < 0.01) in the plasma of treated cows. This is the first report to indicate a potential role for repeated oral administration of LPS around parturition to modulate the profile of plasma metabolites and minerals postpartum.

  19. Analysis of the effects of increasing doses of ionizing radiation to the exteriorized rat ovary on follicular development, atresia, and serum gonadotropin levels

    SciTech Connect

    Jarrell, J.; YoungLai, E.V.; Barr, R.; O'Connell, G.; Belbeck, L.; McMahon, A.

    1986-02-01

    There is increasing interest in the effects of environmental and therapeutic agents on the reproductive system, in particular, the ovary. To study the effects of controlled doses of ionizing radiation to the ovary, Sprague-Dawley rats had their ovaries exteriorized and subjected to increasing doses of radiation. There was a significant increase in ovarian follicular atresia, a significant increase in serum follicle-stimulating hormone levels, but no change in serum luteinizing hormone levels. This experimental protocol may facilitate the testing putative radioprotectants.

  20. Additive insulinogenic action of Opuntia ficus-indica cladode and fruit skin extract and leucine after exercise in healthy males

    PubMed Central

    2013-01-01

    Background Oral intake of a specific extract of Opuntia ficus-indica cladode and fruit skin (OpunDia™) (OFI) has been shown to increase serum insulin concentration while reducing blood glucose level for a given amount of glucose ingestion after an endurance exercise bout in healthy young volunteers. However, it is unknown whether OFI-induced insulin stimulation after exercise is of the same magnitude than the stimulation by other insulinogenic agents like leucine as well as whether OFI can interact with those agents. Therefore, the aims of the present study were: 1) to compare the degree of insulin stimulation by OFI with the effect of leucine administration; 2) to determine whether OFI and leucine have an additive action on insulin stimulation post-exercise. Methods Eleven subjects participated in a randomized double-blind cross-over study involving four experimental sessions. In each session the subjects successively underwent a 2-h oral glucose tolerance test (OGTT) after a 30-min cycling bout at ~70% VO2max. At t0 and t60 during the OGTT, subjects ingested 75 g glucose and capsules containing either 1) a placebo; 2) 1000 mg OFI; 3) 3 g leucine; 4) 1000 mg OFI + 3 g leucine. Blood samples were collected before and at 30-min intervals during the OGTT for determination of blood glucose and serum insulin. Results Whereas no effect of leucine was measured, OFI reduced blood glucose at t90 by ~7% and the area under the glucose curve by ~15% and increased serum insulin concentration at t90 by ~35% compared to placebo (P<0.05). From t60 to the end of the OGTT, serum insulin concentration was higher in OFI+leucine than in placebo which resulted in a higher area under the insulin curve (+40%, P<0.05). Conclusion Carbohydrate-induced insulin stimulation post-exercise can be further increased by the combination of OFI with leucine. OFI and leucine could be interesting ingredients to include together in recovery drinks to resynthesize muscle glycogen faster post

  1. Addition of a third field significantly increases dose to the brachial plexus for patients undergoing tangential whole-breast therapy after lumpectomy

    SciTech Connect

    Stanic, Sinisa; Mathai, Mathew; Mayadev, Jyoti S.; Do, Ly V.; Purdy, James A.; Chen, Allen M.

    2012-07-01

    Our goal was to evaluate brachial plexus (BP) dose with and without the use of supraclavicular (SCL) irradiation in patients undergoing breast-conserving therapy with whole-breast radiation therapy (RT) after lumpectomy. Using the standardized Radiation Therapy Oncology Group (RTOG)-endorsed guidelines delineation, we contoured the BP for 10 postlumpectomy breast cancer patients. The radiation dose to the whole breast was 50.4 Gy using tangential fields in 1.8-Gy fractions, followed by a conedown to the operative bed using electrons (10 Gy). The prescription dose to the SCL field was 50.4 Gy, delivered to 3-cm depth. The mean BP volume was 14.5 {+-} 1.5 cm{sup 3}. With tangential fields alone, the median mean dose to the BP was 0.57 Gy, the median maximum dose was 1.93 Gy, and the irradiated volume of the BP receiving 40, 45, and 50 Gy was 0%. When the third (SCL field) was added, the dose to the BP was significantly increased (P = .01): the median mean dose to the BP was 40.60 Gy, and the median maximum dose was 52.22 Gy. With 3-field RT, the median irradiated volume of the BP receiving 40, 45, and 50 Gy was 83.5%, 68.5%, and 24.6%, respectively. The addition of the SCL field significantly increases dose to the BP. The possibility of increasing the risk of BP morbidity should be considered in the context of clinical decision making.

  2. Leucine-induced activation of translational initiation is partly regulated by the branched-chain {alpha}-keto acid dehydrogenase complex in C2C12 cells

    SciTech Connect

    Nakai, Naoya . E-mail: nakai@hss.osaka-u.ac.jp; Shimomura, Yoshiharu; Tamura, Tomohiro; Tamura, Noriko; Hamada, Koichiro; Kawano, Fuminori; Ohira, Yoshinobu

    2006-05-19

    Branched-chain amino acid leucine has been shown to activate the translational regulators through the mammalian target of rapamycin. However, the leucine's effects are self-limiting because leucine promotes its own disposal by an oxidative pathway. The irreversible and rate-limiting step in the leucine oxidation pathway is catalyzed by the branched-chain {alpha}-keto acid dehydrogenase (BCKDH) complex. The complex contains E1 ({alpha}2{beta}2), E2, and E3 subunits, and its activity is abolished by phosphorylation of the E1{alpha} subunit by BCKDH kinase. The relationship between the activity of BCKDH complex and leucine-mediated activation of the protein translation was investigated using the technique of RNA interference. The activity of BCKDH complex in C2C12 cell was modulated by transfection of small interfering RNA (siRNA) for BCKDH E2 subunit or BCKDH kinase. Transfection of siRNAs decreased the mRNA expression and protein amount of corresponding gene. Suppression of either E2 subunit or kinase produced opposite effects on the cell proliferation and the activation of translational regulators by leucine. Suppression of BCKDH kinase for 48 h resulted in decreasing cell proliferation. In contrast, E2 suppression led to increased amount of total cellular protein. The phosphorylation of p70 S6 kinase by leucine was increased in E2-siRNA transfected C2C12 cells, whereas the leucine's effect was diminished in kinase-siRNA transfected cells. These results suggest that the activation of the translational regulators by leucine was partly regulated by the activity of BCKDH complex.

  3. The increase in animal mortality risk following exposure to sparsely ionizing radiation is not linear quadratic with dose

    SciTech Connect

    Haley, Benjamin M.; Paunesku, Tatjana; Grdina, David J.; Woloschak, Gayle E.; Aravindan, Natarajan

    2015-12-09

    The US government regulates allowable radiation exposures relying, in large part, on the seventh report from the committee to estimate the Biological Effect of Ionizing Radiation (BEIR VII), which estimated that most contemporary exposures- protracted or low-dose, carry 1.5 fold less risk of carcinogenesis and mortality per Gy than acute exposures of atomic bomb survivors. This correction is known as the dose and dose rate effectiveness factor for the life span study of atomic bomb survivors (DDREFLSS). As a result, it was calculated by applying a linear-quadratic dose response model to data from Japanese atomic bomb survivors and a limited number of animal studies.

  4. The increase in animal mortality risk following exposure to sparsely ionizing radiation is not linear quadratic with dose

    DOE PAGES

    Haley, Benjamin M.; Paunesku, Tatjana; Grdina, David J.; ...

    2015-12-09

    The US government regulates allowable radiation exposures relying, in large part, on the seventh report from the committee to estimate the Biological Effect of Ionizing Radiation (BEIR VII), which estimated that most contemporary exposures- protracted or low-dose, carry 1.5 fold less risk of carcinogenesis and mortality per Gy than acute exposures of atomic bomb survivors. This correction is known as the dose and dose rate effectiveness factor for the life span study of atomic bomb survivors (DDREFLSS). As a result, it was calculated by applying a linear-quadratic dose response model to data from Japanese atomic bomb survivors and a limitedmore » number of animal studies.« less

  5. The pharmacokinetics, CNS pharmacodynamics and adverse event profile of brivaracetam after multiple increasing oral doses in healthy men

    PubMed Central

    Rolan, Paul; Sargentini-Maier, Maria Laura; Pigeolet, Etienne; Stockis, Armel

    2008-01-01

    AIMS Brivaracetam is a novel synaptic vesicle protein 2A ligand that has shown potent activity in animal models of epilepsy. This study examined the pharmacokinetics, central nervous system pharmacodynamics and adverse event profile of multiple oral doses of brivaracetam in healthy male subjects. METHODS Three successive panels of 12 healthy male subjects received double-blind brivaracetam 200, 400 or 800 mg day−1 (all doses well above the expected therapeutic range) or placebo (9 : 3), in two divided doses, for 14 days. RESULTS Brivaracetam was rapidly absorbed (tmax∼2 h) and eliminated (t1/2 7–8 h). Volume of distribution was slightly lower than total body water. A small fraction of the dose (5–8%) was excreted unchanged in urine together with significant levels of metabolites, suggesting predominantly metabolic clearance. Based on 6-β-hydroxycortisol/cortisol ratios in urine, there was no evidence of induction of CYP3A4 activity. Saliva and plasma brivaracetam levels were highly correlated. Adverse events were mostly mild to moderate, central nervous system-related and resolved within the first day of treatment. No clinically relevant changes were observed in laboratory tests, vital signs, physical examinations or ECGs. Pharmacodynamic tests showed dose-related sedation and decreased alertness that only persisted at 800 mg daily. CONCLUSIONS Brivaracetam was well tolerated by healthy male volunteers at doses of 200–800 mg daily for 2 weeks, well above the expected clinically effective dose range. Brivaracetam had a favourable pharmacokinetic profile in this population, characterized by rapid absorption, volume of distribution limited to total body water, apparent single-compartment elimination and dose proportionality. WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT The pharmacokinetic profile, metabolism and proof of concept of a single oral dose of brivaracetam have been reported.Previous studies have shown that it was well absorbed, had linear kinetics

  6. Low dose influenza virus challenge in the ferret leads to increased virus shedding and greater sensitivity to oseltamivir.

    PubMed

    Marriott, Anthony C; Dove, Brian K; Whittaker, Catherine J; Bruce, Christine; Ryan, Kathryn A; Bean, Thomas J; Rayner, Emma; Pearson, Geoff; Taylor, Irene; Dowall, Stuart; Plank, Jenna; Newman, Edmund; Barclay, Wendy S; Dimmock, Nigel J; Easton, Andrew J; Hallis, Bassam; Silman, Nigel J; Carroll, Miles W

    2014-01-01

    Ferrets are widely used to study human influenza virus infection. Their airway physiology and cell receptor distribution makes them ideal for the analysis of pathogenesis and virus transmission, and for testing the efficacy of anti-influenza interventions and vaccines. The 2009 pandemic influenza virus (H1N1pdm09) induces mild to moderate respiratory disease in infected ferrets, following inoculation with 106 plaque-forming units (pfu) of virus. We have demonstrated that reducing the challenge dose to 102 pfu delays the onset of clinical signs by 1 day, and results in a modest reduction in clinical signs, and a less rapid nasal cavity innate immune response. There was also a delay in virus production in the upper respiratory tract, this was up to 9-fold greater and virus shedding was prolonged. Progression of infection to the lower respiratory tract was not noticeably delayed by the reduction in virus challenge. A dose of 104 pfu gave an infection that was intermediate between those of the 106 pfu and 102 pfu doses. To address the hypothesis that using a more authentic low challenge dose would facilitate a more sensitive model for antiviral efficacy, we used the well-known neuraminidase inhibitor, oseltamivir. Oseltamivir-treated and untreated ferrets were challenged with high (106 pfu) and low (102 pfu) doses of influenza H1N1pdm09 virus. The low dose treated ferrets showed significant delays in innate immune response and virus shedding, delayed onset of pathological changes in the nasal cavity, and reduced pathological changes and viral RNA load in the lung, relative to untreated ferrets. Importantly, these observations were not seen in treated animals when the high dose challenge was used. In summary, low dose challenge gives a disease that more closely parallels the disease parameters of human influenza infection, and provides an improved pre-clinical model for the assessment of influenza therapeutics, and potentially, influenza vaccines.

  7. Understanding the Recent Increase in Ferritin Levels in United States Dialysis Patients: Potential Impact of Changes in Intravenous Iron and Erythropoiesis-Stimulating Agent Dosing

    PubMed Central

    Zee, Jarcy; Morgenstern, Hal; Nolen, Jacqueline G.; Hakim, Raymond; Kalantar-Zadeh, Kamyar; Zager, Philip; Pisoni, Ronald L.; Port, Friedrich K.; Robinson, Bruce M.

    2015-01-01

    Background and objectives Anemia management changed substantially among dialysis patients in the United States around the time of implementation of the new Centers for Medicare & Medicaid Services bundled payment system and erythropoiesis-stimulating agent (ESA) label change in 2011. Among these, average ferritin levels increased dramatically and have remained high since; this study sought to gain understanding of this sustained rise in ferritin levels. Design, setting, participants, & measurements Trends in mean ferritin, hemoglobin, IV iron dose, and ESA dose from 2009 to 2013 were examined in 9735 patients from 91 United States Dialysis Outcomes and Practice Patterns Study facilities. Linear mixed models were used to assess the extent to which intravenous (IV) iron and ESA dose accounted for patients’ changes in ferritin over time. Results Mean ESA dose and hemoglobin levels declined throughout the study. Mean IV iron dose increased from 210 mg/mo in 2009–2010 to a peak of 280 mg/mo in 2011, then declined back to 200 mg/mo and remained stable from 2012 to 2013. Mean ferritin increased from 601 ng/ml in the third quarter of 2009 to 887 ng/ml in the first quarter of 2012; models suggest that higher IV iron dosing was a primary determinant during 2011, but lower ESA doses contributed to the sustained high ferritin levels thereafter. In a subset of 17 facilities that decreased IV iron dose in 2011, mean ferritin rose by 120 ng/ml to 764 ng/ml, which appeared to be primarily due to ESA reduction. Together, changes in IV iron and ESA doses accounted for 46% of the increase in ferritin over the study period. Conclusions In contrast to expectations, the rise in average IV iron dose did not persist beyond 2011. The sustained rise in ferritin levels in United States dialysis patients after policy changes in 2011, to average levels well in excess of 800 ng/ml, appeared to be partly due to reductions in ESA dosing and not solely IV iron dosing practices. The effect of

  8. Ultrahigh dose-rate FLASH irradiation increases the differential response between normal and tumor tissue in mice.

    PubMed

    Favaudon, Vincent; Caplier, Laura; Monceau, Virginie; Pouzoulet, Frédéric; Sayarath, Mano; Fouillade, Charles; Poupon, Marie-France; Brito, Isabel; Hupé, Philippe; Bourhis, Jean; Hall, Janet; Fontaine, Jean-Jacques; Vozenin, Marie-Catherine

    2014-07-16

    In vitro studies suggested that sub-millisecond pulses of radiation elicit less genomic instability than continuous, protracted irradiation at the same total dose. To determine the potential of ultrahigh dose-rate irradiation in radiotherapy, we investigated lung fibrogenesis in C57BL/6J mice exposed either to short pulses (≤ 500 ms) of radiation delivered at ultrahigh dose rate (≥ 40 Gy/s, FLASH) or to conventional dose-rate irradiation (≤ 0.03 Gy/s, CONV) in single doses. The growth of human HBCx-12A and HEp-2 tumor xenografts in nude mice and syngeneic TC-1 Luc(+) orthotopic lung tumors in C57BL/6J mice was monitored under similar radiation conditions. CONV (15 Gy) triggered lung fibrosis associated with activation of the TGF-β (transforming growth factor-β) cascade, whereas no complications developed after doses of FLASH below 20 Gy for more than 36 weeks after irradiation. FLASH irradiation also spared normal smooth muscle and epithelial cells from acute radiation-induced apoptosis, which could be reinduced by administration of systemic TNF-α (tumor necrosis factor-α) before irradiation. In contrast, FLASH was as efficient as CONV in the repression of tumor growth. Together, these results suggest that FLASH radiotherapy might allow complete eradication of lung tumors and reduce the occurrence and severity of early and late complications affecting normal tissue.

  9. 21 CFR 862.1460 - Leucine aminopeptidase test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... system is a device intended to measure the activity of the enzyme leucine amino-peptidase in serum... diseases such as viral hepatitis and obstructive jaundice. (b) Classification. Class I (general...

  10. Glutathione conjugation dose-dependently increases brain-specific liposomal drug delivery in vitro and in vivo.

    PubMed

    Maussang, David; Rip, Jaap; van Kregten, Joan; van den Heuvel, Angelique; van der Pol, Susanne; van der Boom, Burt; Reijerkerk, Arie; Chen, Linda; de Boer, Marco; Gaillard, Pieter; de Vries, Helga

    2016-06-01

    The blood-brain barrier (BBB) represents a major obstacle for the delivery and development of drugs curing brain pathologies. However, this biological barrier presents numerous endogenous specialized transport systems that can be exploited by engineered nanoparticles to enable drug delivery to the brain. In particular, conjugation of glutathione (GSH) onto PEGylated liposomes (G-Technology(®)) showed to safely enhance delivery of encapsulated drugs to the brain. Yet, understanding of the mechanism of action remains limited and full mechanistic understanding will aid in the further optimization of the technology. In order to elucidate the mechanism of brain targeting by GSH-PEG liposomes, we here demonstrate that the in vivo delivery of liposomal ribavirin is increased in brain extracellular fluid according to the extent of GSH conjugation onto the liposomes. In vitro, using the hCMEC/D3 human cerebral microvascular endothelial (CMEC) cell line, as well as primary bovine and porcine CMEC (and in contrast to non-brain derived endothelial and epithelial cells), we show that liposomal uptake occurs through the process of endocytosis and that the brain-specific uptake is also glutathione conjugation-dependent. Interestingly, the uptake mechanism is an active process that is temperature-, time- and dose-dependent. Finally, early endocytosis events rely on cytoskeleton remodeling, as well as dynamin- and clathrin-dependent endocytosis pathways. Overall, our data demonstrate that the glutathione-dependent uptake mechanism of the G-Technology involves a specific endocytosis pathway indicative of a receptor-mediated mechanism, and supports the benefit of this drug delivery technology for the treatment of devastating brain diseases.

  11. Median infectious dose of human norovirus GII.4 in gnotobiotic pigs is decreased by simvastatin treatment and increased by age

    PubMed Central

    Bui, Tammy; Kocher, Jacob; Li, Yanru; Wen, Ke; Li, Guohua; Liu, Fangning; Yang, Xingdong; LeRoith, Tanya; Tan, Ming; Xia, Ming; Zhong, Weiming; Jiang, Xi

    2013-01-01

    Human noroviruses (NoVs), a major cause of viral gastroenteritis, are difficult to study due to the lack of a cell-culture and a small-animal model. Pigs share with humans the types A and H histo-blood group antigens on the intestinal epithelium and have been suggested as a potential model for studies of NoV pathogenesis, immunity and vaccines. In this study, the effects of age and a cholesterol-lowering drug, simvastatin, on the susceptibility of pigs to NoV infection were evaluated. The median infectious dose (ID50) of a genogroup II, genotype 4 (GII.4) 2006b variant was determined. The ID50 in neonatal (4–5 days of age) pigs was ≤2.74×103 viral RNA copies. In older pigs (33–34 days of age), the ID50 was 6.43×104 but decreased to <2.74×103 in simvastatin-fed older pigs. Evidence of NoV infection was obtained by increased virus load in the intestinal contents, cytopathological changes in the small intestine, including irregular microvilli, necrosis and apoptosis, and detection of viral antigen in the tip of villi in duodenum. This GII.4 variant was isolated in 2008 from a patient from whom a large volume of stool was collected. GII.4 NoVs are continuously subjected to selective pressure by human immunity, and antigenically different GII.4 NoV variants emerge every 1–2 years. The determination of the ID50 of this challenge virus is valuable for evaluation of protection against different GII.4 variants conferred by NoV vaccines in concurrence with other GII.4 variants in the gnotobiotic pig model. PMID:23804568

  12. Low-dose irradiation prior to bone marrow transplantation results in ATM activation and increased lethality in Atm-deficient mice.

    PubMed

    Pietzner, J; Merscher, B M; Baer, P C; Duecker, R P; Eickmeier, O; Fußbroich, D; Bader, P; Del Turco, D; Henschler, R; Zielen, S; Schubert, R

    2016-04-01

    Ataxia telangiectasia is a genetic instability syndrome characterized by neurodegeneration, immunodeficiency, severe bronchial complications, hypersensitivity to radiotherapy and an elevated risk of malignancies. Repopulation with ATM-competent bone marrow-derived cells (BMDCs) significantly prolonged the lifespan and improved the phenotype of Atm-deficient mice. The aim of the present study was to promote BMDC engraftment after bone marrow transplantation using low-dose irradiation (IR) as a co-conditioning strategy. Atm-deficient mice were transplanted with green fluorescent protein-expressing, ATM-positive BMDCs using a clinically relevant non-myeloablative host-conditioning regimen together with TBI (0.2-2.0 Gy). IR significantly improved the engraftment of BMDCs into the bone marrow, blood, spleen and lung in a dose-dependent manner, but not into the cerebellum. However, with increasing doses, IR lethality increased even after low-dose IR. Analysis of the bronchoalveolar lavage fluid and lung histochemistry revealed a significant enhancement in the number of inflammatory cells and oxidative damage. A delay in the resolution of γ-H2AX-expression points to an insufficient double-strand break repair capacity following IR with 0.5 Gy in Atm-deficient splenocytes. Our results demonstrate that even low-dose IR results in ATM activation. In the absence of ATM, low-dose IR leads to increased inflammation, oxidative stress and lethality in the Atm-deficient mouse model.

  13. Dose relations between goal setting, theory-based correlates of goal setting and increases in physical activity during a workplace trial.

    PubMed

    Dishman, Rod K; Vandenberg, Robert J; Motl, Robert W; Wilson, Mark G; DeJoy, David M

    2010-08-01

    The effectiveness of an intervention depends on its dose and on moderators of dose, which usually are not studied. The purpose of the study is to determine whether goal setting and theory-based moderators of goal setting had dose relations with increases in goal-related physical activity during a successful workplace intervention. A group-randomized 12-week intervention that included personal goal setting was implemented in fall 2005, with a multiracial/ethnic sample of employees at 16 geographically diverse worksites. Here, we examined dose-related variables in the cohort of participants (N = 664) from the 8 worksites randomized to the intervention. Participants in the intervention exceeded 9000 daily pedometer steps and 300 weekly minutes of moderate-to-vigorous physical activity (MVPA) during the last 6 weeks of the study, which approximated or exceeded current public health guidelines. Linear growth modeling indicated that participants who set higher goals and sustained higher levels of self-efficacy, commitment and intention about attaining their goals had greater increases in pedometer steps and MVPA. The relation between change in participants' satisfaction with current physical activity and increases in physical activity was mediated by increases in self-set goals. The results show a dose relation of increased physical activity with changes in goal setting, satisfaction, self-efficacy, commitment and intention, consistent with goal-setting theory.

  14. Evaluation of total body weight and body mass index cut-offs for increased cefazolin dose for surgical prophylaxis.

    PubMed

    Hites, Maya; Deprez, Guillaume; Wolff, Fleur; Ickx, Brigitte; Verleije, Anita; Closset, Jean; Loi, Patrizia; Prévost, Jessica; Taccone, Fabio S; Racapé, Judith; Cotton, Frédéric; Jacobs, Frédérique

    2016-12-01

    French and American guidelines recommend increased dosage regimens of cefazolin (CFZ) for surgical prophylaxis in patients with a body mass index (BMI) ≥ 35 kg/m(2) or with a total body weight (TBW) ≥ 120 kg. The objective of this study was to evaluate the accuracy of these cut-offs in identifying patients who require CFZ dose adjustment. A pharmacokinetic study was conducted in patients of varying TBW and BMI who received 2 g of CFZ intravenously for prophylaxis prior to digestive surgery. Adequacy of therapy, defined as a serum concentration of unbound CFZ (fCFZ) ≥ 4 mg/L, was evaluated 180 min (T180) and 240 min (T240) after the start of CFZ infusion. Possible factors associated with insufficient fCFZ levels were also assessed. A P-value of <0.05 was considered statistically significant. A total of 63 patients were included in the study, categorised according to BMI (<35 kg/m(2), 20 patients; and ≥35 kg/m(2), 43 patients) and TBW (<120 kg, 41 patients; and ≥120 kg, 22 patients). All patients had adequate drug levels at T180 but only 40/63 patients (63%) had adequate levels at T240. At T240, therapy was adequate in 15/20 patients (75%) and 25/43 patients (58%) with BMI <35 kg/m(2) and ≥35 kg/m(2), respectively (P = 0.20), and in 28/41 patients (68%) and 12/22 patients (55%) with TBW <120 kg and ≥120 kg, respectively (P = 0.28). No factor associated with insufficient fCFZ was identified. In conclusion, current BMI and TBW cut-offs are poor indicators of which patients could benefit from increased CFZ dosage regimens.

  15. Low-dose growth hormone supplementation increases clinical pregnancy rate in poor responders undergoing in vitro fertilisation.

    PubMed

    Lattes, Karinna; Brassesco, Mario; Gomez, Manuel; Checa, Miguel A

    2015-07-01

    Poor ovarian response (POR) often means low success rates after in vitro fertilisation (IVF). We aim to study the impact of a low-dose growth hormone (GH) supplementation in pregnancy rates in poor responders in a prospective, self-controlled study of 64 poor responders to previous IVF cycles, who failed to achieve pregnancy and were supplemented with low-doses of GH in a subsequent cycle using the same gonadotropin dose and protocol. Our primary endpoint was the clinical pregnancy rate (CPR), considering secondary endpoints, the number of retrieved oocytes, embryos, embryo quality and the proportion of cycles with embryo transfer. CPR in the GH group was 34.4%. Significant differences were observed for the GH group both in the number of top quality embryos (0.64 ± 0.88 versus 1.03 ± 1.17, p < 0.05) and cryopreserved embryos (0.3 ± 0.81 versus 0.85 ± 1.49, p < 0.05). This is, to our knowledge, the first clinical trial to use a low dose of GH as a supplement for IVF in POR patients. Despite this low dose, we achieved excellent success rates in patients with a very poor prognosis, at a reasonable cost and without side effects, which makes this a safe and cost-effective alternative.

  16. Sparing Healthy Tissue and Increasing Tumor Dose Using Bayesian Modeling of Geometric Uncertainties for Planning Target Volume Personalization

    SciTech Connect

    Herschtal, Alan; Te Marvelde, Luc; Mengersen, Kerrie; Foroudi, Farshad; Eade, Thomas; Pham, Daniel; Caine, Hannah; Kron, Tomas

    2015-06-01

    Objective: To develop a mathematical tool that can update a patient's planning target volume (PTV) partway through a course of radiation therapy to more precisely target the tumor for the remainder of treatment and reduce dose to surrounding healthy tissue. Methods and Materials: Daily on-board imaging was used to collect large datasets of displacements for patients undergoing external beam radiation therapy for solid tumors. Bayesian statistical modeling of these geometric uncertainties was used to optimally trade off between displacement data collected from previously treated patients and the progressively accumulating data from a patient currently partway through treatment, to optimally predict future displacements for that patient. These predictions were used to update the PTV position and margin width for the remainder of treatment, such that the clinical target volume (CTV) was more precisely targeted. Results: Software simulation of dose to CTV and normal tissue for 2 real prostate displacement datasets consisting of 146 and 290 patients treated with a minimum of 30 fractions each showed that re-evaluating the PTV position and margin width after 8 treatment fractions reduced healthy tissue dose by 19% and 17%, respectively, while maintaining CTV dose. Conclusion: Incorporating patient-specific displacement patterns from early in a course of treatment allows PTV adaptation for the remainder of treatment. This substantially reduces the dose to healthy tissues and thus can reduce radiation therapy–induced toxicities, improving patient outcomes.

  17. Leucine supplementation chronically improves muscle protein synthesis in older adults consuming the RDA for protein

    PubMed Central

    Casperson, Shanon L.; Sheffield-Moore, Melinda; Hewlings, Susan J.; Paddon-Jones, Douglas

    2013-01-01

    SUMMARY Background & aim Protein-energy supplementation is routinely employed to combat muscle loss. However, success is often compromised by increased satiety, poor palatability, high costs and low compliance. Methods For 2-weeks we supplemented meals of older individuals with leucine (4 g/meal; 3 meals/day; days 2–14). Metabolic studies were performed prior to (Day 1) and following (Day 15) supplementation. Leucine was not provided on metabolic study days. Venous blood and vastus lateralis muscle biopsies were obtained during a primed constant infusion of L-[ring-13C6] phenylalanine. Mixed muscle fractional synthesis rate (FSR), body composition and markers of nutrient signaling (mTOR, 4E-BP1 and p70S6K1 phosphorylation) were measured before and after a low protein/carbohydrate simulated meal. Results The meal modestly increased FSR on Day 1 (postabsorptive: 0.063 ± 0.004 vs. postprandial: 0.075 ± 0.006%/h; p = 0.03), however, two weeks of leucine supplementation increased postabsorptive FSR (p = 0.004) and the response to the meal (p = 0.01) (postabsorptive: 0.074 ± 0.007 vs. postprandial: 0.10 ± 0.007%/h). Changes in FSR were mirrored by increased phosphorylation of mTOR, 4E-BP1 and p70S6K1 (p ≤ 0.1). No change in fat free mass was observed (p > 0.05). Conclusions In older adults, leucine supplementation may improve muscle protein synthesis in response to lower protein meals. PMID:22357161

  18. Radiation Dose Associated with Renal Failure Mortality: A Potential Pathway to Partially Explain Increased Cardiovascular Disease Mortality Observed after Whole-Body Irradiation

    PubMed Central

    Adams, Michael Jacob; Grant, Eric J.; Kodama, Kazunori; Shimizu, Yukiko; Kasagi, Fumiyoshi; Suyama, Akihiko; Sakata, Ritsu; Akahoshi, Masazumi

    2012-01-01

    Whole-body and thoracic ionizing radiation exposure are associated with increased cardiovascular disease (CVD) risk. In atomic bomb survivors, radiation dose is also associated with increased hypertension incidence, suggesting that radiation dose may be associated with chronic renal failure (CRF), thus explaining part of the mechanism for increased CVD. Multivariate Poisson regression was used to evaluate the association of radiation dose with various definitions of chronic kidney disease (CKD) mortality in the Life Span Study (LSS) of atomic bomb survivors. A secondary analysis was performed using a subsample for whom self-reported information on hypertension and diabetes, the two biggest risk factors for CRF, had been collected. We found a significant association between radiation dose and only our broadest definition of CRF among the full cohort. A quadratic dose excess relative risk model [ERR/Gy2 = 0.091 (95% CI: 0.05, 0.198)] fit minimally better than a linear model. Within the subsample, association was also observed only with the broadest CRF definition [ERR/Gy2 = 0.15 (95% CI: 0.02, 0.32)]. Adjustment for hypertension and diabetes improved model fit but did not substantially change the ERR/Gy2 estimate, which was 0.17 (95% CI: 0.04, 0.35). We found a significant quadratic dose relationship between radiation dose and possible chronic renal disease mortality that is similar in shape to that observed between radiation and incidence of hypertension in this population. Our results suggest that renal dysfunction could be part of the mechanism causing increased CVD risk after whole-body irradiation, a hypothesis that deserves further study. PMID:22149958

  19. Fetoplacental deamination and decarboxylation of leucine

    SciTech Connect

    Loy, G.L.; Quick, A.N. Jr.; Hay, W.W. Jr.; Meschia, G.; Battaglia, F.C.; Fennessey, P.V. )

    1990-10-01

    Fetal and placental metabolism of leucine (Leu) and ketoisocaproic acid (KIC) were studied in seven fetal lambs at 132 +/- 1.3-days gestation. Fetal infusions of (1-13C)Leu, (1-14C)Leu, and antipyrine were carried out for 4 h. Uterine and umbilical blood flows were measured using the antipyrine steady-state diffusion technique. Leu and KIC concentrations, (14C)Leu-specific activities, 14CO2, (13C)Leu, and (13C)KIC enrichment (mole percent enrichment) were measured in the maternal artery, uterine vein, and umbilical artery and vein to calculate net fluxes of tracee and tracer molecules between fetus and placenta and between the uteroplacenta and the maternal circulation. There were net Leu and KIC fluxes into the fetus from the placenta with the KIC flux equal to approximately 19% of the combined Leu plus KIC flux. In addition, there was a net KIC flux into the uterine circulation. The fraction of infused tracer Leu escaping the placenta into the mother was small (approximately 6%). By contrast, there was a rapid exchange of tracer Leu carbon between placenta and fetus resulting in a significant flux of labeled KIC from placenta to fetus. Approximately 20% of the infused tracer carbon was converted to CO2 within the fetus. This rate of conversion was greater than 80% of the total fetoplacental conversion rate and significantly higher than the flux of KIC tracer carbon from placenta to fetus. Fetal KIC decarboxylation rate, calculated from the fetal KIC enrichment data, was 2.83 +/- 0.40 mumol.min-1.kg fetus-1 and approximately 60% of the combined net Leu and KIC flux into the fetus from the placenta.

  20. Photodissociation spectroscopy of protonated leucine enkephalin.

    PubMed

    Herburger, Andreas; van der Linde, Christian; Beyer, Martin K

    2017-02-24

    Protonated leucine enkephalin (YGGFL) was studied by ultraviolet photodissociation (UVPD) from 225 to 300 nm utilizing an optical parametric oscillator tunable wavelength laser system (OPO). Fragments were identified by absolute mass measurement in a 9.4 T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS). Bond cleavage was preferred in the vicinity of the two aromatic residues, resulting in high ion abundances for a4, a1, b3, y2 and y1 fragments. a, b and y ions dominated the mass spectrum, and full sequence coverage was achieved for those types. Photodissociation was most effective at the short wavelength end of the studied range, which is assigned to the onset of the La π-π* transition of the tyrosine chromophore, but worked well also at the Lb π-π* chromophore absorption maxima in the 35 000-39 000 cm(-1) region. Several side-chain and internal fragments were observed. H atom loss is observed only above 41 000 cm(-1), consistent with the requirement of a curve crossing to a repulsive (1)πσ* state. It is suggested that the photochemically generated mobile H atom plays a role in further backbone cleavages, similar to the mechanism for electron capture dissociation. The b4 fragment is most intense at the Lb chromophore absorptions, undergoing additional fragmentation at higher photon energies. The high resolution of the FT-ICR MS revealed that out of all x and z-type fragments only x3 and x4 were formed, with low intensity. Other previously reported x- and z-fragments were re-assigned to internal fragments, based on exact mass measurement.

  1. Impact of prolonged leucine supplementation on protein synthesis and lean growth in neonatal pigs.

    PubMed

    Columbus, Daniel A; Steinhoff-Wagner, Julia; Suryawan, Agus; Nguyen, Hanh V; Hernandez-Garcia, Adriana; Fiorotto, Marta L; Davis, Teresa A

    2015-09-15

    Most low-birth weight infants experience extrauterine growth failure due to reduced nutrient intake as a result of feeding intolerance. The objective of this study was to determine whether prolonged enteral leucine supplementation improves lean growth in neonatal pigs fed a restricted protein diet. Neonatal pigs (n = 14-16/diet, 5 days old, 1.8 ± 0.3 kg) were fed by gastric catheter a whey-based milk replacement diet with either a high protein (HP) or restricted protein (RP) content or RP supplemented with leucine to the same level as in the HP diet (RPL). Pigs were fed 40 ml·kg body wt(-1)·meal(-1) every 4 h for 21 days. Feeding the HP diet resulted in greater total body weight and lean body mass compared with RP-fed pigs (P < 0.05). Masses of the longissimus dorsi muscle, heart, and kidneys were greater in the HP- than RP-fed pigs (P < 0.05). Body weight, lean body mass, and masses of the longissimus dorsi, heart, and kidneys in pigs fed the RPL diet were intermediate to RP- and HP-fed pigs. Protein synthesis and mTOR signaling were increased in all muscles with feeding (P < 0.05); leucine supplementation increased mTOR signaling and protein synthesis rate in the longissimus dorsi (P < 0.05). There was no effect of diet on indices of protein degradation signaling in any tissue (P > 0.05). Thus, when protein intake is chronically restricted, the capacity for leucine supplementation to enhance muscle protein accretion in neonatal pigs that are meal-fed milk protein-based diets is limited.

  2. Augmented renal clearance implies a need for increased amoxicillin-clavulanic acid dosing in critically ill children.

    PubMed

    De Cock, Pieter A J G; Standing, Joseph F; Barker, Charlotte I S; de Jaeger, Annick; Dhont, Evelyn; Carlier, Mieke; Verstraete, Alain G; Delanghe, Joris R; Robays, Hugo; De Paepe, Peter

    2015-11-01

    There is little data available to guide amoxicillin-clavulanic acid dosing in critically ill children. The primary objective of this study was to investigate the pharmacokinetics of both compounds in this pediatric subpopulation. Patients admitted to the pediatric intensive care unit (ICU) in whom intravenous amoxicillin-clavulanic acid was indicated (25 to 35 mg/kg of body weight every 6 h) were enrolled. Population pharmacokinetic analysis was conducted, and the clinical outcome was documented. A total of 325 and 151 blood samples were collected from 50 patients (median age, 2.58 years; age range, 1 month to 15 years) treated with amoxicillin and clavulanic acid, respectively. A three-compartment model for amoxicillin and a two-compartment model for clavulanic acid best described the data, in which allometric weight scaling and maturation functions were added a priori to scale for size and age. In addition, plasma cystatin C and concomitant treatment with vasopressors were identified to have a significant influence on amoxicillin clearance. The typical population values of clearance for amoxicillin and clavulanic acid were 17.97 liters/h/70 kg and 12.20 liters/h/70 kg, respectively. In 32% of the treated patients, amoxicillin-clavulanic acid therapy was stopped prematurely due to clinical failure, and the patient was switched to broader-spectrum antibiotic treatment. Monte Carlo simulations demonstrated that four-hourly dosing of 25 mg/kg was required to achieve the therapeutic target for both amoxicillin and clavulanic acid. For patients with augmented renal function, a 1-h infusion was preferable to bolus dosing. Current published dosing regimens result in subtherapeutic concentrations in the early period of sepsis due to augmented renal clearance, which risks clinical failure in critically ill children, and therefore need to be updated. (This study has been registered at Clinicaltrials.gov as an observational study [NCT02456974].).

  3. Augmented Renal Clearance Implies a Need for Increased Amoxicillin-Clavulanic Acid Dosing in Critically Ill Children

    PubMed Central

    Standing, Joseph F.; Barker, Charlotte I. S.; de Jaeger, Annick; Dhont, Evelyn; Carlier, Mieke; Verstraete, Alain G.; Delanghe, Joris R.; Robays, Hugo; De Paepe, Peter

    2015-01-01

    There is little data available to guide amoxicillin-clavulanic acid dosing in critically ill children. The primary objective of this study was to investigate the pharmacokinetics of both compounds in this pediatric subpopulation. Patients admitted to the pediatric intensive care unit (ICU) in whom intravenous amoxicillin-clavulanic acid was indicated (25 to 35 mg/kg of body weight every 6 h) were enrolled. Population pharmacokinetic analysis was conducted, and the clinical outcome was documented. A total of 325 and 151 blood samples were collected from 50 patients (median age, 2.58 years; age range, 1 month to 15 years) treated with amoxicillin and clavulanic acid, respectively. A three-compartment model for amoxicillin and a two-compartment model for clavulanic acid best described the data, in which allometric weight scaling and maturation functions were added a priori to scale for size and age. In addition, plasma cystatin C and concomitant treatment with vasopressors were identified to have a significant influence on amoxicillin clearance. The typical population values of clearance for amoxicillin and clavulanic acid were 17.97 liters/h/70 kg and 12.20 liters/h/70 kg, respectively. In 32% of the treated patients, amoxicillin-clavulanic acid therapy was stopped prematurely due to clinical failure, and the patient was switched to broader-spectrum antibiotic treatment. Monte Carlo simulations demonstrated that four-hourly dosing of 25 mg/kg was required to achieve the therapeutic target for both amoxicillin and clavulanic acid. For patients with augmented renal function, a 1-h infusion was preferable to bolus dosing. Current published dosing regimens result in subtherapeutic concentrations in the early period of sepsis due to augmented renal clearance, which risks clinical failure in critically ill children, and therefore need to be updated. (This study has been registered at Clinicaltrials.gov as an observational study [NCT02456974].) PMID:26349821

  4. Effects of Increasing Doses of Activated Recombinant Factor VII (rFVIIa) on Hemostatic Parameters in Swine

    DTIC Science & Technology

    2005-01-07

    when exposed to por- cine thromboplastin (11, 12). Furthermore, standard in vitro co- agulation tests commonly use as a tissue factor source rabbit...activity in pig blood when measured by in vitro tests than the same dose has in vivo when exposed solely to pig tissue factor. This could compli...on capillary bleed- ing time and in vitro haemostatic parameters using species-spe- cific procedures. The second phase (Injury Phase) was a pilot

  5. Effect of Increasing Doses of Linoleic and α-Linolenic Acids on High-Fructose and High-Fat Diet Induced Metabolic Syndrome in Rats.

    PubMed

    Zhang, Jianan; Wang, Ou; Guo, Yingjian; Wang, Tuo; Wang, Siyi; Li, Guopeng; Ji, Baoping; Deng, Qianchun

    2016-02-03

    Doses and ratio of linoleic acid (LA) and α-linolenic acid (ALA) preventing metabolic syndrome (MS) were investigated. SD rats were fed (i) basal diet, (ii) high-fructose and high-fat diet (HFFD), (iii) HFFD with increasing-dose LA (0.75 energy-% ALA + 3, 6, 9, 12, 15, and 30 energy-% LA), and (iv) HFFD with increasing-dose ALA (6 energy-% LA + 0.3, 0.5, 0.75, 1.5, 2.25, and 3.75 energy-% ALA) for 18 weeks. Results showed 6, 12, 15, and 30 energy-% LA significantly ameliorated central obesity, hyperlipidemia, glucose homeostasis, and leptin status; 0.5 and 0.75 energy-% ALA significantly improved insulin sensitivity, adiponectin, and anti-inflammatory status. Moreover, high intakes of ALA (1.5, 2.25, and 3.75 energy-%) presented a pro-oxidant activity. In conclusion, dose instead of ratio determines the prevention of MS. The optimal doses are 6 energy-% LA and 0.75 energy-% ALA; high intakes of ALA may have side effects.

  6. SU-E-I-82: Improving CT Image Quality for Radiation Therapy Using Iterative Reconstruction Algorithms and Slightly Increasing Imaging Doses

    SciTech Connect

    Noid, G; Chen, G; Tai, A; Li, X

    2014-06-01

    Purpose: Iterative reconstruction (IR) algorithms are developed to improve CT image quality (IQ) by reducing noise without diminishing spatial resolution or contrast. For CT in radiation therapy (RT), slightly increasing imaging dose to improve IQ may be justified if it can substantially enhance structure delineation. The purpose of this study is to investigate and to quantify the IQ enhancement as a result of increasing imaging doses and using IR algorithms. Methods: CT images were acquired for phantoms, built to evaluate IQ metrics including spatial resolution, contrast and noise, with a variety of imaging protocols using a CT scanner (Definition AS Open, Siemens) installed inside a Linac room. Representative patients were scanned once the protocols were optimized. Both phantom and patient scans were reconstructed using the Sinogram Affirmed Iterative Reconstruction (SAFIRE) and the Filtered Back Projection (FBP) methods. IQ metrics of the obtained CTs were compared. Results: IR techniques are demonstrated to preserve spatial resolution as measured by the point spread function and reduce noise in comparison to traditional FBP. Driven by the reduction in noise, the contrast to noise ratio is doubled by adopting the highest SAFIRE strength. As expected, increasing imaging dose reduces noise for both SAFIRE and FBP reconstructions. The contrast to noise increases from 3 to 5 by increasing the dose by a factor of 4. Similar IQ improvement was observed on the CTs for selected patients with pancreas and prostrate cancers. Conclusion: The IR techniques produce a measurable enhancement to CT IQ by reducing the noise. Increasing imaging dose further reduces noise independent of the IR techniques. The improved CT enables more accurate delineation of tumors and/or organs at risk during RT planning and delivery guidance.

  7. ATRAZINE DOES NOT INDUCE GASTROINTESTINAL DISCOMFORT (PICA) IN RATS AT DOSES THAT INCREASE HPA-AXIS ACTIVATION AND CAUSE CONDITIONED TASTE AVERSION.

    EPA Science Inventory

    Previous work has shown that a single oral administration of atrazine (ATR), a chlorotriazine herbicide, induces dose-dependent increases in plasma adrenocorticotropic hormone (ACTH) and serum corticosterone (CORT), with a NOEL equal to 5mg/kg. The mechanism for these effects ...

  8. ATRAZINE DOES NOT INDUCE GASTROINTESTINAL DISCOMFORT (PICA) IN RATS AT DOSES THAT INCREASE ACTH ANDCORTICOSTERONE RELEASE AND CAUSE CONDITIONED TASTE AVERSION.

    EPA Science Inventory

    Previous work has shown that a single oral administration of atrazine (ATR), a chlorotriazine herbicide, induces dose-dependent increases in plasma adrenocorticotropic hormone (ACTH) and serum corticosterone (CORT), with a LOEL of 12.5mg/kg. The mechanism for these effects is unk...

  9. Continuous gamma and neutron irradiation at low doses can increase the number of stromal progenitor cell (CFU-F) in mouse bone marrow

    NASA Astrophysics Data System (ADS)

    Domaratskaya, E. I.; Tsetlin, V. V.; Bueverova, E. I.; Payushina, O. I.; Butorina, N. N.; Khrushchov, N. G.; Starostin, V. I.

    Experimental groups of male and female F1 (CBA × C57Bl/6) mice at the age of 3-4 months were exposed for 10 days to gamma irradiation (total dose 1.5 cGy, dose rate 0.15 cGy/day) or neutron irradiation (neutrons at average energy of 4.5 MeV at a total neutron flux ranging from 10 5 to 10 6 cm -2 and neutron flux density from 1 to 30 cm -2 s -1). These radiation doses were chosen so as to correspond to those received aboard spacecraft. [Mitrikas, V.G., Tsetlin, V.V., 2000. Radiation control onboard the MIR orbital manned station during the 22th solar cycle. Kosm. Issled. 38(2), 113-118.] Gamma irradiation stimulated the proliferation of femoral CFU-F, and their number increased by a factor of 1.5-4.5. The ectopic marrow grafts from γ-irradiated donors also increased in size. However, no changes in CFU-S proliferation rate and their number were observed. Neutron irradiation at a total absorbed dose of 2 × 10 -1 cGy (total neutron flux 2.8 × 10 7 cm -2) produced a 1.5-3-fold increase in the number of femoral CFU-F, but that of CFU-S remained unchanged. At a lower total absorbed dose 0.82 × 10 -2 cGy, total neutron flux 1.3 × 10 6 cm -2, the number of CFU-F remained at the control level. Therefore, the effect of radiation hormesis caused by neutron irradiation was observed at doses much lower than those of gamma irradiation.

  10. The CUG codon is decoded in vivo as serine and not leucine in Candida albicans.

    PubMed Central

    Santos, M A; Tuite, M F

    1995-01-01

    Previous studies have shown that the yeast Candida albicans encodes a unique seryl-tRNA(CAG) that should decode the leucine codon CUG as serine. However, in vitro translation of several different CUG-containing mRNAs in the presence of this unusual seryl-tRNA(CAG) result in an apparent increase in the molecular weight of the encoded polypeptides as judged by SDS-PAGE even though the molecular weight of serine is lower than that of leucine. A possible explanation for this altered electrophoretic mobility is that the CUG codon is decoded as modified serine in vitro. To elucidate the nature of CUG decoding in vivo, a reporter system based on the C. albicans gene (RBP1) encoding rapamycin-binding protein (RBP), coupled to the promoter of the C. albicans TEF3 gene, was utilized. Sequencing and mass-spectrometry analysis of the recombinant RBP expressed in C. albicans demonstrated that the CUG codon was decoded exclusively as serine while the related CUU codon was translated as leucine. A database search revealed that 32 out of the 65 C. albicans gene sequences available have CUG codons in their open reading frames. The CUG-containing genes do not belong to any particular gene family. Thus the amino acid specified by the CUG codon has been reassigned within the mRNAs of C. albicans. We argue here that this unique genetic code change in cellular mRNAs cannot be explained by the 'Codon Reassignment Theory'. Images PMID:7784200

  11. The Effect of Increasing Doses of Saw Palmetto Fruit Extract on Serum PSA Levels: Analysis of the CAMUS Randomized Trial

    PubMed Central

    Andriole, Gerald L.; McCullum-Hill, Christie; Sandhu, Gurdarshan S.; Crawford, E. David; Barry, Michael J.; Cantor, Alan

    2014-01-01

    Purpose Saw palmetto extracts are used for treating lower urinary tract symptoms in men despite level I evidence concluding that saw palmetto was ineffective in reducing lower urinary symptoms. We sought to determine whether higher doses of saw palmetto as studied in CAMUS affect serum PSA levels. Materials and Methods The CAMUS trial was a randomized, placebo-controlled double blind multi-centered North American trial conducted between June 5, 2008 and October 10, 2012 in which 369 men >45 years of age with AUA symptom score ≥ 8 and ≤ 24 were randomly assigned to placebo or dose escalation saw palmetto, which consisted of 320mg for first 24 weeks to 640mg for next 24 weeks to 960mg for last 24 weeks of this 72 week trial. Serum PSA levels (Beckman-Coulter) were obtained at baseline and at weeks 24, 48 and 72 and were compared between treatment groups using the pooled t and Fisher's exact tests. Results Serum PSA levels were similar at baseline for the placebo (1.93 ± 1.59 ng/ml) and saw palmetto groups (2.20 ± 1.95, p = 0.16). Changes in PSA levels over the course of the study were similar: placebo group mean change 0.16 ± 1.08 ng/ml and saw palmetto group mean change 0.23 ± 0.83 ng/ml (p value 0.50). Additionally, no differential effect on serum PSA levels was observed between treatment arms when groups were stratified by baseline PSA values. Conclusions Saw palmetto extract does not affect serum PSA levels more than placebo even at relatively high doses. PMID:23253958

  12. Brassinolide Increases Potato Root Growth In Vitro in a Dose-Dependent Way and Alleviates Salinity Stress

    PubMed Central

    Xia, Shitou; Su, Yi; Wang, Huiqun; Luo, Weigui; Su, Shengying

    2016-01-01

    Brassinosteroids (BRs) are steroidal phytohormones that regulate various physiological processes, such as root development and stress tolerance. In the present study, we showed that brassinolide (BL) affects potato root in vitro growth in a dose-dependent manner. Low BL concentrations (0.1 and 0.01 μg/L) promoted root elongation and lateral root development, whereas high BL concentrations (1–100 μg/L) inhibited root elongation. There was a significant (P < 0.05) positive correlation between root activity and BL concentrations within a range from 0.01 to 100 μg/L, with the peak activity of 8.238 mg TTC·g−1 FW·h−1 at a BL concentration of 100 μg/L. Furthermore, plants treated with 50 μg/L BL showed enhanced salt stress tolerance through in vitro growth. Under this scenario, BL treatment enhanced the proline content and antioxidant enzymes' (superoxide dismutase, peroxidase, and catalase) activity and reduced malondialdehyde content in potato shoots. Application of BL maintain K+ and Na+ homeostasis by improving tissue K+/Na+ ratio. Therefore, we suggested that the effects of BL on root development from stem fragments explants as well as on primary root development are dose-dependent and that BL application alleviates salt stress on potato by improving root activity, root/shoot ratio, and antioxidative capacity in shoots and maintaining K+/Na+ homeostasis in potato shoots and roots. PMID:27803931

  13. The Development of Leucine Dehydrogenase and Formate Dehydrogenase Bifunctional Enzyme Cascade Improves the Biosynthsis of L-tert-Leucine.

    PubMed

    Lu, Jixue; Zhang, Yonghui; Sun, Dongfang; Jiang, Wei; Wang, Shizhen; Fang, Baishan

    2016-11-01

    Leucine dehydrogenase (LDH) and formate dehydrogenase (FDH) were assembled together based on a high-affinity interaction between two different cohesins in a miniscaffoldin and corresponding dockerins in LDH and FDH. The miniscaffoldin with two enzymes was further absorbed by regenerated amorphous cellulose (RAC) to form a bifunctional enzyme complex (miniscaffoldin with LDH and FDH adsorbed by RAC, RSLF) in vitro. The enzymatic characteristics of the bifunctional enzyme complex and free enzymes mixture were systematically compared. The synthesis of L-tert-leucine by the RSLF and free enzyme mixture were compared under different concentrations of enzymes, coenzyme, and substrates. The initial L-tert-leucine production rate by RSLF was enhanced by 2-fold compared with that of the free enzyme mixture. Ninety-one grams per liter of L-tert-leucine with an enantiomeric purity of 99 % e.e. was obtained by RSLF multienzyme catalysis. The results indicated that the bifuntional enzyme complex based on cohesin-dockerin interaction has great potential in the synthesis of L-tert-leucine.

  14. Spectroscopic, thermal and structural studies of new L-leucine and D-leucine complexes with chloranilic acid

    NASA Astrophysics Data System (ADS)

    Pawlukojć, A.; Hetmańczyk, J.; Nowicka-Scheibe, J.; Maurin, Jan K.; Schilf, W.; Rozwadowski, Z.

    2017-04-01

    New molecular complexes of L-leucine and D-leucine with chloranilic acid have been synthesized. Crystal structures of these crystals have been solved; they crystallize in non-centrosymmetric monoclinic P21 space group. In the crystal, leucine molecules exist in protonated form (C6NO2H14+) and chloranilic acid molecules in deprotonated form (C6HCl2O4-). Electronic (UV-Vis) and vibrational absorption (VA) spectra for both materials were collected. Circular dichroism methods such as electronic circular dichroism (ECD) and vibrational circular dichroism (VCD) were used to determine the absolute configuration of new complexes. In both methods a characteristic (-,+) Cotton patterns are observed. In ECD spectra absorption bands are observed at 212 nm for acetonitrile solution and at 202 nm for aqueous solution. VCD spectra (in DMSO-d6 solution) show (-,+) Cotton pattern with strong peaks at 1323 cm-1 (CH rocking mode). Differential scanning calorimetry (DSC) and thermogravimetric (TG) investigations were performed to explore thermal properties of new materials. In DSC curve the decomposition and combustion processes are observed in 220-227 °C. The decomposition process was described by use of TG method and quadruple mass spectrometer (QMS). NMR spectra of pure L-leucine and chloranilic acid as well as L-leucine - chloranilic acid complex in solutions (D2O and DMSO) and in solid state confirm the geometry of molecules in complex both in solution and in solid state.

  15. Ibandronate dose response is associated with increases in bone mineral density and reductions in clinical fractures: results of a meta-analysis.

    PubMed

    Sebba, Anthony I; Emkey, Ronald D; Kohles, Joseph D; Sambrook, Philip N

    2009-03-01

    This meta-analysis pooled data from the four phase III clinical trials of ibandronate to assess the relationship between ibandronate dose, changes in bone mineral density, and rates of both clinical and non-vertebral fractures. Individual patient data from the intent-to-treat population of the BONE, IV fracture prevention, MOBILE, and DIVA studies were included for analysis. The relationship between ibandronate dose and bone mineral density at both the lumbar spine and at the total hip was assessed qualitatively. The relationship between lumbar spine bone mineral density and clinical fracture rate, and the relationship between total hip bone mineral density and non-vertebral fracture rate, were assessed both qualitatively and using mathematical models. A total of 8710 patients were included in this analysis. Both lumbar spine and total hip bone mineral density were observed to increase with increasing ibandronate dose. The incidence of all clinical fractures was observed to decrease as lumbar spine bone mineral density increased. A statistically significant inverse linear relationship was observed between percent change in lumbar spine bone mineral density and the rate of clinical fractures (P=0.005). A non-significant curvilinear relationship was observed between percent change in total hip bone mineral density and non-vertebral fracture rate. Increased ibandronate exposure is associated with increasing gains in the lumbar spine bone mineral density and decreasing clinical fracture rates. A non-linear relationship may exist between increases in the total hip bone mineral density and non-vertebral fracture rate.

  16. Maternal post-absorptive leucine kinetics during late pregnancy in US women with HIV taking antiretroviral therapy: a cross-sectional pilot study

    PubMed Central

    Cade, W. Todd; Singh, Gautam K.; Holland, Mark R.; Reeds, Dominic N.; Overton, E. Turner; Cibulka, Nancy; Bahow, Karen; Presti, Rachel; Stephens, Andrea; Cahill, Alison G.

    2015-01-01

    Background Despite the success of combination antiretroviral therapy (cART) for the prevention of mother to child transmission of HIV, infants exposed to cART in utero frequently are born smaller and have mild cardiac abnormalities. The mechanisms responsible for lower birth weight and cardiac abnormalities in children exposed to cART are unclear but could be related to dysregulation of maternal amino acid metabolism during pregnancy. Previous data in HIV(−) women have shown a relationship between abnormal maternal protein metabolism during pregnancy and low infant birth weight and animal data demonstrate a relationship between altered maternal protein metabolism and increased risk for offspring cardiovascular abnormalities. Objective The objectives of this study were to: characterize post-absorptive maternal leucine kinetics during late pregnancy and examine the relationships between maternal leucine kinetics and offspring birth weight and cardiac function. Design Post-absorptive maternal leucine kinetics (evaluated by using stable isotope tracer methodology) in 16 HIV(+) women receiving cART and 14 HIV(−) US women during the 3rd trimester of pregnancy were compared. Relationships between post-absorptive maternal leucine kinetics, cardiac function (echocardiography) and birth weight were statistically examined. Results Maternal plasma leucine concentration (HIV(−): 82.8 ± 10.7 vs. HIV(+): 72.3 ± 13.5 μM, p=0.06) and leucine oxidation rate (HIV(−): 6.1 ± 1.6 vs. HIV(+): 4.9 ± 1.8 μmol/kgBW/min, p=0.03) were lower in HIV+ women compared to controls. Total leucine turnover rate, non-oxidative leucine disposal rate and post-absorptive maternal glucose and palmitate kinetics did not differ between groups. Left ventricular fractional shortening tended to be lower in children born to HIV(+) compared to controls (HIV(−): 42 ± 1 vs. HIV+: 36 ± 5 %, p=0.08) and associated with lower maternal plasma leucine concentration (r= 0.43, p=0.08). Conclusions

  17. Gamma- and neutron continuous irradiations at low doses can increase stromal progenitor cell (cfu-f) number in mouse bone marrow

    NASA Astrophysics Data System (ADS)

    Domaratskaya, E.; Tsetlin, V.; Bueverova, E.; Payushina, O.; Butorina, N.; Starostin, V.

    Low doses of continuous gamma and neutron irradiation chosen in these experiments corresponded to those aboard a spacecraft (Mitricas, Tsetlin, 2000). F1 (CBAxC57Bl/6) male and female mice at the age of 3-4 months were used. The experimental groups of mice were exposed for 10 days to gamma irradiation (total dose 1.5 cGy, dose rate 0.15 cGy/day) or neutron irradiation (neutrons with energy of 4 MeV at flow in the range from 10-5 to 10-6 n/cm2, flow densities from 1 to 30 n/cm2sec). Gamma irradiation stimulated the proliferative rate of femoral CFU-F and raised their number 1,5-4,5-fold. The size of ectopic marrow transplants from gamma irradiated donors also increased. However, no changes in CFU-S proliferative rate and their number were observed. Neutron irradiation at total absorbed dose of 48x10-3 cGy (total neutron flow 2,8x106 n/cm2) produced a 3-fold increase of femoral CFU-F number, but CFU-S number remained unchanged. If total absorbed dose was lowered to 7x10-3 cGy (total neutron flow 1,3x105 n/cm2) CFU-F number remained at the control level. Therefore, the effect of radiation hormesis that caused by the neutron irradiation was observed at doses much lower than those of gamma irradiation. Supported in part by Russian Ministry of Education (projects ``Scientific Schools'' - 1629.2003.4).

  18. Leucine facilitates the insulin-stimulated glucose uptake and insulin signaling in skeletal muscle cells: involving mTORC1 and mTORC2.

    PubMed

    Liu, Hui; Liu, Rui; Xiong, Yufang; Li, Xiang; Wang, Xiaolei; Ma, Yan; Guo, Huailan; Hao, Liping; Yao, Ping; Liu, Liegang; Wang, Di; Yang, Xuefeng

    2014-08-01

    Leucine, a branched-chain amino acid, has been shown to promote glucose uptake and increase insulin sensitivity in skeletal muscle, but the exact mechanism remains unestablished. We addressed this issue in cultured skeletal muscle cells in this study. Our results showed that leucine alone did not have an effect on glucose uptake or phosphorylation of protein kinase B (AKT), but facilitated the insulin-induced glucose uptake and AKT phosphorylation. The insulin-stimulated glucose uptake and AKT phosphorylation were inhibited by the phosphatidylinositol 3-kinase inhibitor, wortmannin, but the inhibition was partially reversed by leucine. The inhibitor of mammalian target of rapamycin complex 1 (mTORC1), rapamycin, had no effect on the insulin-stimulated glucose uptake, but eliminated the facilitating effect of leucine in the insulin-stimulated glucose uptake and AKT phosphorylation. In addition, leucine facilitation of the insulin-induced AKT phosphorylation was neutralized by knocking down the core component of the mammalian target of rapamycin complex 2 (mTORC2) with specific siRNA. Together, these findings show that leucine can facilitate the insulin-induced insulin signaling and glucose uptake in skeletal muscle cells through both mTORC1 and mTORC2, implicating the potential importance of this amino acid in glucose homeostasis and providing new mechanistic insights.

  19. Modification of the 3H-leucine Incorporation Technique for Quantifying Rates of Bacterial Secondary Production on Decaying Wetland Plant Litter: Effectiveness of Microdialysis.

    NASA Astrophysics Data System (ADS)

    Gillies, J. E.; Francoeur, S. N.; Kuehn, K. A.

    2005-05-01

    The radiolabelled 3H-leucine incorporation technique for quantifying rates of bacterial production has increased in popularity since its original description for bacterioplankton communities. Prior studies addressing incorporation conditions (e.g., substrate saturation) for bacterial communities in other habitats, such as decaying plant litter, have reported a wide range of final leucine concentrations (400nM to 50,000nM) to achieve saturation-level uptake. We assessed the application of the 3H-leucine incorporation procedure for measuring bacterial production on decaying wetland plant litter. Substrate saturation experiments (9 concentrations, 10nM to 50,000nM final leucine) were conducted for bacterial communities colonizing submerged litter of three emergent plant species (Typha angustifolia, Schoenoplectus validus, and Phragmites australis). A modified 3H-leucine protocol was developed by coupling previously described incubation and extraction protocols with microdialysis (500MWCO) of the final radiolabelled protein extract. Incorporation of 3H-leucine into protein exhibited a biphasic saturation curve, with lower Km values ranging from 400nM to 1200nM depending on the plant species studied. Upper Km values ranged from 4000nM to 6000nM. Dialysis of the crude protein extract significantly improved counting precision and the signal-to-noise ratio. These results suggest differential uptake by litter associated microbial assemblages, with lower Km values possibly representing bacterial uptake and higher Km values representing non-bacterial uptake.

  20. Leucine disposal rate for assessment of amino acid metabolism in maintenance hemodialysis patients

    PubMed Central

    Denny, Gerald B.; Deger, Serpil M.; Chen, Guanhua; Bian, Aihua; Sha, Feng; Booker, Cindy; Kesler, Jaclyn T.; David, Sthuthi; Ellis, Charles D.; Ikizler, T. Alp

    2016-01-01

    Background Protein energy wasting (PEW) is common in patients undergoing maintenance hemodialysis (MHD) and closely associated with poor outcomes. Insulin resistance and associated alterations in amino acid metabolism are potential pathways leading to PEW. We hypothesized that the measurement of leucine disposal during a hyperinsulinemic- euglycemic-euaminoacidemic clamp (HEAC) procedure would accurately measure the sensitivity to insulin for its actions on concomitant carbohydrate and protein metabolism in MHD patients. Methods We examined 35 MHD patients and 17 control subjects with normal kidney function by hyperinsulinemic-euglycemic clamp (HEGC) followed by HEAC clamp procedure to obtain leucine disposal rate (LDR) along with isotope tracer methodology to assess whole body protein turnover. Results The glucose disposal rate (GDR) by HEGC was 5.1 ± 2.1 mg/kg/min for the MHD patients compared to 6.3 ± 3.9 mg/kg/min for the controls (p = 0.38). The LDR during HEAC was 0.09 ± 0.03 mg/kg/min for the MHD patients compared to 0.11 ± 0.05 mg/kg/min for the controls (p = 0.009). The LDR level was correlated with whole body protein synthesis (r = 0.25; p = 0.08), with whole body protein breakdown (r = −0.38 p = 0.01) and net protein balance (r = 0.85; p < 0.001) in the overall study population. Correlations remained significant in subgroup analysis. The GDR derived by HEGC and LDR correlated well in the controls (r = 0.79, p < 0.001), but less so in the MHD patients (r = 0.58, p < 0.001). Conclusions Leucine disposal rate reliably measures amino acid utilization in MHD patients and controls in response to high dose insulin. PMID:27413537

  1. The radiolysis and racemization of leucine on proton irradiation

    NASA Technical Reports Server (NTRS)

    Bonner, W. A.; Lemmon, R. M.; Conzett, H. E.

    1982-01-01

    D- and L-Leucine have been subjected to 39-55 percent radiolysis using 0.11 MeV protons, both with the proton beam passing through the sample or being absorbed by it and with quenching the sample immediately on completion of irradiation or after a 21-day interval. Racemization was small (1.1-1.7 percent) and comparable in all cases, suggesting that radioracemization and secondary degradative effects were not important factors in the recent unsuccessful attempts to induce optical activity in DL-Leucine by partial radiolysis using 0-11 MeV longitudinally polarized protons.

  2. Evidence that increased Kcnj6 gene dose is necessary for deficits in behavior and dentate gyrus synaptic plasticity in the Ts65Dn mouse model of Down syndrome.

    PubMed

    Kleschevnikov, Alexander M; Yu, Jessica; Kim, Jeesun; Lysenko, Larisa V; Zeng, Zheng; Yu, Y Eugene; Mobley, William C

    2017-03-22

    Down syndrome (DS), trisomy 21, is caused by increased dose of genes present on human chromosome 21 (HSA21). The gene-dose hypothesis argues that a change in the dose of individual genes or regulatory sequences on HSA21 is necessary for creating DS-related phenotypes, including cognitive impairment. We focused on a possible role for Kcnj6, the gene encoding Kir3.2 (Girk2) subunits of a G-protein-coupled inwardly-rectifying potassium channel. This gene resides on a segment of mouse Chromosome 16 that is present in one extra copy in the genome of the Ts65Dn mouse, a well-studied genetic model of DS. Kir3.2 subunit-containing potassium channels serve as effectors for a number of postsynaptic metabotropic receptors including GABAB receptors. Several studies raise the possibility that increased Kcnj6 dose contributes to synaptic and cognitive abnormalities in DS. To assess directly a role for Kcnj6 gene dose in cognitive deficits in DS, we produced Ts65Dn mice that harbor only 2 copies of Kcnj6 (Ts65Dn:Kcnj6++- mice). The reduction in Kcnj6 gene dose restored to normal the hippocampal level of Kir3.2. Long-term memory, examined in the novel object recognition test with the retention period of 24h, was improved to the level observed in the normosomic littermate control mice (2N:Kcnj6++). Significantly, both short-term and long-term potentiation (STP and LTP) was improved to control levels in the dentate gyrus (DG) of the Ts65Dn:Kcnj6++- mouse. In view of the ability of fluoxetine to suppress Kir3.2 channels, we asked if fluoxetine-treated DG slices of Ts65Dn:Kcnj6+++ mice would rescue synaptic plasticity. Fluoxetine increased STP and LTP to control levels. These results are evidence that increased Kcnj6 gene dose is necessary for synaptic and cognitive dysfunction in the Ts65Dn mouse model of DS. Strategies aimed at pharmacologically reducing channel function should be explored for enhancing cognition in DS.

  3. Differential responses of the incretin hormones GIP and GLP-1 to increasing doses of dietary carbohydrate but not dietary protein in lean rats

    PubMed Central

    Yang, Qing; Kindel, Tammy L.; Tso, Patrick

    2010-01-01

    Previous studies have shown that oral ingestion of nutrients stimulates secretion of the incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1); however, it is unclear whether there is a dose-dependent response between the amount of nutrient ingested and the secretion of the hormones in vivo. Using our lymph fistula rat model, we previously demonstrated that both GIP and GLP-1 responded dose dependently to increasing amounts of infused dietary lipid and that the GLP-1-secreting cells were more sensitive to changes in intestinal lipid content. In the present study, we investigated the dose-dependent relationships between incretin secretion and the two remaining macronutrients, carbohydrate and protein. To accomplish this objective, the major mesenteric lymphatic duct of male Sprague-Dawley rats was cannulated. Each animal received a single bolus (3 ml) of saline, dextrin, whey protein, or casein hydrolysate (0.275, 0.55, 1.1, 2.2, 4.4 kcal) via a surgically inserted duodenal or ileal feeding tube. Lymph was continuously collected for 3 h and analyzed for GIP and GLP-1 content. Both GIP and GLP-1 outputs responded dose dependently to increasing amounts of dietary carbohydrate but not protein. Additionally, we found that the GIP-secreting cells were more sensitive than the GLP-1-secreting cells to changes in intestinal carbohydrate content. PMID:20522638

  4. High-dose green tea polyphenol intake decreases CYP3A expression in a liver-specific manner with increases in blood substrate drug concentrations.

    PubMed

    Ikarashi, Nobutomo; Ogawa, Sosuke; Hirobe, Ryuta; Kusunoki, Yoshiki; Kon, Risako; Ochiai, Wataru; Sugiyama, Kiyoshi

    2016-06-30

    In recent years, the intake of functional foods containing high-doses of green tea polyphenols (GP) has been increasing. In this study, the long-term safety of high-dose GP was assessed from a pharmacokinetic point of view by focusing on the drug-metabolizing enzyme, cytochrome P450 (CYP). Mice were fed a diet containing 3% GP for 4weeks, and the CYP expression levels and activity were determined. The GP-treated group showed a significant decrease in the hepatic CYP3A and an increase in the hepatic CYP2C expression compared with the control group. CYP1A, CYP2D, and CYP2E expression were not different between the GP-treated and the control groups. In the small intestine, there were no differences in the CYP3A protein levels between the groups. The increase in the plasma triazolam concentration in the GP-treated group was observed. Although no changes were found in the hepatic CYP3A levels in mice receiving a diet containing 0.1% GP for 4weeks, a significant decrease was seen in the hepatic CYP3A level in mice receiving a diet containing 3% GP for only 1week. This study revealed that the intake of a high-dose GP results in a liver-specific decrease in the CYP3A expression level. The results also indicated that the effects of GP on CYP3A were not observed following the intake of a low-dose GP. In the future, caution should be taken in cases when functional foods containing a high-dose GP are concomitantly consumed with a CYP3A substrate drug.

  5. Add-on LABA in a separate inhaler as asthma step-up therapy versus increased dose of ICS or ICS/LABA combination inhaler

    PubMed Central

    Colice, Gene; Israel, Elliot; Roche, Nicolas; Postma, Dirkje S.; Guilbert, Theresa W.; van Aalderen, Willem M.C.; Grigg, Jonathan; Hillyer, Elizabeth V.; Thomas, Victoria; Martin, Richard J.

    2016-01-01

    Asthma management guidelines recommend adding a long-acting β2-agonist (LABA) or increasing the dose of inhaled corticosteroid (ICS) as step-up therapy for patients with uncontrolled asthma on ICS monotherapy. However, it is uncertain which option works best, which ICS particle size is most effective, and whether LABA should be administered by separate or combination inhalers. This historical, matched cohort study compared asthma-related outcomes for patients (aged 12–80 years) prescribed step-up therapy as a ≥50% extrafine ICS dose increase or add-on LABA, via either a separate inhaler or a fine-particle ICS/LABA fixed-dose combination (FDC) inhaler. Risk-domain asthma control was the primary end-point in comparisons of cohorts matched for asthma severity and control during the baseline year. After 1:2 cohort matching, the increased extrafine ICS versus separate ICS+LABA cohorts included 3232 and 6464 patients, respectively, and the fine-particle ICS/LABA FDC versus separate ICS+LABA cohorts included 7529 and 15 058 patients, respectively (overall mean age 42 years; 61–62% females). Over one outcome year, adjusted OR (95% CI) for achieving asthma control were 1.25 (1.13–1.38) for increased ICS versus separate ICS+LABA and 1.06 (1.05–1.09) for ICS/LABA FDC versus separate ICS+LABA. For patients with asthma, increased dose of extrafine-particle ICS, or add-on LABA via ICS/LABA combination inhaler, is associated with significantly better outcomes than ICS+LABA via separate inhalers. PMID:27730200

  6. Low-dose rapamycin treatment increases the ability of human regulatory T cells to inhibit transplant arteriosclerosis in vivo.

    PubMed

    Hester, J; Schiopu, A; Nadig, S N; Wood, K J

    2012-08-01

    Regulatory T cells (T(reg)) are currently being tested in clinical trials as a potential therapy in cell and solid organ transplantation. The immunosuppressive drug rapamycin has been shown to preferentially promote T(reg) expansion. Here, we hypothesized that adjunctive rapamycin therapy might potentiate the ability of ex vivo expanded human T(reg) to inhibit vascular allograft rejection in a humanized mouse model of arterial transplantation. We studied the influence of combined treatment with low-dose rapamycin and subtherapeutic T(reg) numbers on the development of transplant arteriosclerosis (TA) in human arterial grafts transplanted into immunodeficient BALB/cRag2(-/-) Il2rg(-/-) mice reconstituted with allogeneic human peripheral blood mononuclear cell. In addition, we assessed the effects of the treatment on the proliferation and apoptosis of naïve/effector T cells. The combined therapy efficiently suppressed T-cell proliferation in vivo and in vitro. Neointima formation in the human arterial allografts was potently inhibited compared with each treatment alone. Interestingly, CD4(+) but not CD8(+) T lymphocytes were sensitive to T(reg) and rapamycin-induced apoptosis in vitro. Our data support the concept that rapamycin can be used as an adjunctive therapy to improve efficacy of T(reg)-based immunosuppressive protocols in clinical practice. By inhibiting TA, T(reg) and rapamycin may prevent chronic transplant dysfunction and improve long-term allograft survival.

  7. Exposure to low UVA doses increases KatA and KatB catalase activities, and confers cross-protection against subsequent oxidative injuries in Pseudomonas aeruginosa.

    PubMed

    Pezzoni, Magdalena; Tribelli, Paula M; Pizarro, Ramón A; López, Nancy I; Costa, Cristina S

    2016-05-01

    Solar UVA radiation is one of the main environmental stress factors for Pseudomonas aeruginosa. Exposure to high UVA doses produces lethal effects by the action of the reactive oxygen species (ROS) it generates. P. aeruginosa has several enzymes, including KatA and KatB catalases, which provide detoxification of ROS. We have previously demonstrated that KatA is essential in defending P. aeruginosa against high UVA doses. In order to analyse the mechanisms involved in the adaptation of this micro-organism to UVA, we investigated the effect of exposure to low UVA doses on KatA and KatB activities, and the physiological consequences. Exposure to UVA induced total catalase activity; assays with non-denaturing polyacrylamide gels showed that both KatA and KatB activities were increased by radiation. This regulation occurred at the transcriptional level and depended, at least partly, on the increase in H2O2 levels. We demonstrated that exposure to low UVA produced a protective effect against subsequent lethal doses of UVA, sodium hypochlorite and H2O2. Protection against lethal UVA depends on katA, whilst protection against sodium hypochlorite depends on katB, demonstrating that different mechanisms are involved in the defence against these oxidative agents, although both genes can be involved in the global cellular response. Conversely, protection against lethal doses of H2O2 could depend on induction of both genes and/or (an)other defensive factor(s). A better understanding of the adaptive response of P. aeruginosa to UVA is relevant from an ecological standpoint and for improving disinfection strategies that employ UVA or solar irradiation.

  8. Comparison of Combined Tofogliflozin and Glargine, Tofogliflozin Added to Insulin, and Insulin Dose-Increase Therapy in Uncontrolled Type 2 Diabetes

    PubMed Central

    Suzuki, Katsunori; Mitsuma, Yurie; Sato, Takaaki; Anraku, Takumi; Hatta, Mariko

    2016-01-01

    Background Some patients with type 2 diabetes mellitus (T2DM) on insulin have poor glycemic control and require add-on therapy to reach target glucose values. Increased insulin doses or the addition of an oral antidiabetic drug (OAD) may improve glycemic control, but many patients fail to achieve target values. The aim of this study was to compare the treatment efficacy and safety of three different therapies in such patients. Methods T2DM outpatients with poor glycemic control (HbA1c ≥ 7.0%) despite insulin therapy (including patients on OADs other than a sodium-glucose cotransporter 2 (SGLT2) inhibitor) were included. The patients had a body mass index (BMI) of ≥ 22 kg/m2 and an estimated glomerular filtration rate (eGFR) of ≥ 45 mL/min/1.73 m2, did not have depletion of endogenous insulin, and had stable glucose levels for 3 months before study entry on insulin therapy. Treatment was continued for 24 weeks with insulin dose-increase therapy, tofogliflozin add-on therapy, or a combination of insulin glargine + tofogliflozin. The primary endpoints were HbA1c, weight, and total insulin dose. Secondary endpoints included fasting plasma glucose (FPG), blood pressure, lipid profiles, and incidence of adverse events. Results At baseline, the participants’ median age was 59.0 years, mean BMI was 28.7 kg/m2, mean eGFR was 89.2 mL/min/1.73 m2, mean HbA1c was 8.7%, and mean FPG was 174.1 mg/dL. The mean duration of insulin therapy was approximately 7 years. The mean daily insulin dose was approximately 40 U in the three groups. Overall, 85% received other background OADs in addition to insulin. Over the 24-week period, HbA1c in the insulin group decreased slightly initially and then plateaued; daily total insulin dose and weight increased, and blood pressure increased slightly. In the insulin + tofogliflozin group and the glargine + tofogliflozin group, HbA1c decreased greatly initially, and this continued over the 24-week period, with HbA1c decreases of -1.0% and

  9. Enteral leucine and protein synthesis in skeletal and cardiac muscle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There are three members of the Branch Chain Amino Acids: leucine, isoleucine, and valine. As essential amino acids, these amino acids have important functions which include a primary role in protein structure and metabolism. It is intriguing that the requirement for BCAA in humans comprise about 40–...

  10. The apo-structure of the leucine sensor Sestrin2 is still elusive

    PubMed Central

    Saxton, Robert A.; Knockenhauer, Kevin E.; Schwartz, Thomas U.; Sabatini, David M.

    2016-01-01

    Sestrin2 is a GATOR2 interacting protein that directly binds leucine and is required for the inhibition of mTORC1 under leucine deprivation, indicating that it is a leucine sensor for the mTORC1 pathway. We recently reported the structure of Sestrin2 in complex with leucine (PDB ID: 5DJ4), and demonstrated that mutations in the leucine-binding pocket alter the affinity of Sestrin2 for leucine and result in a corresponding change in the leucine sensitivity of mTORC1 in cells. A lower resolution structure of human Sestrin2 (PDB ID: 5CUF), which was crystallized in the absence of exogenous leucine, showed Sestrin2 to be in a nearly identical conformation as the leucine-bound structure. Based on this observation, it has been argued that leucine binding does not affect the conformation of Sestrin2 and thus that Sestrin2 may not be a sensor for leucine. Here, we show that simple analysis of the reported “apo”-Sestrin2 structure reveals the clear presence of prominent, unmodeled electron density in the leucine-binding pocket that exactly accommodates the leucine observed in the higher resolution structure. Refining the reported “apo”-structure with leucine eliminates the large FO-FC difference density at this position and improves the working and free R-factors of the model. Consistent with this, our own structure of Sestrin2 crystallized in the absence of exogenous leucine also contains electron density that is best explained by leucine. Thus, the structure of apo-Sestrin2 remains elusive. PMID:27649739

  11. Automatic tube current modulation for whole-body polytrauma CT with immobilization devices: is there an increase in radiation dose and degradation of image quality?

    PubMed

    Euler, André; Stieltjes, Bram; Schindera, Sebastian T

    2017-02-01

    The objective of this study was the assessment of the image quality and radiation dose in polytrauma CT using immobilization devices. An anthropomorphic whole body and a liver phantom were scanned on a 128-slice CT scanner with four different protocols using automatic tube current modulation (120 kVp, 150 ref. mAs; 120 kV, 200 ref. mAs; 140 kVp, 150 ref. mAs; and 140 kVp, 200 ref. mAs) and four different setups (no immobilization device (setup A), vacuum mattress 1 (setup B), vacuum mattress 2 (setup C), and spineboard (setup D)). Qualitative and quantitative image quality parameters and radiation dose were assessed. Image noise increased on average by 6.6, 11.2, and 9.4 %, and CNR decreased by 11.2, 13.9, and 6.5 for setups B, C, and D, respectively, compared with setup A. The CTDIvol increased up to 6 % using immobilization devices. Severe streak artifacts, provoked by the inflation valve of the mattresses were detected at the level of the head and shoulder. Applying immobilization devices for whole-body CT with automatic tube current modulation increases the radiation dose and decreases the quantitative image quality slightly. Severe artifacts, induced by the inflation valve of the mattress, can influence the diagnostic accuracy at the level of the head and shoulder.

  12. Immunization of teenagers with a fifth dose of reduced DTaP-IPV induces high levels of pertussis antibodies with a significant increase in opsonophagocytic activity.

    PubMed

    Aase, Audun; Herstad, Tove Karin; Merino, Samuel; Bolstad, Merete; Sandbu, Synne; Bakke, Hilde; Aaberge, Ingeborg S

    2011-08-01

    Waning vaccine-induced immunity against Bordetella pertussis is observed among adolescents and adults. A high incidence of pertussis has been reported in this population, which serves as a reservoir for B. pertussis. A fifth dose of reduced antigen of diphtheria-tetanus-acellular-pertussis and inactivated polio vaccine was given as a booster dose to healthy teenagers. The antibody activity against B. pertussis antigens was measured prior to and 4 to 8 weeks after the booster by different assays: enzyme-linked immunosorbent assays (ELISAs) of IgG and IgA against pertussis toxin (PT) and filamentous hemagglutinin (FHA), IgG against pertactin (PRN), opsonophagocytic activity (OPA), and IgG binding to live B. pertussis. There was a significant increase in the IgG activity against PT, FHA, and PRN following the booster immunization (P < 0.001). The prebooster sera showed a geometric mean OPA titer of 65.1 and IgG binding to live bacteria at a geometric mean concentration of 164.9 arbitrary units (AU)/ml. Following the fifth dose, the OPA increased to a titer of 360.4, and the IgG concentration against live bacteria increased to 833.4 AU/ml (P < 0.001 for both). The correlation analyses between the different assays suggest that antibodies against FHA and PRN contribute the most to the OPA and IgG binding.

  13. Phosphorylation of hepatic AMP-activated protein kinase and liver kinase B1 is increased after a single oral dose of green tea extract to mice.

    PubMed

    Banerjee, Subhashis; Ghoshal, Sarbani; Porter, Todd D

    2012-12-01

    We have previously shown that green and black tea extracts increase the phosphorylation of AMP-activated protein kinase (AMPK) and HMG-CoA reductase in rat hepatoma cells in culture, concomitant with a decrease in cholesterol synthesis. In the present study, we evaluated the ability of a single oral dose of green or black tea extract to promote the phosphorylation of AMPK, liver kinase B1 (LKB1, an AMPK-kinase), and HMG-CoA reductase in mouse liver. Green tea extract administered by gavage at 50 and 100 mg/kg caused a 2- to 3-fold increase in hepatic AMPK phosphorylation at 3 and 6 hours after dosing and a 1.5- to 2-fold increase in LKB1 phosphorylation at these same time points. The phosphorylation of HMG-CoA reductase at these and later time points was not significantly increased. Black tea administered by gavage at up to 250 mg/kg was ineffective in increasing hepatic AMPK phosphorylation. Both green and black tea extracts increased LKB1 phosphorylation in hepatoma cells in culture at 15 μg/mL, and black tea also increased the phosphorylation of protein kinase A in hepatoma cells. These results suggest that compounds in both tea extracts activate AMPK by activating its upstream kinase, LKB1, and that black tea may do so by first activating protein kinase A, a known kinase for LKB1. Only green tea, at 50 and 100 mg/kg, was able to activate AMPK and LKB1 in mouse liver after oral dosing, suggesting that the polymerized catechins present in black tea do not reach the liver in sufficient concentration to affect AMPK activity.

  14. Cerebral pathological and compensatory mechanisms in the premotor phase of leucine-rich repeat kinase 2 parkinsonism.

    PubMed

    van Nuenen, Bart F L; Helmich, Rick C; Ferraye, Murielle; Thaler, Avner; Hendler, Talma; Orr-Urtreger, Avi; Mirelman, Anat; Bressman, Susan; Marder, Karen S; Giladi, Nir; van de Warrenburg, Bart P C; Bloem, Bastiaan R; Toni, Ivan

    2012-12-01

    Compensatory cerebral mechanisms can delay motor symptom onset in Parkinson's disease. We aim to characterize these compensatory mechanisms and early disease-related changes by quantifying movement-related cerebral function in subjects at significantly increased risk of developing Parkinson's disease, namely carriers of a leucine-rich repeat kinase 2-G2019S mutation associated with dominantly inherited parkinsonism. Functional magnetic resonance imaging was used to examine cerebral activity evoked during internal selection of motor representations, a core motor deficit in clinically overt Parkinson's disease. Thirty-nine healthy first-degree relatives of Ashkenazi Jewish patients with Parkinson's disease, who carry the leucine-rich repeat kinase 2-G2019S mutation, participated in this study. Twenty-one carriers of the leucine-rich repeat kinase 2-G2019S mutation and 18 non-carriers of this mutation were engaged in a motor imagery task (laterality judgements of left or right hands) known to be sensitive to motor control parameters. Behavioural performance of both groups was matched. Mutation carriers and non-carriers were equally sensitive to the extent and biomechanical constraints of the imagined movements in relation to the current posture of the participants' hands. Cerebral activity differed between groups, such that leucine-rich repeat kinase 2-G2019S carriers had reduced imagery-related activity in the right caudate nucleus and increased activity in the right dorsal premotor cortex. More severe striatal impairment was associated with stronger effective connectivity between the right dorsal premotor cortex and the right extrastriate body area. These findings suggest that altered movement-related activity in the caudate nuclei of leucine-rich repeat kinase 2-G2019S carriers might remain behaviourally latent by virtue of cortical compensatory mechanisms involving long-range connectivity between the dorsal premotor cortex and posterior sensory regions. These

  15. Acute exposure to sarin increases blood brain barrier permeability and induces neuropathological changes in the rat brain: dose-response relationships.

    PubMed

    Abdel-Rahman, A; Shetty, A K; Abou-Donia, M B

    2002-01-01

    We hypothesize that a single exposure to an LD(50) dose of sarin induces widespread early neuropathological changes in the adult brain. In this study, we evaluated the early changes in the adult brain after a single exposure to different doses of sarin. Adult male rats were exposed to sarin by a single intramuscular injection at doses of 1, 0.5, 0.1 and 0.01 x LD(50). Twenty-four hours after the treatment, both sarin-treated and vehicle-treated (controls) animals were analyzed for: (i) plasma butyrylcholinesterase (BChE) activity; (ii) brain acetylcholinesterase (AChE) activity, (iii) m2 muscarinic acetylcholine receptor (m2 mAChR) ligand binding; (iv) blood brain barrier (BBB) permeability using [H(3)]hexamethonium iodide uptake assay and immunostaining for endothelial barrier antigen (EBA); and (v) histopathological changes in the brain using H&E staining, and microtubule-associated protein (MAP-2) and glial fibrillary acidic protein immunostaining. In animals treated with 1 x LD(50) sarin, the significant changes include a decreased plasma BChE, a decreased AChE in the cerebrum, brainstem, midbrain and the cerebellum, a decreased m2 mAChR ligand binding in the cerebrum, an increased BBB permeability in the cerebrum, brainstem, midbrain and the cerebellum associated with a decreased EBA expression, a diffuse neuronal cell death and a decreased MAP-2 expression in the cerebral cortex and the hippocampus, and degeneration of Purkinje neurons in the cerebellum. Animals treated with 0.5 x LD(50) sarin however exhibited only a few alterations, which include decreased plasma BChE, an increased BBB permeability in the midbrain and the brain stem but without a decrease in EBA expression, and degeneration of Purkinje neurons in the cerebellum. In contrast, animals treated with 0.1 and 0.01 x LD(50) did not exhibit any of the above changes. However, m2 mAChR ligand binding in the brainstem was increased after exposure to all doses of the sarin.Collectively, the above

  16. TU-CD-304-10: Development and Optimization of “Compton Lens” Collimator Design for Increased Dose Rate in SRS

    SciTech Connect

    Shepard, A; Bender, E

    2015-06-15

    Purpose: To provide a proof of concept for a new collimator design to increase the dose rate at isocenter for stereotactic radiosurgery (SRS) treatments by taking advantage of off axis Compton scattered photons which are attenuated in current collimators. Methods: A fundamentally new collimator design was developed and optimized by introducing a series of slits to a standard block collimator. The introduction of slits allowed for initially off axis radiation that was scattered in the direction of the target to contribute to the target dose. For optimization, the design was broken into two parts: an upper interaction plate where primary scattering occurs, and a lower “Compton slit” region which allows for scattered photons traveling toward the target to reach isocenter. To optimize the design, a series of simulations were performed using MCNP6 in which several key parameters were adjusted and the output was compared to a standard collimator. Key parameters modified included the collimator material, cone size, and interaction plate thickness. The effects of using energies different than the traditional 6 MV beam were also explored. Results: An optimized collimator design utilizing a solid interaction plate with a Cesium-137 beam and a 4 mm cone size resulted in a dose rate increase on the order of 5% relative to standard collimators in use. Currently, designs incorporating a Cesium-137 source are the most feasible due to necessary size and weight concerns for 6 MV beams. Conclusion: Preliminary designs provide a proof of concept and indicate a potential to improve upon the dose rate of current collimators while not largely compromising the sharp dose falloff inherent to SRS. Further optimization into the geometry and positioning of the interaction plate, as well as slit optimization, will likely lead to further dose rate increases than were observed in this study. Funding for this work was provided by the Wisconsin Alumni Research Foundation (WARF). Authors have

  17. Interaction of Prevotella intermedia strain 17 leucine-rich repeat domain protein AdpF with eukaryotic cells promotes bacterial internalization.

    PubMed

    Sengupta, Dipanwita; Kang, Dae-Joong; Anaya-Bergman, Cecilia; Wyant, Tiana; Ghosh, Arnab K; Miyazaki, Hiroshi; Lewis, Janina P

    2014-06-01

    Prevotella intermedia is an oral bacterium implicated in a variety of oral diseases. Although internalization of this bacterium by nonphagocytic host cells is well established, the molecular players mediating the process are not well known. Here, the properties of a leucine-rich repeat (LRR) domain protein, designated AdpF, are described. This protein contains a leucine-rich region composed of 663 amino acid residues, and molecular modeling shows that it folds into a classical curved solenoid structure. The cell surface localization of recombinant AdpF (rAdpF) was confirmed by electron and confocal microscopy analyses. The recombinant form of this protein bound fibronectin in a dose-dependent manner. Furthermore, the protein was internalized by host cells, with the majority of the process accomplished within 30 min. The internalization of rAdpF was inhibited by nystatin, cytochalasin, latrunculin, nocodazole, and wortmannin, indicating that microtubules, microfilaments, and signal transduction are required for the invasion. It is noteworthy that preincubation of eukaryotic cells with AdpF increased P. intermedia 17 internalization by 5- and 10-fold for HeLa and NIH 3T3 fibroblast cell lines, respectively. The addition of the rAdpF protein was also very effective in inducing bacterial internalization into the oral epithelial cell line HN4, as well as into primary cells, including human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs). Finally, cells exposed to P. intermedia 17 internalized the bacteria more readily upon reinfection. Taken together, our data demonstrate that rAdpF plays a role in the internalization of P. intermedia 17 by a variety of host cells.

  18. Interaction of Prevotella intermedia Strain 17 Leucine-Rich Repeat Domain Protein AdpF with Eukaryotic Cells Promotes Bacterial Internalization

    PubMed Central

    Sengupta, Dipanwita; Kang, Dae-Joong; Anaya-Bergman, Cecilia; Wyant, Tiana; Ghosh, Arnab K.; Miyazaki, Hiroshi

    2014-01-01

    Prevotella intermedia is an oral bacterium implicated in a variety of oral diseases. Although internalization of this bacterium by nonphagocytic host cells is well established, the molecular players mediating the process are not well known. Here, the properties of a leucine-rich repeat (LRR) domain protein, designated AdpF, are described. This protein contains a leucine-rich region composed of 663 amino acid residues, and molecular modeling shows that it folds into a classical curved solenoid structure. The cell surface localization of recombinant AdpF (rAdpF) was confirmed by electron and confocal microscopy analyses. The recombinant form of this protein bound fibronectin in a dose-dependent manner. Furthermore, the protein was internalized by host cells, with the majority of the process accomplished within 30 min. The internalization of rAdpF was inhibited by nystatin, cytochalasin, latrunculin, nocodazole, and wortmannin, indicating that microtubules, microfilaments, and signal transduction are required for the invasion. It is noteworthy that preincubation of eukaryotic cells with AdpF increased P. intermedia 17 internalization by 5- and 10-fold for HeLa and NIH 3T3 fibroblast cell lines, respectively. The addition of the rAdpF protein was also very effective in inducing bacterial internalization into the oral epithelial cell line HN4, as well as into primary cells, including human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs). Finally, cells exposed to P. intermedia 17 internalized the bacteria more readily upon reinfection. Taken together, our data demonstrate that rAdpF plays a role in the internalization of P. intermedia 17 by a variety of host cells. PMID:24711565

  19. Exponentially increasing incidences of cutaneous malignant melanoma in Europe correlate with low personal annual UV doses and suggests 2 major risk factors

    PubMed Central

    Merrill, Stephen J; Ashrafi, Samira; Subramanian, Madhan; Godar, Dianne E

    2015-01-01

    For several decades the incidence of cutaneous malignant melanoma (CMM) steadily increased in fair-skinned, indoor-working people around the world. Scientists think poor tanning ability resulting in sunburns initiate CMM, but they do not understand why the incidence continues to increase despite the increased use of sunscreens and formulations offering more protection. This paradox, along with lower incidences of CMM in outdoor workers, although they have significantly higher annual UV doses than indoor workers have, perplexes scientists. We found a temporal exponential increase in the CMM incidence indicating second-order reaction kinetics revealing the existence of 2 major risk factors. From epidemiology studies, we know one major risk factor for getting CMM is poor tanning ability and we now propose the other major risk factor may be the Human Papilloma Virus (HPV) because clinicians find β HPVs in over half the biopsies. Moreover, we uncovered yet another paradox; the increasing CMM incidences significantly correlate with decreasing personal annual UV dose, a proxy for low vitamin D3 levels. We also discovered the incidence of CMM significantly increased with decreasing personal annual UV dose from 1960, when it was almost insignificant, to 2000. UV and other DNA-damaging agents can activate viruses, and UV-induced cytokines can hide HPV from immune surveillance, which may explain why CMM also occurs in anatomical locations where the sun does not shine. Thus, we propose the 2 major risk factors for getting CMM are intermittent UV exposures that result in low cutaneous levels of vitamin D3 and possibly viral infection. PMID:26413188

  20. Environmentally Relevant Chronic Low-Dose Tritium and Gamma Exposures do not Increase Somatic Intrachromosomal Recombination in pKZ1 Mouse Spleen.

    PubMed

    Bannister, Laura; Serran, Mandy; Bertrand, Lindsey; Klokov, Dmitry; Wyatt, Heather; Blimkie, Melinda; Gueguen, Yann; Priest, Nicholas; Jourdain, Jean-René; Sykes, Pamela

    2016-12-01

    The toxicity of tritium is a public health concern given its presence and mobility in the environment. For risk predictions using radiological protection models, it is essential to allocate an appropriate radiation weighting factor (WR). This in turn should be consistent with the observed relative biological effectiveness (RBE) of tritium beta radiation. Although the International Commission on Radiological Protection (ICRP) currently recommends a WR of 1 for the calculation of committed effective dose for X rays, gamma rays and electrons of all energies, including tritium energies, there are concerns that tritium health risks are underestimated and that current regulatory tritium drinking water standards need revision. In this study, we investigated potential cytotoxic and genotoxic effects in mouse spleen after one month and eight months of chronic exposure to low-dose tritiated water (HTO). The dose regimes studied were designed to mimic human chronic consumption of HTO at levels of 10 kBq/l, 1 MBq/l and 20 MBq/l. The total doses from these radiation exposures ranged from 0.01 to 180 mGy. We also compared the biological effects of exposure to HTO with equivalent exposure to external whole-body (60)Co gamma rays. Changes in spleen weight and somatic intrachromosomal recombination (DNA inversions) in spleen tissue of pKZ1(Tg/+) mice were monitored. Our results showed no overall changes in either spleen organ weights and no increase mouse splenic intrachromosomal recombination frequencies, indicating that current drinking water standards for tritium exposure in the form of HTO are likely to be adequately protective against cytotoxic and genotoxic damage in spleen. These results demonstrate no evidence for cytotoxicity or genotoxicity in mouse spleen following chronic exposures to HTO activities (or equivalent gamma doses) up to 20 MBq/L.

  1. Has the sensitivity of soybean cultivars to ozone pollution increased with time? An analysis of published dose-response data

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The rising trend in concentrations of ground-level ozone (O3) – a common air pollutant and phytotoxin – currently being experienced in some world regions represents a threat to agricultural yield. Soybean (Glycine max (L.) Merr.) is an O3-sensitive crop species, and is experiencing increasing globa...

  2. PD-1 inhibitors increase the incidence and risk of pneumonitis in cancer patients in a dose-independent manner: a meta-analysis

    PubMed Central

    Wu, Jiaying; Hong, Dongsheng; Zhang, Xiangnan; Lu, Xiaoyang; Miao, Jing

    2017-01-01

    Therapies that targeted PD-1 have shown remarkable rates of durable clinical responses in patients with various tumor types. However, the extent and knowledge of pulmonary toxicities associated with PD-1 blockade, mainly manifested as pneumonitis, remains obscure. In this study, a total of 6360 subjects from 16 phase II/III clinical trials were pooled for meta-analysis to evaluate the overall incidence and risk of PD-1 inhibitors-related pneumonitis in cancer patients. The incidence of pneumonitis during anti-PD-1 immunotherapy was 2.92% (95%CI: 2.18–3.90%) for all-grade and 1.53% (95%CI: 1.15–2.04%) for high-grade pneumonitis. Compared with routine chemotherapy, PD-1 inhibitors were associated with a significant increased risk of pneumonitis. Moreover, among the types of tumor treated with PD-1 inhibitors, the melanoma patients have the lowest incidence of pneumonitis, while the non-small cell lung cancer (NSCLC) and renal cell carcinoma (RCC) patients have the highest. Furthermore, no significant differences were detected in the incidences of all- and high-grade pneumonitis between high-dose and low-dose groups of PD-1 inhibitors. In conclusion, PD-1 inhibitors were probably associated with an increased risk of pneumonitis in a dose-independent manner, compared with routine chemotherapeutic agents. The frequency and severity of treatment-mediated pneumonitis was quite different in patients with various tumor types. PMID:28272463

  3. Effects of Increasing Doses of UV-B on Main Phenolic Acids Content, Antioxidant Activity and Estimated Biomass in Lavandin (Lavandula x intermedia).

    PubMed

    Usano-Alemany, Jaime; Panjai, Lachinee

    2015-07-01

    Lavandin is a well-known aromatic plant cultivated mainly for its valuable essential oil. Nonetheless, little attention has been paid so far to the quantification of other natural products such as polyphenols. Accordingly, we examined the effect of increasing doses of UV-B radiation on the main phenolic content, antioxidant activity and estimated biomass of one year old lavandin pots compared with pots grown outdoors. Significantly higher total phenolic content and concentration of main polyphenols have been found in outdoor plants. Rosmarinic acid has been described as the major phenolic compound in methanolic extracts (max. 25.9 ± 9.7 mg/g(-1) DW). Furthermore, we found that increasing doses of UV-B promote the plant growth of this species as well as the accumulation of phenolic compounds although with less antioxidant capacity in scavenging DPPH radicals. On the other hand, our results showed a remarkable variability among individual plants regarding the content of major phenolic acids. The application of UV-B doses during plant growth could be a method to promote biomass in this species along with the promotion of higher content of valuable secondary metabolites.

  4. Increased blood oxidative stress in experimental menopause rat model: the effects of vitamin A low-dose supplementation upon antioxidant status in bilateral ovariectomized rats.

    PubMed

    Behr, Guilherme Antônio; Schnorr, Carlos Eduardo; Moreira, José Cláudio Fonseca

    2012-04-01

    Menopause has been reported to be associated with increased oxidative stress and metabolic disorders among women worldwide. Disarrangements in the redox state similar to those observed in women during the decline of ovarian hormonal activity can be obtained experimentally through rat bilateral ovariectomy. The search for alternative treatments to improve life quality in postmenopausal woman is really important. The aim of this study was to evaluate biochemical and oxidative stress parameters that distinguish sham-operated female rats from Wistar rats bilaterally ovariectomized (OVX). Additionally, we have also investigated the effects of retinol palmitate (a vitamin A supplement) low-dose supplementation (500 or 1500 IU/kg/day, during 30 days) upon blood and plasma antioxidant status in OVX rats. Ovariectomy caused an increase in body weight gain, pronounced uterine atrophy, decreased plasma triglycerides and increased total cholesterol levels, and reduced acid uric content. Moreover, we found increased blood peroxidase activities (catalase and glutathione peroxidase), decreased plasma non-enzymatic antioxidant defenses total reactive antioxidant potential and total antioxidant reactivity, decreased protein and non-protein SH levels, accompanied by increased protein oxidative damage (carbonyl). In addition, vitamin A low-dose supplementation was capable to ameliorate antioxidant status in OVX rats, restoring both enzymatic and non-enzymatic defenses, promoting reduction in plasma SH content, and decreasing protein oxidative damage levels. This is the first work in the literature showing that vitamin A at low dose may be beneficial in the treatment of menopause symptoms. Further studies will be made to better understand the effects of vitamin A supplementation in menopause rat model.

  5. The cessation of the long-term exposure to low doses of mercury ameliorates the increase in systolic blood pressure and vascular damage in rats.

    PubMed

    Rizzetti, Danize Aparecida; Torres, João Guilherme Dini; Escobar, Alyne Goulart; da Silva, Taiz Martins; Moraes, Paola Zambelli; Hernanz, Raquel; Peçanha, Franck Maciel; Castro, Marta Miguel; Vassallo, Dalton Valentim; Salaices, Mercedes; Alonso, Maria Jesús; Wiggers, Giulia Alessandra

    2017-02-18

    This study aimed to verify whether a prolonged exposure to low-level mercury promotes haemodynamic disorders and studied the reversibility of this vascular damage. Rats were divided into seven groups: three control groups received saline solution (im) for 30, 60 or 90 days; two groups received HgCl2 (im, first dose, 4.6μg/kg, subsequent doses 0.07μg/kg/day) for 30 or 60 days; two groups received HgCl2 for 30 or 60 days (im, same doses) followed by a 30-day washout period. Systolic blood pressure (SBP) was measured, along with analysis of vascular response to acetylcholine (ACh) and phenylephrine (Phe) in the absence and presence of endothelium, a nitric oxide (NO) synthase inhibitor, an NADPH oxidase inhibitor, superoxide dismutase, a non-selective cyclooxygenase (COX) inhibitor and an AT1 receptor blocker. Reactive oxygen species (ROS) levels and antioxidant power were measured in plasma. HgCl2 exposure for 30 and 60 days: a) reduced the endothelium-dependent relaxation; b) increased the Phe-induced contraction and the contribution of ROS, COX-derived vasoconstrictor prostanoids and angiotensin II acting on AT1 receptors to this response while the NO participation was reduced; c) increased the oxidative stress in plasma; d) increased the SBP only after 60 days of exposure. After the cessation of HgCl2 exposure, SBP, endothelium-dependent relaxation, Phe-induced contraction and the oxidative stress were normalised, despite the persistence of the increased COX-derived prostanoids. These results demonstrated that long-term HgCl2 exposure increases SBP as a consequence of vascular dysfunction; however, after HgCl2 removal from the environment the vascular function ameliorates.

  6. Ultraviolet B radiation increases hairless mouse mast cells in a dose-dependent manner and alters distribution of UV-induced mast cell growth factor.

    PubMed

    Kligman, L H; Murphy, G F

    1996-01-01

    In studies of the effects of chronic UVB irradiation on dermal connective tissue in the hairless mouse, we observed that the number and size of mast cells was increased. Because mast cells are known to be associated with connective tissue remodeling, we examined and quantified the effect of increasing UVB (290-320 nm) doses on this cell. Groups of mice were exposed to filtered FS-40 Westinghouse lamps (290-400 nm: peak irradiance 313 nm) for 1-5 minimal erythema doses (MED) thrice weekly for 10 weeks. Appropriate controls were included. Biopsies, processed for light microscopy, were stained with toluidine blue. Mast cells were counted in 15 high-magnification fields per specimen with upper and lower dermis scored separately. Significant increases in large densely granular mast cells occurred at 2 MED in the lower dermis, in association with a UVB-exacerbated granulomatous reaction. In the upper dermis, mast cells were significantly increased with 3 MED. These findings suggest that mast cells may play a dual role in UV-irradiated skin with those in the lower dermis related to inflammation processes and those in the upper dermis involved in connective tissue modeling. To gain understanding of the mechanism of mast cell recruitment and maturation, we examined the effect of UVB on mast cell growth factor expression. This was enhanced in the epidermis by UVB, with a shift from cytoplasmic staining to membrane-associated or intercellular staining at 2 MED and higher. Dermal dendritic and mononuclear cells also showed increased reactivity.

  7. Induction of extracellular ATP mediates increase in intracellular thioredoxin in RAW264.7 cells exposed to low-dose γ-rays.

    PubMed

    Ohshima, Yasuhiro; Kitami, Akihiro; Kawano, Ayumi; Tsukimoto, Mitsutoshi; Kojima, Shuji

    2011-09-15

    We previously showed that low doses (0.25-0.5 Gy) of γ-rays elevated thioredoxin (Trx-1) in various organs of mice after whole-body irradiation. Also, it is reported that extracellular ATP, which is released in response to various stresses, regulates the expression of intracellular antioxidants through activation of P2 receptors. We have recently found that low-dose γ-rays induce ATP release from the exposed cells. However, it is not yet clear whether the radiation-induced extracellular ATP modulates the cellular redox balance. Here, we investigated whether γ-ray irradiation-induced release of extracellular ATP contributes to the induction of the cellular antioxidant Trx-1, using mouse macrophage-like RAW264.7 cells. Irradiation with γ-rays or exogenously added ATP increased the expression of Trx-1, and in both cases the increase was blocked by pretreatment with an ectonucleotidase, apyrase. Then, the involvement of ATP-dependent reactive oxygen species (ROS) generation in the increase in antioxidant capacity was examined. ATP stimulation promoted the generation of intracellular ROS and also increased Trx-1 expression. The increase in Trx-1 expression was significantly suppressed by pretreatment of the cells with antioxidants. In conclusion, the γ-ray irradiation-induced release of extracellular ATP may, at least in part, contribute to the production of ROS via purinergic signaling, leading to promotion of intracellular antioxidants as an adaptive response to an oxidative stress.

  8. Long-term low-level laser therapy promotes an increase in maximal oxygen uptake and exercise performance in a dose-dependent manner in Wistar rats.

    PubMed

    Perini, Júlia Luiza; Hentschke, Vítor Scotta; Sonza, Anelise; Dal Lago, Pedro

    2016-02-01

    The use of low-level laser therapy (LLLT) represents a new intervention modality that has been explored to enhance exercise performance. The aim of this study was to evaluate the influence of LLLT (GaAIAs-850 nm) at different doses on VO2max and on exercise performance in rats. Male Wistar rats were divided into three groups: "placebo" rats (P-LLLT, n = 10), rats at a dose of 0.315 J per treatment point of LLLT (8.7 J/cm(2)-LLLT, n = 10), and rats at a dose of 2.205 J per treatment point of LLLT (61.2 J/cm(2)-LLLT, n = 10). The LLLT was applied bilaterally at the biceps femoris, gluteus, lateral and medial gastrocnemius, iliopsoas, and adductor longus muscles. One spot in each muscle belly was applied, with a sum of 12 spots in each rat, once a day, for 10 days. All animals performed the maximal exercise test (ET) at a metabolic treadmill for rats, with simultaneous gas analysis. The distance covered was measured during ET, before and after the conclusion of the LLLT protocol. The data were compared by a repeated measures two-way ANOVA followed by the Student-Newman-Keuls post hoc tests (p < .05). The 61.2 J/cm(2)-LLLT group increased VO2basal (~40 %), VO2max (~24 %), VCO2max (~17 %), and distance covered (~34 %) after LLLT application on the skeletal muscle. No significant results were found comparing before and after conditions for the studied variables considering P-LLLT and 8.7 J/cm(2)-LLLT groups. The LLLT promoted in a dose-dependent manner an increase in oxygen consumption uptake and a performance increment of male Wistar rats.

  9. Dietary Leucine Supplementation Improves the Mucin Production in the Jejunal Mucosa of the Weaned Pigs Challenged by Porcine Rotavirus.

    PubMed

    Mao, Xiangbing; Liu, Minghui; Tang, Jun; Chen, Hao; Chen, Daiwen; Yu, Bing; He, Jun; Yu, Jie; Zheng, Ping

    2015-01-01

    The present study was mainly conducted to determine whether dietary leucine supplementation could attenuate the decrease of the mucin production in the jejunal mucosa of weaned pigs infected by porcine rotavirus (PRV). A total of 24 crossbred barrows weaned at 21 d of age were assigned randomly to 1 of 2 diets supplemented with 1.00% L-leucine or 0.68% L-alanine (isonitrogenous control) for 17 d. On day 11, all pigs were orally infused PRV or the sterile essential medium. During the first 10 d of trial, dietary leucine supplementation could improve the feed efficiency (P = 0.09). The ADG and feed efficiency were impaired by PRV infusion (P<0.05). PRV infusion also increased mean cumulative score of diarrhea, serum rotavirus antibody concentration and crypt depth of the jejunal mucosa (P<0.05), and decreased villus height: crypt depth (P = 0.07), goblet cell numbers (P<0.05), mucin 1 and 2 concentrations (P<0.05) and phosphorylated mTOR level (P<0.05) of the jejunal mucosa in weaned pigs. Dietary leucine supplementation could attenuate the effects of PRV infusion on feed efficiency (P = 0.09) and mean cumulative score of diarrhea (P = 0.09), and improve the effects of PRV infusion on villus height: crypt depth (P = 0.06), goblet cell numbers (P<0.05), mucin 1 (P = 0.08) and 2 (P = 0.07) concentrations and phosphorylated mTOR level (P = 0.08) of the jejunal mucosa in weaned pigs. These results suggest that dietary 1% leucine supplementation alleviated the decrease of mucin production and goblet cell numbers in the jejunal mucosa of weaned pigs challenged by PRV possibly via activation of the mTOR signaling.

  10. Leucine-enriched essential amino acids attenuate inflammation in rat muscle and enhance muscle repair after eccentric contraction.

    PubMed

    Kato, Hiroyuki; Miura, Kyoko; Nakano, Sayako; Suzuki, Katsuya; Bannai, Makoto; Inoue, Yoshiko

    2016-09-01

    Eccentric exercise results in prolonged muscle damage that may lead to muscle dysfunction. Although inflammation is essential to recover from muscle damage, excessive inflammation may also induce secondary damage, and should thus be suppressed. In this study, we investigated the effect of leucine-enriched essential amino acids on muscle inflammation and recovery after eccentric contraction. These amino acids are known to stimulate muscle protein synthesis via mammalian target of rapamycin (mTOR), which, is also considered to alleviate inflammation. Five sets of 10 eccentric contractions were induced by electrical stimulation in the tibialis anterior muscle of male SpragueDawley rats (8-9 weeks old) under anesthesia. Animals received a 1 g/kg dose of a mixture containing 40 % leucine and 60 % other essential amino acids or distilled water once a day throughout the experiment. Muscle dysfunction was assessed based on isometric dorsiflexion torque, while inflammation was evaluated by histochemistry. Gene expression of inflammatory cytokines and myogenic regulatory factors was also measured. We found that leucine-enriched essential amino acids restored full muscle function within 14 days, at which point rats treated with distilled water had not fully recovered. Indeed, muscle function was stronger 3 days after eccentric contraction in rats treated with amino acids than in those treated with distilled water. The amino acid mix also alleviated expression of interleukin-6 and impeded infiltration of inflammatory cells into muscle, but did not suppress expression of myogenic regulatory factors. These results suggest that leucine-enriched amino acids accelerate recovery from muscle damage by preventing excessive inflammation.

  11. Increased Hypoxic Dose After Training at Low Altitude with 9h Per Night at 3000m Normobaric Hypoxia.

    PubMed

    Carr, Amelia J; Saunders, Philo U; Vallance, Brent S; Garvican-Lewis, Laura A; Gore, Christopher J

    2015-12-01

    This study examined effects of low altitude training and a live-high: train-low protocol (combining both natural and simulated modalities) on haemoglobin mass (Hbmass), maximum oxygen consumption (VO2max), time to exhaustion, and submaximal exercise measures. Eighteen elite-level race-walkers were assigned to one of two experimental groups; lowHH (low Hypobaric Hypoxia: continuous exposure to 1380 m for 21 consecutive days; n = 10) or a combined low altitude training and nightly Normobaric Hypoxia (lowHH+NHnight: living and training at 1380 m, plus 9 h.night(-1) at a simulated altitude of 3000 m using hypoxic tents; n = 8). A control group (CON; n = 10) lived and trained at 600 m. Measurement of Hbmass, time to exhaustion and VO2max was performed before and after the training intervention. Paired samples t-tests were used to assess absolute and percentage change pre and post-test differences within groups, and differences between groups were assessed using a one-way ANOVA with least significant difference post-hoc testing. Statistical significance was tested at p < 0.05. There was a 3.7% increase in Hbmass in lowHH+NHnight compared with CON (p = 0.02). In comparison to baseline, Hbmass increased by 1.2% (±1.4%) in the lowHH group, 2.6% (±1.8%) in lowHH+NHnight, and there was a decrease of 0.9% (±4.9%) in CON. VO2max increased by ~4% within both experimental conditions but was not significantly greater than the 1% increase in CON. There was a ~9% difference in pre and post-intervention values in time to exhaustion after lowHH+NH-night (p = 0.03) and a ~8% pre to post-intervention difference (p = 0.006) after lowHH only. We recommend low altitude (1380 m) combined with sleeping in altitude tents (3000 m) as one effective alternative to traditional altitude training methods, which can improve Hbmass. Key pointsIn some countries, it may not be possible to perform classical altitude training effectively, due to the low elevation at altitude training venues. An

  12. Leucine acts in the brain to suppress food intake but does not function as a physiological signal of low dietary protein.

    PubMed

    Laeger, Thomas; Reed, Scott D; Henagan, Tara M; Fernandez, Denise H; Taghavi, Marzieh; Addington, Adele; Münzberg, Heike; Martin, Roy J; Hutson, Susan M; Morrison, Christopher D

    2014-08-01

    Intracerebroventricular injections of leucine are sufficient to suppress food intake, but it remains unclear whether brain leucine signaling represents a physiological signal of protein balance. We tested whether variations in dietary and circulating levels of leucine, or all three branched-chain amino acids (BCAAs), contribute to the detection of reduced dietary protein. Of the essential amino acids (EAAs) tested, only intracerebroventricular injection of leucine (10 μg) was sufficient to suppress food intake. Isocaloric low- (9% protein energy; LP) or normal- (18% protein energy) protein diets induced a divergence in food intake, with an increased consumption of LP beginning on day 2 and persisting throughout the study (P < 0.05). Circulating BCAA levels were reduced the day after LP diet exposure, but levels subsequently increased and normalized by day 4, despite persistent hyperphagia. Brain BCAA levels as measured by microdialysis on day 2 of diet exposure were reduced in LP rats, but this effect was most prominent postprandially. Despite these diet-induced changes in BCAA levels, reducing dietary leucine or total BCAAs independently from total protein was neither necessary nor sufficient to induce hyperphagia, while chronic infusion of EAAs into the brain of LP rats failed to consistently block LP-induced hyperphagia. Collectively, these data suggest that circulating BCAAs are transiently reduced by dietary protein restriction, but variations in dietary or brain BCAAs alone do not explain the hyperphagia induced by a low-protein diet.

  13. The influence of 8 weeks of whey-protein and leucine supplementation on physical and cognitive performance.

    PubMed

    Walker, Thomas B; Smith, Jessica; Herrera, Monica; Lebegue, Breck; Pinchak, Andrea; Fischer, Joseph

    2010-10-01

    The purpose of this study was to investigate the ability of whey-protein and leucine supplementation to enhance physical and cognitive performance and body composition. Thirty moderately fit participants completed a modified Air Force fitness test, a computer-based cognition test, and a dual-energy X-ray-absorptiometry scan for body composition before and after supplementing their daily diet for 8 wk with either 19.7 g of whey protein and 6.2 g leucine (WPL) or a calorie-equivalent placebo (P). Bench-press performance increased significantly from Week 1 to Week 8 in the WPL group, whereas the increase in the P group was not significant. Push-up performance increased significantly for WPL, and P showed a nonsignificant increase. Total mass, fat-free mass, and lean body mass all increased significantly in the WPL group but showed no change in the P group. No differences were observed within or between groups for crunches, chin-ups, 3-mile-run time, or cognition. The authors conclude that supplementing with whey protein and leucine may provide an advantage to people whose performance benefits from increased upper body strength and/or lean body mass.

  14. Regulation of the plasma amino acid profile by leucine via the system L amino acid transporter.

    PubMed

    Zhen, Hongmin; Nakamura, Koichi; Kitaura, Yasuyuki; Kadota, Yoshihiro; Ishikawa, Takuya; Kondo, Yusuke; Xu, Minjun; Shimomura, Yoshiharu

    2015-01-01

    Plasma concentrations of amino acids reflect the intracellular amino acid pool in mammals. However, the regulatory mechanism requires clarification. In this study, we examined the effect of leucine administration on plasma amino acid profiles in mice with and without the treatment of 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH) or rapamycin as an inhibitor of system L or mammalian target of rapamycin complex 1, respectively. The elevation of plasma leucine concentration after leucine administration was associated with a significant decrease in the plasma concentrations of isoleucine, valine, methionine, phenylalanine, and tyrosine; BCH treatment almost completely blocked the leucine-induced decrease in plasma amino acid concentrations. Rapamycin treatment had much less effects on the actions of leucine than BCH treatment. These results suggest that leucine regulates the plasma concentrations of branched-chain amino acids, methionine, phenylalanine, and tyrosine, and that system L amino acid transporters are involved in the leucine action.

  15. Supplemental dietary leucine and the skeletal muscle anabolic response to essential amino acids.

    PubMed

    Pasiakos, Stefan M; McClung, James P

    2011-09-01

    Skeletal muscle protein synthesis (MPS) is regulated by a number of dietary factors, to include essential amino acids (EAAs). Leucine, a branched-chain amino acid, has been identified as a stimulator of MPS in many cell culture and animal studies. However, whether supplemental leucine exerts a unique stimulatory effect, as compared to other EAAs, on muscle anabolism in humans has not been clearly demonstrated. A recent study found no improvement in resting MPS in adults who consumed a 10 g EAA supplement providing added leucine (3.5 g leucine) when compared to a control 10 g EAA supplement (1.8 g leucine). These findings suggest that added leucine is unnecessary for the stimulation of MPS when sufficient EAAs are provided; however, the study of supplemental leucine during conditions such as endurance exercise, caloric deprivation, and ageing may be warranted.

  16. HIGH ETHANOL DOSE DURING EARLY ADOLESCENCE INDUCES LOCOMOTOR ACTIVATION AND INCREASES SUBSEQUENT ETHANOL INTAKE DURING LATE ADOLESCENCE

    PubMed Central

    Acevedo, María Belén; Molina, Juan Carlos; Nizhnikov, Michael E.; Spear, Norman E.; Pautassi, Ricardo Marcos

    2011-01-01

    Adolescent initiation of ethanol consumption is associated with subsequent heightened probability of ethanol-use disorders. The present study examined the relationship between motivational sensitivity to ethanol initiation in adolescent rats and later ethanol intake. Experiment 1 determined that ethanol induces locomotor activation shortly after administration but not if tested at a later post-administration interval. In Experiment 2, adolescents were assessed for ethanol-induced locomotor activation on postnatal day 28. These animals were then evaluated for ethanol-mediated conditioned taste aversion and underwent a 16-day-long ethanol intake protocol. Ethanol-mediated aversive effects were unrelated to ethanol locomotor stimulation or subsequent ethanol consumption patterns. Ethanol intake during late adolescence was greatest in animals initiated to ethanol earliest at postnatal day 28. Females that were more sensitive to ethanol’s locomotor-activating effects showed a transient increase in ethanol self-administration. Blood ethanol concentrations during initiation were not related to ethanol-induced locomotor activation. Adolescent rats appeared sensitive to the locomotor-stimulatory effects of ethanol. Even brief ethanol exposure during adolescence may promote later ethanol intake. PMID:20373327

  17. Increase in the astaxanthin synthase gene (crtS) dose by in vivo DNA fragment assembly in Xanthophyllomyces dendrorhous

    PubMed Central

    2013-01-01

    Background Xanthophyllomyces dendrorhous is a basidiomycetous yeast that is relevant to biotechnology, as it can synthesize the carotenoid astaxanthin. However, the astaxanthin levels produced by wild-type strains are low. Although different approaches for promoting increased astaxanthin production have been attempted, no commercially competitive results have been obtained thus far. A promising alternative to facilitate the production of carotenoids in this yeast involves the use of genetic modification. However, a major limitation is the few available molecular tools to manipulate X. dendrorhous. Results In this work, the DNA assembler methodology that was previously described in Saccharomyces cerevisiae was successfully applied to assemble DNA fragments in vivo and integrate these fragments into the genome of X. dendrorhous by homologous recombination in only one transformation event. Using this method, the gene encoding astaxanthin synthase (crtS) was overexpressed in X. dendrorhous and a higher level of astaxanthin was produced. Conclusions This methodology could be used to easily and rapidly overexpress individual genes or combinations of genes simultaneously in X. dendrorhous, eliminating numerous steps involved in conventional cloning methods. PMID:24103677

  18. A sub-threshold dose of pilocarpine increases glutamine synthetase in reactive astrocytes and enhances the progression of amygdaloid-kindling epilepsy in rats.

    PubMed

    Sun, Hong-Liu; Deng, Da-Ping; Pan, Xiao-Hong; Wang, Chao-Yun; Zhang, Xiu-Li; Chen, Xiang-Ming; Wang, Chun-Hua; Liu, Yu-Xia; Li, Shu-Cui; Bai, Xian-Yong; Zhu, Wei

    2016-03-02

    The prognosis of patients exposed to a sub-threshold dose of a proconvulsant is difficult to establish. In this study, we investigated the effect of a single sub-threshold dose of the proconvulsant pilocarpine (PILO) on the progression of seizures that were subsequently induced by daily electrical stimulation (kindling) of the amygdaloid formation. Male Sprague–Dawley rats were each implanted with an electrode in the right basolateral amygdala and an indwelling cannula in the right ventricle. The animals were randomized into groups and were administered one of the following treatments: saline, PILO, saline+L-α-aminoadipic acid (L-AAA; one dosage tested), PILO+L-AAA, or PILO+L-methionine sulfoximine (three dosages tested). Amygdaloid stimulation and electroencephalography were performed once daily. We performed immunohistochemistry and western blot for glial fibrillary acidic protein and glutamine synthetase (GS). We also assayed the enzymic activity of GS in discrete brain regions. An intraperitoneal injection of a sub-threshold PILO dose enhanced the progression of amygdaloid-kindling seizures and was accompanied by an increase in reactive-astrocyte and GS (content and activity) in the hippocampus and piriform cortex. L-AAA and L-methionine sulfoximine, inhibitors of astrocytic and GS function, respectively, abolished the effect of PILO on amygdaloid-kindling seizures. We conclude that one sub-threshold dose of a proconvulsant may enhance the progression of subsequent epilepsy and astrocytic GS may play a role in this phenomenon. Thus, a future therapy for epilepsy could be inhibition of astrocytes and/or GS.

  19. Transition in Survival From Low-Dose Hyper-Radiosensitivity to Increased Radioresistance Is Independent of Activation of ATM SER1981 Activity

    SciTech Connect

    Krueger, Sarah A.; Collis, Spencer J.; Joiner, Michael C.; Wilson, George D.; Marples, Brian

    2007-11-15

    Purpose: The molecular basis of low-dose hyper-radiosensitivity (HRS) is only partially understood. The aim of this study was to define the roles of ataxia telangiectasia mutated (ATM) activity and the downstream ATM-dependent G{sub 2}-phase cell cycle checkpoint in overcoming HRS and triggering radiation resistance. Methods and Materials: Survival was measured using a high-resolution clonogenic assay. ATM Ser1981 activation was measured by Western blotting. The role of ATM was determined in survival experiments after molecular (siRNA) and chemical (0.4 mM caffeine) inhibition and chemical (20 {mu}g/mL chloroquine, 15 {mu}M genistein) activation 4-6 h before irradiation. Checkpoint responsiveness was assessed in eight cell lines of differing HRS status using flow cytometry to quantify the progression of irradiated (0-2 Gy) G{sub 2}-phase cells entering mitosis, using histone H3 phosphorylation analysis. Results: The dose-response pattern of ATM activation was concordant with the transition from HRS to radioresistance. However, ATM activation did not play a primary role in initiating increased radioresistance. Rather, a relationship was discovered between the function of the downstream ATM-dependent early G{sub 2}-phase checkpoint and the prevalence and overcoming of HRS. Four cell lines that exhibited HRS failed to show low-dose (<0.3-Gy) checkpoint function. In contrast, four HRS-negative cell lines exhibited immediate cell cycle arrest for the entire 0-2-Gy dose range. Conclusion: Overcoming HRS is reliant on the function of the early G{sub 2}-phase checkpoint. These data suggest that clinical exploitation of HRS could be achieved by combining radiotherapy with chemotherapeutic agents that modulate this cell cycle checkpoint.

  20. A High-Dose Shiitake Mushroom Increases Hepatic Accumulation of Triacylglycerol in Rats Fed a High-Fat Diet: Underlying Mechanism

    PubMed Central

    Handayani, Dian; Meyer, Barbara J.; Chen, Jiezhong; Brown, Simon H. J.; Mitchell, Todd W.; Huang, Xu-Feng

    2014-01-01

    Shiitake mushroom have been shown to have health benefits including lowering plasma lipids and preventing body weight gain. However, their underlying mechanisms are largely unknown. The study aim was to assess the potential underlying mechanism of Shiitake mushrooms in lowering plasma triacylglycerol (TAG) in rats fed a high fat diet (HFD). Forty Wistar rats were divided into control group were given HFD and treatment group were fed HFD, enriched with Shiitake mushroom powder at a low dose (LD-M): 0.7%, medium dose (MD-M): 2%, or high dose (HD-M): 6% (wt:wt) for six weeks. Diets were isocaloric containing ~50% energy from fat. After six weeks’ dietary intervention, the rats were sacrificed, and blood and tissue samples were collected. The HD-M group showed a significantly higher ratio of liver weight to 100 g body weight (p < 0.05), a more severe hepatic steatosis marker, such as hepatocyte ballooning (p < 0.0001), and more liver triacylglycerol content than LD-M and MD-M (p < 0.05). HD-M also showed a significantly decreased ratio of phosphatidylcholine (PC) to phosphatidylethanolamine (PE) compared to HFD (p < 0.05), however, there were no differences compared to HD-M and MD-M. Our results also showed a positive association between the dosage, liver TAG, and liver ballooning histology. A negative association was found between the mushroom dosage and the ratio of liver PC to PE. This study showed the mechanism of how high-dose Shiitake mushroom (HD-M) prevents obesity by increasing TAG accumulation in the liver, rather than adipose tissue. PMID:24566434

  1. Light-emitting diode therapy (LEDT) before matches prevents increase in creatine kinase with a light dose response in volleyball players.

    PubMed

    Ferraresi, Cleber; Dos Santos, Ricardo Vinicius; Marques, Guilherme; Zangrande, Marcelo; Leonaldo, Roberley; Hamblin, Michael R; Bagnato, Vanderlei Salvador; Parizotto, Nivaldo Antonio

    2015-05-01

    Low-level laser (light) therapy (LLLT) has been applied over skeletal muscles before intense exercise (muscular pre-conditioning) in order to reduce fatigue and muscle damage (measured by creatine kinase, CK) in clinical trials. However, previous exercise protocols do not exactly simulate the real muscle demand required in sports. For this reason, the aim of this randomized and double-blind placebo-controlled trial was to investigate whether light-emitting diode therapy (LEDT) applied over the quadriceps femoris muscles, hamstrings, and triceps surae of volleyball players before official matches could prevent muscle damage (CK) with a dose response, establishing a therapeutic window. A professional male volleyball team (12 athletes) was enrolled in this study, and LEDT was applied before 4 matches during a national championship. LEDT used an array of 200 light-emitting diodes (LEDs) arranged in 25 clusters of 4 infrared LEDs (850 ± 20 nm; 130 mW) and 25 clusters of 4 red LEDs (630 ± 10 nm; 80 mW). Athletes were randomized to receive one of four different total doses over each muscle group in a double-blind protocol: 105 J (20 s), 210 J (40 s), 315 J (60 s), and placebo (no light for 30 s). CK in blood was assessed 1 h before and 24 h after each match. LEDT at 210 J avoided significant increases in CK (+10 %; P = 0.993) as well as 315 J (+31 %, P = 0.407). Placebo (0 J) allowed a significant increase in CK (+53 %; P = 0.012) as well as LEDT at 105 J (+59 %; P = 0.001). LEDT prevented significant increases of CK in blood in athletes when applied before official matches with a light dose response of 210-315 J, suggesting athletes might consider applying LEDT before competition.

  2. Increasing prepartum dietary crude protein using poultry by-product meal dose not influence performance of multiparous Holstein dairy cows.

    PubMed

    Yazdi, M Hossein; Amanlou, H; Mahjoubi, E

    2009-11-15

    The aim of this study was to compare the effects of two levels of Crude Protein (CP) using Poultry by-Product Meal (PBPM) fed during late gestation on the performance, blood metabolites and colostrum composition of Holstein cows. Twenty multiparous cows 26 +/- 6 days before expected calving were assigned randomly to two treatments containing 1) 140 g kg(-1) DM CP (34 g kg(-1) DM PBPM) 2) 160 g kg(-1) DM CP (75 g kg(-1) DM PBPM). The cow's BCS was 3.56 +/- 0.5 on average, at the beginning of the trial. Yields of milk, protein, lactose and fat were not affected by prepartum dietary CP level. Colostrum composition (fat, CP and total solids percents), blood metabolites (Ca, glucose, total protein, albumin, globulin, urea N and cholesterol) and metabolic diseases incidence were not influenced by prepartum dietary CP level. There was no significant difference between treatments in body weight and BCS changes. As expected, blood urea N before calving was higher in the cows fed 160 g kg(-1) DM CP diets (p < 0.002). Serum cholesterol during prepartum (p < 0.03) and postpartum (p < 0.01) periods was significantly lower in 160 g kg(-1) DM CP treatment. In general, although postpartum glucose level increased in cows which received 160 g kg(-1) DM CP in the diet, it seems that there is no other obvious advantages over feeding the 140 g kg(-1) DM CP diet. So feeding this level of CP diet to close up cows is recommended.

  3. Leucine stimulates ASCT2 amino acid transporter expression in porcine jejunal epithelial cell line (IPEC-J2) through PI3K/Akt/mTOR and ERK signaling pathways.

    PubMed

    Zhang, Shihai; Ren, Man; Zeng, Xiangfang; He, Pingli; Ma, Xi; Qiao, Shiyan

    2014-12-01

    Leucine has been shown to influence intestinal protein metabolism, cell proliferation and migration. Furthermore, our previous study demonstrated that branched-chain amino acids could modulate the intestinal amino acid and peptide transporters in vivo. As the possible mechanisms are still largely unknown, in the present work, we studied the transcriptional and translational regulation of leucine on amino acid transporter production in IPEC-J2 cells and the signaling pathways involved. Treatment of IPEC-J2 cells with 7.5 mM leucine enhanced the mRNA expression of the Na(+)-neutral AA exchanger 2 (ASCT2) and 4F2 heavy chain (4F2hc) and caused an increase in ASCT2 protein expression. Leucine also activated phosphorylation of 4E-BP1 and eIF4E through the phosphorylation of mTOR, Akt and ERK signaling pathways in IPEC-J2 cells. Pre-treatment of IPEC-J2 cells with inhibitors of mTOR and Akt (rapamycin and wortmannin) or an inhibitor of ERK (PD098059) for 30 min before leucine treatment attenuated the positive effect of leucine in enhancing the protein abundance of ASCT2. These results demonstrate that leucine could up-regulate the expression of the amino acid transporters (ASCT2) through transcriptional and translational regulation by ERK and PI3K/Akt/mTOR activation.

  4. Determination of Levo-Rotatory Leucine by a Fluorescent Probe

    NASA Astrophysics Data System (ADS)

    Cheng, D.; Zhu, H.

    2015-03-01

    With a certain concentration of Michaelis buffer solution (pH 6.1), and in the presence of CTMAB, the fluorescence intensity of fluorescein, which is quenched by Pd 2+ , can be enhanced after adding a certain content of L-leucine. Thereby, we establish a fluorescence spectrometry method to detect the content of L-leucine, the added amount of which is proportional to the fluorescence enhancement. When the excitation slit width is 3 nm and the emission slit width is 5 nm, we obtain the following results: correlation coefficient R = 0.9981, linear range 0.125-1.375 mg/l, and detection limit 0.00028 mg/l.

  5. Photolysis of rac-leucine with circularly polarized synchrotron radiation.

    PubMed

    Meierhenrich, Uwe J; Filippi, Jean-Jacques; Meinert, Cornelia; Hoffmann, Søren V; Bredehöft, Jan Hendrik; Nahon, Laurent

    2010-06-01

    Amino acids that pass the RNA machinery in living organisms occur in L-configuration. The question on the evolutionary origin of this biomolecular asymmetry remains unanswered to this day. Amino acids were detected in artificially produced interstellar ices, and L-enantiomer-enriched amino acids were identified in CM-type meteorites. This hints at a possible interstellar/circumstellar origin of the amino acids themselves as well as their stereochemical asymmetry. Based upon the current knowledge about the occurrence of circularly-polarized electromagnetic radiation in interstellar environments, we subjected rac-leucine to far-UV circularly-polarized synchrotron radiation. Asymmetric photolysis was followed by an analysis in an enantioselective GC/MS system. Here, we report on an advanced photolysis rate of more than 99% for leucine. The results indicate that high photolysis rates can occur under the chosen conditions, favoring enantioselective photolysis. In 2014, the obtained results will be reexamined by cometary mission Rosetta.

  6. Control of proteinuria with increased doses of agalsidase alfa in a patient with Fabry disease with atypical genotype-phenotype expression.

    PubMed

    Paliouras, Christos; Aperis, Georgios; Lamprianou, Foteini; Ntetskas, Giorgos; Roufas, Konstantinos; Alivanis, Polichronis

    2015-01-01

    Fabry disease is a rare X-linked lysosomal storage disorder of glycosphingolipids, caused by the partial or complete deficiency of the lysosomal enzyme alpha-galactosidase A (a-Gal A). The missense mutation pN215S usually causes a milder form of the disease with isolated cardiac involvement. We report a case of a male Fabry patient with the pN215S mutation and a generalized disease. He suffered a relapse in proteinuria which responded to increased doses of the administered recombinant enzyme. Individualization of enzyme replacement therapy must be considered in selected cases characterized by clinical deterioration.

  7. Artificial leucine rich repeats as new scaffolds for protein design.

    PubMed

    Baabur-Cohen, Hemda; Dayalan, Subashini; Shumacher, Inbal; Cohen-Luria, Rivka; Ashkenasy, Gonen

    2011-04-15

    The leucine rich repeat (LRR) motif that participates in many biomolecular recognition events in cells was suggested as a general scaffold for producing artificial receptors. We describe here the design and first total chemical synthesis of small LRR proteins, and their structural analysis. When evaluating the tertiary structure as a function of different number of repeating units (1-3), we were able to find that the 3-repeats sequence, containing 90 amino acids, folds into the expected structure.

  8. A C-terminal di-leucine motif controls plasma membrane expression of PMCA4b.

    PubMed

    Antalffy, Géza; Pászty, Katalin; Varga, Karolina; Hegedűs, Luca; Enyedi, Agnes; Padányi, Rita

    2013-12-01

    Recent evidences show that the localization of different plasma membrane Ca(2+) ATPases (PMCAs) is regulated in various complex, cell type-specific ways. Here we show that in low-density epithelial and endothelial cells PMCA4b localized mostly in intracellular compartments and its plasma membrane localization was enhanced upon increasing density of cells. In good correlation with the enhanced plasma membrane localization a significantly more efficient Ca(2+) clearance was observed in confluent versus non-confluent HeLa cell cultures expressing mCherry-PMCA4b. We analyzed the subcellular localization and function of various C-terminally truncated PMCA4b variants and found that a truncated mutant PMCA4b-ct24 was mostly intracellular while another mutant, PMCA4b-ct48, localized more to the plasma membrane, indicating that a protein sequence corresponding to amino acid residues 1158-1181 contained a signal responsible for the intracellular retention of PMCA4b in non-confluent cultures. Alteration of three leucines to alanines at positions 1167-1169 resulted in enhanced cell surface expression and an appropriate Ca(2+) transport activity of both wild type and truncated pumps, suggesting that the di-leucine-like motif (1167)LLL was crucial in targeting PMCA4b. Furthermore, upon loss of cell-cell contact by extracellular Ca(2+) removal, the wild-type pump was translocated to the early endosomal compartment. Targeting PMCA4b to early endosomes was diminished by the L(1167-69)A mutation, and the mutant pump accumulated in long tubular cytosolic structures. In summary, we report a di-leucine-like internalization signal at the C-tail of PMCA4b and suggest an internalization-mediated loss of function of the pump upon low degree of cell-cell contact.

  9. Characterization of a leucine aminopeptidase of Babesia gibsoni.

    PubMed

    Jia, H; Terkawi, M A; Aboge, G O; Goo, Y-K; Luo, Y; Li, Y; Yamagishi, J; Nishikawa, Y; Igarashi, I; Sugimoto, C; Fujisaki, K; Xuan, X

    2009-08-01

    Peptidases of parasitic protozoa are currently under intense investigation in order to identify novel virulence factors, drug targets, and vaccine candidates, except in Babesia. Leucine aminopeptidases in protozoa, such as Plasmodium and Leishmania, have been identified to be involved in free amino acid regulation. We report here the molecular and enzymatic characterization, as well as the localization of a leucine aminopeptidase, a member of the M17 cytosolic aminopeptidase family, from B. gibsoni (BgLAP). A functional recombinant BgLAP (rBgLAP) expressed in Escherichia coli efficiently hydrolysed synthetic substrates for aminopeptidase, a leucine substrate. Enzyme activity of the rBgLAP was found to be optimum at pH 8.0 and at 37 degrees C. The substrate profile was slightly different from its homologue in P. falciprum. The activity was also strongly dependent on metal divalent cations, and was inhibited by bestatin, which is a specific inhibitor for metalloprotease. These results indicated that BgLAP played an important role in free amino acid regulation.

  10. Identification and characterization of Paragonimus westermani leucine aminopeptidase.

    PubMed

    Song, Su-Min; Park, Joon-Hyung; Kim, Jin; Kim, Suk-Il; Hong, Yeon-Chul; Kong, Hyun-Hee; Chung, Dong-Il

    2008-09-01

    Paragonimus westermani is a tissue-invading trematode parasite that causes inflammatory lung disease as well as systemic infections including cerebral invasion in carnivorous mammals. While aminopeptidases play important roles in trematodes in the catabolism of host hemoglobin, an essential source of nutrient for the parasite, little is known about aminopeptidase in Paragonimus. Presently, we isolated a cDNA encoding a 58 kDa P. westermani leucine aminopeptidase (PwLAP). Deduced amino acid sequence of PwLAP exhibited significant sequence homology with LAP from Schistosoma spp. and Fasciola hepatica. Biochemical analysis of the recombinant PwLAP protein demonstrated preferential substrate specificity for Leu-NHMec and inhibition by EDTA, 1,10-phenanthroline, and bestatin, which are conserved characteristics of the M17 family of leucine aminopeptidase. PwLAP exhibited relatively higher enzyme activity in the presence of Mn2+ compared to Schistosoma mansoni LAP. Based on the biochemical properties and immunohistochemical analysis, PwLAP is concluded to represent a leucine aminopeptidase. The enzyme is most likely responsible for the catabolism of host hemoglobin, and, hence, represents a potential target of Paragonimus chemotherapy.

  11. Human circulating plasma DNA significantly decreases while lymphocyte DNA damage increases under chronic occupational exposure to low-dose gamma-neutron and tritium β-radiation.

    PubMed

    Korzeneva, Inna B; Kostuyk, Svetlana V; Ershova, Liza S; Osipov, Andrian N; Zhuravleva, Veronika F; Pankratova, Galina V; Porokhovnik, Lev N; Veiko, Natalia N

    2015-09-01

    The blood plasma of healthy people contains cell-fee (circulating) DNA (cfDNA). Apoptotic cells are the main source of the cfDNA. The cfDNA concentration increases in case of the organism's cell death rate increase, for example in case of exposure to high-dose ionizing radiation (IR). The objects of the present research are the blood plasma and blood lymphocytes of people, who contacted occupationally with the sources of external gamma/neutron radiation or internal β-radiation of tritium N = 176). As the controls (references), blood samples of people, who had never been occupationally subjected to the IR sources, were used (N = 109). With respect to the plasma samples of each donor there were defined: the cfDNA concentration (the cfDNA index), DNase1 activity (the DNase1 index) and titre of antibodies to DNA (the Ab DNA index). The general DNA damage in the cells was defined (using the Comet assay, the tail moment (TM) index). A chronic effect of the low-dose ionizing radiation on a human being is accompanied by the enhancement of the DNA damage in lymphocytes along with a considerable cfDNA content reduction, while the DNase1 content and concentration of antibodies to DNA (Ab DNA) increase. All the aforementioned changes were also observed in people, who had not worked with the IR sources for more than a year. The ratio cfDNA/(DNase1×Ab DNA × TM) is proposed to be used as a marker of the chronic exposure of a person to the external low-dose IR. It was formulated the assumption that the joint analysis of the cfDNA, DNase1, Ab DNA and TM values may provide the information about the human organism's cell resistivity to chronic exposure to the low-dose IR and about the development of the adaptive response in the organism that is aimed, firstly, at the effective cfDNA elimination from the blood circulation, and, secondly - at survival of the cells, including the cells with the damaged DNA.

  12. (n-3) Long chain PUFA dose-dependently increase oxygen utilization efficiency and inhibit arrhythmias after saturated fat feeding in rats.

    PubMed

    Pepe, Salvatore; McLennan, Peter L

    2007-11-01

    Fish oil (FO) modifies cardiac membrane phospholipid fatty acid composition to confer increased efficiency of oxygen utilization and antiarrhythmic effects. We tested the capacity of low-dose increments of FO, rich in (n-3) PUFA, to reverse the detrimental pro-arrhythmic and inefficient oxygen usage effects of dietary saturated fat (SAT) [including high ratio of (n-6) PUFA:(n-3) PUFA] during ischemia and reperfusion. Wistar rats were fed an SAT-enriched diet (15.3% fat, including 12% SAT, added by weight) for 6 wk and were then divided into 4 groups (n = 10/group) fed that diet or a 12% fat diet containing 3, 6, or 12% FO in place of SAT for 6 wk. Paced (300/min), erythrocyte-perfused isolated working hearts were subjected to low coronary flow ischemia (15 min) and were then reperfused. At normoxic baseline, external work capacity increased marginally at 6 and 12% FO; however, marked dose-related reductions in oxygen consumption were evident due to FO-dependent reduction in oxygen-energy utilization efficiency and associated reductions in coronary flow and oxygen extraction. Postischemic recovery resulted in lower oxygen consumption, greater oxygen-energy utilization efficiency, reduced coronary release of creatine kinase, and reduced incidence of arrhythmias in all FO groups compared with the SAT group. FO at a dose as low as 3% of total fat dietary supplement effectively reversed the high oxygen requirements and pro-arrhythmic effects of a SAT-rich diet even with continued consumption of SAT (9%) in this ex vivo animal model.

  13. Leucine supplementation does not affect protein turnover and impairs the beneficial effects of endurance training on glucose homeostasis in healthy mice.

    PubMed

    Costa Júnior, José M; Rosa, Morgana R; Protzek, André O; de Paula, Flávia M; Ferreira, Sandra M; Rezende, Luiz F; Vanzela, Emerielle C; Zoppi, Cláudio C; Silveira, Leonardo R; Kettelhut, Isis C; Boschero, Antonio C; de Oliveira, Camila A M; Carneiro, Everardo M

    2015-04-01

    Endurance exercise training as well as leucine supplementation modulates glucose homeostasis and protein turnover in mammals. Here, we analyze whether leucine supplementation alters the effects of endurance exercise on these parameters in healthy mice. Mice were distributed into sedentary (C) and exercise (T) groups. The exercise group performed a 12-week swimming protocol. Half of the C and T mice, designated as the CL and TL groups, were supplemented with leucine (1.5 % dissolved in the drinking water) throughout the experiment. As well known, endurance exercise training reduced body weight and the retroperitoneal fat pad, increased soleus mass, increased VO2max, decreased muscle proteolysis, and ameliorated peripheral insulin sensitivity. Leucine supplementation had no effect on any of these parameters and worsened glucose tolerance in both CL and TL mice. In the soleus muscle of the T group, AS-160(Thr-642) (AKT substrate of 160 kDa) and AMPK(Thr-172) (AMP-Activated Protein Kinase) phosphorylation was increased by exercise in both basal and insulin-stimulated conditions, but it was reduced in TL mice with insulin stimulation compared with the T group. Akt phosphorylation was not affected by exercise but was lower in the CL group compared with the other groups. Leucine supplementation increased mTOR phosphorylation at basal conditions, whereas exercise reduced it in the presence of insulin, despite no alterations in protein synthesis. In trained groups, the total FoxO3a protein content and the mRNA for the specific isoforms E2 and E3 ligases were reduced. In conclusion, leucine supplementation did not potentiate the effects of endurance training on protein turnover, and it also reduced its positive effects on glucose homeostasis.

  14. A comparison between augmentation with olanzapine and increased risperidone dose in acute schizophrenia patients showing early non-response to risperidone.

    PubMed

    Hatta, Kotaro; Otachi, Taro; Sudo, Yasuhiko; Kuga, Hironori; Takebayashi, Hiroshi; Hayashi, Hideaki; Ishii, Ryusuke; Kasuya, Masataka; Hayakawa, Tatsuro; Morikawa, Fumiyoshi; Hata, Kazuya; Nakamura, Mitsuru; Usui, Chie; Nakamura, Hiroyuki; Hirata, Toyoaki; Sawa, Yutaka

    2012-07-30

    We examined whether augmentation with olanzapine would be superior to increased risperidone dose among acute schizophrenia patients showing early non-response to risperidone. We performed a rater-blinded, randomized controlled trial at psychiatric emergency sites. Eligible patients were newly admitted patients with acute schizophrenia. Early response was defined as Clinical Global Impressions-Improvement Scale score ≤3 following 2 weeks of treatment. Early non-responders were allocated to receive either augmentation with olanzapine (RIS+OLZ group) or increased risperidone dose (RIS+RIS group). The 78 patients who completed 2 weeks of treatment were divided into 52 early responders to risperidone and 26 early non-responders to risperidone (RIS+OLZ group, n=13; RIS+RIS group, n=13). No difference in the achievement of ≥50% improvement in Positive and Negative Syndrome Scale total score was observed between RIS+OLZ and RIS+RIS groups. Although time to treatment discontinuation for any cause was significantly shorter in the RIS+RIS group (6.8 weeks [95% confidence interval, 5.2-8.4]) than in early responders to risperidone (8.6 weeks [7.9-9.3]; P=0.018), there was no significant difference between the RIS+OLZ group (7.9 weeks [6.3-9.5]) and early responders to risperidone. Secondary outcomes justify the inclusion of augmentation arms in additional, larger studies comparing strategies for early non-responders.

  15. Chronic exposure to low doses of lipopolysaccharide and high-fat feeding increases body mass without affecting glucose tolerance in female rats

    PubMed Central

    Dudele, Anete; Fischer, Christina W; Elfving, Betina; Wegener, Gregers; Wang, Tobias; Lund, Sten

    2015-01-01

    Obesity-related inflammation may have a causal role in the development of diabetes and insulin resistance, and studies using animal models of chronic experimental endotoxemia have shown the link. However, many studies use only males, and much less is known about the role of obesity-related inflammation in females. Therefore, we addressed how experimentally induced chronic inflammation affects body mass, energy intake, and glucose metabolism in female rats. Adult female Sprague Dawley rats were instrumented with slow release pellets that delivered a constant daily dose of 53 or 207 μg of lipopolysaccharide (LPS) per rat for 60 days. Control rats were instrumented with vehicle pellets. Due to inflammatory nature of high-fat diet (HFD) half of the rats received HFD (60% of calories from lard), while the other half remained on control diet to detect possible interactions between two modes of induced inflammation. Our results showed that chronic LPS administration increased female rat body mass and calorie intake in a dose-dependent manner, and that HFD further exacerbated these effects. Despite these effects, no effects of LPS and HFD were evident on female rat glucose metabolism. Only LPS elevated expression of inflammatory markers in the hypothalamus. To conclude, female rats respond to experimentally induced chronic inflammation by increasing body mass, but do not develop glucose intolerance in the given period of time. PMID:26537342

  16. Leucine is a major regulator of muscle protein synthesis in neonates

    PubMed Central

    Columbus, Daniel A; Fiorotto, Marta L; Davis, Teresa A

    2014-01-01

    Approximately 10% of infants born in the United States are of low birth weight. Growth failure during the neonatal period is a common occurrence in low birth weight infants due to their inability to tolerate full feeds, concerns about advancing amino acid supply, and high nutrient requirements for growth. An improved understanding of the nutritional regulation of growth during this critical period of development is vital for the development of strategies to improve lean growth. Past studies with animal models have demonstrated that muscle protein synthesis is increased substantially following a meal and that this increase is due to the postprandial rise in amino acids as well as insulin, which independently stimulate protein synthesis in a mammalian target of rapamycin (mTOR)-dependent manner. Further studies have elucidated that leucine, in particular, as well as its metabolites, α-ketoisocaproic acid and β-hydroxy-β-methylbutyrate, have unique anabolic properties. Supplementation with leucine, provided either parenterally or enterally, has been shown to enhance muscle protein synthesis in neonatal pigs, making it an ideal candidate for stimulating growth of low birth weight infants. PMID:25408462

  17. Human leucine zipper protein promotes hepatic steatosis via induction of apolipoprotein A-IV.

    PubMed

    Kang, Minsoo; Kim, Jeonghan; An, Hyoung-Tae; Ko, Jesang

    2017-02-28

    The molecular mechanism of stress-induced hepatic steatosis is not well known. Human leucine zipper protein (LZIP) regulates the expression of genes involved in inflammation, cell migration, and stress response. The aim of this study was to determine the regulatory role of LZIP in stress-induced hepatic steatosis. We used a microarray analysis to identify LZIP-induced genes involved in hepatic lipid metabolism. LZIP increased the expression of apolipoprotein A-IV (APOA4) mRNA. In the presence of stress inducer, APOA4 promoter analysis was performed and LZIP-induced lipid accumulation was monitored in mouse primary cells and human tissues. Under Golgi stress conditions, LZIP underwent proteolytic cleavage and was phosphorylated by AKT to protect against proteasome degradation. The stabilized N-terminal LZIP was translocated to the nucleus, where it directly bound to the APOA4 promoter, leading to APOA4 induction. LZIP-induced APOA4 expression resulted in increased absorption of surrounding free fatty acids. LZIP also promoted hepatic steatosis in mouse liver. Both LZIP and APOA4 were highly expressed in human steatosis samples. Our findings indicate that LZIP is a novel modulator of APOA4 expression and hepatic lipid metabolism. LZIP might be a therapeutic target for developing treatment strategies for hepatic steatosis and related metabolic diseases.-Kang, M., Kim, J., An, H.-T., Ko, J. Human leucine zipper protein promotes hepatic steatosis via induction of apolipoprotein A-IV.

  18. Chronic cigarette smoke exposure increases the pulmonary retention and radiation dose of {sup 239}Pu inhaled as {sup 239}PuO{sub 2} by F344 rats

    SciTech Connect

    Finch, G.L.; Lundgren, D.L.; Barr, E.B.; Chen, B.T.; Griffith, W.C.; Hobbs, C.H.; Hoover, M.D.; Nikula, K.J.; Mauderly, J.L.

    1998-12-01

    As a portion of a study to examine how chronic cigarette smoke exposure might alter the risk of lung tumors from inhaled {sup 239}PuO{sub 2} in rats, the effects of smoke exposure on alpha-particle lung dosimetry over the life-span of exposed rats were determined. Male and female rats were exposed to inhaled {sup 239}PuO{sub 2} alone or in combination with cigarette smoke. Animals exposed to filtered air along served as controls for the smoke exposure. Whole-body exposure to mainstream smoke diluted to concentrations of either 100 or 250 mg total particulate matter m{sup {minus}3} began at 6 wk of age and continued for 6 h d{sup {minus}1}, 5 d wk{sup {minus}1}, for 30 mo. A single, pernasal, acute exposure to {sup 239}PuO{sub 2} was given to all rats at 12 wk of age. Exposure to cigarette smoke caused decreased body weight gains in a concentration dependent manner. Lung-to-body weight ratios were increased in smoke-exposed rats. Rats exposed to cigarette smoke before the {sup 239}PuO{sub 2} exposure deposited less {sup 239}Pu in the lung than did controls. Except for male rats exposed to LCS, exposure to smoke retarded the clearance of {sup 239}Pu from the lung compared to control rats through study termination at 870 d after {sup 239}PuO{sub 2} exposure. Radiation doses to lungs were calculated by sex and by exposure group for rats on study for at least 360 d using modeled body weight changes, lung-to-body weight ratios, and standard dosimetric calculations. For both sexes, estimated lifetime radiation doses from the time of {sup 239}PuO{sub 2} exposure to death were 3.8 Gy, 4.4 Gy, or 6.7 Gy for the control, LCS, or HCS exposure groups, respectively. Assuming an approximately linear dose-response relationship between radiation dose and lung neoplasm incidence, approximate increases of 20% or 80% in tumor incidence over controls would be expected in rats exposed to {sup 239}PuO{sub 2} and LCS or {sup 239}PuO{sub 2} and HCS, respectively.

  19. Increased Biological Effective Dose of Radiation Correlates with Prolonged Survival of Patients with Limited-Stage Small Cell Lung Cancer: A Systematic Review

    PubMed Central

    Xu, Xiao; Wang, Bing; Wu, Kan; Deng, Qinghua; Xia, Bing; Ma, Shenglin

    2016-01-01

    Objective Thoracic radiotherapy (TRT) is a critical component of the treatment of limited-stage small cell lung cancer (LS-SCLC). However, the optimal radiation dose/fractionation remains elusive. This study reviewed current evidence and explored the dose-response relationship in patients with LS-SCLC who were treated with radiochemotherapy. Materials and Methods A quantitative analysis was performed through a systematic search of PubMed, Web of Science, and the Cochrane Library. The correlations between the biological effective dose (BED) and median overall survival (mOS), median progression-free survival (mPFS), 1-, 3-, and 5-year overall survival (OS) as well as local relapse (LR) were evaluated. Results In all, 2389 patients in 19 trials were included in this study. Among these 19 trials, seven were conducted in Europe, eight were conducted in Asia and four were conducted in the United States. The 19 trials that were included consisted of 29 arms with 24 concurrent and 5 sequential TRT arms. For all included studies, the results showed that a higher BED prolonged the mOS (R2 = 0.198, p<0.001) and the mPFS (R2 = 0.045, p<0.001). The results also showed that increased BED improved the 1-, 3-, and 5-year OS. A 10-Gy increment added a 6.3%, a 5.1% and a 3.7% benefit for the 1-, 3-, and 5-year OS, respectively. Additionally, BED was negatively correlated with LR (R2 = 0.09, p<0.001). A subgroup analysis of concurrent TRT showed that a high BED prolonged the mOS (p<0.001) and the mPFS (p<0.001), improved the 1-, 3-, and 5-year OS (p<0.001) and decreased the rate of LR (p<0.001). Conclusion This study showed that an increased BED was associated with improved OS, PFS and decreased LR in patients with LS-SCLC who were treated with combined chemoradiotherapy, which indicates that the strategy of radiation dose escalation over a limited time frame is worth exploring in a prospective clinical trial. PMID:27227819

  20. Double dose plateletpheresis by continuous and intermittent flow devices increases platelet-neutrophil complex formation in healthy donors without noticeable neutrophil activation.

    PubMed

    Bilgin, Aynur Ugur; Karadogan, Ihsan; Yilmaz, Ferahnaz Gencay; Undar, Levent

    2007-02-01

    Several reports have demonstrated that during a single plateletpheresis procedure, platelets may form heterotypic aggregates which may predispose certain donors to thrombotic complications. In this study, changes in the expression of neutrophil adhesion molecules (CD11b/CD18, CD50/54, CD62L) and platelet-neutrophil complex (PNC) formation were investigated by a flow cytometric method in healthy donors following a double dose plateletpheresis (DDP) procedure. Our results show that DDP which are carried out by the Fresenius AS.TEC 204 and Haemonetics MCS+ cause a significant increase in PNC formation in donors. Additionally, the Fresenius AS.TEC 204 device caused a decrease in CD62L expression which is a sign of mild neutrophil activation. Although the clinical significance of these laboratory changes is not clear, the occurrence of neutrophil activation and increased PNC formation might predispose certain donors to thrombotic complications following DDP.

  1. Protective effects of leucine on redox status and mitochondrial-related gene abundance in the jejunum of intrauterine growth-retarded piglets during early weaning period.

    PubMed

    Huang, Qiang; Xu, Wen; Bai, Kai-Wen; He, Jin-Tian; Ahmad, Hussain; Zhou, Le; Zhang, Li-Li; Wang, Tian

    2017-04-01

    The aim of this study was to investigate the effects of dietary supplementation with 0.35% l-leucine on redox status and gene abundance relating to mitochondrial biogenesis and function in the jejunum of intrauterine growth-retarded (IUGR) piglets during early weaning period. According to a 2 × 2 factorial arrangement, 16 IUGR and 16 normal body weight (NBW) piglets were fed a basal diet without l-leucine supplementation or a basal diet plus 0.35% l-leucine supplementation from the age of 14 to 35 d. The results showed that compared with NBW piglets, IUGR piglets had a lower (p < 0.05) jejunal DNA concentration, a reduced (p < 0.05) manganese superoxide dismutase (MnSOD) and total antioxidant capability (T-AOC) activities and mitochondrial DNA content in the jejunum. Leucine supplementation increased (p < 0.05) MnSOD and T-AOC activities and decreased (p < 0.05) the malondialdehyde content in the jejunum of IUGR piglets. The mRNA gene abundance of nuclear respiratory factor-1 (NRF1), mitochondrial transcription factor A (TFAM), ATP synthase (ATPs), cytochrome c oxidase V (CcOX V), cytochrome c and glucokinase in the jejunum of IUGR piglets was reduced (p < 0.05) compared with NBW piglets. However, NRF1, peroxisome proliferation-activated receptor gamma coactivator-1 alpha, TFAM, ATPs and CcOX I mRNA gene abundance in the jejunum of IUGR piglets were increased (p < 0.05) by diets supplemented with leucine. These data indicate that leucine supplementation has therapeutic potential for attenuating intestinal oxidative stress and mitochondrial dysfunction in IUGR piglets during the early period of life via increasing enzyme activities and up-regulating mRNA gene abundance.

  2. Sorghum bran in the diet dose dependently increased the excretion of catechins and microbial-derived phenolic acids in female rats.

    PubMed

    Gu, Liwei; House, Suzanne E; Rooney, Lloyd; Prior, Ronald L

    2007-06-27

    Sorghum bran is concentrated with procyanidins (predominately polymers), which may be beneficial for health in humans; however, the bioavailability of procyanidins is not well-understood. Female Sprague-Dawley rats were fed an AIN93G diet containing 0, 5, 10, 20, or 40% Hi-tannin sorghum bran (n = 5-7 for each group) for 50 days. Sorghum bran contained 23.3 mg/g of procyanidins. The urinary excretions of catechin, epicatechin, methylated catechins, and phenolic acids were analyzed using liquid chromatography-tandem mass spectrometry. Sorghum bran dose dependently increased the urinary excretion of catechin (0-2.2 nmol/day) and 3'-O-methylcatechin (0-9.5 nmol/day). Their serum concentrations also increased with dose (range of 0-14 nM for 3'-O-methylcatechin). Among the 14 phenolic acids analyzed, 3,4-dihydroxybenzoic acid, 3-methoxy-4-hydroxybenzoic acid, and 4-hydroxyphenylacetic acid dominated in the serum (1.8-8 micromol/L). In the urine, 3-methoxy-4-hydroxyphenylacetic acid, 3-hydroxyphenylacetic acid, and 3-hydroxyphenylpropionic acid dominated and their excretion increased significantly with the level of sorghum bran in the diet. The summed phenolic acid excretion was 0.8 micromol/day in the control group and increased to 23 micromol/day for 40% sorghum bran group. The hippuric acid excretion ranged from 2.2 to 16.2 micromol/day and peaked in the 10% sorghum bran group. On the basis of chromic oxide, a nonabsorbable marker, total procyanidins and polymers disappeared progressively, and significant degradation occurred in the cecum and colon. Catechins and procyanidins in sorghum were bioavailable; however, bacteria-derived phenolic acids were the predominant metabolites of procyanidins. Procyanidins degraded in the gastrointestinal tract. Depolymerization was not observed.

  3. Development of inducible leucine-rich repeat kinase 2 (LRRK2) cell lines for therapeutics development in Parkinson's disease.

    PubMed

    Huang, Liang; Shimoji, Mika; Wang, Juan; Shah, Salim; Kamila, Sukanta; Biehl, Edward R; Lim, Seung; Chang, Allison; Maguire-Zeiss, Kathleen A; Su, Xiaomin; Federoff, Howard J

    2013-10-01

    The pathogenic mechanism(s) contributing to loss of dopamine neurons in Parkinson's disease (PD) remain obscure. Leucine-rich repeat kinase 2 (LRRK2) mutations are linked, as a causative gene, to PD. LRRK2 mutations are estimated to account for 10% of familial and between 1 % and 3 % of sporadic PD. LRRK2 proximate single nucleotide polymorphisms have also been significantly associated with idiopathic/sporadic PD by genome-wide association studies. LRRK2 is a multidomain-containing protein and belongs to the protein kinase super-family. We constructed two inducible dopaminergic cell lines expressing either human-LRRK2-wild-type or human-LRRK2-mutant (G2019S). Phenotypes of these LRRK2 cell lines were examined with respect to cell viability, morphology, and protein function with or without induction of LRRK2 gene expression. The overexpression of G2019S gene promoted (1) low cellular metabolic activity without affecting cell viability, (2) blunted neurite extension, and (3) increased phosphorylation at S910 and S935. Our observations are consistent with reported general phenotypes in LRRK2 cell lines by other investigators. We used these cell lines to interrogate the biological function of LRRK2, to evaluate their potential as a drug-screening tool, and to investigate screening for small hairpin RNA-mediated LRRK2 G2019S gene knockdown as a potential therapeutic strategy. A proposed LRRK2 kinase inhibitor (i.e., IN-1) decreased LRRK2 S910 and S935 phosphorylation in our MN9DLRRK2 cell lines in a dose-dependent manner. Lentivirus-mediated transfer of LRRK2 G2019S allele-specific small hairpin RNA reversed the blunting of neurite extension caused by LRRK2 G2019S overexpression. Taken together, these inducible LRRK2 cell lines are suitable reagents for LRRK2 functional studies, and the screening of potential LRRK2 therapeutics.

  4. Increased expression of connective tissue growth factor (CTGF) in multiple organs after exposure of non-human primates (NHP) to lethal doses of radiation

    PubMed Central

    Zhang, Pei; Cui, Wanchang; Hankey, Kim G.; Gibbs, Allison M.; Smith, Cassandra P.; Taylor-Howell, Cheryl; Kearney, Sean R.; MacVittie, Thomas J.

    2015-01-01

    Exposure to sufficiently high doses of ionizing radiation is known to cause fibrosis in many different organs and tissues. Connective tissue growth factor (CTGF/CCN2), a member of the CCN family of matricellular proteins, plays an important role in the development of fibrosis in multiple organs. The aim of the present study was to quantify the gene and protein expression of CTGF in a variety of organs from non-human primates (NHP) that were previously exposed to potentially lethal doses of radiation. Tissues from non-irradiated NHP, and NHP exposed to whole thoracic lung irradiation (WTLI) or partial-body irradiation with 5% bone marrow sparing (PBI/BM5) were examined by real-time quantitative reverse transcription PCR, western blot, and immunohistochemistry. Expression of CTGF was elevated in the lung tissues of NHP exposed to WTLI relative to the lung tissues of the non-irradiated NHP. Increased expression of CTGF was also observed in multiple organs from NHP exposed to PBI/BM5 compared to non-irradiated NHP; these included the lung, kidney, spleen, thymus and liver. These irradiated organs also exhibited histological evidence of increased collagen deposition compared to the control tissues. There was significant correlation of CTGF expression with collagen deposition in the lung and spleen of NHP exposed to PBI/BM5. Significant correlations were observed between spleen and multiple organs on CTGF expression and collagen deposition respectively, suggesting possible crosstalk between spleen and other organs. Our data suggest that CTGF levels are increased in multiple organs after radiation exposure and that inflammatory cell infiltration may contribute to the elevated levels of CTGF in multiple organs. PMID:26425899

  5. Increased Expression of Connective Tissue Growth Factor (CTGF) in Multiple Organs After Exposure of Non-Human Primates (NHP) to Lethal Doses of Radiation.

    PubMed

    Zhang, Pei; Cui, Wanchang; Hankey, Kim G; Gibbs, Allison M; Smith, Cassandra P; Taylor-Howell, Cheryl; Kearney, Sean R; MacVittie, Thomas J

    2015-11-01

    Exposure to sufficiently high doses of ionizing radiation is known to cause fibrosis in many different organs and tissues. Connective tissue growth factor (CTGF/CCN2), a member of the CCN family of matricellular proteins, plays an important role in the development of fibrosis in multiple organs. The aim of the present study was to quantify the gene and protein expression of CTGF in a variety of organs from non-human primates (NHP) that were previously exposed to potentially lethal doses of radiation. Tissues from non-irradiated NHP and NHP exposed to whole thoracic lung irradiation (WTLI) or partial-body irradiation with 5% bone marrow sparing (PBI/BM5) were examined by real-time quantitative reverse transcription PCR, western blot, and immunohistochemistry. Expression of CTGF was elevated in the lung tissues of NHP exposed to WTLI relative to the lung tissues of the non-irradiated NHP. Increased expression of CTGF was also observed in multiple organs from NHP exposed to PBI/BM5 compared to non-irradiated NHP; these included the lung, kidney, spleen, thymus, and liver. These irradiated organs also exhibited histological evidence of increased collagen deposition compared to the control tissues. There was significant correlation of CTGF expression with collagen deposition in the lung and spleen of NHP exposed to PBI/BM5. Significant correlations were observed between spleen and multiple organs on CTGF expression and collagen deposition, respectively, suggesting possible crosstalk between spleen and other organs. These data suggest that CTGF levels are increased in multiple organs after radiation exposure and that inflammatory cell infiltration may contribute to the elevated levels of CTGF in multiple organs.

  6. Prebiotic fibres dose-dependently increase satiety hormones and alter Bacteroidetes and Firmicutes in lean and obese JCR:LA-cp rats.

    PubMed

    Parnell, Jill A; Reimer, Raylene A

    2012-02-01

    There is a growing interest in modulating gut microbiota with diet in the context of obesity. The purpose of the present study was to evaluate the dose-dependent effects of prebiotics (inulin and oligofructose) on gut satiety hormones, energy expenditure, gastric emptying and gut microbiota. Male lean and obese JCR:LA-cp rats were randomised to either of the following: lean 0 % fibre (LC), lean 10 % fibre (LF), lean 20 % fibre (LHF), obese 0 % fibre (OC), obese 10 % fibre (OF) or obese 20 % fibre (OHF). Body composition, gastric emptying, energy expenditure, plasma satiety hormone concentrations and gut microbiota (using quantitative PCR) were measured. Caecal proglucagon and peptide YY mRNA levels were up-regulated 2-fold in the LF, OF and OHF groups and 3-fold in the LHF group. Ghrelin O-acyltransferase mRNA levels were higher in obese v. lean rats and decreased in the OHF group. Plasma ghrelin response was attenuated in the LHF group. Microbial species measured in the Bacteroidetes division decreased, whereas those in the Firmicutes increased in obese v. lean rats and improved with prebiotic intake. Bifidobacterium and Lactobacillus increased in the OHF v. OC group. Bacteroides and total bacteria negatively correlated with percentage of body fat and body weight. Enterobacteriaceae increased in conjunction with glucose area under the curve (AUC) and glucagon-like peptide-1 AUC. Bacteroides and total bacteria correlated positively with ghrelin AUC yet negatively with insulin AUC and energy intake (P < 0·05). Several of the mechanisms through which prebiotics act (food intake, satiety hormones and alterations in gut microbiota) are regulated in a dose-dependent manner. The combined effects of prebiotics may have therapeutic potential for obesity.

  7. Prebiotic fibres dose-dependently increase satiety hormones and alter Bacteroidetes and Firmicutes in lean and obese JCR:LA-cp rats

    PubMed Central

    Parnell, Jill A.; Reimer, Raylene A.

    2013-01-01

    There is a growing interest in modulating gut microbiota with diet in the context of obesity. The purpose of the present study was to evaluate the dose-dependent effects of prebiotics (inulin and oligofructose) on gut satiety hormones, energy expenditure, gastric emptying and gut microbiota. Male lean and obese JCR:LA-cp rats were randomised to either of the following: lean 0 % fibre (LC), lean 10 % fibre (LF), lean 20 % fibre (LHF), obese 0 % fibre (OC), obese 10 % fibre (OF) or obese 20 % fibre (OHF). Body composition, gastric emptying, energy expenditure, plasma satiety hormone concentrations and gut microbiota (using quantitative PCR) were measured. Caecal proglucagon and peptide YY mRNA levels were up-regulated 2-fold in the LF, OF and OHF groups and 3-fold in the LHF group. Ghrelin O-acyltransferase mRNA levels were higher in obese v. lean rats and decreased in the OHF group. Plasma ghrelin response was attenuated in the LHF group. Microbial species measured in the Bacteroidetes division decreased, whereas those in the Firmicutes increased in obese v. lean rats and improved with prebiotic intake. Bifidobacterium and Lactobacillus increased in the OHF v. OC group. Bacteroides and total bacteria negatively correlated with percentage of body fat and body weight. Enterobacteriaceae increased in conjunction with glucose area under the curve (AUC) and glucagon-like peptide-1 AUC. Bacteroides and total bacteria correlated positively with ghrelin AUC yet negatively with insulin AUC and energy intake (P<0·05). Several of the mechanisms through which prebiotics act (food intake, satiety hormones and alterations in gut microbiota) are regulated in a dose-dependent manner. The combined effects of prebiotics may have therapeutic potential for obesity. PMID:21767445

  8. Zoledronic acid induces dose-dependent increase of antigen-specific CD8 T-cell responses in combination with peptide/poly-IC vaccine.

    PubMed

    Park, Hye-Mi; Cho, Hyun-Il; Shin, Chang-Ae; Shon, Hyun-Jung; Kim, Tai-Gyu

    2016-03-04

    Zoledronic acid (ZA) is used for treating osteoporosis and for preventing skeletal fractures in cancer patients suffering from myeloma and prostate cancer. It is also reported to directly induce cancer cell apoptosis and indirectly modulate T-cell immune response as an antitumor agent. In this study, the effect of ZA following peptide/polyinosinic-polycytidylic acid (poly-IC) vaccination was investigated in a murine tumor model. The combination of ZA with peptide/poly-IC vaccine showed a synergistic effect on the induction of antigen-specific CD8 T-cell response. Three consecutive intravenous administrations of ZA was defined to induce the highest CD8 T-cell response. Further, total splenocyte counts and antigen-specific CD8 T-cell response gradually increased depending on the dose of ZA. In tumor-bearing mice, ZA showed a dose-dependent decrease of growth and prolonged survival. Treatment with ZA only decreased the number of CD11b(+)Gr1(+) myeloid cells in blood. Our results demonstrate that the use of ZA could improve antitumor immune responses induced by the peptide/poly-IC vaccine.

  9. Vaccination of cattle with a high dose of BCG vaccine 3 weeks after experimental infection with Mycobacterium bovis increased the inflammatory response, but not tuberculous pathology.

    PubMed

    Buddle, Bryce M; Shu, Dairu; Parlane, Natalie A; Subharat, Supatsak; Heiser, Axel; Hewinson, R Glyn; Vordermeier, H Martin; Wedlock, D Neil

    2016-07-01

    A study was undertaken to determine whether BCG vaccination of cattle post-challenge could have an effect on a very early Mycobacterium bovis infection. Three groups of calves (n = 12/group) were challenged endobronchially with M. bovis and slaughtered 13 weeks later to examine for tuberculous lesions. One group had been vaccinated prophylactically with BCG Danish vaccine 21 weeks prior to challenge; a second group was vaccinated with a 4-fold higher dose of BCG Danish 3 weeks post-challenge and the third group, remained non-vaccinated. Vaccination prior to challenge induced only minimal protection with just a significant reduction in the lymph node lesion scores. Compared to the non-vaccinated group, BCG vaccination post-challenge produced no reduction in gross pathology and histopathology, but did result in significant increases in mRNA expression of pro-inflammatory mediators (IFN-γ, IL-12p40, IL-17A, IRF-5, CXCL9, CXCL10, iNOs, and TNF-α) in the pulmonary lymph nodes. Although there was no significant differences in the gross pathology and histopathology between the post-challenge BCG and non-vaccinated groups, the enhanced pro-inflammatory immune responses observed in the post-challenge BCG group suggest caution in the use of high doses of BCG where there is a possibility that cattle may be infected with M. bovis prior to vaccination.

  10. Increasing radiation dose improves immunotherapy outcome and prolongation of tumor dormancy in a subgroup of mice treated for advanced intracerebral melanoma.

    PubMed

    Smilowitz, Henry M; Micca, Peggy L; Sasso, Daniel; Wu, Qian; Dyment, Nathanial; Xue, Crystal; Kuo, Lynn

    2016-02-01

    Previously, we developed a clinically relevant therapy model for advanced intracerebral B16 melanomas in syngeneic mice combining radiation and immunotherapies. Here, 7 days after B16-F10-luc2 melanoma cells were implanted intracerebrally (D7), syngeneic mice with bioluminescent tumors that had formed (1E10(5) to 7E10(6) photons per minute (>1E10(6), large; <1E10(6), small) were segregated into large-/small-balanced subgroups. Then, mice received either radiation therapy alone (RT) or radiation therapy plus immunotherapy (RT plus IT) (single injection of mAbPC61 to deplete regulatory T cells followed by multiple injections of irradiated granulocyte macrophage colony stimulating factor transfected B16-F10 cells) (RT plus IT). Radiation dose was varied (15, 18.75 or 22.5 Gy, given on D8), while immunotherapy was provided similarly to all mice. The data support the hypothesis that increasing radiation dose improves the outcome of immunotherapy in a subgroup of mice. The tumors that were greatly delayed in beginning their progressive growth were bioluminescent in vivo-some for many months, indicating prolonged tumor "dormancy," in some cases presaging long-term cures. Mice bearing such tumors had far more likely received radiation plus immunotherapy, rather than RT alone. Radiotherapy is a very important adjunct to immunotherapy; the greater the tumor debulking by RT, the greater should be the benefit to tumor immunotherapy.

  11. Increasing the susceptibility of the rat 208F fibroblast cell line to radiation-induced apoptosis does not alter its clonogenic survival dose-response.

    PubMed Central

    Aldridge, D. R.; Arends, M. J.; Radford, I. R.

    1995-01-01

    Recent studies have suggested a correlation between the rate and incidence of apoptosis and the radiation response of particular cell lines. However, we found that increasing the rate of induction of apoptosis in the fibroblast line 208F, by transfecting it with human c-myc, did not lead to a change in its clonogenic survival dose-response for either gamma-irradiation or 125I-induced DNA damage. It was also found that expression of mutant (T24) Ha-ras in the 208F line appeared to decrease the level of apoptosis per mitosis after irradiation and inhibited the formation of nucleosomal ladders, but did not affect either the onset of the morphological features of apoptosis or the clonogenic survival dose-response of the cells to either gamma-irradiation or 125I-induced DNA damage. Our findings suggest that it may be incorrect to make predictions about the radiosensitivity of cells based only on knowledge of their mode of death. Images Figure 3 Figure 4 PMID:7880740

  12. Obese non-Hodgkin lymphoma patients tolerate full uncapped doses of chemotherapy with no increase in toxicity, and a similar survival to that seen in nonobese patients.

    PubMed

    Chan, Henry; Jackson, Sharon; McLay, Jessica; Knox, Angela; Lee, Jae; Wang, Sarah; Issa, Samar

    2016-11-01

    The aim of this study is to compare the risk of treatment-related toxicities and long-term survival between obese and nonobese patients with non-Hodgkin lymphoma when treated with full uncapped doses of R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) chemotherapy. A total of 133 patients and 733 cycles of chemotherapy were analyzed. Obese patients did not experience an increased risk of acute treatment-related toxicities (adjusted odds ratio 0.825, p = 0.197), or delayed toxicities (adjusted odds ratio 0.819, p = 0.779). In the subgroup of diffuse large B-cell lymphoma patients (n = 109), treatment response rate was similar between the two body mass index (BMI) groups, and obese patients tended to have superior overall and progression-free survivals, albeit not statistically significant. Full uncapped doses of R-CHOP chemotherapy administered to obese patients with non-Hodgkin lymphoma (NHL) are safe, well tolerated, and do not lead to inferior treatment response or long-term outcomes.

  13. Enhanced production of branched-chain amino acids by Gluconacetobacter europaeus with a specific regional deletion in a leucine responsive regulator.

    PubMed

    Akasaka, Naoki; Ishii, Yuri; Hidese, Ryota; Sakoda, Hisao; Fujiwara, Shinsuke

    2014-12-01

    Vinegar with increased amounts of branched-chain amino acids (BCAAs; valine, leucine and isoleucine) is favorable for human health as BCAAs decrease diet-induced obesity and hyperglycemia. To construct Gluconacetobacter europaeus which produces BCAAs, leucine responsive regulator (GeLrp) is focused and two Gelrp mutants were constructed. Wild-type KGMA0119 didn't produce significant amount of valine (0.13 mM) and leucine (0 mM) and strain KGMA7110 which lacks complete Gelrp accumulated valine (0.48 mM) and leucine (0.11 mM) but showed impaired growth, and it was fully restored in the presence of essential amino acids. Strain KGMA7203 was then constructed with a nonsense mutation at codon Trp132 in the Gelrp, which leads a specific deletion at an estimated ligand-sensing region in the C-terminal domain. KGMA7203 produced greater quantities of valine (0.80 mM) and leucine (0.26 mM) and showed the same growth characteristics as KGMA0119. mRNA levels of BCAAs biosynthesis genes (ilvI and ilvC) and probable BCAAs efflux pump (leuE) were determined by quantitative reverse-transcription PCR. Expression rates of ilvI and ilvC in the two Gelrp disruptants were greater than those in KGMA0119. leuE was highly expressed in KGMA7110 only, suggesting that the accumulation in KGMA7110 culture was caused by increased expression of the biosynthesis genes and abnormal enhanced export of amino acids resulting in impaired cell growth. In contrast, KGMA7203 would achieve the high level production through enhanced expression of the biosynthesis genes without enhancing that for the efflux pump. KGMA7203 was considered advantageous for production of vinegar with higher amounts of valine and leucine.

  14. Activation of mTORC1 by leucine is potentiated by branched-chain amino acids and even more so by essential amino acids following resistance exercise.

    PubMed

    Moberg, Marcus; Apró, William; Ekblom, Björn; van Hall, Gerrit; Holmberg, Hans-Christer; Blomstrand, Eva

    2016-06-01

    Protein synthesis is stimulated by resistance exercise and intake of amino acids, in particular leucine. Moreover, activation of mammalian target of rapamycin complex 1 (mTORC1) signaling by leucine is potentiated by the presence of other essential amino acids (EAA). However, the contribution of the branched-chain amino acids (BCAA) to this effect is yet unknown. Here we compare the stimulatory role of leucine, BCAA, and EAA ingestion on anabolic signaling following exercise. Accordingly, eight trained volunteers completed four sessions of resistance exercise during which they ingested either placebo, leucine, BCAA, or EAA (including the BCAA) in random order. Muscle biopsies were taken at rest, immediately after exercise, and following 90 and 180 min of recovery. Following 90 min of recovery the activity of S6 kinase 1 (S6K1) was greater than at rest in all four trials (Placebo<Leucineincrease in the EAA trial. At this same time point, phosphorylation of eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1) at Thr(37/46) was unaffected by supplementation, while that of Thr(46) alone exhibited a pattern similar to that of S6K1, being 18% higher with EAA than BCAA. However, after 180 min of recovery this difference between EAA and BCAA had disappeared, although with both these supplements the increases were still higher than with leucine (40%, P < 0.05) and placebo (100%, P < 0.05). In summary, EAA ingestion appears to stimulate translation initiation more effectively than the other supplements, although the results also suggest that this effect is primarily attributable to the BCAA.

  15. Crystal Engineering of l-Alanine with l-Leucine Additive using Metal-Assisted and Microwave-Accelerated Evaporative Crystallization

    PubMed Central

    2015-01-01

    In this work, we demonstrated that the change in the morphology of l-alanine crystals can be controlled with the addition of l-leucine using the metal-assisted and microwave accelerated evaporative crystallization (MA-MAEC) technique. Crystallization experiments, where an increasing stoichiometric amount of l-leucine is added to initial l-alanine solutions, were carried out on circular poly(methyl methacrylate) (PMMA) disks modified with a 21-well capacity silicon isolator and silver nanoparticle films using microwave heating (MA-MAEC) and at room temperature (control experiments). The use of the MA-MAEC technique afforded for the growth of l-alanine crystals with different morphologies up to ∼10-fold faster than those grown at room temperature. In addition, the length of l-alanine crystals was systematically increased from ∼380 to ∼2000 μm using the MA-MAEC technique. Optical microscope images revealed that the shape of l-alanine crystals was changed from tetragonal shape (without l-leucine additive) to more elongated and wire-like structures with the addition of the l-leucine additive. Further characterization of l-alanine crystals was undertaken by Fourier transform infrared (FT-IR) spectroscopy, Raman spectroscopy and powder X-ray diffraction (PXRD) measurements. In order to elucidate the growth mechanism of l-alanine crystals, theoretical simulations of l-alanine’s morphology with and without l-leucine additive were carried out using Materials Studio software in conjunction with our experimental data. Theoretical simulations revealed that the growth of l-alanine’s {011} and {120} crystal faces were inhibited due to the incorporation of l-leucine into these crystal faces in selected positions. PMID:24839404

  16. Crystal Engineering of l-Alanine with l-Leucine Additive using Metal-Assisted and Microwave-Accelerated Evaporative Crystallization.

    PubMed

    Mojibola, Adeolu; Dongmo-Momo, Gilles; Mohammed, Muzaffer; Aslan, Kadir

    2014-05-07

    In this work, we demonstrated that the change in the morphology of l-alanine crystals can be controlled with the addition of l-leucine using the metal-assisted and microwave accelerated evaporative crystallization (MA-MAEC) technique. Crystallization experiments, where an increasing stoichiometric amount of l-leucine is added to initial l-alanine solutions, were carried out on circular poly(methyl methacrylate) (PMMA) disks modified with a 21-well capacity silicon isolator and silver nanoparticle films using microwave heating (MA-MAEC) and at room temperature (control experiments). The use of the MA-MAEC technique afforded for the growth of l-alanine crystals with different morphologies up to ∼10-fold faster than those grown at room temperature. In addition, the length of l-alanine crystals was systematically increased from ∼380 to ∼2000 μm using the MA-MAEC technique. Optical microscope images revealed that the shape of l-alanine crystals was changed from tetragonal shape (without l-leucine additive) to more elongated and wire-like structures with the addition of the l-leucine additive. Further characterization of l-alanine crystals was undertaken by Fourier transform infrared (FT-IR) spectroscopy, Raman spectroscopy and powder X-ray diffraction (PXRD) measurements. In order to elucidate the growth mechanism of l-alanine crystals, theoretical simulations of l-alanine's morphology with and without l-leucine additive were carried out using Materials Studio software in conjunction with our experimental data. Theoretical simulations revealed that the growth of l-alanine's {011} and {120} crystal faces were inhibited due to the incorporation of l-leucine into these crystal faces in selected positions.

  17. The Role of Leucine-Rich Repeat Containing Protein 10 (LRRC10) in Dilated Cardiomyopathy

    PubMed Central

    Brody, Matthew J.; Lee, Youngsook

    2016-01-01

    Leucine-rich repeat containing protein 10 (LRRC10) is a cardiomyocyte-specific member of the Leucine-rich repeat containing (LRRC) protein superfamily with critical roles in cardiac function and disease pathogenesis. Recent studies have identified LRRC10 mutations in human idiopathic dilated cardiomyopathy (DCM) and Lrrc10 homozygous knockout mice develop DCM, strongly linking LRRC10 to the molecular etiology of DCM. LRRC10 localizes to the dyad region in cardiomyocytes where it can interact with actin and α-actinin at the Z-disc and associate with T-tubule components. Indeed, this region is becoming increasingly recognized as a signaling center in cardiomyocytes, not only for calcium cycling, excitation-contraction coupling, and calcium-sensitive hypertrophic signaling, but also as a nodal signaling hub where the myocyte can sense and respond to mechanical stress. Disruption of a wide range of critical structural and signaling molecules in cardiomyocytes confers susceptibility to cardiomyopathies in addition to the more classically studied mutations in sarcomeric proteins. However, the molecular mechanisms underlying DCM remain unclear. Here, we review what is known about the cardiomyocyte functions of LRRC10, lessons learned about LRRC10 and DCM from the Lrrc10 knockout mouse model, and discuss ongoing efforts to elucidate molecular mechanisms whereby mutation or absence of LRRC10 mediates cardiac disease. PMID:27536250

  18. A leucine zipper motif determines different functions in a DNA replication protein.

    PubMed Central

    Garcia de Viedma, D; Giraldo, R; Rivas, G; Fernández-Tresguerres, E; Diaz-Orejas, R

    1996-01-01

    RepA is the replication initiator protein of the Pseudomonas plasmid pPS10 and is also able to autoregulate its own synthesis. Here we report a genetic and functional analysis of a leucine zipper-like (LZ) motif located at the N-terminus of RepA. It is shown that the LZ motif modulates the equilibrium between monomeric and dimeric forms of the protein and that monomers of RepA interact with sequences at the origin of replication, oriV, while dimers are required for interactions of RepA at the repA promoter. Further, different residues of the LZ motif are seen to have different functional roles. Leucines at the d positions of the putative alpha-helix are relevant in the formation of RepA dimers required for transcriptional autoregulation. They also modulate other RepA-RepA interactions that result in cooperative binding of protein monomers to the origin of replication. The residues at the b/f positions of the putative helix play no relevant role in RepA-RepA interactions. These residues do not affect RepA autoregulation but do influence replication, as demonstrated by mutants that, without affecting binding to oriV, either increase the host range of the plasmid or are inactive in replication. It is proposed that residues in b/f positions play a relevant role in interactions between RepA and host replication factors. Images PMID:8631313

  19. Sestrin2 is a leucine sensor for the mTORC1 pathway.

    PubMed

    Wolfson, Rachel L; Chantranupong, Lynne; Saxton, Robert A; Shen, Kuang; Scaria, Sonia M; Cantor, Jason R; Sabatini, David M

    2016-01-01

    Leucine is a proteogenic amino acid that also regulates many aspects of mammalian physiology, in large part by activating the mTOR complex 1 (mTORC1) protein kinase, a master growth controller. Amino acids signal to mTORC1 through the Rag guanosine triphosphatases (GTPases). Several factors regulate the Rags, including GATOR1, aGTPase-activating protein; GATOR2, a positive regulator of unknown function; and Sestrin2, a GATOR2-interacting protein that inhibits mTORC1 signaling. We find that leucine, but not arginine, disrupts the Sestrin2-GATOR2 interaction by binding to Sestrin2 with a dissociation constant of 20 micromolar, which is the leucine concentration that half-maximally activates mTORC1. The leucine-binding capacity of Sestrin2 is required for leucine to activate mTORC1 in cells. These results indicate that Sestrin2 is a leucine sensor for the mTORC1 pathway.

  20. Leucine Leucine-37 Uses Formyl Peptide Receptor–Like 1 to Activate Signal Transduction Pathways, Stimulate Oncogenic Gene Expression, and Enhance the Invasiveness of Ovarian Cancer Cells

    PubMed Central

    Coffelt, Seth B.; Tomchuck, Suzanne L.; Zwezdaryk, Kevin J.; Danka, Elizabeth S.; Scandurro, Aline B.

    2009-01-01

    Emerging evidence suggests that the antimicrobial peptide, leucine leucine-37 (LL-37), could play a role in the progression of solid tumors. LL-37 is expressed as the COOH terminus of human cationic antimicrobial protein-18 (hCAP-18) in ovarian, breast, and lung cancers. Previous studies have shown that the addition of LL-37 to various cancer cell lines in vitro stimulates proliferation, migration, and invasion. Similarly, overexpression of hCAP-18/LL-37 in vivo accelerates tumor growth. However, the receptor or receptors through which these processes are mediated have not been thoroughly examined. In the present study, expression of formyl peptide receptor–like 1 (FPRL1) was confirmed on ovarian cancer cells. Proliferation assays indicated that LL-37 does not signal through a G protein–coupled receptor, such as FPRL1, to promote cancer cell growth. By contrast, FPRL1 was required for LL-37–induced invasion through Matrigel. The peptide stimulated mitogen-activated protein kinase and Janus-activated kinase/signal transducers and activators of transcription signaling cascades and led to the significant activation of several transcription factors, through both FPRL1-dependent and FPRL1-independent pathways. Likewise, expression of some LL-37–stimulated genes was attenuated by the inhibition of FPRL1. Increased expression of CXCL10, EGF, and PDGF-BB as well as other soluble factors was confirmed from conditioned medium of LL-37–treated cells. Taken together, these data suggest that LL-37 potentiates a more aggressive behavior from ovarian cancer cells through its interaction with FPRL1. PMID:19491199

  1. Leucine leucine-37 uses formyl peptide receptor-like 1 to activate signal transduction pathways, stimulate oncogenic gene expression, and enhance the invasiveness of ovarian cancer cells.

    PubMed

    Coffelt, Seth B; Tomchuck, Suzanne L; Zwezdaryk, Kevin J; Danka, Elizabeth S; Scandurro, Aline B

    2009-06-01

    Emerging evidence suggests that the antimicrobial peptide, leucine leucine-37 (LL-37), could play a role in the progression of solid tumors. LL-37 is expressed as the COOH terminus of human cationic antimicrobial protein-18 (hCAP-18) in ovarian, breast, and lung cancers. Previous studies have shown that the addition of LL-37 to various cancer cell lines in vitro stimulates proliferation, migration, and invasion. Similarly, overexpression of hCAP-18/LL-37 in vivo accelerates tumor growth. However, the receptor or receptors through which these processes are mediated have not been thoroughly examined. In the present study, expression of formyl peptide receptor-like 1 (FPRL1) was confirmed on ovarian cancer cells. Proliferation assays indicated that LL-37 does not signal through a G protein-coupled receptor, such as FPRL1, to promote cancer cell growth. By contrast, FPRL1 was required for LL-37-induced invasion through Matrigel. The peptide stimulated mitogen-activated protein kinase and Janus-activated kinase/signal transducers and activators of transcription signaling cascades and led to the significant activation of several transcription factors, through both FPRL1-dependent and FPRL1-independent pathways. Likewise, expression of some LL-37-stimulated genes was attenuated by the inhibition of FPRL1. Increased expression of CXCL10, EGF, and PDGF-BB as well as other soluble factors was confirmed from conditioned medium of LL-37-treated cells. Taken together, these data suggest that LL-37 potentiates a more aggressive behavior from ovarian cancer cells through its interaction with FPRL1.

  2. Knockout of leucine aminopeptidase in Toxoplasma gondii using CRISPR/Cas9.

    PubMed

    Zheng, Jun; Jia, Honglin; Zheng, Yonghui

    2015-02-01

    Leucine aminopeptidases of the M17 peptidase family represent ideal drug targets for therapies directed against the pathogens Plasmodium, Babesia and Trypanosoma. Previously, we characterised Toxoplasma gondii leucine aminopeptidase and demonstrated its role in regulating the levels of free amino acids. In this study, we evaluated the potential of T. gondii leucine aminopeptidase as a drug target in T. gondii by a knockout method. Existing knockout methods for T. gondii have many drawbacks; therefore, we developed a new technique that takes advantage of the CRISPR/Cas9 system. We first chose a Cas9 target site in the gene encoding T. gondii leucine aminopeptidase and then constructed a knockout vector containing Cas9 and the single guide RNA. After transfection, single tachyzoites were cloned in 96-well plates by limiting dilution. Two transfected strains derived from a single clone were cultured in Vero cells, and then subjected to expression analysis by western blotting. The phenotypic analysis revealed that knockout of T. gondii leucine aminopeptidase resulted in inhibition of attachment/invasion and replication; both the growth and attachment/invasion capacity of knockout parasites were restored by complementation with a synonymously substituted allele of T. gondii leucine aminopeptidase. Mouse experiments demonstrated that T. gondii leucine aminopeptidase knockout somewhat reduced the pathogenicity of T. gondii. An enzymatic activity assay showed that T. gondii leucine aminopeptidase knockout reduced the processing of a leucine aminopeptidase-specific substrate in T. gondii. The absence of leucine aminopeptidase activity could be slightly compensated for in T. gondii. Overall, T. gondii leucine aminopeptidase knockout influenced the growth of T. gondii, but did not completely block parasite development, virulence or enzymatic activity. Therefore, we conclude that leucine aminopeptidase would be useful only as an adjunctive drug target in T. gondii.

  3. Systematic Review and Meta-Analysis of Randomised Trials to Ascertain Fatal Gastrointestinal Bleeding Events Attributable to Preventive Low-Dose Aspirin: No Evidence of Increased Risk

    PubMed Central

    Dolwani, Sunil; Graziano, J. Michael; Lanas, Angel; Longley, Marcus; Phillips, Ceri J.; Roberts, Stephen E.; Soon, Swee S.; Steward, Will

    2016-01-01

    Background Aspirin has been shown to lower the incidence and the mortality of vascular disease and cancer but its wider adoption appears to be seriously impeded by concerns about gastrointestinal (GI) bleeding. Unlike heart attacks, stroke and cancer, GI bleeding is an acute event, usually followed by complete recovery. We propose therefore that a more appropriate evaluation of the risk-benefit balance would be based on fatal adverse events, rather than on the incidence of bleeding. We therefore present a literature search and meta-analysis to ascertain fatal events attributable to low-dose aspirin. Methods In a systematic literature review we identified reports of randomised controlled trials of aspirin in which both total GI bleeding events and bleeds that led to death had been reported. Principal investigators of studies in which fatal events had not been adequately described were contacted via email and asked for further details. A meta-analyses was then performed to estimate the risk of fatal gastrointestinal bleeding attributable to low-dose aspirin. Results Eleven randomised trials were identified in the literature search. In these the relative risk (RR) of ‘major’ incident GI bleeding in subjects who had been randomised to low-dose aspirin was 1.55 (95% CI 1.33, 1.83), and the risk of a bleed attributable to aspirin being fatal was 0.45 (95% CI 0.25, 0.80). In all the subjects randomised to aspirin, compared with those randomised not to receive aspirin, there was no significant increase in the risk of a fatal bleed (RR 0.77; 95% CI 0.41, 1.43). Conclusions The majority of the adverse events caused by aspirin are GI bleeds, and there appears to be no valid evidence that the overall frequency of fatal GI bleeds is increased by aspirin. The substantive risk for prophylactic aspirin is therefore cerebral haemorrhage which can be fatal or severely disabling, with an estimated risk of one death and one disabling stroke for every 1,000 people taking aspirin

  4. Kinetics and conformational stability studies of recombinant leucine aminopeptidase.

    PubMed

    Hernández-Moreno, Ana V; Villaseñor, Francisco; Medina-Rivero, Emilio; Pérez, Néstor O; Flores-Ortiz, Luis F; Saab-Rincón, Gloria; Luna-Bárcenas, Gabriel

    2014-03-01

    Leucine aminopeptidase from Vibrio proteolyticus is a broad specificity N-terminal aminopeptidase that is widely used in pharmaceutical processes where the removal of N-terminal residues in recombinant proteins is required. We previously reported the expression of a heterologous construction of the mature protein fused to a 6-histidine tag that presents a reasonable refolding rate for its use at industrial level. Here, we investigate this recombinant leucine aminopeptidase (rLAP) to explain the gain of activity observed when incubated at 37 °C after its production. Unfolding transitions of rLAP as a function of urea concentration were monitored by circular dichroism (CD) and fluorescence (FL) spectroscopy exhibiting single transitions by both techniques. Free energy change for unfolding measured by CD and FL spectroscopy are 2.8 ± 0.4 and 3.7 ± 0.4 kcal mol(-1), respectively. Thermal stability conformation of rLAP is 2.6 ± 0.1 and 6.1 kcal mol(-1) for CD and Nano-Differential Scanning Calorimetry (Nano-DSC), respectively. Enzyme activity was assessed with L-leucine-p-nitroanilide (L-pNA) as substrate. The catalytic efficiency was 3.87 ± 0.10 min(-1) μM(-1) at 37 °C and pH 8.0. Kinetic and conformation studies show differences between the enzyme native and rLAP; however rLAP is selective and specific to remove N-terminal groups from amino acids.

  5. Histochemistry of leucine aminonaphthylamidase (LAN) in rainbow trout (Salmo gairdneri)

    USGS Publications Warehouse

    Bouck, Gerald R.

    1979-01-01

    The histochemistry of leucine aminonaphthylamidase (LAN) was studied in frozen tissue sections of rainbow trout both in yearling and adult fish. Age of fish had relatively little effect upon the results. The most intense LAN color production was in epithelial cells of midgut, pyloric ceca, hindgut, and in some segments of kidney tubules. Lower levels of LAN were evident in liver cells of Kupffer, and still lower or slight levels of LAN activity were found in blood cells, muscle, nerve, connective tissue, gonad, and pancreas. The results indicate that LAN might be useful in assessing histotoxicity to LAN-rich areas of the body.

  6. Histochemistry of leucine aminoaphthylamidase (LAN) in rainbow trout (Salmo gairdneri)

    USGS Publications Warehouse

    Bouck, Gerald R.

    1979-01-01

    The histochemistry of leucine aminonaphthylamidase (LAN) was studied in frozen tissue sections of rainbow trout both in yearling and adult fish. Age of fish had relatively little effect upon the results. The most intense LAN color production was in epithelial cells of midgut, pyloric ceca, hindgut, and in some segments of kidney tubules. Lower levels of LAN were evident in liver cells of Kupffer, and still lower or slight levels of LAN activity were found in blood cells, muscle, nerve, connective tissue, gonad, and pancreas. The results indicate that LAN might be useful in assessing histotoxicity to LAN-rich areas of the body.

  7. Novel leucine ureido derivatives as inhibitors of aminopeptidase N (APN).

    PubMed

    Ma, Chunhua; Jin, Kang; Cao, Jiangying; Zhang, Lei; Li, Xiaoguang; Xu, Wenfang

    2013-04-01

    Aminopeptidase N (APN/CD13) over expressed on tumor cells, plays a critical role in tumor invasion, metastasis, and tumor angiogenesis. Here we described the design, synthesis and preliminary activity studies of novel leucine ureido derivatives as aminopeptidase N (APN/CD13) inhibitors. The results showed that compound 8c had the most potent inhibitory activity against APN with the IC50 value to 0.06 ± 0.041 μM, which could be used for further anticancer agent research.

  8. "Leucine aminopeptidase" (neutral arylamidase) in sheep sera: improved resolution with gradient gel electrophoresis.

    PubMed

    Manwell, C; Baker, C M

    1986-01-01

    Electrophoretic resolution of the heterogeneity of sheep serum "leucine aminopeptidase" is greatly improved by the use of gradients of acrylamide polymer, together with enzyme localisation involving L-alanyl beta-naphthylamide and cobaltous ion. The improved resolution contradicts an earlier claim of the existence of only two patterns of individual variation in the heterogeneity of sheep serum "leucine aminopeptidase", with one pattern completely dominant to the other. While the sheep enzyme is unusual among mammalian serum "leucine aminopeptidases" in its complex heterogeneity, it does conform to the typical mammalian pattern of codominant individual variation. The complexity of sheep serum "leucine aminopeptidase" is useful in the study of sheep evolution.

  9. Dose-response relationship between weekly resistance training volume and increases in muscle mass: A systematic review and meta-analysis.

    PubMed

    Schoenfeld, Brad J; Ogborn, Dan; Krieger, James W

    2017-06-01

    The purpose of this paper was to systematically review the current literature and elucidate the effects of total weekly resistance training (RT) volume on changes in measures of muscle mass via meta-regression. The final analysis comprised 34 treatment groups from 15 studies. Outcomes for weekly sets as a continuous variable showed a significant effect of volume on changes in muscle size (P = 0.002). Each additional set was associated with an increase in effect size (ES) of 0.023 corresponding to an increase in the percentage gain by 0.37%. Outcomes for weekly sets categorised as lower or higher within each study showed a significant effect of volume on changes in muscle size (P = 0.03); the ES difference between higher and lower volumes was 0.241, which equated to a percentage gain difference of 3.9%. Outcomes for weekly sets as a three-level categorical variable (<5, 5-9 and 10+ per muscle) showed a trend for an effect of weekly sets (P = 0.074). The findings indicate a graded dose-response relationship whereby increases in RT volume produce greater gains in muscle hypertrophy.

  10. Comparative analysis of pharmacological treatments with N-acetyl-DL-leucine (Tanganil) and its two isomers (N-acetyl-L-leucine and N-acetyl-D-leucine) on vestibular compensation: Behavioral investigation in the cat.

    PubMed

    Tighilet, Brahim; Leonard, Jacques; Bernard-Demanze, Laurence; Lacour, Michel

    2015-12-15

    Head roll tilt, postural imbalance and spontaneous nystagmus are the main static vestibular deficits observed after an acute unilateral vestibular loss (UVL). In the UVL cat model, these deficits are fully compensated over 6 weeks as the result of central vestibular compensation. N-Acetyl-dl-leucine is a drug prescribed in clinical practice for the symptomatic treatment of acute UVL patients. The present study investigated the effects of N-acetyl-dl-leucine on the behavioral recovery after unilateral vestibular neurectomy (UVN) in the cat, and compared the effects of each of its two isomers N-acetyl-L-leucine and N-acetyl-D-leucine. Efficacy of these three drug treatments has been evaluated with respect to a placebo group (UVN+saline water) on the global sensorimotor activity (observation grids), the posture control (support surface measurement), the locomotor balance (maximum performance at the rotating beam test), and the spontaneous vestibular nystagmus (recorded in the light). Whatever the parameters tested, the behavioral recovery was strongly and significantly accelerated under pharmacological treatments with N-acetyl-dl-leucine and N-acetyl-L-leucine. In contrast, the N-acetyl-D-leucine isomer had no effect at all on the behavioral recovery, and animals of this group showed the same recovery profile as those receiving a placebo. It is concluded that the N-acetyl-L-leucine isomer is the active part of the racemate component since it induces a significant acceleration of the vestibular compensation process similar (and even better) to that observed under treatment with the racemate component only.

  11. Volumetric-modulated arc stereotactic body radiotherapy for prostate cancer: dosimetric impact of an increased near-maximum target dose and of a rectal spacer

    PubMed Central

    Naccarato, Stefania; Stavrev, Pavel; Stavreva, Nadejda; Fersino, Sergio; Giaj Levra, Niccolò; Mazzola, Rosario; Mancosu, Pietro; Scorsetti, Marta; Alongi, Filippo

    2015-01-01

    Objective: In volumetric-modulated arc therapy (VMAT) prostate stereotactic body radiotherapy (SBRT), dose coverage of the planning target volume (PTV) becomes challenging when the sparing of rectum, bladder and urethra is strictly pursued. Our current 35-Gy-in-five-fraction plans only assure 33.2 Gy to ≥95% PTV (V33.2PTV ≥ 95%). Looking for an improved V33.2PTV, increased near-maximum target dose (D2%) and prostate–rectum spacer insertion were tested. Methods: For 11 patients, two VMAT plans, with D2% ≤ 37.5 Gy (Hom) or D2% ≤ 40.2 Gy (Het), on each of two CT studies, before or after spacer insertion, were computed. All plans assured V33.2PTV ≥95%, and <1 cm3 of rectum, bladder and urethra receiving ≥35 Gy. By hypothesis testing, several dose–volume metrics for target coverage and rectal sparing were compared across the four groups of plans. The impact of spacer insertion on the fractions of rectum receiving more than 18, 28 and 32 Gy (VXr) was further tested by linear correlation analysis. Results: By hypothesis testing, the increased D2% was associated with improvements in target coverage, whereas spacer insertion was associated with improvements in both target coverage and rectal VXr. By linear correlation analysis, spacer insertion was related to the reductions in rectal VXr for X ≥ 28 Gy. Conclusion: A slightly increased D2% or the use of spacer insertion was each able to improve V33.2PTV. Their combined use assured V33.2PTV ≥ 98% to all our patients. Spacer insertion was further causative for improvements in rectal sparing. Advances in knowledge: For VMAT plans in prostate SBRT, the distinct dosimetric usefulness of increased D2% and of the use of spacer insertion were validated in terms of target coverage and rectal sparing. PMID:26235142

  12. GCN2 contributes to mTORC1 inhibition by leucine deprivation through an ATF4 independent mechanism

    PubMed Central

    Averous, Julien; Lambert-Langlais, Sarah; Mesclon, Florent; Carraro, Valérie; Parry, Laurent; Jousse, Céline; Bruhat, Alain; Maurin, Anne-Catherine; Pierre, Philippe; Proud, Christopher G.; Fafournoux, Pierre

    2016-01-01

    It is well known that the GCN2 and mTORC1 signaling pathways are regulated by amino acids and share common functions, in particular the control of translation. The regulation of GCN2 activity by amino acid availability relies on the capacity of GCN2 to sense the increased levels of uncharged tRNAs upon amino acid scarcity. In contrast, despite recent progress in the understanding of the regulation of mTORC1 by amino acids, key aspects of this process remain unsolved. In particular, while leucine is well known to be a potent regulator of mTORC1, the mechanisms by which this amino acid is sensed and control mTORC1 activity are not well defined. Our data establish that GCN2 is involved in the inhibition of mTORC1 upon leucine or arginine deprivation. However, the activation of GCN2 alone is not sufficient to inhibit mTORC1 activity, indicating that leucine and arginine exert regulation via additional mechanisms. While the mechanism by which GCN2 contributes to the initial step of mTORC1 inhibition involves the phosphorylation of eIF2α, we show that it is independent of the downstream transcription factor ATF4. These data point to a novel role for GCN2 and phosphorylation of eIF2α in the control of mTORC1 by certain amino acids. PMID:27297692

  13. High dose trans-10,cis-12 CLA increases lean body mass in hamsters, but elevates levels of plasma lipids and liver enzyme biomarkers.

    PubMed

    Liu, Xiaoran; Joseph, Shama V; Wakefield, Andrew P; Aukema, Harold M; Jones, Peter J H

    2012-01-01

    The current study examined the efficacy of graded doses of c9,t11 and t10,c12 CLA isomers on body composition, energy expenditure, hepatic and serum lipid liver biomarkers in hamsters. Animals (n = 105) were randomized to seven treatments (control, 1, 2, 3% of c9,t11; 1, 2, 3% of t10,c12) for 28 days. After 28 days treatment, 1-3% of t10,c12 lowered (p < 0.05) body fat mass compared to the control group. The 1-3% t10,c12 and 3% c9,t11 fed groups showed higher (p < 0.05) lean mass compared to other groups. We observed unfavorable changes in plasma total cholesterol and non-HDL cholesterol levels in animals fed with 3% t10,c12 CLA isomers. The 2%, 3% t10,c12 groups presented elevated (p < 0.05) ALT levels. The present data suggest that a diet enriched with more than 2% t10, c12 led to liver malfunction and poses unfavorable changes on plasma lipid profiles. The 1% t10,c12 CLA lowered (p < 0.05) body fat mass and increased (p < 0.05) lean body mass. The c9,t11 CLA has less potent actions than t10,c12 CLA. We conclude that the actions of CLA on energy and lipid metabolism are form and dose dependent in the hamster model.

  14. Radiation dose to the brachial plexus in nasopharyngeal carcinoma treated with intensity-modulated radiation therapy: An increased risk of an excessive dose to the brachial plexus adjacent to gross nodal disease

    PubMed Central

    FENG, GUOSHENG; LU, HEMING; LIANG, YUAN; CHEN, HUASHENG; SHU, LIUYANG; LU, SHUI; ZHU, JIANFANG; GAO, WEIWEI

    2012-01-01

    This retrospective study aimed to evaluate the dose to the brachial plexus in patients with nasopharyngeal carcinoma (NPC) treated with intensity-modulated radiation therapy (IMRT). Twenty-eight patients were selected and the brachial plexus was delineated retrospectively. Brachial plexus adjacent/not adjacent to nodes were defined and abbreviated as BPAN and BPNAN, respectively. Dose distribution was recalculated and a dose-volume histogram was generated based on the original treatment plan. The maximum dose to the left brachial plexus was 59.12–78.47 Gy, and the percentage of patients receiving the maximum dose exceeding 60, 66 and 70 Gy was 96.4, 57.1 and 25.0%, respectively; the maximum dose to the right brachial plexus was 59.74–80.31 Gy, and the percentage of patients exposed to a maximum dose exceeding 60, 66 and 70 Gy was 96.4, 64.3 and 39.3%, respectively. For the left brachial plexus, the maximum doses to the BPANs and the BPNANs were 72.84±3.91 and 64.81±3.47 Gy, respectively (p<0.001). For the right brachial plexus, the maximum doses to the BPANs and the BPNANs were 72.91±4.74 and 64.91±3.52 Gy, respectively (p<0.001). The difference between the left BPANs and the left BPNANs was statistically significant not only for V60 (3.60 vs. 1.01 cm3, p=0.028) but also for V66 (1.26 vs. 0.11 cm3, p=0.046). There were significant differences in V60 (3.68 vs. 1.16 cm3, p<0.001) and V66 (1.83 vs. 1.23 cm3, p=0.012) between the right BPANs and the right BPNANs. In conclusion, a large proportion of patients were exposed to the maximum dose to the brachial plexus exceeding the Radiation Therapy Oncology Group-recommended restraints when the brachial plexus was not outlined. The BPANs are at a significantly higher risk of receiving an excessive radiation dose when compared to the BPNANs. A further study is underway to test whether brachial plexus contouring assists in the dose reduction to the brachial plexus for IMRT optimization. PMID:22970028

  15. Sodium orthovanadate associated with pharmacological doses of ascorbate causes an increased generation of ROS in tumor cells that inhibits proliferation and triggers apoptosis

    SciTech Connect

    Günther, T-hat nia Mara Fischer; Kviecinski, Maicon Roberto; Baron, Carla Cristine; Felipe, Karina Bettega; Farias, Mirelle Sifroni; Ourique da Silva, Fabiana; Bücker, Nádia Cristina Falcão; Pich, Claus Tröger; Ferreira, Eduardo Antonio; Filho, Danilo Wilhelm; Verrax, Julien; Calderon, Pedro Buc; Pedrosa, Rozangela Curi

    2013-01-18

    Graphical abstract: -- Abstract: Pharmacological doses of ascorbate were evaluated for its ability to potentiate the toxicity of sodium orthovanadate (Na{sub 3}VO{sub 4}) in tumor cells. Cytotoxicity, inhibition of cell proliferation, generation of ROS and DNA fragmentation were assessed in T24 cells. Na{sub 3}VO{sub 4} was cytotoxic against T24 cells (EC{sub 50} = 5.8 μM at 24 h), but in the presence of ascorbate (100 μM) the EC{sub 50} fell to 3.3 μM. Na{sub 3}VO{sub 4} plus ascorbate caused a strong inhibition of cell proliferation (up to 20%) and increased the generation of ROS (4-fold). Na{sub 3}VO{sub 4} did not directly cleave plasmid DNA, at this aspect no synergism was found occurring between Na{sub 3}VO{sub 4} and ascorbate once the resulting action of the combination was no greater than that of both substances administered separately. Cells from Ehrlich ascites carcinoma-bearing mice were used to determine the activity of antioxidant enzymes, the extent of the oxidative damage and the type of cell death. Na{sub 3}VO{sub 4} alone, or combined with ascorbate, increased catalase activity, but only Na{sub 3}VO{sub 4} plus ascorbate increased superoxide dismutase activity (up to 4-fold). Oxidative damage on proteins and lipids was higher due to the treatment done with Na{sub 3}VO{sub 4} plus ascorbate (2–3-fold). Ascorbate potentiated apoptosis in tumor cells from mice treated with Na{sub 3}VO{sub 4}. The results indicate that pharmacological doses of ascorbate enhance the generation of ROS induced by Na{sub 3}VO{sub 4} in tumor cells causing inhibition of proliferation and apoptosis. Apoptosis induced by orthovanadate and ascorbate is closer related to inhibition on Bcl-xL and activation of Bax. Our data apparently rule out a mechanism of cell demise p53-dependent or related to Cdk2 impairment.

  16. Prophylactic activity of increasing doses of intravenous histamine in refractory migraine: Retrospective observations of a series of patients with migraine without aura

    PubMed Central

    Pietrini, Umberto; De Luca, Massimo; Del Bene, Enrico; De Cesaris, Francesco; Bertinotti, Luca; Colangelo, Nicola; Moggi Pignone, Alberto

    2004-01-01

    Background: Histamine is thought to play a pivotal role in the modulation of peripheral and central pain. The administration of increasing doses of histamine may lead to desensitization of receptors of histamine types 1 and 2, causing meningeal vasodilation, and to depletion of neuropeptides in the trigeminal ganglion, thus inhibiting the initiation of migraine. Objective: In this study, the efficacy and tolerability of increasing doses of IV histamine in migraine prophylaxis were investigated. Methods: This single-center, open-label, retrospective, controlled study was conducted at the Headache Center (Department of Internal Medicine, University of Florence, Villa Monna Tessa, Italy). Patients included in the study had 3 to 6 migraines without aura per month that were refractory to common symptomatic and prophylactic agents in the 6 months preceding the study. Patients were treated with IV histamine hydrochloride for 21 days starting with a dosage of 0.5 mg/d and increasing to 4.0 mg/d. To assess the efficacy of the treatment, these patients were matched for age; sex; and frequency, duration, and severity of attacks with untreated migraineurs. Clinical benefit was defined as ⩽ 1 migraine of mild intensity per month. Tolerability was assessed during the hospitalization period, and patients were instructed to contact the Headache Center to report any adverse effects after hospital discharge. Results: The histamine group comprised 47 patients (40 women, 7 men; mean [SD] age, 42.0 [8.6] years) and the control group comprised 23 patients (20 women, 3 men; mean [SD] age, 38.8 [8.4] years). The histamine-treated patients showed a clinical benefit lasting for a mean of 10.4 (4.2) months, while the patients in the control group showed a clinical benefit of 3.8 (1.9) months. The difference in the duration of the clinical benefit between the 2 groups was 6.6 months (95% CI, 5.15-7.99). Adverse effects consisted of flushing, heat sensation during infusion, headache, and

  17. Simultaneous determination of leucine, isoleucine and valine in Beagle dog plasma by HPLC-MS/MS and its application to a pharmacokinetic study.

    PubMed

    Wang, Ting; Xie, Huiru; Chen, Xu; Jiang, Xuehua; Wang, Ling

    2015-10-10

    Leucine (Leu), isoleucine (Ile) and valine (Val) are three branched-chain amino acids (BCAAs), which have been widely used as dietary supplements for professional athletes and patients with liver failure or catabolic diseases. To date, no pharmacokinetic studies of BCAAs in vivo useful for the assessment of clinical effect following daily intake has been reported. Thus in this study, an HPLC-MS/MS method for simultaneous determination of Leu, Ile and Val in Beagle dog plasma using homoarginine as the internal standard was developed and validated in terms of specificity, linearity, precision, accuracy, and stability. This assay method was then applied to a pharmacokinetic study of BCAAs in dogs following oral administration of 0.25 g/kg and 0.50 g/kg BCAAs. The HPLC-MS/MS method was found to be sensitive and reproducible for quantification of BCAAs in dog plasma and successfully applied to the pharmacokinetic study. All these BCAAs were well absorbed with a substantial increase in the plasma concentration after a baseline modification. No statistical significance was identified in different gender group and no drug accumulation was observed following multiple doses.

  18. Mesenchymal Stem Cells Within Gelatin/CaSO4 Scaffolds Treated Ex Vivo with Low Doses of BMP-2 and Wnt3a Increase Bone Regeneration.

    PubMed

    Aquino-Martínez, Rubén; Rodríguez-Carballo, Edgardo; Gámez, Beatriz; Artigas, Natalia; Carvalho-Lobato, Patricia; Manzanares-Céspedes, Maria Cristina; Rosa, Jose Luis; Ventura, Francesc

    2016-01-01

    The delivery of osteogenic factors is a proven therapeutic strategy to promote bone regeneration. Bone morphogenetic proteins (BMPs) constitute a family of cytokines with well-known osteogenic and bone regenerative abilities. However, clinical uses of BMPs require high doses that have been associated with complications such as osteolysis, ectopic bone formation, or hematoma formation. In the present work, we sought to improve bone tissue engineering through an approach that combines the use of bone marrow-derived mesenchymal stem cells (BMMSCs), composite scaffolds, and osteoinductive agents. We employed a composite gelatin/CaSO4 scaffold that allows for an early expansion of seeded BMMSCs, which is followed by an increased level of osteogenic differentiation after 10 days in culture. Furthermore, this scaffold enhanced bone formation by BMMSCs in a mouse model of critical-sized calvarial defect. More importantly, our results demonstrate that ex vivo pretreatment of BMMSCs with low amounts of BMP-2 (2 nM) and Wnt3a (50 ng/mL) for 24 h cooperatively increases the expression of osteogenic markers in vitro and bone regeneration in the critical-sized calvarial defect mouse model. These data provide a strong rationale for the development of an ex vivo cooperative use of BMP-2 and Wnt3a. Osteogenic factor cooperation might be applied to reduce the required amount of growth factors while obtaining higher therapeutic effects.

  19. Increased phytotoxic O3 dose accelerates autumn senescence in an O3-sensitive beech forest even under the present-level O3

    NASA Astrophysics Data System (ADS)

    Kitao, Mitsutoshi; Yasuda, Yukio; Kominami, Yuji; Yamanoi, Katsumi; Komatsu, Masabumi; Miyama, Takafumi; Mizoguchi, Yasuko; Kitaoka, Satoshi; Yazaki, Kenichi; Tobita, Hiroyuki; Yoshimura, Kenichi; Koike, Takayoshi; Izuta, Takeshi

    2016-09-01

    Ground-level ozone (O3) concentrations are expected to increase over the 21st century, especially in East Asia. However, the impact of O3 has not been directly assessed at the forest level in this region. We performed O3 flux-based risk assessments of carbon sequestration capacity in an old cool temperate deciduous forest, consisting of O3-sensitive Japanese beech (Fagus crenata), and in a warm temperate deciduous and evergreen forest dominated by O3-tolerant Konara oak (Quercus serrata) based on long-term CO2 flux observations. On the basis of a practical approach for a continuous estimation of canopy-level stomatal conductance (Gs), higher phytotoxic ozone dose above a threshold of 0 uptake (POD0) with higher Gs was observed in the beech forest than that in the oak forest. Light-saturated gross primary production, as a measure of carbon sequestration capacity of forest ecosystem, declined earlier in the late growth season with increasing POD0, suggesting an earlier autumn senescence, especially in the O3-sensitive beech forest, but not in the O3-tolerant oak forest.

  20. Increased phytotoxic O3 dose accelerates autumn senescence in an O3-sensitive beech forest even under the present-level O3

    PubMed Central

    Kitao, Mitsutoshi; Yasuda, Yukio; Kominami, Yuji; Yamanoi, Katsumi; Komatsu, Masabumi; Miyama, Takafumi; Mizoguchi, Yasuko; Kitaoka, Satoshi; Yazaki, Kenichi; Tobita, Hiroyuki; Yoshimura, Kenichi; Koike, Takayoshi; Izuta, Takeshi

    2016-01-01

    Ground-level ozone (O3) concentrations are expected to increase over the 21st century, especially in East Asia. However, the impact of O3 has not been directly assessed at the forest level in this region. We performed O3 flux-based risk assessments of carbon sequestration capacity in an old cool temperate deciduous forest, consisting of O3-sensitive Japanese beech (Fagus crenata), and in a warm temperate deciduous and evergreen forest dominated by O3-tolerant Konara oak (Quercus serrata) based on long-term CO2 flux observations. On the basis of a practical approach for a continuous estimation of canopy-level stomatal conductance (Gs), higher phytotoxic ozone dose above a threshold of 0 uptake (POD0) with higher Gs was observed in the beech forest than that in the oak forest. Light-saturated gross primary production, as a measure of carbon sequestration capacity of forest ecosystem, declined earlier in the late growth season with increasing POD0, suggesting an earlier autumn senescence, especially in the O3-sensitive beech forest, but not in the O3-tolerant oak forest. PMID:27601188

  1. One-step biosynthesis of α-ketoisocaproate from L-leucine by an Escherichia coli whole-cell biocatalyst expressing an L-amino acid deaminase from Proteus vulgaris.

    PubMed

    Song, Yang; Li, Jianghua; Shin, Hyun-dong; Du, Guocheng; Liu, Long; Chen, Jian

    2015-07-28

    This work aimed to develop a whole-cell biotransformation process for the production of α-ketoisocaproate from L-leucine. A recombinant Escherichia coli strain was constructed by expressing an L-amino acid deaminase from Proteus vulgaris. To enhance α-ketoisocaproate production, the reaction conditions were optimized as follows: whole-cell biocatalyst 0.8 g/L, leucine concentration 13.1 g/L, temperature 35 °C, pH 7.5, and reaction time 20 h. Under the above conditions, the α-ketoisocaproate titer reached 12.7 g/L with a leucine conversion rate of 97.8%. In addition, different leucine feeding strategies were examined to increase the α-ketoisocaproate titer. When 13.1 g/L leucine was added at 2-h intervals (from 0 to 22 h, 12 addition times), the α-ketoisocaproate titer reached 69.1 g/L, while the leucine conversion rate decreased to 50.3%. We have developed an effective process for the biotechnological production of α-ketoisocaproate that is more environmentally friendly than the traditional petrochemical synthesis approach.

  2. Effect of post-exercise protein-leucine feeding on neutrophil function, immunomodulatory plasma metabolites and cortisol during a 6-day block of intense cycling.

    PubMed

    Nelson, Andre R; Jackson, Lara; Clarke, Jim; Stellingwerff, Trent; Broadbent, Suzanne; Rowlands, David S

    2013-09-01

    Whey protein and leucine ingestion following exercise increases muscle protein synthesis and could influence neutrophil function during recovery from prolonged intense exercise. We examined the effects of whey protein and leucine ingestion post-exercise on neutrophil function and immunomodulators during a period of intense cycling. In a randomized double-blind crossover, 12 male cyclists ingested protein/leucine/carbohydrate/fat (LEUPRO 20/7.5/89/22 g h(-1), respectively) or isocaloric carbohydrate/fat control (CON 119/22 g h(-1)) beverages for 1-3 h post-exercise during 6 days of high-intensity training. Blood was taken pre- and post-exercise on days 1, 2, 4 and 6 for phorbol myristate acetate (PMA)-stimulated neutrophil superoxide (O2 (-)) production, immune cell counts, amino acid and lipid metabolism via metabolomics, hormones (cortisol, testosterone) and cytokines (interleukin-6, interleukin-10). During recovery on day 1, LEUPRO ingestion increased mean concentrations of plasma amino acids (glycine, arginine, glutamine, leucine) and myristic acid metabolites (acylcarnitines C14, myristoylcarnitine; and C14:1-OH, hydroxymyristoleylcarnitine) with neutrophil priming capacity, and reduced neutrophil O2 production (15-17 mmol O2 (-) cell(-1) ± 90 % confidence limits 20 mmol O2 (-) cell(-1)). On day 2, LEUPRO increased pre-exercise plasma volume (6.6 ± 3.8 %) but haematological effects were trivial. LEUPRO supplementation did not substantially alter neutrophil elastase, testosterone, or cytokine concentrations. By day 6, however, LEUPRO reduced pre-exercise cortisol 21 % (±15 %) and acylcarnitine C16 (palmitoylcarnitine) during exercise, and increased post-exercise neutrophil O2 (-) (33 ± 20 mmol O2 (-) cell(-1)), relative to control. Altered plasma amino acid and acylcarnitine concentrations with protein-leucine feeding might partly explain the acute post-exercise reduction in neutrophil function and increased exercise-stimulated neutrophil oxidative burst on

  3. Signal Increase on Unenhanced T1-Weighted Images in the Rat Brain After Repeated, Extended Doses of Gadolinium-Based Contrast Agents

    PubMed Central

    Jost, Gregor; Lenhard, Diana Constanze; Sieber, Martin Andrew; Lohrke, Jessica; Frenzel, Thomas; Pietsch, Hubertus

    2016-01-01

    Objectives In this prospective preclinical study, we evaluated T1-weighted signal intensity in the deep cerebellar nuclei (CN) and globus pallidus (GP) up to 24 days after repeated administration of linear and macrocyclic gadolinium-based contrast agents (GBCAs) using homologous imaging and evaluation methods as in the recently published retrospective clinical studies. In a second part of the study, cerebrospinal fluid (CSF) spaces were evaluated for contrast enhancement by fluid-attenuated magnetic resonance imaging (MRI). Materials and Methods Sixty adult male Wistar-Han rats were randomly divided into a control and 5 GBCA groups (n = 10 per group). The administered GBCAs were gadodiamide, gadopentetate dimeglumine, and gadobenate dimeglumine (linear GBCAs) as well as gadobutrol and gadoterate meglumine (macrocyclic GBCAs) and saline (control). Over a period of 2 weeks, the animals received 10 intravenous injections at a dose of 2.5 mmol Gd/kg body weight, each on 5 consecutive days per week. Before GBCA administration, as well as 3 and 24 days after the last injection, a whole-brain MRI was performed using a standard T1-weighted 3-dimensional turbo spin echo sequence on a clinical 1.5 T scanner. The ratios of signal intensities in deep CN to pons (CN/Po) and GP to thalamus (GP/Th) were determined. For the evaluation of the CSF spaces, 18 additional rats were randomly divided into 6 groups (n = 3 per group) that received the same GBCAs as in the first part of the study. After MR cisternography for anatomical reference, a fluid-attenuated inversion recovery sequence was performed before and 1 minute after intravenous injection of a dose of 1 mmol Gd/kg body weight GBCA or saline. Results A significantly increased signal intensity ratio of CN/Po was observed 3 and 24 days after the last injection of gadodiamide and gadobenate dimeglumine. No significant changes were observed between the 2 time points. Gadopentetate dimeglumine injection led to a moderately elevated

  4. Contrarily to whey and high protein diets, dietary free leucine supplementation cannot reverse the lack of recovery of muscle mass after prolonged immobilization during ageing.

    PubMed

    Magne, Hugues; Savary-Auzeloux, Isabelle; Migné, Carole; Peyron, Marie-Agnès; Combaret, Lydie; Rémond, Didier; Dardevet, Dominique

    2012-04-15

    During ageing, immobilization periods increase and are partially responsible of sarcopaenia by inducing a muscle atrophy which is hardly recovered from. Immobilization-induced atrophy is due to an increase of muscle apoptotic and proteolytic processes and decreased protein synthesis. Moreover, previous data suggested that the lack of muscle mass recovery might be due to a defect in protein synthesis response during rehabilitation. This study was conducted to explore protein synthesis during reloading and leucine supplementation effect as a nutritional strategy for muscle recovery. Old rats (22–24 months old) were subjected to unilateral hindlimb casting for 8 days (I8) and allowed to recover for 10–40 days (R10–R40). They were fed a casein (±leucine) diet during the recovery. Immobilized gastrocnemius muscles atrophied by 20%, and did not recover even at R40. Amount of polyubiquitinated conjugates and chymotrypsin- and trypsin-like activities of the 26S proteasome increased. These changes paralleled an ‘anabolic resistance' of the protein synthesis at the postprandial state (decrease of protein synthesis, P-S6 and P-4E-BP1). During the recovery, proteasome activities remained elevated until R10 before complete normalization and protein synthesis was slightly increased. With free leucine supplementation during recovery, if proteasome activities were normalized earlier and protein synthesis was higher during the whole recovery, it nevertheless failed in muscle mass gain. This discrepancy could be due to a ‘desynchronization' between the leucine signal and the availability of amino acids coming from casein digestion. Thus, when supplemented with leucine-rich proteins (i.e. whey) and high protein diets, animals partially recovered the muscle mass loss.

  5. Contrarily to whey and high protein diets, dietary free leucine supplementation cannot reverse the lack of recovery of muscle mass after prolonged immobilization during ageing

    PubMed Central

    Magne, Hugues; Savary-Auzeloux, Isabelle; Migné, Carole; Peyron, Marie-Agnès; Combaret, Lydie; Rémond, Didier; Dardevet, Dominique

    2012-01-01

    During ageing, immobilization periods increase and are partially responsible of sarcopaenia by inducing a muscle atrophy which is hardly recovered from. Immobilization-induced atrophy is due to an increase of muscle apoptotic and proteolytic processes and decreased protein synthesis. Moreover, previous data suggested that the lack of muscle mass recovery might be due to a defect in protein synthesis response during rehabilitation. This study was conducted to explore protein synthesis during reloading and leucine supplementation effect as a nutritional strategy for muscle recovery. Old rats (22–24 months old) were subjected to unilateral hindlimb casting for 8 days (I8) and allowed to recover for 10–40 days (R10–R40). They were fed a casein (±leucine) diet during the recovery. Immobilized gastrocnemius muscles atrophied by 20%, and did not recover even at R40. Amount of polyubiquitinated conjugates and chymotrypsin- and trypsin-like activities of the 26S proteasome increased. These changes paralleled an ‘anabolic resistance’ of the protein synthesis at the postprandial state (decrease of protein synthesis, P-S6 and P-4E-BP1). During the recovery, proteasome activities remained elevated until R10 before complete normalization and protein synthesis was slightly increased. With free leucine supplementation during recovery, if proteasome activities were normalized earlier and protein synthesis was higher during the whole recovery, it nevertheless failed in muscle mass gain. This discrepancy could be due to a ‘desynchronization’ between the leucine signal and the availability of amino acids coming from casein digestion. Thus, when supplemented with leucine-rich proteins (i.e. whey) and high protein diets, animals partially recovered the muscle mass loss. PMID:22351629

  6. Differential effects of long-term leucine infusion on tissue protein synthesis in neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leucine is unique among the amino acids in its ability to promote protein synthesis by activating translation initiation via the mammalian target of rapamycin (mTOR) pathway. Previously, we showed that leucine infusion acutely stimulates protein synthesis in fast-twitch glycolytic muscle of neonatal...

  7. Prolonged stimulation of protein synthesis by leucine is dependent on amino acid availability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leucine is unique among the amino acids in its ability to enhance protein synthesis by activating translation initiation (Kimball and Jefferson, 2005). Our laboratory has shown that raising leucine to postprandial levels, whilst keeping all other amino acids at the post absorptive, level acutely st...

  8. Genetically encoded FRET-based nanosensor for in vivo measurement of leucine.

    PubMed

    Mohsin, Mohd; Abdin, M Z; Nischal, Lata; Kardam, Hemant; Ahmad, Altaf

    2013-12-15

    Besides fundamental role in protein synthesis, leucine has metabolic roles as energy substrates, precursors for synthesis of other amino acids and as a modulator of muscle protein synthesis via the insulin-signaling pathway. Leucine concentration in cell and tissue is temporally dynamic as the metabolism of leucine is regulated through multiple enzymes and transporters. Assessment of cell-type specific activities of transporters and enzymes by physical fractionation is extremely challenging. Therefore, a method of reporting leucine dynamics at the cellular level is highly desirable. Given this, we developed a series of genetically encoded nanosensors for real-time in vivo measurement of leucine at cellular level. A leucine binding periplasmic binding protein (LivK) of Escherichia coli K12 was flanked with CFP (cyan fluorescent protein) and YFP (yellow fluorescent protein) at N-terminus and C-terminus, respectively. The constructed nanosensors allowed in vitro determination of fluorescence resonance energy transfer (FRET) changes in a concentration-dependent manner. These sensors were found to be specific to leucine, and stable to pH-changes within a physiological range. Genetically encoded sensors can be targeted to a specific cell type, and allow dynamic measurement of leucine concentration in bacterial and yeast cells.

  9. Complete genome sequence of Corynebacterium glutamicum CP, a Chinese l-leucine producing strain.

    PubMed

    Gui, Yongli; Ma, Yuechao; Xu, Qingyang; Zhang, Chenglin; Xie, Xixian; Chen, Ning

    2016-02-20

    Here, we report the complete genome sequence of Corynebacterium glutamicum CP, an industrial l-leucine producing strain in China. The whole genome consists of a circular chromosome and a plasmid. The comparative genomics analysis shows that there are many mutations in the key enzyme coding genes relevant to l-leucine biosynthesis compared to C. glutamicum ATCC 13032.

  10. Kinetic Behavior of Leucine and Other Amino Acids Modulating Cognitive Performance via mTOR Pathway

    DTIC Science & Technology

    2011-12-02

    Acid BBB Transporter Type Structure Leucine L1 Essential Neutral Non-polar (hydrophobic) Branched chain Aspartic Acid Acidic X- Acidic Polar Glutamine...compared with other tissues. • Effects of leucine on other amino acids were analyzed. Those measured were aspartic acid , glutamic acid, serine, histidine

  11. Post-meal responses of elongation factor 2 (eEF2) and adenosine monophosphate-activated protein kinase (AMPK) to leucine and carbohydrate supplements for regulating protein synthesis duration and energy homeostasis in rat skeletal muscle.

    PubMed

    Wilson, Gabriel J; Moulton, Christopher J; Garlick, Peter J; Anthony, Tracy G; Layman, Donald K

    2012-11-13

    Previous research demonstrates that the anabolic response of muscle protein synthesis (MPS) to a meal is regulated at the level of translation initiation with signals derived from leucine (Leu) and insulin to activate mTORC1 signaling. Recent evidence suggests that the duration of the meal response is limited by energy status of the cell and inhibition of translation elongation factor 2 (eEF2). This study evaluates the potential to extend the anabolic meal response with post-meal supplements of Leu or carbohydrates. Adult (~256 g) male Sprague-Dawley rats were food deprived for 12 h, then either euthanized before a standard meal (time 0) or at 90 or 180 min post-meal. At 135 min post-meal, rats received one of five oral supplements: 270 mg leucine (Leu270), 80:40:40 mg leucine, isoleucine, and valine (Leu80), 2.63 g carbohydrates (CHO2.6), 1 g carbohydrates (CHO1.0), or water (Sham control). Following the standard meal, MPS increased at 90 min then declined to pre-meal baseline at 180 min. Rats administered Leu270, Leu80, CHO2.6, or CHO1.0 maintained elevated rates of MPS at 180 min, while Sham controls declined from peak values. Leu80 and CHO1.0 treatments maintained MPS, but with values intermediate between Sham controls and Leu270 and CHO2.6 supplements. Consistent with MPS findings, the supplements maintained elongation activity and cellular energy status by preventing increases in AMP/ATP and phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), acetyl-CoA carboxylase ACC and eEF2. The impact of the supplements on MPS and cellular energy status was in proportion to the energy content within the individual treatments (i.e., Leu270 > Leu80; CHO2.6 > CHO1.0), but the Leu supplements produced a disproportionate anabolic stimulation of MPS, eEF2 and energy status with significantly lower energy content. In summary, the incongruity between MPS and translation initiation at 180 min reflects a block in translation elongation due to reduced

  12. Molecular Recognition of Muramyl Dipeptide Occurs in the Leucine-rich Repeat Domain of Nod2.

    PubMed

    Lauro, Mackenzie L; D'Ambrosio, Elizabeth A; Bahnson, Brian J; Grimes, Catherine Leimkuhler

    2017-04-14

    Genetic mutations in the innate immune receptor nucleotide-binding oligomerization domain-containing 2 (Nod2) have demonstrated increased susceptibility to Crohn's disease, an inflammatory bowel disease that is hypothesized to be accompanied by changes in the gut microbiota. Nod2 responds to the presence of bacteria, specifically a fragment of the bacterial cell wall, muramyl dipeptide (MDP). The proposed site of this interaction is the leucine-rich repeat (LRR) domain. Surface plasmon resonance and molecular modeling were used to investigate the interaction of the LRR domain with MDP. A functional and pure LRR domain was obtained from Escherichia coli expression in high yield. The LRR domain binds to MDP with high affinity, with a KD of 212 ± 24 nM. Critical portions of the receptor were determined by mutagenesis of putative binding residues. Fragment analysis of MDP revealed that both the peptide and carbohydrate portion contribute to the binding interaction.

  13. Control of Neural Circuit Formation by Leucine-Rich Repeat Proteins

    PubMed Central

    de Wit, Joris; Ghosh, Anirvan

    2014-01-01

    The function of neural circuits depends on the precise connectivity between populations of neurons. Increasing evidence indicates that disruptions in excitatory or inhibitory synapse formation or function lead to excitation/inhibition (E/I) imbalances and contribute to neurodevelopmental and psychiatric disorders. Leucine-rich repeat (LRR)-containing surface proteins have emerged as key organizers of excitatory and inhibitory synapses. Distinct LRR proteins are expressed in different cell types and interact with key pre- and postsynaptic proteins. These protein interaction networks allow LRR proteins to coordinate pre- and postsynaptic elements during synapse formation and differentiation, pathway-specific synapse development, and synaptic plasticity. LRR proteins thus play a critical role in organizing synaptic connections into functional neural circuits, and their dysfunction may contribute to neuropsychiatric disorders. PMID:25131359

  14. The local application of a flavonoid, (-)-epicatechin, increases the spiking of globus pallidus neurons in a dose-dependent manner and diminishes the catalepsy induced by haloperidol.

    PubMed

    Alatorre, Alberto; Oviedo-Chávez, Aldo; Villalobos, Nelson; Ríos, Alain; Barrientos, Rafael; Querejeta, Enrique

    2015-02-01

    Flavonoids are natural substances obtained from plants. Most flavonoids cross the blood-brain barrier and exert a wide range of effects on the central nervous system. These actions have been attributed to the modulation of GABA-A receptors. Although motor systems in the central nervous system express a high density of GABA-A receptors, physiological studies about the effects of flavonoids on motor nuclei are scarce. Among the nuclei of the basal ganglia, the globus pallidus is potentially important for the processing of information related to movement. The electrical activity of globus pallidus neurons depends on the GABAergic fibers coming from the striatum and recurrent collateral fibers. It is known that the basal activity of the globus pallidus is modified by blocking dopaminergic receptors. In the present work, we analyzed the effects of the local application of a flavonoid, (-)-epicatechin, on the spiking of globus pallidus neurons in chloral hydrate-anesthetized rats and determined whether (-)-epicatechin applied bilaterally to the globus pallidus can modify the catalepsy induced by systemic administration of haloperidol. The results showed that (-)-epicatechin increased the basal firing of globus pallidus neurons in a dose-dependent manner and antagonized the inhibitory effect of GABA. Bilateral infusion of (-)-epicatechin to the globus pallidus diminished the catalepsy induced by haloperidol.

  15. Thermodynamic analysis of the heterodimerization of leucine zippers of Jun and Fos transcription factors

    SciTech Connect

    Seldeen, Kenneth L.; McDonald, Caleb B.; Deegan, Brian J.

    2008-10-31

    Jun and Fos are components of the AP1 family of transcription factors and bind to the promoters of a diverse multitude of genes involved in critical cellular responses such as cell growth and proliferation, cell cycle regulation, embryonic development and cancer. Here, using the powerful technique of isothermal titration calorimetry, we characterize the thermodynamics of heterodimerization of leucine zippers of Jun and Fos. Our data suggest that the heterodimerization of leucine zippers is driven by enthalpic forces with unfavorable entropy change at physiological temperatures. Furthermore, the basic regions appear to modulate the heterodimerization of leucine zippers and may undergo at least partial folding upon heterodimerization. Large negative heat capacity changes accompanying the heterodimerization of leucine zippers are consistent with the view that leucine zippers do not retain {alpha}-helical conformations in isolation and that the formation of the native coiled-coil {alpha}-helical dimer is attained through a coupled folding-dimerization mechanism.

  16. Leucine partially protects muscle mass and function during bed rest in middle-aged adults1,2

    PubMed Central

    English, Kirk L; Mettler, Joni A; Ellison, Jennifer B; Mamerow, Madonna M; Arentson-Lantz, Emily; Pattarini, James M; Ploutz-Snyder, Robert; Sheffield-Moore, Melinda; Paddon-Jones, Douglas

    2016-01-01

    Background: Physical inactivity triggers a rapid loss of muscle mass and function in older adults. Middle-aged adults show few phenotypic signs of aging yet may be more susceptible to inactivity than younger adults. Objective: The aim was to determine whether leucine, a stimulator of translation initiation and skeletal muscle protein synthesis (MPS), can protect skeletal muscle health during bed rest. Design: We used a randomized, double-blind, placebo-controlled trial to assess changes in skeletal MPS, cellular signaling, body composition, and skeletal muscle function in middle-aged adults (n = 19; age ± SEM: 52 ± 1 y) in response to leucine supplementation (LEU group: 0.06 g ∙ kg−1 ∙ meal−1) or an alanine control (CON group) during 14 d of bed rest. Results: Bed rest decreased postabsorptive MPS by 30% ± 9% (CON group) and by 10% ± 10% (LEU group) (main effect for time, P < 0.05), but no differences between groups with respect to pre-post changes (group × time interactions) were detected for MPS or cell signaling. Leucine protected knee extensor peak torque (CON compared with LEU group: −15% ± 2% and −7% ± 3%; group × time interaction, P < 0.05) and endurance (CON compared with LEU: −14% ± 3% and −2% ± 4%; group × time interaction, P < 0.05), prevented an increase in body fat percentage (group × time interaction, P < 0.05), and reduced whole-body lean mass loss after 7 d (CON compared with LEU: −1.5 ± 0.3 and −0.8 ± 0.3 kg; group × time interaction, P < 0.05) but not 14 d (CON compared with LEU: −1.5 ± 0.3 and −1.0 ± 0.3 kg) of bed rest. Leucine also maintained muscle quality (peak torque/kg leg lean mass) after 14 d of bed-rest inactivity (CON compared with LEU: −9% ± 2% and +1% ± 3%; group × time interaction, P < 0.05). Conclusions: Bed rest has a profoundly negative effect on muscle metabolism, mass, and function in middle-aged adults. Leucine supplementation may partially protect muscle health during relatively

  17. Neonatal Immunization with a Single IL-4/Antigen Dose Induces Increased Antibody Responses after Challenge Infection with Equine Herpesvirus Type 1 (EHV-1) at Weanling Age

    PubMed Central

    Wagner, Bettina; Perkins, Gillian; Babasyan, Susanna; Freer, Heather; Keggan, Alison; Goodman, Laura B.; Glaser, Amy; Torsteinsdóttir, Sigurbjorg; Svansson, Vilhjálmur; Björnsdóttir, Sigríður

    2017-01-01

    Neonatal foals respond poorly to conventional vaccines. These vaccines typically target T-helper (Th) cell dependent B-cell activation. However, Th2-cell immunity is impaired in foals during the first three months of life. In contrast, neonatal basophils are potent interleukin-4 (IL-4) producers. The purpose of this study was to develop a novel vaccine triggering the natural capacity of neonatal basophils to secrete IL-4 and to evaluate if vaccination resulted in B-cell activation and antibody production against EHV-1 glycoprotein C (gC). Neonatal vaccination was performed by oral biotinylated IgE (IgE-bio) treatment at birth followed by intramuscular injection of a single dose of streptavidin-conjugated gC/IL-4 fusion protein (Sav-gC/IL-4) for crosslinking of receptor-bound IgE-bio (group 1). Neonates in group 2 received the intramuscular Sav-gC/IL-4 vaccine only. Group 3 remained non-vaccinated at birth. After vaccination, gC antibody production was not detectable. The ability of the vaccine to induce protection was evaluated by an EHV-1 challenge infection after weaning at 7 months of age. Groups 1 and 2 responded to EHV-1 infection with an earlier onset and overall significantly increased anti-gC serum antibody responses compared to control group 3. In addition, group 1 weanlings had a decreased initial fever peak after infection indicating partial protection from EHV-1 infection. This suggested that the neonatal vaccination induced a memory B-cell response at birth that was recalled at weanling age after EHV-1 challenge. In conclusion, early stimulation of neonatal immunity via the innate arm of the immune system can induce partial protection and increased antibody responses against EHV-1. PMID:28045974

  18. Low-dose exposure to bisphenol A in combination with fructose increases expression of genes regulating angiogenesis and vascular tone in juvenile Fischer 344 rat cardiac tissue

    PubMed Central

    Klint, Helén; Karimullina, Elina; Rönn, Monika; Lind, Lars; Lind, P. Monica

    2017-01-01

    Objectives Epidemiological studies report associations between exposure to the high-volume chemical and endocrine disruptor bisphenol A (BPA) and cardiovascular disorders, but there is a lack of experimental studies addressing the mechanisms of action of BPA on the cardiovascular system. In the present study, effects on markers for cardiovascular function of exposure to BPA and fructose in vivo in rat cardiac tissues, and of BPA exposure in human cardiomyocytes in vitro, were investigated. Materials Juvenile female Fischer 344 rats were exposed to 5, 50, and 500 μg BPA/kg bodyweight/day in their drinking water from 5 to 15 weeks of age, in combination with 5% fructose. Further, cultured human cardiomyocytes were exposed to 10 nM BPA to 1 × 104 nM BPA for six hours. Expression of markers for cardiovascular function and BPA target receptors was investigated using qRT-PCR. Results Exposure to 5 μg BPA/kg bodyweight/day plus fructose increased mRNA expression of Vegf, Vegfr2, eNos, and Ace1 in rat heart. Exposure of human cardiomyocytes to 1 × 104 nM BPA increased mRNA expression of eNOS and ACE1, as well as IL-8 and NFκβ known to regulate inflammatory response. Conclusions:. Low-dose exposure of juvenile rats to BPA and fructose induced up-regulation of expression of genes controlling angiogenesis and vascular tone in cardiac tissues. The observed effects of BPA in rat heart were in line with our present and previous studies of BPA in human endothelial cells and cardiomyocytes. These findings may aid in understanding the mechanisms of the association between BPA exposure and cardiovascular disorders reported in epidemiological studies. PMID:27622962

  19. Effect of leucine uptake on hepatic and skeletal muscle gene expression in rats: a microarray analysis

    PubMed Central

    Cheon, Wookwang

    2015-01-01

    [Purpose] This study was performed to explore the physiological functions of leucine by exploring genes with leucine-dependent variability using DNA microarray. [Methods] Sprague-Dawley rats (n = 20) were separated into a HPD (30% High Protein Diet, n = 10) group and a NPD (0% Non Protein Diet, n = 10) group and fed a protein diet for 2 weeks. At the end of the 2-week period, the rats were fasted for 12-16 hours, further separated into subgroups within the HPD (Saline, n = 5, Leucine, n = 5) and NPD (Saline, n = 5, Leucine, n = 5) groups and administered with a leucine solution. The liver and muscles were harvested after 2 hours for RNA extraction. RNA purification from the isolated muscles and target gene identification using DNA chip were performed. The target gene was determined based on the results of the DNA chip experiment, and mRNA expression of the target gene was analyzed using Real-Time PCR. [Results] In the skeletal muscle, 27 genes were upregulated while 52 genes were down regulated after leucine administration in the NPD group. In the liver, 160 genes were up-regulated while 126 were down-regulated. The per2 gene was one of the genes with leucine-dependent induction in muscles and liver. [Conclusion] This study was performed to explore the physiological functions of leucine, however, a large number of genes showed variability. Therefore, it was difficult to definitively identify the genes linked with a particular physiological function. Various nutritional effects of leucine were observed. High variability in cytokines, receptors, and various membrane proteins were observed, which suggests that leucine functions as more than a nutrient. The interpretation may depend on investigators’ perspectives, therefore, discussion with relevant experts and the BCAA (Branched-Chain Amino Acids) society may be needed for effective utilization of this data. PMID:26244133

  20. Glutamate formation via the leucine-to-glutamate pathway of rat pancreas.

    PubMed

    Schachter, David; Buteau, Jean

    2014-06-01

    The leucine-to-glutamate (Leu→Glu) pathway, which metabolizes the carbon atoms of l-leucine to form l-glutamate, was studied by incubation of rat tissue segments with l-[U-(14)C]leucine and estimation of the [(14)C]glutamate formed. Metabolism of the leucine carbon chain occurs in most rat tissues, but maximal activity of the Leu→Glu pathway for glutamate formation is limited to the thoracic aorta and pancreas. In rat aorta, the Leu→Glu pathway functions to relax the underlying smooth muscle; its functions in the pancreas are unknown. This report characterizes the Leu→Glu pathway of rat pancreas and develops methods to examine its functions. Pancreatic segments effect net formation of glutamate on incubation with l-leucine, l-glutamine, or a mix of 18 other plasma amino acids at their concentrations in normal rat plasma. Glutamate formed from leucine remains mainly in the tissue, whereas that from glutamine enters the medium. The pancreatic Leu→Glu pathway uses the leucine carbons for net glutamate formation; the α-amino group is not used; the stoichiometry is as follows: 1 mol of leucine yields 2 mol of glutamate (2 leucine carbons per glutamate) plus 2 mol of CO2. Comparison of the Leu→Glu pathway in preparations of whole pancreatic segments, isolated acini, and islets of Langerhans localizes it in the acini; relatively high activity is found in cultures of the AR42J cell line and very little in the INS-1 832/13 cell line. Pancreatic tissue glutamate concentration is homeostatically regulated in the range of ∼1-3 μmol/g wet wt. l-Valine and leucine ethyl, benzyl, and tert-butyl esters inhibit the Leu→Glu pathway without decreasing tissue total glutamate.

  1. Proteomic analysis of post-nuclear supernatant fraction and percoll-purified membranes prepared from brain cortex of rats exposed to increasing doses of morphine

    PubMed Central

    2014-01-01

    Background Proteomic analysis was performed in post-nuclear supernatant (PNS) and Percoll-purified membranes (PM) prepared from fore brain cortex of rats exposed to increasing doses of morphine (10–50 mg/kg) for 10 days. Results In PNS, the 10 up (↑)- or down (↓)-regulated proteins exhibiting the largest morphine-induced change were selected, excised manually from the gel and identified by MALDI-TOF MS/MS: 1-(gi|148747414, Guanine deaminase), ↑2.5×; 2-(gi|17105370, Vacuolar-type proton ATP subunit B, brain isoform), ↑2.6×; 3-(gi|1352384, Protein disulfide-isomerase A3), ↑3.4×; 4-(gi|40254595, Dihydropyrimidinase-related protein 2), ↑3.6×; 5-(gi|149054470, N-ethylmaleimide sensitive fusion protein, isoform CRAa), ↑2.0×; 6-(gi|42476181, Malate dehydrogenase, mitochondrial precursor), ↑1.4×; 7-(gi|62653546, Glyceraldehyde-3-phosphate dehydrogenase), ↑1.6×; 8-(gi|202837, Aldolase A), ↑1.3×; 9-(gi|31542401, Creatine kinase B-type), ↓0.86×; 10-(gi|40538860, Aconitate hydratase, mitochondrial precursor), ↑1.3×. The identified proteins were of cytoplasmic (1, 4, 5, 7, 9), cell membrane (2), endoplasmic reticulum (3) and mitochondrial (6, 8, 10) origin and 9 of them were significantly increased, 1.3-3.6×. The 4 out of 9 up-regulated proteins (4, 6, 7, 10) were described as functionally related to oxidative stress; the 2 proteins participate in genesis of apoptotic cell death. In PM, the 18 up (↑)- or down (↓)-regulated proteins were identified by LC-MS/MS and were of plasma membrane [Brain acid soluble protein, ↓2.1×; trimeric Gβ subunit, ↓2.0x], myelin membrane [MBP, ↓2.5×], cytoplasmic [Internexin, ↑5.2×; DPYL2, ↑4.9×; Ubiquitin hydrolase, ↓2.0×; 60S ribosomal protein, ↑2.7×; KCRB, ↓2.6×; Sirtuin-2, ↑2.5×; Peroxiredoxin-2, ↑2.2×; Septin-11, ↑2.2×; TERA, ↑2.1×; SYUA, ↑2.0×; Coronin-1A, ↓5.4×] and mitochondrial [Glutamate dehydrogenase 1, ↑2.7×; SCOT1, ↑2.2×; Prohibitin, ↑2.2

  2. The addition of granulocyte-colony stimulating factor shifts the dose limiting toxicity and markedly increases the maximum tolerated dose and activity of the kinesin spindle protein inhibitor SB-743921 in patients with relapsed or refractory lymphoma: results of an international, multicenter phase I/II study.

    PubMed

    O'Connor, Owen A; Gerecitano, John; Van Deventer, Henrik; Hainsworth, John; Zullo, Kelly M; Saikali, Khalil; Seroogy, Joseph; Wolff, Andrew; Escandón, Rafael

    2015-01-01

    This was a phase I study of SB-743921 (SB-921) in patients with relapsed/refractory lymphoma. Previous studies established that neutropenia was the only dose limiting toxicity (DLT). The primary objective was to determine the DLT, maximum tolerated dose (MTD) and efficacy of SB-921 with and without granulocyte-colony stimulating factor (G-CSF). Sixty-eight patients were enrolled, 42 without G-CSF, 26 with G-CSF. In the cohort without G-CSF, SB-921 doses ranged from 2 to 7 mg/m(2), with 6 mg/m(2) being the MTD. In the cohort with G-CSF support, doses of 6-10 mg/m(2) were administered, with 9 mg/m(2) being the MTD, representing a 50% increase in dose density. Fifty-six patients were evaluable for efficacy. Four of 55 patients experienced a partial response (three in Hodgkin lymphoma and one in non-Hodgkin lymphoma, all at doses ≥ 6 mg/m(2)); 19 patients experienced stable disease, 33 patients developed progression of disease. G-CSF shifted the DLT from neutropenia to thrombocytopenia, allowing for a 50% increase in dose density. Responses were seen at higher doses with G-CSF support.

  3. SU-F-BRF-14: Increasing the Accuracy of Dose Calculation On Cone-Beam Imaging Using Deformable Image Registration in the Case of Prostate Translation

    SciTech Connect

    Fillion, O; Gingras, L; Archambault, L

    2014-06-15

    Purpose: Artifacts can reduce the quality of dose re-calculations on CBCT scans during a treatment. The aim of this project is to correct the CBCT images in order to allow for more accurate and exact dose calculations in the case of a translation of the tumor in prostate cancer. Methods: Our approach is to develop strategies based on deformable image registration algorithms using the elastix software (Klein et al., 2010) to register the treatment planning CT on a daily CBCT scan taken during treatment. Sets of images are provided by a 3D deformable phantom and comprise two CT and two CBCT scans: one of both with the reference anatomy and the others with known deformations (i.e. translations of the prostate). The reference CT is registered onto the deformed CBCT and the deformed CT serves as the control for dose calculation accuracy. The planned treatment used for the evaluation of dose calculation is a 2-Gy fraction prescribed at the location of the reference prostate and assigned to 7 rectangular fields. Results: For a realistic 0.5-cm translation of the prostate, the relative dose discrepancy between the CBCT and the CT control scan at the prostate's centroid is 8.9 ± 0.8 % while dose discrepancy between the registered CT and the control scan lessens to −2.4 ± 0.8 %. For a 2-cm translation, clinical indices like the V90 and the D100 are more accurate by 0.7 ± 0.3 % and 8.0 ± 0.5 cGy respectively when using registered CT than when using CBCT for dose calculation. Conclusion: The results show that this strategy gives doses in agreement within a few percents with those from calculations on actual CT scans. In the future, various deformations of the phantom anatomy will allow a thorough characterization of the registration strategies needed for more complex anatomies.

  4. Effects of dietary leucine supplementation on the hepatic mitochondrial biogenesis and energy metabolism in normal birth weight and intrauterine growth-retarded weanling piglets

    PubMed Central

    Su, Weipeng; Xu, Wen; Zhang, Hao; Ying, Zhixiong; Zhou, Le; Zhang, Lili

    2017-01-01

    BACKGROUND/OBJECTIVES The study was conducted to evaluate the effects of dietary leucine supplementation on mitochondrial biogenesis and energy metabolism in the liver of normal birth weight (NBW) and intrauterine growth-retarded (IUGR) weanling piglets. MATERIALS/METHODS A total of sixteen pairs of NBW and IUGR piglets from sixteen sows were selected according to their birth weight. At postnatal day 14, all piglets were weaned and fed either a control diet or a leucine-supplemented diet for 21 d. Thereafter, a 2 × 2 factorial experimental design was used. Each treatment consisted of eight replications with one piglet per replication. RESULTS Compared with NBW piglets, IUGR piglets had a decreased (P < 0.05) hepatic adenosine triphosphate (ATP) content. Also, IUGR piglets exhibited reductions (P < 0.05) in the activities of hepatic mitochondrial pyruvate dehydrogenase (PDH), citrate synthase (CS), α-ketoglutarate dehydrogenase (α-KGDH), malate dehydrogenase (MDH), and complexes I and V, along with decreases (P < 0.05) in the concentration of mitochondrial DNA (mtDNA) and the protein expression of hepatic peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α). Dietary leucine supplementation increased (P < 0.05) the content of ATP, and the activities of CS, α-KGDH, MDH, and complex V in the liver of piglets. Furthermore, compared to those fed a control diet, piglets given a leucine-supplemented diet exhibited increases (P < 0.05) in the mtDNA content and in the mRNA expressions of sirtuin 1, PGC-1α, nuclear respiratory factor 1, mitochondrial transcription factor A, and ATP synthase, H+ transporting, mitochondrial F1 complex, β polypeptide in liver. CONCLUSIONS Dietary leucine supplementation may exert beneficial effects on mitochondrial biogenesis and energy metabolism in NBW and IUGR weanling piglets. PMID:28386385

  5. Comparative biochemical and functional properties of two leucine aminopeptidases of Clonorchis sinensis.

    PubMed

    Kang, Jung-Mi; Ju, Hye-Lim; Ju, Jung-Won; Sohn, Woon-Mok; Kim, Tong-Soo; Bahk, Young-Yil; Hong, Sung-Jong; Na, Byoung-Kuk

    2012-01-01

    Leucine aminopeptidases (LAP; EC 3.4.11.1) are a group of metalloexopeptidases, which catalyze the sequential removal of leucine amino acids from the N-termini of the polypeptides or proteins. In this study, we identified two novel genes that encode LAPs of Clonorchis sinensis (CsLAP1 and CsLAP2) and characterized their biochemical and functional properties. Multiple sequence alignment of the deduced amino acid sequences of CsLAP1 and CsLAP2 with those of other organisms revealed that typical metal-binding coordinating and active site residues for LAPs were well conserved in CsLAP1 and CsLAP2. Recombinant CsLAP1 and CsLAP2 showed similar biochemical properties such as pH optima at pH 8.0 and stability at neutral pHs. Both enzymes were specifically inhibited by bestatin and showed preferential substrate specificity for Leu-MCA. However, the enzymes differed in that they required different metal ions for maximum activity. Expressions of CsLAP1 and CsLAP2 were detected throughout the various developmental stages of C. sinensis, and their transcription levels increased gradually in accordance with the maturation of the parasite. Both enzymes were identified in soluble worm extract of C. sinensis, but not in excretory and secretory products. Immunolocalization studies showed that both enzymes were co-localized to the intestinal epithelial cells and gastrodermis of the parasite. These results collectively suggest that CsLAP1 and CsLAP2 are synthesized in the intestinal epithelial and gastrodermal cells of C. sinensis and may be involved in the final digestion of peptides that hydrolyzed within intestinal lumen followed by absorbed into gastrodermal cells of the parasite.

  6. Delivering a “Dose of Hope”: A Faith-Based Program to Increase Older African Americans’ Participation in Clinical Trials

    PubMed Central

    Omer, Saad B; Parker, Kimberly; Bolton, Marcus; Schamel, Jay; Shapiro, Eve; Owens, Lauren; Saint-Victor, Diane; Boggavarapu, Sahithi; Braxton, Nikia; Archibald, Matthew; Kalokhe, Ameeta S; Horton, Takeia; Root, Christin M; Fenimore, Vincent L; Anderson, Aaron M

    2015-01-01

    Background Underrepresentation of older-age racial and ethnic minorities in clinical research is a significant barrier to health in the United States, as it impedes medical research advancement of effective preventive and therapeutic strategies. Objective The objective of the study was to develop and test the feasibility of a community-developed faith-based intervention and evaluate its potential to increase the number of older African Americans in clinical research. Methods Using a cluster-randomized design, we worked with six matched churches to enroll at least 210 persons. We provided those in the intervention group churches with three educational sessions on the role of clinical trials in addressing health disparity topics, and those in the comparison group completed surveys at the same timepoints. All persons enrolled in the study received ongoing information via newsletters and direct outreach on an array of clinical studies seeking participants. We evaluated the short-, mid-, and longer-term effects of the interventional program on clinical trial-related outcomes (ie, screening and enrollment). Results From 2012 to 2013, we enrolled a balanced cohort of 221 persons in the program. At a 3-month follow-up, mean intention to seek information about clinical trials was higher than baseline in both treatment (mu=7.5/10; sigma=3.1) and control arms (mu=6.6/10; sigma=3.3), with the difference more pronounced in the treatment arm. The program demonstrated strong retention at 3-month (95.4%, 211/221) and 6-month timepoints (94.1%, 208/221). Conclusions The “Dose of Hope” program addressed an unmet need to reach an often overlooked audience of older African Americans who are members of churches and stimulate their interest in clinical trial participation. The program demonstrated its appeal in the delivery of effective messages and information about health disparities, and the role of clinical research in addressing these challenges. PMID:26036841

  7. A single dose of vortioxetine, but not ketamine or fluoxetine, increases plasticity-related gene expression in the rat frontal cortex.

    PubMed

    du Jardin, Kristian Gaarn; Müller, Heidi Kaastrup; Sanchez, Connie; Wegener, Gregers; Elfving, Betina

    2016-09-05

    Ketamine is a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist that has been shown to induce a rapid antidepressant effect in treatment-resistant patients. Vortioxetine is a multimodal-acting antidepressant that exert its therapeutic activity through serotonin (5-hydroxytryptamine; 5-HT) reuptake inhibition and modulation of several 5-HT receptors. In clinical trials, vortioxetine improves depression symptoms and cognitive dysfunction. Neuroplasticity as well as serotonergic and glutamatergic signaling attain significant roles in depression pathophysiology and antidepressant responses. Here, we investigate the effects of ketamine and vortioxetine on gene expression related to serotonergic and glutamatergic neurotransmission as well as neuroplasticity and compare them to those of the selective serotonin reuptake inhibitor fluoxetine. Rats were injected with fluoxetine (10mg/kg), ketamine (15mg/kg), or vortioxetine (10mg/kg) at 2, 8, 12, or 27h prior to harvesting of the frontal cortex and hippocampus. mRNA levels were measured by real-time quantitative polymerase chain reaction (qPCR). The main finding was that vortioxetine enhanced plasticity-related gene expression (Mtor, Mglur1, Pkcα, Homer3, Spinophilin, and Synapsin3) in the frontal cortex at 8h after a single dose. Ingenuity pathway analysis of this subset of data identified a biological network that was engaged by vortioxetine and is plausibly associated with neuroplasticity. Transcript levels had returned to baseline levels 12h after injection. Only minor effects on gene expression were found for ketamine or fluoxetine. In conclusion, acute vortioxetine, but not fluoxetine or ketamine, transiently increased plasticity-related gene expression in the frontal cortex. These effects may be ascribed to the direct 5-HT receptor activities of vortioxetine.

  8. Residues threonine 346 and leucine 352 are critical for the proper function of Bacillus kaustophilus leucine aminopeptidase.

    PubMed

    Chi, Meng-Chun; Huang, Hsien-Ben; Liu, Jai-Shin; Wang, Wen-Ching; Liang, Wan-Chi; Lin, Long-Liu

    2006-07-01

    The importance of Thr-346 and Leu-352 residues in Bacillus kaustophilus leucine aminopeptidase (BkLAP) was explored by site-directed mutagenesis. The impact of substitutions at these positions was evaluated with His6-BkLAP fusion proteins expressed in Escherichia coli. Substitution of Thr-346 with Tyr, Arg, and Leu, respectively, resulted in a dramatic reduction in LAP activity. A complete loss of activity was observed in L352E and L352R variants with the exception of L352 V, which retained approximately 60% of the wild-type activity. Zinc content analysis and protein modeling suggested that Thr-346 and Leu-352 of BkLAP play a role in maintaining the coordination environment for the zinc-binding residues.

  9. Functional diversification within the family of B-GATA transcription factors through the leucine-leucine-methionine domain.

    PubMed

    Behringer, Carina; Bastakis, Emmanouil; Ranftl, Quirin L; Mayer, Klaus F X; Schwechheimer, Claus

    2014-09-01

    The transcription of the Arabidopsis (Arabidopsis thaliana) GATA transcription factors GATA, NITRATE-INDUCIBLE, CARBON METABOLISM-INVOLVED (GNC) and GNC-LIKE (GNL)/CYTOKININ-RESPONSIVE GATA FACTOR1 is controlled by several growth regulatory signals including light and the phytohormones auxin, cytokinin, and gibberellin. To date, GNC and GNL have been attributed functions in the control of germination, greening, flowering time, floral development, senescence, and floral organ abscission. GNC and GNL belong to the 11-member family of B-class GATA transcription factors that are characterized to date solely by their high sequence conservation within the GATA DNA-binding domain. The degree of functional conservation among the various B-class GATA family members is not understood. Here, we identify and examine B-class GATAs from Arabidopsis, tomato (Solanum lycopersicon), Brachypodium (Brachypodium distachyon), and barley (Hordeum vulgare). We find that B-class GATAs from these four species can be subdivided based on their short or long N termini and the presence of the 13-amino acid C-terminal leucine-leucine-methionine (LLM) domain with the conserved motif LLM. Through overexpression analyses and by complementation of a gnc gnl double mutant, we provide evidence that the length of the N terminus may not allow distinguishing between the different B-class GATAs at the functional level. In turn, we find that the presence and absence of the LLM domain in the overexpressors has differential effects on hypocotyl elongation, leaf shape, and petiole length, as well as on gene expression. Thus, our analyses identify the LLM domain as an evolutionarily conserved domain that determines B-class GATA factor identity and provides a further subclassification criterion for this transcription factor family.

  10. Leucine deprivation inhibits proliferation and induces apoptosis of human breast cancer cells via fatty acid synthase

    PubMed Central

    Xiao, Fei; Wang, Chunxia; Yin, Hongkun; Yu, Junjie; Chen, Shanghai; Fang, Jing; Guo, Feifan

    2016-01-01

    Substantial studies on fatty acid synthase (FASN) have focused on its role in regulating lipid metabolism and researchers have a great interest in treating cancer with dietary manipulation of amino acids. In the current study, we found that leucine deprivation caused the FASN-dependent anticancer effect. Here we showed that leucine deprivation inhibited cell proliferation and induced apoptosis of MDA-MB-231 and MCF-7 breast cancer cells. In an in vivo tumor xenograft model, the leucine-free diet suppressed the growth of human breast cancer tumors and triggered widespread apoptosis of the cancer cells. Further study indicated that leucine deprivation decreased expression of lipogenic gene FASN in vitro and in vivo. Over-expression of FASN or supplementation of palmitic acid (the product of FASN action) blocked the effects of leucine deprivation on cell proliferation and apoptosis in vitro and in vivo. Moreover, leucine deprivation suppressed the FASN expression via regulating general control non-derepressible (GCN)2 and sterol regulatory element-binding protein 1C (SREBP1C). Taken together, our study represents proof of principle that anticancer effects can be obtained with strategies to deprive tumors of leucine via suppressing FASN expression, which provides important insights in prevention of breast cancer via metabolic intervention. PMID:27579768

  11. L-Leucine for gold nanoparticles synthesis and their cytotoxic effects evaluation.

    PubMed

    Berghian-Grosan, Camelia; Olenic, Liliana; Katona, Gabriel; Perde-Schrepler, Maria; Vulcu, Adriana

    2014-11-01

    This work reports the preparation of water-soluble leucine capped gold nanoparticles by two single-step synthesis methods. The first procedure involves a citrate reduction approach where the citrate is used as reducing agent and leucine as capping/stabilizing agent. Different sizes of gold nanoparticles, citrate reduced and stabilized by leucine, Leu-AuNPs-C, with the mean diameters in the range of 21-56 nm, were obtained by varying the macroscopic parameters such as: concentration of the gold precursor solution, Au (III):citrate molar ratio and leucine pH. In the second procedure, leucine acts both as reducing and stabilizing agent, allowing us to obtain spherical gold nanoparticles, Leu-AuNPs, with a majority of 80 % (with the mean diameter of 63 nm). This proves that leucine is an appropriate reductant for the formation of water-soluble and stable gold nanoparticles colloids. The characterization of the leucine coated gold nanoparticles was carried out by TEM, UV-Vis and FT-IR analysis. The cytotoxic effect of Leu-AuNPs-C and Leu-AuNPs was also evaluated.

  12. Lack of protective efficacy in buffaloes vaccinated with Fasciola gigantica leucine aminopeptidase and peroxiredoxin recombinant proteins.

    PubMed

    Raina, O K; Nagar, Gaurav; Varghese, Anju; Prajitha, G; Alex, Asha; Maharana, B R; Joshi, P

    2011-06-01

    Gene coding for leucine aminopeptidase (LAP), a metalloprotease, was identified in the tropical liver fluke, Fasciola gigantica; that on sequence analysis showed a close homology (98.6%) with leucine aminopeptidase of the temperate liver fluke, Fasciola hepatica. The recombinant leucine aminopeptidase protein was expressed in Escherichia coli. F. gigantica peroxiredoxin, a hydrogen peroxide scavenger and an immunomodulating protein, was also cloned and expressed in E. coli. A vaccination trial in buffaloes was conducted with these two recombinant proteins, with 150 and 300 μg of leucine aminopeptidase and a cocktail of 150 μg each of recombinant leucine aminopeptidase and peroxiredoxin in three groups, respectively. Both Th1- and Th2-associated humoral immune responses were elicited to immunization with these antigens. A challenge study with 400 metacercariae did not show a significant protection in terms of reduction in the worm burden (8.4%) or anti-fecundity/embryonation effect in the immunized groups, as to the non-immunized control animals. Our observations in this buffalo vaccination trial are contrary to the earlier promise shown by leucine aminopeptidase of F. hepatica as a leading candidate vaccine molecule. Identification of leucine aminopeptidase gene and evaluation of the protein for its protective efficacy in buffaloes is the first scientific report on this protein in F. gigantica.

  13. Design and characterization of short antimicrobial peptides using leucine zipper templates with selectivity towards microorganisms.

    PubMed

    Ahmad, Aqeel; Azmi, Sarfuddin; Srivastava, Saurabh; Kumar, Amit; Tripathi, Jitendra Kumar; Mishra, Nripendra N; Shukla, Praveen K; Ghosh, Jimut Kanti

    2014-11-01

    Design of antimicrobial peptides with selective activity towards microorganisms is an important step towards the development of new antimicrobial agents. Leucine zipper sequence has been implicated in cytotoxic activity of naturally occurring antimicrobial peptides; moreover, this motif has been utilized for the design of novel antimicrobial peptides with modulated cytotoxicity. To understand further the impact of substitution of amino acids at 'a' and/or 'd' position of a leucine zipper sequence of an antimicrobial peptides on its antimicrobial and cytotoxic properties four short peptides (14-residue) were designed on the basis of a leucine zipper sequence without or with replacement of leucine residues in its 'a' and 'd' positions with D-leucine or alanine or proline residue. The original short leucine zipper peptide (SLZP) and its D-leucine substituted analog, DLSA showed comparable activity against the tested Gram-positive and negative bacteria and the fungal strains. The alanine substituted analog (ASA) though showed appreciable activity against the tested bacteria, it showed to some extent lower activity against the tested fungi. However, the proline substituted analog (PSA) showed lower activity against the tested bacterial or fungal strains. Interestingly, DLSA, ASA and PSA showed significantly lower cytotoxicity than SLZP against both human red blood cells (hRBCs) and murine 3T3 cells. Cytotoxic and bactericidal properties of these peptides matched with peptide-induced damage/permeabilization of mammalian cells and bacteria or their mimetic lipid vesicles suggesting cell membrane could be the target of these peptides. As evidenced by tryptophan fluorescence and acrylamide quenching studies the peptides showed similarities either in interaction or in their localization within the bacterial membrane mimetic negatively charged lipid vesicles. Only SLZP showed localization inside the mammalian membrane mimetic zwitterionic lipid vesicles. The results show

  14. Acoustic dose and acoustic dose-rate.

    PubMed

    Duck, Francis

    2009-10-01

    Acoustic dose is defined as the energy deposited by absorption of an acoustic wave per unit mass of the medium supporting the wave. Expressions for acoustic dose and acoustic dose-rate are given for plane-wave conditions, including temporal and frequency dependencies of energy deposition. The relationship between the acoustic dose-rate and the resulting temperature increase is explored, as is the relationship between acoustic dose-rate and radiation force. Energy transfer from the wave to the medium by means of acoustic cavitation is considered, and an approach is proposed in principle that could allow cavitation to be included within the proposed definitions of acoustic dose and acoustic dose-rate.

  15. Proline/arginine-rich end leucine-rich repeat protein N-terminus is a novel osteoclast antagonist that counteracts bone loss.

    PubMed

    Rucci, Nadia; Capulli, Mattia; Ventura, Luca; Angelucci, Adriano; Peruzzi, Barbara; Tillgren, Viveka; Muraca, Maurizio; Heinegård, Dick; Teti, Anna

    2013-09-01

    (hbd) PRELP is a peptide corresponding to the N-terminal heparin binding domain of the matrix protein proline/arginine-rich end leucine-rich repeat protein (PRELP). (hbd) PRELP inhibits osteoclastogenesis entering pre-fusion osteoclasts through a chondroitin sulfate- and annexin 2-dependent mechanism and reducing the nuclear factor-κB transcription factor activity. In this work, we hypothesized that (hbd) PRELP could have a pharmacological relevance, counteracting bone loss in a variety of in vivo models of bone diseases induced by exacerbated osteoclast activity. In healthy mice, we demonstrated that the peptide targeted the bone and increased trabecular bone mass over basal level. In mice treated with retinoic acid to induce an acute increase of osteoclast formation, the peptide consistently antagonized osteoclastogenesis and prevented the increase of the serum levels of the osteoclast-specific marker tartrate-resistant acid phosphatase. In ovariectomized mice, in which osteoclast activity was chronically enhanced by estrogen deficiency, (hbd) PRELP counteracted exacerbated osteoclast activity and bone loss. In mice carrying osteolytic bone metastases, in which osteoclastogenesis and bone resorption were enhanced by tumor cell-derived factors, (hbd) PRELP reduced the incidence of osteolytic lesions, both preventively and curatively, with mechanisms involving impaired tumor cell homing to bone and tumor growth in the bone microenvironment. Interestingly, in tumor-bearing mice, (hbd) PRELP also inhibited breast tumor growth in orthotopic sites and development of metastatic disease in visceral organs, reducing cachexia and improving survival especially when administered preventively. (hbd) PRELP was retained in the tumor tissue and appeared to affect tumor growth by interacting with the microenvironment rather than by directly affecting the tumor cells. Because safety studies and high-dose treatments revealed no adverse effects, (hbd) PRELP could be employed as a

  16. Leucine modulates the effect of Walker factor, a proteolysis-inducing factor-like protein from Walker tumours, on gene expression and cellular activity in C2C12 myotubes.

    PubMed

    Gonçalves, Estela Maria; Salomão, Emilianne Miguel; Gomes-Marcondes, Maria Cristina Cintra

    2013-10-01

    Cancer-cachexia causes severe weight loss, particularly from the wasting of skeletal muscle, which occurs due to increased protein catabolism and/or decreased protein synthesis. The muscle protein degradation observed in cancer patients is mediated by a specific cytokine, proteolysis-inducing factor (PIF), which is produced by the tumour. This protein increases the ubiquitin-proteasome pathway activity, and the synthesis of muscle protein in these patients can be affected by several factors, including nutrient-related signalling. Some nutrients, such as leucine, can decrease the ubiquitin-proteasome pathway activity and increase the skeletal muscle protein content in cachectic animals. In this study, we investigated the effects of leucine on cell viability, morphology, functional proteasome activity, enzymatic activity, and protein synthesis and degradation in C2C12 myotubes exposed to the proteolysis-inducing factor (PIF)-like protein purified from Walker tumour-bearing rats. Walker factor (WF) had no cytotoxic effects on myotube cells and morphological characteristics were not altered in the presence of WF and/or leucine. However, increased alkaline phosphatase activity was observed. At higher WF concentrations, chymotrypsin-like activity, cathepsin B activity and 20S proteasome gene expression increased. Treating myotubes with leucine before exposure to WF causes leads to a decrease in proteasome activity as well as the activity of chymotrypsin and cathepsin enzymes. Total protein synthesis decreased in WF-treated cells concomitantly as protein degradation increased. After leucine exposure, the observed effects of WF were minimal or even reverted in some cases. Taken together, these results suggest an important modulatory effect for leucine on the effects of WF in C2C12 myotube cells.

  17. Increasing Radiation Therapy Dose Is Associated With Improved Survival in Patients Undergoing Stereotactic Body Radiation Therapy for Stage I Non–Small-Cell Lung Cancer

    SciTech Connect

    Koshy, Matthew; Malik, Renuka; Weichselbaum, Ralph R.; Sher, David J.

    2015-02-01

    Purpose: To determine the comparative effectiveness of different stereotactic body radiation therapy (SBRT) dosing regimens for early-stage non–small-cell lung cancer, using a large national database, focusing on the relative impact of dose as a function of tumor stage. Methods and Materials: The study included patients in the National Cancer Database from 2003 to 2006 with T1-T2N0M0 inoperable lung cancer (n=498). The biologically effective dose (BED) was calculated according to the linear quadratic formula using an α/β ratio of 10. High versus lower-dose (HD vs LD) SBRT was defined as a calculated BED above or below 150 Gy. Overall survival was estimated using Kaplan-Meier methods and Cox proportional hazard regression. Results: The 5 most common dose fractionation schemes (percentage of cohort) used were 20 Gy × 3 (34%), 12 Gy × 4 (16%), 18 Gy × 3 (10%), 15 Gy × 3 (10%), and 16 Gy × 3 (4%). The median calculated BED was 150 Gy (interquartile range 106-166 Gy). The 3-year overall survival (OS) for patients who received HD versus LD was 55% versus 46% (log–rank P=.03). On subset analysis of the T1 cohort there was no association between calculated BED and 3-year OS (61% vs 60% with HD vs LD, P=.9). Among the T2 cohort, patients receiving HD experienced superior 3-year OS (37% vs 24%, P=.01). On multivariable analysis, factors independently prognostic for mortality were female gender (hazard ratio [HR] 0.76, P=.01), T2 tumor (HR 1.99, P=.0001), and HD (HR 0.68, P=.001). Conclusions: This comparative effectiveness analysis of SBRT dose for patients with stage I non–small-cell lung cancer suggests that higher doses (>150 Gy BED) are associated with a significant survival benefit in patients with T2 tumors.

  18. Bifunctional role of leucine 300 of firefly luciferase in structural rigidity.

    PubMed

    Yousefi, Farzad; Ataei, Farangis; Mortazavi, Mojtaba; Hosseinkhani, Saman

    2017-03-18

    Firefly luciferase is susceptible to thermal inactivation, thereby its intracellular half-life decreased. Previous reports indicated that L(300)R mutation (LRR mutant) in E(354)R/Arg(356) double mutant (ERR mutant) from Lampyris turkestanicus luciferase has increased its thermal stability and rigidity through induction of some ionic bonds with Asp 270 and 271. Disruption of the deduced ionic bonds in an ultra-rigid mutant of firefly luciferase did not reverse the flexibility of the protein. In this study, we investigated the effects of this residue to find the truth behind an extraordinary increase in thermal stability and rigidity of luciferase after replacement of leucine 300 by arginine based on previous reports. For this purpose, L(300)R, L(300)K and L(300)E mutations were performed to compare the effects of these mutations on the native firefly luciferase. In spite of increase of intrinsic fluorescence of the mutants a slight increase in thermostability and retention of kinetic properties was observed. Based on our results, we can conclude that L(300)R mutation in LRR mutant accompanying with alteration in a flexible loop (352-359) increased thermostability and rigidity of luciferase.

  19. Intensity-Modulated Radiotherapy Might Increase Pneumonitis Risk Relative to Three-Dimensional Conformal Radiotherapy in Patients Receiving Combined Chemotherapy and Radiotherapy: A Modeling Study of Dose Dumping

    SciTech Connect

    Vogelius, Ivan S.; Westerly, David C.; Cannon, George M.; Mackie, Thomas R.; Mehta, Minesh P.; Sugie, Chikao; Bentzen, Soren M.

    2011-07-01

    Purpose: To model the possible interaction between cytotoxic chemotherapy and the radiation dose distribution with respect to the risk of radiation pneumonitis. Methods and Materials: A total of 18 non-small-cell lung cancer patients previously treated with helical tomotherapy at the University of Wisconsin were selected for the present modeling study. Three treatment plans were considered: the delivered tomotherapy plans; a three-dimensional conformal radiotherapy (3D-CRT) plan; and a fixed-field intensity-modulated radiotherapy (IMRT) plan. The IMRT and 3D-CRT plans were generated specifically for the present study. The plans were optimized without adjusting for the chemotherapy effect. The effect of chemotherapy was modeled as an independent cell killing process by considering a uniform chemotherapy equivalent radiation dose added to all voxels of the organ at risk. The risk of radiation pneumonitis was estimated for all plans using the Lyman and the critical volume models. Results: For radiotherapy alone, the critical volume model predicts that the two IMRT plans are associated with a lower risk of radiation pneumonitis than the 3D-CRT plan. However, when the chemotherapy equivalent radiation dose exceeds a certain threshold, the radiation pneumonitis risk after IMRT is greater than after 3D-CRT. This threshold dose is in the range estimated from clinical chemoradiotherapy data sets. Conclusions: Cytotoxic chemotherapy might affect the relative merit of competing radiotherapy plans. More work is needed to improve our understanding of the interaction between chemotherapy and the radiation dose distribution in clinical settings.

  20. The Effects of Leucine, Zinc, and Chromium Supplements on Inflammatory Events of the Respiratory System in Type 2 Diabetic Rats

    PubMed Central

    Kolahian, Saeed; Sadri, Hassan; Shahbazfar, Amir Ali; Amani, Morvarid; Mazadeh, Anis; Mirani, Mehdi

    2015-01-01

    Diabetes mellitus is a major cause of serious micro- and macrovascular diseases that affect nearly every system in the body, including the respiratory system. Non-enzymatic protein glycation due to hyperglycaemic stress has fundamental implications due to the large capillary network and amount of connective tissue in the lung. The current study was designed to determine whether leucine, zinc, and chromium supplementations influence the function and histological structure of the respiratory tract in a rat model of type 2 diabetes. Seventy-seven rats were divided into eleven groups, consisting of 7 animals each. One group served as negative control and insulin and glibenclamide were used as positive control drugs. Thus, eight groups received the nutritional supplements alone or in combination with each other. Nutritional supplements and glibenclamide were added to the drinking water and neutral protamine Hagedorn insulin was subcutaneously injected during the 4 weeks of treatment period. The induction of type 2 diabetes in the rats caused an infiltration of mononuclear cells and edema in the submucosa of the trachea and lung, severe fibrosis around the vessels and airways, and perivascular and peribronchial infiltration of inflammatory cells and fibrin. In the diabetic group, the total inflammation score and Reid index significantly increased. Diabetes induction significantly reduced the total antioxidant status and elevated the lipid peroxidation products in the serum, lung lavage and lung tissue of the diabetic animals. Treatment with nutritional supplements significantly decreased the histopathological changes and inflammatory indices in the diabetic animals. Supplementation of diabetic rats with leucine, zinc, and chromium, alone and in combination, significantly increased the total antioxidant status and lipid peroxidation level in the diabetic animals. The nutritional supplements improved the enzymatic antioxidant activity of catalase, glutathione peroxidase

  1. The Effects of Leucine, Zinc, and Chromium Supplements on Inflammatory Events of the Respiratory System in Type 2 Diabetic Rats.

    PubMed

    Kolahian, Saeed; Sadri, Hassan; Shahbazfar, Amir Ali; Amani, Morvarid; Mazadeh, Anis; Mirani, Mehdi

    2015-01-01

    Diabetes mellitus is a major cause of serious micro- and macrovascular diseases that affect nearly every system in the body, including the respiratory system. Non-enzymatic protein glycation due to hyperglycaemic stress has fundamental implications due to the large capillary network and amount of connective tissue in the lung. The current study was designed to determine whether leucine, zinc, and chromium supplementations influence the function and histological structure of the respiratory tract in a rat model of type 2 diabetes. Seventy-seven rats were divided into eleven groups, consisting of 7 animals each. One group served as negative control and insulin and glibenclamide were used as positive control drugs. Thus, eight groups received the nutritional supplements alone or in combination with each other. Nutritional supplements and glibenclamide were added to the drinking water and neutral protamine Hagedorn insulin was subcutaneously injected during the 4 weeks of treatment period. The induction of type 2 diabetes in the rats caused an infiltration of mononuclear cells and edema in the submucosa of the trachea and lung, severe fibrosis around the vessels and airways, and perivascular and peribronchial infiltration of inflammatory cells and fibrin. In the diabetic group, the total inflammation score and Reid index significantly increased. Diabetes induction significantly reduced the total antioxidant status and elevated the lipid peroxidation products in the serum, lung lavage and lung tissue of the diabetic animals. Treatment with nutritional supplements significantly decreased the histopathological changes and inflammatory indices in the diabetic animals. Supplementation of diabetic rats with leucine, zinc, and chromium, alone and in combination, significantly increased the total antioxidant status and lipid peroxidation level in the diabetic animals. The nutritional supplements improved the enzymatic antioxidant activity of catalase, glutathione peroxidase

  2. Glucocorticoid Induced Leucine Zipper inhibits apoptosis of cardiomyocytes by doxorubicin

    SciTech Connect

    Aguilar, David; Strom, Joshua; Chen, Qin M.

    2014-04-01

    Doxorubicin (Dox) is an indispensable chemotherapeutic agent for the treatment of various forms of neoplasia such as lung, breast, ovarian, and bladder cancers. Cardiotoxicity is a major concern for patients receiving Dox therapy. Previous work from our laboratory indicated that glucocorticoids (GCs) alleviate Dox-induced apoptosis in cardiomyocytes. Here we have found Glucocorticoid-Induced Leucine Zipper (GILZ) to be a mediator of GC-induced cytoprotection. GILZ was found to be induced in cardiomyocytes by GC treatment. Knocking down of GILZ using siRNA resulted in cancelation of GC-induced cytoprotection against apoptosis by Dox treatment. Overexpressing GILZ by transfection was able to protect cells from apoptosis induced by Dox as measured by caspase activation, Annexin V binding and morphologic changes. Western blot analyses indicate that GILZ overexpression prevented cytochrome c release from mitochondria and cleavage of caspase-3. When bcl-2 family proteins were examined, we found that GILZ overexpression causes induction of the pro-survival protein Bcl-xL. Since siRNA against Bcl-xL reverses GC induced cytoprotection, Bcl-xL induction represents an important event in GILZ-induced cytoprotection. Our data suggest that GILZ functions as a cytoprotective gene in cardiomyocytes. - Highlights: • Corticosteroids act as a cytoprotective agent in cardiomyocytes • Corticosteroids induce GILZ expression in cardiomyocytes • Elevated GILZ results in resistance against apoptosis induced by doxorubicin • GILZ induces Bcl-xL protein without inducing Bcl-xL mRNA.

  3. Do non-targeted effects increase or decrease low dose risk in relation to the linear-non-threshold (LNT) model?

    PubMed

    Little, M P

    2010-05-01

    In this paper we review the evidence for departure from linearity for malignant and non-malignant disease and in the light of this assess likely mechanisms, and in particular the potential role for non-targeted effects. Excess cancer risks observed in the Japanese atomic bomb survivors and in many medically and occupationally exposed groups exposed at low or moderate doses are generally statistically compatible. For most cancer sites the dose-response in these groups is compatible with linearity over the range observed. The available data on biological mechanisms do not provide general support for the idea of a low dose threshold or hormesis. This large body of evidence does not suggest, indeed is not statistically compatible with, any very large threshold in dose for cancer, or with possible hormetic effects, and there is little evidence of the sorts of non-linearity in response implied by non-DNA-targeted effects. There are also excess risks of various types of non-malignant disease in the Japanese atomic bomb survivors and in other groups. In particular, elevated risks of cardiovascular disease, respiratory disease and digestive disease are observed in the A-bomb data. In contrast with cancer, there is much less consistency in the patterns of risk between the various exposed groups; for example, radiation-associated respiratory and digestive diseases have not been seen in these other (non-A-bomb) groups. Cardiovascular risks have been seen in many exposed populations, particularly in medically exposed groups, but in contrast with cancer there is much less consistency in risk between studies: risks per unit dose in epidemiological studies vary over at least two orders of magnitude, possibly a result of confounding and effect modification by well known (but unobserved) risk factors. In the absence of a convincing mechanistic explanation of epidemiological evidence that is, at present, less than persuasive, a cause-and-effect interpretation of the reported

  4. Computer-aided design and activity prediction of leucine aminopeptidase inhibitors

    NASA Astrophysics Data System (ADS)

    Grembecka, J.; Sokalski, W. A.; Kafarski, P.

    2000-08-01

    The Ligand Design (LUDI) approach has been used in order to design leucine aminopeptidase inhibitors, predict their activity and analyze their interactions with the enzyme. The investigation was based on the crystal structure of bovine lens leucine aminopeptidase (LAP) complexed with its inhibitor - the phosphonic acid analogue of leucine (LeuP). More than 50 potential leucine aminopeptidase inhibitors have been obtained, including the most potent aminophosphonic LAP inhibitors with experimentally known activity, which have been the subject of more detailed studies. A reasonable agreement between theoretical and experimental activities has been obtained for most of the studied inhibitors. Our results confirm that LUDI is a powerful tool for the design of enzyme inhibitors as well as in the prediction of their activity. In addition, for inhibitor-active site interactions dominated by the electrostatic effects it is possible to improve binding energy estimates by using a more accurate description of inhibitor charge distribution.

  5. Leucine incorporation into the membranellar bands of regenerating and nonregenerating stentor.

    PubMed

    de Terra, N

    1966-07-29

    Autoradiographs of membranellar bands isolated from regenerating and nonregenerating stentors labeled with tritiated leucine indicate that the synthesis of proteins of the oral apparatus is continuous in Stentor and is not initiated at the time of regeneration.

  6. Up-regulation of leucine aminopeptidase-A in cadmium-treated tomato roots.

    PubMed

    Boulila-Zoghlami, Latifa; Gallusci, Philippe; Holzer, Frances M; Basset, Gilles J; Djebali, Whabi; Chaïbi, Wided; Walling, Linda L; Brouquisse, Renaud

    2011-10-01

    The effects of cadmium (Cd) on aminopeptidase (AP) activities and Leucine-AP (LAP) expression were investigated in the roots of tomato (Solanum lycopersicum L., var Ibiza) plants. Three-week-old plants were grown for 10 days in the presence of 0.3-300 μM Cd and compared to control plants grown in the absence of Cd. AP activities were measured using six different p-nitroanilide (p-NA) substrates. Leu, Met, Arg, Pro and Lys hydrolyzing activities increased in roots of Cd-treated plants, while Phe-pNA cleavage was not enhanced after Cd treatments. The use of peptidase inhibitors showed that most of the Leu-pNA hydrolyzing activity was related to one or several metallo-APs. Changes in Lap transcripts, protein and activities were measured in the roots of 0 and 30-μM Cd-treated plants. LapA transcript levels increased in Cd-treated roots, whereas LapN RNAs levels were not modified. To assess amount of Leu-pNA hydrolyzing activity associated with the hexameric LAPs, LAP activity was measured following immunoprecipitation with a LAP polyclonal antiserum. LAP activity increased in Cd-treated roots. There was a corresponding increase in LAP-A protein levels detected in 2D-immunoblots. The role of LAP-A in the proteolytic response to Cd stress is discussed.

  7. Differential Assimilation of Inorganic Carbon and Leucine by Prochlorococcus in the Oligotrophic North Pacific Subtropical Gyre

    PubMed Central

    Björkman, Karin M.; Church, Matthew J.; Doggett, Joseph K.; Karl, David M.

    2015-01-01

    The light effect on photoheterotrophic processes in Prochlorococcus, and primary and bacterial productivity in the oligotrophic North Pacific Subtropical Gyre was investigated using 14C-bicarbonate and 3H-leucine. Light and dark incubation experiments were conducted in situ throughout the euphotic zone (0–175 m) on nine expeditions to Station ALOHA over a 3-year period. Photosynthetrons were also used to elucidate rate responses in leucine and inorganic carbon assimilation as a function of light intensity. Taxonomic group and cell-specific rates were assessed using flow cytometric sorting. The light:dark assimilation rate ratios of leucine in the top 150 m were ∼7:1 for Prochlorococcus, whereas the light:dark ratios for the non-pigmented bacteria (NPB) were not significant different from 1:1. Prochlorococcus assimilated leucine in the dark at per cell rates similar to the NPB, with a contribution to the total community bacterial production, integrated over the euphotic zone, of approximately 20% in the dark and 60% in the light. Depth-resolved primary productivity and leucine incorporation showed that the ratio of Prochlorococcus leucine:primary production peaked at 100 m then declined steeply below the deep chlorophyll maximum (DCM). The photosynthetron experiments revealed that, for Prochlorococcus at the DCM, the saturating irradiance (Ek) for leucine incorporation was reached at approximately half the light intensity required for light saturation of 14C-bicarbonate assimilation. Additionally, high and low red fluorescing Prochlorococcus populations (HRF and LRF), co-occurring at the DCM, had similar Ek values for their respective substrates, however, maximum assimilation rates, for both leucine and inorganic carbon, were two times greater for HRF cells. Our results show that Prochlorococcus contributes significantly to bacterial production estimates using 3H-leucine, whether or not the incubations are conducted in the dark or light, and this should be

  8. Differential Assimilation of Inorganic Carbon and Leucine by Prochlorococcus in the Oligotrophic North Pacific Subtropical Gyre.

    PubMed

    Björkman, Karin M; Church, Matthew J; Doggett, Joseph K; Karl, David M

    2015-01-01

    The light effect on photoheterotrophic processes in Prochlorococcus, and primary and bacterial productivity in the oligotrophic North Pacific Subtropical Gyre was investigated using (14)C-bicarbonate and (3)H-leucine. Light and dark incubation experiments were conducted in situ throughout the euphotic zone (0-175 m) on nine expeditions to Station ALOHA over a 3-year period. Photosynthetrons were also used to elucidate rate responses in leucine and inorganic carbon assimilation as a function of light intensity. Taxonomic group and cell-specific rates were assessed using flow cytometric sorting. The light:dark assimilation rate ratios of leucine in the top 150 m were ∼7:1 for Prochlorococcus, whereas the light:dark ratios for the non-pigmented bacteria (NPB) were not significant different from 1:1. Prochlorococcus assimilated leucine in the dark at per cell rates similar to the NPB, with a contribution to the total community bacterial production, integrated over the euphotic zone, of approximately 20% in the dark and 60% in the light. Depth-resolved primary productivity and leucine incorporation showed that the ratio of Prochlorococcus leucine:primary production peaked at 100 m then declined steeply below the deep chlorophyll maximum (DCM). The photosynthetron experiments revealed that, for Prochlorococcus at the DCM, the saturating irradiance (E k) for leucine incorporation was reached at approximately half the light intensity required for light saturation of (14)C-bicarbonate assimilation. Additionally, high and low red fluorescing Prochlorococcus populations (HRF and LRF), co-occurring at the DCM, had similar E k values for their respective substrates, however, maximum assimilation rates, for both leucine and inorganic carbon, were two times greater for HRF cells. Our results show that Prochlorococcus contributes significantly to bacterial production estimates using (3)H-leucine, whether or not the incubations are conducted in the dark or light, and this should

  9. Geraniol attenuates α-synuclein expression and neuromuscular impairment through increase dopamine content in MPTP intoxicated mice by dose dependent manner.

    PubMed

    Rekha, Karamkolly R; Selvakumar, Govindasamy P; Santha, Karunanithi; Inmozhi Sivakamasundari, Ramu

    2013-11-01

    Parkinson's disease (PD) is characterized by progressive loss of dopamine (DA) neurons in the nigrostriatal system and by the presence of Lewy bodies (LB), proteinaceous inclusions mainly composed of filamentous α-synuclein (α-Syn) aggregates. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was adopted to generate PD models in C57BL/6 mice. In the present study, we investigated the effect of geraniol (GE) against α-Syn aggregation on MPTP induced mouse model of PD in dose dependant manner. When pretreatment of GE improved neuromuscular impairment, TH expressions and decreases α-Syn expressions in MPTP intoxicated PD mice by dose dependent manner. In addition, we confirmed that sub-chronic administration of MPTP in mice leads to permanent neuromuscular deficits and depletion of dopamine and its metabolites. Our results suggest that GE is beneficial for the treatment of PD associated with neuromuscular disability and LB aggregation.

  10. Increased pattern transfer fidelity ZEP 520A during reactive ion etching through chemical modifications by additional dosing of the electron beam resist.

    SciTech Connect

    Czaplewski, D. A.; Ocola, L. E.

    2011-03-01

    This article describes a postdevelopment, additional electron exposure to enhance the etch selectivity and improve pattern transfer fidelity of an electron beam resist, ZEP 520A, through chemical changes of the resist. After the critical features were patterned and developed, the resist was exposed at 5 kV accelerating voltage to a second dose of electrons ranging from 300 to 300,000 {micro}C/cm{sup 2}. The etch rate of the resist decreased by approximately 25% in a CHF{sub 3} and O{sub 2} plasma. More critically, the fidelity of the pattern transfer was improved. Infrared and Raman spectroscopies were used to characterize the resist before and after electron beam exposure for doses up to 3000 {micro}C/cm{sup 2}. The carbonyl bonding in the polymer showed significant changes after electron beam exposure that can be associated with improvement in the etch performance of this resist.

  11. Increasing Dose of Autologous Bone Marrow Mononuclear Cells Transplantation Is Related to Stroke Outcome: Results from a Pooled Analysis of Two Clinical Trials

    PubMed Central

    Escudero, Irene; Zapata, Elena; de la Torre Laviana, Francisco Javier; Carmona, Magdalena; Piñero, Pilar; Bustamante, Alejandro; Lebrato, Lucía; Cabezas, Juan Antonio; Gonzalez, Alejandro; de Freitas, Grabriel R.; Montaner, Joan

    2016-01-01

    Background and Purpose. BM-MNC transplantation improves recovery in experimental models of ischemic stroke. Clinical trials are ongoing to test efficacy in stroke patients. However, whether cell dose is related to outcomes is not known. Methods. We performed a pooling data analysis of two pilot clinical trials with autologous BM-MNCs transplantation in ischemic stroke patients. Cell dose and route were analyzed to evaluate their relation to good outcome (m-Rankin scale [mRS] score 0–2) at 6 months. Results. Twenty-two patients were included. A median of 153 × 106 (±121 × 106) BM-MNCs was injected. Intra-arterial route was used in 77.3% of cases. A higher number of cells injected were associated with better outcomes at 180 days (390 × 106 [320–422] BM-MNCs injected in those patients with mRS of 0–2 at 6 months versus 130 × 106 [89–210] in those patients with mRS 3–6, p = 0.015). In the intra-arterially treated patients, a strong correlation between dose of cells and disability was found (r = −0.63, p = 0.006). A cut point of 310 × 106 injected cells predicted good outcome with 80% sensitivity and 88.2% specificity. Conclusions. Similar to preclinical studies, a higher dose of autologous BM-MNC was related to better outcome in stroke patients, especially when more than 310 × 106 cells are injected. Further interventional studies are warranted to confirm these data. PMID:27525011

  12. K-Ras mutant fraction in A/J mouse lung increases as a function of benzo[a]pyrene dose

    EPA Science Inventory

    K-Ras mutant fraction (MF) was measured to examine the default assumption of low dose linearity in the benzo[a]pyrene (B[a]P) mutational response. Groups of ten male A/J mice (7-9 weeks-old) received a single i.p. injection of 0, 0.05, 0.5, 5, or 50 mg/kg B[a]P, and were sacrifi...

  13. Results of autologous transplantation in lymphoma are not improved by increasing the dose of etoposide in the BEAM regimen: a single-centre sequential-cohort study.

    PubMed

    Martín, A; Caballero, M D; Pérez-Simón, J A; López-Holgado, N; Mateos, M V; Cañizo, M C; Miguel, J F San

    2004-10-01

    We have undertaken a retrospective sequential-cohort analysis of 131 lymphoma patients treated with the BEAM regimen and autologous stem cell transplantation, to compare BEAM at standard doses (sBEAM; n = 67 from May 1990 to April 1995) and BEAM with escalated etoposide dose from 800 to 1600 mg/m(2) (eBEAM; n = 64 from May 1995 to June 1999). Transplant-related mortality and incidence of secondary malignancies were similar in both groups. Disease progression was significantly lower in indolent lymphoma (IL) patients receiving eBEAM (7 vs 43%), although survival was comparable due to a higher toxic mortality in the eBEAM group. The 5-year event-free survival and overall survival were better in Hodgkin's disease (HD) patients treated with eBEAM (70 and 77%, respectively) compared to sBEAM (58 and 69%, respectively), but the difference was not statistically significant. In aggressive lymphomas, no difference was detected between groups. Our results indicate that while escalation of the etoposide doses in the BEAM conditioning regimen does not appear to improve outcome, encouraging results in IL and HD may warrant further studies.

  14. Metabolic mechanism for L-leucine-induced metabolome to eliminate Streptococcus iniae.

    PubMed

    Du, Chao-Chao; Yang, Manjun; Li, Min-Yi; Yang, Jun; Peng, Bo; Li, Hui; Peng, Xuan-Xian

    2017-03-07

    Crucial metabolites that modulate hosts' metabolome to eliminate bacterial pathogens have been documented, but the metabolic mechanisms are largely unknown. The present study explores the metabolic mechanism for L-leucine-induced metabolome to eliminate Streptococcus iniae in tilapia. GC-MS based metabolomics was used to investigate tilapia liver metabolic profile in the presence of exogenous L-leucine. Thirty-seven metabolites of differential abundance were determined, and eleven metabolic pathways were enriched. Pattern recognition analysis identified serine and proline as crucial metabolites, which are the two metabolites identified in survived tilapias during S. iniae infection, suggesting the two metabolites play crucial roles in L-leucine-induced elimination of the pathogen by the host. Exogenous L-serine reduces mortality of tilapias infected by S. iniae, providing a robust proof for supporting the conclusion. Furthermore, exogenous serine elevates expression of genes Il-1β and Il-8 in tilapia spleen, but not TNFα, CXCR4 and Mx, suggesting the metabolite promotes a phagocytosis role of macrophages, which is consistent with the finding that L-leucine promotes macrophages to kill both Gram-positive and negative bacterial pathogens. Therefore, the ability of phagocytosis enhanced by exogenous L-leucine is partly attributed to elevation of serine. These results demonstrate a metabolic mechanism by which exogenous L-leucine modulates tilapias' metabolome to enhance innate immunity and eliminate pathogens.

  15. How to find a leucine in a haystack? Structure, ligand recognition and regulation of leucine-aspartic acid (LD) motifs.

    PubMed

    Alam, Tanvir; Alazmi, Meshari; Gao, Xin; Arold, Stefan T

    2014-06-15

    LD motifs (leucine-aspartic acid motifs) are short helical protein-protein interaction motifs that have emerged as key players in connecting cell adhesion with cell motility and survival. LD motifs are required for embryogenesis, wound healing and the evolution of multicellularity. LD motifs also play roles in disease, such as in cancer metastasis or viral infection. First described in the paxillin family of scaffolding proteins, LD motifs and similar acidic LXXLL interaction motifs have been discovered in several other proteins, whereas 16 proteins have been reported to contain LDBDs (LD motif-binding domains). Collectively, structural and functional analyses have revealed a surprising multivalency in LD motif interactions and a wide diversity in LDBD architectures. In the present review, we summarize the molecular basis for function, regulation and selectivity of LD motif interactions that has emerged from more than a decade of research. This overview highlights the intricate multi-level regulation and the inherently noisy and heterogeneous nature of signalling through short protein-protein interaction motifs.

  16. Leucine supplementation of a chronically restricted protein and energy diet enhances mTOR pathway activation but not muscle protein synthesis in neonatal pigs.

    PubMed

    Manjarín, Rodrigo; Columbus, Daniel A; Suryawan, Agus; Nguyen, Hanh V; Hernandez-García, Adriana D; Hoang, Nguyet-Minh; Fiorotto, Marta L; Davis, Teresa

    2016-01-01

    Suboptimal nutrient intake represents a limiting factor for growth and long-term survival of low-birth weight infants. The objective of this study was to determine if in neonates who can consume only 70 % of their protein and energy requirements for 8 days, enteral leucine supplementation will upregulate the mammalian target of rapamycin (mTOR) pathway in skeletal muscle, leading to an increase in protein synthesis and muscle anabolism. Nineteen 4-day-old piglets were fed by gastric tube 1 of 3 diets, containing (kg body weight(-1) · day(-1)) 16 g protein and 190 kcal (CON), 10.9 g protein and 132 kcal (R), or 10.8 g protein + 0.2 % leucine and 136 kcal (RL) at 4-h intervals for 8 days. On day 8, plasma AA and insulin levels were measured during 6 post-feeding intervals, and muscle protein synthesis rate and mTOR signaling proteins were determined at 120 min post-feeding. At 120 min, leucine was highest in RL (P < 0.001), whereas insulin, isoleucine and valine were lower in RL and R compared to CON (P < 0.001). Compared to RL and R, the CON diet increased (P < 0.01) body weight, protein synthesis, phosphorylation of S6 kinase (p-S6K1) and 4E-binding protein (p-4EBP1), and activation of eukaryotic initiation factor 4 complex (eIF4E · eIF4G). RL increased (P ≤ 0.01) p-S6K1, p-4EBP1 and eIF4E · eIF4G compared to R. In conclusion, when protein and energy intakes are restricted for 8 days, leucine supplementation increases muscle mTOR activation, but does not improve body weight gain or enhance skeletal muscle protein synthesis in neonatal pigs.

  17. Changes in labeling of soluble and solubilized rat brain proteins using (3H)-leucine as precursor during a learning experiment.

    PubMed

    Popov, N; Schulzeck, S; Schmidt, S; Matthies, H

    1975-01-01

    This paper deals with an electrophoretic study on soluble and solubilized rat brain proteins during a brightness discrimination. After intraventricular injection of L-[3H]-leucine the most pronounced increase in relative specific activity was found for several soluble acidic protein-band complexes as well as for solubilized slow-migrating non-myelin proteins obtained from hippocampus of trained animals over the data obtained for active and passive controls.

  18. Leucine supplementation of a chronically restricted protein and energy diet enhances mTOR pathway activation but not muscle protein synthesis in neonatal pigs

    PubMed Central

    Suryawan, Agus; Nguyen, Hanh V.; Hernandez-García, Adriana D.; Hoang, Nguyet-Minh; Fiorotto, Marta L.; Davis, Teresa

    2016-01-01

    Suboptimal nutrient intake represents a limiting factor for growth and long-term survival of low-birth weight infants. The objective of this study was to determine if in neonates who can consume only 70 % of their protein and energy requirements for 8 days, enteral leucine supplementation will upregulate the mammalian target of rapamycin (mTOR) pathway in skeletal muscle, leading to an increase in protein synthesis and muscle anabolism. Nineteen 4-day-old piglets were fed by gastric tube 1 of 3 diets, containing (kg body weight−1·day−1) 16 g protein and 190 kcal (CON), 10.9 g protein and 132 kcal (R), or 10.8 g protein + 0.2 % leucine and 136 kcal (RL) at 4-h intervals for 8 days. On day 8, plasma AA and insulin levels were measured during 6 post-feeding intervals, and muscle protein synthesis rate and mTOR signaling proteins were determined at 120 min post-feeding. At 120 min, leucine was highest in RL (P < 0.001), whereas insulin, isoleucine and valine were lower in RL and R compared to CON (P < 0.001). Compared to RL and R, the CON diet increased (P < 0.01) body weight, protein synthesis, phosphorylation of S6 kinase (p-S6K1) and 4E-binding protein (p-4EBP1), and activation of eukaryotic initiation factor 4 complex (eIF4E·eIF4G). RL increased (P ≤ 0.01) p-S6K1, p-4EBP1 and eIF4E · eIF4G compared to R. In conclusion, when protein and energy intakes are restricted for 8 days, leucine supplementation increases muscle mTOR activation, but does not improve body weight gain or enhance skeletal muscle protein synthesis in neonatal pigs. PMID:26334346

  19. Expression and regulation of glucocorticoid-induced leucine zipper in the developing anterior pituitary gland.

    PubMed

    Ellestad, Laura E; Malkiewicz, Stefanie A; Guthrie, H David; Welch, Glenn R; Porter, Tom E

    2009-02-01

    The expression profile of glucocorticoid-induced leucine zipper (GILZ) in the anterior pituitary during the second half of embryonic development in the chick is consistent with in vivo regulation by circulating corticosteroids. However, nothing else has been reported about the presence of GILZ in the neuroendocrine system. We sought to characterize expression and regulation of GILZ in the chicken embryonic pituitary gland and determine the effect of GILZ overexpression on anterior pituitary hormone levels. Pituitary GILZ mRNA levels increased during embryogenesis to a maximum on the day of hatch, and decreased through the first week after hatch. GILZ expression was rapidly upregulated by corticosterone in embryonic pituitary cells. To determine whether GILZ regulates hormone gene expression in the developing anterior pituitary, we overexpressed GILZ in embryonic pituitary cells and measured mRNA for the major pituitary hormones. Exogenous GILZ increased prolactin mRNA above basal levels, but not as high as that in corticosterone-treated cells, indicating that GILZ may play a small role in lactotroph differentiation. The largest effect we observed was a twofold increase in FSH beta subunit in cells transfected with GILZ but not treated with corticosterone, suggesting that GILZ may positively regulate gonadotroph development in a manner not involving glucocorticoids. In conclusion, this is the first report to characterize avian GILZ and examine its regulation in the developing neuroendocrine system. We have shown that GILZ is upregulated by glucocorticoids in the embryonic pituitary gland and may regulate expression of several pituitary hormones.

  20. Acute depletion of plasma glutamine increases leucine oxidation in prednisone-treated humans.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To determine whether depletion in plasma glutamine worsens the catabolic response to corticosteroids, seven healthy volunteers received oral prednisone for 6 days on two separate occasions, at least 2 weeks apart, and in random order. On the sixth day of each treatment course, they received 5 h intr...

  1. Low-dose radiation from 18F-FDG PET does not increase cancer frequency or shorten latency but reduces kidney disease in cancer-prone Trp53+/- mice

    DOE PAGES

    Taylor, Kristina; Lemon, Jennifer A.; Phan, Nghi; ...

    2014-05-28

    There is considerable interest in the health effects associated with low-level radiation exposure from medical imaging procedures. Concerns in the medical community that increased radiation exposure from imaging procedures may increase cancer risk among patients are confounded by research showing that low-dose radiation exposure can extend lifespan by increasing the latency period of some types of cancer. The most commonly used radiopharmaceutical for positron emission tomography (PET) scans is 2-[18F] fluoro-2-deoxy-d-glucose (18F-FDG), which exposes tissue to a low-dose, mixed radiation quality: 634 keV β+ and 511 keV γ-rays. The goal of this research was to investigate how modification of cancermore » risk associated with exposure to low-dose ionising radiation in cancer-prone Trp53+/- mice is influenced by radiation quality from PET. At 7-8 weeks of age, Trp53+/- female mice were exposed to one of five treatments: 0 Gy, 10 mGy γ-rays, 10 mGy 18F-FDG, 4 Gy γ-rays, 10 mGy 18F-FDG + 4 Gy γ-rays (n > 185 per group). The large 4-Gy radiation dose significantly reduced the lifespan by shortening the latency period of cancer and significantly increasing the number of mice with malignancies, compared with unirradiated controls. The 10 mGy γ-rays and 10 mGy PET doses did not significantly modify the frequency or latency period of cancer relative to unirradiated mice. Similarly, the PET scan administered prior to a large 4-Gy dose did not significantly modify the latency or frequency of cancer relative to mice receiving a dose of only 4 Gy. The relative biological effectiveness of radiation quality from 18F-FDG, with respect to malignancy, is approximately 1. Furthermore, when non-cancer endpoints were studied, it was found that the 10-mGy PET group had a significant reduction in kidney lesions (P < 0.021), indicating that a higher absorbed dose (20 ± 0.13 mGy), relative to the whole-body average, which occurs in specific tissues, may not be detrimental.« less

  2. Low-dose radiation from 18F-FDG PET does not increase cancer frequency or shorten latency but reduces kidney disease in cancer-prone Trp53+/- mice

    SciTech Connect

    Taylor, Kristina; Lemon, Jennifer A.; Phan, Nghi; Boreham, Douglas R.

    2014-05-28

    There is considerable interest in the health effects associated with low-level radiation exposure from medical imaging procedures. Concerns in the medical community that increased radiation exposure from imaging procedures may increase cancer risk among patients are confounded by research showing that low-dose radiation exposure can extend lifespan by increasing the latency period of some types of cancer. The most commonly used radiopharmaceutical for positron emission tomography (PET) scans is 2-[18F] fluoro-2-deoxy-d-glucose (18F-FDG), which exposes tissue to a low-dose, mixed radiation quality: 634 keV β+ and 511 keV γ-rays. The goal of this research was to investigate how modification of cancer risk associated with exposure to low-dose ionising radiation in cancer-prone Trp53+/- mice is influenced by radiation quality from PET. At 7-8 weeks of age, Trp53+/- female mice were exposed to one of five treatments: 0 Gy, 10 mGy γ-rays, 10 mGy 18F-FDG, 4 Gy γ-rays, 10 mGy 18F-FDG + 4 Gy γ-rays (n > 185 per group). The large 4-Gy radiation dose significantly reduced the lifespan by shortening the latency period of cancer and significantly increasing the number of mice with malignancies, compared with unirradiated controls. The 10 mGy γ-rays and 10 mGy PET doses did not significantly modify the frequency or latency period of cancer relative to unirradiated mice. Similarly, the PET scan administered prior to a large 4-Gy dose did not significantly modify the latency or frequency of cancer relative to mice receiving a dose of only 4 Gy. The relative biological effectiveness of radiation quality from 18F-FDG, with respect to malignancy, is approximately 1. Furthermore, when non-cancer endpoints were studied, it was found that the 10-mGy PET group had a significant reduction in kidney lesions (P < 0.021), indicating that a higher absorbed dose (20 ± 0.13 mGy), relative to the whole-body average

  3. Low-dose radiation from 18F-FDG PET does not increase cancer frequency or shorten latency but reduces kidney disease in cancer-prone Trp53+/- mice.

    PubMed

    Taylor, Kristina; Lemon, Jennifer A; Phan, Nghi; Boreham, Douglas R

    2014-07-01

    There is considerable interest in the health effects associated with low-level radiation exposure from medical imaging procedures. Concerns in the medical community that increased radiation exposure from imaging procedures may increase cancer risk among patients are confounded by research showing that low-dose radiation exposure can extend lifespan by increasing the latency period of some types of cancer. The most commonly used radiopharmaceutical for positron emission tomography (PET) scans is 2-[(18)F] fluoro-2-deoxy-d-glucose ((18)F-FDG), which exposes tissue to a low-dose, mixed radiation quality: 634 keV β+ and 511 keV γ-rays. The goal of this research was to investigate how modification of cancer risk associated with exposure to low-dose ionising radiation in cancer-prone Trp53+/- mice is influenced by radiation quality from PET. At 7-8 weeks of age, Trp53+/- female mice were exposed to one of five treatments: 0 Gy, 10 mGy γ-rays, 10 mGy (18)F-FDG, 4 Gy γ-rays, 10 mGy (18)F-FDG + 4 Gy γ-rays (n > 185 per group). The large 4-Gy radiation dose significantly reduced the lifespan by shortening the latency period of cancer and significantly increasing the number of mice with malignancies, compared with unirradiated controls. The 10 mGy γ-rays and 10 mGy PET doses did not significantly modify the frequency or latency period of cancer relative to unirradiated mice. Similarly, the PET scan administered prior to a large 4-Gy dose did not significantly modify the latency or frequency of cancer relative to mice receiving a dose of only 4 Gy. The relative biological effectiveness of radiation quality from (18)F-FDG, with respect to malignancy, is approximately 1. However; when non-cancer endpoints were studied, it was found that the 10-mGy PET group had a significant reduction in kidney lesions (P < 0.021), indicating that a higher absorbed dose (20 ± 0.13 mGy), relative to the whole-body average, which occurs in specific tissues, may not be detrimental.

  4. Terminate lung cancer (TLC) study-A mixed-methods population approach to increase lung cancer screening awareness and low-dose computed tomography in Eastern Kentucky.

    PubMed

    Cardarelli, Roberto; Reese, David; Roper, Karen L; Cardarelli, Kathryn; Feltner, Frances J; Studts, Jamie L; Knight, Jennifer R; Armstrong, Debra; Weaver, Anthony; Shaffer, Dana

    2017-02-01

    For low dose CT lung cancer screening to be effective in curbing disease mortality, efforts are needed to overcome barriers to awareness and facilitate uptake of the current evidence-based screening guidelines. A sequential mixed-methods approach was employed to design a screening campaign utilizing messages developed from community focus groups, followed by implementation of the outreach campaign intervention in two high-risk Kentucky regions. This study reports on rates of awareness and screening in intervention regions, as compared to a control region.

  5. Differential activities of glucocorticoid-induced leucine zipper protein isoforms.

    PubMed

    Soundararajan, Rama; Wang, Jian; Melters, Daniël; Pearce, David

    2007-12-14

    Glucocorticoid-induced leucine zipper protein (GILZ) is expressed in both epithelial and immune tissues and modulates a variety of cellular functions, including proliferation and epithelial sodium channel (ENaC) activity. A number of reports have described various GILZ activities, focusing on a single isoform with molecular mass of approximately 17 kDa, now termed GILZ1. In GILZ immunoblots using a newly developed antiserum, we detected multiple species in extracts from cultured kidney cells. Mass spectrometric analysis revealed that one of these represented a previously uncharacterized distinct isoform of GILZ, GILZ2. Rapid amplification of cDNA ends was used to clone cDNAs corresponding to four isoforms, which, in addition to GILZ1 and GILZ2, included new isoforms GILZ3 and GILZ4. Heterologous expression of these four GILZ isoforms in cultured cells revealed striking functional differences. Notably, GILZ1 was the only isoform that significantly stimulated ENaC-mediated Na+ current in a kidney collecting duct cell line, although GILZ2 and GILZ3 also stimulated ENaC surface expression in HEK 293 cells. GILZ1 and GILZ3, and to a lesser extent GILZ2, inhibited ERK phosphorylation. Interestingly, GILZ4, which had no effect on either ENaC or ERK, potently suppressed cellular proliferation, as did GILZ1, but not GILZ2 or GILZ3. Finally, rat and mouse tissues all expressed multiple GILZ species but varied in the relative abundance of each. These data suggest that multiple GILZ isoforms are expressed in most cells and tissues and that these play distinct roles in regulating key cellular functions, including proliferation and ion transport. Furthermore, GILZ inhibition of ERK appears to play an essential role in stimulation of cell surface ENaC but not in inhibition of proliferation.

  6. Distribution and Evolution of Yersinia Leucine-Rich Repeat Proteins

    PubMed Central

    Hu, Yueming; Huang, He; Hui, Xinjie; Cheng, Xi; White, Aaron P.

    2016-01-01

    Leucine-rich repeat (LRR) proteins are widely distributed in bacteria, playing important roles in various protein-protein interaction processes. In Yersinia, the well-characterized type III secreted effector YopM also belongs to the LRR protein family and is encoded by virulence plasmids. However, little has been known about other LRR members encoded by Yersinia genomes or their evolution. In this study, the Yersinia LRR proteins were comprehensively screened, categorized, and compared. The LRR proteins encoded by chromosomes (LRR1 proteins) appeared to be more similar to each other and different from those encoded by plasmids (LRR2 proteins) with regard to repeat-unit length, amino acid composition profile, and gene expression regulation circuits. LRR1 proteins were also different from LRR2 proteins in that the LRR1 proteins contained an E3 ligase domain (NEL domain) in the C-terminal region or an NEL domain-encoding nucleotide relic in flanking genomic sequences. The LRR1 protein-encoding genes (LRR1 genes) varied dramatically and were categorized into 4 subgroups (a to d), with the LRR1a to -c genes evolving from the same ancestor and LRR1d genes evolving from another ancestor. The consensus and ancestor repeat-unit sequences were inferred for different LRR1 protein subgroups by use of a maximum parsimony modeling strategy. Structural modeling disclosed very similar repeat-unit structures between LRR1 and LRR2 proteins despite the different unit lengths and amino acid compositions. Structural constraints may serve as the driving force to explain the observed mutations in the LRR regions. This study suggests that there may be functional variation and lays the foundation for future experiments investigating the functions of the chromosomally encoded LRR proteins of Yersinia. PMID:27217422

  7. Biomimetic Enamel Regeneration Mediated by Leucine-Rich Amelogenin Peptide.

    PubMed

    Kwak, S Y; Litman, A; Margolis, H C; Yamakoshi, Y; Simmer, J P

    2017-01-01

    We report here a novel biomimetic approach to the regeneration of human enamel. The approach combines the use of inorganic pyrophosphate (PPi) to control the onset and rate of enamel regeneration and the use of leucine-rich amelogenin peptide (LRAP), a nonphosphorylated 56-amino acid alternative splice product of amelogenin, to regulate the shape and orientation of growing enamel crystals. This study builds on our previous findings that show LRAP can effectively guide the formation of ordered arrays of needle-like hydroxyapatite (HA) crystals in vitro and on the known role mineralization inhibitors, like PPi, play in the regulation of mineralized tissue formation. Acid-etched enamel surfaces of extracted human molars, cut perpendicular or parallel to the direction of the enamel rods, were exposed to a PPi-stabilized supersaturated calcium phosphate (CaP) solution containing 0 to 0.06 mg/mL LRAP for 20 h. In the absence of LRAP, PPi inhibition was reversed by the presence of etched enamel surfaces and led to the formation of large, randomly distributed plate-like HA crystals that were weakly attached, regardless of rod orientation. In the presence of 0.04 mg/mL LRAP, however, densely packed mineral layers, comprising bundles of small needle-like HA crystals, formed on etched surfaces that were cut perpendicular to the enamel rods. These crystals were strongly attached, and their arrangement reflected to a significant degree the underlying enamel prism pattern. In contrast, under the same conditions with LRAP, little to no crystal formation was found on enamel surfaces that were cut parallel to the direction of the enamel rods. These results suggest that LRAP preferentially interacts with ab surfaces of mature enamel crystals, inhibiting their directional growth, thus selectively promoting linear growth along the c-axis of enamel crystals. The present findings demonstrate a potential for the development of a new approach to regenerate enamel structure and properties.

  8. Dose‐dependent increases in p70S6K phosphorylation and intramuscular branched‐chain amino acids in older men following resistance exercise and protein intake

    PubMed Central

    D'Souza, Randall F.; Markworth, James F.; Figueiredo, Vandre C.; Della Gatta, Paul A.; Petersen, Aaron C.; Mitchell, Cameron J.; Cameron‐Smith, David

    2014-01-01

    Abstract Resistance exercise and whey protein supplementation are effective strategies to activate muscle cell anabolic signaling and ultimately promote increases in muscle mass and strength. In the current study, 46 healthy older men aged 60–75 (69.0 ± 0.55 years, 85.9 ± 1.8 kg, 176.8 ± 1.0 cm) performed a single bout of unaccustomed lower body resistance exercise immediately followed by ingestion of a noncaloric placebo beverage or supplement containing 10, 20, 30, or 40 g of whey protein concentrate (WPC). Intramuscular amino acid levels in muscle biopsy samples were measured by Gas Chromatography–Mass Spectrometry (GC‐MS) at baseline (before exercise and WPC supplementation) plus at 2 h and 4 h post exercise. Additionally, the extent of p70S6K phosphorylation at Thr389 in muscle biopsy homogenates was assessed by western blot. Resistance exercise alone reduced intramuscular branch chain amino acid (BCAA; leucine, isoleucine, and valine) content. Supplementation with increasing doses of whey protein prevented this fall in muscle BCAAs during postexercise recovery and larger doses (30 g and 40 g) significantly augmented postexercise muscle BCAA content above that observed following placebo ingestion. Additionally, the fold change in the phosphorylation of p70S6K (Thr389) at 2 h post exercise was correlated with the dose of whey protein consumed (r = 0.51, P < 001) and was found to be significantly correlated with intramuscular leucine content (r = 0.32, P = 0.026). Intramuscular BCAAs, and leucine in particular, appear to be important regulators of anabolic signaling in aged human muscle during postexercise recovery via reversal of exercise‐induced declines in intramuscular BCAAs. PMID:25107987

  9. Modifications in rat testicular morphology and increases in IFN-gamma serum levels by the oral administration of subtoxic doses of mercuric chloride.

    PubMed

    Penna, Salvador; Pocino, Marisol; Marval, Maria Josefina; Lloreta, José; Gallardo, Luis; Vila, Joan

    2009-01-01

    Mercury induces structural and functional damage in several organs, however the effects of subtoxic doses of the metal on the male reproductive system are not well defined. In order to analyze testicular and epididymal morphological alterations and changes in IL-4 or IFN-gamma serum levels, adult male Sprague-Dawley rats received 0.01, 0.05 or 0.1 microg/ml of mercuric chloride (HgCl(2)) in deionized water for 1 to 7 months by oral route. Controls received deionized water alone. Twenty rats, separated in four groups of five animals each, were used per time of exposure. Progressive degenerative lesions consisting of lack of germ cell cohesion and desquamation, arrest at spermatocyte stage and hypospermatogenesis were observed in seminiferous epithelium by light and electron microscopy. Leydig cells showed cytoplasmic vacuolation and nuclear signs of cell death. Loss of peritubular cell aggregation was evidenced in the epididymis. Mercury accumulation was detected in both organs by mass spectroscopy. Rats showed enhanced IFN-gamma serum levels as compared to controls but only reached significance after 7 months of mercury administration. Subtoxic doses of inorganic mercury could lead to reproductive and immunological alterations. The results demonstrate that sublethal concentrations of mercuric chloride are enough to induce morphological and ultrastructural modifications in male reproductive organs. These contribute to functional alterations of spermatogenesis with arrest at spermatocyte stage, hypospermatogenesis and possibly impaired steroidogenesis which together could affect male fertility.

  10. Association between the transcriptional levels of Htr-1a and tryptophan hydroxylase-1 in the hippocampus and the antifatigue effects of leucine on rats with postoperative fatigue.

    PubMed

    Wu, Tiantian; Chen, Jing; Zhu, Jiang; Yu, Zhen

    2014-11-01

    Leucine (Leu), a branched-chain amino acid (BCAA), is widely used in clinical practice following severe burns, gastrointestinal surgery, trauma and sepsis. In the present study, the antifatigue effects of BCAAs on a postoperative fatigue (POF) rat model, induced by 70% intestinal resection, were investigated. Leu (16.5 g/l) was administered intraperitoneally at a dose of 18 ml/kg/day. The fatigue level and antifatigue effects of Leu were evaluated by open-field testing on day 1, 3, 5 and 7 after surgery. In addition, mRNA specimens were extracted and measured using a quantitative polymerase chain reaction method. The open-field test results indicated that Leu exhibited a significant antifatigue effect. The total distance travelled and the number of times the rats passed from the outermost grids of an open-top case were greatly improved in the Leu treatment group when compared with the POF model group. With the exception of the normal group, the mRNA expression levels of Htr-1a exhibited a similar trend in all other groups, reaching a climax on day 3 and 5, while being restored to a normal level on day 7. With regard to the Leu intervention group, the mRNA expression level of Htr-1a decreased significantly on day 3 and 5 following surgery. The mRNA expression levels of tryptophan hydroxylase-1 were unchanged in this short time period; however, the levels were increased gradually in the Leu treatment group. Therefore, Leu exhibited an apparent antifatigue effect on various 5-hydroxytryptamine-associated genes.

  11. Vascular accumulation of the small leucine-rich proteoglycan decorin in CADASIL.

    PubMed

    Lee, Soo Jung; Zhang, Xiaojie; Wang, Michael M

    2014-09-10

    Small penetrating brain artery thickening is a major feature of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). Although affected fibrotic arteries of CADASIL have been shown to accumulate collagen, other components that compose pathological arterial walls remain incompletely characterized. We investigated the expression of decorin (DCN), the first collagen-binding small leucine-rich proteoglycan identified, in CADASIL. DCN was markedly upregulated in pathologically affected leptomeningeal and small penetrating arteries in CADASIL and was notably weaker in normal arteries from control brains. DCN protein was localized principally to the media and adventitia and only occasionally expressed in the intima. Immunoblotting of brain lysates showed a three-fold increase of DCN in CADASIL brains (compared with controls). Messenger RNA encoding DCN was five-fold increased in CADASIL. We conclude that DCN is the first identified proteoglycan to be identified in CADASIL arteries and may accumulate through transcriptional mechanisms. Additional studies are warranted to determine whether DCN localizes broadly to pathological small vessels in other cerebrovascular disorders.

  12. The role of leucine and its metabolites in protein and energy metabolism.

    PubMed

    Duan, Yehui; Li, Fengna; Li, Yinghui; Tang, Yulong; Kong, Xiangfeng; Feng, Zemeng; Anthony, Tracy G; Watford, Malcolm; Hou, Yongqing; Wu, Guoyao; Yin, Yulong

    2016-01-01

    Leucine (Leu) is a nutritionally essential branched-chain amino acid (BCAA) in animal nutrition. It is usually one of the most abundant amino acids in high-quality protein foods. Leu increases protein synthesis through activation of the mammalian target of rapamycin (mTOR) signaling pathway in skeletal muscle, adipose tissue and placental cells. Leu promotes energy metabolism (glucose uptake, mitochondrial biogenesis, and fatty acid oxidation) to provide energy for protein synthesis, while inhibiting protein degradation. Approximately 80 % of Leu is normally used for protein synthesis, while the remainder is converted to α-ketoisocaproate (α-KIC) and β-hydroxy-β-methylbutyrate (HMB) in skeletal muscle. Therefore, it has been hypothesized that some of the functions of Leu are modulated by its metabolites. Both α-KIC and HMB have recently received considerable attention as nutritional supplements used to increase protein synthesis, inhibit protein degradation, and regulate energy homeostasis in a variety of in vitro and in vivo models. Leu and its metabolites hold great promise to enhance the growth and health of animals (including humans, birds and fish).

  13. The role and effects of glucocorticoid-induced leucine zipper in the context of inflammation resolution.

    PubMed

    Vago, Juliana P; Tavares, Luciana P; Garcia, Cristiana C; Lima, Kátia M; Perucci, Luiza O; Vieira, Érica L; Nogueira, Camila R C; Soriani, Frederico M; Martins, Joilson O; Silva, Patrícia M R; Gomes, Karina B; Pinho, Vanessa; Bruscoli, Stefano; Riccardi, Carlo; Beaulieu, Elaine; Morand, Eric F; Teixeira, Mauro M; Sousa, Lirlândia P

    2015-05-15

    Glucocorticoid (GC)-induced leucine zipper (GILZ) has been shown to mediate or mimic several actions of GC. This study assessed the role of GILZ in self-resolving and GC-induced resolution of neutrophilic inflammation induced by LPS in mice. GILZ expression was increased during the resolution phase of LPS-induced pleurisy, especially in macrophages with resolving phenotypes. Pretreating LPS-injected mice with trans-activator of transcription peptide (TAT)-GILZ, a cell-permeable GILZ fusion protein, shortened resolution intervals and improved resolution indices. Therapeutic administration of TAT-GILZ induced inflammation resolution, decreased cytokine levels, and promoted caspase-dependent neutrophil apoptosis. TAT-GILZ also modulated the activation of the survival-controlling proteins ERK1/2, NF-κB and Mcl-1. GILZ deficiency was associated with an early increase of annexin A1 (AnxA1) and did not modify the course of neutrophil influx induced by LPS. Dexamethasone treatment resolved inflammation and induced GILZ expression that was dependent on AnxA1. Dexamethasone-induced resolution was not altered in GILZ(-/-) mice due to compensatory expression and action of AnxA1. Our results show that therapeutic administration of GILZ efficiently induces a proapoptotic program that promotes resolution of neutrophilic inflammation induced by LPS. Alternatively, a lack of endogenous GILZ during the resolution of inflammation is compensated by AnxA1 overexpression.

  14. In Situ Ratiometric Quantitative Tracing of Intracellular Leucine Aminopeptidase Activity via an Activatable Near-Infrared Fluorescent Probe.

    PubMed

    Gu, Kaizhi; Liu, Yajing; Guo, Zhiqian; Lian, Cheng; Yan, Chenxu; Shi, Ping; Tian, He; Zhu, Wei-Hong

    2016-10-03

    Leucine aminopeptidase (LAP), one of the important proteolytic enzymes, is intertwined with the progress of many pathological disorders as a well-defined biomarker. To explore fluorescent aminopeptidase probe for quantitative detection of LAP distribution and dynamic changes, herein we report a LAP-targeting near-infrared (NIR) fluorescent probe (DCM-Leu) for ratiometric quantitative trapping of LAP activity in different kinds of living cells. DCM-Leu is composed of a NIR-emitting fluorophore (DCM) as a reporter and l-leucine as a triggered moiety, which are linked together by an amide bond specific for LAP cleavage. High contrast on the ratiometric NIR fluorescence signal can be achieved in response to LAP activity, thus enabling quantification of endogenous LAP with "build-in calibration" as well as minimal background interference. Its ratiometric NIR signal can be blocked in a dose-dependent manner by bestatin, an LAP inhibitor, indicating that the alteration of endogenous LAP activity results in these obviously fluorescent signal responses. It is worth noting that DCM-Leu features striking characteristics such as a large Stokes shift (∼205 nm), superior selectivity, and strong photostability responding to LAP. Impressively, not only did we successfully exemplify DCM-Leu in situ ratiometric trapping and quantification of endogenous LAP activity in various types of living cells, but also, with the aid of three-dimensional confocal imaging, the intracellular LAP distribution is clearly observed from different perspectives for the first time, owing to the high signal-to-noise of ratiometric NIR fluorescent response. Collectively, these results demonstrate preclinical potential value of DCM-Leu serving as a useful NIR fluorescent probe for early detection of LAP-associated disease and screening inhibitor.

  15. Simulations of potentials of mean force for separating a leucine zipper dimer and the basic region of a basic region leucine zipper dimer.

    PubMed

    Cukier, Robert I

    2014-09-04

    Basic region leucine zipper (bZIP) transcription factors involved in DNA recognition are dimeric proteins. The monomers consist of two subdomains, a leucine zipper sequence responsible for dimerization and a highly basic DNA recognition sequence. Leucine zippers are strongly dimerized, and in a bZIP, the basic region can, in the absence of DNA, undergo extensive relative monomer-to-monomer fluctuations. In this work, LZ and bZIP potentials of mean force (PMFs), which provide free energies along reaction coordinates, are simulated with a distance replica exchange method. The method uses restraint potentials to provide sampling along a reaction coordinate and enhances configuration space exploration by exchanging information between neighboring restraint potential configurations. Restraint potentials that are constructed from sums over a number of atom distances are employed. Their use requires a modification of the Weighted Histogram Analysis Method (WHAM) procedure to combine and unbias the data from the different restraint-potential-biased window densities to provide a PMF. These methods are first used to obtain a PMF for separating a leucine zipper (GCN4-p1) of the yeast transcriptional activator GCN4. The PMF indicates a very strong binding free energy that only weakens when the monomers are separated by about 12 Å, which is about 6 Å beyond their bound, dimer equilibrium distance. PMFs are also obtained for separating the basic subdomain monomer parts of the GCN4 bZIP transcriptional factor, in the absence of DNA. In a monomer separation range spanning the open, crystal-based structure to closer configurations, the basic subdomain PMF is quite flat, implying essentially thermal sampling in this distance range. A PMF generated starting from a "collapsed" state, taken from a previous simulation ( J. Phys. Chem. B 2012 , 116 , 6071 ), where collapsed refers to the feature that the basic subdomain monomers are also effectively dimerized, shows that this state is

  16. Salt bridges destabilize a leucine zipper designed for maximized ion pairing between helices.

    PubMed

    Phelan, Paul; Gorfe, Alemayehu A; Jelesarov, Ilian; Marti, Daniel N; Warwicker, James; Bosshard, Hans Rudolf

    2002-03-05

    Interhelical salt bridges are common in leucine zippers and are thought to stabilize the coiled coil conformation. Here we present a detailed thermodynamic investigation of the designed, disulfide-linked leucine zipper AB(SS) whose high-resolution NMR structure shows six interhelical ion pairs between heptad positions g of one helix and e' of the other helix but no ion pairing within single helices. The average pK(a) value of the Glu side chain carboxyl groups of AB(SS) is slightly higher than the pK(a) of a freely accessible Glu in an unfolded peptide [Marti, D. N., Jelesarov, I., and Bosshard, H. R. (2000) Biochemistry 39, 12804-12818]. This indicates that the salt bridges are destabilizing, a prediction we now have confirmed by determining the pH +/- stability profile of AB(SS). Circular dichroism-monitored unfolding by urea and by heating and differential scanning calorimetry show that the coiled coil conformation is approximately 5 kJ/mol more stable when salt bridges are broken by protonation of the carboxyl side chains. Using guanidinium chloride as the denaturant, the increase in the free energy of unfolding on protonation of the carboxyl side chains is larger, approximately 17 kJ/mol. The discrepancy between urea and guanidinium chloride unfolding can be ascribed to the ionic nature of guanidinium chloride, which screens charge-charge interactions. This work demonstrates the difficulty of predicting the energetic contribution of salt bridges from structural data alone even in a case where the ion pairs are seen in high-resolution NMR structures. The reason is that the contribution to stability results from a fine balance between energetically favorable Coulombic attractions and unfavorable desolvation of charges and conformational constraints of the residues involved in ion pairing. The apparent discrepancy between the results presented here and mutational studies indicating stabilization by salt bridges is discussed and resolved. An explanation is

  17. Fusarium moniliforme extract fed before a single dose of diethylnitrosamine increases the numbers of placental glutathione S-transferase positive hepatocytes in rat liver

    SciTech Connect

    Lebepe, S.; Hendrich, S. )

    1991-03-11

    The carcinogenic potential of an alcohol:water (1:1) extract of Fusarium moniliforme (FUSX), containing 20 ppm fumonisin B{sub 1} was assayed. Groups of six 5-week-old female F344/N rats were fed a semipurified diet, with and without FUSX. A dose of initiating agent, diethylnitrosamine, was given orally. Placental glutathione S-transferase-positive (PGST(+)) hepatocytes were detected by immunohistochemistry and counted on 5 frozen hepatic sections/rat, as an endpoint to assess early stages of carcinogenesis. FUSX had significant co-initiating activity. Fusarium moniliforme infection of feed has been shown to promote hepatocarcinogenesis, and may pose a cocarcinogenic risk even during short-term, low-level exposure.

  18. Separation by high-performance liquid chromatography of (3R)- and (3S)-beta-leucine as diastereomeric derivatives.

    PubMed

    Aberhart, D J; Cotting, J A; Lin, H J

    1985-11-15

    For studies on the coenzyme B12-dependent enzyme, leucine-2,3-aminomutase, (3R)- and (3S)-beta-leucines were synthesized. The 10-camphorsulfonamide p-nitrobenzyl esters could be resolved by normal-phase HPLC. A much better separation was obtained by reversed-phase HPLC of the diastereomeric derivatives obtained by treatment of -leucine with Marfey's reagent (N2-(5-fluoro-2,4-dinitrophenyl)-L-alaninamide).

  19. Effect of L-Tryptophan and L-Leucine on Gut Hormone Secretion, Appetite Feelings and Gastric Emptying Rates in Lean and Non-Diabetic Obese Participants: A Randomized, Double-Blind, Parallel-Group Trial

    PubMed Central

    Meyer-Gerspach, Anne Christin; Häfliger, Simon; Meili, Julian; Doody, Alison; Rehfeld, Jens F; Drewe, Jürgen; Beglinger, Christoph; Wölnerhanssen, Bettina

    2016-01-01

    Background/Objectives Gut hormones such as cholecystokinin (CCK) and glucagon-like peptide-1 (GLP-1) play a role as satiation factors. Strategies to enhance satiation peptide secretion could provide a therapeutic approach for obesity. Carbohydrates and lipids have been extensively investigated in relation to peptide release. In contrast, the role of proteins or amino acids is less clear. Our aim was to compare the effects of the amino acids L-tryptophan (L-trp) and L-leucine (L-leu) separately on gastric emptying and gut peptide secretion. Participants/Methods The study was conducted as a randomized (balanced), double-blind, parallel-group trial. A total of 10 lean and 10 non-diabetic obese participants were included. Participants received intragastric loads of