Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0432222 (SEM)
 47,337 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The primed, continuous intravenous infusion of amino acids labeled with 13C together with measurement of isotopic enrichment in plasma is commonly used to study amino acid metabolism. However, a less invasive, oral infusion that also produces an isotopic steady state in CO2 and urine would be useful, particularly for pediatric studies. We measured the 13C enrichments of expired CO2, plasma and urine free phenylalanine and lysine and estimated flux and oxidation rates in adult humans (n = 12) who received a 4-h oral, primed, equal dose infusion of either L-[1-13C]phenylalanine, L-[1-13C]lysine (D-lysine = 1.6%) or L-[1-13C]lysine (D-lysine </= 0.2%). Steady fed state conditions were established by feeding subjects eight hourly meals beginning 4 h before the start of the oral infusion protocol. Isotopic plateau in CO2, plasma and urine was achieved within 120 min of phenylalanine or lysine infusion. At isotopic plateau, the mean ratio of plasma to urine enrichment was 1.0 +/- 0.04 (SEM), 0. 39 +/- 0.03 and 0.97 +/- 0.02 for L-[1-13C]phenylalanine, L-[1-13C]lysine (D-lysine = 1.6%) and L-[1-13C]lysine (D-lysine</= 0. 2%), respectively. There was good agreement between isotopic enrichment in plasma and urine for L-[1-13C]phenylalanine and L-[1-13C]lysine (D-lysine </= 0.2%). However, use of L-[1-13C]lysine (D-lysine = 1.6%) resulted in significantly higher enrichment in urine, probably due to renal tubular discrimination of D-lysine. Mean flux rates for phenylalanine and lysine were consistent with the literature. Thus, the oral, primed, equal dose infusion produces the isotopic steady-state condition required for amino acid flux and oxidation determination within an 8-h period.
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PMID:Development of a minimally invasive protocol for the determination of phenylalanine and lysine kinetics in humans during the fed state. 980 42

The activation of the neutrophil respiratory burst is a two-step process involving an initial 'priming' phase followed by a 'triggering' event. The biochemical mechanisms which underlie these events are yet to be fully elucidated, but the evidence suggests a crucial role for stimulus-induced tyrosine phosphorylation. The enhanced tyrosine phosphorylation observed upon triggering primed cells may reflect an increase in tyrosine kinase activity or a reduction in the levels of the opposing phosphotyrosine phosphatases (PTPases). We have investigated the latter by examining the possibility that lipopolysaccharide (LPS)-induced priming of the neutrophil respiratory burst involves the suppression of cellular PTPase activity. Purified human neutrophils were incubated for 60 min with and without LPS. Priming of the respiratory burst was confirmed by fMet-Leu-Phe-induced cytochrome c reduction. The level of PTPase activity was assessed by dephosphorylation of [32P]RR-src peptide as substrate. Pretreatment of human neutrophils with 200 ng/ml LPS induced a 2.9 +/- 0.3 (mean +/- SEM, n = 3, P = 0.022) fold increase in the fMet-Leu-Phe-triggered respiratory burst. In the same cells, LPS did not induce a significant change in the total cellular PTPase activity (1.02 +/- 0.02-fold, mean +/- SEM, n = 3, P = 0.63). Similarly, stimulation of neutrophils with fMet-Leu-Phe or phorbol myristate acetate did not significantly affect the cellular PTPase activity (P = 0.94 and 0.68, respectively). Our results suggest that suppression of PTPase activity is not the mechanism underlying the priming and/or triggering of the neutrophil respiratory burst.
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PMID:Lipopolysaccharide-induced priming of the human neutrophil is not associated with a change in phosphotyrosine phosphatase activity. 1039 19

We measured N-formyl-methionyl-leucyl-phenylalanine-induced reactive oxygen species production by neutrophils from three patients with acute promyelocytic leukemia during treatment with all-trans retinoic acid using a luminol-enhanced chemiluminescence assay. The maximum level of reactive oxygen species production during all-trans retinoic acid treatment was 58.8 +/- 2.3 x 10(4) (mean +/- SEM) counted photons per seconds (cps), which was significantly higher (p<0.0001) than that of neutrophils from health volunteers (13.3 +/- 2.3 x 10(4) cps).
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PMID:Reactive oxygen species production of neutrophils in patients with acute promyelocytic leukemia during treatment with all-trans retinoic acid. 1057 82

To evaluate whether increased levels of reactive oxygen species (ROS) are involved in the pathogenesis of essential hypertension (EH) and non-insulin-dependent diabetes mellitus (NIDDM), both resting and stimulated levels of intracellular ROS were measured in lymphocytes from patients with EH (n = 10), NIDDM (n = 16) and age-matched healthy individuals (control subjects, n = 19). ROS was monitored with the dye, dihydrorhodamine-123 (DHR; 1 micromol/L) in the presence or absence of superoxide dismutase (superoxide scavenger), sodium azide (singlet oxygen/hydrogen peroxide scavenger), genistein (tyrosine kinase inhibitor), or bisindolylmaleimide (protein kinase C inhibitor). Simultaneous monitoring of cytosolic [Ca2+]i was done with fura-2. Resting ROS levels were significantly higher in NIDDM (4.71+/-0.25 nmol/10(6) cells; mean +/- SEM, P<.05) compared with EH (4.03+/-0.22 nmol/10(6) cells) or controls (4.05+/-0.15 nmol/10(6) cells). The formyl-Met-Leu-Phenylalanine-(fMLP)-induced ROS generation was significantly higher in NIDDM (21.92+/-2.23 nmol/10(6) cells; P<.05) compared with EH (14.58+/-1.90 nmol/10(6) cells) or control (16.06+/-1.22 nmol/10(6) cells). The fMLP-induced ROS increase was significantly reduced in the presence of sodium azide in all groups (P<.01) but was largely unaffected in the presence of SOD. Genistein and bisindolylmaleimide significantly inhibited the fMLP-induced ROS in all groups. The fMLP-induced [Ca2+]i increase was significantly higher in NIDDM (71+/-12 nmol/L, P <.01) compared with EH (42+/-4 nmol/L) and control subjects (35+/-3 nmol/L). Phytohemagglutinin was more effective in increasing [Ca2+]i than ROS. It is concluded that ROS may play a role in the metabolic syndrome of NIDDM but not in EH.
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PMID:Reactive oxygen species in essential hypertension and non-insulin-dependent diabetes mellitus. 1061 78

The purpose of this study was to establish, using a flooding dose of L-[ring 2, 6-(3)H] phenylalanine, whether feeding pigs diets that induce high endogenous gut nitrogen losses (ENL) also increases protein synthesis rates in (PSR) the visceral organs. Twelve 18-kg Yorkshire barrows with catheters in the right and left jugular veins were fed for 3 wk either casein-cornstarch- (CC) or barley-canola meal- (BCM) based diets formulated to a similar digestible energy /crude protein ratio and designed to induce either low or high ENL, respectively. Pigs were infused with 10 mL/kg body weight of a 150 mmol. L(-1) phenylalanine solution containing 230 MBq. L(-1) labeled phenylalanine for 12 min and killed 20 min later. Plasma phenylalanine specific radioactivity (SRA) rose to a plateau value within 3 min of starting the infusion and did not change (P > 0.10) thereafter. Fractional rates of protein synthesis (K(s), %/d) based on SRA in plasma- or intracellular-free phenylalanine did not differ (P > 0.10) in all tissues except pancreas (P < 0.05). Diet affected K(s )in liver (P < 0.01) and colon (P < 0.05) but not in pancreas, duodenum, jejunum and cecum. Based on plasma-free phenylalanine SRA, liver K(s)were 85.4 +/- 11.0 vs. 60.5 +/- 5.2 (mean +/- SEM) in CC- and BCM-fed pigs, respectively; these values were 82.3 +/- 4.7 vs. 98.2 +/- 5.8 in the colon. The absolute amount of protein synthesis (g.d(-1)) was higher in the liver (P < 0.05) and pancreas (P < 0. 05) of the CC pigs compared to BCM pigs. No dietary effects were observed in all other organs (P > 0.10). The present results suggest that feeding growing pigs a BCM diet that induces high ENL does not affect PSR in the small intestine of growing pigs from which >50% of ENL originates.
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PMID:Endogenous gut nitrogen losses in growing pigs are not caused by increased protein synthesis rates in the small intestine. 1070 86

We examined the effects of dietary fat type (10% of either soybean oil, S, or beef tallow, T)(3) and xylanase supplementation (-, without; +, with 1 g of Avizyme 1300 per kg diet) in rye- based diets (56%) on tissue protein synthesis in male broilers. Birds were injected with a large flooding dose of a phenylalanine solution (150 mmol/L, 38 atom percentage excess [(15) N] phenylalanine) and tissues were obtained after a 10-min incorporation period. [(15) N]-enrichment in tissue free phenylalanine and tissue protein bound phenylalanine were measured by gas-chromatography mass-spectrometry and by gas-chromatography combustion isotope ratio mass-spectrometry, respectively in order to calculate tissue specific fractional rates of protein synthesis (k(s)). The k(s) (%/d) in (S-), (S+), (T-) and (T+)-fed birds were 56, 64, 84 and 61 (SEM = 3.7) in duodenum, 51, 52, 75 and 58 in jejunum (SEM = 3.1), 66, 67, 105 and 68 (SEM =7.0) in jejunal mucosa cells, 53, 56, 68 and 50 (SEM = 3.7) in ileum and 52, 45, 118 and 39 (SEM = 20.2) in pancreas, respectively. Significant fat, enzyme or interaction effects in these tissues were mainly caused by the elevated k(s) in (T-)-fed birds which was closely associated with intestinal viscosity. We conclude that the effect of soluble nonstarch polysaccharides (NSP) and of NSP-hydrolyzing enzymes may be explained partially by modification in tissue protein synthesis of the intestinal tract.
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PMID:Replacement of soybean oil with tallow in rye-based diets without xylanase increases protein synthesis in small intestine of broilers. 1073 37

Heat and alkali treatment of food may increase the concentrations of protein-bound D-amino acids and cross-links such as lysinoalanine (LAL). To examine how protein treatment affects digestibility, purified test meals [total protein 150 g/kg dry matter (DM), 0.44 MJ/(kg BW(0.75). d)] were prepared, containing (g/kg DM) casein, 75; beta-lactoglobulin, 50; or wheat protein, 40. Each was (15)N-labeled. Test proteins were used either in their native form or after treatment for 6 or 24 h at 65 degrees C, pH 10.5-11.5. Each meal was fed to nine adult miniature pigs (twofold complete cross-classification). Chyme was collected continuously over 33 h postprandially via T-fistulas in the terminal ileum, and digestibilities of test proteins and individual L- and D-amino acids were calculated on the basis of recovery of (15)N and the respective amino acids in the chyme. Treatment of casein, beta-lactoglobulin or wheat protein for 24 h increased levels of D-amino acid residues. L-Asparagine and aspartate (L-Asx) were particularly susceptible; 14. 7 +/- 0.4, 11.7 +/- 0.2 and 11.0 +/- 0.9%, respectively, underwent racemization. LAL levels increased in parallel; 11.7 +/- 0.3, 13.6 +/- 0 and 14.8 +/- 0.0%, respectively, of total lysine was converted to LAL. At the same time, prececal protein digestibility was decreased by 13.4 +/- 2.3, 15.3 +/- 1.4 and 17.8 +/- 1.2% units, respectively (P < 0.05; mean +/- SEM, n = 9). Digestibility of individual L-amino acids decreased by 10-15%, but L-amino acids prone to peptic cleavage, such as L-phenylalanine and L-tyrosine, were not affected. Digestibilities of D-amino acids and LAL were approximately 35%. It seems that mainly D-amino acids, and to a lesser extent LAL, were responsible for lower digestibility by interfering with peptic cleavage.
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PMID:Protein-bound D-amino acids, and to a lesser extent lysinoalanine, decrease true ileal protein digestibility in minipigs as determined with (15)N-labeling. 1091 20

In order to elucidate the role of mitogen-activated protein kinase kinase (MEK-1/2) in 5-lipoxygenase (5-LO) activation we studied the N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced 5-LO translocation in human blood neutrophils (PMNs). In non-primed, Ca(2+)-repleted PMNs, fMLP consistently stimulated MEK-1/2 phosphorylation, but induced 5-LO translocation and product formation (430+/-128 pmol; SEM, n=13) only in 13 of 18 PMN preparations from different healthy donors. In fMLP-responsive cells, the MEK-1/2 inhibitor PD098059 (50 microM) attenuated MEK phosphorylation and abolished 5-LO activation at the translocation step. The fMLP-mediated 5-LO product formation was also sensitive to MEK inhibition by U0126 and to p38 inhibition by SB203580. But in contrast to PD098059, U0126 at 10 microM and SB203580 at 20-50 microM impaired 5-LO activity in the cell-free assay setting, suggesting direct actions of higher concentrations of U0126 and SB203580 on 5-LO apart from MEK and p38 inhibition, respectively. These data show that fMLP initiates 5-LO product formation in non-primed, Ca(2+)-repleted human blood PMNs from healthy donors, and that MEK signaling is pivotal, but not sufficient for 5-LO activation in response to the receptor agonist fMLP.
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PMID:MEK-1/2 inhibition prevents 5-lipoxygenase translocation in N-formylpeptide-challenged human neutrophils. 1109 Nov 39

We used RT-PCR to clone monoamine transporters from Macaca mulatta, Macaca fasicularis and Saimiri sciureus (dopamine transporter; DAT) and Macaca mulatta (norepinephrine transporter; NET and serotonin transporter; SERT). Monkey DAT, NET and SERT proteins were >98% homologous to human and, when expressed in HEK-293 cells, displayed drug affinities and uptake kinetics that were highly correlated with monkey brain or human monoamine transporters. In contrast to reports of other species, we discovered double (leucine for phenylalanine 143 and arginine for glutamine 509; Variant I) and single (proline for leucine 355; Variant II) amino acid variants of DAT. Variant I displayed dopamine transport kinetics and binding affinities for various DAT blockers (including cocaine) versus [3H] CFT (WIN 35, 428) that were identical to wild-type DAT (n=7 drugs; r(2)=0.991). However, we detected a six-fold difference in the affinity of cocaine versus [3H] cocaine between Variant I (IC(50): 488+/-102 nM, SEM, n=3) and wild-type DAT (IC(50): 79+/-8.2 nM, n=3, P<0.05). Variant II was localized intracellularly in HEK-293 cells, as detected by confocal microscopy, and had very low levels of binding and dopamine transport. Also discovered was a novel exon 5 splice variant of NET that displayed very low levels of transport and did not bind cocaine. With NetPhos analysis, we detected a number of highly conserved putative phosphorylation sites on extracellular as well as intracellular loops of the DAT, NET, and SERT, which may be functional for internalized transporters. The homology and functional similarity of human and monkey monoamine transporters further support the value of primates in investigating the role of monoamine transporters in substance abuse mechanisms, neuropsychiatric disorders and development of diagnostic and therapeutic agents.
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PMID:Cloning of dopamine, norepinephrine and serotonin transporters from monkey brain: relevance to cocaine sensitivity. 1122 67

In vitro matured (IVM) and fertilized (IVF) putative Day 1 zygotes (Day 0 = IVF) were allocated randomly to culture in formulations based on Synthetic Oviduct Fluid (SOF) medium and identified on the basis of their contrasting principal supplements, which were 10% v/v steer serum (SS; n = 558) or 4 mg/mL crystalline BSA (SBSA; n = 531) or 3 mg/mL polyvinyl alcohol (SPVA; n = 607) in 9 replicates. SBSA and SPVA also contained 10 microg/mL non-essential amino acids, while the former was further supplemented with 20 microL/mL essential amino acids and the latter with 0.5 mmol/L sodium citrate and 5 ng/mL epidermal growth factor. Zygotes were cultured in 20 microL drops (4 zygotes per drop) until Day 8 in an atmosphere of 5% CO2, 5% O2 and 90% N2 at 39 degrees C and droplets were renewed every 48 hours. The incidence of zygote cleavage was lower (P < 0.05) in SS (mean +/- SEM = 61 +/- 3%) than in SBSA (76 +/- 3%) but not in SPVA (72 +/- 4%) up to Day 3. The SPVA generated a lower yield of blastocysts on Day 7 (12 +/- 2%; P < 0.001) and by Day 8 (21 +/- 4%; P < 0.01) than did SS (33 +/- 3%; 40 +/- 3%) and SBSA (30 +/- 3%; 37 +/- 4%). Cell numbers (n) and diameters (d) of blastocysts on Day 8 were greater (P < 0.001; Replicates 1 to 5) in embryos from SBSA (n, 156 +/- 9; d, 203 +/- 4 microm) than in those from SS (n, 81 +/- 4; d, 177 +/- 3 microm) and SPVA (n, 76 +/- 5; d, 167 +/- 3 microm). Embryos produced in SS incorporated less 3H-phenylalanine into PCA-precipitable protein (replicates 6 to 9; log10 dpm = 3.03 +/- 0.04) than did embryos cultured in SBSA (3.21 +/- 0.03; P < 0.001) or in SPVA (3.14 +/- 0.03; NS). In conclusion, blastocyst yield was poor in SPVA, but the embryos had metabolic activities similar to those of embryos produced in SBSA. Blastocyst yields from SS were not compromised but their capacity for de novo protein synthesis was reduced significantly.
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PMID:Development and de novo protein synthetic activity of bovine embryos produced in vitro in different culture systems. 1123 85


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