RESUMO
NEW FINDINGS: What is the central question of this study? Low dose carbon monoxide (CO) inhalation plays a role in regulating proteins involved in glucose metabolism; does low dose CO improve glucose and insulin responses to an oral glucose tolerance test in overweight adults? What is the main finding and its importance? Five days of intermittent CO inhalation does not alter the glucose or insulin responses to ingestion of a glucose bolus in overweight adults. Low dose CO is utilized in various physiological assessment procedures; these findings allow researchers and clinicians to utilize these procedures without concern of altering glucose metabolism. ABSTRACT: Low dose carbon monoxide (CO) inhalation upregulates several proteins important for glucose metabolism. Such changes could be clinically significant and may be relevant to those who use CO as a research tool. We hypothesized that low dose CO inhalation would improve glucose and insulin responses to an oral glucose bolus in overweight humans. Eleven young adults (5 men, 6 women; body mass index: 25-35 kg m-2 ) were included in this randomized, placebo-controlled, single-blinded crossover study. Following screening, participants completed two 7-day protocols with a 4-week washout. Twenty-four hours prior to and following five consecutive days of either once daily CO (men: 1.2 ml (kg body mass)-1 ; women: 1.0 ml (kg body mass)-1 ) or placebo (room air) inhalation, participants underwent oral glucose tolerance tests (OGTT). For key outcome variables, there were no significant main effects or interactions across condition or time point (mean ± SD), including fasting glucose (mg dl-1 : pre-placebo: 85.2 ± 10.1; post-placebo: 82.9 ± 10.6; pre-CO: 83.6 ± 7.7; post-CO: 84.0 ± 9.0), 2 h post glucose (mg dl-1 : pre-placebo: 100.9 ± 20.0; post-placebo: 98.7 ± 13.1; pre-CO: 94.2 ± 23.2; post-CO: 94.4 ± 14.9), or the Matsuda index (pre-placebo: 16.1 ± 11.5; post-placebo: 20.3 ± 24.7; pre-CO: 15.6 ± 15.3; post-CO: 17.5 ± 16.8). In conclusion, 5 days of low dose CO administration did not influence glucose and insulin responses to an OGTT in overweight adults. Low dose CO inhalation is utilized in a variety of physiological assessment procedures; these findings allow researchers to utilize these procedures without concern of altering glucose metabolism.
Assuntos
Monóxido de Carbono/administração & dosagem , Glucose/metabolismo , Sobrepeso/tratamento farmacológico , Adulto , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Índice de Massa Corporal , Estudos Cross-Over , Jejum/metabolismo , Feminino , Teste de Tolerância a Glucose/métodos , Humanos , Insulina/metabolismo , Resistência à Insulina/fisiologia , Masculino , Sobrepeso/metabolismo , Método Simples-Cego , Adulto JovemRESUMO
This year, 2019, marks the centennial of embryologist E. E. Just's discovery of what is known as the fast block to polyspermy. Just's observation of the subtle changes that occur at the egg's surface during fertilization (and in experimental parthenogenesis) led him to postulate that the egg, and indeed every cell, possesses a property he called independent irritability, which represents the cell's ability to respond in a physiologically-relevant way to a variety of signals or triggers. In this paper, I argue that Just's concept of independent irritability informed his contemporary Johannes Holtfreter as Holtfreter attempted to explain the phenomena of embryonic induction and competence and that Holtfreter, in turn, influenced Marc Kirschner and John Gerhart in their formulation of the theory of facilitated variation. Just's influence is especially evident in Gerhart and Kirschner's presentations of what they call weak linkage-a property of living systems that allows core processes and components to be mixed and matched in different ways to generate novel traits. Unfortunately, the connection between Holtfreter's work and Just's has remained hidden. This paper gives examples of phenomena that exhibit weak linkage, and it lays out the case that Just's concept of independent irritability, through Holtfreter, Gerhart, and Kirschner, has broadly infiltrated modern cell and developmental biology.
Assuntos
Biologia Celular/história , Biologia do Desenvolvimento/história , Poliploidia , Interações Espermatozoide-Óvulo/fisiologia , Animais , Indução Embrionária/fisiologia , Feminino , História do Século XX , História do Século XXI , Masculino , Oócitos/fisiologia , Espermatozoides/fisiologiaRESUMO
OBJECTIVE: To assess prospectively the effectiveness of lacosamide (LCM) added to levetiracetam (LEV) after down-titration of a concomitant sodium channel blocker (SCB) among patients with focal epilepsy not adequately controlled on LEV and SCB. METHODS: In this open-label trial, LCM was initiated at 100 mg/day and up-titrated to 200-600 mg/day over 9 weeks; SCB down-titration started when LCM dose reached 200 mg/day. Patients remained on stable LCM/LEV doses for 12 weeks' maintenance (21-week treatment period). The primary outcome was retention rate on LCM. RESULTS: Due to recruitment challenges, fewer than the planned 300 patients participated in the trial, resulting in the trial being underpowered. Overall, 120 patients (mean age 39.7 years) started and 93 completed the trial. The most frequently used SCBs were lamotrigine (39.2%), carbamazepine (30.8%) and oxcarbazepine (27.5%). Eighty-four patients adhered to protocol and discontinued their SCB after cross-titration, but there was insufficient evidence for 36 patients. Retention rate was 73.3% (88/120) for all patients and 83.3% (70/84) for those with evidence of SCB discontinuation. Seizure freedom for patients completing maintenance was 14.0% (13/93). Discontinuation due to adverse events (6.7%) and lack of efficacy (3.3%) occurred primarily during cross-titration. Most frequently reported adverse events during treatment were dizziness (23.3%), headache (15.0%) and fatigue (8.3%). CONCLUSIONS: In patients with uncontrolled seizures on LEV/SCB, the LCM/LEV combination appeared to be effective and well tolerated. A cross-titration schedule-flexible LCM up-titration, concomitant SCB down-titration and stable background LEV-could present a feasible and practical approach to initiating LCM while minimizing pharmacodynamic interactions with a SCB.
Assuntos
Acetamidas/uso terapêutico , Anticonvulsivantes/uso terapêutico , Epilepsias Parciais/tratamento farmacológico , Piracetam/análogos & derivados , Bloqueadores dos Canais de Sódio/uso terapêutico , Acetamidas/efeitos adversos , Adulto , Anticonvulsivantes/administração & dosagem , Anticonvulsivantes/efeitos adversos , Esquema de Medicação , Quimioterapia Combinada , Feminino , Humanos , Lacosamida , Levetiracetam , Masculino , Pessoa de Meia-Idade , Piracetam/administração & dosagem , Piracetam/efeitos adversos , Piracetam/uso terapêutico , Bloqueadores dos Canais de Sódio/administração & dosagem , Bloqueadores dos Canais de Sódio/efeitos adversosRESUMO
Our purpose was to determine if using an individual's power-specific gross efficiency improves the accuracy of estimating energy expenditure from cycling power. 30 subjects performed a graded cycling test to develop 4 gross efficiencies: individual power-specific gross efficiencies, a group mean power-specific gross efficiency, individual fixed gross efficiencies, and a group mean fixed gross efficiency. Energy expenditure was estimated from power using these different gross efficiencies and compared to measured energy expenditure during moderate- and hard-intensity constant-power and 2 variable-power cycling bouts. Estimated energy expenditures using individual or group mean power-specific gross efficiencies were not different from measured energy expenditure across all cycling bouts (p>0.05). To examine the intra-individual variability of the estimates, absolute difference scores (absolute value of estimated minus measured energy expenditure) were compared, where values closer to zero represent more accurate individual estimates. The absolute difference score using individual power-specific gross efficiencies was significantly lower compared to the other gross efficiencies across all cycling bouts (p<0.01). Significant and strong correlations (r≥0.97, p<0.001) were found across all cycling bouts between estimated and measured energy expenditures using individual power-specific gross efficiencies. In conclusion, using an individual's power-specific gross efficiency significantly improves their energy expenditure estimate across different power outputs.
Assuntos
Ciclismo/fisiologia , Metabolismo Energético , Teste de Esforço , Adulto , Feminino , Frequência Cardíaca , Humanos , Ácido Láctico/sangue , Masculino , Consumo de Oxigênio , Adulto JovemRESUMO
Carbon monoxide (CO) rebreathing procedures are used to assess hemoglobin mass (Hbmass) but recent evidence suggests that CO is a signaling molecule that may alter physiological functions. We examined the effects of 10 days of intermittent, low-dose CO inhalation on Hbmass, aerobic performance predictors, and peak-power exercise tolerance. 18 recreationally-active men were randomized to either CO or placebo inhalation groups in a single-blind, pre-post parallel-groups trial. Primary outcomes were assessed before and after an intervention period during which subjects inhaled a bolus of 1.2 ml kg(-1) CO or placebo (room air) for 30 s, once per day on 10 days over a 12-day period. Cycling tests were performed >16 h following CO inhalation to exclude acute effects of CO exposure. CO inhalation elevated carboxyhemoglobin by 4.4±0.4% (mean±SD) following each exposure. Compared to placebo, chronic CO inhalation did not significantly alter Hbmass (p=0.99), peak oxygen uptake (p=0.59), peak power output (p=0.10), submaximal oxygen uptake (p=0.91), submaximal RER (p=0.22), lactate threshold (p=0.65), or peak-power exercise tolerance (p=0.60). In conclusion, our data support the ability to perform repeated measurements of Hbmass using CO rebreathing over a 12-day period without altering physiological responses.
Assuntos
Desempenho Atlético/fisiologia , Monóxido de Carbono/administração & dosagem , Tolerância ao Exercício/fisiologia , Hemoglobinas/metabolismo , Administração por Inalação , Adulto , Ciclismo/fisiologia , Carboxihemoglobina/metabolismo , Exercício Físico/fisiologia , Teste de Esforço , Humanos , Masculino , Consumo de Oxigênio/fisiologia , Método Simples-Cego , Adulto JovemRESUMO
Carbon monoxide, a gas known most widely for its toxic effects at high doses, is receiving increased attention for its role as a physiological signaling molecule and potential therapeutic agent when administered in low doses. We sought to quantify any changes to oxygen consumption and energy expenditure during submaximal exercise after low-dose CO inhalation. 9 active individuals completed 4 graded submaximal exercise tests, with each test occurring during a separate visit. For their first exercise test, subjects inhaled CO or room air (1.2 mL·kg(-1) body mass) in a randomized, subject-blind fashion. A second test was repeated 24 h later when the inhaled gas should have cleared the system. Subjects repeated study procedures with the alternate dose after a washout period of at least 2 days. Low-dose CO administration did not affect oxygen consumption or energy expenditure during submaximal exercise immediately or 24 h following its administration. Increases in heart rate, blood [lactate], and perceived exertion were observed following acute CO inhalation but these effects were absent after 24 h. The results of this study suggest that low-dose CO administration does not influence the energetics of submaximal exercise, but it acutely increases the relative intensity associated with absolute workloads below the lactate threshold.
Assuntos
Monóxido de Carbono/administração & dosagem , Metabolismo Energético/fisiologia , Exercício Físico/fisiologia , Consumo de Oxigênio/fisiologia , Administração por Inalação , Adulto , Monóxido de Carbono/sangue , Carboxihemoglobina/metabolismo , Metabolismo Energético/efeitos dos fármacos , Teste de Esforço , Frequência Cardíaca , Hemoglobinometria , Humanos , Masculino , Consumo de Oxigênio/efeitos dos fármacos , Percepção/fisiologia , Esforço Físico/fisiologia , Método Simples-CegoRESUMO
Renowned biologist Ernest Everett Just (1883-1941) was an outspoken advocate for the classical embryologist's view of the cell; he believed that all the parts of the cell, but especially the cytoplasm, have important roles to play in the process of development, whereby a one-celled zygote becomes a many-celled animal. In opposition to geneticist Thomas Hunt Morgan, Just formulated a hypothesis for how the cell works in development, one that gave a more dominant role to cytoplasmic (instead of nuclear) factors. This paper argues that, in creating his hypothesis, Just applied insights from the African American intellectual community in which he was immersed, much as Charles Darwin applied insights from British political economist Thomas R. Malthus in formulating his theory of evolution by natural selection. This in no way diminishes the scientific validity of Just's (or Darwin's) hypothesis. Rather, it highlights Just's creativity and, as such, points to the importance of having diversity in science.
RESUMO
Recently, we showed that the fused chorismate-utilizing enzyme from the antibiotic-producing soil bacterium Streptomyces venezuelae is an anthranilate synthase (designated SvAS), not a 2-amino-2-deoxyisochorismate (ADIC) synthase, as was predicted based on its amino acid sequence similarity to the phenazine biosynthetic enzyme PhzE (an ADIC synthase). Here, we report the characterization of SvAS using steady-state kinetics, gel filtration chromatography, and laser light scattering. The recombinant His-tagged enzyme has Michaelis constants Km with respect to substrates chorismate and glutamine of 8.2 ± 0.2 µM and 0.84 ± 0.05 mM, respectively, and a catalytic rate constant k cat of 0.57 ± 0.02 s(-1) at 30 °C. Unlike most other anthranilate synthases, SvAS does not utilize ammonia as a substrate. The enzyme is competitively but non-cooperatively inhibited by tryptophan (K i = 11.1 ± 0.1 µM) and is active as a monomer. The finding that SvAS is a monomer jibes with the variety of association modes that have been observed for anthranilate synthases from different microorganisms, and it identifies the enzyme's minimal functional unit as a single TrpE-TrpG pair.
Assuntos
Antranilato Sintase/química , Catálise , Streptomyces/enzimologia , Sequência de Aminoácidos/genética , Antranilato Sintase/genética , Cinética , Estrutura Terciária de Proteína , Especificidade por Substrato , TriptofanoRESUMO
Ernest Everett Just (1883-1941) was an African American embryologist of international standing whose research interests lay in the area of fertilization and early development in marine invertebrates. Perhaps best known for his discovery of the dynamical and structural blocks to polyspermy that sweep over the egg upon fertilization, E. E. Just also was the first to associate cell surface changes with stages of embryonic development. He was deeply familiar with the natural history of the animals whose eggs he studied, and his knowledge of natural settings led him to emphasize the importance of using laboratory conditions that closely match those in nature. Based on more than 30 years of work, he came to believe that it was the cell surface that played the most critical role in development, heredity, and evolution. He promoted a holistic view of cells and organisms in opposition to the gene-centric view that was becoming more prevalent with the rise of genetics, but rejected the vitalism espoused by some biologists of his era, calling instead for "a physics and chemistry in a new dimension superimposed upon the now known physics and chemistry" to account for biological phenomena. Just's incisive critique of genetic reductionism finds echoes in contemporary multiscale, systems approaches in biology. His speculations on the relationship between developmental and evolutionary mechanisms resonate with today's evolutionary developmental biology. After a brief biographical sketch, this paper outlines and discusses some of Just's scientific contributions, and shows how his ideas remain relevant today.
Assuntos
Invertebrados/embriologia , Invertebrados/genética , Óvulo/fisiologia , Interações Espermatozoide-Óvulo , Animais , Organismos Aquáticos , Evolução Biológica , Desenvolvimento Embrionário , História do Século XIX , História do Século XX , PartenogêneseRESUMO
As part of an overall project to characterize the streptomycin phosphotransferase enzyme APH(6)-Id, which confers bacterial resistance to streptomycin, we cloned, expressed, purified, and characterized the enzyme. When expressed in Escherichia coli, the recombinant enzyme increased by up to 70-fold the minimum inhibitory concentration needed to inhibit cell growth. Size-exclusion chromatography gave a molecular mass of 31.4 ± 1.3 kDa for the enzyme, showing that it functions as a monomer. Activity was assayed using three methods: (1) an HPLC-based method that measures the consumption of streptomycin over time; (2) a spectrophotometric method that utilizes a coupled assay; and (3) a radioenzymatic method that detects production of (32)P-labeled streptomycin phosphate. Altogether, the three methods demonstrated that streptomycin was consumed in the APH(6)-Id-catalyzed reaction, ATP was hydrolyzed, and streptomycin phosphate was produced in a substrate-dependent manner, demonstrating that APH(6)-Id is a streptomycin phosphotransferase. Steady-state kinetic analysis gave the following results: K(m)(streptomycin) of 0.38 ± 0.13 mM, K(m)(ATP) of 1.03 ± 0.1 mM, V(max) of 3.2 ± 1.1 µmol/min/mg, and k(cat) of 1.7 ± 0.6 s(-1). Our study demonstrates that APH(6)-Id is a bona fide streptomycin phosphotransferase, functions as a monomer, and confers resistance to streptomycin.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/química , Fosfotransferases (Aceptor do Grupo Álcool)/química , Estreptomicina/farmacologia , Antibacterianos/química , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Farmacorresistência Bacteriana , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Cinética , Testes de Sensibilidade Microbiana , Fosfotransferases (Aceptor do Grupo Álcool)/biossíntese , Fosfotransferases (Aceptor do Grupo Álcool)/isolamento & purificação , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Streptomyces/enzimologia , Estreptomicina/químicaRESUMO
Matrix metalloproteinase-9 is a proteolytic enzyme capable of degrading proteins of the muscle extracellular matrix. Systemic levels of MMP-9 or its inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1), have the potential to serve as blood markers of exercise-induced muscle damage. The purpose of this study was to determine if an eccentrically-dominated task, downhill running (DHR), produces changes in plasma MMP-9 or TIMP-1 and examine the relationship between MMP-9/TIMP-1 levels and indirect indicators of muscle damage. Subjects were sedentary (SED, n=12) or had a history of concentrically-biased training (CON, n=9). MMP-9 and TIMP-1 were measured before (Pre-Ex), immediately after (Post-Ex), and 1-, 2-, 4-, and 7-days post-DHR (-10°), and compared to discomfort ratings, creatine kinase activity and strength loss. At 1-day Post-Ex, discomfort increased (5.6 ± 7.8 to 45.5 ± 19.9 mm; 0-100 mm scale), strength decreased (-6.9 ± 1.6%) and CK increased (162.9 ± 177.2%). MMP-9 was modestly but significantly increased at Post-Ex in both CONC and SED (32.7 ± 33.6%) and at 4-days in SED (66.9 ± 88.1%), Individual responses were variable, however. There were no correlations between MMPs and discomfort ratings, plasma CK or strength. While plasma MMP-9 changes may be detectable in the systemic circulation after DHR, they are small and do not correspond to other markers of damage.
Assuntos
Metaloproteinase 9 da Matriz/sangue , Músculo Esquelético/lesões , Corrida/fisiologia , Inibidor Tecidual de Metaloproteinase-1/sangue , Adulto , Biomarcadores/sangue , Creatina Quinase/sangue , Feminino , Humanos , Perna (Membro)/fisiologia , Masculino , Força Muscular/fisiologia , Adulto JovemRESUMO
Exercise efficiency at low power outputs, energetically comparable to daily living activities, can be influenced by homeostatic perturbations (e.g., weight gain/loss). However, an appropriate efficiency calculation for low power outputs used in these studies has not been determined. Fifteen active subjects (seven females, eight males) performed 14, 5-min cycling trials: two types of seated rest (cranks vertical and horizontal), passive (motor-driven) cycling, no-chain cycling, no-load cycling, cycling at low (10, 20, 30, 40 W), and moderate (50, 60, 80, 100, 120 W) power outputs. Mean delta efficiency was 57% for low power outputs compared to 41.3% for moderate power outputs. Means for gross (3.6%) and net (5.7%) efficiencies were low at the lowest power output. At low power outputs, delta and work efficiency values exceeded theoretical values. In conclusion, at low power outputs, none of the common exercise efficiency calculations gave values comparable to theoretical muscle efficiency. However, gross efficiency and the slope and intercept of the metabolic power vs mechanical power output regression provide insights that are still valuable when studying homeostatic perturbations.
Assuntos
Ciclismo/fisiologia , Eficiência/fisiologia , Exercício Físico/fisiologia , Consumo de Oxigênio/fisiologia , Adulto , Feminino , Humanos , Masculino , Adulto JovemRESUMO
In the current study, we developed a HPLC method to quantitatively measure the permeability of the BpT-based chelators, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT) and 2-benzoylpyridine 4-allyl-3-thiosemicarbazone (Bp4aT), across human colorectal adenocarcinoma (Caco-2) monolayers as a model of gut absorption. In aqueous solution, Bp4eT and Bp4aT formed inter-convertible Z and E isomers that were resolved by HPLC. Peak area was linear with respect to chelator concentration. Acceptable within-day and between-day precision (<22%) and accuracy (85-115% of true values) were obtained over a range of 1.0-100µM for Bp4eT and 1.5-300µM for Bp4aT. Limits of detection were 0.3µM and 1µM for Bp4eT and Bp4aT, respectively, while corresponding limits of quantification were 1µM and 5µM. Both chelators showed significant ability to chelate iron in THP-1 cells using a calcein-based assay and no apparent cytotoxicity was observed within 24h. Ratios of the apical to basolateral and basolateral to apical transport for Bp4eT were 1.10 and 0.89 at 100µM and 300µM respectively, indicating equal bi-directional movement of the compounds. Similarly, ratios were 0.77 and 0.92 for Bp4aT, respectively. This study demonstrates that Bp4eT and Bp4aT can be efficiently transported through Caco-2 cells and can potentially be formulated for oral delivery.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Quelantes de Ferro/análise , Quelantes de Ferro/farmacocinética , Tiossemicarbazonas/análise , Tiossemicarbazonas/farmacocinética , Células CACO-2 , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Ferro/metabolismo , Quelantes de Ferro/química , Quelantes de Ferro/farmacologia , Isomerismo , Isoquinolinas/metabolismo , Limite de Detecção , Modelos Biológicos , Reprodutibilidade dos Testes , Tiossemicarbazonas/química , Tiossemicarbazonas/farmacologiaRESUMO
Ernest E. Just (1883-1941) is best known for his discovery of the "wave of negativity" that sweeps of the sea urchin egg during fertilization, and his elucidation of what are known as the fast and slow blocks to polyspermy. Just's contemporary Johannes Holtfreter (1901-1992) is known for his pioneering work in amphibian morphogenesis, which helped to lay the foundation for modern vertebrate developmental biology. This paper, after briefly describing the life and scientific contributions of Just, argues that his work and ideas strongly influenced two of the concepts for which Holtfreter is best known: tissue affinity and autoneuralization (or autoinduction). Specifically, this paper argues that, first, Just's experiments demonstrating developmental stage-specific changes in the adhesiveness of the blastomeres of cleavage embryos helped lay the foundation for Holtfreter's concept of tissue affinity and, second, Just's notion of the intrinsic irritability of the egg cell, which is evident in experimental parthenogenesis, strongly informed Holtfreter's concept of the nonspecific induction of neural tissue formation in amphibian gastrula ectoderm explants, a phenomenon known as autoinduction. Acknowledgment of these contributions by Just in no way diminishes the importance of Holtfreter's groundbreaking work. It does, however, extend the impact of Just's work into the area of embryo morphogenesis. It connects Just to Holtfreter and positions his work as an antecedent to embryo research that continues to this day.
Assuntos
Embriologia/história , Europa (Continente) , História do Século XX , Humanos , Estados UnidosRESUMO
The chloramphenicol producer Streptomyces venezuelae contains an enzyme, SvTrpEG, that has a high degree of amino acid sequence similarity to the phenazine biosynthetic enzyme PhzE of certain species of Pseudomonas. PhzE has the sequence signature of an anthranilate synthase, but recent evidence indicates that it catalyzes the production of 2-amino-2-deoxyisochorismate [corrected] (ADIC), an intermediate in the two-step anthranilate synthase reaction, not anthranilate. In order to determine if SvTrpEG is likewise an ADIC synthase, we have cloned the gene for SvTrpEG, expressed the recombinant enzyme in Escherichia coli, and purified the enzyme. Analysis of the SvTrpEG-catalyzed reaction mixture using UV-visible spectrophotometry, fluorescence spectrometry, and high-performance liquid chromatography shows that the product of the reaction is anthranilate, not ADIC. Our results therefore reveal that, despite its sequence similarity to PhzE, SvTrpEG is an anthranilate synthase, not an ADIC synthase.
Assuntos
Antranilato Sintase/genética , Genes Bacterianos/genética , Streptomyces/enzimologia , Streptomyces/genética , Antranilato Sintase/metabolismo , Sequência de Bases , Catálise , Ácido Corísmico/metabolismo , Clonagem Molecular , Eletroforese em Gel de Poliacrilamida , Escherichia coli/enzimologia , Escherichia coli/genética , Cinética , Dados de Sequência Molecular , Plasmídeos/genética , Reação em Cadeia da Polimerase , Pseudomonas/enzimologia , Pseudomonas/genética , Salmonella/enzimologia , Salmonella/genética , Termodinâmica , Transformação GenéticaRESUMO
The magnitude and pattern of the hematocrit (Hct), hemoglobin (Hb), and plasma volume (PV) responses during and upon recovery from two resistance training protocols based on either a ten-repetition maximum (10 RM) or five-repetition maximum (5 RM) resistance was examined. Twelve college-aged male resistance exercise trainers were equally divided between the protocols and performed at least four workouts prior to testing to determine the 10 RM or 5 RM for each exercise set. Each protocol included three sets of nine exercises. The 10-RM session used one-minute rest periods between sets, and two minutes between exercises. The 5-RM session employed three-minute rest periods between sets and exercises. A catheter inserted in the forearm allowed for venous blood sampling after twenty minutes supine rest, the last set of each exercise, and at fifteen and thirty minutes of recovery. Control conditions were included for posture (P) and limb motion (U). Loaded exercise (L) was significantly different from U and P controls for Hct, Hb, and PV responses. For 10 RM and 5 RM respectively, the mean change from rest was 6.2 (+/- 0.9) and 3.5 (+/- 0.4) % for Hct, 2.2 (+/- 0.2) and 1.2 (+/- 0.1) gm . dl (-1) for Hb, and - 22.6 (+/- 2.3) and - 13.0 (+/- 1.2) % for PV. The main effect for protocol was significant for Hct (p = 0.0006) and Hb (p = 0.0033), with 10-RM changes being greater than 5 RM. The greatest increase in Hct and Hb occurred after the first set for both protocols. An increase in Hct and Hb during the protocol was observed for the 10 RM, but not the 5 RM. During recovery, Hct and Hb were elevated above rest for 10 RM, but not 5 RM. PV decreases mirrored Hct and Hb in pattern of change and significance. The data demonstrate that magnitude and pattern of Hct, Hb, and PV was dependent on the type of resistance training protocol.
Assuntos
Plasma/metabolismo , Levantamento de Peso/fisiologia , Adulto , Hematócrito , Hemoglobinas/análise , Humanos , Masculino , Estados UnidosRESUMO
First put forth in June 2005, the altered nuclear transfer-oocyte assisted reprogramming (ANT-OAR) proposal has been promoted as an ethically-acceptable alternative to the embryo-destructive methods now used to obtain embryonic stem cells. According to its proponents, the goal of ANT-OAR is to use the cloning process to create a pluripotent stem cell. This would be achieved through overexpression of the transcription factor Nanog (or a hypothetical substitute) both in the enucleated egg cell and in the somatic cell prior to transfer of its nucleus. Although the ethical acceptability of ANT-OAR has been publicly debated, its scientific feasibility has not. This paper aims to help rectify this situation. It argues that ANT-OAR, as currently conceived, cannot realistically work. It presents evidence from the scientific literature showing that Nanog cannot single-handedly establish pluripotency in cells, but rather works together with a network of other transcription factors to maintain pluripotency. It argues that ANT-OAR is based on a flawed understanding of stem cell biology, and emphasizes that, in this debate about embryonic stem cells, scientists must strive to accurately and realistically assess the feasibility of the embryo research strategies they propose.
Assuntos
Reprogramação Celular/fisiologia , Pesquisas com Embriões , Bases de Conhecimento , Técnicas de Transferência Nuclear , Oócitos/citologia , Projetos de Pesquisa , Animais , Fator de Transcrição CDX2 , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/fisiologia , Pesquisas com Embriões/ética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/fisiologia , Humanos , Proteína Homeobox Nanog , Células-Tronco Pluripotentes/citologiaRESUMO
Discovered in the 1940s by Selman Waksman, the aminoglycoside antibiotic streptomycin is clinically important in the treatment of tuberculosis worldwide. However, strains of Mycobacterium tuberculosis and other pathogenic bacteria have become resistant to streptomycin. One mechanism by which this can occur is through the action of phosphotransferases that attach a phosphate group to position 6 of the streptidine ring of streptomycin, thereby inactivating it. Two such phosphotransferases are APH(6)-Ia from producer strain Streptomyces griseus, and APH(6)-Id found in animal, plant and human pathogenic isolates. Here, we report the subcloning and expression in Escherichia coli of soluble recombinant APH(6)-Ia and Id enzymes. Sequencing of aph(6)-Ia revealed a one-nucleotide disagreement with the published sequence, such that the amino acid at position 262 is an alanine instead of a serine. The sequence of aph(6)-Id is identical to that of the gene found in transposon Tn5393 of plant pathogen Erwinia amylovora. The successful expression of soluble forms of these enzymes now paves the way for experiments to study their structure and function by using site-directed mutagenesis.