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Isotopically Nonstationary Metabolic Flux Analysis (INST-MFA) of Photosynthesis and Photorespiration in Plants.
Ma F, Jazmin LJ, Young JD, Allen DK
(2017) Methods Mol Biol 1653: 167-194
MeSH Terms: Amino Acids, Arabidopsis, Carbon Dioxide, Carbon Isotopes, Chlorophyll, Chloroplasts, Isotope Labeling, Mass Spectrometry, Metabolic Flux Analysis, Metabolic Networks and Pathways, Oxygen, Oxygen Consumption, Photosynthesis, Plant Leaves, Ribulose-Bisphosphate Carboxylase, Starch, Sucrose
Show Abstract · Added September 11, 2017
Photorespiration is a central component of photosynthesis; however to better understand its role it should be viewed in the context of an integrated metabolic network rather than a series of individual reactions that operate independently. Isotopically nonstationary C metabolic flux analysis (INST-MFA), which is based on transient labeling studies at metabolic steady state, offers a comprehensive platform to quantify plant central metabolism. In this chapter, we describe the application of INST-MFA to investigate metabolism in leaves. Leaves are an autotrophic tissue, assimilating CO over a diurnal period implying that the metabolic steady state is limited to less than 12 h and thus requiring an INST-MFA approach. This strategy results in a comprehensive unified description of photorespiration, Calvin cycle, sucrose and starch synthesis, tricarboxylic acid (TCA) cycle, and amino acid biosynthetic fluxes. We present protocols of the experimental aspects for labeling studies: transient CO labeling of leaf tissue, sample quenching and extraction, mass spectrometry (MS) analysis of isotopic labeling data, measurement of sucrose and amino acids in vascular exudates, and provide details on the computational flux estimation using INST-MFA.
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17 MeSH Terms
Effect of CO on Peroxynitrite-Mediated Bacteria Killing: Response to Tsikas et al.
Gobert AP, Wilson KT
(2017) Trends Microbiol 25: 602-603
MeSH Terms: Carbon Dioxide, Helicobacter pylori, Humans, Peroxynitrous Acid
Added June 29, 2017
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4 MeSH Terms
End-Tidal Carbon Dioxide as a Prognostic Feature in Pulmonary Arterial Hypertension.
Welch CE, Brittain EL, Newman AL, Robbins IM, Pugh ME, Newman JH, Hemnes AR
(2017) Ann Am Thorac Soc 14: 896-902
MeSH Terms: Adult, Aged, Carbon Dioxide, Female, Humans, Hypertension, Pulmonary, Male, Middle Aged, Point-of-Care Testing, Prognosis, Proportional Hazards Models, Prospective Studies, Survival Analysis, Tennessee, Walk Test
Show Abstract · Added June 7, 2018
RATIONALE - Pulmonary arterial hypertension (PAH) is characterized in part by increased dead space ventilation, which can be estimated noninvasively at the bedside by measurement of end-tidal CO (ETco).
OBJECTIVES - Prior work has demonstrated that ETco is lower in patients with PAH than in control patients, but whether ETco has prognostic value is unknown. We hypothesized that lower measurements of ETco in patients with PAH correlate with worse long-term outcomes.
METHODS - Patients with PAH seen in our referral clinic were prospectively recruited for ETco measurement between September 2009 and February 2010. Vital status as of July 2015 was documented using medical records and the Social Security Death Index.
RESULTS - Eighty-two patients were followed for a median of 60 months. Twenty-six patients died, and two were lost to follow-up. Patients who died were more likely to be older (58.5 ± 14.9 vs. 47.6 ± 12.2 yr; P < 0.05) and to have shorter 6-minute walk distance (296 ± 127 vs. 401 ± 92 m; P < 0.05). Mean ETco in survivors was 30.5 ± 4.8 mm Hg, whereas mean ETco in patients who died was 27.1 ± 4.2 mm Hg (P = 0.004). After stratification by median baseline ETco of 29 mm Hg, survival in each group was analyzed. Patients with lower ETco had shorter survival (P = 0.006). Cox regression analysis with ETco as a continuous variable revealed the hazard ratio to be 0.88 (95% confidence interval, 0.80-0.97; P = 0.006). In 52 patients with more than one measurement a median of 17 months apart, ETco was unchanged.
CONCLUSIONS - Our single-center data suggest that lower ETco is associated with shorter survival and that ETco is stable over time in patients with PAH.
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MeSH Terms
Randomized Trial of Video Laryngoscopy for Endotracheal Intubation of Critically Ill Adults.
Janz DR, Semler MW, Lentz RJ, Matthews DT, Assad TR, Norman BC, Keriwala RD, Ferrell BA, Noto MJ, Shaver CM, Richmond BW, Zinggeler Berg J, Rice TW, Facilitating EndotracheaL intubation by Laryngoscopy technique and apneic Oxygenation Within the ICU Investigators and the Pragmatic Critical Care Research Group
(2016) Crit Care Med 44: 1980-1987
MeSH Terms: Academic Medical Centers, Aged, Carbon Dioxide, Critical Illness, Female, Hospital Mortality, Humans, Intensive Care Units, Intubation, Intratracheal, Laryngoscopy, Length of Stay, Male, Middle Aged, Oxygen, Prospective Studies, Respiration, Artificial, Time Factors, Video Recording
Show Abstract · Added March 30, 2020
OBJECTIVE - To evaluate the effect of video laryngoscopy on the rate of endotracheal intubation on first laryngoscopy attempt among critically ill adults.
DESIGN - A randomized, parallel-group, pragmatic trial of video compared with direct laryngoscopy for 150 adults undergoing endotracheal intubation by Pulmonary and Critical Care Medicine fellows.
SETTING - Medical ICU in a tertiary, academic medical center.
PATIENTS - Critically ill patients 18 years old or older.
INTERVENTIONS - Patients were randomized 1:1 to video or direct laryngoscopy for the first attempt at endotracheal intubation.
MEASUREMENTS AND MAIN RESULTS - Patients assigned to video (n = 74) and direct (n = 76) laryngoscopy were similar at baseline. Despite better glottic visualization with video laryngoscopy, there was no difference in the primary outcome of intubation on the first laryngoscopy attempt (video 68.9% vs direct 65.8%; p = 0.68) in unadjusted analyses or after adjustment for the operator's previous experience with the assigned device (odds ratio for video laryngoscopy on intubation on first attempt 2.02; 95% CI, 0.82-5.02, p = 0.12). Secondary outcomes of time to intubation, lowest arterial oxygen saturation, complications, and in-hospital mortality were not different between video and direct laryngoscopy.
CONCLUSIONS - In critically ill adults undergoing endotracheal intubation, video laryngoscopy improves glottic visualization but does not appear to increase procedural success or decrease complications.
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MeSH Terms
A Phox2b BAC Transgenic Rat Line Useful for Understanding Respiratory Rhythm Generator Neural Circuitry.
Ikeda K, Takahashi M, Sato S, Igarashi H, Ishizuka T, Yawo H, Arata S, Southard-Smith EM, Kawakami K, Onimaru H
(2015) PLoS One 10: e0132475
MeSH Terms: Animals, Carbon Dioxide, Chemoreceptor Cells, Chromosomes, Artificial, Bacterial, Homeodomain Proteins, Humans, Oxygen, Pulmonary Gas Exchange, Rats, Rats, Transgenic, Respiratory Rate, Transcription Factors, Trapezoid Body
Show Abstract · Added September 28, 2015
The key role of the respiratory neural center is respiratory rhythm generation to maintain homeostasis through the control of arterial blood pCO2/pH and pO2 levels. The neuronal network responsible for respiratory rhythm generation in neonatal rat resides in the ventral side of the medulla and is composed of two groups; the parafacial respiratory group (pFRG) and the pre-Bötzinger complex group (preBötC). The pFRG partially overlaps in the retrotrapezoid nucleus (RTN), which was originally identified in adult cats and rats. Part of the pre-inspiratory (Pre-I) neurons in the RTN/pFRG serves as central chemoreceptor neurons and the CO2 sensitive Pre-I neurons express homeobox gene Phox2b. Phox2b encodes a transcription factor and is essential for the development of the sensory-motor visceral circuits. Mutations in human PHOX2B cause congenital hypoventilation syndrome, which is characterized by blunted ventilatory response to hypercapnia. Here we describe the generation of a novel transgenic (Tg) rat harboring fluorescently labeled Pre-I neurons in the RTN/pFRG. In addition, the Tg rat showed fluorescent signals in autonomic enteric neurons and carotid bodies. Because the Tg rat expresses inducible Cre recombinase in PHOX2B-positive cells during development, it is a potentially powerful tool for dissecting the entire picture of the respiratory neural network during development and for identifying the CO2/O2 sensor molecules in the adult central and peripheral nervous systems.
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13 MeSH Terms
Routine clinical evaluation of cerebrovascular reserve capacity using carbogen in patients with intracranial stenosis.
Donahue MJ, Dethrage LM, Faraco CC, Jordan LC, Clemmons P, Singer R, Mocco J, Shyr Y, Desai A, O'Duffy A, Riebau D, Hermann L, Connors J, Kirshner H, Strother MK
(2014) Stroke 45: 2335-41
MeSH Terms: Adult, Aged, Brain, Carbon Dioxide, Cerebrovascular Circulation, Constriction, Pathologic, Female, Humans, Intracranial Arteriosclerosis, Male, Middle Aged, Moyamoya Disease, Oxygen
Show Abstract · Added February 19, 2015
BACKGROUND AND PURPOSE - A promising method for identifying hemodynamic impairment that may serve as a biomarker for stroke risk in patients with intracranial stenosis is cerebrovascular reactivity (CVR) mapping using noninvasive MRI. Here, abilities to measure CVR safely in the clinic using hypercarbic hyperoxic (carbogen) gas challenges, which increase oxygen delivery to tissue, are investigated.
METHODS - In sequence with structural and angiographic imaging, blood oxygenation level-dependent carbogen-induced CVR scans were performed in patients with symptomatic intracranial stenosis (n=92) and control (n=10) volunteers, with a subgroup of patients (n=57) undergoing cerebral blood flow-weighted pseudocontinuous arterial spin labeling CVR. Subjects were stratified for 4 substudies to evaluate relationships between (1) carbogen and hypercarbic normoxic CVR in healthy tissue (n=10), (2) carbogen cerebral blood flow CVR and blood oxygenation level-dependent CVR in intracranial stenosis patients (n=57), (3) carbogen CVR and clinical measures of disease in patients with asymmetrical intracranial atherosclerotic (n=31) and moyamoya (n=29) disease, and (4) the CVR scan and immediate and longer-term complications (n=92).
RESULTS - Noninvasive blood oxygenation level-dependent carbogen-induced CVR values correlate with (1) lobar hypercarbic normoxic gas stimuli in healthy tissue (R=0.92; P<0.001), (2) carbogen-induced cerebral blood flow CVR in patients with intracranial stenosis (R=0.30-0.33; P<0.012), and (3) angiographic measures of disease severity both in atherosclerotic and moyamoya patients after appropriate processing. No immediate stroke-related complications were reported in response to carbogen administration; longer-term neurological events fell within the range for expected events in this patient population.
CONCLUSIONS - Carbogen-induced CVR elicited no added adverse events and provided a surrogate marker of cerebrovascular reserve consistent with intracranial vasculopathy.
© 2014 American Heart Association, Inc.
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13 MeSH Terms
Controlled colonic insufflation by a remotely triggered capsule for improved mucosal visualization.
Pasricha T, Smith BF, Mitchell VR, Fang B, Brooks ER, Gerding JS, Washington MK, Valdastri P, Obstein KL
(2014) Endoscopy 46: 614-8
MeSH Terms: Animals, Capsule Endoscopy, Carbon Dioxide, Colon, Colonoscopy, Feasibility Studies, Female, Insufflation, Intestinal Mucosa, Swine
Show Abstract · Added April 12, 2016
BACKGROUND AND STUDY AIMS - Capsule endoscopy is an attractive alternative to colorectal cancer screening by conventional colonoscopy, but is currently limited by compromised mucosal visibility because of the lack of safe, controlled colonic insufflation. We have therefore developed a novel system of untethered, wireless-controlled carbon dioxide (CO2) insufflation for use in colonic capsule endoscopy, which this study aims to assess in vivo.
MATERIAL AND METHODS - This observational, nonsurvival, in vivo study used five Yorkshire-Landrace cross swine. A novel insufflation capsule was placed in the porcine colons, and we recorded volume of insufflation, time, force, visualization, and a pathologic assessment of the colon.
RESULTS - The mean (standard deviation [SD]) diameter of insufflation was 32.1 (3.9) mm. The volume of CO2 produced successfully allowed complete endoscopic visualization of the mucosa and safe proximal passage of the endoscope. Pathologic examination demonstrated no evidence of trauma caused by the capsule.
CONCLUSIONS - These results demonstrate the feasibility of a novel method of controlled colonic insufflation via an untethered capsule in vivo. This technological innovation addresses a critical need in colon capsule endoscopy.
© Georg Thieme Verlag KG Stuttgart · New York.
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10 MeSH Terms
Isotopically nonstationary MFA (INST-MFA) of autotrophic metabolism.
Jazmin LJ, O'Grady JP, Ma F, Allen DK, Morgan JA, Young JD
(2014) Methods Mol Biol 1090: 181-210
MeSH Terms: Carbon Dioxide, Carbon Isotopes, Enzyme Assays, Isotope Labeling, Kinetics, Metabolic Flux Analysis, Phosphoenolpyruvate Carboxylase, Photosynthesis, Plant Leaves, Plant Proteins, Plants, Seeds, Starch, Sugar Phosphates, Tandem Mass Spectrometry, Transaminases
Show Abstract · Added January 23, 2015
Metabolic flux analysis (MFA) is a powerful approach for quantifying plant central carbon metabolism based upon a combination of extracellular flux measurements and intracellular isotope labeling measurements. In this chapter, we present the method of isotopically nonstationary (13)C MFA (INST-MFA), which is applicable to autotrophic systems that are at metabolic steady state but are sampled during the transient period prior to achieving isotopic steady state following the introduction of (13)CO2. We describe protocols for performing the necessary isotope labeling experiments, sample collection and quenching, nonaqueous fractionation and extraction of intracellular metabolites, and mass spectrometry (MS) analysis of metabolite labeling. We also outline the steps required to perform computational flux estimation using INST-MFA. By combining several recently developed experimental and computational techniques, INST-MFA provides an important new platform for mapping carbon fluxes that is especially applicable to autotrophic organisms, which are not amenable to steady-state (13)C MFA experiments.
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16 MeSH Terms
Stability of cerebral metabolism and substrate availability in humans during hypoxia and hyperoxia.
Ainslie PN, Shaw AD, Smith KJ, Willie CK, Ikeda K, Graham J, Macleod DB
(2014) Clin Sci (Lond) 126: 661-70
MeSH Terms: Adult, Analysis of Variance, Blood Flow Velocity, Blood Glucose, Brain, Carbohydrate Metabolism, Carbon Dioxide, Cerebrovascular Circulation, Energy Metabolism, Female, Humans, Hyperoxia, Hypoxia, Lactic Acid, Male, Oxygen, Partial Pressure, Time Factors, Young Adult
Show Abstract · Added October 20, 2015
Characterization of the influence of oxygen availability on brain metabolism is an essential step toward a better understanding of brain energy homoeostasis and has obvious clinical implications. However, how brain metabolism depends on oxygen availability has not been clearly examined in humans. We therefore assessed the influence of oxygen on CBF (cerebral blood flow) and CMRO2 (cerebral metabolic rates for oxygen) and carbohydrates. PaO2 (arterial partial pressure of oxygen) was decreased for 15 min to ~60, ~44 and ~35 mmHg [to target a SaO2 (arterial oxygen saturation) of 90, 80 and 70% respectively], and elevated to ~320 and ~430 mmHg. Isocapnia was maintained during each trial. At the end of each stage, arterial-jugular venous differences and volumetric CBF were measured to directly calculate cerebral metabolic rates. During progressive hypoxaemia, elevations in CBF were correlated with the reductions in both SaO2 (R2=0.54, P<0.05) and CaO2 (arterial oxygen content) (R2=0.57, P<0.05). Despite markedly reduced CaO2, cerebral oxygen delivery was maintained by increased CBF. Cerebral metabolic rates for oxygen, glucose and lactate remained unaltered during progressive hypoxia. Consequently, cerebral glucose delivery was in excess of that required, and net lactate efflux increased slightly in severe hypoxia, as reflected by a small increase in jugular venous lactate. Progressive hyperoxia did not alter CBF, CaO2, substrate delivery or cerebral metabolism. In conclusion, marked elevations in CBF with progressive hypoxaemia and related reductions in CaO2 resulted in a well-maintained cerebral oxygen delivery. As such, cerebral metabolism is still supported almost exclusively by carbohydrate oxidation during severe levels of hypoxaemia.
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19 MeSH Terms
Apical oscillations in amnioserosa cells: basolateral coupling and mechanical autonomy.
Jayasinghe AK, Crews SM, Mashburn DN, Hutson MS
(2013) Biophys J 105: 255-65
MeSH Terms: Animals, Biomechanical Phenomena, Carbon Dioxide, Cell Polarity, Cell Separation, Drosophila melanogaster, Embryo, Nonmammalian, Mechanical Phenomena, Models, Biological
Show Abstract · Added September 26, 2013
Holographic laser microsurgery is used to isolate single amnioserosa cells in vivo during early dorsal closure. During this stage of Drosophila embryogenesis, amnioserosa cells undergo oscillations in apical surface area. The postisolation behavior of individual cells depends on their preisolation phase in these contraction/expansion cycles: cells that were contracting tend to collapse quickly after isolation; cells that were expanding do not immediately collapse, but instead pause or even continue to expand for ∼40 s. In either case, the postisolation apical collapse can be prevented by prior anesthetization of the embryos with CO2. These results suggest that although the amnioserosa is under tension, its cells are subjected to only small elastic strains. Furthermore, their postisolation apical collapse is not a passive elastic relaxation, and both the contraction and expansion phases of their oscillations are driven by intracellular forces. All of the above require significant changes to existing computational models.
Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.
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9 MeSH Terms