Measurement Of Oxygen Consumption Research Articles

Analysis of oxygen consumption rates in zebrafish reveals differences based on sex, age and physical activity recovery.

Introduction: Mitochondrial dysfunction is linked to a variety of human diseases. Understanding the dynamic alterations in mitochondrial respiration at various stages of development is important to our understanding of disease progression. Zebrafish provide a system for investigating mitochondrial function and alterations during different life stages. The purpose of this study was to investigate our ability to measure mitochondrial oxygen consumption rates in zebrafish embryos, larvae, and adults as an indicator of mitochondrial function. Methods: Basal respiration of entire zebrafish embryos (5dpf), larvae (0.6-0.9cm), young adults (3-month-old), and old adults (12-month-old) was measured using an Oroboros Oxygraph, with a stirrer speed of 26rpm. For embryos and larvae, "leak" respiration (plus oligomycin), maximum respiration (plus uncoupler), non-mitochondrial respiration (plus inhibitors), and complex IV activity were also measured. To induce physical activity in adult fish, the stirrer speed was increased to 200rpm. Results and Discussion: We demonstrate the ability to accurately measure respiration rates in zebrafish at various ages using the Oroboros Oxygraph. When comparing zebrafish embryos to larvae, embryos have a higher maximum respiration. Three-month-old zebrafish males have higher basal respiration than females, while 12-month-old zebrafish females exhibit greater rates of respiration than males and younger females. When the stirrer speed was increased, respiration rates decrease, but with differences depending on sex. This study demonstrates a simple and accessible method to assess zebrafish physiology by mitochondrial oxygen consumption measurements in an unmodified Oroboros Oxygraph. The method should facilitate studies to understand the intricate interplay between mitochondrial function, development, and aging.

Continuous Monitoring of Entropy Production and Entropy Flow in Humans Exercising under Heat Stress.

Complex living systems, such as the human organism, are characterized by their self-organized and dissipative behaviors, where irreversible processes continuously produce entropy internally and export it to the environment; however, a means by which to measure human entropy production and entropy flow over time is not well-studied. In this article, we leverage prior experimental data to introduce an experimental approach for the continuous measurement of external entropy flow (released to the environment) and internal entropy production (within the body), using direct and indirect calorimetry, respectively, for humans exercising under heat stress. Direct calorimetry, performed with a whole-body modified Snellen calorimeter, was used to measure the external heat dissipation from the change in temperature and relative humidity between the air outflow and inflow, from which was derived the rates of entropy flow of the body. Indirect calorimetry, which measures oxygen consumption and carbon dioxide production from inspired and expired gases, was used to monitor internal entropy production. A two-compartment entropy flow model was used to calculate the rates of internal entropy production and external entropy flow for 11 middle-aged men during a schedule of alternating exercise and resting bouts at a fixed metabolic heat production rate. We measured a resting internal entropy production rate of (0.18 ± 0.01) W/(K·m2) during heat stress only, which is in agreement with published measurements. This research introduces an approach for the real-time monitoring of entropy production and entropy flow in humans, and aims for an improved understanding of human health and illness based on non-equilibrium thermodynamics.

Analysis of Individual V˙O2max Responses during a Cardiopulmonary Exercise Test and the Verification Phase in Physically Active Women.

This study aimed to investigate the test-retest reliability, mean, and individual responses in the measurement of maximal oxygen consumption (V˙O2max) during a cardiopulmonary exercise test (CPET) and the verification phase during cycle ergometry in women. Nine women (22 ± 2 yrs, 166.0 ± 4.5 cm, 58.6 ± 7.7 kg) completed a CPET, passively rested for 5 min, and then completed a verification phase at 90% of peak power output to determine the highest V˙O2 from the CPET (V˙O2CPET) and verification phase (V˙O2verification) on 2 separate days. Analyses included a two-way repeated measures ANOVA, intraclass correlation coefficients (ICC2,1), standard errors of the measurement (SEM), minimal differences (MD), and coefficients of variation (CoV). There was no test (test 1 versus test 2) × method (CPET vs. verification phase) interaction (p = 0.896) and no main effect for method (p = 0.459). However, test 1 (39.2 mL·kg-1·min-1) was significantly higher than test 2 (38.3 mL·kg-1·min-1) (p = 0.043). The V˙O2CPET (ICC = 0.984; CoV = 1.98%; SEM = 0.77 mL·kg-1·min-1; MD = 2.14 mL·kg-1·min-1) and V˙O2verification (ICC = 0.964; CoV = 3.30%; SEM = 1.27 mL·kg-1·min-1; MD = 3.52 mL·kg-1·min-1) demonstrated "excellent" reliability. Two subjects demonstrated a test 1 V˙O2CPET that exceeded the test 2 V˙O2CPET, and one subject demonstrated a test 1 V˙O2verification that exceeded the test 2 V˙O2verification by more than the respective CPET and verification phase MD. One subject demonstrated a V˙O2CPET that exceeded the V˙O2verification, and one subject demonstrated a V˙O2verification that exceeded the V˙O2CPET by more than the MD. These results demonstrate the importance of examining the individual responses in the measurement of the V˙O2max and suggest that the MD may be a useful threshold to quantify real individual changes in V˙O2.

Energy utilization and requirement of broiler breeders during the production phase

This study aimed to evaluate energy utilization and propose models for metabolizable and net energy requirements in broiler breeders during the egg production phase. Sixty Cobb500 broiler breeders aged between 29 and 65 wk were randomly assigned to 3 feeding levels. At each age, 6 birds were adapted for 8 d to 3 levels of metabolizable energy intake (MEi), established based on the amount of feed allocated: 1) the amount of feed recommended in the guideline, 2) 25% above, and 3) 25% below. The birds were housed in respirometry chambers for 6 d (1 adaptation, 4 feeding state, 1 fasting) to measure oxygen consumption and carbon dioxide production to calculate heat production (HP) and fasting HP (FHP). Daily measurements of feed intake, egg weight, egg production, and total excreta were recorded. Variables of MEi, HP, and retained energy (RE) in the egg were calculated, while RE in the body and its partitioning into fat and protein in the egg and body were calculated from MEi, total HP (THP), and RE in the egg. Statistical analysis involved linear regression of multiple factors with MEi and age (categorical) as the independent variables. Pearson correlation analysis was conducted to investigate the relationship between visceral mass and the evaluated variables. The study proposed mixed models for developing models of energy requirements for both metabolizable (ME) and net systems (NE). The study found that FHP (average 259 ± 20.08 kJ/kg0.75*d) remained constant throughout the production cycle regardless of the MEi level (P > 0.05). The efficiency of energy utilization for depositing protein and fat in the body changed with the bird's age. The lower error model was considered to select ME requirements for maintenance, egg, and gain efficiencies, disregarding the effect of age. The efficiencies were 0.89, 0.78, and 0.80 for maintenance, gain, and egg production, respectively. The NE was unaffected by age and showed a lower error than the ME model. The NE system was found to be more accurate in expressing the energy requirements of broiler breeders.

Quantification of regional CMRO2 in human brain using dynamic 17O-MRI at 3T

ObjectiveTo investigate the feasibility of cerebral metabolic rate of oxygen consumption (CMRO2) measurements with MRI at 3 Tesla in different brain regions. MethodsCMRO2 represents a key indicator of the physiological state of brain tissue. Dynamic 17O-MRI with inhalation of isotopically enriched 17O gas has been used to quantify global CMRO2 in brain white (WM) and gray matter (GM). However, global CMRO2 can only reflect the overall oxygen metabolism of the brain and cannot provide enough information on local tissue oxygen metabolism. To investigate the feasibility of determination of regional CMRO2 at a clinical 3 T MRI system, CMRO2 values in frontal, parietal and occipital WM and GM were determined in 5 healthy volunteers and compared to evaluate the regional differences of oxygen metabolism in WM and GM. Additionally, regional CMRO2 values were determined in deep brain structures including thalamus, dorsal striatum, caudate nucleus and insula cortex and in the cerebella, and compared with literature values from 15O-PET studies. ResultsIn cortical GM the determined CMRO2 values were in good agreement with the literature, whereas values in WM were about 32–48% higher than literature values. Regional analysis revealed a significantly higher CMRO2 in the occipital GM compared to the frontal and parietal GM. By contrast, no significant difference of CMRO2 was observed across the WM. In addition, CMRO2 in deep brain structures was lower compared to literature values and in the cerebella a good hemispheric symmetry of the tissue oxygen metabolism was found. ConclusionDynamic 17O-MRI enables direct, non-invasive determination of regional CMRO2 in brain structures in healthy volunteers at 3T.

Tissue oxygen partial pressure as a viability metric for ex vivo brain tissue slices

BackgroundDespite the prevalent use of the ex vivo brain slice preparation in neurophysiology research, a reliable method for judging tissue viability ― and thus suitability of a slice for inclusion in an experiment ― is lacking. The utility of indirect electrophysiological measures of tissue health is model-specific and needs to be used cautiously. In this study, we verify a more direct test of slice viability, based on tissue oxygen consumption rate. New methodWe hypothesised that the minimum intra-slice partial pressure of oxygen (pO2min) would correlate with tissue oxygen consumption rate, providing an accessible method for reliably assessing tissue viability status. Using mouse brain cortex slices, we measured tissue oxygen consumption rate using a Fick’s law diffusion–consumption model applied to full intra-tissue pO2 profiles and compared this to pO2min and 2,3,5-triphenol tetrazolium chloride (TTC) viability staining. ResultsTissue pO2min correlated strongly with oxygen consumption rate in both neurophysiological active and quiescent tissue (in “no-magnesium” and “normal” artificial cerebrospinal fluid, respectively) (R2 =49.7% and 42.1%, respectively). Both correlated with TTC viability stain. Oxygen consumption rate was positively related to the frequency of seizure-like event activity in no-magnesium artificial cerebrospinal fluid (R2 = 44.8%). Comparison with existing methodsWhile measurement of tissue oxygen levels and oxygen consumption is not new, intra-tissue pO2min is a novel approach to assess brain slice viability. ConclusionThe results confirm that tissue oxygen minimum pO2min is a robust metric for estimating tissue viability status — the lower the pO2min, the healthier the tissue.

Loxenatide attenuates ROS-mediated vascular endothelial progenitor cell damage and mitochondrial dysfunction via SIRT3/Foxo3 signaling pathway.

Diabetes mellitus (DM), becomes a main public health issue worldwide due to the rapid increase in DM patient numbers. The dysfunction of endothelial progenitor cells (EPCs) in DM patients plays a critical role in endothelial repair and the progression of DM-related vascular complications. Loxenatide is an a glucagon-like peptide 1 receptor agonist, which is used to control glycemic in type 2 diabetes patients. However, the role of Loxenatide in EPCs remains to be investigated. EPCs were isolated, characterized, and treated with Loxenatide, high-glucose, or 3-TYP. quantitative real-time polymerase chain reaction, flow cytometry, western blot, and cell counting kit-8 assay were employed to validate the expression of gene and protein expressions and cell viability, respectively. Application of Seahorse XFp to measure oxygen consumption and mitochondrial membrane potential (MMP) were measured by Seahorse XFp and MMP assay. Loxenatide attenuated high-glucose-induced reactive oxygen species (ROS) production and mitochondrial-dependent apoptosis of EPCs in a concentration-dependent manner. The EPC mitochondrial respiration dysfunction induced by high glucose was also repressed by the loxenatide treatment. The protection effect of Loxenatide on EPCs against high-glucose was applied by activating the sirtuin 3 (SIRT3)/Foxo3 signaling pathway. We demonstrated the regulatory role of Loxenatide in mitochondrial dysfunction and apoptosis of EPCs. We elucidated that Loxenatide protects EPC from high-glucose-induced apoptosis via ROS-mediated mitochondrial pathway through the SIRT3/Foxo3 signing pathway. This may provide a new therapeutical target for the treatment of DM-related vascular complications.

Clinical metabolic monitoring to inform metabolic dysregulation in sepsis.

The objective of this study was to offer further evidence of the utility of metabolic monitoring in early recognition of sepsis. Metabolic derangement in sepsis is of increasing interest. Sepsis was redefined as a dysregulated host response to infection, and studies have since emerged advising that disrupted metabolic pathways in sepsis may interfere with the host's ability to convert oxygen to useable energy. Indirect calorimetry (IC) is a metabolic monitoring technology that measures oxygen consumption (V02) and resting energy expenditure (REE). IC offers clinically important, specific information in terms of patient's metabolic state and has been shown to differentiate patients with sepsis from those without. Additionally, IC is more specific than predictive equations used as the established standard for clinical nutrition. Data for this retrospective descriptive study were obtained from chart review of records of critically ill patients who received metabolic monitoring while under the care of the nutrition support team. Data were retrieved from January through March of 2020. The cases included were from January 2018 through January 2020. Variables included key demographics, sepsis diagnosis and specific metabolic variables of cellular respiration and energy expenditure. For this all-male sample (N = 56), mean age was 56 years (±17.5). Significant differences were noted in V02 between the two groups of cases (sepsis and non-sepsis); (p = .026, Cohen's d = 0.618); and REE (p = .032, Cohen's d = 0.607). A strong association was found between V02 and sepsis (Eta 0.981). REE as measured by IC was statistically more specific than predictive equation (p < .001, Cohen's d = 0.527). VO2 and REE were significantly altered in subjects with sepsis in this study, demonstrating that IC may be a useful tool in identifying sepsis. This study was based on an earlier pilot which yielded similar results. Indirect calorimetry can be easily performed clinically, offering specific metabolic information that can be helpful in the determination of a diagnosis of sepsis. No patient or public contribution to this manuscript. The study design, analysis of retrospective data, and manuscript preparation were completed entirely by the authors. Sepsis remains one of the number one killers of hospitalized patients globally Current means of identifying sepsis remain overly sensitive and under-specific, making recognition of sepsis challenging for Emergency Clinicians Metabolic monitoring can be done easily in the clinical setting by nurses and respiratory therapists. Metabolic monitoring has the capability of offering further information specific to the identification of sepsis, and to further understanding of the altered metabolic phenotype of patients with sepsis.

Changes in pulmonary function in patients with breast cancer before and after radiation therapy

Background: The respiratory system is negatively affected by radiation exposure during the comprehensive treatment of breast cancer (BC), which has an effect on the treatment effectiveness and patients’ quality of life.Objective: To evaluate changes in pulmonary ventilation and diffusion characteristics and ventilation effectiveness in patients with BC before and after radiation therapy (RT).Methods: Our prospective cohort study included 232 women with BC admitted to the Meshalkin National Medical Research Center for an adjuvant RT after surgical treatment. Before and after the RT we conducted pulmonary function tests: body plethysmography, lung diffusion capacity testing, and cardiopulmonary exercise testing with measurement of peak oxygen consumption. We used multivariate regression analysis to determine predictors of pulmonary function decline during RT (decrease in the lung diffusion capacity &gt; 5% compared with baseline).Results: Pulmonary function tests prior to RT demonstrated that 15% of the patients with BC had pulmonary ventilation disorders, and 25% of the patients had pulmonary gas exchange abnormalities (most of them were newly diagnosed). In the general group following RT, we saw a significant decrease in the lung diffusion capacity (P = .044) and an increased incidence of obstructive breathing patterns among the examined patients (P = .036). Other pulmonary ventilation parameters, its effectiveness, and reserve capacity of the respiratory system after RT did not change significantly in the study group and corresponded to the reference values. The postradiation pulmonary function was negatively affected by the reduced level of pulmonary diffusion capacity at baseline (≤ 70% of the reference value) [OR 1.15 (1.03-1.34), P = .004], abnormal breathing pattern prior to RT [OR 1.08 (1.01-1.15), P = .029], involvement of regional lymph nodes (N1-3) [OR 1.10 (1.04-1.26), P = .003].Conclusion: Early postradiation respiratory disorders in patients with BC are marked by a decrease in the lung diffusion capacity and an increased incidence of obstructive patterns of pulmonary ventilation. The factors negatively affecting changes in pulmonary ventilation and diffusion characteristics after RT were the baseline pulmonary gas exchange disorders, obstructive breathing pattern prior to RT, and radiation exposure to lymph nodes. Received 28 December 2022. Revised 26 May 2023. Accepted 30 May 2023. Funding: The study did not have sponsorship. Conflict of interest: The authors declare no conflict of interest. Contribution of the authorsConception and study design: O.V. Kamenskaya, V.V. Lomivorotov Data collection and analysis: S.S. Porotnikova, E.A. Samoilova, V.G. Tihomirov Statistical analysis: I.Yu. Loginova, A.S. Klinkova Drafting the article: I.Yu. Loginova, O.V. Kamenskaya Critical revision of the article: S.E. Krasilnikov, A.M. Chernyavskiy Final approval of the version to be published: O.V. Kamenskaya, I.Yu. Loginova, A.S. Klinkova, S.S. Porotnikova, E.A. Samoilova, V.G. Tihomirov, S.E. Krasilnikov, V.V. Lomivorotov, A.M. Chernyavskiy

235-OR: Myc Activity Is Required for Mitochondrial Function and Glucose-Stimulated Insulin Secretion (GSIS) in Pancreatic ß-Cells

Glucose enhances mitochondrial function, insulin secretion and Myc expression in β-cells. β-cell-specific Myc knockout mice show glucose intolerance, hypoinsulinemia and lack of adaptive β-cell mass expansion following high-fat diet feeding. However, whether Myc regulates GSIS and mitochondrial function in β-cells is unknown. Here, we tested the effects of the Myc activity inhibitor 10058-F4 (1RH) in GSIS, mitochondrial function and metabolism in islets using islet perifusion, Seahorse and metabolomics/transcriptomics approaches. Mouse and human islets incubated 6h with 40µM 1RH displayed impaired phase 1 and phase 2 insulin secretion induced by 11mM glucose. Insulin secretion was not affected in the presence of 25mM KCl suggesting that Myc is required for glucose- but not membrane depolarization-induced insulin secretion. Since adequate mitochondrial function is essential for GSIS, we measured oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) to analyze mitochondrial respiration and glycolytic flux in these islets. 1RH significantly reduced OCR, ECAR and glucose-induced ATP production. Conversely, the Myc inducer harmine (10µM), increased basal OCR, ATP production, and ECAR and partially reversed 1RH effects on mitochondrial function. RNAseq of mouse islets treated with 1RH in 11mM glucose revealed reduced expression of both oxidative phosphorylation and β-cell signature genes but enhanced gene expression of glycolysis, ROS, fatty acid metabolism and autophagy/mitophagy pathways. Metabolomics analysis of mouse islets treated with 1RH in 11mM glucose confirmed the reduction in ATP and highlighted the accumulation of L-palmitoylcarnitine and palmitic acid suggesting inefficient β-oxidation. In conclusion, Myc is required for GSIS and mitochondrial function in β-cells. Impaired Myc action can lead to unbalanced metabolism and autophagy/mytophagy leading to β-cell dysfunction. Disclosure G.Lu: None. R.Kang: None. P.Diaz-pozo: None. J.Lee: None. M.Li: None. V.M.Victor: None. D.Scott: None. A.Garcia-ocana: Consultant; Sun Pharmaceutical Industries Ltd. Funding National Institutes of Health (R01DK126450)

Real-Time Measurement of the Mitochondrial Bioenergetic Profile of Neutrophils.

Neutrophils are the first line of defense and the most abundant leukocytes in humans. These effector cells perform functions such as phagocytosis and oxidative burst, and create neutrophil extracellular traps (NETs) for microbial clearance. New insights into the metabolic activities of neutrophils challenge the early concept that they primarily rely on glycolysis. Precise measurement of metabolic activities can unfold different metabolic requirements of neutrophils, including the tricarboxylic acid (TCA) cycle (also known as the Krebs cycle), oxidative phosphorylation (OXPHOS), pentose phosphate pathway (PPP), and fatty acid oxidation (FAO) under physiological conditions and in disease states. This paper describes a step-by-step protocol and prerequirements to measure oxygen consumption rate (OCR) as an indicator of mitochondrial respiration on mouse bone marrow-derived neutrophils, human blood-derived neutrophils, and the neutrophil-like HL60 cell line, using metabolic flux analysis on a metabolic extracellular flux analyzer. This method can be used for quantifying the mitochondrial functions of neutrophils under normal and disease conditions.

Measured Oxygen Consumption During Pediatric Cardiac Catheterization is More Accurate than Assumed Oxygen Consumption.

When calculating cardiac index (C.I.) by the Fick method, oxygen consumption (VO2) is often unknown, so assumed values are typically used. This practiceintroduces a knownsource of inaccuracy into the calculation. Using a measured VO2 (mVO2) from the CARESCAPE E-sCAiOVX module provides an alternative that may improve accuracy of C.I. calculations. Our aim is to validate this measurement in a general pediatric catheterization population and compare its accuracywith assumed VO2 (aVO2).mVO2 was recorded for all patients undergoing cardiac catheterization with general anesthesia and controlled ventilation during the study period. mVO2 was compared to the reference VO2 (refVO2) determined by the reverse Fick method using cardiac MRI (cMRI) or thermodilution (TD) as a reference standard for measurement of C.I. when available. 193 VO2 measurements were obtained, including 71 with a corresponding cMRI or TD measure of cardiac index for validation. mVO2 demonstrated satisfactory concordance and correlation with the TD- or cMRI-derived refVO2 (ρc = 0.73, r2 = 0.63) with a mean bias of -3.2% (SD ± 17.3%). Assumed VO2 demonstrated much weaker concordance and correlation with refVO2 (ρc = 0.28, r2 = 0.31) with a mean bias of + 27.5% (SD ± 30.0%). Subgroup analysis of patients < 36months of age demonstrated that error in mVO2 was not significantly different from that observed in older patients. Many previously reported prediction models for assuming VO2 performed poorly in this younger age range. Measured oxygen consumption using the E-sCAiOVX module is significantly more accurate than assumed VO2 when compared to TD- or cMRI-derived VO2 in a pediatric catheterization lab.

Interest of ergospirometry in the management of thoracic oncology surgery

Abstract Funding Acknowledgements Type of funding sources: None. Background A cardiopulmonary exercise testing (CPET) is recommended before a resection of a primary pulmonary nodule by surgical approach for patients at risk (with abnormal spirometry). The maximal oxygen consumption measurement (VO2max) evaluates the morbidity/mortality and allows to orient the surgical choice towards a lobectomy or a cuneiform resection. However, the VO2 max can be influenced by some parameters as patient’s motivation, overweight, …. Purpose The objective of this work is therefore to identify other complementary predictive factors of cardiopulmonary exercise test (CPET) in at-risk patients population with abnormal pulmonary function tests. Methods 211 subjects who underwent thoracic surgery for non-small cell lung cancer (NSCLC) between 2016 and 2020 at Erasmus Hospital and performed a CPET were analyzed and divided into 2 groups (G1 At Risk: n = 159: 10 mL/min/kg &amp;lt; VO2max &amp;lt; 20 mL/min/kg; G2 Control: n = 52: VO2max ≥ 20 mL/min/kg). Anthropological, biometric, clinical, and intraoperative data were collected based on the medical record. Results The risk group (G1) had a reduced VO2max associated with a higher VE/VCO2 slope. However, the 30-day and 1-year mortality as well as the number of minor and major complications were comparable to the control group (G2) because of the screening and the operative choices. All groups combined, VO2max was predictive of major complications (OR = 0.98, p = 0.04) and VE/VCO2 slope emerged as a predictor of total and minor postoperative complications (OR = 1.07, p = 0.0054 and respectively OR = 1.05, p = 0.046). Patients with a slope ≥ 34.5 had a higher incidence of total complications (OR = 3.16, p = 0.0004). In a subgroup with medium operative risk (VO2max between 15 and 20 mL/min/kg), VE/VCO2 slope was predictive of total complications (OR = 1.09, p = 0.02) whereas VO2max was predictive of major complications (OR = 0.97, p = 0.04).

C47 ADAPTATION OF THE PULMONARY CIRCULATION AND OF THE RIGHT HEART IN INTRACARDIAC INVETERATE SHUNTS: UNDER PRESSURE OR LET IT FLOW?

Abstract Congenital heart disease can be associated with pulmonary hypertension (PH) development and heart right remodelling. Nevertheless, the adaptation of the pulmonary circulation and of the right heart can be extremely heterogeneous, so that an accurate diagnostic work up may be advisable when an intracardiac shunt is suspected. Case Presentation A 53 years old woman, with no relevant clinical history and asymptomatic, was incidentally found to have clinical, ECG and echocardiographic signs of PH, and was referred to our Center. Diagnostic and Therapeutic Work Up Echocardiography showed: normal left ventricle (LV 3D volumes 88/35 ml), mild left atrial dilatation (39 ml/m2); severe right chambers dilatation (RV 3D 347/124 ml, EF 3D 62%, AD 116 ml/m2), moderate tricuspid regurgitation with a regurgitant jet velocity of 2.9 m/s. Right heart catheterization showed normal pulmonary pressures at rest (mPAP 20, PAWP 5 mmHg), extremely high pulmonary blood flow (Qp 29 L/min, calculated with the direct Fick method), with normal pulmonary vascular resistance (PVR 0.5 WU). During the oxymetric run, a step up in hemoglobin saturation between superior vena cava and right atrium (from 74 to 97%) was found, with a Qp/Qs ratio of 5.5. During exercise, Qp reserve was normal, suggesting a normal contractile reserve function of RV despite the extreme dilatation. Additionally, the patient did not develop exercise PH and exercise capacity was normal, based on a measured oxygen consumption (VO2) at peak of 24 ml/kg/min (104% of predicted). Angio–CT of the chest was performed, confirming the presence of a sinus venosus atrial septal defect. The patient was then referred for surgical correction. Conclusion Echocardiography signs are useful to direct correctly diagnostic work up. A severe RV dilatation with normal systolic function in an asymptomatic middle age woman should raise the possibility of an intracardiac shunt. Despite a large intracardiac shunt with a high Qp/Qs ratio for decades, PVR was normal, confirming that individual predisposition may play a role in the development of PH in pre–tricuspid shunts. Exercise test showed an optimal RV reserve, suggesting a low cardiac surgery risk.

Sediment oxygen consumption in Antarctic subglacial environments

AbstractOxygen consumption in aquatic sediments is an indicator of overall biological activity of the ecosystem. As such, rates of sedimentary oxygen utilization are well documented for much of the open oceans and freshwater lakes. However, there are few direct measurements of sedimentary oxygen consumption from Antarctic subglacial aquatic sediments. We report the first microsensor oxygen profiles and derived sedimentary oxygen consumption rates from beneath the Ross Ice Shelf and a subglacial lake beneath the West Antarctic Ice Sheet. Rates of oxygen consumption in these two environments are relatively low, but comparable to those reported from ice‐free polar oceans and oligotrophic Arctic lakes. Our study demonstrates the presence of oxygen within Antarctic subglacial aquatic sediments and its importance for oxygen‐consuming microorganisms living in these ecosystems.

Ru(II) Oxygen Sensors for Co(III) Complexes and Amphotericin B Antifungal Activity Detection by Phosphorescence Optical Respirometry.

The measurement of oxygen consumption is an important element in the understanding of an organism's metabolic state. Oxygen is also a phosphorescence quencher, which allows the evaluation of phosphorescence emitted by oxygen sensors. Two Ru(II)-based oxygen-sensitive sensors were used to study the effect of chemical compounds [(1) = [CoCl2(dap)2]Cl, and (2) = [CoCl2(en)2]Cl (AmB = amphotericin B) against reference and clinical strains of Candida albicans. The tris-[(4,7-diphenyl-1,10-phenanthroline)ruthenium(II)] chloride ([Ru(DPP)3]Cl2) (Box) adsorbed onto the DavisilTM silica gel was embedded in the silicone rubber Lactite NuvaSil® 5091 and the coating on the bottom of 96-well plates. The water-soluble oxygen sensor (BsOx = tris-[(4,7-diphenyl-1,10-phenanthrolinedisulphonic acid disodium)ruthenium(II)] chloride 'x' hydrate = {Ru[DPP(SO3Na)2]3}Cl2 = water molecules were omitted in the BsOx formula) was synthesized and characterized by RP-UHPLC, LCMS, MALDI, elemental analysis, ATR, UV-Vis, 1H NMR, and TG/IR techniques. The microbiological studies were performed in the environment of RPMI broth and blood serum. Both Ru(II)-based sensors turned out to be useful in the study of the activity of Co(III) complexes and the commercial antifungal drug amphotericin B. In addition, a new activity of the oxygen sensor, the soluble Ru(II) complex BsOx, was demonstrated, which is a mixture with amphotericin B that caused a significant increase in its antifungal activity. Thus, it is also possible to demonstrate the synergistic effect of compounds active against the microorganisms under study.

Investigating respiratory simplification caused by prenatal RAGE upregulation

Receptors for advanced glycation end-products (RAGE) are multi-ligand cell surface receptors of the immunoglobulin superfamily predominantly expressed by lung epithelium. Previous experiments showed that RAGE upregulation throughout pregnancy caused significant fetal lung hypoplasia via elevated apoptosis and misregulation of the transcription factors thyroid transcription factor 1 (TTF1) and forkhead box protein A2 (Fox-A2). Our current research focused on characterizing the contributors to these abnormalities. We confirmed poor development of murine lungs through a time course of RAGE upregulation from conception to sacrifice on embryonic day (E)15.5, E16.5, E17.5, E18.5, and specific upregulation from E15.5 to E18.5 only. H&amp;E staining confirmed significant simplification. We hypothesized that this malformation was in part due to aberrant levels of inflammation and apoptosis. Blotting revealed significantly increased levels of phosphorylated AKT and ERK1/2 in RAGE TG pups which cause NF-κB mediated transcription of inflammatory genes. Due to potential roles for these signaling intermediates, we sought to assess RAGE-mediated impact on mitochondrial efficiency. Oxidative phosphorylation was assessed in lungs from RAGE TG pups and controls by measuring oxygen consumption per unit mass when lungs were exposed to the substrates malate, glutamate, ADP, succinate, and FCCP which activate sequential components of the electron transport chain. We discovered significantly increased levels of oxidative phosphorylation in transgenic pups compared to controls. Levels of ATP and reactive oxidative species (ROS) in relation to altered oxidative phosphorylation and inflammation provided additional mechanistic clarity. These results support the importance of proper RAGE regulation in the development of lung tissue and elucidate mechanisms possibly causing such damage. This work was supported by funding from the National Institutes of Health (NIH 1R15-HL152257, PRR and JAA). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

IFNγ Augments the Oxygen Consumption Rate and Metabolism in Monocytes Partially Through P2X7

BackgroundInterferon-gamma (IFNγ) is a pleiotropic cytokine crucial for immune responses to intracellular pathogens. IFNγ causes transcriptional changes that lead to increased production of reactive oxygen species (ROS) which promote microbial killing. IFNγ increases the oxygen consumption rate (OCR), ROS production, as well as NAD salvage, all through transcription regulation by STAT1. The ATP-gated, purinergic pore P2X7 has been identified as an ion channel that allows for the influx of Ca2+ and Na+ and the efflux of K+, as well as playing a role in the processing and release of proinflammatory cytokines in macrophages. P2X7 appears to play a role in IFNγ-induced metabolic reprogramming in monocytes as well as the increase in ROS production. MethodsWe used a Seahorse Extracellular Flux analyzer to measure oxygen consumption (as an indicator of ROS production), medium acidification, and kinetics in human monocytes in response to IFNγ or inhibitors before and after PMA activation. We used small molecule inhibitors to block NAD salvage and P2X7 pathways. We also used flow cytometry to measure protein expression, using anti-P2X7 antibodies. ResultsIFNγ activation led to doubling of expression level of the purinergic pore P2X7 on monocytes as shown by flow cytometry. Inhibition of the STAT1-dependent nicotinamide phosphoribosyltransferase (NAMPT) by daporinad led to a reduction of OCR by 67%. Addition of NAD to NAMPT blocked cells circumvented the blockade and restored normal OCR. Interestingly, inhibition of P2X7 by AZ-11645373 in the setting of NADcomplemented NAMPT blockade led to ~50% reduction of OCR. In the absence of NAMPT inhibition, P2X7 blockade alone leads to about 15% reduction in OCR; in the setting of IFNγ-stimulated OCR, the reduction was 25%. ConclusionsP2X7 expression increases in monocytes following IFNγ-activation, and its patency facilitates NAD-dependent OCR. In the setting of NAMPT blockade, P2X7 plays an important role in trafficking NAD or its metabolites into cells and allowing oxygen consumption following IFNγ stimulation. Blockade of P2X7 leads to reduced OCR, which diminished even further upon IFNγ stimulation and NAMPT blockade. The ability to augment OCR through P2X7 may contribute to control of intracellular pathogens.

PO-01-124 THE CALCIUM HANDLING MACHINERY IS REMODELED IN FRIEDREICH’S ATAXIA

Friedreich's ataxia (FA) is an inherited neurodegenerative disorder that causes progressive nervous system damage. FA is caused by a deficiency in the frataxin protein due to an expansion of GAA repeats in the first intron of the frataxin gene. Reduced frataxin protein expression is thought to negatively affect mitochondrial function, leading to increased oxidative damage. Although FA is considered a neurodegenerative disorder, it is not completely understood why most patients present with left ventricular systolic dysfunction and die from cardiac events. The objective of our study is to determine whether the abnormal calcium handling machinery is a molecular mechanism that perpetuates cardiac dysfunction in FA. We used the frataxin knock-out (KO) mouse model of FA as well as human heart samples from donors with FA and from unaffected donors. The expression of calcium handling machinery proteins was assessed with proteomics and western blot. In left ventricular myocytes from KO and WT mice, the IonOptix system was used for calcium imaging, the seahorse assay was utilized to measure oxygen consumption rate (OCR) and confocal imaging was used to quantify the mitochondrial membrane potential (Δψm), and reactive oxygen species (ROS). ECG and echocardiography were used to assess cardiac function in the mice. In the proteomic analysis and western blot, SERCA2, Ryr2 and CaMKII were significantly downregulated, and the mitochondrial calcium handling proteins voltage-dependent anion channel and mitochondrial calcium uniporter were significantly upregulated in left ventricular tissue from humans with FA and from KO mice. On the ECG, the RR, PR, QRS, and QTc intervals were significantly longer in the KO mice. Echocardiography showed that the ejection fraction and fractional shortening were significantly decreased and left ventricular wall thickness and diameter were significantly increased in the KO mice. Moreover, the calcium transient decay was significantly prolonged in ventricular myocytes from KO mice compared to WT. Additionally, the Δψm was significantly depolarized, the OCR was significantly decreased, and ROS levels were significantly elevated in left ventricular myocytes from KO compared to WT. The development of left ventricular contractile dysfunction in FA is associated with reduced expression of calcium handling proteins, abnormal calcium cycling, and mitochondrial dysfunction.

Sustained mitochondrial dysfunction in male, but not female Spontaneous Hypertensive Rats (SHR) corresponds with delayed renal recovery from ischemic injury

Acute kidney injury (AKI) due to ischemia-reperfusion (IR) is a serious and frequent complication in hospital settings with high mortality rates. The mechanisms mediating renal IR injury and leading to increased risk of later developing cardiovascular and renal diseases in either sex remain poorly understood. We previously reported that there is sex difference in renal recovery from IR where males have a delayed recovery vs females. Mitochondrial dysfunction plays a pivotal role in the pathogenesis of AKI in males, resulting in abnormal kidney repair and disease progression. However, the impact of IR on mitochondrial function in females has not been examined. The goal of the current study was to test the hypothesis that IR results in greater sustained mitochondrial dysfunction in males vs. females corresponds to delayed recovery of renal function in males. 13-week-old male and female SHR were subjected to sham or 30-minute warm bilateral IR. Subsets of rats were euthanized 1- or 7-days post-IR (n=6/group). Blood was collected to measure kidney function. The right kidney was dissected and processed for electron microscopy (EM) to assess mitochondrial structure, histological analysis of renal structure, and measurement of mitochondrial DNA (mtDNA) copy number via RT-PCR. Tubular cells were isolated from the left kidney to measure oxygen consumption rate (OCR) to assess mitochondrial respiration using Seahorse XFe24 Extracellular Flex Analyzer. IR increased plasma creatinine (Pcr) in male and female SHR vs. respective sham controls 1-day post-IR (PIR =0.0001; Psex✕IR=0.2). Pcr remained elevated in male, but not female, SHR 7 days post-IR (Pcr: PIR =0.03; Psex✕IR=0.04). OCR was reduced in male and female tubules 1 day post IR (PIR =0.001; Psex=0.94) and remained reduced only in tubules from males 7 days post-IR (PIR =0.0001; Psex✕IR=0.0001). Consistent with decreased OCR, EM revealed swollen round mitochondria, reduced mitochondria density with complete loss of cristae membranes vs. sham controls 1-day post-IR in both sexes, which was sustained 7 days post-IR only in males. Mitochondrial DNA copy number and peroxisome proliferator-activator receptor-y coactivator (PGC-1α) mRNA expression was also reduced 1-day post-IR in both sexes (PIR =0.05; Psex✕IR=0.59) and remained down only in males 7 days post-IR (PIR =0.02; Psex✕IR=0.001). In conclusion, our data demonstrate that there is sex difference in mitochondria after renal IR. Males have sustained mitochondrial dysfunction with delayed renal recovery compared to females. Further studies will be carried out to better understand the direct impact of mitochondria dysfunction on renal recovery post-IR in male and females. This work was supported by career development grant from AHA (20CDA35310362) to R Mohamed; National Institutes of Health (1P01HL134604-01 and R01 HL127091 to J. C. Sullivan. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.