scholarly journals ASSESSMENT OF A MICROPLATE SYSTEM FOR MEASURING INDIVIDUAL REAL-TIME RESPIRATION IN SMALL MODEL ORGANISMS OF AGING

2019 ◽  
Vol 3 (Supplement_1) ◽  
pp. S918-S919
Author(s):  
Ashley N Turner ◽  
Jessica M Hoffman ◽  
Mickie L Powell ◽  
Melissa J Sammy ◽  
Douglas R Moellering ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Real Time ◽  
Mitochondrial Function ◽  
Genetic Model ◽  
Living Organism ◽  
Preliminary Evidence ◽  
Model Organisms ◽  
Zebrafish Embryos ◽  
Clinical Genetic ◽  
Consumption Rates

Abstract The ability to measure oxygen consumption rates of a living organism in real-time provides an indirect method of monitoring dynamic changes in metabolism reflecting organismal level mitochondrial function. In this study, we assessed the Loligo Systems microplate system for measuring individual respiration in small organisms. This included adult nematodes (Caenorhabditis elegans, N2), zebrafish embryos (Danio rerio, AB), and adult fruit flies (Drosophila melanogaster, w1118). Organisms were placed inside 80 µL glass chambers on a 24-well microplate atop a 24-channel optical fluorescence oxygen reading device. Adult nematodes and zebrafish embryos were in liquid culture, M9 buffer and egg water respectively, and the adult flies were in room air. The microplate and reader were placed inside an incubator for temperature control. A silicone gasket with a thin liner was used to seal the chambers. Reference standard oxygen consumption (respiration) of single and multiple adult nematodes (n=1–4 animals/well), zebrafish embryos (n=1–4 animals/well), and adult flies (n=1–2 animals/well) in the microplate system were achieved. Significant differences across numbers of animals/well and by sex were observed. Validation experiments of the oxygen consumption rates measured in C. elegans in parallel with Seahorse extracellular flux (XF) experiments are underway. The Loligo Systems microplate system offers a non-invasive, non-destructive method to measure real-time respiration in smaller organisms. These data provide preliminary evidence for utility of the system for a variety of biomedical applications that relate to organismal and mitochondrial function/dysfunction, including research in the basic biology of aging in these highly-utilized, pre-clinical, genetic model organisms.

scholarly journals A Multivariate Metabolomics Method for Estimating Platelet Mitochondrial Oxygen Consumption Rates in Patients with Sepsis

Metabolites ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 139
Author(s):  
Marc R. McCann ◽  
Cora E. McHugh ◽  
Maggie Kirby ◽  
Theodore S. Jennaro ◽  
Alan E. Jones ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Mitochondrial Function ◽  
Regression Models ◽  
Consumption Rate ◽  
P Value ◽  
Linear Regression Models ◽  
Mitochondrial Oxygen Consumption ◽  
Non Invasive ◽  
Consumption Rates ◽  
Multiple Linear Regression Models

Background: Sepsis-induced alterations in mitochondrial function contribute to organ dysfunction and mortality. Measuring mitochondrial function in vital organs is neither feasible nor practical, highlighting the need for non-invasive approaches. Mitochondrial function may be reflected in the concentrations of metabolites found in platelets and whole blood (WB) samples. We proposed to use these as alternates to indirectly estimate platelet mitochondrial oxygen consumption rate (mOCR) in sepsis patients. Methods: We determined the relationships between platelet mOCR and metabolites in both platelets and WB, as measured by quantitative 1H-NMR metabolomics. The associations were identified by building multiple linear regression models with stepwise forward-backward variable selection. We considered the models to be significant with an ANOVA test (p-value ≤ 0.05) and a positive predicted-R2. Results: The differences in adjusted-R2 and ANOVA p-values (platelet adj-R2: 0.836 (0.0003), 0.711 (0.0004) vs. WB adj-R2: 0.428 (0.0079)) from the significant models indicate the platelet models were more associated with platelet mOCR. Conclusions: Our data suggest there are groups of metabolites in WB (leucine, acetylcarnitine) and platelets (creatine, ADP, glucose, taurine) that are associated with platelet mOCR. Thus, WB and platelet metabolites could be used to estimate platelet mOCR.

scholarly journals NFκB Regulates Muscle Development and Mitochondrial Function

2018 ◽  
Vol 75 (4) ◽  
pp. 647-653 ◽  
Author(s):  
Joseph M Valentine ◽  
Mengyao E Li ◽  
Steven E Shoelson ◽  
Ning Zhang ◽  
Robert L Reddick ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Life Span ◽  
Muscle Mass ◽  
Mitochondrial Function ◽  
Muscle Development ◽  
Tissue Mass ◽  
Lower Oxygen ◽  
Consumption Rates ◽  
Nfκb Pathway ◽  
Gastrocnemius Muscles

Abstract Nuclear factor (NF)κB is a transcription factor that controls immune and inflammatory signaling pathways. In skeletal muscle, NFκB has been implicated in the regulation of metabolic processes and tissue mass, yet its affects on mitochondrial function in this tissue are unclear. To investigate the role of NFκB on mitochondrial function and its relationship with muscle mass across the life span, we study a mouse model with muscle-specific NFκB suppression (muscle-specific IκBα super-repressor [MISR] mice). In wild-type mice, there was a natural decline in muscle mass with aging that was accompanied by decreased mitochondrial function and mRNA expression of electron transport chain subunits. NFκB inactivation downregulated expression of PPARGC1A, and upregulated TFEB and PPARGC1B. NFκB inactivation also decreased gastrocnemius (but not soleus) muscle mass in early life (1–6 months old). Lower oxygen consumption rates occurred in gastrocnemius and soleus muscles from young MISR mice, whereas soleus (but not gastrocnemius) muscles from old MISR mice displayed increased oxygen consumption compared to age-matched controls. We conclude that the NFκB pathway plays an important role in muscle development and growth. The extent to which NFκB suppression alters mitochondrial function is age dependent and muscle specific. Finally, mitochondrial function and muscle mass are tightly associated in both genotypes and across the life span.

scholarly journals Measuring mitochondrial respiration in intact single muscle fibers

2012 ◽  
Vol 302 (6) ◽  
pp. R712-R719 ◽  
Author(s):  
Rosemary A. Schuh ◽  
Kathryn C. Jackson ◽  
Ramzi J. Khairallah ◽  
Christopher W. Ward ◽  
Espen E. Spangenburg
Keyword(s):  
Skeletal Muscle ◽  
Oxygen Consumption ◽  
Mitochondrial Function ◽  
Muscle Fibers ◽  
Peak Oxygen Consumption ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Total Oxygen ◽  
Novel Approach ◽  
Consumption Rates

Measurement of mitochondrial function in skeletal muscle is a vital tool for understanding regulation of cellular bioenergetics. Currently, a number of different experimental approaches are employed to quantify mitochondrial function, with each involving either mechanically or chemically induced disruption of cellular membranes. Here, we describe a novel approach that allows for the quantification of substrate-induced mitochondria-driven oxygen consumption in intact single skeletal muscle fibers isolated from adult mice. Specifically, we isolated intact muscle fibers from the flexor digitorum brevis muscle and placed the fibers in culture conditions overnight. We then quantified oxygen consumption rates using a highly sensitive microplate format. Peak oxygen consumption rates were significantly increased by 3.4-fold and 2.9-fold by simultaneous stimulation with the uncoupling agent, carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP), and/or pyruvate or palmitate exposure, respectively. However, when calculating the total oxygen consumed over the entire treatment, palmitate exposure resulted in significantly more oxygen consumption compared with pyruvate. Further, as proof of principle for the procedure, we isolated fibers from the mdx mouse model, which has known mitochondrial deficits. We found significant reductions in initial and peak oxygen consumption of 51% and 61% compared with fibers isolated from the wild-type (WT) animals, respectively. In addition, we determined that fibers isolated from mdx mice exhibited less total oxygen consumption in response to the FCCP + pyruvate stimulation compared with the WT mice. This novel approach allows the user to make mitochondria-specific measures in a nondisrupted muscle fiber that has been isolated from a whole muscle.

scholarly journals A cellular isolation system for real-time single-cell oxygen consumption monitoring

2008 ◽  
Vol 5 (suppl_2) ◽  
Author(s):  
Joe Dragavon ◽  
Tim Molter ◽  
Cody Young ◽  
Tim Strovas ◽  
Sarah McQuaide ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Single Cell ◽  
Real Time ◽  
Physiological State ◽  
Isolation System ◽  
Consumption Rates ◽  
Actuator System ◽  
The University ◽  
Oxygen Measurements

The development of a cellular isolation system (CIS) that enables the monitoring of single-cell oxygen consumption rates in real time is presented. The CIS was developed through a multidisciplinary effort within the Microscale Life Sciences Center (MLSC) at the University of Washington. The system comprises arrays of microwells containing Pt-porphyrin-embedded polystyrene microspheres as the reporter chemistry, a lid actuator system and a gated intensified imaging camera, all mounted on a temperature-stabilized confocal microscope platform. Oxygen consumption determination experiments were performed on RAW264.7 mouse macrophage cells as proof of principle. Repeatable and consistent measurements indicate that the oxygen measurements did not adversely affect the physiological state of the cells measured. The observation of physiological rates in real time allows studies of cell-to-cell heterogeneity in oxygen consumption rate to be performed. Such studies have implications in understanding the role of mitochondrial function in the progression of inflammatory-based diseases, and in diagnosing and treating such diseases.

The Effect of Real-Time Feedback Using a Smartwatch on the Clinical Behavior of Novice Student Clinicians

2017 ◽  
Vol 2 (11) ◽  
pp. 79-90
Author(s):  
Courtney G. Scott ◽  
Trina M. Becker ◽  
Kenneth O. Simpson
Keyword(s):  
Real Time ◽  
Rating Scales ◽  
Qualitative Interviews ◽  
Communication Disorders ◽  
Preliminary Evidence ◽  
Single Subject ◽  
Clinical Behavior ◽  
Stimulus Response ◽  
Treatment Data ◽  
Computer Monitors

The use of computer monitors to provide technology-based written feedback during clinical sessions, referred to as “bug-in-the-eye” (BITi) feedback, recently emerged in the literature with preliminary evidence to support its effectiveness (Carmel, Villatte, Rosenthal, Chalker & Comtois, 2015; Weck et al., 2016). This investigation employed a single-subject, sequential A-B design with two participants to observe the effects of implementing BITi feedback using a smartwatch on the clinical behavior of student clinicians (SCs). Baseline and treatment data on the stimulus-response-consequence (S-R-C) contingency completion rates of SCs were collected using 10 minute segments of recorded therapy sessions. All participants were students enrolled in a clinical practicum experience in a communication disorders and sciences (CDS) program. A celeration line, descriptive statistics, and stability band were used to analyze the data by slope, trend, and variability. Results demonstrated a significant correlative relationship between BITi feedback with a smartwatch and an increase in positive clinical behaviors. Based on qualitative interviews and exit rating scales, SCs reported BITi feedback was noninvasive and minimally distracting. Preliminary evidence suggests BITi feedback with a smartwatch may be an effective tool for providing real-time clinical feedback.

scholarly journals The Impact of Exercise Serum on Selected Parameters of CD4+ T Cell Metabolism

Immuno ◽  
2021 ◽  
Vol 1 (3) ◽  
pp. 119-131
Author(s):  
Jana Palmowski ◽  
Kristina Gebhardt ◽  
Thomas Reichel ◽  
Torsten Frech ◽  
Robert Ringseis ◽  
...  
Keyword(s):  
T Cells ◽  
Oxygen Consumption ◽  
T Cell ◽  
Real Time ◽  
Cd4 T Cells ◽  
Cell Metabolism ◽  
Cd4 T Cell ◽  
Autologous Serum ◽  
Cell Phenotype ◽  
The Impact

CD4+ T cells are sensitive to peripheral changes of cytokine levels and metabolic substrates such as glucose and lactate. This study aimed to analyze whether factors released after exercise alter parameters of human T cell metabolism, specifically glycolysis and oxidative phosphorylation. We used primary human CD4+ T cells activated in the presence of autologous serum, which was collected before (CO) and after a 30-min exercise intervention (EX). In the course of activation, cells and supernatants were analyzed for cell viability and diameter, real-time oxygen consumption by using PreSens Technology, mRNA expression of glycolytic enzymes and complexes of the electron transport chain by real-time PCR, glucose, and lactate levels in supernatants, and in vitro differentiation by flow cytometry. EX did not alter T cell phenotype, viability, or on-blast formation. Similarly, no difference between CO and EX were found for CD4+ T cell activation and cellular oxygen consumption. In contrast, higher levels of glucose were found after 48 h activation in EX conditions. T cells activated in autologous exercise serum expressed lower HK1 mRNA and higher IFN-γ receptor 1. We suggest that the exercise protocol used was not sufficient to destabilize the immune metabolism of T cells. Therefore, more intense and prolonged exercise should be used in future studies.

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