24 research outputs found

    DataSheet_1_13C-Metabolic flux analysis detected a hyperoxemia-induced reduction of tricarboxylic acid cycle metabolism in granulocytes during two models of porcine acute subdural hematoma and hemorrhagic shock.pdf

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    IntroductionSupplementation with increased inspired oxygen fractions has been suggested to alleviate the harmful effects of tissue hypoxia during hemorrhagic shock (HS) and traumatic brain injury. However, the utility of therapeutic hyperoxia in critical care is disputed to this day as controversial evidence is available regarding its efficacy. Furthermore, in contrast to its hypoxic counterpart, the effect of hyperoxia on the metabolism of circulating immune cells remains ambiguous. Both stimulating and detrimental effects are possible; the former by providing necessary oxygen supply, the latter by generation of excessive amounts of reactive oxygen species (ROS). To uncover the potential impact of increased oxygen fractions on circulating immune cells during intensive care, we have performed a 13C-metabolic flux analysis (MFA) on PBMCs and granulocytes isolated from two long-term, resuscitated models of combined acute subdural hematoma (ASDH) and HS in pigs with and without cardiovascular comorbidity.MethodsSwine underwent resuscitation after 2 h of ASDH and HS up to a maximum of 48 h after HS. Animals received normoxemia (PaO2 = 80 – 120 mmHg) or targeted hyperoxemia (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation, thereafter PaO2 as in the control group). Blood was drawn at time points T1 = after instrumentation, T2 = 24 h post ASDH and HS, and T3 = 48 h post ASDH and HS. PBMCs and granulocytes were isolated from whole blood to perform electron spin resonance spectroscopy, high resolution respirometry and 13C-MFA. For the latter, we utilized a parallel tracer approach with 1,2-13C2 glucose, U-13C glucose, and U-13C glutamine, which covered essential pathways of glucose and glutamine metabolism and supplied redundant data for robust Bayesian estimation. Gas chromatography-mass spectrometry further provided multiple fragments of metabolites which yielded additional labeling information. We obtained precise estimations of the fluxes, their joint credibility intervals, and their relations, and characterized common metabolic patterns with principal component analysis (PCA).Results13C-MFA indicated a hyperoxia-mediated reduction in tricarboxylic acid (TCA) cycle activity in circulating granulocytes which encompassed fluxes of glutamine uptake, TCA cycle, and oxaloacetate/aspartate supply for biosynthetic processes. We further detected elevated superoxide levels in the swine strain characterized by a hypercholesterolemic phenotype. PCA revealed cell type-specific behavioral patterns of metabolic adaptation in response to ASDH and HS that acted irrespective of swine strains or treatment group.ConclusionIn a model of resuscitated porcine ASDH and HS, we saw that ventilation with increased inspiratory O2 concentrations (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation) did not impact mitochondrial respiration of PBMCs or granulocytes. However, Bayesian 13C-MFA results indicated a reduction in TCA cycle activity in granulocytes compared to cells exposed to normoxemia in the same time period. This change in metabolism did not seem to affect granulocytes’ ability to perform phagocytosis or produce superoxide radicals. </p

    Table_2_13C-Metabolic flux analysis detected a hyperoxemia-induced reduction of tricarboxylic acid cycle metabolism in granulocytes during two models of porcine acute subdural hematoma and hemorrhagic shock.xlsx

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    IntroductionSupplementation with increased inspired oxygen fractions has been suggested to alleviate the harmful effects of tissue hypoxia during hemorrhagic shock (HS) and traumatic brain injury. However, the utility of therapeutic hyperoxia in critical care is disputed to this day as controversial evidence is available regarding its efficacy. Furthermore, in contrast to its hypoxic counterpart, the effect of hyperoxia on the metabolism of circulating immune cells remains ambiguous. Both stimulating and detrimental effects are possible; the former by providing necessary oxygen supply, the latter by generation of excessive amounts of reactive oxygen species (ROS). To uncover the potential impact of increased oxygen fractions on circulating immune cells during intensive care, we have performed a 13C-metabolic flux analysis (MFA) on PBMCs and granulocytes isolated from two long-term, resuscitated models of combined acute subdural hematoma (ASDH) and HS in pigs with and without cardiovascular comorbidity.MethodsSwine underwent resuscitation after 2 h of ASDH and HS up to a maximum of 48 h after HS. Animals received normoxemia (PaO2 = 80 – 120 mmHg) or targeted hyperoxemia (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation, thereafter PaO2 as in the control group). Blood was drawn at time points T1 = after instrumentation, T2 = 24 h post ASDH and HS, and T3 = 48 h post ASDH and HS. PBMCs and granulocytes were isolated from whole blood to perform electron spin resonance spectroscopy, high resolution respirometry and 13C-MFA. For the latter, we utilized a parallel tracer approach with 1,2-13C2 glucose, U-13C glucose, and U-13C glutamine, which covered essential pathways of glucose and glutamine metabolism and supplied redundant data for robust Bayesian estimation. Gas chromatography-mass spectrometry further provided multiple fragments of metabolites which yielded additional labeling information. We obtained precise estimations of the fluxes, their joint credibility intervals, and their relations, and characterized common metabolic patterns with principal component analysis (PCA).Results13C-MFA indicated a hyperoxia-mediated reduction in tricarboxylic acid (TCA) cycle activity in circulating granulocytes which encompassed fluxes of glutamine uptake, TCA cycle, and oxaloacetate/aspartate supply for biosynthetic processes. We further detected elevated superoxide levels in the swine strain characterized by a hypercholesterolemic phenotype. PCA revealed cell type-specific behavioral patterns of metabolic adaptation in response to ASDH and HS that acted irrespective of swine strains or treatment group.ConclusionIn a model of resuscitated porcine ASDH and HS, we saw that ventilation with increased inspiratory O2 concentrations (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation) did not impact mitochondrial respiration of PBMCs or granulocytes. However, Bayesian 13C-MFA results indicated a reduction in TCA cycle activity in granulocytes compared to cells exposed to normoxemia in the same time period. This change in metabolism did not seem to affect granulocytes’ ability to perform phagocytosis or produce superoxide radicals. </p

    Table_1_13C-Metabolic flux analysis detected a hyperoxemia-induced reduction of tricarboxylic acid cycle metabolism in granulocytes during two models of porcine acute subdural hematoma and hemorrhagic shock.xlsx

    No full text
    IntroductionSupplementation with increased inspired oxygen fractions has been suggested to alleviate the harmful effects of tissue hypoxia during hemorrhagic shock (HS) and traumatic brain injury. However, the utility of therapeutic hyperoxia in critical care is disputed to this day as controversial evidence is available regarding its efficacy. Furthermore, in contrast to its hypoxic counterpart, the effect of hyperoxia on the metabolism of circulating immune cells remains ambiguous. Both stimulating and detrimental effects are possible; the former by providing necessary oxygen supply, the latter by generation of excessive amounts of reactive oxygen species (ROS). To uncover the potential impact of increased oxygen fractions on circulating immune cells during intensive care, we have performed a 13C-metabolic flux analysis (MFA) on PBMCs and granulocytes isolated from two long-term, resuscitated models of combined acute subdural hematoma (ASDH) and HS in pigs with and without cardiovascular comorbidity.MethodsSwine underwent resuscitation after 2 h of ASDH and HS up to a maximum of 48 h after HS. Animals received normoxemia (PaO2 = 80 – 120 mmHg) or targeted hyperoxemia (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation, thereafter PaO2 as in the control group). Blood was drawn at time points T1 = after instrumentation, T2 = 24 h post ASDH and HS, and T3 = 48 h post ASDH and HS. PBMCs and granulocytes were isolated from whole blood to perform electron spin resonance spectroscopy, high resolution respirometry and 13C-MFA. For the latter, we utilized a parallel tracer approach with 1,2-13C2 glucose, U-13C glucose, and U-13C glutamine, which covered essential pathways of glucose and glutamine metabolism and supplied redundant data for robust Bayesian estimation. Gas chromatography-mass spectrometry further provided multiple fragments of metabolites which yielded additional labeling information. We obtained precise estimations of the fluxes, their joint credibility intervals, and their relations, and characterized common metabolic patterns with principal component analysis (PCA).Results13C-MFA indicated a hyperoxia-mediated reduction in tricarboxylic acid (TCA) cycle activity in circulating granulocytes which encompassed fluxes of glutamine uptake, TCA cycle, and oxaloacetate/aspartate supply for biosynthetic processes. We further detected elevated superoxide levels in the swine strain characterized by a hypercholesterolemic phenotype. PCA revealed cell type-specific behavioral patterns of metabolic adaptation in response to ASDH and HS that acted irrespective of swine strains or treatment group.ConclusionIn a model of resuscitated porcine ASDH and HS, we saw that ventilation with increased inspiratory O2 concentrations (PaO2 = 200 – 250 mmHg for 24 h after treatment initiation) did not impact mitochondrial respiration of PBMCs or granulocytes. However, Bayesian 13C-MFA results indicated a reduction in TCA cycle activity in granulocytes compared to cells exposed to normoxemia in the same time period. This change in metabolism did not seem to affect granulocytes’ ability to perform phagocytosis or produce superoxide radicals. </p

    Table_1_Mental but no bio-physiological long-term habituation to repeated social stress: A study on soldiers and the influence of mission abroad.docx

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    Soldiers regularly participate in missions abroad and subjectively adapt to this situation. However, they have an increased lifetime cardiovascular risk compared to other occupational groups. To test the hypothesis that foreign deployment results in different stress habituation patterns, we investigated long-term psychological and bio-physiological stress responses to a repeated social stress task in healthy soldiers with and without foreign deployment. Ninety-one female and male soldiers from the BEST study (German armed forces deployment and stress) participated three times in the Trier Social Stress Test for groups (TSST-G) prior to, 6–8 weeks after and 1 year after the mission abroad and were compared to a control group without foreign deployment during the study period. They completed the State-Trait-Anxiety Inventory scale (STAI), the Primary Appraisal Secondary Appraisal questionnaire (PASA) and the Multidimensional Mood State Questionnaire (MDBF). Salivary cortisol and α-amylase, blood pressure, heart rate and heart rate variability were determined. Soldiers showed mental habituation over the three times with a significant decrease after the TSST-G in anxiousness (STAI) and cognitive stress appraisal (PASA), they were calmer and reported better mood (MDBF). Prior to the social stress part, the mood (MDBF) declined significantly. None of the biological and physiological markers showed any adaptation to the TSST-G. Mission abroad did not significantly influence any measured psychobiological marker when compared to soldiers without foreign deployment. Foreign deployment does not result in alterations in psychobiological social stress response patterns over 1 year after mission abroad which indicates that adaptation to acute social stress is highly maintained in healthy soldiers. The discrepancy between subjective perception and objective stress response has numerous clinical implications and should receive more attention.</p

    A CRHR1 antagonist prevents synaptic loss and memory deficits in a trauma-induced delirium-like syndrome

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    Older patients with severe physical trauma are at high risk of developing neuropsychiatric syndromes with global impairment of cognition, attention, and consciousness. We employed a thoracic trauma (TxT) mouse model and thoroughly analyzed age-dependent spatial and temporal posttraumatic alterations in the central nervous system. Up to 5 days after trauma, we observed a transient 50% decrease in the number of excitatory synapses specifically in hippocampal pyramidal neurons accompanied by alterations in attention and motor activity and disruption of contextual memory consolidation. In parallel, hippocampal corticotropin-releasing hormone (CRH) expression was highly upregulated, and brain-derived neurotrophic factor (BDNF) levels were significantly reduced. In vitro experiments revealed that CRH application induced neuronal autophagy with rapid lysosomal degradation of BDNF via the NF-κB pathway. The subsequent synaptic loss was rescued by BDNF as well as by specific NF-κB and CRH receptor 1 (CRHR1) antagonists. In vivo, the chronic application of a CRHR1 antagonist after TxT resulted in reversal of the observed histological, molecular, and behavioral alterations. The data suggest that neuropsychiatric syndromes (i.e., delirium) after peripheral trauma might be at least in part due to the activation of the hippocampal CRH/NF-κB/BDNF pathway, which results in a dramatic loss of synaptic contacts. The successful rescue by stress hormone receptor antagonists should encourage clinical trials focusing on trauma-induced delirium and/or other posttraumatic syndromes

    Enriched Air Nitrox Breathing Reduces Venous Gas Bubbles after Simulated SCUBA Diving: A Double-Blind Cross-Over Randomized Trial

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    <div><p>Objective</p><p>To test the hypothesis whether enriched air nitrox (EAN) breathing during simulated diving reduces decompression stress when compared to compressed air breathing as assessed by intravascular bubble formation after decompression.</p><p>Methods</p><p>Human volunteers underwent a first simulated dive breathing compressed air to include subjects prone to post-decompression venous gas bubbling. Twelve subjects prone to bubbling underwent a double-blind, randomized, cross-over trial including one simulated dive breathing compressed air, and one dive breathing EAN (36% O<sub>2</sub>) in a hyperbaric chamber, with identical diving profiles (28 msw for 55 minutes). Intravascular bubble formation was assessed after decompression using pulmonary artery pulsed Doppler.</p><p>Results</p><p>Twelve subjects showing high bubble production were included for the cross-over trial, and all completed the experimental protocol. In the randomized protocol, EAN significantly reduced the bubble score at all time points (cumulative bubble scores: 1 [0–3.5] vs. 8 [4.5–10]; P < 0.001). Three decompression incidents, all presenting as cutaneous itching, occurred in the air versus zero in the EAN group (P = 0.217). Weak correlations were observed between bubble scores and age or body mass index, respectively.</p><p>Conclusion</p><p>EAN breathing markedly reduces venous gas bubble emboli after decompression in volunteers selected for susceptibility for intravascular bubble formation. When using similar diving profiles and avoiding oxygen toxicity limits, EAN increases safety of diving as compared to compressed air breathing.</p><p>Trial Registration</p><p><a href="http://www.isrctn.com/ISRCTN31681480" target="_blank">ISRCTN 31681480</a></p></div

    Exposure to 100% Oxygen Abolishes the Impairment of Fracture Healing after Thoracic Trauma

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    <div><p>In polytrauma patients a thoracic trauma is one of the most critical injuries and an important trigger of post-traumatic inflammation. About 50% of patients with thoracic trauma are additionally affected by bone fractures. The risk for fracture malunion is considerably increased in such patients, the pathomechanisms being poorly understood. Thoracic trauma causes regional alveolar hypoxia and, subsequently, hypoxemia, which in turn triggers local and systemic inflammation. Therefore, we aimed to unravel the role of oxygen in impaired bone regeneration after thoracic trauma. We hypothesized that short-term breathing of 100% oxygen in the early post-traumatic phase ameliorates inflammation and improves bone regeneration. Mice underwent a femur osteotomy alone or combined with blunt chest trauma 100% oxygen was administered immediately after trauma for two separate 3 hour intervals. Arterial blood gas tensions, microcirculatory perfusion and oxygenation were assessed at 3, 9 and 24 hours after injury. Inflammatory cytokines and markers of oxidative/nitrosative stress were measured in plasma, lung and fracture hematoma. Bone healing was assessed on day 7, 14 and 21. Thoracic trauma induced pulmonary and systemic inflammation and impaired bone healing. Short-term exposure to 100% oxygen in the acute post-traumatic phase significantly attenuated systemic and local inflammatory responses and improved fracture healing without provoking toxic side effects, suggesting that hyperoxia could induce anti-inflammatory and pro-regenerative effects after severe injury. These results suggest that breathing of 100% oxygen in the acute post-traumatic phase might reduce the risk of poorly healing fractures in severely injured patients.</p></div

    Cytokine/chemokine concentrations in lung homogenates.

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    <p>Data represent medians and quartiles. Specimen numbers for each group are depicted.</p><p>*p < 0.05 and</p><p>**p < 0.01 vs F</p><p>#p < 0.05 vs. F+TXT.</p><p>Cytokine/chemokine concentrations in lung homogenates.</p

    Immunohistological stainings of fractured femurs for markers of inflammation, nitrosative stress and vascularization.

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    <p>Left panels are representative slides of mice with isolated fracture, middle panels of animals with additional TXT, and right panels of mice with fracture, TXT and O<sub>2</sub> treatment. Images indicate cortical bone proximal to the fracture gap and periosteal callus, macrophage staining show the marrow cavity. (A-C) Neutrophil staining 3 days post-injury, (D-F) macrophage staining on day 7, (G-L) IL-6 and IL-10 staining on day 7, and (M-R) nitrotyrosine and PECAM-1 staining on day 14. Scale bars: 100 μm.</p

    Tissue composition of fracture calli 14 and 21 days after injury.

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    <p>Callus composition of mice 14 and 21 days post-injury. (A) Mice with TXT displayed significantly more cartilage in comparison to O<sub>2</sub> treated mice after 14 days. (B) Analysis after 21 days did not reveal intergroup differences. (C-E) Representative Safranin-O stained callus sections 14 days after injury. Markedly more cartilage (stained red) was observed in F+TXT mice compared to the other groups. TOT = total osseous tissue, Cg = cartilage, FT = fibrous tissue. Scale bars: 500 μm. Data represent medians and quartiles. Specimen numbers for each group are depicted. *p<0.05.</p
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