Porcine blood cell and brain tissue energy metabolism: Effects of “early life stress”

Background: Early Life Stress (ELS) may exert long-lasting biological effects, e.g., on PBMC energy metabolism and mitochondrial respiration. Data on its effect on brain tissue mitochondrial respiration is scarce, and it is unclear whether blood cell mitochondrial activity mirrors that of brain tissue. This study investigated blood immune cell and brain tissue mitochondrial respiratory activity in a porcine ELS model.Methods: This prospective randomized, controlled, animal investigation comprised 12 German Large White swine of either sex, which were weaned at PND (postnatal day) 28–35 (control) or PND21 (ELS). At 20–24 weeks, animals were anesthetized, mechanically ventilated and surgically instrumented. We determined serum hormone, cytokine, and “brain injury marker” levels, superoxide anion (O2•¯) formation and mitochondrial respiration in isolated immune cells and immediate post mortem frontal cortex brain tissue.Results: ELS animals presented with higher glucose levels, lower mean arterial pressure. Most determined serum factors did not differ. In male controls, TNFα and IL-10 levels were both higher than in female controls as well as, no matter the gender in ELS animals. MAP-2, GFAP, and NSE were also higher in male controls than in the other three groups. Neither PBMC routine respiration and brain tissue oxidative phosphorylation nor maximal electron transfer capacity in the uncoupled state (ETC) showed any difference between ELS and controls. There was no significant relation between brain tissue and PBMC, ETC, or brain tissue, ETC, and PBMC bioenergetic health index. Whole blood O2•¯ concentrations and PBMC O2•¯ production were comparable between groups. However, granulocyte O2•¯ production after stimulation with E. coli was lower in the ELS group, and this effect was sex-specific: increased O2•¯ production increased upon stimulation in all control animals, which was abolished in the female ELS swine.Conclusion: This study provides evidence that ELS i) may, gender-specifically, affect the immune response to general anesthesia as well as O2•¯ radical production at sexual maturity, ii) has limited effects on brain and peripheral blood immune cell mitochondrial respiratory activity, and iii) mitochondrial respiratory activity of peripheral blood immune cells and brain tissue do not correlate

Cystathionine-γ-lyase expression is associated with mitochondrial respiration during sepsis-induced acute kidney injury in swine with atherosclerosis

Sepsis is associated with disturbed glucose metabolism and reduced mitochondrial activity and biogenesis, ultimately leading to multiple organ dysfunction, e.g., acute kidney injury (AKI). Cystathionine-γ-lyase (CSE), the major cardiovascular source of endogenous H2S release, is implicated in the regulation of glucose metabolism and mitochondrial activity through a PGC1α-dependent mechanism, and critical for kidney function. Atherosclerosis is associated with mitochondrial dysfunction and reduced CSE expression. Thus, the aim of this post hoc study was to test the hypothesis whether there is an interplay between CSE expression and kidney dysfunction, mitochondrial activity, and oxidative/nitrosative stress in porcine septic AKI with underlying coronary artery disease.Methods: This study is a post hoc analysis of material from anesthetized and instrumented swine with a high fat diet-induced hypercholesterolemia and atherosclerosis undergoing faecal peritonitis-induced septic shock or sham procedure and intensive care (comprising fluid resuscitation and continuous i.v. noradrenaline (NoA) infusion) for 24 h. Glucose metabolism was quantified from blood 13C6-glucose and expiratory 13CO2/12CO2 isotope enrichment during 13C6-glucose infusion. Mitochondrial activity was determined by high- resolution respirometry. CSE and PGC1α expression, as well as nitrotyrosine formation and albumin extravasation, were quantified by immunohistochemistry of formalin-fixed kidney paraffin sections.Results: Sepsis was associated with lactic acidosis (p = 0.004) and AKI (50% fall of creatinine clearance (CrCl), p = 0.019). While both whole-body glucose production (p = 0.004) and oxidation (p = 0.006) were increased, kidney tissue mitochondrial respiration was reduced (p = 0.028), coinciding with decreased CSE (p = 0.003) and PGC1α (p = 0.003) expression. Albumin extravasation (p = 0.011) and nitrotyrosine formation (p = 0.008) were increased in septic kidneys.Conclusions: Sepsis-induced AKI is associated with disturbed mitochondrial respiration and biogenesis, which may be aggravated by oxidative and nitrosative stress. Our results confirm previous data in murine septic shock and porcine hemorrhage and resuscitation on the crucial role of CSE for barrier integrity and kidney functio

Interaction of the hydrogen sulfide system with the oxytocin system in the injured mouse heart

Both the hydrogen sulfide/cystathionine-γ-lyase (H2S/CSE) and oxytocin/oxytocin receptor (OT/OTR) systems have been reported to be cardioprotective. H2S can stimulate OT release, thereby affecting blood volume and pressure regulation. Systemic hyper-inflammation after blunt chest trauma is enhanced in cigarette smoke (CS)-exposed CSE−/− mice compared to wildtype (WT). CS increases myometrial OTR expression, but to this point, no data are available on the effects CS exposure on the cardiac OT/OTR system. Since a contusion of the thorax (Txt) can cause myocardial injury, the aim of this post hoc study was to investigate the effects of CSE−/− and exogenous administration of GYY4137 (a slow release H2S releasing compound) on OTR expression in the heart, after acute on chronic disease, of CS exposed mice undergoing Txt.Methods: This study is a post hoc analysis of material obtained in wild type (WT) homozygous CSE−/− mice after 2-3 weeks of CS exposure and subsequent anesthesia, blast wave-induced TxT, and surgical instrumentation for mechanical ventilation (MV) and hemodynamic monitoring. CSE−/− animals received a 50 μg/g GYY4137-bolus after TxT. After 4h of MV, animals were exsanguinated and organs were harvested. The heart was cut transversally, formalin-fixed, and paraffin- embedded. Immunohistochemistry for OTR, arginine-vasopressin-receptor (AVPR), and vascular endothelial growth factor (VEGF) was performed with naïve animals as native controls.Results: CSE−/− was associated with hypertension and lower blood glucose levels, partially and significantly restored by GYY4137 treatment, respectively. Myocardial OTR expression was reduced upon injury, and this was aggravated in CSE−/−. Exogenous H2S administration restored myocardial protein expression to WT levels.Conclusions: This study suggests that cardiac CSE regulates cardiac OTR expression, and this effect might play a role in the regulation of cardiovascular function

An exploratory study investigating the effect of targeted hyperoxemia in a randomized controlled trial in a long-term resuscitated model of combined acute subdural hematoma and hemorrhagic shock in cardiovascular healthy pigs

Severe physical injuries and associated traumatic brain injury and/or hemorrhagic shock (HS) remain leading causes of death worldwide, aggravated by accompanying extensive inflammation. Retrospective clinical data indicated an association between mild hyperoxemia and improved survival and outcome. However, corresponding prospective clinical data, including long-term resuscutation, are scarce. Therefore, the present study explored the effect of mild hyperoxemia for 24 hours in a prospective randomized controlled trial in a long-term resuscitated model of combined acute subdural hematoma (ASDH) and HS. ASDH was induced by injecting 0.1 ml × kg−1 autologous blood into the subdural space and HS was triggered by passive removal of blood. After 2 hours, the animals received full resuscitation, including retransfusion of the shed blood and vasopressor support. During the first 24 hours, the animals underwent targeted hyperoxemia (PaO2 = 200 – 250 mmHg) or normoxemia (PaO2 = 80 – 120 mmHg) with a total observation period of 55 hours after the initiation of ASDH and HS. Survival, cardiocirculatory stability, and demand for vasopressor support were comparable between both groups. Likewise, humoral markers of brain injury and systemic inflammation were similar. Multimodal brain monitoring, including microdialysis and partial pressure of O2 in brain tissue, did not show significant differences either, despite a significantly better outcome regarding the modified Glasgow Coma Scale 24 hours after shock that favors hyperoxemia. In summary, the present study reports no deleterious and few beneficial effects of mild targeted hyperoxemia in a clinically relevant model of ASDH and HS with long-term resuscitation in otherwise healthy pigs. Further beneficial effects on neurological function were probably missed due to the high mortality in both experimental groups. The present study remains exploratory due to the unavailability of an a priori power calculation resulting from the lack of necessary data

The effect of sodium thiosulfate on immune cell metabolism during porcine hemorrhage and resuscitation

Introduction Sodium thiosulfate (Na2S2O3), an H2S releasing agent, was shown to be organ-protective in experimental hemorrhage. Systemic inflammation activates immune cells, which in turn show cell type-specific metabolic plasticity with modifications of mitochondrial respiratory activity. Since H2S can dose-dependently stimulate or inhibit mitochondrial respiration, we investigated the effect of Na2S2O3 on immune cell metabolism in a blinded, randomized, controlled, long-term, porcine model of hemorrhage and resuscitation. For this purpose, we developed a Bayesian sampling-based model for 13C isotope metabolic flux analysis (MFA) utilizing 1,2-13C2-labeled glucose, 13C6-labeled glucose, and 13C5-labeled glutamine tracers. Methods After 3 h of hemorrhage, anesthetized and surgically instrumented swine underwent resuscitation up to a maximum of 68 h. At 2 h of shock, animals randomly received vehicle or Na2S2O3 (25 mg/kg/h for 2 h, thereafter 100 mg/kg/h until 24 h after shock). At three time points (prior to shock, 24 h post shock and 64 h post shock) peripheral blood mononuclear cells (PBMCs) and granulocytes were isolated from whole blood, and cells were investigated regarding mitochondrial oxygen consumption (high resolution respirometry), reactive oxygen species production (electron spin resonance) and fluxes within the metabolic network (stable isotope-based MFA). Results PBMCs showed significantly higher mitochondrial O2 uptake and lower O 2 • − production in comparison to granulocytes. We found that in response to Na2S2O3 administration, PBMCs but not granulocytes had an increased mitochondrial oxygen consumption combined with a transient reduction of the citrate synthase flux and an increase of acetyl-CoA channeled into other compartments, e.g., for lipid biogenesis. Conclusion In a porcine model of hemorrhage and resuscitation, Na2S2O3 administration led to increased mitochondrial oxygen consumption combined with stimulation of lipid biogenesis in PBMCs. In contrast, granulocytes remained unaffected. Granulocytes, on the other hand, remained unaffected. O 2 • − concentration in whole blood remained constant during shock and resuscitation, indicating a sufficient anti-oxidative capacity. Overall, our MFA model seems to be is a promising approach for investigating immunometabolism; especially when combined with complementary methods

The effect of targeted hyperoxemia in a randomized controlled trial employing a long-term resuscitated, model of combined acute subdural hematoma and hemorrhagic shock in swine with coronary artery disease: An exploratory, hypothesis-generating study

Controversial evidence is available regarding suitable targets for the arterial O2 tension (PaO2) after traumatic brain injury and/or hemorrhagic shock (HS). We previously demonstrated that hyperoxia during resuscitation from hemorrhagic shock attenuated cardiac injury and renal dysfunction in swine with coronary artery disease. Therefore, this study investigated the impact of targeted hyperoxemia in a long-term, resuscitated model of combined acute subdural hematoma (ASDH)-induced brain injury and HS. The prospective randomized, controlled, resuscitated animal investigation consisted of 15 adult pigs. Combined ASDH plus HS was induced by injection of 0.1 ml/kg autologous blood into the subdural space followed by controlled passive removal of blood. Two hours later, resuscitation was initiated comprising re-transfusion of shed blood, fluids, continuous i.v. noradrenaline, and either hyperoxemia (target PaO2 200 – 250 mmHg) or normoxemia (target PaO2 80 – 120 mmHg) during the first 24 h of the total of 54 h of intensive care. Systemic hemodynamics, intracranial and cerebral perfusion pressures, parameters of brain microdialysis and blood biomarkers of brain injury did not significantly differ between the two groups. According to the experimental protocol, PaO2 was significantly higher in the hyperoxemia group at the end of the intervention period, i.e., at 24 h of resuscitation, which coincided with a higher brain tissue PO2. The latter persisted until the end of observation period. While neurological function as assessed using the veterinary Modified Glasgow Coma Score progressively deteriorated in the control group, it remained unaffected in the hyperoxemia animals, however, without significant intergroup difference. Survival times did not significantly differ in the hyperoxemia and control groups either. Despite being associated with higher brain tissue PO2 levels, which were sustained beyond the intervention period, targeted hyperoxemia exerted neither significantly beneficial nor deleterious effects after combined ASDH and HS in swine with pre-existing coronary artery disease. The unavailability of a power calculation and, thus, the limited number of animals included, are the limitations of the study

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

H2S in Critical Illness—A New Horizon for Sodium Thiosulfate?

Ever since the discovery of endogenous H2S and the identification of its cytoprotective properties, efforts have been made to develop strategies to use H2S as a therapeutic agent. The ability of H2S to regulate vascular tone, inflammation, oxidative stress, and apoptosis might be particularly useful in the therapeutic management of critical illness. However, neither the inhalation of gaseous H2S, nor the administration of inorganic H2S-releasing salts or slow-releasing H2S-donors are feasible for clinical use. Na2S2O3 is a clinically approved compound with a good safety profile and is able to release H2S, in particular under hypoxic conditions. Pre-clinical studies show promise for Na2S2O3 in the acute management of critical illness. A current clinical trial is investigating the therapeutic potential for Na2S2O3 in myocardial infarct. Pre-eclampsia and COVID-19 pneumonia might be relevant targets for future clinical trials