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

Abstract

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.

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

“…We isolated PBMCs and granulocytes from whole blood samples to detect changes in immunometabolism in response to combined ASDH and HS and hyperoxemia as previously described ( 20 , 25 ). Blood was drawn at three different time points over the course of the experiment: T1: after (neuro)surgical instrumentation following a stabilization period; T2: 24 h after ASDH and HS, i.e.…”

“…Mitochondrial respiration of isolated cells was derived from the measurement of O 2 consumption with high resolution respirometry. We used the Oroboros-2K (Oroboros Instruments, Innsbruck, Austria) for this purpose, which is a device for simultaneous recording of the O 2 concentration in two parallel chambers ( 20 , 25 ). Briefly, chambers were calibrated with 2 mL of mitochondrial respiration medium MiR05 (composition available in Supplementary Table 1 ) adjusted to pH 7.1 with KOH and equilibrated with 21% O 2 at 37°C.…”

“…Immediately after arterial blood withdrawal at the same time points as for cell isolation, 25 µL of whole blood were mixed 1:1 (v/v) with CMH spin probe solution containing 400 µM CMH spin probe (1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine), 25 µM deferoxamine, and 5 µM diethyldithiocarbamate solved in Krebs-HEPES-Buffer (KHB) (Noxygen, Elzach, Germany). As previously described ( 20 , 25 ), the mixture was transferred to a 50 µL glass capillary, sealed, and placed in an EMXnano ESR spectrometer (Bruker, Billerica, MA, USA). After a 5 min incubation period at 37°C, the measurement was started with device settings detailed in the Supplementary File .…”

Bayesian 13C-Metabolic Flux Analysis of Parallel Tracer Experiments in Granulocytes: A Directional Shift within the Non-Oxidative Pentose Phosphate Pathway Supports Phagocytosis

“…1559, approval 29 October 2021) and in compliance with the National Institute of Health Guidelines on the Use of Laboratory Animals and the European Union’s “Directive 2010/63/EU on the protection of animals used for scientific purposes”. Arterial blood from adult, human-sized German landrace swine was collected immediately after anesthesia and instrumentation as described in detail by Münz et al [ 22 ]. Directly after sampling, granulocytes were isolated from the lithium–heparin blood using Ficoll density centrifugation.…”

Ex Vivo 13C-Metabolic Flux Analysis of Porcine Circulating Immune Cells Reveals Cell Type-Specific Metabolic Patterns and Sex Differences in the Pentose Phosphate Pathway

“…1559, approval 29 October 2021) and is in compliance with the National Institute of Health Guidelines on the Use of Laboratory Animals and the European Union "Directive 2010/63/EU on the protection of animals used for scientific purposes". According to the 3R principle (replacement, reduction, and refinement in animal research), the present study is a post hoc analysis of material available from animal experiments recently described by Münz et al [35]. A total of 12 healthy, human-sized, sexually mature German landrace pigs (range body weight 63-102 kg) of either sex (each n = 6) between the ages of 21 and 27 weeks had been included in that study.…”