Respiration In Peripheral Blood Mononuclear Cells Research Articles

Mitochondrial function in peripheral blood cells across the human lifespan

Mitochondrial dysfunction is considered a hallmark of aging. Up to now, a gradual decline of mitochondrial respiration with advancing age has mainly been demonstrated in human muscle tissue. A handful of studies have examined age-related mitochondrial dysfunction in human blood cells, and only with small sample sizes and mainly in platelets. In this study, we analyzed mitochondrial respiration in peripheral blood mononuclear cells (PBMCs) and platelets from 308 individuals across the human lifespan (0–86 years). In regression analyses, with adjustment for false discovery rate (FDR), we found age-related changes in respiratory measurements to be either small or absent. The main significant changes were an age-related relative decline in complex I-linked respiration and a corresponding rise of complex II-linked respiration in PBMCs. These results add to the understanding of mitochondrial dysfunction in aging and to its possible role in immune cell and platelet senescence.

Central obesity is detrimental to anti-inflammatory, phenotype, and exhaustion markers in mononuclear cells - A cross-sectional study

To investigate the role of central obesity on immunometabolic response in peripheral blood mononuclear cells (PBMCs) from normal weight and overweight/obese young men. Eighteen individuals were classified as normal weight (NW; n=9 - age: 25±5 and BMI: 21.4±1.7) and overweight/obese (OW; n=9 - age: 29±7 and BMI: 29.2±2.7). The body composition was evaluated by dual-energy x-ray absorptiometry (DXA), waist circumference, and visceral and subcutaneous fat depots by ultrasound. Physical activity levels, metabolic parameters, immune phenotypic characterization, cytokine production by lipopolysaccharide (LPS) -stimulated whole blood cells and LPS or phorbol 12-myristate 13-acetate (PMA)-stimulated PBMC, and mitochondrial respiration in PBMCs were evaluated. Expression of AMP-activated protein kinase (AMPK), peroxisome proliferator-activated receptor gamma (PPAR-γ), nuclear factor-kappa B (NF-κB), toll-like receptor 4 (TLR-4), hypoxia-inducible factor-1 alpha (HIF-1α), and adrenergic receptor beta 1 and 2 (AR-β1 and β2) genes were evaluated in cultured PBMC using quantitative real-time polymerase chain reaction (qRT-PCR). Individuals with overweight/obese (OW) presented higher glucose (P=0.009) and leptin (P=0.010) than individuals with normal weight (NW). PBMCs of OW under stimulation with LPS presented a lower production of interleukin-10 (IL-10) (P=0.011) and macrophage inflammatory protein-1alpha (MIP-1α) (P=0.048) than NW. Mitochondrial respiration rates were not different between NW and OW subjects. Cultured PBMCs in LPS-stimulated condition indicated higher gene expression of AR-β2 in OW, while PMA-stimulated PBMCs presented lower expression of AMPK (P=0.002) and higher expression of NF-κB (P=<0.0001) than NW. OW presented higher numbers of CD3+CD4+ T cells (P=0.009) and higher expression of programmed cell death protein 1 (PD-1) in CD8+ T cells (P=0.001) than NW. Central obesity promoted reductions in interleukin 10 production response and increase in AR-β2 expressions in mitogen-stimulated PBMCs. Furthermore, central obesity altered the phenotype of PBMCs, also increasing the expression of PD-1 exhaustion markers in young adults.

245-OR: Impaired Mitochondrial Respiration in Peripheral Blood Mononuclear Cells Is Associated with Increased Obesity Classified by Waist-to-Hip Ratio in Nondiabetes EGAT Population

This study aimed to determine the impact of waist-to-hip ratio (WHR)-classified obesity and diabetes (DM) status on mitochondrial respiration in peripheral blood mononuclear cells (PBMCs) using a large cohort from the Electricity Generation Authority of Thailand (EGAT) during 2017-2018. The 1954 participants were included in this study with an average age of 67.1±7.7 years old. The mitochondrial respiration in PBMCs obtained from fresh blood samples was analyzed by the Seahorse Flux Analyzer. The participants were classified based on DM status and WHR including 1) non-DM non-obese, 2) non-DM obese, 3) DM non-obese and 4) DM obese. We found that the non-DM obese group had lower basal respiration (BR), maximal respiration, spare respiratory capacity (SRC), and proton leak (PL), compared to those of the non-DM non-obese group (Figure 1). The BR, SRC, and PL of non-DM obese group were not significantly different from those of the DM group with or without adjusting with age. All of these findings suggest that WHR-classified obesity in people with non-DM is associated with impaired mitochondrial respiration in PBMCs at a level comparable to that in people with DM. Thus, not only DM, but also obesity, may play an important role on impaired mitochondrial respiration in PBMCs. Disclosure T.Attachaipanich: None. S.C.Chattipakorn: None. S.Sriwichaiin: None. N.Apaijai: None. S.Kerdphoo: None. N.Thongmung: None. P.Vathesatogkit: None. C.Kitiyakara: None. P.Sritara: None. N.Chattipakorn: None.

Mitochondrial Respiration in Response to Iron Deficiency Anemia: Comparison of Peripheral Blood Mononuclear Cells and Liver.

Iron is an essential component for metabolic processes, including oxygen transport within hemoglobin, tricarboxylic acid (TCA) cycle activity, and mitochondrial energy transformation. Iron deficiency can thus lead to metabolic dysfunction and eventually result in iron deficiency anemia (IDA), which affects approximately 1.5 billion people worldwide. Using a rat model of IDA induced by phlebotomy, we studied the effects of IDA on mitochondrial respiration in peripheral blood mononuclear cells (PBMCs) and the liver. Furthermore, we evaluated whether the mitochondrial function evaluated by high-resolution respirometry in PBMCs reflects corresponding alterations in the liver. Surprisingly, mitochondrial respiratory capacity was increased in PBMCs from rats with IDA compared to the controls. In contrast, mitochondrial respiration remained unaffected in livers from IDA rats. Of note, citrate synthase activity indicated an increased mitochondrial density in PBMCs, whereas it remained unchanged in the liver, partly explaining the different responses of mitochondrial respiration in PBMCs and the liver. Taken together, these results indicate that mitochondrial function determined in PBMCs cannot serve as a valid surrogate for respiration in the liver. Metabolic adaptions to iron deficiency resulted in different metabolic reprogramming in the blood cells and liver tissue.

Assessment of mitochondrial respiratory capacity using minimally invasive and noninvasive techniques in persons with spinal cord injury.

Muscle biopsies are the gold standard to assess mitochondrial respiration; however, biopsies are not always a feasible approach in persons with spinal cord injury (SCI). Peripheral blood mononuclear cells (PBMCs) and near-infrared spectroscopy (NIRS) may alternatively be predictive of mitochondrial respiration. The purpose of the study was to evaluate whether mitochondrial respiration of PBMCs and NIRS are predictive of respiration of permeabilized muscle fibers after SCI. Twenty-two individuals with chronic complete and incomplete motor SCI between 18-65 years old were recruited to participate in the current trial. Using high-resolution respirometry, mitochondrial respiratory capacity was measured for PBMCs and muscle fibers of the vastus lateralis oxidizing complex I, II, and IV substrates. NIRS was used to assess mitochondrial capacity of the vastus lateralis with serial cuff occlusions and electrical stimulation. Positive relationships were observed between PBMC and permeabilized muscle fibers for mitochondrial complex IV (r = 0.86, P < 0.0001). Bland-Altman displayed agreement for complex IV (MD = 0.18, LOA = -0.86 to 1.21), between PBMCs and permeabilized muscles fibers. No significant relationships were observed between NIRS mitochondrial capacity and respiration in permeabilized muscle fibers. This is the first study to explore and support the agreement of less invasive clinical techniques for assessing mitochondrial respiratory capacity in individuals with SCI. The findings will assist in the application of PBMCs as a viable alternative for assessing mitochondrial health in persons with SCI in future clinical studies.

Lisinopril Mitigates Radiation-Induced Mitochondrial Defects in Rat Heart and Blood Cells.

The genetic bases and disparate responses to radiotherapy are poorly understood, especially for cardiotoxicity resulting from treatment of thoracic tumors. Preclinical animal models such as the Dahl salt-sensitive (SS) rat can serve as a surrogate model for salt-sensitive low renin hypertension, common to African Americans, where aldosterone contributes to hypertension-related alterations of peripheral vascular and renal vascular function. Brown Norway (BN) rats, in comparison, are a normotensive control group, while consomic SSBN6 with substitution of rat chromosome 6 (homologous to human chromosome 14) on an SS background manifests cardioprotection and mitochondrial preservation to SS rats after injury. In this study, 2 groups from each of the 3 rat strains had their hearts irradiated (8 Gy X 5 fractions). One irradiated group was treated with the ACE-inhibitor lisinopril, and a separate group in each strain served as nonirradiated controls. Radiation reduced cardiac end diastolic volume by 9-11% and increased thickness of the interventricular septum (11-16%) and left ventricular posterior wall (14-15%) in all 3 strains (5-10 rats/group) after 120 days. Lisinopril mitigated the increase in posterior wall thickness. Mitochondrial function was measured by the Seahorse Cell Mitochondrial Stress test in peripheral blood mononuclear cells (PBMC) at 90 days. Radiation did not alter mitochondrial respiration in PBMC from BN or SSBN6. However, maximal mitochondrial respiration and spare capacity were reduced by radiation in PBMC from SS rats (p=0.016 and 0.002 respectively, 9-10 rats/group) and this effect was mitigated by lisinopril (p=0.04 and 0.023 respectively, 9-10 rats/group). Taken together, these results indicate injury to the heart by radiation in all 3 strains of rats, although the SS rats had greater susceptibility for mitochondrial dysfunction. Lisinopril mitigated injury independent of genetic background.

Mitochondrial Respiration Defects in Single-Ventricle Congenital Heart Disease.

Background: Congenital heart disease (CHD) with single-ventricle (SV) physiology is now survivable with a three-stage surgical course ending with Fontan palliation. However, 10-year transplant-free survival remains at 39–50%, with ventricular dysfunction progressing to heart failure (HF) being a common sequela. For SV-CHD patients who develop HF, undergoing the surgical course would not be helpful and could even be detrimental. As HF risk cannot be predicted and metabolic defects have been observed in Ohia SV-CHD mice, we hypothesized that respiratory defects in peripheral blood mononuclear cells (PBMCs) may allow HF risk stratification in SV-CHD.Methods: SV-CHD (n = 20), biventricular CHD (BV-CHD; n = 16), or healthy control subjects (n = 22) were recruited, and PBMC oxygen consumption rate (OCR) was measured using the Seahorse Analyzer. Respiration was similarly measured in Ohia mouse heart tissue.Results: Post-Fontan SV-CHD patients with HF showed higher maximal respiratory capacity (p = 0.004) and respiratory reserve (p < 0.0001), parameters important for cell stress adaptation, while the opposite was found for those without HF (reserve p = 0.037; maximal p = 0.05). This was observed in comparison to BV-CHD or healthy controls. However, respiration did not differ between SV patients pre- and post-Fontan or between pre- or post-Fontan SV-CHD patients and BV-CHD. Reminiscent of these findings, heart tissue from Ohia mice with SV-CHD also showed higher OCR, while those without CHD showed lower OCR.Conclusion: Elevated mitochondrial respiration in PBMCs is correlated with HF in post-Fontan SV-CHD, suggesting that PBMC respiration may have utility for prognosticating HF risk in SV-CHD. Whether elevated respiration may reflect maladaptation to altered hemodynamics in SV-CHD warrants further investigation.

Altered mitochondrial metabolism in peripheral blood cells from patients with inborn errors of β-oxidation.

Inborn errors of mitochondrial fatty acid oxidation (FAO), such as medium‐chain acyl‐CoA dehydrogenase deficiency (MCAD) and very long‐chain acyl‐CoA dehydrogenase deficiency (VLCAD) affects cellular function and whole‐body metabolism. Carnitine uptake deficiency (CUD) disturbs the transportation of fatty acids into the mitochondria, but when treated is a mild disease without significant effects on FAO. For improved clinical care of VLCAD in particular, estimation of FAO severity could be important. We have investigated whether the oxygen consumption rate (OCR) of peripheral blood mononuclear cells (PBMCs) obtained from patients with MCAD, VLCAD, and CUD can be used to study cellular metabolism in patients with FAO defects and to determine the severity of FAO impairment. PBMCs were isolated from patients with VLCAD (n = 9), MCAD (n = 5–7), and CUD (n = 5). OCR was measured within 6‐hours of venous puncture using the Seahorse XFe96. The PBMCs were exposed to glucose alone or with caprylic acid (C8:0) or palmitic acid (C16:0). OCR was significantly lower in cells from patients with β‐oxidation deficiencies (MCAD and VLCAD) compared to CUD at basal conditions. When exposed to C16:0, OCR in VLCAD cells was unchanged, whereas OCR in MCAD cells increased but not to the levels observed in CUD. However, C8:0 did not increase OCR, as would be expected, in VLCAD cells. There was no clear relationship between clinical severity level and OCR. In patients with β‐oxidation deficiencies, changes of mitochondrial respiration in PBMCs are detectable, which indicate that PBMCs have translational potential for studies of β‐oxidation defects. However, further studies are warranted.

Mitochondrial respiration and redox protein expression in peripheral blood mononuclear cells from Non‐Hispanic Black and White Males

Background Non-Hispanic Black (NHB) adults exhibit a higher prevalence of cardiovascular disease (CVD) compared to Non-Hispanic White (NHW) adults. Vascular oxidative stress may be one contributing factor in these racial disparities. Peripheral blood mononuclear cells (PBMCs) are primary contributors to systemic oxidative stress and there are documented racial disparities in the PBMC-derived reactive oxygen species (ROS) production. However, it is unclear if the racial differences in PBMC-derived ROS is associated with alterations in mitochondrial function. Therefore, the purpose of this study was to investigate potential racial differences in cellular metabolism and redox protein expression in PBMCs from NHW and NHB adults. Methods Commercial PBMCs (n=10 cell lines; 5 NHW & 5 NHB males) were cultured 14-18 hours prior to cell respiration and protein harvest. Basal and maximal respiration were measured using a Clark electrode (Oxytherm, Hanstech) and then calculated coupling efficiency and leak respiration. Western blotting was used to measure protein expression for the antioxidant proteins: superoxide dismutase (SOD) isoforms 1 and 2, as well as the acetylated (i.e., inactivated) SOD2 (AcSOD2) and the NAD-dependent deacetylase sirtuin-3 (Sirt3). The pro-oxidant complex NADPH oxidase subunits p47phox and gp91phox were also assessed. Welch unpaired t-tests were used to assess the racial differences in respiration measures and independent t-tests were used for protein expression. Results For the respiration measures, NHW had significantly higher levels of basal respiration (Fig. 1A, p=0.048) and leak respiration (Fig. 1D, p=0.016), while having a lower coupling efficiency (Fig. 1C, p=0.003). There was no difference in maximal respiration (Fig. 1B, p=0.188). No baseline racial differences were found in PBMC expression for the redox regulating proteins: SOD1 (Fig. 2A, p=0.42), SOD2 (Fig. 2B, p=0.40), AcSOD2 (Fig. 2C, p=0.45), Sirt3 (Fig. 2D, p=0.45), p47phox (Fig 3A, p=0.23), or gp91phox (Fig. 3B, p=0.86). Conclusion The lower coupling efficiency and higher leak respiration in PBMCs from NHW may be beneficial in decreasing ROS production and overall oxidative damage. While we found no racial differences in the redox regulating proteins, additional data are needed, particularly in PBMCs from female participants.

Cardiometabolic risk factors are associated with immune cell mitochondrial respiration in humans.

Modifiable cardiometabolic risk factors induce the release of proinflammatory cytokines and reactive oxygen species from circulating peripheral blood mononuclear cells (PBMCs), resulting in increased cardiovascular disease risk and compromised immune health. These changes may be driven by metabolic reprogramming of PBMCs, resulting in reduced mitochondrial respiration; however, this has not been fully tested. We aimed to determine the independent associations between cardiometabolic risk factors including BMI, blood pressure, fasting glucose, and plasma lipids with mitochondrial respiration in PBMCs isolated from generally healthy individuals (n = 21) across the adult lifespan (12 men/9 women; age, 56 ± 21 yr; age range, 22-78 yr; body mass index, 27.9 ± 5.7 kg/m2; blood pressure, 123 ± 16/72 ± 10 mmHg; glucose, 90 ± 14 mg/dL; low-density lipoprotein cholesterol (LDL-C), 111 ± 22 mg/dL; and high-density lipoprotein cholesterol (HDL-C), 62 ± 16 mg/dL). PBMCs were isolated from whole blood by density-dependent centrifugation and used to assess mitochondrial function by respirometry. Primary outcomes included basal and maximal oxygen consumption rate (OCR), which were subsequently used to determine spare respiratory capacity and OCR metabolic potential. After we corrected for systolic blood pressure (SBP), diastolic blood pressure (DBP), and blood glucose, LDL-C was negatively associated with maximal respiration (r = -0.56, P = 0.016), spare respiratory capacity (r = -0.58, P = 0.012), and OCR metabolic potential (r = -0.71, P = 0.0011). In addition, SBP was negatively associated with OCR metabolic potential (r = -0.62, P = 0.0056) after we corrected for DBP, blood glucose, and LDL-C. These data suggest a link between blood cholesterol, SBP, and mitochondrial health that may provide insight into how cardiometabolic risk factors contribute to impaired immune cell function.NEW & NOTEWORTHY Independent of other cardiometabolic risk factors, low-density lipoprotein cholesterol, and systolic blood pressure were found to be negatively associated with several parameters of mitochondrial respiration in peripheral blood mononuclear cells of healthy adults. These data suggest that low-density lipoprotein cholesterol and systolic blood pressure may induce metabolic reprogramming of immune cells, contributing to increased cardiovascular disease risk and impaired immune health.

Low-intensity exercise stimulates bioenergetics and increases fat oxidation in mitochondria of blood mononuclear cells from sedentary adults.

AimExercise training induces adaptations in muscle and other tissue mitochondrial metabolism, dynamics, and oxidative phosphorylation capacity. Mitochondrial fatty acid oxidation was shown to be pivotal for the anti‐inflammatory status of immune cells. We hypothesize that exercise training can exert effects influence mitochondrial fatty acid metabolism in peripheral blood mononuclear cells (PBMCs). The aim was to investigate the effect of exercise on the fatty acid oxidation‐dependent respiration in PBMCs.DesignTwelve fasted or fed volunteers first performed incremental‐load exercise tests to exhaustion on a cycle ergometer to determine the optimal workload ensuring maximal health benefits in volunteers with a sedentary lifestyle. In addition, the same volunteers performed 60 min of low‐intensity constant‐load exercise.ResultsIn the incremental‐load exercise, the maximal whole‐body fat oxidation rate measured by indirect calorimetry was reached at the fasted state already at a 50 W workload. At the 75–175 W workloads, the contribution of fat oxidation significantly decreased to only 11%, the heart rate increased to 185 BPM, and the study participants reached exhaustion. These results show that low‐intensity exercise (50W) is optimal for maximal whole‐body fat utilization. After low‐intensity exercise, the ROUTINE mitochondrial respiration, as well as fatty acid oxidation‐dependent respiration in PBMCs at LEAK and OXPHOS states, were significantly increased by 31%, 65%, and 76%, respectively. In addition, during 60 min of low‐intensity (50W) exercise, a 2‐fold higher lipolysis rate was observed and 13.5 ± 0.9 g of fat was metabolized, which was 57% more than the amount of fat that was metabolized during the incremental‐load exercise.ConclusionsIn individuals with a sedentary lifestyle participating in a bicycle ergometry exercise program, maximal lipolysis and whole‐body fat oxidation rate is reached in a fasted state during low‐intensity exercise. For the first time, it was demonstrated that low‐intensity exercise improves bioenergetics and increases fatty acid oxidation in PBMCs and may contribute to the anti‐inflammatory phenotype.

Mitochondrial Dysfunction is Associated With an Immune Paralysis Phenotype in Pediatric Sepsis.

Immune dysregulation is a defining feature of sepsis, but the role for mitochondria in the development of immunoparalysis in pediatric sepsis is not known. We sought to determine if mitochondrial dysfunction measured in peripheral blood mononuclear cells (PBMCs) is associated with immunoparalysis and systemic inflammation in children with sepsis. Prospective observational study. Single-academic pediatric intensive care unit (PICU). One hundred sixty-one children with sepsis/septic shock and 18 noninfected PICU controls. Mitochondrial respiration in PBMCs, markers of immune function, and plasma cytokines were measured on days 1 to 2 (T1), 3 to 5 (T2), and 8 to 14 (T3) after sepsis recognition, and once for controls. Immunoparalysis was defined as whole-blood ex vivo lipopolysaccharide-induced tumor necrosis factor-alpha (TNF-α) ≤200 pg/mL or monocyte human leukocyte antigen-DR ≤30%. Mitochondrial respiration was lower in children with versus without immunoparalysis measured at the same timepoint. Mitochondrial respiration measured early (at T1 and T2) was also lower in those with immunoparalysis at T2 and T3, respectively. Although most patients with immunoparalysis exhibited low mitochondrial respiration, this metabolic finding was not specific to the immunoparalysis phenotype. Plasma cytokines, including IL-8, IL-10, TNF-α, and MCP-1, were highest in the subset of sepsis patients with immune paralysis or low mitochondrial respiration at T1. Children with sepsis had lower PBMC mitochondrial respiration when immunoparalysis was present compared with those without immunoparalysis. The subsets with immune paralysis and low mitochondrial respiration exhibited the highest levels of systemic inflammation.

Autistic Siblings with Novel Mutations in Two Different Genes: Insight for Genetic Workups of Autistic Siblings and Connection to Mitochondrial Dysfunction.

The prevalence of autism spectrum disorder (ASD) is high, yet the etiology of this disorder is still uncertain. Advancements in genetic analysis have provided the ability to identify potential genetic changes that may contribute to ASD. Interestingly, several genetic syndromes have been linked to metabolic dysfunction, suggesting an avenue for treatment. In this case study, we report siblings with ASD who had similar initial phenotypic presentations. Whole exome sequencing (WES) revealed a novel c.795delT mutation in the WDR45 gene affecting the girl, which was consistent with her eventual progression to a Rett-like syndrome phenotype including seizures along with a stereotypical cyclic breathing pattern. Interestingly, WES identified that the brother harbored a novel heterozygous Y1546H variant in the DEP domain-containing protein 5 (DEPDC5) gene, consistent with his presentation. Both siblings underwent a metabolic workup that demonstrated different patterns of mitochondrial dysfunction. The girl demonstrated statistically significant elevations in mitochondrial activity of complex I + III in both muscle and fibroblasts and increased respiration in peripheral blood mononuclear cells (PBMCs) on Seahorse Extracellular Flux analysis. The boy demonstrates a statistically significant decrease in complex IV activity in buccal epithelium and decreased respiration in PBMCs. These cases highlight the differences in genetic abnormalities even in siblings with ASD phenotypes as well as highlights the individual role of novel mutations in the WDR45 and DEPDC5 genes. These cases demonstrate the importance of advanced genetic testing combined with metabolic evaluations in the workup of children with ASD.

Mitochondrial respiration in peripheral blood mononuclear cells correlates with depressive subsymptoms and severity of major depression.

Mitochondrial dysfunction might have a central role in the pathophysiology of depression. Phenotypically, depression is characterized by lack of energy, concentration problems and fatigue. These symptoms might be partially explained by reduced availability of adenosine triphosphate (ATP) as a consequence of impaired mitochondrial functioning. This study investigated mitochondrial respiration in peripheral blood mononuclear cells (PBMCs), an established model to investigate the pathophysiology of depression. Mitochondrial respiration was assessed in intact PBMCs in 22 individuals with a diagnosis of major depression (MD) compared with 22 healthy age-matched controls using high-resolution respirometry. Individuals with MD showed significantly impaired mitochondrial functioning: routine and uncoupled respiration as well as spare respiratory capacity, coupling efficiency and ATP turnover-related respiration were significantly lower in the MD compared with the control group. Furthermore, mitochondrial respiration was significantly negatively correlated with the severity of depressive symptoms, in particular, with loss of energy, difficulties concentrating and fatigue. The results suggest that mitochondrial dysfunction contributes to the biomolecular pathophysiology of depressive symptoms. The decreased immune capability observed in MD leading to a higher risk of comorbidities could be attributable to impaired energy supply due to mitochondrial dysfunction. Thus mitochondrial respiration in PBMCs and its functional consequences might be an interesting target for new therapeutical approaches in the treatment of MD and immune-related comorbidities.

Mitochondrial dysfunction during sepsis: Still more questions than answers*

Mitochondrial dysfunction during sepsis: Still more questions than answers*