Mitochondrial phenotypes in purified human immune cell subtypes and cell mixtures

Abstract

Using a high-throughput mitochondrial phenotyping platform to quantify multiple mitochondrial features among molecularly-defined immune cell subtypes, we quantify the natural variation in citrate synthase, mitochondrial DNA copy number (mtDNAcn), and respiratory chain enzymatic activities in human neutrophils, monocytes, B cells, and naïve and memory T lymphocyte subtypes. In mixed peripheral blood mononuclear cells (PBMCs) from the same individuals, we show to what extent mitochondrial measures are confounded by both cell type distributions and contaminating platelets. Cell subtype-specific measures among women and men spanning 4 decades of life indicate potential age- and sex-related differences, including an age-related elevation in mtDNAcn, which are masked or blunted in mixed PBMCs. Finally, a proof-of-concept, repeated-measures study in a single individual validates cell type differences and also reveals week-to-week changes in mitochondrial activities. Larger studies are required to validate and mechanistically extend these findings. These mitochondrial phenotyping data build upon established immunometabolic differences among leukocyte sub-populations, and provide foundational quantitative knowledge to develop interpretable blood-based assays of mitochondrial health.

Extracellular flux analyses reveal differences in mitochondrial PBMC metabolism between high-fit and low-fit females

“…Differences in sampling time might underlie these different observations, as Liepinsh et al ( 25 ) sampled directly after exercise while we sampled 21 h after exercise. Since PBMC subsets are metabolically different ( 17 , 18 ) and substantial shifts in PBMC subsets have been described after acute exercise ( 27 – 29 ), the increased mitochondrial PBMC function as observed by Liepinsh et al could possibly be related to large differences in analyzed PBMC subsets between the two timepoints, whereas in our study PBMC composition was similar at baseline and after the recent bout of exercise. Second, differences in the metabolic substrates provided to the PBMCs during metabolic analysis might play a role.…”

“…The CD3 + T cells are composed of CD4 + and CD8 + T cells, roughly in a 2:1 ratio. Different PBMC subsets do not only provoke different immune responses but they can also display distinct metabolic states ( 16 – 18 ). Importantly, apart from providing energy substrates and metabolic building blocks for immune cell proliferation and synthesis of macromolecules (e.g., cytokines), metabolic pathways have recently been shown to not only act as a consequence but also as driver of immune cell differentiation ( 16 ), which further increases the relevance of monitoring cellular metabolism as a biomarker for health.…”

“…Since a single exercise bout often elicits a transient proinflammatory response whereas regular bouts of exercise induce an anti-inflammatory state ( 6 , 26 ), the impact of regular physical activity (long-term effect) and a single-exercise session (short-term effect) on PBMC metabolism could differ, yet this has not been studied in detail. Furthermore, it is not yet clear whether the metabolic responses in PBMCs upon these exercise interventions are primarily related to alterations in cellular energy metabolism per se, or that these responses reflect changes in PBMC composition, since changes in PBMC subset frequencies have been demonstrated in response to exercise ( 27 – 29 ), and PBMC subsets are not only immunologically but also metabolically distinct ( 17 , 18 ). Better understanding of the variation in PBMC subsets and overall PBMC composition between healthy donors and their contribution to the overall metabolic outcomes in PBMCs is therefore also needed.…”

Personality traits are consistently associated with blood mitochondrial DNA copy number estimated from genome sequences in two genetic cohort studies

“…It should also be noted that although some past studies assayed for mtDNAcn from whole blood, they did not adjust for the effect of platelets, white blood cells, and other leukocytes subgroups on mtDNAcn as we do in this study. mtDNAcn estimates differ amongst the various blood cell types (62), and this significantly impacts whole-blood mtDNAcn estimates and should be adjusted for in association models (also see Discussion below). mtDNAcn and personality are both predictors of mortality (25,36,63), thus making our finding of an association between the two even more intriguing.…”

Influence of Immune Cell Subtypes on Mitochondrial Measurements in Peripheral Blood Mononuclear Cells From Children with Sepsis

“…A recent study of healthy adults demonstrated that mitochondrial phenotypes measured in PBMCs are confounded by variable composition of immune cell subtypes and urged caution in using PBMCs to assess mitochondrial changes within and between individuals (20). However, there are no data about the influence of immune cell subtypes on mitochondrial phenotypes measured in PBMCs from children.…”