Bioenergetic analysis of human peripheral blood mononuclear cells

Human leucocytes perform energy-demanding tasks such as cytokine production and migration. These processes rely heavily on glycolysis to supply ATP quickly. While animal studies have explored this, human data remains limited. Researchers have long known that glycolysis supports immune responses, but its role in human peripheral blood mononuclear cells (MNCs) is less clear. This gap motivated a study to assess the bioenergetic profiles of human MNCs. The study aimed to determine how glycolysis and oxidative phosphorylation contribute to immune function in humans. Cryopreservation is widely used in translational research, but its impact on bioenergetic measurements is uncertain. This uncertainty drove the investigation into whether cryopreserved MNCs can reliably reflect bioenergetic health. No prior work had resolved the differences between fresh and cryopreserved MNCs in this context.

The study aimed to clarify the roles of glycolysis and oxidative phosphorylation in human MNCs. It focused on how these energy systems support cytokine production during immune activation. The researchers wanted to determine if cryopreserved MNCs are suitable for bioenergetic analysis. They also sought to assess the translational potential of bioenergetic profiling in clinical settings. The study compared fresh and cryopreserved MNCs from the same donors. It tested the impact of cryopreservation on glycolytic and oxidative phosphorylation profiles. The researchers examined cytokine responses to lipopolysaccharide (LPS) stimulation. They aimed to establish a reliable method for measuring bioenergetic health in human immune cells.

The study used human peripheral blood mononuclear cells (MNCs) obtained from donors. Both fresh and cryopreserved MNCs were analyzed for bioenergetic profiles. The researchers optimized cell numbers and concentrations of activators and inhibitors. They measured glycolysis and oxidative phosphorylation using standardized protocols. Cytokine production was assessed after LPS stimulation. 2-deoxy-D-glucose was used to inhibit glycolysis and test its role in cytokine release. The team compared results from fresh and resuscitated MNCs from the same individuals. They also evaluated bioenergetic differences between pregnant and non-pregnant women.

Glycolysis was essential for LPS-induced cytokine production in MNCs. Inhibition of glycolysis with 2-deoxy-D-glucose significantly reduced IL-1β, IL-6, and TNF-α levels. Fresh and resuscitated MNCs showed similar cytokine responses to LPS. However, cryopreservation significantly altered oxidative phosphorylation measurements. Some glycolytic parameters were also affected by cryopreservation. The bioenergetic health index was higher in pregnant women compared to non-pregnant women. Basal glycolysis and glycolytic capacity were lower in pregnant individuals. These findings suggest that cryopreserved MNCs may not accurately reflect bioenergetic health.

The study shows that glycolysis supports cytokine production in human MNCs. Cryopreserved MNCs are not suitable for bioenergetic analysis due to altered profiles. The bioenergetic health index can be measured in human MNCs using fresh samples. Pregnancy increases the bioenergetic health index but reduces glycolytic capacity. These findings highlight the importance of using fresh cells for accurate measurements. The study confirms the translational potential of bioenergetic profiling in immune research. More detailed analysis of specific leucocyte populations is needed. The researchers propose that discrete cell types may have distinct bioenergetic roles.