Immunometabolic adaptation in monocytes underpins functional changes during pregnancy.

Metabolic heterogeneity is a determinant of immune cell function. The normal physiological metabolic reprogramming of pregnancy that ensures the fuel requirements of mother and baby are met, might also underpin changes in immunity that occur with pregnancy and manifest as altered responses to pathogens and changes to autoimmune disease symptoms. Using peripheral blood from pregnant women at term, we reveal that monocytes lose M2-like and gain M1-like properties accompanied by reductions in mitochondrial mass, maximal respiration, and cardiolipin content in pregnancy; glycolysis is unperturbed. We establish that muramyl dipeptide (MDP)-stimulated cytokine production relies on oxidative metabolism, then show in pregnancy reduced cytokine production in response to MDP but not LPS. Overall, mitochondrially centered metabolic capabilities of late gestation monocytes are down-regulated revealing natural plasticity in monocyte phenotype and function that could reveal targets for improving pregnancy outcomes but also yield alternative therapeutic approaches to diverse metabolic and/or immune-mediated diseases beyond pregnancy.

Loss of mitochondrial pyruvate carrier 1 supports proline-dependent proliferation and collagen biosynthesis in ovarian cancer.

The pyruvate transporter MPC1 (mitochondrial pyruvate carrier 1) acts as a tumour-suppressor, loss of which correlates with a pro-tumorigenic phenotype and poor survival in several tumour types. In high-grade serous ovarian cancers (HGSOC), patients display copy number loss of MPC1 in around 78% of cases and reduced MPC1 mRNA expression. To explore the metabolic effect of reduced expression, we demonstrate that depleting MPC1 in HGSOC cell lines drives expression of key proline biosynthetic genes; PYCR1, PYCR2 and PYCR3, and biosynthesis of proline. We show that altered proline metabolism underpins cancer cell proliferation, reactive oxygen species (ROS) production, and type I and type VI collagen formation in ovarian cancer cells. Furthermore, exploring The Cancer Genome Atlas, we discovered the PYCR3 isozyme to be highly expressed in a third of HGSOC patients, which was associated with more aggressive disease and diagnosis at a younger age. Taken together, our study highlights that targeting proline metabolism is a potential therapeutic avenue for the treatment of HGSOC.

Canagliflozin impairs T cell effector function via metabolic suppression in autoimmunity.

Augmented T cell function leading to host damage in autoimmunity is supported by metabolic dysregulation, making targeting immunometabolism an attractive therapeutic avenue. Canagliflozin, a type 2 diabetes drug, is a sodium glucose co-transporter 2 (SGLT2) inhibitor with known off-target effects on glutamate dehydrogenase and complex I. However, the effects of SGLT2 inhibitors on human T cell function have not been extensively explored. Here, we show that canagliflozin-treated T cells are compromised in their ability to activate, proliferate, and initiate effector functions. Canagliflozin inhibits T cell receptor signaling, impacting on ERK and mTORC1 activity, concomitantly associated with reduced c-Myc. Compromised c-Myc levels were encapsulated by a failure to engage translational machinery resulting in impaired metabolic protein and solute carrier production among others. Importantly, canagliflozin-treated T cells derived from patients with autoimmune disorders impaired their effector function. Taken together, our work highlights a potential therapeutic avenue for repurposing canagliflozin as an intervention for T cell-mediated autoimmunity.

The dynamic inflammatory profile of pregnancy can be monitored using a novel lipid-based mass spectrometry technique.

The lipid environment changes throughout pregnancy both physiologically with emergent insulin resistance and pathologically e.g., gestational diabetes mellitus (GDM). Novel mass spectrometry (MS) techniques applied to minimally processed blood might lend themselves to monitoring changing lipid profiles to inform care decisions across pregnancy. In this study we use an intact-sandwich, MALDI-ToF MS method to identify phosphatidylcholine (PC) and lysophosphatidylcholine (LPC) species and calculate their ratio as an indicator of inflammation. Plasma and sera were prepared from venous blood of non-pregnant women (aged 18-40) and pregnant women at 16 weeks, 28 weeks (including GDM-positive women), and 37+ weeks (term) of gestation alongside umbilical cord blood (UCB). Women with a normal menstrual cycle and age-matched men provided finger-prick derived capillary sera at 6 time-points over a month. Serum rather than plasma was preferable for PC/LPC measurement. As pregnancy progresses, an anti-inflammatory phenotype dominates the maternal circulation, evidenced by increasing PC/LPC ratio. In contrast, the PC/LPC ratio of UCB was aligned to that of non-pregnant donors. BMI had no significant effect on the PC/LPC ratio, but GDM-complicated pregnancies had significantly lower PC/LPC at 16 weeks of gestation. To further translate the use of the PC/LPC ratio clinically, the utility of finger-prick blood was evaluated; no significant difference between capillary versus venous serum was found and we revealed the PC/LPC ratio oscillates with the menstrual cycle. Overall, we show that the PC/LPC ratio can be measured simply in human serum and has the potential to be used as a time-efficient and less invasive biomarker of (mal)adaptative inflammation.

Immunometabolic adaptation and immune plasticity in pregnancy and the bi-directional effects of obesity.

Mandatory maternal metabolic and immunological changes are essential to pregnancy success. Parallel changes in metabolism and immune function make immunometabolism an attractive mechanism to enable dynamic immune adaptation during pregnancy. Immunometabolism is a burgeoning field with the underlying principle being that cellular metabolism underpins immune cell function. With whole body changes to the metabolism of carbohydrates, protein and lipids well recognised to occur in pregnancy and our growing understanding of immunometabolism as a determinant of immunoinflammatory effector responses, it would seem reasonable to expect immune plasticity during pregnancy to be linked to changes in the availability and handling of multiple nutrient energy sources by immune cells. While studies of immunometabolism in pregnancy are only just beginning, the recognised bi-directional interaction between metabolism and immune function in the metabolic disorder obesity might provide some of the earliest insights into the role of immunometabolism in immune plasticity in pregnancy. Characterised by chronic low-grade inflammation including in pregnant women, obesity is associated with numerous adverse outcomes during pregnancy and beyond for both mother and child. Concurrent changes in metabolism and immunoinflammation are consistently described but any causative link is not well established. Here we provide an overview of the metabolic and immunological changes that occur in pregnancy and how these might contribute to healthy versus adverse pregnancy outcomes with special consideration of possible interactions with obesity.

Maternal body mass index is associated with an altered immunological profile at 28 weeks of gestation.

Healthy pregnancy is accompanied by various immunological and metabolic adaptations. Maternal obesity has been implicated in adverse pregnancy outcomes such as miscarriage, preeclampsia, and gestational diabetes mellitus (GDM), while posing a risk to the neonate. There is a lack of knowledge surrounding obesity and the maternal immune system. The objective of this study was to consider if immunological changes in pregnancy are influenced by maternal obesity. Peripheral blood was collected from fasted GDM-negative pregnant women at 26-28 weeks of gestation. Analysis was done using immunoassay, flow cytometry, bioenergetics analysis, and cell culture. The plasma profile was significantly altered with increasing BMI, specifically leptin (r = 0.7635), MCP-1 (r = 0.3024), and IL-6 (r = 0.4985). Circulating leukocyte populations were also affected with changes in the relative abundance of intermediate monocytes (r = -0.2394), CD4:CD8 T-cell ratios (r = 0.2789), and NKT cells (r = -0.2842). Monocytes analysed in more detail revealed elevated CCR2 expression and decreased mitochondrial content with increased BMI. However, LPS-stimulated cytokine production and bioenergetic profile of PBMCs were not affected by maternal BMI. The Th profile skews towards Th17 with increasing BMI; Th2 (r = -0.3202) and Th9 (r = -0.3205) cells were diminished in maternal obesity, and CytoStim™-stimulation exacerbates IL-6 (r = 0.4166), IL-17A (r = 0.2753), IL-17F (r = 0.2973), and IL-22 (r = 0.2257) production with BMI, while decreasing IL-4 (r = -0.2806). Maternal obesity during pregnancy creates an inflammatory microenvironment. Successful pregnancy requires Th2-biased responses yet increasing maternal BMI favours a Th17 response that could be detrimental to pregnancy. Further research should investigate key populations of cells identified here to further understand the immunological challenges that beset pregnant women with obesity.

Does Altered Cellular Metabolism Underpin the Normal Changes to the Maternal Immune System during Pregnancy?

Pregnancy is characterised by metabolic changes that occur to support the growth and development of the fetus over the course of gestation. These metabolic changes can be classified into two distinct phases: an initial anabolic phase to prepare an adequate store of substrates and energy which are then broken down and used during a catabolic phase to meet the energetic demands of the mother, placenta and fetus. Dynamic readjustment of immune homeostasis is also a feature of pregnancy and is likely linked to the changes in energy substrate utilisation at this time. As cellular metabolism is increasingly recognised as a key determinant of immune cell phenotype and function, we consider how changes in maternal metabolism might contribute to T cell plasticity during pregnancy.

Fructose reprogrammes glutamine-dependent oxidative metabolism to support LPS-induced inflammation.

Fructose intake has increased substantially throughout the developed world and is associated with obesity, type 2 diabetes and non-alcoholic fatty liver disease. Currently, our understanding of the metabolic and mechanistic implications for immune cells, such as monocytes and macrophages, exposed to elevated levels of dietary fructose is limited. Here, we show that fructose reprograms cellular metabolic pathways to favour glutaminolysis and oxidative metabolism, which are required to support increased inflammatory cytokine production in both LPS-treated human monocytes and mouse macrophages. A fructose-dependent increase in mTORC1 activity drives translation of pro-inflammatory cytokines in response to LPS. LPS-stimulated monocytes treated with fructose rely heavily on oxidative metabolism and have reduced flexibility in response to both glycolytic and mitochondrial inhibition, suggesting glycolysis and oxidative metabolism are inextricably coupled in these cells. The physiological implications of fructose exposure are demonstrated in a model of LPS-induced systemic inflammation, with mice exposed to fructose having increased levels of circulating IL-1ß after LPS challenge. Taken together, our work underpins a pro-inflammatory role for dietary fructose in LPS-stimulated mononuclear phagocytes which occurs at the expense of metabolic flexibility.

Macrophage Plasticity in Reproduction and Environmental Influences on Their Function.

Macrophages are key components of the innate immune system and exhibit extensive plasticity and heterogeneity. They play a significant role in the non-pregnant cycling uterus and throughout gestation they contribute to various processes underpinning reproductive success including implantation, placentation and parturition. Macrophages are also present in breast milk and impart immunomodulatory benefits to the infant. For a healthy pregnancy, the maternal immune system must adapt to prevent fetal rejection and support development of the semi-allogenic fetus without compromising host defense. These functions are dependent on macrophage polarization which is governed by the local tissue microenvironmental milieu. Disruption of this microenvironment, possibly by environmental factors of infectious and non-infectious origin, can affect macrophage phenotype and function and is linked to adverse obstetric outcomes, e.g. spontaneous miscarriage and preterm birth. Determining environmental influences on cellular and molecular mechanisms that control macrophage polarization at the maternal-fetal interface and the role of this in pregnancy complications could support approaches to alleviating adverse pregnancy outcomes.

Production and regulation of interleukin-1 family cytokines at the materno-fetal interface.

IL-1 family members regulate innate immune responses, are produced by gestation-associated tissues, and have a role in healthy and adverse pregnancy outcomes. To better understand their role at the materno-fetal interface we used a human tissue explant model to map lipopolysaccharide (LPS)-stimulated production of IL-1α, IL-1ß, IL-18, IL-33, IL-1Ra, IL-18BPa, ST2 and IL-1RAcP by placenta, choriodecidua and amnion. Caspase-dependent processing of IL-1α, IL-1ß, IL-18, and IL-33 and the ability of IL-1α, IL-1ß, IL-18, and IL-33 to regulate the production of IL-1RA, IL-18BPa, ST2 and IL-1RAcP was also determined. LPS acted as a potent inducer of IL-1 family member expression especially in the placenta and choriodecidua with the response by the amnion restricted to IL-1ß. Caspases-1, 4 and 8 contributed to LPS-stimulated production of IL-1ß and IL-18, whereas calpain was required for IL-1α production. Exogenous administration of IL-1α, IL-1ß, IL-18, and IL-33 lead to differential expression of IL-1Ra, IL-18BPa, ST2 and IL-1RAcP across all tissues examined. Most notable were the counter-regulatory effect of LPS on IL-1ß and IL-1Ra in the amnion and the broad responsiveness of the amnion to IL-1 family cytokines for increased production of immunomodulatory peptides and soluble receptors. The placenta and membranes vary not only in their output of various IL-1 family members but also in their counter-regulatory mechanisms through endogenous inhibitory peptides, processing enzymes and soluble decoy receptors. This interactive network of inflammatory mediators likely contributes to innate defence mechanisms at the materno-fetal interface to limit, in particular, the detrimental effects of microbial invasion.