Modelling Metabolic Shifts during Cardiomyocyte Differentiation, Iron Deficiency and Transferrin Rescue Using Human Pluripotent Stem Cells

Cardiomyocytes rely on specialised metabolism to meet the high energy demand of the heart. During heart development, metabolism matures and shifts from the predominant utilisation of glycolysis and glutamine oxidation towards lactate and fatty acid oxidation. Iron deficiency (ID) leads to cellular metabolism perturbations. However, the exact alterations in substrate metabolism during ID are poorly defined. Using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM), the present study investigated changes in major metabolic substrate utilisation in the context of ID or upon transferrin rescue. Typically, during hiPSC-CM differentiation, the greatest increase in total metabolic output and rate was seen in fatty acid metabolism. When ID was induced, hiPSC-CMs displayed increased reliance on glycolytic metabolism, and six TCA cycle, five amino acid, and four fatty acid substrates were significantly impaired. Transferrin rescue was able to improve TCA cycle substrate metabolism, but the amino acid and fatty acid metabolism remained perturbed. Replenishing iron stores partially reverses the adverse metabolic changes that occur during ID. Understanding the changes in metabolic substrate utilisation and their modification may provide potential for discovery of new biomarkers and therapeutic targets in cardiovascular diseases.

Evolutionary engineering of Wickerhamomyces subpelliculosus and Kazachstania gamospora for baking

The conventional baker’s yeast, Saccharomyces cerevisiae , is an indispensable baking workhorse of all times. Its monopoly coupled to its major drawbacks such as streamlined carbon substrate utilisation base and a poor ability to withstand a number of baking associated stresses prompt the need to search for alternative yeasts to leaven bread in the era of increasingly complex consumer lifestyles. Our previous work identified the inefficient baking attributes of Wickerhamomyces subpelliculosus and Kazachstania gamospora as well as preliminarily observations of improving fermentative capacity of potential alternative baker’s yeasts using evolutionary engineering. Here we report the characterisation and improvement in baking traits in five out of six independently evolved lines incubated for longer time and passaged for at least 60 cycles relative to their parental strains as well as the conventional baker’s yeast. In addition, evolved clones produced bread with a higher loaf volume when compared to bread baked with either ancestral strain or the control conventional baker’s yeast. Remarkably, our approach improved the yeasts’ ability to withstand baking associated stresses, a key baking trait exhibited poorly in both the conventional baker’s yeast and their ancestral strains. W. subpelliculosus evolved the best characteristics attractive for alternative baker’s yeasts as compared to the evolved K. gamospora strains. These results demonstrate the robustness of evolutionary engineering in development of alternative baker’s yeasts.

The Impact of Decaffeinated Green Tea Extract on Fat Oxidation, Body Composition and Cardio-Metabolic Health in Overweight, Recreationally Active Individuals

This study investigated the effect of decaffeinated green tea extract (dGTE), with or without antioxidant nutrients, on fat oxidation, body composition and cardio-metabolic health measures in overweight individuals engaged in regular exercise. Twenty-seven participants (20 females, 7 males; body mass: 77.5 ± 10.5 kg; body mass index: 27.4 ± 3.0 kg·m2; peak oxygen uptake (V.O2peak): 30.2 ± 5.8 mL·kg−1·min−1) were randomly assigned, in a double-blinded manner, either: dGTE (400 mg·d−1 (−)-epigallocatechin−3-gallate (EGCG), n = 9); a novel dGTE+ (400 mg·d−1 EGCG, quercetin (50 mg·d−1) and α-lipoic acid (LA, 150 mg·d−1), n = 9); or placebo (PL, n = 9) for 8 weeks, whilst maintaining standardised, aerobic exercise. Fat oxidation (‘FATMAX’ and steady state exercise protocols), body composition, cardio-metabolic and blood measures (serum glucose, insulin, leptin, adiponectin, glycerol, free fatty acids, total cholesterol, high [HDL-c] and low-density lipoprotein cholesterol [LDL-c], triglycerides, liver enzymes and bilirubin) were assessed at baseline, week 4 and 8. Following 8 weeks of dGTE+, maximal fat oxidation (MFO) significantly improved from 154.4 ± 20.6 to 224.6 ± 23.2 mg·min−1 (p = 0.009), along with a 22.5% increase in the exercise intensity at which fat oxidation was deemed negligible (FATMIN; 67.6 ± 3.6% V.O2peak, p = 0.003). Steady state exercise substrate utilisation also improved for dGTE+ only, with respiratory exchange ratio reducing from 0.94 ± 0.01 at week 4, to 0.89 ± 0.01 at week 8 (p = 0.004). This corresponded with a significant increase in the contribution of fat to energy expenditure for dGTE+ from 21.0 ± 4.1% at week 4, to 34.6 ± 4.7% at week 8 (p = 0.006). LDL-c was also lower (normalised fold change of −0.09 ± 0.06) for dGTE+ by week 8 (p = 0.038). No other significant effects were found in any group. Eight weeks of dGTE+ improved MFO and substrate utilisation during exercise, and lowered LDL-c. However, body composition and cardio-metabolic markers in healthy, overweight individuals who maintained regular physical activity were largely unaffected by dGTE.

A model for determining cardiac mitochondrial substrate utilisation using stable 13C-labelled metabolites

Introduction Relative oxidation of different metabolic substrates in the heart varies both physiologically and pathologically, in order to meet metabolic demands under different circumstances. 13C labelled substrates have become a key tool for studying substrate use—yet an accurate model is required to analyse the complex data produced as these substrates become incorporated into the Krebs cycle. Objectives We aimed to generate a network model for the quantitative analysis of Krebs cycle intermediate isotopologue distributions measured by mass spectrometry, to determine the 13C labelled proportion of acetyl-CoA entering the Krebs cycle. Methods A model was generated, and validated ex vivo using isotopic distributions measured from isolated hearts perfused with buffer containing 11 mM glucose in total, with varying fractions of universally labelled with 13C. The model was then employed to determine the relative oxidation of glucose and triacylglycerol by hearts perfused with 11 mM glucose and 0.4 mM equivalent Intralipid (a triacylglycerol mixture). Results The contribution of glucose to Krebs cycle oxidation was measured to be 79.1 ± 0.9%, independent of the fraction of buffer glucose which was U-13C labelled, or of which Krebs cycle intermediate was assessed. In the presence of Intralipid, glucose and triglyceride were determined to contribute 58 ± 3.6% and 35.6 ± 0.8% of acetyl-CoA entering the Krebs cycle, respectively. Conclusion These results demonstrate the accuracy of a functional model of Krebs cycle metabolism, which can allow quantitative determination of the effects of therapeutics and pathology on cardiac substrate metabolism.