Redox ratio in the left ventricle of the growth restricted fetus is positively correlated with cardiac output.

Intrauterine growth restriction (IUGR) is a result of limited substrate supply to the developing fetus in utero, and can be caused by either placental, genetic or environmental factors. Babies born IUGR can have poor long-term health outcomes, including being at higher risk of developing cardiovascular disease. Limited substrate supply in the IUGR fetus not only changes the structure of the heart but may also affect metabolism and function of the developing heart. We have utilised two imaging modalities, two-photon microscopy and phase-contrast MRI (PC-MRI), to assess alterations in cardiac metabolism and function using a sheep model of IUGR. Two-photon imaging revealed that the left ventricle of IUGR fetuses (at 140-141 d GA) had a reduced optical redox ratio, suggesting a reliance on glycolysis for ATP production. Concurrently, the use of PC-MRI to measure foetal left ventricular cardiac output (LVCO) revealed a positive correlation between LVCO and redox ratio in IUGR, but not control fetuses. These data suggest that altered heart metabolism in IUGR fetuses is indicative of reduced cardiac output, which may contribute to poor cardiac outcomes in adulthood.

In utero substrate restriction by placental insufficiency or maternal undernutrition decreases optical redox ratio in foetal perirenal fat.

Intrauterine growth restriction (IUGR) can result from reduced delivery of substrates, including oxygen and glucose, during pregnancy and may be caused by either placental insufficiency or maternal undernutrition. As a consequence of IUGR, there is altered programming of adipose tissue and this can be associated with metabolic diseases later in life. We have utilised two sheep models of IUGR, placental restriction and late gestation undernutrition, to determine the metabolic effects of growth restriction on foetal perirenal adipose tissue (PAT). Two-photon microscopy was employed to obtain an optical redox ratio, which gives an indication of cell metabolism. PAT of IUGR foetuses exhibited higher metabolic activity, altered lipid droplet morphology, upregulation of cytochrome c oxidase subunit genes and decreased expression of genes involved in growth and differentiation. Our results indicate that there are adaptations in PAT of IUGR foetuses that might be protective and ensure survival in response to an IUGR insult.

Stabilization and scale-up of photosynthesis-driven ω-hydroxylation of nonanoic acid methyl ester by two-liquid phase whole-cell biocatalysis.

Photoautotrophic organisms are promising hosts for biocatalytic oxyfunctionalizations because they supply reduction equivalents as well as O2 via photosynthetic water oxidation. Thus far, research on photosynthesis-driven bioprocesses mainly focuses on strain development and the proof of principle in small-scale biocatalytic reaction setups. This study investigates the long-term applicability of the previously developed cyanobacterial strain Synechocystis sp. PCC 6803_BGT harboring the alkane monooxygenase system AlkBGT catalyzing terminal alkyl group oxyfunctionalization. For the regiospecific ω-hydroxylation of nonanoic acid methyl ester (NAME), this biocatalyst showed light intensity-independent hydroxylation activity and substantial hydrolysis of NAME to nonanoic acid. Substrate mass transfer limitation, substrate hydrolysis, as well as reactant toxicity were overcome via in situ substrate supply by means of a two-liquid phase system. The application of diisononyl phthalate as organic carrier solvent enabled 1.7-fold increased initial specific activities (5.6 ± 0.1 U/gCDW ) and 7.6-fold increased specific yields on biomass (3.8 ± 0.1 mmolH-NAME /gCDW ) as compared with single aqueous phase biotransformations. Finally, the whole-cell biotransformation system was successfully scaled from glass tubes to a stirred-tank photobioreactor. This is the first study reporting the application of the two-liquid phase concept for efficient phototrophic whole-cell biocatalysis.

Enhancement of substrate supply and ido expression to improve 4-hydroxyisoleucine production in recombinant Corynebacterium glutamicum ssp. lactofermentum.

4-Hydroxyisoleucine (4-HIL) has potential value in treating diabetes. L-isoleucine dioxygenase (IDO) catalyzes the hydroxylation of L-isoleucine (Ile) to form 4-HIL with the concomitant oxidation of α-ketoglutarate (α-KG) and oxygen consumption. In our previous study, by expressing the ido gene in the Ile producer Corynebacterium glutamicum ssp. lactofermentum SN01, 4-HIL was de novo-synthesized from glucose without adding either Ile or α-KG. In this study, synergistically improving the substrates supply and IDO activity was applied to enhance the de novo biosynthesis of 4-HIL. Deletion of aceA and blocking of the glyoxylate pathway effectively enhanced α-KG supply and Ile synthesis and thus improved 4-HIL production to 69.47 ± 2.18 mM, 18.9% higher than in the original strain. Coexpression of mqo with ido further improved Ile synthesis but decreased 4-HIL production, partially due to the inadequate activity of IDO. Coexpression of another gene, ido3, with mqo and ido efficiently promoted IDO activity, thus improving 4-HIL production to 91.54 ± 8.29 mM. Further expression of vgb and promotion of the oxygen uptake rate did not change the 4-HIL titer significantly but increased the 4-HIL production rate in the first 72 h of fermentation. After fermentation in the optimized medium, 4-HIL production by the final strains increased to 112-117 mM, indicating these strains are promising candidates for producing 4-HIL. These results demonstrate that synergistically promoting substrate supply and improving IDO activity are efficient approaches to enhance 4-HIL production in C. glutamicum.

Development of in situ product removal strategies in biocatalysis applying scaled-down unit operations.

An experimental platform based on scaled-down unit operations combined in a plug-and-play manner enables easy and highly flexible testing of advanced biocatalytic process options such as in situ product removal (ISPR) process strategies. In such a platform, it is possible to compartmentalize different process steps while operating it as a combined system, giving the possibility to test and characterize the performance of novel process concepts and biocatalysts with minimal influence of inhibitory products. Here the capabilities of performing process development by applying scaled-down unit operations are highlighted through a case study investigating the asymmetric synthesis of 1-methyl-3-phenylpropylamine (MPPA) using ω-transaminase, an enzyme in the sub-family of amino transferases (ATAs). An on-line HPLC system was applied to avoid manual sample handling and to semi-automatically characterize ω-transaminases in a scaled-down packed-bed reactor (PBR) module, showing MPPA as a strong inhibitor. To overcome the inhibition, a two-step liquid-liquid extraction (LLE) ISPR concept was tested using scaled-down unit operations combined in a plug-and-play manner. Through the tested ISPR concept, it was possible to continuously feed the main substrate benzylacetone (BA) and extract the main product MPPA throughout the reaction, thereby overcoming the challenges of low substrate solubility and product inhibition. The tested ISPR concept achieved a product concentration of 26.5 gMPPA · L-1 , a purity up to 70% gMPPA · gtot-1 and a recovery in the range of 80% mol · mol-1 of MPPA in 20 h, with the possibility to increase the concentration, purity, and recovery further. Biotechnol. Bioeng. 2017;114: 600-609. © 2016 Wiley Periodicals, Inc.

Integrated omics approaches provide strategies for rapid erythromycin yield increase in Saccharopolyspora erythraea.

BACKGROUND: Omics approaches have significantly increased our understanding of biological systems. However, they have had limited success in explaining the dramatically increased productivity of commercially important natural products by industrial high-producing strains, such as the erythromycin-producing actinomycete Saccharopolyspora erythraea. Further yield increase is of great importance but requires a better understanding of the underlying physiological processes. RESULTS: To reveal the mechanisms related to erythromycin yield increase, we have undertaken an integrated study of the genomic, transcriptomic, and proteomic differences between the wild type strain NRRL2338 (WT) and the industrial high-producing strain ABE1441 (HP) of S. erythraea at multiple time points of a simulated industrial bioprocess. 165 observed mutations lead to differences in gene expression profiles and protein abundance between the two strains, which were most prominent in the initial stages of erythromycin production. Enzymes involved in erythromycin biosynthesis, metabolism of branched chain amino acids and proteolysis were most strongly upregulated in the HP strain. Interestingly, genes related to TCA cycle and DNA-repair were downregulated. Additionally, comprehensive data analysis uncovered significant correlations in expression profiles of the erythromycin-biosynthetic genes, other biosynthetic gene clusters and previously unidentified putative regulatory genes. Based on this information, we demonstrated that overexpression of several genes involved in amino acid metabolism can contribute to increased yield of erythromycin, confirming the validity of our systems biology approach. CONCLUSIONS: Our comprehensive omics approach, carried out in industrially relevant conditions, enabled the identification of key pathways affecting erythromycin yield and suggests strategies for rapid increase in the production of secondary metabolites in industrial environment.

Changing appetites: the adaptive advantages of fuel choice.

Cells are capable of metabolizing a variety of carbon substrates, including glucose, fatty acids, ketone bodies, and amino acids. Cellular fuel choice not only fulfills specific biosynthetic needs, but also enables programmatic adaptations to stress conditions beyond compensating for changes in nutrient availability. Emerging evidence indicates that specific switches from utilization of one substrate to another can have protective or permissive roles in disease pathogenesis. Understanding the molecular determinants of cellular fuel preference may provide insights into the homeostatic control of stress responses, and unveil therapeutic targets. Here, we highlight overarching themes encompassing cellular fuel choice; its link to cell fate and function; its advantages in stress protection; and its contribution to metabolic dependencies and maladaptations in pathological conditions.

Intensive medicine - Guidelines on Parenteral Nutrition, Chapter 14.

In intensive care patients parenteral nutrition (PN) should not be carried out when adequate oral or enteral nutrition is possible. Critically ill patients without symptoms of malnutrition, who probably cannot be adequately nourished enterally for a period of <5 days, do not require full PN but should be given at least a basal supply of glucose. Critically ill patients should be nourished parenterally from the beginning of intensive care if they are unlikely to be adequately nourished orally or enterally even after 5-7 days. Critically ill and malnourished patients should, in addition to a possible partial enteral nutrition, be nourished parenterally. Energy supply should not be constant, but should be adapted to the stage, the disease has reached. Hyperalimentation should be avoided at an acute stage of disease in any case. Critically ill patients should be given, as PN, a mixture consisting of amino acids (between 0.8 and 1.5 g/kg/day), carbohydrates (around 60% of the non-protein energy) and fat (around 40% of the non-protein energy) as well as electrolytes and micronutrients.