Computational method for inferring objective function of glycerol metabolism in Klebsiella pneumoniae

Flux balance analysis (FBA) is an effective tool in the analysis of metabolic network. It can predict the flux distribution of engineered cells, whereas the accurate prediction depends on the reasonable objective function. In this work, we propose two nonlinear bilevel programming models on anaerobic glycerol metabolism in Klebsiella pneumoniae (K. pneumoniae) for 1,3-propanediol (1,3-PD) production. One intends to infer the metabolic objective function, and the other is to analyze the robustness of the objective function. In view of the models' characteristic an improved genetic algorithm is constructed to solve them, where some techniques are adopted to guarantee all chromosomes are feasible and move quickly towards the global optimal solution. Numerical results reveal some interesting conclusions, e.g., biomass production is the main force to drive K. pneumoniae metabolism, and the objective functions, which are obtained in term of several different groups of flux distributions, are similar.     

基元通量模式预测酵母生长现象

本文通过EFM预测了基因突变后的酵母细胞生长现象, 模拟预测结果和实验结果吻合很好; 与FBA方法得到的模拟结果相比较, EFM方法能更好地把基因突变和其表型(生长)联系起来.     

Zooming-in on cancer metabolic rewiring with tissue specific constraint-based models

The metabolic rearrangements occurring in cancer cells can be effectively investigated with a Systems Biology approach supported by metabolic network modeling. We here present tissue-specific constraint-based core models for three different types of tumors (liver, breast and lung) that serve this purpose. The core models were extracted and manually curated from the corresponding genome-scale metabolic models in the Human Metabolic Atlas database with a focus on the pathways that are known to play a key role in cancer growth and proliferation. Along similar lines, we also reconstructed a core model from the original general human metabolic network to be used as a reference model.A comparative Flux Balance Analysis between the reference and the cancer models highlighted both a clear distinction between the two conditions and a heterogeneity within the three different cancer types in terms of metabolic flux distribution. These results emphasize the need for modeling approaches able to keep up with this tumoral heterogeneity in order to identify more suitable drug targets and develop effective treatments. According to this perspective, we identified key points able to reverse the tumoral phenotype toward the reference one or vice-versa.     

SPECIES DIFFERENCES IN RESPONSE TO PHOTOHEMOLYTIC AGENTS

Species differences in red blood cell susceptibility to the photohemolytic agents chlorpromazine, menadione and tetracycline were examined in mouse, rat, dog, and human blood. Menadione and tetracycline (25 microM) hemolyzed mouse but not dog, rat, or human red blood cells (RBC) when irradiated with UV light but not in the dark. Chlorpromazine (25 microM) produced a photohemolytic response in all four species with mouse and rat RBC lysing fastest followed by human then dog cells. Investigations into the nature of these species differences suggested that the size of mouse RBC may contribute to its high sensitivity to photohemolytic agents. An investigation of the effect of UV light on key antioxidant enzymes revealed species differences in enzyme inactivation. These data suggest that mouse RBC may be particularly vulnerable to phototoxic agents, especially those compounds which produce active oxygen species and, therefore, may prove more useful than human RBC as a model for predicting phototoxic potential of some chemical entities.     

Metabonomics Study of the Hematopoietic Effect of Medicinal Wine Maoji Jiu on a Blood Deficiency Rat Model by Ultra-High-Performance Liquid Chromatography Coupled to Quadrupole Time-of-Flight Mass Spectrometry and a Pattern Recognition Approach

Maoji Jiu (MJ) is a kind of medicinal wine that has been widely used by Chinese people for many years to nourish and promote blood circulation. The purpose of this study was to investigate the hematopoietic effect of MJ on the metabolism of blood deficient rats and to explore the underlying hematopoietic regulation mechanisms. Blood deficiency model rats were induced by subcutaneous injection of N-acetylphenylhydrazine (APH) and intraperitoneal injection of cyclophosphamide (CTX). The plasma metabolic fingerprints of blood deficiency model rats with and without MJ treatment were obtained by using metabonomics based on ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC–QTOF/MS). Orthogonal partial least squares-discriminant analysis (OPLS–DA) was used to evaluate the hematopoietic effect of MJ and identify potential biomarkers in the plasma of blood deficiency model rats. The levels of white blood cells (WBC), red blood cells (RBC) and hemoglobin (HGB) and the activity of antioxidant capacity showed a recovery trend to the control group after MJ treatment, while the dose of 10 mL/kg showed the best effect. In this study, thirteen potential biomarkers were identified, which were mainly related to seven metabolic pathways, including linoleic acid metabolism, d-glutamine and d-glutamate metabolism, alanine, aspartate and glutamate metabolism, tryptophan metabolism, pyrimidine metabolism, porphyrin and chlorophyll metabolism and arginine biosynthesis. Metabolomics was applied frequently to reflect the physiological and metabolic state of organisms comprehensively, indicating that the rapid plasma metabonomics may be a potentially powerful tool to reveal the efficacy and enriching blood mechanism of MJ.     

The NMR added value to the green foodomics perspective: Advances by machine learning to the holistic view on food and nutrition

Food is a complex matter, literally. From production to functionalization, from nutritional quality engineering to predicting effects on health, the interest in finding an efficient physicochemical characterization of food has boomed in recent years. The sheer complexity of characterizing food and its interaction with the human organism has however made the use of data driven approaches in modeling a necessity. High-throughput techniques, such as nuclear magnetic resonance (NMR) spectroscopy, are well suited for omics data production and, coupled with machine learning, are paving a promising way of modeling food–human interaction. The foodomics approach sets the framework for omic data integration in food studies, in which NMR experiments play a key role. NMR data can be used to assess nutritional qualities of food, helping the design of functional and sustainable sources of nutrients; detect biomarkers of intake and study how they impact the metabolism of different individuals; study the kinetics of compounds in foods or their by-products to detect pathological conditions; and improve the efficiency of in silico models of the metabolic network.     

Heat flux and the calorimetric-respirometric ratio as measures of catabolic flux in mammalian cells

It is advocated that cellular heat flow rate (Ø = dQ/dt, where Q is heat) be expressed as an intensive quantity specific to cell size (X) and termed heat flux (J_Ø/X). It has been the practice to cite such data on a ‘per cell’ basis, but it would be preferable to use biomass (cellular volume or mass). This quantity is shown to be a measure of metabolic activity and, more accurately, catabolic rate coupled to the demand for ATP in anabolic processes and work in the cell. Recent developments in flow microcalorimetry and dielectric spectroscopy reveal that heat flux can be measured on-line, with the potential of industrial use as a control variable in the growth of hybridoma and genetically engineered cells. This is because the enthalpy change of growth can be regarded as a unique kind of stoichiometric coefficient directly related to the mass coefficients in the growth reaction. This can be verified by an enthalpy balance comparing data for material fluxes of catabolites with the value for heat flux. Information revealed by the stoichiometric growth equation can be used to improve medium design.

The ratio of heat flux to oxygen consumption (flux) is known as the calorimetric-respirometric (CR) ratio. It detects anaerobic processes when the value is more negative than −450 (±5%) kJ mol⁻¹ O₂. These processes are found in cells growing under fully aerobic conditions, because glycolysis provides biosynthetic precursors with lactate as the byproduct. It is suggested that the CR ratio would be a powerful on-line control variable for the growth of animal cells in bioreactors.     

Colored Corn: An Up-Date on Metabolites Extraction,Health Implication,and Potential Use

Colored (orange, pink, red, purple, and blue) corn strongly attracted attention on its healthy properties mainly due to its anthocyanin and carotenoid composition which is also responsible for its pigmentation. The present review summarized the recent updates on the extraction and chemical characterization of the main plant secondary metabolites present in colored seeds, kernel, cob, husk, and silk. The main approaches used to stabilize the extracts have been discussed as well as their food and non-food uses. Both in vitro and in vivo (animal models) studies on the different effects (antibacterial, antimutagenic, antioxidant, and anti-inflammatory activities, effects on metabolic syndrome, diabetes, glucose and lipidic metabolism, and neuroprotection) of pigmented extracts on animal and human health have been summarized.     

Computational prediction of metabolism: sites, products, SAR, P450 enzyme dynamics, and mechanisms

Metabolism of xenobiotics remains a central challenge for the discovery and development of drugs, cosmetics, nutritional supplements, and agrochemicals. Metabolic transformations are frequently related to the incidence of toxic effects that may result from the emergence of reactive species, the systemic accumulation of metabolites, or by induction of metabolic pathways. Experimental investigation of the metabolism of small organic molecules is particularly resource demanding; hence, computational methods are of considerable interest to complement experimental approaches. This review provides a broad overview of structure- and ligand-based computational methods for the prediction of xenobiotic metabolism. Current computational approaches to address xenobiotic metabolism are discussed from three major perspectives: (i) prediction of sites of metabolism (SOMs), (ii) elucidation of potential metabolites and their chemical structures, and (iii) prediction of direct and indirect effects of xenobiotics on metabolizing enzymes, where the focus is on the cytochrome P450 (CYP) superfamily of enzymes, the cardinal xenobiotics metabolizing enzymes. For each of these domains, a variety of approaches and their applications are systematically reviewed, including expert systems, data mining approaches, quantitative structure-activity relationships (QSARs), and machine learning-based methods, pharmacophore-based algorithms, shape-focused techniques, molecular interaction fields (MIFs), reactivity-focused techniques, protein-ligand docking, molecular dynamics (MD) simulations, and combinations of methods. Predictive metabolism is a developing area, and there is still enormous potential for improvement. However, it is clear that the combination of rapidly increasing amounts of available ligand- and structure-related experimental data (in particular, quantitative data) with novel and diverse simulation and modeling approaches is accelerating the development of effective tools for prediction of in vivo metabolism, which is reflected by the diverse and comprehensive data sources and methods for metabolism prediction reviewed here. This review attempts to survey the range and scope of computational methods applied to metabolism prediction and also to compare and contrast their applicability and performance.     

Toxic Metal Species and ‘Endogenous’ Metalloproteins at the Blood–Organ Interface: Analytical and Bioinorganic Aspects

Globally, human exposure to environmental pollutants causes an estimated 9 million deaths per year and it could also be implicated in the etiology of diseases that do not appear to have a genetic origin. Accordingly, there is a need to gain information about the biomolecular mechanisms that causally link exposure to inorganic environmental pollutants with distinct adverse health effects. Although the analysis of blood plasma and red blood cell (RBC) cytosol can provide important biochemical information about these mechanisms, the inherent complexity of these biological matrices can make this a difficult task. In this perspective, we will examine the use of metalloentities that are present in plasma and RBC cytosol as potential exposure biomarkers to assess human exposure to inorganic pollutants. Our primary objective is to explore the principal bioinorganic processes that contribute to increased or decreased metalloprotein concentrations in plasma and/or RBC cytosol. Furthermore, we will also identify metabolites which can form in the bloodstream and contain essential as well as toxic metals for use as exposure biomarkers. While the latter metal species represent useful biomarkers for short-term exposure, endogenous plasma metalloproteins represent indicators to assess the long-term exposure of an individual to inorganic pollutants. Based on these considerations, the quantification of metalloentities in blood plasma and/or RBC cytosol is identified as a feasible research avenue to better understand the adverse health effects that are associated with chronic exposure of various human populations to inorganic pollutants. Exposure to these pollutants will likely increase as a consequence of technological advances, including the fast-growing applications of metal-based engineering nanomaterials.     

Role of Human Liver Microsomes in In Vitro Metabolism of Drugs—A Review

Drug metabolism studies are essential and necessary during the evaluation of drugs. This review discusses the in vitro human liver models to estimate the drug metabolic fates in vivo. Different approaches are provided and emphasis is placed on the potential of human liver microsomes for drug metabolism and inhibition studies. The methodology for these studies using human liver microsomes, applications of human liver microsomes, and the drugs studied by human liver microsomes are listed. Human liver microsomes represent a critical experimental model for the evaluation of drug metabolites with a high probability of clinical success.     

~(13)C代谢通量分析

代谢通量分析(metabolic flux analysis,MFA)是通过确定代谢网络中代谢流分布来表征细胞代谢状态的强有力的工具。鉴于计量学代谢通量分析在处理复杂代谢网络时表现出的局限性,发展了以13C标记实验为基础的13C MFA。本文介绍了13C MFA的原理与方法,总结和评述了13C MFA在实验与数据分析方面的最新进展以及MFA在功能基因组研究中的重要地位,同时对代谢通量分析的发展前景进行了展望。     

Electrokinetic transport of red blood cells in microcapillaries

Electrokinetic flow of a suspension of erythrocytes (red blood cells, RBCs) in 20 num cylindrical fused-silica capillaries is examined in the present work. Flow direction anomalies are observed experimentally and tentatively explained by the development of a pH gradient between the cathode well and the anode well due to electrolysis reactions at the electrodes. This pH gradient alters the local zeta potentials of both the capillary and the RBC and thus the local electroendosmotic liquid flow (EOF) velocities and RBC electrophoretic (EP) velocities. The two velocities are opposite in direction but with EOF dominating such that the RBC moves toward the cathode, opposite to the anode migration observed in bulk conditions. The opposing zeta potentials also lead to RBC aggregation at the anode end for low fields less than 25 V/cm. As the electroendosmotic velocity decreases at the anode end due to decreasing pH, pressure-driven back flow develops to oppose the original EOF at the remaining portions of the capillary ensuring constant fluid flux. When the anode EOF velocity is smaller in magnitude than the EP velocity, reversal of blood cell transport is observed after a short transient time in which a pH gradient forms. RBC velocities and pH dependencies on electric field and MgCl(2) concentration are presented along with data showing the accumulation of charge separation across the capillary. Also, a short-term solution to the pH gradient formation is presented that could help thwart development of pH gradients in micro-devices at lower voltages.     

Cholesterol-rich membrane coatings for interaction studies in capillary electrophoresis: application to red blood cell lipid extracts

The purpose was to develop a stable biological membrane coating for CE useful for membrane interaction studies. The effect of cholesterol (chol) on the stability of dipalmitoylphosphatidylcholine (DPPC) and sphingomyelin (SM) coatings was studied. In addition, a fused-silica capillary for CE was coated with human red blood cell (RBC) ghost lipids. Liposomes prepared of DPPC/SM with and without chol or RBC ghost lipids were flushed through the capillary and the stability of the coating was measured electrophoretically. Similar mixtures of DPPC/SM with and without chol were further studied by differential scanning calorimetry. The presence of phosphatidylcholine as a basic component in the coating solution of DPPC/SM/chol was found to be essential to achieve a good and stable coating. The results also confirmed the stability of coatings obtained with solutions of DPPC with 0-30 mol% of chol and SM in different ratios, which more closely resemble natural membranes. Finally, the electrophoretic measurements revealed that a stable coating is formed when capillaries are coated with liposomes of RBC ghost lipids.     

Theoretical aspects of C metabolic flux analysis with sole quantification of carbon dioxide labeling

The potential of using sole respirometric CO2 labeling measurement for 13C metabolic flux analysis was investigated by metabolic simulations. For this purpose a model was created, considering all CO2 forming and consuming reactions in the central catabolic and anabolic pathways. To facilitate the interpretation of the simulation results, the underlying metabolic network was parameterized by physiologically meaningful flux parameters such as flux partitioning ratios at metabolic branch points and reaction reversibilities. For real case flux scenarios of the industrial amino acid producer Corynebacterium glutamicum and different commercially available (13)C-labeled tracer substrates, observability and output sensitivity towards key flux parameters was investigated. Metabolic net fluxes in the central metabolism, involving, e.g. glycolysis, pentose phosphate pathway, tricarboxylic acid cycle, anaplerotic carboxylation, and glyoxylate pathway were found to be determinable by the respirometric approach using a combination of [1-13C] and [6-13C] glucose in two parallel studies. The reversibilities of bidirectional reactions influence the isotopic labeling of CO2 only to a negligible degree. On one hand, they therefore cannot be determined. On the other hand, their precise values are not required for the quantification of net fluxes. Computer-aided optimal experimental design was carried out to predict the quality of the information from the respirometric tracer experiments and identify suitable tracer substrates. A combination of [1-13C] and [6-13C] glucose in two parallel studies was found to yield a similar quality of information as compared to an approach with mass spectrometric labeling analysis of secreted products. The quality of information can be further increased by additional studies with [1,2-13C2] or [1,6-13C2] glucose. Respirometric tracer studies with sole labeling analysis of CO2 are therefore promising for 13C metabolic flux analysis.     

Theoretical aspects of 13C metabolic flux analysis with sole quantification of carbon dioxide labeling

The potential of using sole respirometric CO2 labeling measurement for 13C metabolic flux analysis was investigated by metabolic simulations. For this purpose a model was created, considering all CO2 forming and consuming reactions in the central catabolic and anabolic pathways. To facilitate the interpretation of the simulation results, the underlying metabolic network was parameterized by physiologically meaningful flux parameters such as flux partitioning ratios at metabolic branch points and reaction reversibilities. For real case flux scenarios of the industrial amino acid producer Corynebacterium glutamicum and different commercially available (13)C-labeled tracer substrates, observability and output sensitivity towards key flux parameters was investigated. Metabolic net fluxes in the central metabolism, involving, e.g. glycolysis, pentose phosphate pathway, tricarboxylic acid cycle, anaplerotic carboxylation, and glyoxylate pathway were found to be determinable by the respirometric approach using a combination of [1-13C] and [6-13C] glucose in two parallel studies. The reversibilities of bidirectional reactions influence the isotopic labeling of CO2 only to a negligible degree. On one hand, they therefore cannot be determined. On the other hand, their precise values are not required for the quantification of net fluxes. Computer-aided optimal experimental design was carried out to predict the quality of the information from the respirometric tracer experiments and identify suitable tracer substrates. A combination of [1-13C] and [6-13C] glucose in two parallel studies was found to yield a similar quality of information as compared to an approach with mass spectrometric labeling analysis of secreted products. The quality of information can be further increased by additional studies with [1,2-13C2] or [1,6-13C2] glucose. Respirometric tracer studies with sole labeling analysis of CO2 are therefore promising for 13C metabolic flux analysis.     

Direct measurement of the impact of impaired erythrocyte deformability on microvascular network perfusion in a microfluidic device

The ability of red blood cells (RBCs, erythrocytes) to deform and pass through capillaries is essential for continual flow of blood in the microvasculature, which ensures an adequate supply of oxygen and nutrients, prompt removal of metabolic waste products, transport of drugs and hormones, and traffic of circulating cells to and from all living tissues. This paper presents a novel tool for evaluating the impact of impaired deformability of RBCs on the flow of blood in the microvasculature by directly measuring perfusion of a test microchannel network with dimensions and topology similar to the real microcirculation. The measurement of microchannel network perfusion is compared with RBC filtration -- a conventional assay of RBC deformability. In contrast to RBC filterability, network perfusion depends linearly on RBC deformability modulated by graded exposure to glutaraldehyde, showing a higher sensitivity to small changes of deformability. The direct measurement of microchannel network perfusion represents a new concept for the field of blood rheology and should prove beneficial for basic science and clinical applications.     

A sensitive and specific method for the determination of total ribavirin in human red blood cells by liquid chromatography-tandem mass spectrometry

A sensitive and specific method using high-performance liquid chromatography (LC)-tandem mass spectrometry (MS) for the analysis of total ribavirin in human red blood cells (RBC) is developed and validated. The method involves the addition of an internal standard and perchloric acid, the conversion of ribavirin phosphorylated metabolites to ribavirin, purification with a solid-phase exchange cartridge, and LC-MS-MS analysis. The MS-MS is selected to monitor m/z 245-113 for ribavirin and m/z 250-113 for [13C]ribavirin using positive electrospray ionization. The calibration curve is linear over a concentration of 100-10,000 ng/mL with a limit of quantitation of 100 ng/mL. Mean interassay accuracy for quality control (QC) at 100, 1000, and 10,000 ng/mL are 101.8%, 99.4%, and 98.8%, respectively. Mean interassay precision (%CV) for QC at 100, 1000, and 10,000 ng/mL are 5.0%, 5.0%, and 2.5%, respectively. Extractibility of total ribavirin from RBC is confirmed with RBC obtained from a [(14)C]ribavirin-dosed monkey. The method is used to determine the free and total ribavirin concentration in human RBC obtained from hepatitis C patients treated with ribavirin.     

Covalent binding of polyethylene glycol to the surface of red blood cells as detected and followed up by cell electrophoresis and rheological methods

Cyanuric chloride activated polyethylene glycol (PEG)-5000 was covalently coupled to murine and human red blood cells (pegylated RBC). Our purpose was to camouflage RBC receptors, which is necessary for parasite invasion, a process essential to sustain parasitemia. Cell electrophoretic mobility analysis (CEM) of pegylated RBC distinguished a new population of cells bearing characteristic CEM. Pegylation of RBC also modified their rheological properties, which were documented by evaluation of cell deformability (based on cell transit time through calibrated micropores) and cell aggregation (as measured by ultrasonic interferometry). Homologous transfusion of pegylated RBC into murine malaria-infected mice had no significant effect on the cerebral malaria death rate in Plasmodium berghei-infected mice, but it reduced the peripheral blood parasitemia by a factor 2 while in Plasmodium yoelii infected mice, the parasitemia was dramatically reduced by a factor of 4. These experiments demonstrate that transfusion of pegylated RBC may inhibit peripheral parasitemia. Cell electrophoresis appears to be a useful tool to allow in vivo detection and to investigate the fate of transfused pegylated RBC.