A Systems Biology Approach to Iron Metabolism

Iron is critical to the survival of almost all living organisms. However, inappropriately low or high levels of iron are detrimental and contribute to a wide range of diseases. Recent advances in the study of iron metabolism have revealed multiple intricate pathways that are essential to the maintenance of iron homeostasis. Further, iron regulation involves processes at several scales, ranging from the subcellular to the organismal. This complexity makes a systems biology approach crucial, with its enabling technology of computational models based on a mathematical description of regulatory systems. Systems biology may represent a new strategy for understanding imbalances in iron metabolism and their underlying cause

A Boolean probabilistic model of metabolic adaptation to oxygen in relation to iron homeostasis and oxidative stress

<p>Abstract</p> <p>Background</p> <p>In aerobically grown cells, iron homeostasis and oxidative stress are tightly linked processes implicated in a growing number of diseases. The deregulation of iron homeostasis due to gene defects or environmental stresses leads to a wide range of diseases with consequences for cellular metabolism that remain poorly understood. The modelling of iron homeostasis in relation to the main features of metabolism, energy production and oxidative stress may provide new clues to the ways in which changes in biological processes in a normal cell lead to disease.</p> <p>Results</p> <p>Using a methodology based on probabilistic Boolean modelling, we constructed the first model of yeast iron homeostasis including oxygen-related reactions in the frame of central metabolism. The resulting model of 642 elements and 1007 reactions was validated by comparing simulations with a large body of experimental results (147 phenotypes and 11 metabolic flux experiments). We removed every gene, thus generating <it>in silico </it>mutants. The simulations of the different mutants gave rise to a remarkably accurate qualitative description of most of the experimental phenotype (overall consistency > 91.5%). A second validation involved analysing the anaerobiosis to aerobiosis transition. Therefore, we compared the simulations of our model with different levels of oxygen to experimental metabolic flux data. The simulations reproducted accurately ten out of the eleven metabolic fluxes. We show here that our probabilistic Boolean modelling strategy provides a useful description of the dynamics of a complex biological system. A clustering analysis of the simulations of all <it>in silico </it>mutations led to the identification of clear phenotypic profiles, thus providing new insights into some metabolic response to stress conditions. Finally, the model was also used to explore several new hypothesis in order to better understand some unexpected phenotypes in given mutants.</p> <p>Conclusions</p> <p>All these results show that this model, and the underlying modelling strategy, are powerful tools for improving our understanding of complex biological problems.</p

Computational Techniques for the Structural and Dynamic Analysis of Biological Networks

The analysis of biological systems involves the study of networks from different omics such as genomics, transcriptomics, metabolomics and proteomics. In general, the computational techniques used in the analysis of biological networks can be divided into those that perform (i) structural analysis, (ii) dynamic analysis of structural prop- erties and (iii) dynamic simulation. Structural analysis is related to the study of the topology or stoichiometry of the biological network such as important nodes of the net- work, network motifs and the analysis of the flux distribution within the network. Dy- namic analysis of structural properties, generally, takes advantage from the availability of interaction and expression datasets in order to analyze the structural properties of a biological network in different conditions or time points. Dynamic simulation is useful to study those changes of the biological system in time that cannot be derived from a structural analysis because it is required to have additional information on the dynamics of the system. This thesis addresses each of these topics proposing three computational techniques useful to study different types of biological networks in which the structural and dynamic analysis is crucial to answer to specific biological questions. In particu- lar, the thesis proposes computational techniques for the analysis of the network motifs of a biological network through the design of heuristics useful to efficiently solve the subgraph isomorphism problem, the construction of a new analysis workflow able to integrate interaction and expression datasets to extract information about the chromo- somal connectivity of miRNA-mRNA interaction networks and, finally, the design of a methodology that applies techniques coming from the Electronic Design Automation (EDA) field that allows the dynamic simulation of biochemical interaction networks and the parameter estimation

Time-dependent structural transformation analysis to high-level Petri net model with active state transition diagram

<p>Abstract</p> <p>Background</p> <p>With an accumulation of <it>in silico </it>data obtained by simulating large-scale biological networks, a new interest of research is emerging for elucidating how living organism functions over time in cells.</p> <p>Investigating the dynamic features of current computational models promises a deeper understanding of complex cellular processes. This leads us to develop a method that utilizes structural properties of the model over all simulation time steps. Further, user-friendly overviews of dynamic behaviors can be considered to provide a great help in understanding the variations of system mechanisms.</p> <p>Results</p> <p>We propose a novel method for constructing and analyzing a so-called <it>active state transition diagram </it>(ASTD) by using time-course simulation data of a high-level Petri net. Our method includes two new algorithms. The first algorithm extracts a series of subnets (called <it>temporal subnets</it>) reflecting biological components contributing to the dynamics, while retaining positive mathematical qualities. The second one creates an ASTD composed of unique temporal subnets. ASTD provides users with concise information allowing them to grasp and trace how a key regulatory subnet and/or a network changes with time. The applicability of our method is demonstrated by the analysis of the underlying model for circadian rhythms in <it>Drosophila</it>.</p> <p>Conclusions</p> <p>Building ASTD is a useful means to convert a hybrid model dealing with discrete, continuous and more complicated events to finite time-dependent states. Based on ASTD, various analytical approaches can be applied to obtain new insights into not only systematic mechanisms but also dynamics.</p

Focussed MeV-Ion Micro- and Nano-Beams in the Life Sciences: Selected Applications

This work presents the development of a sub-micron nuclear microprobe for applications in the life sciences. It includes quantitative trace element analysis with sub-micron spatial resolution, 2D- and 3D-microscopy of density distributions and the targeted irradiation of living cells with counted single ions. The analytical methods base on particle induced X-ray emission spectrometry (PIXE), Rutherford backscattering spectrometry (RBS), scanning transmission ion microscopy (STIM) and STIM-tomography. The specific development of the existing nuclear microprobe LIPSION led to an improved performance of the capabilities for trace element analysis. For sub-micron analysis the spatial resolution could be improved to 300 nm at a sensitivity of about 1 µg/g for metal ions in biological matrices; for a resolution of 1 µm the sensitivity was improved to 200 ng/g (3 µmol/l). This habilitation thesis comprises a short general introduction including the motivation to utilize focussed high energy ion beams, an overview on the applications and actual research fields. The introduction is followed by the basic principles of the equipments and analytical methods. An estimation of the limits of resolution for element analytical and single ion techniques is given for the Leipzig system. Thereafter, selected studies from different research areas are presented. The first presented application is a study from environmental air pollution research. It is demonstrated that the microscopic elemental analysis of single aerosol particles can be used to assess the contributions from different sources. A further example is the analysis of the distribution of nanoparticles in skin cross-sections for a risk assessment of the applications of nanosized physical UV-filters in cosmetic products. The risk assessment is followed by the micro-analysis of trace elements, especially of bound metal ions, in brain sections on the cellular and sub-cellular level. After this the application of focussed MeV ion beams in low dose radiobiological research is presented. Finally, the analysis of 3D-density distributions by proton micro-tomography is demonstrated. A summary concludes on the applications and gives an outlook to further applications and methodological developments. The appendix comprises the relevant publications of the author.Die vorliegende Arbeit etabliert für Anwendungen in den Lebenswissenschaften den Einsatz hochfokussierter MeV-Ionenstrahlen für nuklear-mikroskopische Methoden der quantitativen Spurenelementanalyse, der 2D- und 3D-Dichtemikroskopie sowie für die gezielte Bestrahlung einzelner lebender Zellen für radiobiologische Experimente. Zur Anwendung kamen die Methoden ortsaufgelöste Protonen induzierte Röntgenfluoreszenzanalyse (particle induced X-ray emission - PIXE), Spektrometrie rückgestreuter Ionen (Rutherford backscattering spectrometry - RBS) und Rastertransmissionsionenmikroskopie (scanning transmission ion microscopy - STIM). Durch eine gezielte Weiterentwicklung des bestehenden Ionenstrahlmikroskops, der Hochenergie Ionennanosonde LIPSION, konnte die Ortsauflösung für Spurenelementanalyse auf unter 300 nm verbessert werden, beziehungsweise die Sensitivität für Metallionen in biologischen Proben auf unter 200 ng/g (3 µmol/l) bei einer Ortsauflösung von 1 µm verbessert werden. Die Habilitationsschrift umfasst eine kurze allgemeine Einleitung einschließlich der Motivation für den Einsatz fokussierter MeV-Ionenstrahlen sowie einen Überblick über die Anwendungsgebiete und aktuellen Forschungsschwerpunkte. Danach werden kurz die Grundlagen der Technik und Methoden vorgestellt, gefolgt von einer Abschätzung der A\u7fuflösungsgrenzen für Elementanalysen und Einzelionentechniken. Danach werden ausgewählte Anwendungen aus verschiedenen Forschungsgebieten vorgestellt. Das erstes Beispiel ist aus der Umweltforschung. Es wird dargestellt, wie mittels ortsaufgelöster Elementspektroskopie eine Abschätzung der Feinstaubbelastung nach Beiträgen einzelner Verursacherquellen erfolgen kann. Dann folgt als Beispiel eine ortsaufgelöste Analyse der Verteilung von Nanopartikeln aus Sonnencremes in Hautquerschnitten zur Risikoabschätzung der Anwendungen von Nanotechnologie in kosmetischen Produkten. Desweiteren werden Studien der Spurenelementverteilung, speziell der von gebundenen Metallionen, in Hirnschnitten auf zellulärer und subzellulärer Ebene erläutert. Das anschließende Beispiel erläutert die Anwendung niedriger Energiedosen in der Radiobiologie anhand des Beschusses einzelner lebender Zellen mit abgezählten einzelnen Ionen. Als letztes Beispiel wird die Anwendung hochfokussierter Ionenstrahlen für die Mikrotomographie gezeigt. Abschließend folgt eine zusammenfassende Bewertung der vorgestellten Anwendungen mit einem Ausblick auf weitere Anwendungen und methodische Entwicklungen. Der Arbeit sind die relevanten Veröffentlichungen mit Beteiligung des Autors als Anhang beigefügt

Not all questions fit in beakers - direct and indirect toxic effects of metal mixtures and the application of ecotoxicological experiments to derive better water quality standards and predict recovery after abandoned mine reclamation

2019 Fall.Includes bibliographical references.Per author permission, embargo lifted Jan.3, 2020. ac.Aqueous discharges from abandoned metal mines include complex mixtures of physical and chemical stressors. Consequently, identifying mechanisms and causal relationships between acid mine drainage (AMD) and community responses in the field is challenging. In addition to the direct toxicological effects associated with elevated concentrations of metals and reduced pH, mining activities influence aquatic organisms indirectly through physical alterations of habitat, including increased sedimentation, turbidity and substrate embeddedness. Although direct toxicity can sometimes be effectively studied in the laboratory, the indirect toxicity of toxicants rarely manifests into a measurable endpoint in the short duration and limited ecological realism of traditional laboratory toxicity experiments. The installation of a mine effluent treatment plant near Blackhawk Colorado (USA), had potential to remove the majority of aqueous metals from a mountain stream heavily degraded by Acid Mine Drainage (AMD). To investigate direct and indirect effects of Acid Mine Drainage (AMD) a series of field biomonitoring, field experiments, and mesocosm experiments were conducted. These studies quantified the relative importance of chemical (direct) and physical (indirect) stressors associated with AMD discharges and predicted recovery potential for dominant macroinvertebrate taxa. Ferric Fe is often a dominant toxicant present in AMD but is largely believed to be non-toxic to aquatic life. Results of toxicity tests reported here suggest that the current USEPA chronic Fe criterion is underprotective and that the current criterion should be reduced to 25% of its current level (251 µg/L). These studies demonstrated additional risk to aquatic insects and periphyton in metal mixtures that included ferric Fe. Responses were primarily a result of indirect physical effects associated with Fe oxide deposition rather than direct toxicity. All aquatic insects hatch as nearly microscopic organisms and small size classes were consistently the most sensitive in numerous experiments. Sampling small age classes in nature and conducting toxicity trials with small age classes is difficult and therefore these studies are lacking from the scientific literature. Failure to characterize sensitivity of early size classes may lead to gross overestimation of tolerance. Mesocosm experiments conducted using natural benthic communities provide a unique opportunity to quantify the relative importance of these indirect physical effects

Mitochondrial Iron Uptake Through Mitoferrin2 Sensitizes Human Head and Neck Squamous Carcinoma Cells to Photodynamic Therapy

Photodynamic therapy (PDT) is an FDA-approved, minimally invasive treatment modality that utilizes light in the presence of oxygen to activate photosensitizing agents to produce cell death. Phthalocyanine 4 (Pc 4), a second generation photosensitizer has shown efficacy in vitro, in vivo and in a phase I clinical trial. Pc 4 localizes primarily to mitochondria and endoplasmic reticulum, where it causes apoptotic cell death during PDT. Previously, our laboratory showed that photo sensitizers that localize to lysosomes are more effective in killing cancer cells than ones directed to mitochondria after PDT. Here, we investigated the interactions between lysosomes and mitochondria in promoting the efficiency of PDT cell killing efficiency. Three head and neck cancer cell lines (UMSCC1, UMSCC14A and UMSCC22A) were exposed to Pc 4-PDT. The 3 cell lines responded differently: UMSCC1 and UMSCC14A cells were more resistant, whereas UMSCC22A cells were more sensitive to Pc 4-PDT. In non-erythroid cells, the mitochondrial iron transporter mitoferrin2 (Mfrn2) localizes on the mitochondrial inner membrane and transports iron from the cytosol into the mitochondria. PDT-sensitive cells expressed higher Mfrn2 mRNA and protein levels compared to PDT-resistant cells. High Mfrn2 expressing cells showed higher rates of mitochondrial Fe2+ uptake compared to low Mfrn2 expressing cells. Bafilomycin, an inhibitor of the vacuolar proton pump of lysosomes and endosomes that releases lysosomal iron to the cytosol, enhanced PDT-induced cell killing of both resistant and sensitive cells. Inhibition of the divalent metal transporter 1 (DMT1) on lysosomal membranes by ferristatin markedly protected high Mfrn2 expressing cells against bafilomycin-enhanced PDT toxicity, suggesting that iron release after bafilomycin occurs via DMT1. Iron chelators and the inhibitor of the mitochondrial Ca2+ (and Fe2+) uniporter, Ru360, protected against PDT plus bafilomycin toxicity. Knockdown of Mfrn2 in UMSCC22A cells decreased the rate of mitochondrial Fe2+ uptake and delayed PDT plus bafilomycin-induced mitochondrial depolarization and cell killing. Conversely, increased expression of Mfrn2 in low Mfrn2 expressing UMSCC1 cells increased PDT plus bafilomycin-induced killing. Chloroquine, which also releases iron from lysosomes, significantly delayed tumor regrowth in high Mfm2 expressing tumors after PDT. Taken together, the data suggest that lysosomal iron release and mitochondrial iron uptake through Mfrn2 act synergistically to induce PDT-mediated and iron-dependent mitochondrial dysfunction and subsequent cell killing. Furthermore, Mfm2 expression levels in tumors might be utilized as a biomarker predicting response to PDT in head and neck cancers

Mitigation of Radiation-Induced Bone Loss by Dried Plum

In recent years, much attention has been given to the effects of radiation on bone deterioration. Past research has demonstrated that radiation acts to alter the balance between osteoblasts and osteoclasts, promoting a net osteoclastic activity in the affected bone tissue, but specific molecular mechanisms remain unknown. Gene expression of many osteoclast markers is upregulated early in response to radiation, leading to bone resorption. This problem is especially prominent in space, as astronauts are regularly exposed to full body ionizing radiation that, over extended periods of time, may lead to significant bone loss. Research suggests that radiation leads to an inflammatory response in bone tissue, which leads to oxidative stress damage and increased osteoclast activity. Numerous natural compounds have been studied in vitro and have been observed to reduce the gene expression of bone resorption genes and their protein derivatives. We have attempted to take a closer look at the mechanisms by which radiation impairs bone health by examining the microarchitecture of mouse bones and the gene expression of osteoclast, osteoblast, and osteocyte markers. Much research has been devoted to studying osteoclastic activity because this is believed to play the most influential role in bone loss. Our gene expression findings show that radiation increases bone resorption and oxidative stress. Oxidative damage analysis indicated a higher level of malondialdehyde (MDA) in the irradiated, control diet samples compared to non-irradiated mice on the control diet and suggested that dried plum may protect bones by a systemic reduction in oxidative damage. Physical characterization results that we obtained from microCT demonstrate that a dried plum diet increased the bone mass compared to the control diet, but failed to show an effect from radiation on bone. The microCT data collected is not sufficient to confirm that dried plum has a radio-protective effect in vertebrae. Although at this stage, we have limited data to fully understand the mechanisms by which dried plum protects bone, we show that dried plum can increase bone mass in vertebrae and systemically reduces MDA levels in circulation. Our research increases the current medical and biological understanding of bone physiology in response to radiation and proposed dietary countermeasures, and is of relevance to astronauts in extended space missions, cancer patients, and patients with osteoporosis

Fertilization and nutrient use efficiency in Mediterranean environments:Proceedings of the 28th International Symposium of CIEC the International Scientific Centre for Fertilizers

CIEC, the “Centre International des Engrais Chimiques” - International Scientific Centre for Fertilizers, is the oldest scientific organization solemnly dedicated to fertilizers and fertilization. Founded in 1933, CIEC is a non-profit, non-governmental organization. It has been organized as a task force with membership on invitation only, with the scope to mobilize scientistis working in the frontline of plant nutrition, soil science, fertilizers, and fertilization areas, to present and disseminate their knowledge, towards understanding the new developments in the aforementioned fields of science and technology. With 17 World Congresses, and 28 International Symposia dedicated to Fertilizers and Fertilization in its history, CIEC continues its journey through the modern concepts, and approaches, innovations of this area. The 28th International Symposium of CIEC is thematically dedicated to issues of plant nutrition, soil science, fertilizers, and fertilization, with special emphasis to Mediterranean environments. The event is hosted by the PlanTerra Institute for Plant Nutrition and Soil Quality of the Agricultural University of Athens and the Hellenic Fertilizers Association, at the Agriculture University of Athens, Greece