Ionic and molecular diffusion in cementitious materials

The work described in this thesis is an attempt to provide improved understanding of the effects of several factors affecting diffusion in hydrated cement pastes and to aid the prediction of ionic diffusion processes in cement-based materials. Effect of pore structure on diffusion was examined by means of comparative diffusion studies of quaternary ammonium ions with different ionic radii. Diffusivities of these ions in hydrated pastes of ordinary portland cement with or without addition of fly ash were determined by a quasi-steady state technique. The restriction of the pore geometry on diffusion was evaluated from the change of diffusivity in response to the change of ionic radius. The pastes were prepared at three water-cement ratios, 0.35, 0.50 and 0.65. Attempts were made to study the effect of surface charge or the electrochemical double layer at the pore/solution interface on ionic diffusion. An approach was to evaluate the zeta potentials of hydrated cement pastes through streaming potential measurements. Another approach was the comparative studies of the diffusion kinetics of chloride and dissolved oxygen in hydrated pastes of ordinary portland cement with addition of 0 and 20% fly ash. An electrochemical technique for the determination of oxygen diffusivity was also developed. Non-steady state diffusion of sodium potassium, chloride and hydroxyl ions in hydrated ordinary portland cement paste of water-cement ratio 0.5 was studied with the aid of computer-modelling. The kinetics of both diffusion and ionic binding were considered for the characterization of the concentration profiles by Fick's first and second laws. The effect of the electrostatic interactions between ions on the overall diffusion rates was also considered. A general model concerning the prediction of ionic diffusion processes in cement-based materials has been proposed.

A molecular dynamics model of the atomic structure of dysprosium alumino-phosphate glass

Molecular dynamics (MD) has been used to identify the relative distribution of dysprosium in the phosphate glass DyAl0.30P3.05O9.62. The MD model has been compared directly with experimental data obtained from neutron diffraction to enable a detailed comparison beyond the total structure factor level. The MD simulation gives Dy ... Dy correlations at 3.80(5) and 6.40(5) angstrom with relative coordination numbers of 0.8(1) and 7.3(5), thus providing evidence of minority rare-earth clustering within these glasses. The nearest neighbour Dy-O peak occurs at 2.30 angstrom with each Dy atom having on average 5.8 nearest neighbour oxygen atoms. The MD simulation is consistent with the phosphate network model based on interlinked PO4 tetrahedra where the addition of network modifiers Dy3+ depolymerizes the phosphate network through the breakage of P-(O)-P bonds whilst leaving the tetrahedral units intact. The role of aluminium within the network has been taken into explicit account, and A1 is found to be predominantly (78 tetrahedrally coordinated. In fact all four A1 bonds are found to be to P (via an oxygen atom) with negligible amounts of Al-O-Dy bonds present. This provides an important insight into the role of Al additives in improving the mechanical properties of these glasses.

Inhibition of pyruvate kinase M2 by reactive oxygen species contributes to the development of pulmonary arterial hypertension

Aims: Pulmonary arterial hypertension [1] is a proliferative disorder associated with enhanced proliferation and suppressed apoptosis of pulmonary artery smooth muscle cells (PASMCs). Reactive oxygen species (ROS) is implicated in the development of PAH and regulates the vascular tone and functions. However, which cellular signaling mechanisms are triggered by ROS in PAH is still unknown. Hence, here we wished to characterize the signaling mechanisms triggered by ROS. Methods and Results: By Western blots, we showed that increased intracellular ROS caused inhibition of the glycolytic pyruvate kinase M2 (PKM2) activity through promoting the phosphorylation of PKM2. Monocrotaline (MCT)-induced rats developed severe PAH and right ventricular hypertrophy, with a significant increase in the P-PKM2 and decrease in pyruvate kinase activity which could be attenuated with the treatments of PKM2 activators, FBP and l-serine. The antioxidant NAC, apocynin and MnTBAP had the similar protective effects in the development of PAH. In vitro assays confirmed that inhibition of PKM2 activity could modulate the flux of glycolytic intermediates in support of cell proliferation through the increased pentose phosphate pathway (PPP). Increased ROS and decreased PKM2 activity also promoted the Cav1.2 expression and intracellular calcium. Conclusion: Our data provide new evidence that PKM2 makes a critical regulatory contribution to the PAHs for the first time. Decreased pyruvate kinase M2 activity confers additional advantages to rat PASMCs by allowing them to sustain anti-oxidant responses and thereby support cell survival in PAH. It may become a novel treatment strategy in PAH by using of PKM2 activators.

Monte Carlo and molecular dynamics simulations of methane in potassium montmorillonite clay hydrates at elevated pressures and temperatures

The structure and dynamics of methane in hydrated potassium montmorillonite clay have been studied under conditions encountered in sedimentary basin and compared to those of hydrated sodium montmorillonite clay using computer simulation techniques. The simulated systems contain two molecular layers of water and followed gradients of 150 barkm-1 and 30 Kkm-1 up to a maximum burial depth of 6 km. Methane particle is coordinated to about 19 oxygen atoms, with 6 of these coming from the clay surface oxygen. Potassium ions tend to move away from the center towards the clay surface, in contrast to the behavior observed with the hydrated sodium form. The clay surface affinity for methane was found to be higher in the hydrated K-form. Methane diffusion in the two-layer hydrated K-montmorillonite increases from 0.39×10-9 m2s-1 at 280 K to 3.27×10-9 m2s-1 at 460 K compared to 0.36×10-9 m2s-1 at 280 K to 4.26×10-9 m2s-1 at 460 K in Na-montmorillonite hydrate. The distributions of the potassium ions were found to vary in the hydrates when compared to those of sodium form. Water molecules were also found to be very mobile in the potassium clay hydrates compared to sodium clay hydrates. © 2004 Elsevier Inc. All All rights reserved.

Impact of co-adsorbed oxygen on crotonaldehyde adsorption over gold nanoclusters:a computational study

Crotonaldehyde (2-butenal) adsorption over gold sub-nanometer particles, and the influence of co-adsorbed oxygen, has been systematically investigated by computational methods. Using density functional theory, the adsorption energetics of crotonaldehyde on bare and oxidised gold clusters (Au , d = 0.8 nm) were determined as a function of oxygen coverage and coordination geometry. At low oxygen coverage, sites are available for which crotonaldehyde adsorption is enhanced relative to bare Au clusters by 10 kJ mol. At higher oxygen coverage, crotonaldehyde is forced to adsorb in close proximity to oxygen weakening adsorption by up to 60 kJ mol relative to bare Au. Bonding geometries, density of states plots and Bader analysis, are used to elucidate crotonaldehyde bonding to gold nanoparticles in terms of partial electron transfer from Au to crotonaldehyde, and note that donation to gold from crotonaldehyde also becomes significant following metal oxidation. At high oxygen coverage we find that all molecular adsorption sites have a neighbouring, destabilising, oxygen adatom so that despite enhanced donation, crotonaldehyde adsorption is always weakened by steric interactions. For a larger cluster (Au, d = 1.1 nm) crotonaldehyde adsorption is destabilized in this way even at a low oxygen coverage. These findings provide a quantitative framework to underpin the experimentally observed influence of oxygen on the selective oxidation of crotyl alcohol to crotonaldehyde over gold and gold-palladium alloys. © 2014 the Partner Organisations.

Complexity of classical dynamics of molecular systems. II. Finite statistical complexity of a water-Na+ system

The computational mechanics approach has been applied to the orientational behavior of water molecules in a molecular dynamics simulated water–Na + system. The distinctively different statistical complexity of water molecules in the bulk and in the first solvation shell of the ion is demonstrated. It is shown that the molecules undergo more complex orientational motion when surrounded by other water molecules compared to those constrained by the electric field of the ion. However the spatial coordinates of the oxygen atom shows the opposite complexity behavior in that complexity is higher for the solvation shell molecules. New information about the dynamics of water molecules in the solvation shell is provided that is additional to that given by traditional methods of analysis.

Molecular dynamics simulation of methane in sodium montmorillonite clay hydrates at elevated pressures and temperatures

Computer simulation has been used to study the structure and dynamics of methane in hydrated sodium montmorillonite clays under conditions encountered in sedimentary basins. Systems containing approximately one, two, three and four molecular layers of water have followed gradients of 150 bar km-1 and 30Kkm-1, to a maximum burial depth of 6 km (900 bar and 460 K). Methane is coordinated to approximately 19 oxygen atoms, of which typically 6 are provided by the clay surface. Only in the three- and four-layer hydrates is methane able to leave the clay surface. Diffusion depends strongly on the porosity (water content) and burial depth: self-diffusion coefficients are in the range 0.12 × 10-9m2s-1 for water and 0.04 × 10−9m2s−1 < D < 8.64 × 10−9m2s−1 for methane. Bearing in mind that porosity decreases with burial depth, it is estimated that maximum diffusion occurs at around 3 km. This is in good agreement with the known location of methane reservoirs in sedimentary basins.

Peptide binding to HLA-DP proteins at pH 5.0 and pH 7.0:a quantitative molecular docking study

Background: HLA-DPs are class II MHC proteins mediating immune responses to many diseases. Peptides bind MHC class II proteins in the acidic environment within endosomes. Acidic pH markedly elevates association rate constants but dissociation rates are almost unchanged in the pH range 5.0 - 7.0. This pH-driven effect can be explained by the protonation/deprotonation states of Histidine, whose imidazole has a pKa of 6.0. At pH 5.0, imidazole ring is protonated, making Histidine positively charged and very hydrophilic, while at pH 7.0 imidazole is unprotonated, making Histidine less hydrophilic. We develop here a method to predict peptide binding to the four most frequent HLA-DP proteins: DP1, DP41, DP42 and DP5, using a molecular docking protocol. Dockings to virtual combinatorial peptide libraries were performed at pH 5.0 and pH 7.0. Results: The X-ray structure of the peptide - HLA-DP2 protein complex was used as a starting template to model by homology the structure of the four DP proteins. The resulting models were used to produce virtual combinatorial peptide libraries constructed using the single amino acid substitution (SAAS) principle. Peptides were docked into the DP binding site using AutoDock at pH 5.0 and pH 7.0. The resulting scores were normalized and used to generate Docking Score-based Quantitative Matrices (DS-QMs). The predictive ability of these QMs was tested using an external test set of 484 known DP binders. They were also compared to existing servers for DP binding prediction. The models derived at pH 5.0 predict better than those derived at pH 7.0 and showed significantly improved predictions for three of the four DP proteins, when compared to the existing servers. They are able to recognize 50% of the known binders in the top 5% of predicted peptides. Conclusions: The higher predictive ability of DS-QMs derived at pH 5.0 may be rationalised by the additional hydrogen bond formed between the backbone carbonyl oxygen belonging to the peptide position before p1 (p-1) and the protonated ε-nitrogen of His 79β. Additionally, protonated His residues are well accepted at most of the peptide binding core positions which is in a good agreement with the overall negatively charged peptide binding site of most MHC proteins. © 2012 Patronov et al.; licensee BioMed Central Ltd.

The role of redox signaling in the molecular mechanism of tamoxifen resistance in breast cancer.

The emergence of tamoxifen or aromatase inhibitor resistance is a major problem in the treatment of breast cancer. The molecular signaling mechanism of antiestrogen resistance is not clear. Understanding the mechanisms by which resistance to these agents arise could have major clinical implications for preventing or circumventing it. Therefore, in this dissertation we have investigated the molecular mechanisms underlying antiestrogen resistance by studying the contributions of reactive oxygen species (ROS)-induced redox signaling pathways in antiestrogen resistant breast cancer cells. Our hypothesis is that the conversion of breast tumors to a tamoxifen-resistant phenotype is associated with a progressive shift towards a pro-oxidant environment of cells as a result of oxidative stress. The hypothesis of this dissertation was tested in an in vitro 2-D cell culture model employing state of the art biochemical and molecular techniques, including gene overexpression, immunoprecipitation, Western blotting, confocal imaging, ChIP, Real-Time RT-PCR, and anchorage-independent cell growth assays. We observed that tamoxifen (TAM) acts like both an oxidant and an antioxidant. Exposure of tamoxifen resistant LCC2 cell to TAM or 17 beta-estradiol (E2) induced the formation of reactive oxidant species (ROS). The formation of E2-induced ROS was inhibited by co-treatment with TAM, similar to cells pretreated with antioxidants. In LCC2 cells, treatments with either E2 or TAM were capable of inducing cell proliferation which was then inhibited by biological and chemical antioxidants. Exposure of LCC2 cells to tamoxifen resulted in a decrease in p27 expression. The LCC2 cells exposed to TAM showed an increase in p27 phosphorylation on T157 and T187. Conversely, antioxidant treatment showed an increase in p27 expression and a decrease in p27 phosphorylation on T157 and T187 in TAM exposed cells which were similar to the effects of Fulvestrant. In line with previous studies, we showed an increase in the binding of cyclin E-Cdk2 and in the level of p27 in TAM exposed cells that overexpressed biological antioxidants. Together these findings highly suggest that lowering the oxidant state of antiestrogen resistant LCC2 cells, increases LCC2 susceptibility to tamoxifen via the cyclin dependent kinase inhibitor p27.

Multi-objective optimization of the molecular structure of refrigerants

The aim of this work was to develop a new methodology, which can be used to design new refrigerants that are better than the currently used refrigerants. The methodology draws some parallels with the general approach of computer aided molecular design. However, the mathematical way of representing the molecular structure of an organic compound and the use of meta models during the optimization process make it different. In essence, this approach aimed to generate molecules that conform to various property requirements that are known and specified a priori. A modified way of mathematically representing the molecular structure of an organic compound having up to four carbon atoms, along with atoms of other elements such as hydrogen, oxygen, fluorine, chlorine and bromine, was developed. The normal boiling temperature, enthalpy of vaporization, vapor pressure, tropospheric lifetime and biodegradability of 295 different organic compounds, were collected from open literature and data bases or estimated. Surrogate models linking the previously mentioned quantities with the molecular structure were developed. Constraints ensuring the generation of structurally feasible molecules were formulated and used in commercially available optimization algorithms to generate molecular structures of promising new refrigerants. This study was intended to serve as a proof-of-concept of designing refrigerants using the newly developed methodology.

Molecular Insights of CD4+ T Cell Differentiation, Effector Formation and Helper Function

CD4+ T cells play a crucial in the adaptive immune system. They function as the central hub to orchestrate the rest of immunity: CD4+ T cells are essential governing machinery in antibacterial and antiviral responses by facilitating B cell affinity maturation and coordinating the innate and adaptive immune systems to boost the overall immune outcome; on the contrary, hyperactivation of the inflammatory lineages of CD4+ T cells, as well as the impairments of suppressive CD4+ regulatory T cells, are the etiology of various autoimmunity and inflammatory diseases. The broad role of CD4+ T cells in both physiological and pathological contexts prompted me to explore the modulation of CD4+ T cells on the molecular level.

microRNAs (miRNAs) are small RNA molecules capable of regulating gene expression post-transcriptionally. miRNAs have been shown to exert substantial regulatory effects on CD4+ T cell activation, differentiation and helper function. Specifically, my lab has previously established the function of the miR-17-92 cluster in Th1 differentiation and anti-tumor responses. Here, I further analyzed the role of this miRNA cluster in Th17 differentiation, specifically, in the context of autoimmune diseases. Using both gain- and loss-of-function approaches, I demonstrated that miRNAs in miR-17-92, specifically, miR-17 and miR-19b in this cluster, is a crucial promoter of Th17 differentiation. Consequently, loss of miR-17-92 expression in T cells mitigated the progression of experimental autoimmune encephalomyelitis and T cell-induced colitis. In combination with my previous data, the molecular dissection of this cluster establishes that miR-19b and miR-17 play a comprehensive role in promoting multiple aspects of inflammatory T cell responses, which underscore them as potential targets for oligonucleotide-based therapy in treating autoimmune diseases.

To systematically study miRNA regulation in effector CD4+ T cells, I devised a large-scale miRNAome profiling to track in vivo miRNA changes in antigen-specific CD4+ T cells activated by Listeria challenge. From this screening, I identified that miR-23a expression tightly correlates with CD4+ effector expansion. Ectopic expression and genetic deletion strategies validated that miR-23a was required for antigen-stimulated effector CD4+ T cell survival in vitro and in vivo. I further determined that miR-23a targets Ppif, a gatekeeper of mitochondrial reactive oxygen species (ROS) release that protects CD4+ T cells from necrosis. Necrosis is a type of cell death that provokes inflammation, and it is prominently triggered by ROS release and its consequent oxidative stress. My finding that miR-23a curbs ROS-mediated necrosis highlights the essential role of this miRNA in maintaining immune homeostasis.

A key feature of miRNAs is their ability to modulate different biological aspects in different cell populations. Previously, my lab found that miR-23a potently suppresses CD8+ T cell cytotoxicity by restricting BLIMP1 expression. Since BLIMP1 has been found to inhibit T follicular helper (Tfh) differentiation by antagonizing the master transcription factor BCL6, I investigated whether miR-23a is also involved in Tfh differentiation. However, I found that miR-23a does not target BLIMP1 in CD4+ T cells and loss of miR-23a even fostered Tfh differentiation. This data indicate that miR-23a may target other pathways in CD4+ T cells regarding the Tfh differentiation pathway.

Although the lineage identity and regulatory networks for Tfh cells have been defined, the differentiation path of Tfh cells remains elusive. Two models have been proposed to explain the differentiation process of Tfh cells: in the parallel differentiation model, the Tfh lineage is segregated from other effector lineages at the early stage of antigen activation; alternatively, the sequential differentiation model suggests that naïve CD4+ T cells first differentiate into various effector lineages, then further program into Tfh cells. To address this question, I developed a novel in vitro co-culture system that employed antigen-specific CD4+ T cells, naïve B cells presenting cognate T cell antigen and BAFF-producing feeder cells to mimic germinal center. Using this system, I were able to robustly generate GC-like B cells. Notably, well-differentiated Th1 or Th2 effector cells also quickly acquired Tfh phenotype and function during in vitro co-culture, which suggested a sequential differentiation path for Tfh cells. To examine this path in vivo, under conditions of classical Th1- or Th2-type immunizations, I employed a TCRβ repertoire sequencing technique to track the clonotype origin of Tfh cells. Under both Th1- and Th2- immunization conditions, I observed profound repertoire overlaps between the Teff and Tfh populations, which strongly supports the proposed sequential differentiation model. Therefore, my studies establish a new platform to conveniently study Tfh-GC B cell interactions and provide insights into Tfh differentiation processes.

(Table 1) Organic carbon, hydrogen and oxygen index and hydrocarbons at DSDP Hole 93-603B

C2-C8 hydrocarbon concentrations (about 35 compounds identified, including saturated, aromatic, and olefinic compounds) from 27 shipboard-sealed, deep-frozen core samples of DSDP Hole 603B off the east coast of North America were determined by a gas-stripping/thermovaporization method. Total yields representing the hydrocarbons dissolved in the pore water and adsorbed on the mineral surfaces of the sediments vary from 22 to 2400 ng/g of dryweight sediment. Highest yields are measured in the two black shale samples of Core 603B-34 (hydrogen index of 360 and 320 mg/g Corg, respectively). In organic-carbon-normalized units these samples have hydrocarbon contents of 12,700 and 21,500 ng/g Corg, respectively, indicating the immaturity of their kerogens. Unusually high organic-carbonnormalized yields are associated with samples that are extremely lean in organic carbon. It is most likely that they are enriched by small amounts of migrated light hydrocarbons. This applies even to those samples with high organic-carbon contents (1.3-2.2%) of Sections 603B-28-4, 603B-29-1, 603B-49-2, and 603B-49-3, because they have an extremely low hydrocarbon potential (hydrogen index between 40 and 60 mg/g Corg). Nearly all samples were found to be contaminated by varying amounts of acetone that is used routinely in large quantities on board ship during core-cutting procedures. Therefore, 48 samples from the original set of 75 collected had to be excluded from the present study.

(Table 1) Summary of low-molecular-weight hydrocarbon concentrations, organic carbon contents, Rock-Eval pyrolysis and lithology at DSDP Leg 79 Holes

A series of core samples taken during Cruise 79 of Glomar Challenger, drilling offshore Morocco (Mazagan Plateau), is analyzed for their low-molecular-weight hydrocarbon contents. Fifty-four samples from DSDP Holes 544A, 545, 547A, and 547B, deep frozen on board immediately after recovery, are studied by a hydrogen-stripping/thermovaporization technique combined with capillary gas chromatography. Thirty-eight compounds in the C2-C8 molecular range, including saturated, olefinic, and aromatic hydrocarbons, are identified. Because of large differences in organic carbon contents, the total C2-C8 hydrocarbon concentrations vary from about 20 to 1500 ng/g dry sediment weight in the whole sample series. Organic-carbon normalized values are about 3.2 x 10**4 ng/g Corg for Lithologic Subunits IIIA and IIIB at Site 545 (Cenomanian to Aptian) and 1.0 x 10**5 ng/g Corg for Unit V at Site 547 (Cenomanian to Albian) reflecting the slightly more advanced maturity stage at the latter site. Values exceeding 10**5 ng/g Corg (Site 545) and 2 x 10**5 ng/g Corg (Site 547) are associated with samples that are very lean in organic carbon and are generally rich in carbonate. These samples are enriched by small amounts of gaseous hydrocarbons. A detailed study of individual hydrocarbon concentrations, plotted against depth, reveal additional indications for migration phenomena. At Site 547, for instance, the most mobile hydrocarbons studied (e.g., ethane) appear to migrate by diffusion or a related process from more than 700 m depth toward the surface.

The adaptive response of humans to exercise: Impact of exercise intensity, fibre-type specific responses and molecular patterns of response

In an attempt to improve the current understanding of the adaptive response to exercise in humans, this dissertation performed a series of studies designed to examine the impact of training intensity and mode on aerobic capacity and performance, fibre-type specific adaptations to training, and individual patterns of response across molecular, morphological and genetic factors. Project #1 determined that training intensity, session dose, baseline VO2max and total training volume do not influence the magnitude of change in VO2max by performing a meta-regression, and meta-analysis of 28 different studies. The intensity of training had no effect on the magnitude of increase in maximal oxygen uptake in young healthy participants, but similar adaptations were achieved with lower training doses following high intensity training. Project # 2 determined the acute molecular response, and training-induced adaptations in aerobic performance, aerobic capacity and muscle phenotype following high-intensity interval training (HIT) or endurance exercise (END). The acute molecular response (fibre recruitment and signal activation) and training-induced adaptations in aerobic capacity, aerobic performance, and muscle phenotype were similar following HIT and END. Project # 3 examined the impact of baseline muscle morphology and molecular characteristics on the training response, and if muscle adaptations are coordinated. The muscle phenotype of individuals who experience the largest improvements (high responders) were lower before training for some muscle characteristics and molecular adaptations were coordinated within individual participants. Project # 4 examined the impact of 2 different intensities of HIT on the expression of nuclear and mitochondrial encoded genes targeted by PGC-1α. A systematic upregulation of nuclear and mitochondrial encoded genes was not present in the early recovery period following acute HIT, but the expression of mitochondrial genes were coordinated at an individual level. Collectively, results from the current dissertation contribute to our understanding of the molecular mechanisms influencing skeletal muscle and whole-body adaptive responses to acute exercise and training in humans.

Molecular Characterization and Localization of the NAD(P)H Oxidase Components gp91-phox and p22-phox in Endothelial Cells

The production of reactive oxygen species (ROS) within endothelial cells may have several effects, including alterations in the activity of paracrine factors, gene expression, apoptosis, and cellular injury. Recent studies indicate that a phagocyte-type NAD(P)H oxidase is a major source of endothelial ROS. In contrast to the high-output phagocytic oxidase, the endothelial enzyme has much lower biochemical activity and a different substrate specificity (NADH.NADPH). In the present study, we (1) cloned and characterized the cDNA and predicted amino acid structures of the 2 major subunits of rat coronary microvascular endothelial cell NAD(P)H oxidase, gp91-phox and p22-phox; (2) undertook a detailed comparison with phagocytic NADPH oxidase sequences; and (3) studied the subcellular location of these subunits in endothelial cells. Although these studies revealed an overall high degree of homology (.90%) between the endothelial and phagocytic oxidase subunits, the endothelial gp91-phox sequence has potentially important differences in a putative NADPH-binding domain and in putative glycosylation sites. In addition, the subcellular location of the endothelial gp91-phox and p22-phox subunits is significantly different from that reported for the neutrophil oxidase, in that they are predominantly intracellular and collocated in the vicinity of the endoplasmic reticulum. This first detailed characterization of gp91-phox and p22-phox structure and location in endothelial cells provides new data that may account, in part, for the differences in function between the phagocytic and endothelial NAD(P)H oxidases.