Increasing antibiotic resistance has created a need to develop rapid and reliable methods to identify bacteria and provide pertinent information to ensure suitable antibiotics or sugar therapeutics can be chosen for treatment. Carbohydrate structures attached to proteins on host cell surfaces provide a binding point for many pathogens, including bacteria. These structures can be mimicked using single monosaccharides glycosylated to alpha-hemolysin (alpha-HL). Alpha-HL is a beta-barrel pore-forming toxin secreted by Staphylococcus aureus that forms an SDS stable heptamer, which can be expressed by coupled in vitro transcription and translation and purified by polyacrylamide gel electrophoresis. The purified heptamers can be reconstituted into planar lipid bilayers and studied at the single channel level. Through single channel recordings the effects of sugar-linker lengths, different glycans and the interaction between the ‘Glycopore’ and sugar binding molecules can be studied. The glycopore, therefore, acts as a scaffold for analysing protein-sugar interactions. Studies in this thesis have focused on the synthesis of carbohydrates for site-selective protein glycosylation; cloning and in vitro transcription translation of alpha-HL monomers; and glycosylation and oligomerisation of alpha-HL to form glycopores suitable for lectin-binding studies. Lectins DC-SIGN and FimH have been expressed in Escherichia coli and these lectins as well as others have been screened using alpha-HL glycopores. The glycopores have also been investigated with bacteria in serum in a controlled molecule-specific manner using single-channel electrical recording. In this work glycosylated alpha-HL-monomers have been found to form stable heptamers which can be formed by oligomerisation on red blood cell membranes. The purified glycopores were reconstituted into planar lipid bilayers and studied at the single-channel level. Through single-channel recordings an optimised glycopore has been shown to be effective in distinguishing lectins alone and in a mixture and has afforded qualitative and quantitative information about the binding interactions between carbohydrates and sugar binding proteins. Furthermore, the glycopore has been used to sense bacteria which may provide an insight into modes of bacterial infection. In addition, a multivalent glycopore has been formed which has proved preliminary information about the effects of multivalency in lectin binding. The design and synthesis of non-beta-lactam antibiotic candidates and their evaluation has also been carried out.

Careful treatment of water molecules in ligand-protein interactions is required in many cases if the correct binding pose is to be identified for molecular docking. Water can form complex bridging networks and can play a critical role in dictating the binding mode of ligands. A particularly striking example of this can be found in the ionotropic glutamate receptors (iGluRs), a family of ligand gated ion channels that are responsible for a majority of the fast synaptic neurotransmission in the central nervous system that are thought to be essential in memory and learning. Thus, pharmacological intervention at these neuronal receptors is a valuable therapeutic strategy. This thesis relies on various computational studies and X-ray crystallography to investigate the role of ligand-water mediated interactions in iGluRs bound to glutamate and α-amino-3-hydroxy-5-methyl-4- isoxazole-propionic acid (AMPA). Comparative molecular dynamics (MD) simulations of each subtype of iGluRs bound to glutamate revealed that crystal water positions were reproduced and that all but one water molecule, W5, in the binding site can be rearranged or replaced with water molecules from the bulk. Further density functional theory calculations (DFT) have been used to confirm the MD results and characterize the energetics of W5 and another water molecule implicated in influencing the dynamics of a proposed switch in these receptors. Additional comparative studies on the AMPA subtypes of iGluRs show that each step of the calculation must be considered carefully if the results are to be meaningful. Crystal structures of two ligands, glutamate and AMPA revealed two distinct modes of binding when bound to an AMPA subtype of iGluRs, GluA2. The difference is related to the position of water molecules within the binding pocket. DFT calculations investigated the interaction energies and polarisation effects resulting in a prediction of the correct binding mode for glutamate. For AMPA alternative modes of binding have similar interaction energies as a result of a higher internal energy than glutamate. A combined MD and X-ray crystallographic study investigated the binding of the ligand AMPA in the AMPA receptor subtypes. Analysis of the binding pocket show that AMPA is not preserved in the crystal bound mode and can instead adopt an alternative mode of binding. This involves a displacement of a key water molecule followed by AMPA adopting the pose seen by glutamate. Thus, this thesis makes use of various studies to assess the energetics and dynamics of water molecules in iGluRs. The resulting data provides additional information on the importance of water molecules in mediating ligand interactions as well as identifying key water molecules that can be useful in the de novo design of new selective drugs against iGluRs.

This thesis explores roles of 2-oxoglutarate-dependent (2OG) oxygenases as interfaces that modulate steps in the flow of genetic information in cells in response to oxygen availability. Chapter 1 introduces mechanistic, biochemical and physiological aspects of major subfamilies of 2OG oxygenases, and their established regulatory roles in cells. In addition, structural and functional aspects of the ribosome and the translation process are discussed, with a focus on post-translational ribosome modifications. Chapter 2 investigates histone demethylases, which mediate chromatin-dependent regulation of gene expression and provides proof-of-concept for the rational, structure-guided design of small-molecules for selective inhibition of 2OG oxygenases with roles in cancer and inflammatory disease. Chapter 3 suggests regulatory roles for ten-eleven-translocation (TET)- catalysed DNA hydroxylation; calorimetric and thermal analyses reveal a duplex-stabilizing effect of the epigenetic 5-methylcytosine mark that is reversed upon conversion to 5- hydroxymethylcytosine (also termed the ‘sixth’ DNA base), raising the possibility that 2OG oxygenase catalysis might affect transcription via biophysical effects. Chapter 4 investigates fluoride release assays as a technology to enable medicinal chemistry studies on 2OG oxygenases with roles in fat mass regulation and obesity, cancer and inflammation; studies on the ALKBH5 enzyme show that it is a hypoxically upregulated 2OG oxygenase with a substrate preference distinct from previously characterized ALKBH enzymes. Chapter 5 identifies OGFOD1 as a 2OG-dependent ribosomal protein hydroxylase. OGFOD1 catalysis is conserved from yeast to humans. OGFOD1 catalyses formation of trans-3- hydroxy-L-proline in a highly conserved loop of ribosomal protein S23 proximal to the ribosomal decoding centre, possibly to modulate the interactions of eukaryotic ribosomes with tRNA, mRNA and translation factors in an oxygen-dependent manner. OGFOD1 is the functionally most well-conserved protein-modifying 2OG oxygenase; likewise, ribosomal protein S23 hydroxylation is the most well-conserved post-translational ribosome modification in eukaryotes. Some cell lines require OGFOD1 for proliferation, and scaffolds for OGFOD1- selective inhibitors are developed for use as potential antiproliferative agents and probes for cellular function. Chapter 6 shows the development of assays to investigate whether OGFOD1 catalysis affects ribosome assembly and function, including processivity, accuracy of initiation, elongation and termination, in yeast and mammalian cell lines. Chapter 7 concludes that ribosome hydroxylation might present an additional layer of regulatory complexity by which 2OG oxygenases could enable cells to respond to fluctuating oxygen levels.

The first identfied 2-oxoglutarate (2OG) dependent oxygenase was a collagen modifying enzyme in the work by Hutton et al. in 1967. Subsequent work has revealed that 2OG dependent oxygenases are a large family with diverse biological roles. With small molecule substrates, these enzymes catalyse a wide range of oxidative reactions, including those that form part of antibiotic biosynthetic pathways. The currently accepted consensus mechanism for catalysis by 2OG-dependent oxygenases is based on crystallographic data, kinetics and on quantum chemical calculations. The consensus mechanism involves oxidative decarboxylation of 2OG by reaction with an oxygen molecule producing CO₂, succinate and a reactive oxidising species that reacts with the 'prime' substrate. Deacetoxycephalosporin C synthase (DAOCS) is a 2OG-dependent oxygenase involved in cephalosporin biosynthesis. The mechanism of DAOCS is of particular interest because it has recently been proposed to be different from the consensus mechanism. The new mechanism proposal from Valeg ard et al. is primarily based on high-resolution crystallographic data with support from steady-state kinetic experiments and quantum-chemical calculations. The work in discussed in this thesis aimed to test the proposal of Valegård et al. by using a combination of spectroscopic and spectrometric methods analysing enzyme-substrate interactions. Substrate binding was investigated using both protein-observe (Chapter 3) and ligand-observe (Chapter 4.1 and 4.2) methods. Preliminary UV-visible data on enzyme-substrates complex formation was also obtained. The strength of substrate and cosubstrate binding was characterised through dissociation constant measurement. An activity assay (Chapter 2) that allows for direct and simultaneous monitoring of 2OG decarboxylation and penicillin ring expansion was optimised. Both the ligand-observe and protein-observe binding experiments as well as the preliminary UV-visible data indicate that the formation of a ternary complex between DAOCS, 2OG and the penicillin substrate is viable. The activity assay conclusively showed that in the presence of unnatural substrates, such as penicillin G, 2OG oxidation is significantly uncoupled from penicillin oxidation. Uncoupled turnover does not occur in the presence of the natural substrate, penicillin N, which is an aspect that should be considered in the analysis of the steady-state kinetic data. Overall, the results provide evidence that, the consensus mechanism for 2OG-dependent oxygenases is viable for DAOCS, at least in the presence of the natural substrate, penicillin N. It is possible that in the presence of an unnatural substrate, the catalytic process undergoes a more complex mechanism, possibly with the direct involvement of reducing agents in the system.

Dans cette thèse, nous avons développé des outils pour simuler des imageslongitudinales réalistes de cerveau présentant de l’atrophie ou de lacroissance. Cette méthode a été spécifiquement élaborée pour simuler leseffets de la maladie d’Alzheimer sur le cerveau. Elle se fonde sur un modèle dedéformation du cerveau qui décrit les effets biomécaniques d’une perte detissue due à une carte d’atrophie prescrite. Nous avons élaboré une méthodepour interpoler et extrapoler les images longitudinales d’un patient en simulantdes images avec une carte d’atrophie spécifique au sujet. Cette méthode a étéutilisée pour interpoler des acquisitions temporelles d’Images par RésonnanceMagnétique (IRM) de 46 patients souffrant de la maladie d’Alzheimer. Pour cefaire, des cartes d’atrophie sont estimées pour chaque patient, d’après deuxacquisitions IRM temporelles distinctes. Les IRM cliniques présentent du bruitet des artefacts. De plus, les acquisitions longitudinales présentent desvariations d’intensité d’une image à l’autre. Nous avons donc élaboré uneméthode qui combine le modèle de déformation du cerveau, ainsi que lesdifférentes images cliniques disponibles d’un patient donné, afin de simuler lesvariations d’intensité des acquisitions longitudinale. Pour finir, les outils desimulation d’images réalistes développés au cours de cette thèse sont mis àdisposition en open-source. / This thesis develops a framework to simulate realistic longitudinal brainimages with atrophy (and potentially growth), particularly in the case ofAlzheimer's Disease (AD). The core component of the framework is a braindeformation model: a carefully designed biomechanics-based tissue loss modelto simulate the deformations having the prescribed atrophy. The thesispresents a method to interpolate or extrapolate longitudinal images of asubject by simulating images with subject-specific atrophy patterns. Themethod was used to simulate interpolated time-point Magnetic ResonanceImages (MRIs) of 46 AD patients by prescribing atrophy estimated for eachpatient from the available two time-point MRIs. Real MRIs have noise andimage acquisition artefacts, and real longitudinal images have variation ofintensity characteristics among the individual images. In this thesis, wepresent a method that uses our brain deformation model and different availableimages of a subject to add realistic variations of intensities in the syntheticlongitudinal images. Finally, the software developed during the thesis tosimulate realistic longitudinal brain images with our brain deformation modelis released open-source.

Les modèles actuels de prédiction de la sensation thermique et du confort thermique ainsi que les solutions visant à améliorer l’état de bien-être thermique des occupants d’un bâtiment sont insuffisants. Ils ne prennent pas assez en compte les différences interindividuelles de jugement thermosensoriel. Pourtant, ces différences, souvent associées à la sensibilité thermique de chaque individu, existent mais restent inexpliquées sur le plan physiologique. Ces travaux de thèse, qui se sont déroulés en deux étapes, ont pour objectif d'identifier les causes physiologiques potentielles des différences interindividuelles du ressenti thermique, à travers des expérimentations multiparamétriques basées sur des mesurées cutanées. Toutes les mesures ont été réalisées après 30 minutes d’acclimatation en environnement thermique contrôlé. La première étape, exploratoire, a permis d’analyser à la fois l’activité neurosensorielle, les propriétés thermo-vasculaires et les propriétés du film hydrolipidique cutané de deux groupes présentant des sensibilités au froid distinctes (selon leur sensation thermique déclarée). Ainsi, les expérimentations ont montré qu’il était plus pertinent d’analyser davantage les propriétés cutanées thermiques et hydriques (reliées aux mécanismes de thermorégulation) plutôt que l’activité neurosensorielle des volontaires pour caractériser les différences interindividuelles de jugement thermosensoriel. Elles ont également mis en évidence la nécessité de contrôler les facteurs non thermiques des environnements et de sélectionner rigoureusement les sujets. La deuxième étape s’est focalisée sur l’analyse des propriétés thermo-vasculaires et des propriétés du film hydrolipidique de deux groupes de sensibilité au froid. Pour cela, 13 femmes ont été confrontées à 6 environnements de températures modérées comprises entre 17°C et 30°C (avec 2 transitions chaudes et 2 transitions froides) et les groupes ont été construits à partir du degré de frilosité déclaré par les sujets. Des différences sur les paramètres cutanées ont alors pu être relevées entre les deux groupes. Le résultat le plus significatif est que les individus dits « frileux » présentent une activité microcirculatoire plus intense sur les joues avec une vasoconstriction plus forte au froid et une vasodilatation plus forte au chaud que l’autre groupe « non sensible au froid » (p=0,002 d’après le test de l’ANCOVA pour l’effet des groupes). De plus, il a été montré que l’approche multiparamétrique (introduction de variables non thermiques comme variables prédictives) ainsi que la prise en compte des sensibilités thermiques individuelles améliorent la prédiction du confort thermique surtout pour le groupe « frileux » (+ 6,4 %). / Current models for predicting thermal sensation and thermal comfort as well as the solutions to improve the state of thermal well-being of the occupants of a building are insufficient. They do not sufficiently take into account interindividual differences of thermosensory judgment. However, these differences, often associated with thermal sensitivity of each person, exist but remain unexplained physiologically. This work, divided into two stages, is intended to identify the potential physiological causes of interindividual differences of thermal feeling through multiparametric experiments based on skin measurements. All measurements were performed after 30 minutes of acclimatization in controlled environment. The exploratory phase allowed to analyze both neurosensory activity, thermo-vascular properties and properties of the skin hydrolipidic film of two groups with different cold sensitivities (depending on their declared thermal sensation). For example, experiments have shown that it was more appropriate to analyse thermal and hydric skin properties (related to thermoregulation mechanisms) rather than neurosensory activity of volunteers to characterize interindividual differences of thermosensory judgment. They have also highlighted the need to control the non-thermal factors of environments and rigorously select subjects. The second step focused on the analysis of thermo-vascular properties and properties of the hydrolipidic film of two groups of different cold sensitivity. Thirteen women have faced in 6 environments of moderate temperatures between 17 ° C and 30 ° C (with 2 warm transitions and 2 cold transitions). Groups were built according to their degree of cold sensitivity. Differences in skin parameters have been found between the two groups. The most significant result is that cold-sensitive individuals have a more intense microcirculatory activity on cheeks with a stronger vasoconstriction in cold environments and a stronger vasodilation in warm environement than the non cold-sensitive group (p = 0. 002 according to ANCOVA test for groups effect). In addition, it has been shown that the multi-parametric approach (introduction of non-thermal parameters as predictors) as well as taking into account individual thermal sensitivities improve the prediction of thermal comfort especially for the cold-sensitive group (+ 6.4%).

Ion channels are fundamental molecules in the nervous system that catalyze the flux of ions across the cell membrane. There are mounting evidences suggesting that the kinetic properties of ion channels undergo activity-dependent changes in various pathophysiological conditions. Here such activity-dependent changes were studied in case of two different ion channels; the rat brain derived voltage-gated Na+ channel, rNav1.2 and the human background leak K+ channel, hTREK1 using the single channel patch-clamp technique. Our results on the voltage-gated Na+ channel (Chapter III) illustrated that sustained membrane depolarization, as seen in pathophysiological conditions like epilepsy, induced a defined non-linear variation in the unitary conductance, activation, inactivation and recovery kinetic properties of the channel. Signal processing tools attributed a pseudo-oscillatory nature to the non-linearity observed in the channel properties. Prolonged membrane depolarization also induced a “molecular memory” phenomenon, characterized by clustering of dwell time events and strong autocorrelation in the dwell time series. The persistence of such molecular memory was found to be dependent on the duration of depolarization. Similar plastic changes were observed in case of the hTREK1 channel in presence of saturating concentrations of agonist, trichloroethanol (TCE) (Chapter IV). TREK1 channel behaves similar to single enzyme molecules with a single binding site for the substrate K+ ion whereas TCE acts as an allosteric activator of the channel. We observed that with increasing concentration of TCE (10 M to 10 mM) the catalytic turnover rate exhibited progressive departure from monoexponential to multi-exponential distribution suggesting the presence of ‘dynamic disorder’ analogous to single enzyme molecules. In addition, we observed the induction of strong correlation in successive waiting times and flux intensities, exemplified by distinct mode switching between high and low flux activity, which implied the induction of memory in single ion channel. Our observation of such molecular memory in two different ion channels in different experimental conditions highlights the importance and generality of the phenomenon which is normally hidden under the ensemble behaviour of ion channels. In the final part of the work (chapter V) we observed strong negative cooperativity and half-of-sites saturation kinetics in the interaction of local anesthetic, lidocaine with hTREK1 channel. We also mapped the specific anesthetic binding site in the c-terminal domain of the channel. Further, single channel analysis and the heterodimer studies enabled us to propose a model for this interaction and provide a plausible paradigm for the inhibitory action of lidocaine on hTREK1.

Processes during the meroplanktonic phase regulate population dynamics for many marine benthic invertebrates. I examined changes in vertical distribution of different meroplanktonic larvae in a coastal embayment during a stable period, at high temporal frequencies and spatial resolutions. Plankton samples were collected at 6 depths (3, 6, 9, 12, 18, 24 m) using a pump, every 2-h over a 36- and a 25-h period, during a spring and neap tide, respectively, concurrently with measures of temperature, salinity, fluorescence and current velocity. For 10 gastropod taxa, larval vertical distribution was mostly related to the thermal structure of the water column. Each of 7 taxonomic groups was found either exclusively near the surface, associated with the fluorescence maximum, or showed diel changes in distribution. These larvae that occupy different depths in the water column exhibit different dispersal potentials. / Biogeographical data contained in this thesis will be submitted to the Oceanographic Biogeographic Information System (OBIS) and may be accessed on-line at http://www.iobis.org

Dans un contexte de changement climatique global et d'évolution locale de l'usage des terres, le devenir des paysages culturels des Alpes françaises, façonnés au cours des siècles par les interactions mutuelles entre sociétés et environnement, apparaît incertain. Dans le même temps, les écosystèmes qui les constituent abritent une biodiversité riche et sont à l'origine de nombreuses ressources naturelles et fonctions écologiques dont bénéficient les populations humaines. Ces ressources et fonctions sont conceptualisées sous le terme de « services écosystémiques » et font aujourd'hui l'objet d'une attention accrue dans la gestion et la protection des ressources environnementales, au même titre que la biodiversité. L'identification des facteurs liés à leur maintien, en termes écologiques, socio-culturels et politiques, est une étape nécessaire à leur gestion durable, bien qu'encore insuffisamment explorée. Mon projet de thèse visait à accroître la compréhension des interactions positives (synergies) et négatives (antagonismes) entre services écosystémiques et biodiversité via une approche multidimensionnelle du socio-écosystème des Alpes françaises. - Le Chapitre I propose une approche biophysique quantitative et spatialisée de la multifonctionnalité des écosystèmes. Suite à une étape de modélisation, les patrons spatiaux de synergie et d'antagonisme entre services et biodiversité ont été explorés statistiquement et reliés à des enjeux de gouvernance actuels à différentes échelles. Ce travail a permis d'identifier les bouquets de services écosystémiques représentatifs des différentes conditions biogéographiques, de gestion et de d'hétérogénéité du paysage représentées dans le massif. - Cette approche est complétée dans le Chapitre II par une représentation qualitative des relations d'influence entre services écosystémiques et biodiversité, ainsi que de leurs liens avec d'autres variables écologiques et sociales. Nous avons considéré explicitement les dimensions multiples englobées par le concept de service écosystémique (leurs ‘facettes') et proposons un cadre conceptuel pour en cartographier les réseaux d'influence. Ce cadre a servi de base à l'analyse d'un processus consultatif que nous avons mené auprès d'acteurs du territoire. Les analyses ont mis en lumière leur perception globale des relations d'influence importantes au sein du socio-écosystème. - Afin de mieux comprendre les régulations sociales appliquées à la gestion environnementale, nous testons dans le Chapitre III une méthodologie d'analyse de l'efficacité environnementale d'instruments de gouvernance. Notre analyse a privilégié un nombre restreint d'instruments qui encadrent actuellement les interactions entre agriculture, tourisme et biodiversité. L'utilisation d'un ensemble d'indicateurs de performance et d'adéquation avec le cadre socio-culturel et de gouvernance a souligné l'articulation complexe des instruments entre eux et a abouti à la production d'une synthèse pour les décideurs (‘policy brief'). - Le Chapitre IV explore enfin certains enjeux conceptuels et éthiques de la recherche dans le domaine des services écosystémiques. Après une réflexion générale sur les relations entre science et société, je propose une évaluation réflexive et personnelle des projets de recherche auxquels j'ai contribué. Pour conclure, je propose une vision transversale du socio-écosystème alpin mettant en lumière les enjeux majeurs identifiés par les différentes analyses. / In the context of global climate change and local land use changes, the future of the French Alps cultural landscapes, shaped through long-lasting and mutual interactions between human and their environment, appears uncertain. Simultaneously, the ecosystems constituting alpine landscapes host a rich biodiversity and provide the many natural resources and ecological functions that benefit to human societies. These resources and functions are conceptualised as “ecosystem services” and currently attract an increasing attention for the management and the conservation of environmental resources, along with biodiversity. Identifying the variables linked to their maintenance, in ecological, socio-cultural and political terms, is a necessary step of their sustainable management, and yet is still under-explored. My PhD project aimed at increasing the understanding of positive (synergies) and negative (trade-offs) interactions among ecosystem services and biodiversity through a multi-dimensional approach of the French Alps social-ecological system. - In Chapter I, I present a quantitative and spatially explicit biophysical assessment of ecosystem multifunctionality. After a modelling step, we explored spatial patterns of trade-offs and synergies among ecosystem services and biodiversity using a series of statistical analyses of increasing complexity. Results were structured to provide insights for sound environmental governance at multiple scales. We identified various bundles of ecosystem services representative of the different conditions across the French Alps massif in terms of biogeography, management and landscape heterogeneity. - This approach is complemented in Chapter II by a qualitative representation of influence relationships among ecosystem services and biodiversity that also accounts for additional ecological and social variables. We explicitly considered the multiple dimensions encompassed by the ecosystem service concept (their ‘facets') and proposed an innovative conceptual framework to represent their influence networks. This framework was applied to analyse a consultative process that we carried out with stakeholders of regional expertise. This analysis highlighted their general perception of important influence relationships in the alpine social ecological system. - In order to better understand social regulations linked to environmental governance, we test in Chapter III a methodology for assessing the environmental effectiveness of policy instruments. We concentrated on a restricted set of instruments regulating the interactions between biodiversity, agriculture and outdoor tourism. The consideration of multiple indicators assessing the performance and the fit with the socio-cultural and governance setting highlighted the complex articulation of instruments within the broader policy mix. Results were synthesised in a policy brief targeting regional decision-makers. - Chapter IV is conceived as my personal exploration of the conceptual and ethical issues linked to research on ecosystem services. Following some general thinking on the relations between environmental sciences and society, I conducted a personal reflexive assessment of the research projects I contributed to. To conclude, I propose a synthetic vision of the alpine social-ecological system and discuss the major issues revealed throughout the analyses.

Healthy bone is bombarded with many different mechanical strain derived signals during normal daily activities. One of these signals is present as a direct connective tissue strain on the cells. However, there is also the presence of an electrically charged streaming potential during this straining. The electrical potential is created from the movement of charged fluid through the small bone porosities. To date, little focus has been applied to elucidating the possible synergistic effects of these two stimulants. The aim of this project was to evaluate the effects of mechanical strain and indirect electrical stimulation upon the development of bone forming osteoblast cells and any possible synergistic effects of the two stimulants. This aim was achieved by using a novel device, designed and developed with the capability of creating a cell substrate surface strain along with an exogenous electrical stimulant individually or at the same time. Proliferation and differentiation were determined as a measure of cellular development. The indirect electrical stimulation was achieved through the use of a pulsed electromagnetic field (PEMF) while the mechanical strain was produced from dynamic stretching of a deformable cell substrate. Strain and strain rate were modelled from recent studies proposing that relatively high frequency, low strain osteogenic mechanical stimulants are more indicative of what healthy bone would be experiencing during normal activities. The PEMF signal mimicked a clinically available bone growth stimulator signal. Results showed a PEMF stimulus on monolayers of SaOS-2 and MG-63 osteoblast-like cells leads to a depression in proliferation. A concomitant increase in alkaline phosphatase production was also observed for the SaOS-2 cultures, but not for the MG-63 cell line. It was hypothesised that this was due to the MG-63's lack of phenotypic maturity compared to the SaOS-2 cells. Mechanical strain of the cell substrate alone, at a relatively high frequency (5Hz) but small strain, did not significantly effect either cell proliferation or differentiation for the MG-63 cells. However, when the electrical and mechanical stimulants were combined a significant increase in cellular differentiation occurred with MG-63 cultures, revealing a possible synergistic effect of these two stimulants on the development of bone cells.