Synthesis, Modification, and Application of Siloxane Materials for Environmental Sensing

As human populations and resource consumption increase, it is increasingly important to monitor the quality of our environment. While laboratory instruments offer useful information, portable, easy to use sensors would allow environmental analysis to occur on-site, at lower cost, and with minimal operator training. We explore the synthesis, modification, and applications of modified polysiloxane in environmental sensing. Multiple methods of producing modified siloxanes were investigated. Oligomers were formed by using functionalized monomers, producing siloxane materials containing silicon hydride, methyl, and phenyl side chains. Silicon hydride-functionalized oligomers were further modified by hydrosilylation to incorporate methyl ester and naphthyl side chains. Modifications to the siloxane materials were also carried out using post-curing treatments. Methyl ester-functionalized siloxane was incorporated into the surface of a cured poly(dimethylsiloxane) film by siloxane equilibration. The materials containing methyl esters were hydrolyzed to reveal carboxylic acids, which could later be used for covalent protein immobilization. Finally, the siloxane surfaces were modified to incorporate antibodies by covalent, affinity, and adsorption-based attachment. These modifications were characterized by a variety of methods, including contact angle, attenuated total reflectance Fourier transform infrared spectroscopy, dye labels, and 1H nuclear magnetic resonance spectroscopy. The modified siloxane materials were employed in a variety of sensing schemes. Volatile organic compounds were detected using methyl, phenyl, and naphthyl-functionalized materials on a Fabry-Perot interferometer and a refractometer. The Fabry-Perot interferometer was found to detect the analytes upon siloxane extraction by deformation of the Bragg reflectors. The refractometer was used to determine that naphthyl-functionalized siloxanes had elevated refractive indices, rendering these materials more sensitive to some analytes. Antibody-modified siloxanes were used to detect biological analytes through a solid phase microextraction-mediated enzyme linked immunosorbent assay (SPME ELISA). The SPME ELISA was found to have higher analyte sensitivity compared to a conventional ELISA system. The detection scheme was used to detect Escherichia coli at 8500 CFU/mL. These results demonstrate the variety of methods that can be used to modify siloxanes and the wide range of applications of modified siloxanes has been demonstrated through chemical and biological sensing schemes.

Proceedings before the Metropolitan Board of Health on the application of The Metropolitan Gas Light Co. for a modification of the order of the board, concerning the manufacture of gas at the foot of West 42d Street, New York.

Mode of access: Internet.

Targeted Gene Repression Technologies for Regenerative Medicine, Genomics, and Gene Therapy

Gene regulation is a complex and tightly controlled process that defines cell function in physiological and abnormal states. Programmable gene repression technologies enable loss-of-function studies for dissecting gene regulation mechanisms and represent an exciting avenue for gene therapy. Established and recently developed methods now exist to modulate gene sequence, epigenetic marks, transcriptional activity, and post-transcriptional processes, providing unprecedented genetic control over cell phenotype. Our objective was to apply and develop targeted repression technologies for regenerative medicine, genomics, and gene therapy applications. We used RNA interference to control cell cycle regulation in myogenic differentiation and enhance the proliferative capacity of tissue engineered cartilage constructs. These studies demonstrate how modulation of a single gene can be used to guide cell differentiation for regenerative medicine strategies. RNA-guided gene regulation with the CRISPR/Cas9 system has rapidly expanded the targeted repression repertoire from silencing single protein-coding genes to modulation of genes, promoters, and other distal regulatory elements. In order to facilitate its adaptation for basic research and translational applications, we demonstrated the high degree of specificity for gene targeting, gene silencing, and chromatin modification possible with Cas9 repressors. The specificity and effectiveness of RNA-guided transcriptional repressors for silencing endogenous genes are promising characteristics for mechanistic studies of gene regulation and cell phenotype. Furthermore, our results support the use of Cas9-based repressors as a platform for novel gene therapy strategies. We developed an in vivo AAV-based gene repression system for silencing endogenous genes in a mouse model. Together, these studies demonstrate the utility of gene repression tools for guiding cell phenotype and the potential of the RNA-guided CRISPR/Cas9 platform for applications such as causal studies of gene regulatory mechanisms and gene therapy.

RNA Localization and Translational Regulation on the Endoplasmic Reticulum

mRNA localization is emerging as a critical cellular mechanism for the spatiotemporal regulation of protein expression and serves important roles in oogenesis, embryogenesis, cell fate specification, and synapse formation. Signal sequence-encoding mRNAs are localized to the endoplasmic reticulum (ER) membrane by either of two mechanisms, a canonical mechanism of translation on ER-bound ribosomes (signal recognition particle pathway), or a poorly understood direct ER anchoring mechanism. In this study, we identify that the ER integral membrane proteins function as RNA-binding proteins and play important roles in the direct mRNA anchoring to the ER. We report that one of the ER integral membrane RNA-binding protein, AEG-1 (astrocyte elevated gene-1), functions in the direct ER anchoring and translational regulation of mRNAs encoding endomembrane transmembrane proteins. HITS-CLIP and PAR-CLIP analyses of the AEG-1 mRNA interactome of human hepatocellular carcinoma cells revealed a high enrichment for mRNAs encoding endomembrane organelle proteins, most notably encoding transmembrane proteins. AEG-1 binding sites were highly enriched in the coding sequence and displayed a signature cluster enrichment downstream of encoded transmembrane domains. In overexpression and knockdown models, AEG-1 expression markedly regulates translational efficiency and protein functions of two of its bound transcripts, MDR1 and NPC1. This study reveals a molecular mechanism for the selective localization of mRNAs to the ER and identifies a novel post-transcriptional gene regulation function for AEG-1 in membrane protein expression.

Synthesis, Modification, and Application of Siloxane Materials for Environmental Sensing

As human populations and resource consumption increase, it is increasingly important to monitor the quality of our environment. While laboratory instruments offer useful information, portable, easy to use sensors would allow environmental analysis to occur on-site, at lower cost, and with minimal operator training. We explore the synthesis, modification, and applications of modified polysiloxane in environmental sensing. Multiple methods of producing modified siloxanes were investigated. Oligomers were formed by using functionalized monomers, producing siloxane materials containing silicon hydride, methyl, and phenyl side chains. Silicon hydride-functionalized oligomers were further modified by hydrosilylation to incorporate methyl ester and naphthyl side chains. Modifications to the siloxane materials were also carried out using post-curing treatments. Methyl ester-functionalized siloxane was incorporated into the surface of a cured poly(dimethylsiloxane) film by siloxane equilibration. The materials containing methyl esters were hydrolyzed to reveal carboxylic acids, which could later be used for covalent protein immobilization. Finally, the siloxane surfaces were modified to incorporate antibodies by covalent, affinity, and adsorption-based attachment. These modifications were characterized by a variety of methods, including contact angle, attenuated total reflectance Fourier transform infrared spectroscopy, dye labels, and 1H nuclear magnetic resonance spectroscopy. The modified siloxane materials were employed in a variety of sensing schemes. Volatile organic compounds were detected using methyl, phenyl, and naphthyl-functionalized materials on a Fabry-Perot interferometer and a refractometer. The Fabry-Perot interferometer was found to detect the analytes upon siloxane extraction by deformation of the Bragg reflectors. The refractometer was used to determine that naphthyl-functionalized siloxanes had elevated refractive indices, rendering these materials more sensitive to some analytes. Antibody-modified siloxanes were used to detect biological analytes through a solid phase microextraction-mediated enzyme linked immunosorbent assay (SPME ELISA). The SPME ELISA was found to have higher analyte sensitivity compared to a conventional ELISA system. The detection scheme was used to detect Escherichia coli at 8500 CFU/mL. These results demonstrate the variety of methods that can be used to modify siloxanes and the wide range of applications of modified siloxanes has been demonstrated through chemical and biological sensing schemes.

Polymer microspheres: synthesis, characterisation and post-functionalisation by olefin metathesis

Porous polymer particles are used in an extraordinarily wide range of advanced and everyday applications, from combinatorial chemistry, solid-phase organic synthesis and polymer-supported reagents, to environmental analyses and the purification of drinking water. The installation and exploitation of functional chemical handles on the particles is often a prerequisite for their successful exploitation, irrespective of the application and the porous nature of the particles. New methodology for the chemical modification of macroreticular polymers is the primary focus of the work presented in this thesis. Porous polymer microspheres decorated with a diverse range of functional groups were synthesised by the post-polymerisation chemical modification of beaded polymers via olefin cross metathesis. The polymer microspheres were prepared by the precipitation polymerisation of divinylbenzene in porogenic (pore-forming) solvents; the olefin cross-metathesis (CM) functionalisation reactions exploited the pendent (polymer-bound) vinyl groups that were not consumed by polymerisation. Olefin CM reactions involving the pendent vinyl groups were performed in dichloromethane using second-generation Grubbs catalyst (Grubbs II), and a wide range of coupling partners used. The results obtained indicate that high quality, porous polymer microspheres synthesised by precipitation polymerisation in near-θ solvents can be functionalised by olefin CM under very mild conditions to install a diverse range of chemical functionalities into a common polydivinylbenzene precursor. Gel-type polymer microspheres were prepared by the precipitation copolymerisation reaction of divinylbenzene and allyl methacrylate in neat acetonitrile. The unreacted pendent vinyl groups that were not consumed by polymerisation were subjected to internal and external olefin metathesis-based hypercrosslinking reactions. Internal hypercrosslinking was carried out by using ring-closing metathesis (RCM) reactions in toluene using Grubbs II catalyst. Under these conditions, hypercrosslinked (HXL) polymers with specific surface areas around 500 m2g-1 were synthesised. External hypercrosslinking was attempted by using CM/RCM in the presence of a multivinyl coupling partner in toluene using second-generation Hoveyda-Grubbs catalyst. The results obtained indicate that no HXL polymers were obtained. However, during the development of this methodology, a new type of polymerisation was discovered with tetraallylorthosilicate as monomer.

Effect of remote ischaemic preconditioning in cardiac dysfunction and end-organ injury following cardiac surgery with cardiopulmonary bypass in children: A translational approach investigating clinical outcome and myocardial molecular biology

Congenital heart disease (CHD) is the most common birth defect, causing an important rate of morbidity and mortality. Treatment of CHD requires surgical correction in a significant percentage of cases which exposes patients to cardiac and end organ injury. Cardiac surgical procedures often require the utilisation of cardiopulmonary bypass (CPB), a system that replaces heart and lungs function by diverting circulation into an external circuit. The use of CPB can initiate potent inflammatory responses, in addition a proportion of procedures require a period of aortic cross clamp during which the heart is rendered ischaemic and is exposed to injury. High O2 concentrations are used during cardiac procedures and when circulation is re-established to the heart which had adjusted metabolically to ischaemia, further injury is caused in a process known as ischaemic reperfusion injury (IRI). Several strategies are in place in order to protect the heart during surgery, however injury is still caused, having detrimental effects in patients at short and long term. Remote ischaemic preconditioning (RIPC) is a technique proposed as a potential cardioprotective measure. It consists of exposing a remote tissue bed to brief episodes of ischaemia prior to surgery in order to activate protective pathways that would act during CPB, ischaemia and reperfusion. This study aimed to assess RIPC in paediatric patients requiring CHD surgical correction with a translational approach, integrating clinical outcome, marker analysis, cardiac function parameters and molecular mechanisms within the cardiac tissue. A prospective, single blinded, randomized, controlled trial was conducted applying a RIPC protocol to randomised patients through episodes of limb ischaemia on the day before surgery which was repeated right before the surgery started, after anaesthesia induction. Blood samples were obtained before surgery and at three post-operative time points from venous lines, additional pre and post-bypass blood samples were obtained from the right atrium. Myocardial tissue was resected during the ischaemic period of surgery. Echocardiographic images were obtained before the surgery started after anaesthetic induction and the day after surgery, images were stored for later off line analysis. PICU surveillance data was collected including ventilation parameters, inotrope use, standard laboratory analysis and six hourly blood gas analysis. Pre and post-operative quantitation of markers in blood specimens included cardiac troponin I (cTnI) and B-type natriuretic peptide (BNP), inflammatory mediators including interleukins IL-6, IL-8, IL-10, tumour necrosis factor (TNF-α), and the adhesion molecules ICAM-1 and VCAM-1; the renal marker Cystatin C and the cardiovascular markers asymmetric dymethylarginine (ADMA) and symmetric dymethylarginine (SDMA). Nitric oxide (NO) metabolites and cyclic guanosine monophosphate (cGMP) were measured before and after bypass. Myocardial tissue was processed at baseline and after incubation at hyperoxic concentration during four hours in order to mimic surgical conditions. Expression of genes involved in IRI and RIPC pathways was analysed including heat shock proteins (HSPs), toll like receptors (TLRs), transcription factors nuclear factor κ-B (NF- κ-B) and hypoxia inducible factor 1 (HIF-1). The participation of hydrogen sulfide enzymatic genes, apelin and its receptor were explored. There was no significant difference according to group allocation in any of the echocardiographic parameters. There was a tendency for higher cTnI values and inotropic score in control patients post-operatively, however this was not statistically significant. BNP presented no significant difference according to group allocation. Inflammatory parameters tended to be higher in the control group, however only TNF- α was significantly higher. There was no difference in levels of Cystatin C, NO metabolites, cGMP, ADMA or SDMA. RIPC patients required shorter PICU stay, all other clinical and laboratory analysis presented no difference related to the intervention. Gene expression analysis revealed interesting patterns before and after incubation. HSP-60 presented a lower expression at baseline in tissue corresponding to RIPC patients, no other differences were found. This study provided with valuable descriptive information on previously known and newly explored parameters in the study population. Demographic characteristics and the presence of cyanosis before surgery influenced patterns of activity in several parameters, numerous indicators were linked to the degree of injury suffered by the myocardium. RIPC did not reduce markers of cardiac injury or improved echocardiographic parameters and it did not have an effect on end organ function; some effects were seen in inflammatory responses and gene expression analysis. Nevertheless, an important clinical outcome indicator, PICU length of stay was reduced suggesting benefit from the intervention. Larger studies with more statistical power could determine if the tendency of lower injury and inflammatory markers linked to RIPC is real. The present results mostly support findings of larger multicentre trials which have reported no cardiac benefit from RIPC in paediatric cardiac surgery.

Regulatory governance of ‘training markets’, ‘market failure’, and ‘quasi’ markets: historical dimensions of the post-initial training market in the Netherlands

This article examines regulatory governance of the post-initial training market in The Netherlands. From an historical perspective on policy formation processes, it examines market formation in terms of social, economic, and cultural factors in the development of provision and demand for post-initial training; the roles of stakeholders in the longterm construction of regulatory governance of the market; regulation of and public providers; policy responses to market failure; and tripartite division of responsibilities between the state, social partners, commercial and publicly-funded providers. Historical description and analysis examine policy narratives of key stakeholders with reference to: a) influence of societal stakeholders on regulatory decision-making; b) state regulation of the post-initial training market; c) public intervention regulating the market to prevent market failure; d) market deregulation, competition, employability and individual responsibility; and, e) regulatory governance to prevent ‘allocative failure’ by the market in non-delivery of post-initial training to specific target groups, particularly the low-qualified. Dominant policy narratives have resulted in limited state regulation of the supply-side, a tripartite system of regulatory governance by the state, social partners and commercial providers as regulatory actors. Current policy discourses address interventions on the demand-side to redistribute structures of opportunity throughout the life courses of individuals. Further empirical research from a comparative historical perspective is required to deepen contemporary understandings of regulatory governance of markets and the commodification of adult learning in knowledge societies and information economies. (DIPF/Orig.)

Bifidobacterium longum 1714 as a translational psychobiotic: modulation of stress, electrophysiology and neurocognition in healthy volunteers

The emerging concept of psychobiotics—live microorganisms with a potential mental health benefit—represents a novel approach for the management of stress-related conditions. The majority of studies have focused on animal models. Recent preclinical studies have identified the B. longum 1714 strain as a putative psychobiotic with an impact on stress-related behaviors, physiology and cognitive performance. Whether such preclinical effects could be translated to healthy human volunteers remains unknown. We tested whether psychobiotic consumption could affect the stress response, cognition and brain activity patterns. In a within-participants design, healthy volunteers (N=22) completed cognitive assessments, resting electroencephalography and were exposed to a socially evaluated cold pressor test at baseline, post-placebo and post-psychobiotic. Increases in cortisol output and subjective anxiety in response to the socially evaluated cold pressor test were attenuated. Furthermore, daily reported stress was reduced by psychobiotic consumption. We also observed subtle improvements in hippocampus-dependent visuospatial memory performance, as well as enhanced frontal midline electroencephalographic mobility following psychobiotic consumption. These subtle but clear benefits are in line with the predicted impact from preclinical screening platforms. Our results indicate that consumption of B. longum 1714 is associated with reduced stress and improved memory. Further studies are warranted to evaluate the benefits of this putative psychobiotic in relevant stress-related conditions and to unravel the mechanisms underlying such effects.

Le stress post-traumatique chez l'agresseur – intérêt pour le clinicien

Introduction : Cet article présente un cas de psychothérapie de stress post-traumatique chez l’agresseur. Ce syndrome clinique est bien décrit dans la littérature, mais notre recherche ne nous a permis de trouver que 2 rapports de cas témoignant de son traitement en psychothérapie. Le traitement de cette population présente plusieurs défis. Un effort a dû être déployé pour réviser une littérature complexe. D’autres contraintes à notre démarche touchaient l’identification d’un sujet éligible, l’adaptation du protocole de psychothérapie à ce sujet et à au contexte de la pratique clinique, ainsi que l’administration du protocole en tenant compte de contraintes organisationnelles spécifiques. Méthode : Toutes les étapes du projet ont été dirigées par le premier auteur. Une revue de la littérature sur l’état de stress post-traumatique chez l’agresseur a été réalisée sous la supervision du deuxième auteur. Elle a été présentée au congrès annuel de l’Association des Médecins Psychiatres du Québec (AMPQ) de 2011. Cette expérience a permis une sensibilisation aux manifestations de ce syndrome clinique dans la pratique psychiatrique. Grâce à cela, un sujet hospitalisé a été identifié comme présentant des symptômes suggestifs d’état de stress post-traumatique chez l’agresseur. Une psychothérapie a été planifiée et administrée, à partir des connaissances acquises lors de la revue de la littérature, sous la supervision du troisième auteur. Le sujet a été sélectionné par échantillon de convenance. Les données colligées concernant l’évolution clinique du sujet sont de nature qualitative. La méthode d’intervention est une thérapie cognitivo-comportementale intensive, chez un patient hospitalisé. Une attention particulière a été portée au vécu subjectif de l’agresseur, tel que suggéré par la littérature. Un suivi a eu lieu en clinique externe par la suite, permettant de colliger des données supplémentaires. Résultats : L’évolution du sujet a été positive, se concluant par le désir de devenir un membre productif de la société. Conclusion : Un protocole de thérapie cognitivo-comportementale visant l’état de stress post-traumatique en général peut être adapté à la situation de l’agresseur. L’exercice d’être attentif à l’expérience subjective de l’agresseur permet de dépasser certains défis que présente son traitement. La psychothérapie peut être adaptée à la situation du patient hospitalisé en psychiatrie. L’ensemble de la démarche illustrée dans cet article représente un exemple concret d’application de principes scientifiques au traitement d’un sujet dont la présentation clinique ne correspond pas à un diagnostic pour lequel il existe un traitement bien établi. Nous invitons le lecteur clinicien à aborder les défis présentés par un cas clinique en formulant une question de recherche pertinente, qui pourra être mise à l’épreuve pendant la phase de traitement du patient. De plus, nous croyons que la rédaction d’une histoire de cas, organisée en fonction de ce principe, constitue un outil didactique précieux dans la formation en psychiatrie.

Using an evidence-based online module to improve parents’ ability to manage their child with Developmental Coordination Disorder

Background : Developmental coordination disorder (DCD) is a prevalent neurodevelopmental disorder. Best practices include raising parents’ awareness and building capacity but few interventions incorporating these best practices are documented. Objective : To examine whether an evidence-based online module can increase the perceived knowledge and skills of parents of children with DCD, and lead to behavioral changes when managing their child’s health condition. Methods : A mixed-methods, before-after design guided by the theory of planned behavior was employed. Data about the knowledge, skills and behaviors of parents of children with DCD were collected using questionnaires prior to completing the module, immediately after, and three months later. Paired T-tests, sensitivity analyses and thematic analyses were performed on data as appropriate. Results: One hundred-sixteen, 81 and 58 participants respectively completed the three questionnaires. For knowledge and skills, post- and follow-up scores were significantly higher than baseline scores (p<0.01). Fifty-two (64%) participants reported an intention to change behavior post-intervention and 29 (50%) participants had tried recommended strategies at follow-up. Three themes emerged to describe parents’ behavioral change: sharing information, trialing strategies and changing attitudes. Factors influencing parents’ ability to implement these behavioral changes included clear recommendations, time, and ‘right’ attitude. Perceived outcomes associated with the parental behavioral changes involved improvement in well-being for the children at school, at home, and for the family as a whole. Conclusions : The online module increased parents’ self-reported knowledge and skills in DCD management. Future research should explore its impacts on children’s long-term outcomes.

The Coming of Post-reflexive Society: Commodification and Language in Digital Capitalism

Language is a unique aspect of human communication because it can be used to discuss itself in its own terms. For this reason, human societies potentially have superior capacities of co-ordination, reflexive self-correction, and innovation than other animal, physical or cybernetic systems. However, this analysis also reveals that language is interconnected with the economically and technologically mediated social sphere and hence is vulnerable to abstraction, objectification, reification, and therefore ideology – all of which are antithetical to its reflexive function, whilst paradoxically being a fundamental part of it. In particular, in capitalism, language is increasingly commodified within the social domains created and affected by ubiquitous communication technologies. The advent of the so-called ‘knowledge economy’ implicates exchangeable forms of thought (language) as the fundamental commodities of this emerging system. The historical point at which a ‘knowledge economy’ emerges, then, is the critical point at which thought itself becomes a commodified ‘thing’, and language becomes its “objective” means of exchange. However, the processes by which such commodification and objectification occurs obscures the unique social relations within which these language commodities are produced. The latest economic phase of capitalism – the knowledge economy – and the obfuscating trajectory which accompanies it, we argue, is destroying the reflexive capacity of language particularly through the process of commodification. This can be seen in that the language practices that have emerged in conjunction with digital technologies are increasingly non-reflexive and therefore less capable of self-critical, conscious change.