The intracellular second messenger InsP₃ is a vital molecule in the regulation of Ca²⁺ signalling. Ca²⁺ mediates a wide range of cellular activities from fertilisation and cell differentiation through to apoptoisis. Using X-ray crystal structure data and molecular modelling, a series of novel InsP₃ analogues were designed as selective InsP₃R-antagonists. Two novel synthetic routes have been developed for the synthesis of these analogues. The first route uses a Ferrier-II rearrangement to provide enantiopure inositol intermediates, whereas, the second route employs a diastereomeric resolution to obtain the enantiopure inositols. The successful synthesis of InsP₃ and a series of 5-position modified analogues are reported herein.

Myo-inositol is ubiquitous in nature and is found at the structural core of a diverse range of biologically important derivatives, including phosphatidylinositols, inositol phosphates and mycothiol. The synthesis of myo-inositol derivatives is notoriously difficult due to the need to control both regio- and enantioselectivity. As a result, synthetic routes to derivatives of this type are often lengthy and low yielding. The first biosynthetic step in the production of all myo-inositol metabolites is the isomerisation of D-glucose 6- phosphate to L-myo-inositol 1-phosphate as mediated by L-myo-inositol 1-phosphate synthase (INO1). For the protozoan parasite Trypanosoma brucei, INO1 is essential for survival and its version of the enzyme (TbINO1) has a high turnover. This makes TbINO1 an attractive candidate for the biocatalytic production of L-myo-inositol 1- phosphate, and a potential starting point for drastically shortened syntheses of important myo-inositol derivatives. The production of L-myo-inositol 1-phosphate by TbINO1 has been optimised to achieve complete conversion in reaction conditions that facilitate product isolation. Due to problems with an in-batch process, the TbINO1 enzyme was immobilised and the process was transferred to a flow system. This has allowed for production of significant quantities of L-myo-inositol 1-phosphate with a high level of purity. L-myo-inositol 1- phosphate obtained from the flow system has been used to prepare mycothiol glycosylation acceptor, 1,2,4,5,6-penta-O-acetyl-D-myo-inositol, in a concise synthesis with a greatly improved yield over the literature.

Lactobacillus casei BL23 expresses two enzymes encoded by the genes iolG1 and iolG2. They have been putatively assigned as myo-inositol dehydrogenases by sequence comparison. The enzyme catalyzes the reversible conversion of myo-inositol to scyllo-inosose and the concurrent reduction of NAD+ to NADH. iolG1 was subsequently determined to be a myo-inositol dehydrogenase but iolG2 was determined to be a scyllo-inositol dehydrogenase. Sequence analysis and kinetics by themselves did not provide insight as to why the enzymes are functionally different. This manuscript provides a structural rationalization for the differences in stereoisomer selectivity by X- ray crystal structure analysis and comparison. High resolution apo, binary, and ternary crystal structures for iolG1 and iolG2 wild type enzymes were determined. For iolG1 the ternary structures were determined for myo-inositol and d-chiro-inositol and for iolG2 the scyllo-inositol bound structure was determined. The high resolution structure information revealed the composition of their respective active sites and showed that subtle differences in critical amino acids for each enzyme define the orientation of the inositol stereoisomer for inline transfer of a hydride to NAD+. Mutagenesis studies of a closely related myo-inositol dehydrogenase from Bacillus subtilis were carried out. The wild type structure for BsIDH had already been determined and characterized. A portion of the results in this manuscript briefly explore structures of dehydrogenase mutants which validate the structural role of residues involved in cofactor selectivity

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal.

<p>Denna rapport avhandlar en tio veckors period på Otto Bock Scandinavia AB i Norrköping. Där analyserades en manuell testutrusning för funktionstest av myoelektroder, för att mäta muskelspänningar. Myoelektroderna används till att styra gripfunktionen hos handproteser, för patienter som förlorat en del av sin arm. Analysen ska resultera i att ge ett förslag på en automatiserad test av elektroderna. En stor del av rapporten består av studier kring hur testmetoderna fungerar och elektrodernas användning och funktion. Slutligen behandlas även ett förslag på en automatiserad test för elektroderna. </p> / <p>The present report concerns a ten weeks period at Otto Bock Scandinavia AB in Norrköping. An analyse of a manual test equipment for testing myoelectrodes, who is used to measure muscle potential in the arm. The myoelectrodes are used to control a grip function on hand prostheses, which is used by persons who has lost their lower arm. The analyse should result in a proposal of an automation of the manual test equipment for the electrodes. </p><p>A significant part of the report discusses the function of test methods and who the electrodes are used for and their function. Finally, discusses a proposal on an automated test for the electrodes.</p>

Polarized light has the ability to increase the specificity of the investigation of biomedical samples and is finding greater utilization in the fields of medical diagnostics, sensing, and measurement. In particular, this dissertation focuses on the application of polarized light to address a major obstacle in the development of an optical based polarimetric non-invasive glucose detector that has the potential to improve the quality of life and prolong the life expectancy of the millions of people afflicted with the disease diabetes mellitus. By achieving the mapping of the relative variations in rabbit corneal birefringence, it is hoped that the understanding of the results contained herein will facilitate the development of techniques to eliminate the effects of changing corneal birefringence on polarimetric glucose measurement through the aqueous humor of the eye. This dissertation also focuses on the application of polarized light to address a major downside of cardiovascular biomechanics research, which is the utilization of toxic chemicals to prepare samples for histological examination. To this end, a polarization microscopy image processing technique is applied to non-stained cardiovascular samples as a means to eliminate, for certain cardiac samples, the necessity for staining using toxic chemicals. The results from this work have the potential to encourage more investigators to join the field of cardiac biomechanics, which studies the remodeling processes responsible for cardiovascular diseases such as myocardial infarct (heart attacks) and congestive heart failure. Cardiovascular disease is epidemic, particularly amongst the population group older than 65 years, and the number of people affected by this disease is expected to increase appreciably as the baby boomer generation transitions into this older, high risk population group. A better understanding of the responsible mechanisms for cardiac tissue remodeling will facilitate the development of better prevention and treatment regimens by improving the early detection and diagnosis of this disease.

myo-lnositol (mIns), a natural component of the human diet and essential precursor of several signalling pathways, including that of G protein-coupled receptors, has also been shown to be effective in the treatment of psychiatric disorders such as depression, obsessive compulsive disorder and panic disorder. Most likely since mlns is a simple isomer of glucose, no serious side effects have been reported with its use, even at high oral doses of mlns. Previous studies suggest that the therapeutic action of mlns may include reduced serotonin 5HTzA and muscarinic acetylcholine receptor function. An important signal transduction system that may possibly be involved in the mechanism of action of antidepressants is phosphoinositide (PI) turnover. In this signalling system PI-phospholipase C (PLCpl), that is implicated in the in the mechanism of action of antidepressants and anxiolytics, is activated. The mechanism of action of mlns, however, still remains elusive and needs further investigation. In this study a possible modulatory role of 24-hour pre-treatment of human neuroblastoma cell line (SH-SY5Y) with mlns on mRNA levels and protein expression of phospholipase C-p1 (PLCP1) and glycogen synthase kinase 3P (GSK3p) was investigated. The effects of mlns were also compared to that of other prototype antidepressants, such as fluoxetine (a selective serotonin reuptake inhibitor), imipramine (a tricyclic antidepressant), lithium and another drug with potential antidepressant effects, sildenafil (phosphodiesterase 5-type (PDE5) inhibitor). Real-time reverse transcription Polymerase Chain Reaction (RTPCR) was performed in order to investigate the mRNA levels, while protein expression in membranes and the cytosol fraction of cells were quantified with Western blots. The expression of PLCPl was decreased after pre-treatments with imipramine or myoinositol in combination with fluoxetine. In addition, sildenafil alone or in combination with myo-inositol, also decreased the expression of membrane-bound PLCp1. However, a 24- hour pre-treatment with lithium did not alter PLCPl expression significantly. Determined mRNA levels for the expression of PLCPl were consistent in these findings, except for the inhibition of the mRNA for the expression of PLCPl also after lithium treatment. The reduced PLCpl mRNA levels after lithium pre-treatment may suggest the involvement of posttranscriptional modification (or delayed translational effects) of PLCpl after lithium treatment. The data from the current study suggest that antidepressant action may include downregulation of PLCPl expression and that modulators of the nitric oxidecGMP pathway (e.g. sildenafil as a PDE5 inhibitor) may exhibit similar properties. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2005.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

myo-lnositol (mIns), a natural component of the human diet and essential precursor of several signalling pathways, including that of G protein-coupled receptors, has also been shown to be effective in the treatment of psychiatric disorders such as depression, obsessive compulsive disorder and panic disorder. Most likely since mlns is a simple isomer of glucose, no serious side effects have been reported with its use, even at high oral doses of mlns. Previous studies suggest that the therapeutic action of mlns may include reduced serotonin 5HTzA and muscarinic acetylcholine receptor function. An important signal transduction system that may possibly be involved in the mechanism of action of antidepressants is phosphoinositide (PI) turnover. In this signalling system PI-phospholipase C (PLCpl), that is implicated in the in the mechanism of action of antidepressants and anxiolytics, is activated. The mechanism of action of mlns, however, still remains elusive and needs further investigation. In this study a possible modulatory role of 24-hour pre-treatment of human neuroblastoma cell line (SH-SY5Y) with mlns on mRNA levels and protein expression of phospholipase C-p1 (PLCP1) and glycogen synthase kinase 3P (GSK3p) was investigated. The effects of mlns were also compared to that of other prototype antidepressants, such as fluoxetine (a selective serotonin reuptake inhibitor), imipramine (a tricyclic antidepressant), lithium and another drug with potential antidepressant effects, sildenafil (phosphodiesterase 5-type (PDE5) inhibitor). Real-time reverse transcription Polymerase Chain Reaction (RTPCR) was performed in order to investigate the mRNA levels, while protein expression in membranes and the cytosol fraction of cells were quantified with Western blots. The expression of PLCPl was decreased after pre-treatments with imipramine or myoinositol in combination with fluoxetine. In addition, sildenafil alone or in combination with myo-inositol, also decreased the expression of membrane-bound PLCp1. However, a 24- hour pre-treatment with lithium did not alter PLCPl expression significantly. Determined mRNA levels for the expression of PLCPl were consistent in these findings, except for the inhibition of the mRNA for the expression of PLCPl also after lithium treatment. The reduced PLCpl mRNA levels after lithium pre-treatment may suggest the involvement of posttranscriptional modification (or delayed translational effects) of PLCpl after lithium treatment. The data from the current study suggest that antidepressant action may include downregulation of PLCPl expression and that modulators of the nitric oxidecGMP pathway (e.g. sildenafil as a PDE5 inhibitor) may exhibit similar properties. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2005.

Denna rapport avhandlar en tio veckors period på Otto Bock Scandinavia AB i Norrköping. Där analyserades en manuell testutrusning för funktionstest av myoelektroder, för att mäta muskelspänningar. Myoelektroderna används till att styra gripfunktionen hos handproteser, för patienter som förlorat en del av sin arm. Analysen ska resultera i att ge ett förslag på en automatiserad test av elektroderna. En stor del av rapporten består av studier kring hur testmetoderna fungerar och elektrodernas användning och funktion. Slutligen behandlas även ett förslag på en automatiserad test för elektroderna. / The present report concerns a ten weeks period at Otto Bock Scandinavia AB in Norrköping. An analyse of a manual test equipment for testing myoelectrodes, who is used to measure muscle potential in the arm. The myoelectrodes are used to control a grip function on hand prostheses, which is used by persons who has lost their lower arm. The analyse should result in a proposal of an automation of the manual test equipment for the electrodes. A significant part of the report discusses the function of test methods and who the electrodes are used for and their function. Finally, discusses a proposal on an automated test for the electrodes.