Temozolomide (TMZ) is a DNA methylating agent currently used for the treatment of several types of glial tumours, including glioblastomas (GBM). While improved patient response has been observed with concomitant TMZ and radiotherapy followed by adjuvant TMZ treatment following surgical resection in GBM, increased TMZ treatment showed no benefits in overall survival. This indicates that intrinsic factors such as DNA injury and repair may be crucial to patient response. The first purpose of this research was to develop an assay to quantitatively detect N7-methylguanine (N7-MeG) and O6-methylguanine (O6-MeG) using Liquid Chromatography and Mass Spectrometry (LC/MS). We then were able to detect N7-MeG and O6-MeG in human glial tumours and normal tissue regions of 31 patients with grade II to grade IV gliomas treated neoadjuvantly with 75 mg/m2/day for 14 days prior to surgery. In addition, N7-MeG and O6-MeG levels were also assessed in tumour regions of 6 untreated control patients. In all, 114 treated tumour samples, 12 treated normal and 19 untreated tumour samples were analyzed. There were significantly higher N7-MeG and O6-MeG levels detected in treated patient tumour samples when compared to untreated tumour samples. We observed intra- and inter-patient variability in N7-MeG and O6-MeG levels in all grades. In treated tumour samples, a negative correlation was seen between N7-MeG and O6-MeG levels across all grades. Increased N7-MeG levels and decreased O6-MeG levels were observed with increased malignancy. Astrocytoma samples showed significantly higher O6-MeG/N7-MeG ratios, than oligodendrogliomas, anaplastic astrocytomas and GBM tissues. No correlation was observed between MGMT protein repair activity and N7-MeG and O6-MeG levels. Samples with methylated MGMT promoters however showed significantly higher O6-MeG and O6-MeG/N7-MeG levels when compared to samples with unmethylated promoters. These results are the first in the literature to detect N7-MeG and O6-MeG induced DNA injury in human gliomas and human normal tissues and provides an experimental paradigm to help in the determination of patient response to TMZ. / Le témozolomide (TMZ) est un agent alkylant actuellement utilisé pour traiter plusieurs types de tumeurs cérebrales, dont les glioblastomes (GBM). Suivant une résection chirurgicale de la tumeur, les patients atteints de GBM sont ensuite traités par radiothérapie standard et chimiothérapie avec le TMZ. Malgré les progrès observés chez ces patients, une augmentation de la dose de TMZ lors de la chimiothérapie n'a pas démontré d'amélioration du devenir des patients. Ceci pourrait indiquer que l'accumulation des lésions cytotoxiques et leurs réparations sont importantes pour l'évolution des tumeurs ou la survie du patient. Dans cette étude, nous avons pour objectif de développer une méthode sensible et stable pour la détection quantitative du N7-methylguanine (N7-MeG) et O6-methylguanine (O6-MeG) par chromatographie liquide et spectrométrie de masse. Chez 31 patients atteints de tumeurs gliales de grade II à IV, nous avons été capable de détecter le niveau de N7-MeG et O6-MeG dans les échantillons des substances grises, blanches et de plusieurs régions du tissus cancéreux. Ces patients ont été traités quotidiennement avec 75 mg/m2/de TMZ pendant 14 jours avant d'être opérés. Nous avons aussi détectés N7-MeG et O6-MeG dans plusieurs régions de tissus tumoraux de 6 patients non-traités. Au total, nous avons analysés au total 114 échantillons de tumeurs traitées, 12 échantillons de tissus cérébraux traités et 19 échantillons de tumeurs non-traitées. Suite à cette analyse, nous avons observé des niveaux de N7-MeG et O6-MeG supérieurs dans les échantillons des patients traités comparativement aux patients non-traités. D'importantes variations du niveau intra- et inter-patients de N7-MeG et O6-MeG ont été détectées. Dans les échantillons traités, nous avons observé une corrélation négative entre les niveaux de N7-MeG et O6-MeG. Nous avons observé qu'une croissance de la malignité est associée avec une augmentation du niveau de N7-MeG et une diminution du niveau de O6-MeG. De plus, nous avons noté que le ratio de O6-MeG/N7-MeG est significativement plus élevé dans les échantillons d'astrocytomes traités en comparaison avec les oligodendrogliomes, astrocytomes anaplastiques et GBM traités. Nous n'avons pas observé de corrélation entre le niveau de O6-MeG ou de O6-MeG/N7-MeG et l'activité réparative du MGMT. Toutefois, nous avons pu détecter des niveaux de O6-MeG et O6-MeG/N7-MeG plus élevés dans les échantillons avec un promoteur de MGMT méthylé en comparaison avec les échantillons non-méthylés. Ces résultats démontrent pour la première fois l'accumulation des lésions N7-MeG et O6-MeG due au traitement par le TMZ dans des tissus cérébraux et des tumeurs gliales.

Human cartilage does not have the capacity for parent-like regeneration; instead, following injury, there is a programmed attempt at regeneration that ultimately results in fibrocartilage formation. This lack of intrinsic repair has been attributed to the avascular state of the tissue and chondrocyte dedifferentiation towards a cell incapable of type II collagen production and with altered responsiveness to the structural and regulatory mediators within its microenvironment. For several decades, debate has existed regarding the role Transforming Growth Factor-Beta (TGF-P) plays in modulating articular cartilage. Blocking of TGF-p signaling in mice by a dominant negative type II TGF-p receptor and transgenic knockouts of Smad 3, a central mediator of TGF-p signaling, result in osteoarthritic-like phenotypes, whereas local up-regulation of TGF-p promotes cartilage healing in degenerative joint disease models. Despite TGF-p being implicated as a key player In the regulation of chondrocyte phenotype and extracellular matrix (ECM), little is known about TGF-p action in human chondrocytes. These investigations have established a critical link between variation in TGF-p receptor expression at the cell surface and TGF-p action in cartilage. In addition to the TGF-p signaling receptors, the presence of betaglycan and RIIB (a spliced variant of the type II signaling receptor) was confirmed on human chondrocytes. Moreover, the expression of three novel TGF-p receptors was identified, namely Sol RI (a soluble form of the type I receptor), Alk-l, and endoglin. These receptors formed a variety of heteromeric complexes and regulated TGF-p signaling. More importantly, RIIB and endoglin were demonstrated to regulate type II collagen levels and evidence was provided that they likely represent chondrocyte phenotypic markers. A critical link between endoglin expression, cell phenotype, TGF-~ responsiveness, and ECM was established. These results demonstrate the formation of TGF-~ receptor heteromeric complexes of various subtype composition on chondrocytes suggesting that such complexes may regulate TGF-~ signaling pathways in those cells. Cell surface TGF-~ binding proteins, acting as phenotypic markers in human cartilage, are potential modulators of complex interactions between the cell and its microenvironment. Therefore, novel TGF-~ receptors may provide an avenue to regulate the effects ofTGF-~ locally and establish new insights into cartilage regeneration or repair.

Aging is the consequence ofan array of phenotypic variations that appear to involve intrinsic or constitutional properties in all cells and systems, including qualitative and quantitative alterations in development, maturational structure and function. The aging process in bone involves a set ofchanges in bone cells differentiation, interaction and premature death. Osteoblasts are the cells most affected during the aging process in bone due to their complex mechanisms ofdifferentiation, their interaction with honnones and growth factors and their progression to apoptosis.

Multiple Organ Dysfunction Syndrome (MODS) is associated with high mortality in patients admitted to the surgical intensive care unit (SICU). MODS begins with a systemic response described as Systemic Inflammatory Response Syndrome (SIRS). Studies on SIRS patients may provide an insight into the mechanisms by which SIRS progresses to MODS. In this thesis, the interactions between circulating polymorphonuclear neutrophils (PMNs) from patients with SIRS and endothelial cells (ECs) from human umbilical veins were measured in order to elucidate the mechanism for PMN adhesion and subsequent cytotoxicity of the ECs. PMNs from patients with SIRS were compared to PMNs from pre-operative surgical patients without SIRS and with healthy control subjects, in vitro. The results showed that PMNs adherence to ECs increased progressively from healthy controls to patients with SIRS. PMN-HUVE cytotoxicity, however, did not show this trend. PMNs from SIRS patients treated with lipopolysaccharide, unlike PMNs from patients without SIRS or healthy controls, showed no increase in PMN-EC adhesion. The results also showed that EC activation with TNF-$ alpha$ and Il-1$ beta$ led to high levels of PMN-EC adhesion and cytotoxicity, whereas PMN treatment with lipopolysaccharide played a lesser role. Autologous plasma provided significant protection from PMN mediated EC damage. From this data I conclude that activation of the EC by cytokines associated with SIRS is far more important in promoting PMN-EC adhesion and subsequent cytotoxicity than PMN stimulation with lipopolysaccharide and that there are host factors in plasma that modulate this response.

A major obstacle to islet transplantation is β-cell death following isolation. Isolation exposes islets to various stresses including endoplasmic reticulum (ER) stress inducers; therefore, the role of ER stress signaling in isolated islet apoptosis was investigated. Activation of eIF2α and JNK1 and XBP1 splicing followed by an increase in caspase-3 activity were observed in isolated human islets. Since the absence of protein-tyrosine phosphatase 1B (PTP1B) was previously shown to reduce ER stress-induced signaling and apoptosis in fibroblasts, the role of PTP1B in ER stress signaling was investigated in β-cells. While encouraging data emerged, using an inhibitor and miRNA targeting PTP1B, a conclusive link between PTP1B inhibition and improved β-cell survival has not yet been seen. This study provides the first evidence that ER stress signaling may influence isolated islet apoptosis and could point to novel therapeutic approaches in islet transplantation. / Un obstacle majeur lors de la transplantation des îlots pancréatiques est la perte des cellules β lors de la procédure d'isolation. En effet, lors de ce processus, les îlots sont exposés à divers stress cellulaires incluant ceux qui induisent un stress au niveau du réticulum endoplasmique (RE). Cette étude porte donc sur la signalisation menant à l'apoptose en réponse au stress du RE sur les îlots isolés. L'activation d'eIF2α, de JNK1 et de l'épissage de XBP1 qui est suivi par une augmentation de l'activité de la caspase-3 fut observées sur des îlots isolés chez l'humain. L'absence de la protéine tyrosine phosphatase 1B (PTP1B) avait précédemment été démontrée comme pouvant contribuer à la diminution de la signalisation déclenchée par le stress du RE et l'apoptose chez les fibroblastes. Malgré des résultats encourageants concernant l'utilisation d'un inhibiteur et d'un miRNA qui ciblent PTP1B, un lien concluant entre l'inhibition de cette enzyme et l'amélioration de la survie des cellules β n'a pas été observé Cette étude fournit la première évidence qui clarifie le rôle de la signalisation induite par le stress du RE lors de l'apoptose des îlots pancréatiques. De plus, elle pourrait résulter en une nouvelle approche thérapeutique pour augmenter la survie des cellules β lors de la transplantation d'îlots.

The effects of reversible fetal tracheal occlusion (TO), and antenatal glucocorticoids on lung growth, structure, surfactant levels, and function were assessed in a lamb hypoplastic lung model of congenital diaphragmatic hernia (CDH). CDH, CDH+TO, CDH+TO+release of the tracheal occlusion one week before delivery (TR), and unoperated twin controls were compared. TO+/-TR partially normalized the hypoplastic lungs of CDH: they accelerated growth of both lungs and led to structural maturity. Only TO thinned the medial area of small pulmonary arteries closer to control values. Despite TO, TR, and glucocorticoids, lungs from lambs with CDH have dysfunctional type II cells with decreased surfactant levels. Nonetheless, CDH+TO lambs showed normal oxygenation, ventilation, and compliance over untreated CDH, with a clear survival advantage over an eight hour resuscitation. TR one week before delivery had no added benefit in terms of lung function. It appears that surfactant independent mechanisms such as pulmonary growth and structural changes are of foremost importance in relating to improved compliance, oxygenation, and ventilation of CDH+TO animals.

Context. Exacerbations of chronic obstructive lung disease (COPD) that result in acute respiratory failure are a significant health care problem with high morbidity and mortality. Recent reports have shown that preventing intubation with the use of non-invasive positive pressure ventilation significantly decreases the morbidity and mortality associated with respiratory failure in this population. / Objective. To evaluate whether patients with COPD and acute respiratory failure treated with non-invasive continuous positive airway pressure (CPAP) have improved clinical outcomes after adjusting for potentially confounding covariates when compared to similar patients who were intubated or treated with medical therapy. / Design and setting. Comparison of three retrospective cohorts, one in which CPAP was routinely available and two in which CPAP were not available for the management of this type of patient. Data were collected on all patients admitted to two ICU's in tertiary care teaching hospitals during the time periods defined by the cohorts. / Population. Two hundred and thirty-eight patients with chronic obstructive lung disease and acute respiratory failure admitted to two intensive care units between January 15th 1985 to December 31st 1995. (Abstract shortened by UMI.)

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No description available.