Human neutrophil elastase (HNE) is a uniquely destructive serine protease with the ability to unleash a wave of proteolytic activity by destroying the inhibitors of other proteases. Although this phenomenon forms an important part of the innate immune response to invading pathogens, it is responsible for the collateral host tissue damage observed in chronic conditions such as chronic obstructive pulmonary disease (COPD), and in more acute disorders such as the lung injuries associated with COVID-19 infection. Previously, a combinatorially selected activity-based probe revealed an unexpected substrate preference for oxidised methionine, which suggests a link to oxida-tive pathogen clearance by neutrophils. Here we use oxidised model substrates and inhibitors to confirm this observation and to show that neutrophil elastase is specifically selective for the di-oxygenated methionine sulfone rather than the mono-oxygenated methionine sulfoxide. We also posit a critical role for ordered solvent in the mechanism of HNE discrimination between the two oxidised forms methionine residue. Preference for the sulfone form of oxidised methionine is especially significant. While both host and pathogens have the ability to reduce methionine sulfoxide back to methionine, a biological pathway to reduce methionine sulfone is not known. Taken to-gether, these data suggest that the oxidative activity of neutrophils may create rapidly cleaved elas-tase “super substrates” that directly damage tissue, while initiating a cycle of neutrophil oxidation that increases elastase tissue damage and further neutrophil recruitment.

No description available.

The cancer stem cell (CSC) concept is still very controversial; therefore identification and isolation of this specific population remain challenging. A variety of putative markers have been described and measurement of high aldehyde dehydrogenase (ALDH) activity has been defined as a characteristic of stem cells (SCs). In this study, a library of novel small molecules (1,4-di-substituted acetalanthraquinones, AAQs), containing an acetal group as protected aldehyde functionality, was designed with the aim of probing affinity for ALDH metabolism and demonstrating their potential as molecular fluorescent probes to identify CSCs. The AAQs were shown to be subjective to acidic hydrolysis using 2M HCl at 37ºC; however compounds containing secondary or tertiary amine functionalities in their sidechain were only partly hydrolysed at 70 ºC. Metabolism studies were conducted using cytosolic fractions from rat liver enriched in ALDHs, yeast ALDH and human recombinant ALDH1A1. Some evidence was demonstrated which linked ALDH metabolism with aldehyde functionalities of hydrolysed AAQs (HAAQs). The AAQs were shown to emit far-red fluorescence (600-750 nm). A close relationship between structure modifications and alteration of cellular localisation, with gained specificity for selected sub-cellular compartments were achieved when assessed in A549 and U-2 OS cell lines. Thermal DNA denaturation and chemosensitivity assays were used to obtain information about DNA binding properties and cytotoxicity of AAQs and HAAQ congeners. All compounds were shown to be weak*to*moderately binding to DNA, and symmetrical 1,4-di-substituted compounds were shown to be non*toxic (IC50 = 100 :/! with non-symmetrical analogues generating IC50 values in the 1-100 :/ range. No fundamental variation in the biological activity was observed when comparing AAQs with HAAQs in the A549 (+ALDH) and MCF7 (-ALDH) cell lines. A pilot investigation revealed that aberrant gene methylation was cell-type dependent for three ALDH isoforms (1A1, 2, 3A1). Decitabine treatment led to enhanced protein expression for ALDH1A1 (A549), ALDH2 (MCF7) and ALDH3A1 (A549). In contrast, the protein level was reduced for ALDH1A1 in HT29 cells after decitabine treatment. ALDH1A1, ALDH2 and ALDH3A1 were highly expressed in prostate cell lines, with expression linked to promoter methylation. In contrast, low levels of DNA methylation were found in primary prostate cancer cells and benign prostatic hyperplasia. Interestingly, ALDH1A1, considered a SC marker, was found to be expressed at low levels in CD133+/ α2β1hi stem cell fraction and upregulated in CD133-= α2β1lo differentiated prostate cancer cells. In summary, the results in this thesis demonstrate the complexity and tumour type specificity of ALDH expression. This creates challenges for the development of selective probes for CSC isolation, such as the AAQs discussed in this thesis. Although inconclusive results were obtained in regard to AAQs and their potential in targeting ALDHs, selected AAQs were shown to reveal interesting biological features highlighting them as potential non-invasive cytometric probes for tracking molecular interactions in live cells. / EPSRC, Biostatus / The full text was made available at the end of the re-embargo period, 1st September 2017.

No description available.

No description available.

O carcinoma epidermóide é uma neoplasia que pode ter origem do revestimento mucoso de vários sítios das vias aerodigestivas superiores, sendo a língua o sítio primário com maior incidência. Entre os fatores de risco para a doença estão a idade, as mutações genômicas, o hábito tabagista e principalmente o consumo de etanol. O etanol é considerado um agente cocarcinogênico no processo de desenvolvimento do câncer de boca. Por outro lado, o acetaldeído, subproduto da oxidação do etanol, é tóxico e participa diretamente na carcinogênese. Assim, polimorfismos genéticos que alteram a oxidação de etanol para acetaldeído promovendo seu acúmulo podem alterar o risco de câncer oral. Os resultados sugerem que pacientes portadores do polimorfismo do gene ADH1C Ile350Val possuem maior risco de tornarem-se etilistas crônicos (OR=2,0199), mas o risco de desenvolverem câncer não é alterado quando comparado aos não portadores. Já os portadores dos polimorfismos nos genes ADH1B Arg47His (OR=0,3445), CY2E1 (ins) (OR=0,3261) e ALDH2 (GA) (OR=0,4811) apresentaram menores riscos de desenvolverem câncer oral, mas estes polimorfismos não estavam associados ao risco de tornarem-se etilistas crônicos. Observou-se também uma possível interação entre a baixa atividade da enzima ALDH2 e a expressão do gene CYP2E1 como um fator protetor no desenvolvimento do câncer de boca. Entretanto, há necessidade de mais estudos para comprovar esses achados / Squamous cell carcinoma is a neoplasm that may originate from the mucosal tissue from various sites of the upper aerodigestive tract, the tongue being the primary site with the highest incidence. Among the risk factors for the disease are age, genomic mutations, smoking habit, and especially the consumption of ethanol. Ethanol is considered a co-carcinogenic agent in the development of oral cancer. Moreover, acetaldehyde, ethanol oxidation product, is toxic and is directly involved in carcinogenesis. Thus, genetic polymorphisms that alter the oxidation of ethanol to acetaldehyde by promoting its accumulation can alter the risk of oral cancer. The results suggest that patients with the ADH1C Ile350Val polymorphism have increased risk of becoming chronic drinkers (OR = 2.0199), but the risk of developing cancer is not changed when compared to non carriers. Since the carriers of polymorphisms in genes ADH1B Arg47His (OR = 0.3445), CY2E1 (ins) (OR =0.3261) and ALDH2 (GA) (OR = 0.4811) lower risk of developing oral cancer, but these polymorphisms were not associated with risk of becoming chronic drinkers .There was also a possible interaction between the low activity of the enzymeALDH2 and CYP2E1 gene expression as a protective factor in the development of oral cancer. However, we need more studies to confirm these findings

Le Dimethyl Ampal Thiolester (DIMATE) est un inhibiteur des aldéhydes déshydrogénases (ALDHs) de type 1 et 3. L’intérêt croissant au cours de ces dernières années pour ces enzymes intra cytoplasmiques que sont les ALDHs, s’explique par leurs utilisations comme marqueurs pour distinguer les cellules souches, saines ou cancéreuses, au sein de différents tissus, comme le tissu hématopoïétique. Le traitement des Leucémies Aiguës Myéloïdes demeure une problématique clinique majeure. En effet, malgré un taux de rémission complète moyen d’environ 70% avec les traitements conventionnels, la survie moyenne des patients porteurs d’une LAM n’excède pas les 50% à 5 ans. A ce titre, le DIMATE, semble être un médicament d’avenir. Le DIMATE présente une toxicité majeure sur plusieurs lignées leucémiques humaines et sur des cellules souches leucémiques issues de patients. De manière remarquable, le DIMATE à ces doses anti-leucémiques présente une innocuité quasi-totale sur les cellules souches hématopoïétiques saines. In vivo, chez la souris, le DIMATE permet d’éradiquer spécifiquement les cellules leucémiques humaines xénogreffées et présente en monothérapie une efficacité similaire à l’association Cytarabine + Daunorubicine qui constitue le standard thérapeutique actuel. Ces résultats encourageants vont servir de support conceptuel à la mise en place prochaine d’essais cliniques. / The Dimethyl Ampal Thiolester (DIMATE) is a type 1 and 3 Aldehydes Dehydrogenases (ALDHs) inhibitor as an innovating treatment for AML. Interest in ALDH is due to its activity as a marker for identification of stem cell in different tissues. The different species of ALDHs control the levels of the endogenous apoptogenic aldehydes. Cancer cells protect themselves from the apoptogenic effect of these aldehydes by the ALDHs that oxidize them to their non-apoptogenic carboxylic acids. The vast majority of patients with AML achieve complete remission (CR) after standard induction chemotherapy. However, the majority subsequently relapses and dies of the disease. Therefore, AML remains a clinical challenge and new therapies are urgently needed. For this, DIMATE appears as a promising drug. In vitro, DIMATE is a powerful ALDH inhibitor and has a major cytotoxic activity on human AML cell lines. Moreover, DIMATE is highly active against leukemic population enriched in LSCs, but, unlike conventional chemotherapy, DIMATE is not toxic for healthy hematopoietic stem cells which retained after treatment their self-renewing and multi-lineage differentiation capacity. In immunodeficient mice, xenografted with human leukemic cells, DIMATE eradicates specifically human AML cells and spares healthy mouse hematologic cells. Moreover, DIMATE showed the same efficiency than the association Daunorubicin + Cyrabine, which is considered as the gold standard for AML induction. Results from our work open new therapeutic perspectives in AML and provide a conceptual support for initiation of a phase I-II clinical trials, but also innovating cellular therapy.

Avaliou-se o efeito de tratamentos químicos, revestimentos comestíveis e irradiação na conservação de mamões processados minimamente. Após seleção, lavagem e sanitização, os mamões foram descascados e cortados em meias rodelas, as quais foram submetidas a diferentes tratamentos, embaladas e armazenadas a 5±1°C e 90±2%UR. Os mamões PM foram avaliados após 1, 3, 6, 9, 12 e 15 dias. As análises microbiológicas foram fundamentadas na quantificação de coliformes totais e termotolerantes, bactérias psicrotróficas, bolores e leveduras, assim como na verificação da presença de Salmonella. As avaliações físico-químicas basearam-se na determinação da concentração de CO2 no interior das embalagens, perda de massa, cor, firmeza, sólidos solúv eis, acidez titulável, ratio e pH. As características sensoriais aparência, aroma, sabor e textura foram avaliad as por meio de testes com escala hedônica. No primeiro experimento os tratamentos foram: controle, aldeído cinâmico a 0,1%, cloreto de cálcio a 0,75% e combinação de aldeído cinâmico a 0,1% com cloreto de cálcio a 0,75%. O uso de tratamentos químicos em mamões PM resultou em: maior controle de coliformes totais para os mamões PM tratados com aldeído cinâmico e com a combinação de aldeído cinâmico e cloreto de cálcio; menor concentração de CO2 e maior manutenção da firmeza para mamões PM tratados com a combinação de aldeído cinâmico e cloreto de cálcio; e maior concentração de CO2 para os mamões PM tratados apenas com aldeído cinâmico. A imersão nos tratamentos químicos resultou em maior descoloração da polpa dos mamões PM e redução do teor de sólidos solúveis ao longo do armazenamento. No segundo experimento foram utilizados os tr atamentos: controle, amido de arroz a 3%, alginato de sódio a 0,5% e carboximetilcelulose a 0,25%. O uso desses três tipos de revestimento resultou em maior controle de coliformes totais do que o observado no controle. Mamões PM revestidos com amido de arroz e carboximetilcelulose apresentaram redução e aumento da concentração de CO2, respectivamente. Os mamões PM revestidos apresentaram menores teores de sólidos solúveis e seus valores de pH se tornaram menores após 9 dias de armazenamento refrigerado. O r evestimento com carboximetilcelulose proporcionou maior firmeza da polpa no 15° dia. No terceiro experimento foram utilizados os tratamentos: controle, radiação nas doses de 2kGy e 4kGy. O uso de radiação gama em mamões PM resultou em: maior controle de coliformes totais; menor concentração de CO2 nos mamões PM tratados com 2kGy; maior concentração de CO2 e maior descoloração da polpa nos mamões PM tratados com 4kGy; redução da firmeza nos mamões PM tratados com 2kGy e 4kGy; ligeira redução do teor de sólidos solúveis e pequenas variações da acidez titulável em todos os tratamentos. As características sensoriais dos mamões PM tratados com radiação gama não diferiram significativamente do controle durante os 15 dias de armazenamento / The effect of chemical treatments, edible coatings , and irradiation on fresh-cut papaya conservation was evaluated. After selection, washing, and sanitation the papayas were peeled and cut into half slices, which were submitted to different treatments, packed, and stored at 5±1°C and 90±2%RU. The fresh-cut papayas were evaluated after 1, 3, 6, 9, 12, and 15 days. The microbiological analyses were based on the count of total coliform, thermotolerant and psychrotrophic bacteria, molds and yeasts, as well as on the presence of Salmonella. The physicochemical evaluations were based on the determination of CO2 concentration inside the package, weight loss, color, firmness, soluble solids, titratable acidity, ratio, and pH. The sensory characteristics appearance, aroma, flavor, and texture were evaluated using a hedonic scale. In the first experiment, the treatments tested we re: control, cinnamic aldehyde 0.1%, calcium chloride 0.75%, and the combination of cinnamic aldehyde 0.1% and calcium chloride 0.75%. Using chemical treatments to preserve fresh cut papaya resulted in: higher control of total coliforms in fresh-cut papayas treated with cinnamic aldehyde and with the combination of cinnamic aldehyde and calcium chloride; lower CO2 concentration and increased maintenance of firmness in freshcut papayas treated with the combination of cinnamic aldehyde and calcium chloride; and increased in the CO2 concentration in fresh-cut papayas treated only with cinnamic aldehyde. Immersion in chemical treatments caused higher pulp discoloration and reduction in solu ble solids during storage. In the second experiment, the treatments tested were: control, rice starch 3%, sodium alginate 0.5%, and carboxymethylcellulose 0.25%. The use of these three coatings resulted in higher control of total coliforms compared to the control treatment. The fresh-cut papayas coated with rice starch and carboxymethylcellulose presented reduction and increase in the CO2 concentration, respectively. Coated fresh-cut papayas presented lower soluble solids and pH values were lower after 9 days of cold storage. Carboxymethylcellulose coating increased firmness maintenance at day 15. In the third experiment, the following treatments were used: control, radiation at the doses of 2kGy and 4kGy. The use of Gamma radiation in fresh-cut papaya resulted in: higher control of total coliforms; lower CO2 concentration in fresh-cut treated with 2kGy; increased CO2 concentration and increased pulp discoloration in fresh-cut papayas treated with 4kGy; reduction in firmness in fresh-cut papayas treated with 2kGy and 4kGy; slight reduction in soluble solids and small changes in titratable acidity in all treatments. The sensory characteristics of fresh-cut papayas treated with gamma radiation did not significantly differ from the control during the 15 days of storage

Traumatic brain injury (TBI) represents a significant health crisis. To date there are no FDA-approved pharmacotherapies available to prevent the neurologic deficits caused by TBI. Following TBI, dysfunctional mitochondria generate reactive oxygen and nitrogen species, initiating lipid peroxidation (LP) and the formation of LP-derived neurotoxic aldehydes, which bind mitochondrial proteins, exacerbating dysfunction and opening of the mitochondrial permeability pore (mPTP), resulting in extrusion of mitochondrial sequestered calcium into the cytosol, and initiating a downstream cascade of calpain activation, spectrin degradation, neurodegeneration and neurologic impairment. As central mediators of the TBI secondary injury cascade, mitochondria and LP-derived neurotoxic aldehydes make promising therapeutic targets. In fact, Cyclosporine A (CsA), an FDA-approved immunosuppressant capable of inhibiting mPTP has been shown to be neuroprotective in experimental TBI. Additionally, phenelzine (PZ), an FDA-approved non-selective irreversible monoamine oxidase inhibitor (MAOI) class antidepressant has also been shown to be neuroprotective in experimental TBI due to the presence of a hydrazine (-NH-NH2) moiety allowing for the scavenging of LP-derived neurotoxic aldehydes. The overall goal of this dissertation is to further examine the neuroprotective effects of the mPTP inhibitor, CsA, and the LP-derived neurotoxic aldehyde scavenger, PZ, using a severe controlled cortical impact injury (CCI) model in 3-month old male Sprague-Dawley rats. First, the effects of CsA on cortical synaptic and non-synaptic mitochondria, two heterogeneous populations, are examined. Our results indicate that compared to non-synaptic mitochondria, synaptic mitochondria sustain greater damage 24h following CCI and are protected to a greater degree by CsA. Second, the neuroprotective effects of a novel 72h continuous subcutaneous infusion of CsA combined with PZ are compared to monotherapy. Following CCI, our results indicate that individually both CsA and PZ attenuate modification of mitochondrial proteins by LP-derived neurotoxic aldehydes, PZ is able to maintain mitochondrial respiratory control ratio and cytoskeletal integrity, but together, PZ and CsA, are unable to improve and in some cases negate monotherapy neuroprotective effects. Finally, the effects of PZ (MAOI, aldehyde scavenger), pargyline (PG, MAOI, non-aldehyde scavenger) and hydralazine (HZ, non-MAOI, aldehyde scavenger) are compared. Our results indicate that PZ, PG, and HZ are unable to improve CCI-induced deficits to learning and memory as measured by Morris water maze (post-CCI D3-7). Of concern, PZ animals lost a significant amount of weight compared to all other group, possibly due to MAOI effects. In fact, in uninjured cortical tissue, PZ administration leads to a significant increase in norepinephrine and serotonin. Additionally, although PZ, PG, and HZ did not lead to a statistically significant improvement in cortical tissue sparing 8 days following CCI, the HZ group saw a 10% improvement over vehicle. Overall, these results indicate that pharmacotherapies which improve mitochondrial function and decrease lipid peroxidation should continue to be pursued as neuroprotective approaches to TBI. However, further pursuit of LP-derived aldehyde scavengers for clinical use in TBI may require the development of hydrazine (-NH-NH2)-compounds which lack additional confounding mechanisms of action.

Récemment, les exigences en chimie ont évolué rapidement, car le développement durable a retenu plus d'attention. Les principes de la chimie verte ont encouragé les chimistes à développer des produits chimiques et des procédés qui réduisent ou éliminent les substances dangereuses. Les travaux de recherche décrits dans cette thèse portent sur le développement de nouveaux systèmes réducteurs en utilisant des hypophosphites comme substituts aux agents réducteurs toxiques traditionnels.Pour atteindre cet objectif, les nitriles aromatiques ont été réduits en aldéhydes correspondants par la formation du gaz de l’hydrogène et de nanoparticules de nickel en combinant un précurseur de nickel avec de l'hypophosphite de calcium en présence d'une base dans un milieu biphasique. De plus, les nitriles aromatiques ont été réduits en amines primaires en utilisant de l'hypophosphite de calcium et le catalyseur hétérogène palladium sur le carbone. La nature du catalyseur métallique, les additifs, les solvants, la température et les concentrations ont été étudiés en détail.D'autre part, l'amination réductrice directe des cétones aliphatiques et aromatiques a été réalisée pour la première fois en utilisant du palladium hétérogène sur du carbone et de l'hypophosphite d'ammonium qui agit comme une source d'ammoniac et un agent réducteur en même temps. Au cours de l'optimisation, des différents paramètres ont été étudiés / Recently, requirements in chemistry are changing fast, since sustainable development has retained more attention. Green chemistry principles have promoted chemists to develop chemical products and processes that reduce or eliminate hazardous substances. The research work described in this thesis is focused on the development of new reducing systems using hypophosphites as substitutes for traditional toxic reducing agents.In order to achieve this goal, aromatic nitriles were reduced into the corresponding aldehydes by the formation of hydrogen gas and nickel nanoparticles upon combining a nickel precursor with calcium hypophosphite in the presence of base in a biphasic medium. Moreover, aromatic nitriles were reduced into primary amines using calcium hypophosphite and the heterogeneous catalyst palladium on carbon. The nature of the metal catalyst, additives, solvents, temperature, and concentrations were studied in details.On the other hand, the well-known direct reductive amination of aliphatic and aromatic ketones was done for the first time using heterogeneous palladium on carbon, and ammonium hypophosphite which acts as a source of ammonia and as a reducing agent at the same time. During optimization different parameters were studied