Defective p53 response and apoptosis associated with an ataxia-telangiectasia-like phenotype

Ataxia-telangiectasia mutated (ATM), the protein defective in ataxia-telangiectasia, plays a central role in DNA damage response and signaling to cell cycle checkpoints. We describe here a cell line from a patient with an ataxia-telangiectasia-like clinical phenotype defective in the p53 response to radiation but with normal ATM activation and efficient downstream phosphorylation of other ATM substrates. No mutations were detected in ATM cDNA. A normal level of interaction between p53 and peptidyl-prolyl-isomerase Pin1 suggests that posttranslational modification was intact in these cells but operating at reduced level. Defective p53 stabilization was accompanied by defective induction of p53 effector genes and failure to induce apoptosis in response to DNA-damaging agents. Continued association between p53 and murine double minute-2 (Mdm2) occurred in irradiated ATL2ABR cells in response to DNA damage, and incubation with Mdm2 antagonists, nutlins, increased the stabilization of p53 and its transcriptional activity but failed to induce apoptosis. These results suggest that ATM-dependent stabilization of p53 and induction of apoptosis by radiation involve an additional factor(s) that is defective in ATL2ABR cells.

Inhibition of S/G(2) phase CDK4 reduces mitotic fidelity

Cyclin-dependent kinase 4 (CDK4)/cyclin D has a key role in regulating progression through late G(1) into S phase of the cell cycle. CDK4-cyclin D complexes then persist through the latter phases of the cell cycle, although little is known about their potential roles. We have developed small molecule inhibitors that are highly selective for CDK4 and have used these to define a role for CDK4-cyclin D in G(2) phase. The addition of the CDK4 inhibitor or small interfering RNA knockdown of cyclin D3, the cyclin D partner, delayed progression through G(2) phase and mitosis. The G(2) phase delay was independent of ATM/ATR and p38 MAPK but associated with elevated Wee1. The mitotic delay was because of failure of chromosomes to migrate to the metaphase plate. However, cells eventually exited mitosis, with a resultant increase in cells with multiple or micronuclei. Inhibiting CDK4 delayed the expression of the chromosomal passenger proteins survivin and borealin, although this was unlikely to account for the mitotic phenotype. These data provide evidence for a novel function for CDK4-cyclin D3 activity in S and G(2) phase that is critical for G(2)/M progression and the fidelity of mitosis.

Operation of polymer electrolyte membrane fuel cells with dry feeds: Design and operating strategies

The operation of polymer electrolyte membrane fuel cells (PEMFCs) with dry feeds has been examined with different fuel cell flow channel designs as functions of pressure, temperature and flow rate. Auto-humidified (or self-humidifying) PEMFC operation is improved at higher pressures and low gas velocities where axial dispersion enhances back-mixing of the product water with the dry feed. We demonstrate auto-humidified operation of the channel-less, self-draining fuel cell, based on a stirred tank reactor; data is presented showing auto-humidified operation from 25 to 115 degrees C at 1 and 3 atm. Design and operating requirements are derived for the auto-humidified operation of the channel-less, self-draining fuel cell. The auto-humidified self-draining fuel cell outperforms a fully humidified serpentine flow channel fuel cell at high current densities. The new design offers substantial benefits for simplicity of operation and control including: the ability to self-drain reducing flooding, the ability to uniformly disperse water removing current gradients and the ability to operate on dry feeds eliminating the need for humidifiers. Additionally, the design lends itself well to a modular design concept. (c) 2005 Elsevier B.V. All rights reserved.

The complexity of p53 stabilization and activation

A number of proteins are activated by stress stimuli but none so spectacularly or with the degree of complexity as the tumour suppressor p53 (human p53 gene or protein). Once stabilized, p53 is responsible for the transcriptional activation of a series of proteins involved in cell cycle control, apoptosis and senescence. This protein is present at low levels in resting cells but after exposure to DNA-damaging agents and other stress stimuli it is stabilized and activated by a series of post-translational modifications that free it from MDM2 (mouse double minute 2 but used interchangeably to denote human also), a ubiquination ligase that ubiquitinates it prior to proteasome degradation. The stability of p53 is also influenced by a series of other interacting proteins. In this review, we discuss the post-translational modifications to p53 in response to different stresses and the consequences of these changes.

Inhibition of transforming growth factor-beta 1 signaling attenuates ataxia telanglectasia mutated activity in response to genotoxic stress

Ionizing radiation causes DNA damage that elicits a cellular program of damage control coordinated by the kinase activity of ataxia telangiectasia mutated protein (ATM). Transforming growth factor beta (TGF beta)-1, which is activated by radiation, is a potent and pleiotropic mediator of physiologic and pathologic processes. Here we show that TGF beta inhibition impedes the canonical cellular DNA damage stress response. Irradiated Tgf beta 1 nail murine epithelial cells or human epithelial cells treated with a small-molecule inhibitor of TGF beta type I receptor kinase exhibit decreased phosphorylation of Chk2, Rad17, and p53; reduced gamma H2AX radiation-induced foci; and increased radiosensitivity compared with TGF beta competent cells. We determined that loss of TGF beta signaling in epithelial cells truncated ATM autophosphorylation and significantly reduced its kinase activity, without affecting protein abundance. Addition of TGF beta restored functional ATM and downstream DNA damage responses. These data reveal a heretofore undetected critical link between the microenvironment and ATM, which directs epithelial cell stress responses, cell fate, and tissue integrity. Thus, Tgf beta 1, in addition to its role in homoeostatic growth control, plays a complex role in regulating responses to genotoxic stress, the failure of which would contribute to the development of cancer; conversely, inhibiting TGF beta may be used to advantage in cancer therapy.

REABSORÇÃO CONDILAR INTERNA DA ARTICULAÇÃO TEMPOROMANDIBULAR NO PACIENTE ADOLESCENTE

A reabsorção condilar interna no adolescente (RCIA) é uma doença progressiva que afeta a articulação temporomandibular (ATM) e que pode resultar em maloclusão, deformidade facial, disfunção de ATM e dor. O aparecimento desta doença ocorre entre os 10 e os 15 anos, sendo mais freqüente em adolescentes do sexo feminino. Esses pacientes apresentam sinais clínicos característicos como: ângulo do plano oclusal e do plano mandibular aumentados, retrusão progressiva da mandíbula e maloclusão Classe II, com ou sem mordida aberta. Nos exames de imagem (tomografia computadorizada e ressonância magnética), observa-se reabsorção interna dos côndilos e deslocamento dos discos articulares da ATM. Tendo em vista a dificuldade em determinar a causa da RCIA e a importância em eliminar a dor e melhorar a função mastigatória, este estudo se propôs a avaliar as alterações promovidas no posicionamento mandibular, a sintomatologia dolorosa e a estabilidade do tratamento da RCIA, utilizando o protocolo de tratamento (Dr. Larry M. Wolford) em pacientes adolescentes submetidos a cirurgia de reposicionamento do disco articular e cirurgia ortognática, realizadas no mesmo ato cirúrgico. Neste estudo retrospectivo, foram avaliadas as telerradiografias laterais adquiridas na avaliação inicial (T1), pré-cirúrgica (T2), pós-cirúrgica imediata (T3) e de maior acompanhamento, com pelo menos um ano pós-cirurgia (T4) e questionários de dor e função mandibular de uma amostra de 24 pacientes com maloclusão de Classe II (20 do sexo feminino e 4 do sexo masculino) diagnosticados com RCIA, os quais foram submetidos ao protocolo de tratamento composto pela cirurgia de reposicionamento do disco articular da ATM concomitantemente à cirurgia ortognática. O protocolo de tratamento utilizado mostrou-se bastante positivo, uma vez que a estabilidade foi adquirida e uma melhora significativa nos níveis de dor e função mandibular foram atingidos. O tratamento dos pacientes diagnosticados com RCIA, através do protocolo de tratamento proposto, é um procedimento estável e atua diretamente em um ganho na redução da dor.

EFEITOS DOS APARELHOS TWIN FORCE BITE CORRECTOR E FORSUS FATIGUE RESISTANT DEVICE NA JUNÇÃO TEMPOROMANDIBULAR E NAS VIAS AÉREAS

O objetivo deste estudo prospectivo foi verificar os efeitos dos aparelhos Twin Force Bite Corrector e Forsus Fatigue Resistant Device na junção temporomandibular e nas vias aéreas. Para tanto, foram selecionados 21 pacientes adultos jovens, má oclusão de Classe II 1ª. divisão, bilateral, mínimo de 1/2 Classe II, padrão de crescimento horizontal. Para tanto, a amostra foi dividida em 2 grupos: grupo A, com média de idade inicial de 14,55 anos, tratado com o aparelho ortopédico fixo Twin ForceR por um período médio de 3,7 meses e o grupo B com média de idade inicial de 15,45 anos, tratado com o aparelho Forsus, por um período médio de 7,4 meses, ambos associados à aparelhagem fixa com mecânica straight-wire, prescrição MBT. Os pacientes do grupo A realizaram TCFC em três tempos: T1A (inicial - sem instalação do aparelho), T2A (no dia da i nstalação do aparelho ortopédico fixo), T3A (após a remoção do aparelho ortopédico fixo). Já o grupo 2 realizou TCFC em dois tempos: T2B (no dia da instalação do aparelho ortopédico fixo) e T3B (após a remoção do aparelho ForsusTM). Após a obtenção das imagens 3Dforam realizados cortes sagitais e coronais na ATM para avaliação do tamanho do côndilo e avaliação dos espaços articulares e vias aéreas. Para comparação intergrupos foi utilizado o teste "t" independente e correlação de Pearson e para comparação intragrupos, análise de variância a um critério e teste t pareado. O nível de significância adotado foi de p<0,05%. Não houve alteração significante na avaliação intergrupos, para as dimensões condilares, espaços articulares, e vias aéreas. Por outro lado, na avaliação intragrupo, houve alteração significante no comprimento efetivo da mandíbula (Co-Gn) para o grupo B, supostamente devido a um maior tempo de tratamento, pois houve correlação significante da largura coronal esquerda e direita com Co-Gn, embora clinicamente tenha sido irrelevante. Sendo assim, conclui-se que não houve diferença entre o desempenho dos dois aparelhos estudados na remodelação óssea condilar em pacientes jovens adultos e não houve alteração das vias aéreas após o tratamento com os aparelhos ortopédicos funcionais fixos e Forsus e Twin Force.

Ultrasonic field and temperature sensor based on short in-fibre Bragg gratings

The authors demonstrate that in-fibre Bragg gratings may be successfully used to measure megahertz acoustic fields if the grating length is sufficiently short and the optical fibre is appropriately desensitised. A noise-limited pressure resolution of 4.5 × 10 –3 atm vHz was found. The capability to simultaneously act as a temperature sensor is also demonstrated.

Combined ultrasound and temperature sensor using a fibre Bragg grating

We demonstrate the feasibility of using in-fibre Bragg gratings to measure MHz acoustic fields and temperature simultaneously. We achieved a noise-limited pressure resolution of ˜4.5×10-4 Atm/vHz and a temperature resolution of 0.2°C.

Packet switching in wide area broadband private networks

B-ISDN is a universal network which supports diverse mixes of service, applications and traffic. ATM has been accepted world-wide as the transport technique for future use in B-ISDN. ATM, being a simple packet oriented transfer technique, provides a flexible means for supporting a continuum of transport rates and is efficient due to possible statistical sharing of network resources by multiple users. In order to fully exploit the potential statistical gain, while at the same time provide diverse service and traffic mixes, an efficient traffic control must be designed. Traffic controls which include congestion and flow control are a fundamental necessity to the success and viability of future B-ISDN. Congestion and flow control is difficult in the broadband environment due to the high speed link, the wide area distance, diverse service requirements and diverse traffic characteristics. Most congestion and flow control approaches in conventional packet switched networks are reactive in nature and are not applicable in the B-ISDN environment. In this research, traffic control procedures mainly based on preventive measures for a private ATM-based network are proposed and their performance evaluated. The various traffic controls include CAC, traffic flow enforcement, priority control and an explicit feedback mechanism. These functions operate at call level and cell level. They are carried out distributively by the end terminals, the network access points and the internal elements of the network. During the connection set-up phase, the CAC decides the acceptance or denial of a connection request and allocates bandwidth to the new connection according to three schemes; peak bit rate, statistical rate and average bit rate. The statistical multiplexing rate is based on a `bufferless fluid flow model' which is simple and robust. The allocation of an average bit rate to data traffic at the expense of delay obviously improves the network bandwidth utilisation.

A new strategy for selecting and evaluating physical solvents for gas absorption

This work describes how the physical properties of a solvent affect the design variables of a physical gas absorption process. The role of every property in determining the capital and the running cost of a process has been specified. Direct mathematical relationships have been formulated between every item of capital or running cost and the properties which are related to that item. The accuracy of the equations formulated has been checked by comparing their outcome with some actual design data. A good agreement has been found. The equations formulated may be used to evaluate on the basis of economics any suggested new solvents. A group of solvents were selected for evaluation. Their physical properties were estimated or collected as experimental data. The selected ones include three important solvents, the first is polyethylene glycol dimethyl ether (Selexol) which represents the currently most successful one, The other two solvents are acetonyl acetone (B2) and n-formyl morpholine which have been suggested previously as potential credible alternatives to the current ones. The important characteristics of: acetonyl acetone are its high solubility and its low viscosity, while the n-formyl morpholine is characterised by its low vapour pressure and its high selectivity. It was found that acetonyl acetone (B2) is the most attractive solvent for commercial applications particularly for process configurations that:include heat exchangers and strippers. The effect of the process configuration on the selected solvent was investigated in detail and it was found that there is no universal solvent which is the best for any process configuration, but that there is a best solvent for a given process configuration. In previous work, acetonyl acetone was suggested as a commercially promising physical solvent. That suggestion was not fully based on experimental measurement of all the physical properties. The viscosity of acetonyl acetone and its solubility at 1 atm were measured but the vapour pressure and the solubility of C02 and CH4 at high pressure were predicted. In this work, the solubilities of C02, CH4 and C3H8 in acetenyl acetone were measured for a partial pressure range of (2 ~ 22) bar at 25°C, The vapour pressure of this solvent was also measured, and the Antoine equation was formulated from tbe experimental data. The experimental data were found to be not In agreement with the predicted ones, so acetonyl acetone was re-evaluated according to the experimental data. It was found that this solvent can be recommended for further trials in a pilot plant study or for small scale commercial units.

The use of context in the classification of urban aerial imagery

Urban regions present some of the most challenging areas for the remote sensing community. Many different types of land cover have similar spectral responses, making them difficult to distinguish from one another. Traditional per-pixel classification techniques suffer particularly badly because they only use these spectral properties to determine a class, and no other properties of the image, such as context. This project presents the results of the classification of a deeply urban area of Dudley, West Midlands, using 4 methods: Supervised Maximum Likelihood, SMAP, ECHO and Unsupervised Maximum Likelihood. An accuracy assessment method is then developed to allow a fair representation of each procedure and a direct comparison between them. Subsequently, a classification procedure is developed that makes use of the context in the image, though a per-polygon classification. The imagery is broken up into a series of polygons extracted from the Marr-Hildreth zero-crossing edge detector. These polygons are then refined using a region-growing algorithm, and then classified according to the mean class of the fine polygons. The imagery produced by this technique is shown to be of better quality and of a higher accuracy than that of other conventional methods. Further refinements are suggested and examined to improve the aesthetic appearance of the imagery. Finally a comparison with the results produced from a previous study of the James Bridge catchment, in Darleston, West Midlands, is made, showing that the Polygon classified ATM imagery performs significantly better than the Maximum Likelihood classified videography used in the initial study, despite the presence of geometric correction errors.

Evaluation of remote sensing for the detection of landfill gas and leachate in an urban environment

The technique of remote sensing provides a unique view of the earth's surface and considerable areas can be surveyed in a short amount of time. The aim of this project was to evaluate whether remote sensing, particularly using the Airborne Thematic Mapper (ATM) with its wide spectral range, was capable of monitoring landfill sites within an urban environment with the aid of image processing and Geographical Information Systems (GIS) methods. The regions under study were in the West Midlands conurbation and consisted of a large area in what is locally known as the Black Country containing heavy industry intermingled with residential areas, and a large single active landfill in north Birmingham. When waste is collected in large volumes it decays and gives off pollutants. These pollutants, landfill gas and leachate (a liquid effluent), are known to be injurious to vegetation and can cause stress and death. Vegetation under stress can exhibit a physiological change, detectable by the remote sensing systems used. The chemical and biological reactions that create the pollutants are exothermic and the gas and leachate, if they leave the waste, can be warmer than their surroundings. Thermal imagery from the ATM (daylight and dawn) and thermal video were obtained and used to find thermal anomalies on the area under study. The results showed that vegetation stress is not a reliable indicator of landfill gas migration, as sites within an urban environment have a cover too complex for the effects to be identified. Gas emissions from two sites were successfully detected by all the thermal imagery with the thermal ATM being the best. Although the results were somewhat disappointing, recent technical advancements in the remote sensing systems used in this project would allow geo-registration of ATM imagery taken on different occasions and the elimination of the effects of solar insolation.

Investigation of EDFA power transients in circuit-switched and packet-switched optical networks

Erbium-doped fibre amplifiers (EDFA’s) are a key technology for the design of all optical communication systems and networks. The superiority of EDFAs lies in their negligible intermodulation distortion across high speed multichannel signals, low intrinsic losses, slow gain dynamics, and gain in a wide range of optical wavelengths. Due to long lifetime in excited states, EDFAs do not oppose the effect of cross-gain saturation. The time characteristics of the gain saturation and recovery effects are between a few hundred microseconds and 10 milliseconds. However, in wavelength division multiplexed (WDM) optical networks with EDFAs, the number of channels traversing an EDFA can change due to the faulty link of the network or the system reconfiguration. It has been found that, due to the variation in channel number in the EDFAs chain, the output system powers of surviving channels can change in a very short time. Thus, the power transient is one of the problems deteriorating system performance. In this thesis, the transient phenomenon in wavelength routed WDM optical networks with EDFA chains was investigated. The task was performed using different input signal powers for circuit switched networks. A simulator for the EDFA gain dynamicmodel was developed to compute the magnitude and speed of the power transients in the non-self-saturated EDFA both single and chained. The dynamic model of the self-saturated EDFAs chain and its simulator were also developed to compute the magnitude and speed of the power transients and the Optical signal-to-noise ratio (OSNR). We found that the OSNR transient magnitude and speed are a function of both the output power transient and the number of EDFAs in the chain. The OSNR value predicts the level of the quality of service in the related network. It was found that the power transients for both self-saturated and non-self-saturated EDFAs are close in magnitude in the case of gain saturated EDFAs networks. Moreover, the cross-gain saturation also degrades the performance of the packet switching networks due to varying traffic characteristics. The magnitude and the speed of output power transients increase along the EDFAs chain. An investigation was done on the asynchronous transfer mode (ATM) or the WDM Internet protocol (WDM-IP) traffic networks using different traffic patterns based on the Pareto and Poisson distribution. The simulator is used to examine the amount and speed of the power transients in Pareto and Poisson distributed traffic at different bit rates, with specific focus on 2.5 Gb/s. It was found from numerical and statistical analysis that the power swing increases if the time interval of theburst-ON/burst-OFF is long in the packet bursts. This is because the gain dynamics is fast during strong signal pulse or with long duration pulses, which is due to the stimulatedemission avalanche depletion of the excited ions. Thus, an increase in output power levelcould lead to error burst which affects the system performance.

Cytotoxic activity of Fagonia cretica against human breast cancer cells

In many parts of the world, plants are directly utilised for their medicinal properties. Traditional medicine from Pakistan, India and the Far East is well documented and its history is embedded in folklore. It has been documented that an aqueous extract of the desert shrub, Fagonia cretica, is a popular treatment for breast cancer in Pakistan. The administration of an aqueous extract of Fagonia cretica is reported effective at reducing tumour size and improving the quality of life of breast cancer patients, is well tolerated and does not exhibit adverse effects like vomiting, diarrhoea or alopecia which are common side effects of standard cytotoxic therapy. In the past, many pharmacologically active and chemotherapeutic compounds have been isolated from plants which subsequently have proven to be successful in clinical trials and been used as primary compounds in therapeutic regimes. Fagonia cretica has historical use as a treatment for breast cancer, yet there is little scientific evidence which shows chemotherapeutic potential towards breast tumours. Preparation and analysis of an aqueous extract of Fagonia cretica may reveal novel chemotherapeutic agents that can be used to effectively target cancer cells. An understanding of the mechanism of any activity may improve our understanding of cancer cell biology and reveal novel therapeutic targets. This thesis describes for the first time that an aqueous extract of Fagonia cretica shows potent in vitro cytotoxic activity towards breast cancer epithelial cell lines which was not seen towards normal mammary epithelial cells. Elucidation and characterisation of the cytotoxic mechanism was undertaken by analysing DNA damage, cell cycle status, apoptosis, metabolic state and expression of transcription factors and their targets. Finally, methods for the isolation and identification of active compound(s) were developed using various chromatographic techniques. An aqueous extract of Fagonia cretica was able to reduce cell viability significantly in two phenotypically different breast cancer cell lines (MCF-7 and MDA-MB-231). This activity was markedly reduced in normal mammary epithelial cells (HMEpC). Further investigation into the mode of action revealed that extract treatment induced cell cycle arrest and apoptosis in both MCF-7 and MDA-MB-231 cell lines. This coincided with the formation of DNA double stranded breaks and the DNA repair marker ?-H2AX. In MCF-7 cells, ATM/ATR activation resulted in increased p53 expression and of its transcriptional targets p21 and bax, suggesting a role for a p53-mediated response. Furthermore, inhibition of extract-induced p53 expression with siRNA reduced the cytotoxic effect against MCF-7 cells. Extract treatment was also associated with increased FOXO3a expression in MCF-7 and MDA-MB-231 cells. In the absence of functional p53, siRNA knockdown of extract-induced FOXO3a expression was completely abrogated, suggesting that FOXO3a plays a vital role in extract-induced cytotoxicity. Isolation and characterisation of the active compound(s) within the extract was attempted using liquid chromatography and mass spectrometry in conjunction with a cell viability assay. Multiple fractionations generated an active fraction that contained four major compounds as detected by mass spectrometry. However, none of these compounds were identified structurally or chemically due to constraints within the methodology.