L’entérotoxine STb d’Escherichia coli affecte les jonctions serrées des cellules intestinales épithéliales

La toxine thermostable d’E.coli (STb) est une cause de diarrhée chez l’homme et l’animal. STb se lie au sulfatide, son récepteur, puis s’internalise. Dans le cytoplasme, par une cascade d’événements, STb déclenche l’ouverture des canaux ioniques permettant la sécrétion des ions et la perte d’eau menant à la diarrhée. Les jonctions serrées forment une barrière physique intercellulaire dans les cellules épithéliales intestinales, contrôlant ainsi le flux paracellulaire des ions et de l’eau. Les jonctions serrées sont affectées par divers pathogènes et par leurs toxines. À ce jour, l’effet de STb sur les jonctions serrées n’a pas été étudié. L’étude entreprise visait à explorer l’effet de STb sur les jonctions serrées et la barrière épithéliale des cellules intestinales. Des cellules épithéliales intestinales du colon humain (T84) ont été traitées pendant 24h soit avec la toxine STb purifiée soit avec une souche d’E.coli exprimant STb. La résistance transépithéliale (TER), le flux de marqueurs paracellulaires et la microscopie confocale ont été utilisés pour analyser les effets de STb sur les jonctions serrées. Les monocouches traitées par la souche E.coli exprimant STb et la toxine STb purifiée ont manifesté une forte réduction de TER (p<0.0001) parallèlement à une augmentation significative de la perméabilité paracellulaire à l’Albumine de Sérum Bovin marqué avec l’IsoThioCyanate Fluoroscéine, BSA-FITC (p<0.0001) comparativement aux cellules non traitées et aux cellules traitées par une souche d’E.coli commensale non-toxinogène. L’augmentation de la perméabilité paracellulaire induite par STb a été associée à une dissolution générale et une condensation des fibres de stress centrales des filaments d’actine. Le réarrangement des filaments d’actine a été accompagné par une redistribution et une fragmentation des protéines des jonctions serrées dont l’occludine, la claudine-1 et la Zonula Occludens-1. Les mêmes modifications on été observées après l’intoxication des cellules T84 avec un octapeptide synthétique retrouvé dans la séquence de STb correspondant à une séquence consensus de la toxine ZOT de Vibrio cholerae, impliquée dans la réorganisation des jonctions serrées. Cet effet n’a pas été observé lorsque les cellules ont été traitées avec un octapeptide synthétique comportant les mêmes acides aminés mais distribués de façon aléatoire ou avec la toxine mutée (D30V). Nos résultats montrent pour la première fois que STb induit le dysfonctionnement de la barrière épithéliale intestinale en modifiant la distribution des protéines des jonctions serrées. Ces résultats ouvrent une nouvelle voie pour la compréhension de la pathogenèse de diarrhée causée par la toxine STb.

Persistence and photochemical decay of springtime total ozone anomalies in the Canadian Middle Atmosphere Model

The persistence and decay of springtime total ozone anomalies over the entire extratropics (midlatitudes plus polar regions) is analysed using results from the Canadian Middle Atmosphere Model (CMAM), a comprehensive chemistry-climate model. As in the observations, interannual anomalies established through winter and spring persist with very high correlation coefficients (above 0.8) through summer until early autumn, while decaying in amplitude as a result of photochemical relaxation in the quiescent summertime stratosphere. The persistence and decay of the ozone anomalies in CMAM agrees extremely well with observations, even in the southern hemisphere when the model is run without heterogeneous chemistry (in which case there is no ozone hole and the seasonal cycle of ozone is quite different from observations). However in a version of CMAM with strong vertical diffusion, the northern hemisphere anomalies decay far too rapidly compared to observations. This shows that ozone anomaly persistence and decay does not depend on how the springtime anomalies are created or on their magnitude, but reflects the transport and photochemical decay in the model. The seasonality of the long-term trends over the entire extratropics is found to be explained by the persistence of the interannual anomalies, as in the observations, demonstrating that summertime ozone trends reflect winter/spring trends rather than any change in summertime ozone chemistry. However this mechanism fails in the northern hemisphere midlatitudes because of the relatively large impact, compared to observations, of the CMAM polar anomalies. As in the southern hemisphere, the influence of polar ozone loss in CMAM increases the midlatitude summertime loss, leading to a relatively weak seasonal dependence of ozone loss in the Northern Hemisphere compared to the observations.

Rhyacian (2.23-2.20 Ga) juvenile accretion in the southern Sao Francisco craton, Brazil: Geochemical and isotopic evidence from the Serrinha magmatic suite, Mineiro belt

The Serrinha magmatic suite (Mineiro belt) crops out in the southern edge of the Sao Francisco craton, comprising the Brito quartz-diorite, Brumado de Cima and Brumado de Baixo granodiorites, granophyres and felsic sub-volcanic and volcanic rocks, part of which intruded into the Nazareno greenstone belt. The suite rocks have petrographic features that are consistent with magma supercooling due to the low water content combined with volatile loss, leading to crystallization of quartz and alkaline feldspar at the rims of plagioclase phenocrysts (granophyric intergrowth). The investigated rocks are sub-alkaline, calc-alkaline and show low content in rare earth elements. The U-Pb zircon crystallization ages for the Brumado de Cima granodiorite [2227 +/- 22 (23) Ma] and a coeval granophyre [2211 +/- 22 (23) Ma], coupled with available single-zircon Pb evaporation ages for the Brito and Brumado de Baixo plutons, are significantly older than the ""Minas orogeny"" (ca. 2100-2050 Ga) of Quadrilatero Ferrifero area, eastward from the Serrinha suite. Our data establish an early Rhyacian event tectonically linked with the evolution of the Mineiro belt. The bulk Nd isotopic signature [low negative to positive epsilon(Nd(t)) values] of the Serrinha samples are consistent with the important role of Paleoproterozoic mantle components in the magma genesis. The integrated geologic, geochemical and isotopic information suggests that Paleoproterozoic evolution of the Mineiro belt initiated in a passive continental margin basin with deposition of the Minas Supergroup at ca. 2500 Ma. This stage was succeeded by outboard rupture of the oceanic lithosphere with development and coalescence of progressively younger magmatic arcs during Rhyacian time. One of the earliest arcs formed the Serrinha suite. The tectonic collage of the Serrinha and Ritapolis (2190-2120 Ma) arcs produced the NE-SW Lenheiro shear zone, resulting in mylonitization and recrystallization of both the granitoid intrusions and host rocks. As a matter of fact juxtaposition of distinct magmatic units in age and origin took place along the Lenheiros structure in this sector of the Mineiro belt. (C) 2009 Elsevier Ltd. All rights reserved.

A theory of weekly specials

Multiproduct retailers facing similar costs and serving the same public commonly announce different weekly specials. These promotional prices also seem to evolve randomly over the weeks. Here, weekly specials are viewed as the strategic outcome of an oligopolistic price competition among multiproduct retail stores facing nonconvex costs. Existence of an equilibrium in mixed strategies is proven. ldentical stores serving the same public will never charge the same price vector with probability one (cross-store price dispersion). Mixed strategies can generate random price dispersion over time in the repeated version of the mode!.

Efeitos cardiovascular e metabólico da reposição volêmica com solução salina hipertônica 7,5% associada ou não ao hidroxietilamido 130/0,4 em gatas com hipovolemia induzida

A hipovolemia é caracterizada por uma perda de fluido corpóreo, cursando com inadequado fluxo circulatório e consequentemente lesão tecidual. Neste trabalho, objetivou-se comparar a expansão volêmica resultante da administração de solução salina hipertônica (NaCl 7,5%), isolada ou em associação com hidroxietilamido 130/0,4 (HES 130/0,4), em gatas com hipovolemia induzida, sob anestesia geral inalatória com isofluorano. Foram utilizadas 12 gatas, sem raça definida, adultas, com massa corporal média de 3,07±0,56kg. Os animais foram anestesiados com isofluorano e, após a preparação cirúrgica, foram mantidos em 1CAM sob ventilação controlada. Após a estabilização do plano anestésico, foram avaliados os parâmetros basais. em ato contínuo, iniciou-se a fase de hipovolemia, por meio da retirada de 30ml kg-1 de sangue da artéria femoral. Após 60 minutos da estabilização do quadro de hipovolemia, foi realizada nova mensuração dos dados, alocando-se os animais aleatoriamente em dois grupos: GSH (grupo solução hipertônica, n=6), que receberam, na fase de expansão volêmica, NaCl 7,5% isolada, na dose de 4ml kg-1, e GSHC (grupo salina hipertônica associado ao coloide, n=6), que receberam NaCl 7,5%, na mesma dose citada, em associação com HES 130/0,4, na dose de 30ml kg-1. Após realização do tratamento, foram avaliados novamente os efeitos cardiovasculares e hemogasométricos por 120 minutos. As pressões arteriais média (PAM), sistólica (PAS) e diastólica (PAD) foram maiores logo após a expansão volêmica (T0) para o GSH. de T45 até T120, as PAM, PAS e PAD foram maiores para o GSHC, em comparação com o GSH. A pressão venosa central foi maior no GSHC até T60. Não foram observadas diferenças entre grupos para frequência cardíaca e respiratória, íons sódio e potássio, déficit de base, bicarbonato, saturação de oxigênio na hemoglobina, glicose, PaCO2, PaO2 e pH. Conclui-se que a administração de NaCl 7,5% isoladamente aumenta rapidamente a PAM, PAS e PAD em gatos com hipovolemia induzida, mantendo esse efeito por apenas 30 minutos, enquanto que a administração de hidroxietilamido 130/0,4 associado à NaCl 7,5% promove reestabelecimento mais tardio (após 30 minutos), porém mais duradouro (até 120 minutos) da PAM, PAS e PAD em gatas com hipovolemia induzida. A administração de HES 130/0,4 associada à NaCl 7,5% promove aumento acentuado da PVC por até 60 minutos após a administração.

Valutazione preoperatoria del rischio chirurgico in pazienti onco-geriatrici (studio PREOP)

La maggior parte dei pazienti che vengono sottoposti a interventi chirurgici per tumori solidi hanno un’età superiore a 70 anni1. Circa il 30% di questi pazienti vengono considerati “fragili”. Questi infatti presentano numerose comorbidità ed hanno un più elevato rischio di sviluppare complicanze postoperatorie con perdita della riserva funzionale residua. Per questo non esistono sistemi semplici di screening che permettano ai medici responsabili del trattamento di identificare questi pazienti con aumentato rischio postoperatorio. Identificare i pazienti a rischio è infatti il primo passo nel processo attraverso il quale è possibile prevenire in necessarie complicanze postoperatorie come delirio, eventi cardiovascolari e perdita della funzionalità complessiva con conseguente perdita di autonomia. Scopo di questo studio è quello di confrontare l’accuratezza nella previsione di mortalità e morbidità a 30 giorni dei tre test preditivi “Groningen Frailty Index” (GFI); “Vulnerable Elders Survey” (VES-13); “timed up and go test” con alcune componenti del Preoperative Assessment of Cancer in the Elderly (PACE). Lo studio verrà effettuato sui pazienti con età maggiore di 70 anni che dovranno essere sottoposti a intervento chirurgico in anestesia generale per la presenza di una neoplasia solida.

A systematic review of organizational workplace wellness programs and their financial and nonfinancial incentives

Workplace wellness programs have revealed immense beneficial results for both the employer and employee. Examples of results include decrease in absenteeism, turnover rate, medical claims and increases in employee satisfaction, productivity, and return on investment. However, the approach taken when implementing requires greater attention since such programs and the financial and/or non-financial incentives chosen have shown to significantly impact employee participation thus the amount of savings the organization experiences. A systematic review was conducted to evaluate the overall effectiveness of workplace wellness programs on employee health status and lifestyle change, recognize the majority types of returns observed by such programs, and identify whether financial or non-financial incentives created a greater effect on the employee. Overall employee health status improvement occurred when participating in wellness programs. The dominant indirect benefit for the organization was employee weight loss leading to a decrease in absenteeism and direct benefits included decreases in medical claims and increases in return on investment. In general, factors such as rate of participation and health status changes were most influenced when a financial incentives was provided in the wellness program. The basis of providing a program with effective incentives resides from efforts made by the employer and their efforts to play a role on every level of the organization regarding planning, implementing, and strategizing the most optimal approach for creating changes for the employees' wellbeing and productivity, thus the organizations overall returns.^

Tin oxide based composites derived using electrostatic spray deposition technique as anodes for lithium-ion batteries

Recent advances in the electric & hybrid electric vehicles and rapid developments in the electronic devices have increased the demand for high power and high energy density lithium ion batteries. Graphite (theoretical specific capacity: 372 mAh/g) used in commercial anodes cannot meet these demands. Amorphous SnO2 anodes (theoretical specific capacity: 781 mAh/g) have been proposed as alternative anode materials. But these materials have poor conductivity, undergo a large volume change during charging and discharging, large irreversible capacity loss leading to poor cycle performances. To solve the issues related to SnO2 anodes, we propose to synthesize porous SnO2 composites using electrostatic spray deposition technique. First, porous SnO2/CNT composites were fabricated and the effects of the deposition temperature (200, 250, 300 °C) & CNT content (10, 20, 30, 40 wt %) on the electrochemical performance of the anodes were studied. Compared to pure SnO2 and pure CNT, the composite materials as anodes showed better discharge capacity and cyclability. 30 wt% CNT content and 250 °C deposition temperature were found to be the optimal conditions with regard to energy capacity whereas the sample with 20% CNT deposited at 250 °C exhibited good capacity retention. This can be ascribed to the porous nature of the anodes and the improvement in the conductivity by the addition of CNT. Electrochemical impedance spectroscopy studies were carried out to study in detail the change in the surface film resistance with cycling. By fitting EIS data to an equivalent circuit model, the values of the circuit components, which represent surface film resistance, were obtained. The higher the CNT content in the composite, lower the change in surface film resistance at certain voltage upon cycling. The surface resistance increased with the depth of discharge and decreased slightly at fully lithiated state. Graphene was also added to improve the performance of pure SnO2 anodes. The composites heated at 280 °C showed better energy capacity and energy density. The specific capacities of as deposited and post heat-treated samples were 534 and 737 mAh/g after 70 cycles. At the 70th cycle, the energy density of the composites at 195 °C and 280 °C were 1240 and 1760 Wh/kg, respectively, which are much higher than the commercially used graphite electrodes (37.2–74.4 Wh/kg). Both SnO2/CNTand SnO2/grapheme based composites with improved energy densities and capacities than pure SnO2 can make a significant impact on the development of new batteries for electric vehicles and portable electronics applications.

Exploring business models to provide a foundation for enhanced eye care services in high street optometric practice

High street optometric practices are for-profit businesses. They mostly provide sight testing and eye examination services and sell optical products, such as spectacles and contact lenses. The sight testing services are often sold at a vastly reduced price and profits are generated primarily through high margin spectacle sales, in a loss leading strategy. Published literature highlights weaknesses in this strategy as it forms a barrier to widening the scope of services provided within optometric practices. This includes specialist non-refraction based services, such as shared care. In addition this business strategy discourages investment in advanced diagnostic equipment and higher professional qualifications. The aim of this thesis was to develop a greater understanding of the traditional loss-leading strategy. The thesis also aimed to assess the plausibility of alternative business models to support the development of specialist non-refraction services within high street optometric practice. This research was based on a single independent optometric practice that specialises in advanced retinal imaging and offers a broad range of shared care services. Specialist non-refraction based services were found to be poor generators of spectacle sales likely due to patient needs and presenting concerns. Alternative business strategies to support these services included charging more realistic professional fees via cost-based pricing and monthly payment plans. These strategies enabled specialist services to be more self-sustainable with less reliance on cross-subsidy from spectacle sales. Furthermore, improving operational efficiency can increase stand-alone profits for specialist services.Practice managers may be reluctant to increase professional fees due to market pressures and confidence. However, this thesis found that patients were accepting of increased professional fees. Practice managers can implement alternative business models to enhance eye care provision in high street optometric practices. These alternative business models also improve revenues and profits generated via clinical services and improve patient loyalty.

Degradação de Ocratoxina a: estudo de processo e toxicidade

A ocratoxina A (OTA), micotoxina encontrada em diferentes níveis e em diversas matrizes, apresenta efeitos carcinogênicos, nefrotóxicos e teratogênicos. O desenvolvimento de métodos capazes de diminuir esta contaminação a níveis permitidos pela legislação é incentivado e os processos biológicos utilizados envolvem o uso de enzimas e/ou microrganismos para degradação da OTA e são preferenciais pela especificidade, bem como pelas condições brandas para a detoxificação. O objetivo do trabalho foi estudar a ação de carboxipeptidase A nos níveis e na toxicidade de OTA, visando aplicar a técnica para detoxificar farinhas de trigo. Primeiramente foi estimado o risco de exposição à ocratoxina A pelo consumo de farinhas de trigo. Para isso foram estabelecidas condições de determinação de OTA em farinhas de trigo, empregando técnicas de estatística multivariada para definir os principais interferentes na extração de OTA pelo método de QuEChERS e detecção em CLAE-FL. O método validado permitiu a avaliação da ocorrência natural em 20 amostras de farinha de trigo, estando estas contaminadas na faixa de 0,22 a 0,85 µg.kg-1 , apresentando um valor de ingestão diária de 0,08 ngOTA.dia-1 .kgmassacorpórea -1 e uma disponibilidade de 94,4%. Em seguida foi realizada a padronização da extração de carboxipeptidase A em biomassa de Rhizopus oryzae que consistiu em agitação ultrassônica durante 30 minutos numa potencia fixa de 150 W e 40 kHz e a triagem de agentes biológicos para degradação de OTA. Para o estudo da degradação in vitro de OTA, método de extração e detecção de OTA e OTα em CLAEFL foi validado e o processo de degradação foi realizado com Rhizopus oryzae e Trichoderma reesei, obtendo-se uma redução máxima de 63,5% e 57,7%, respectivamente. A degradação apresentou uma correlação alta (R>0,9) e significativa (p<0,05) com a produção de Otα, indicando que ocorreu a produção de enzimas capazes de hidrolisar a micotoxina, por exemplo, a carboxipeptidase A. O estudo da toxicidade de OTA e seu metabólito OTα foi realizado em neutrófilos humanos, onde foi observado a ausência de efeito tóxico de OTα. Também foi determinado o mecanismo de toxicidade de OTA pelo aumento de Ca2+ intracelular pela liberação a partir das reservas internas. Esta liberação, subsequentemente, provoca uma cascata de eventos, nomeadamente: a produção de espécies reativas, depleção de ATP, perda de ΔΨm, levando à morte por necrose. Para reduzir o risco de exposição à micotoxina pela ingestão de matéria prima contaminada, carboxipeptidase A extraída de diferentes fontes foi aplicada na hidrólise de OTA em farinha de trigo para posterior determinação do conteúdo residual de OTA e OTα, empregando método validado. O estudo mostrou uma redução de OTA entre 16,8 e 78,5% e produção de OTα entre 2 a 8,2 ng.g-1 . As carboxipeptidases mais promissoras para degradação foram as provenientes de Rhizopus e Trichoderma e a carboxipeptidase comercial. Ficou demonstrado que se pode recomendar a aplicação de enzimas proteolíticas, tipo carboxipeptidase, para reduzir o risco de exposição à micotoxina quando utilizada matéria prima contaminada, por exemplo, farinha de trigo para diferentes processos. A transformação de OTA para OTα e seus efeitos na redução da toxicidade da micotoxina corroboram com esta afirmação.

Tin Oxide Based Composites Derived Using Electrostatic Spray Deposition Technique as Anodes for Li-Ion Batteries

Recent advances in the electric & hybrid electric vehicles and rapid developments in the electronic devices have increased the demand for high power and high energy density lithium ion batteries. Graphite (theoretical specific capacity: 372 mAh/g) used in commercial anodes cannot meet these demands. Amorphous SnO2 anodes (theoretical specific capacity: 781 mAh/g) have been proposed as alternative anode materials. But these materials have poor conductivity, undergo a large volume change during charging and discharging, large irreversible capacity loss leading to poor cycle performances. To solve the issues related to SnO2 anodes, we propose to synthesize porous SnO2 composites using electrostatic spray deposition technique. First, porous SnO2/CNT composites were fabricated and the effects of the deposition temperature (200,250, 300 oC) & CNT content (10, 20, 30, 40 wt %) on the electrochemical performance of the anodes were studied. Compared to pure SnO2 and pure CNT, the composite materials as anodes showed better discharge capacity and cyclability. 30 wt% CNT content and 250 oC deposition temperature were found to be the optimal conditions with regard to energy capacity whereas the sample with 20% CNT deposited at 250 oC exhibited good capacity retention. This can be ascribed to the porous nature of the anodes and the improvement in the conductivity by the addition of CNT. Electrochemical impedance spectroscopy studies were carried out to study in detail the change in the surface film resistance with cycling. By fitting EIS data to an equivalent circuit model, the values of the circuit components, which represent surface film resistance, were obtained. The higher the CNT content in the composite, lower the change in surface film resistance at certain voltage upon cycling. The surface resistance increased with the depth of discharge and decreased slightly at fully lithiated state. Graphene was also added to improve the performance of pure SnO2 anodes. The composites heated at 280 oC showed better energy capacity and energy density. The specific capacities of as deposited and post heat-treated samples were 534 and 737 mAh/g after 70 cycles. At the 70th cycle, the energy density of the composites at 195 °C and 280 °C were 1240 and 1760 Wh/kg, respectively, which are much higher than the commercially used graphite electrodes (37.2-74.4 Wh/kg). Both SnO2/CNTand SnO2/grapheme based composites with improved energy densities and capacities than pure SnO2 can make a significant impact on the development of new batteries for electric vehicles and portable electronics applications.

A resistant genetic background leading to incomplete penetrance of intestinal neoplasia and reduced loss of heterozygosity in ApcMin/+ mice

Previous studies of Min/+ (multiple intestinal neoplasia) mice on a sensitive genetic background, C57BL/6 (B6), showed that adenomas have lost heterozygosity for the germ-line ApcMin mutation in the Apc (adenomatous polyposis coli) gene. We now report that on a strongly resistant genetic background, AKR/J (AKR), Min-induced adenoma multiplicity is reduced by about two orders of magnitude compared with that observed on the B6 background. Somatic treatment with a strong mutagen increases tumor number in AKR Min/+ mice in an age-dependent manner, similar to results previously reported for B6 Min/+ mice. Immunohistochemical analyses indicate that Apc expression is suppressed in all intestinal tumors from both untreated and treated AKR Min/+ mice. However, the mechanism of Apc inactivation in AKR Min/+ mice often differs from that observed for B6 Min/+ mice. Although loss of heterozygosity is observed in some tumors, a significant percentage of tumors showed neither loss of heterozygosity nor Apc truncation mutations. These results extend our understanding of the effects of genetic background on Min-induced tumorigenesis in several ways. First, the AKR strain carries modifiers of Min in addition to Mom1. This combination of AKR modifiers can almost completely suppress spontaneous intestinal tumorigenesis associated with the Min mutation. Second, even on such a highly resistant genetic background, tumor formation continues to involve an absence of Apc function. The means by which Apc function is inactivated is affected by genetic background. Possible scenarios are discussed.

Recycling endosome membrane incorporation into the leading edge regulates lamellipodia formation and macrophage migration

In comparison to our knowledge of the recycling of adhesion receptors and actin assembly, exactly how the cell controls its surface membrane to form a lamellipodium during migration is poorly understood. Here, we show the recycling endosome membrane is incorporated into the leading edge of a migrating cell to expand lamellipodia membrane. We have identified the SNARE complex that is necessary for fusion of the recycling endosome with the cell surface, as consisting of the R-SNARE VAMP3 on the recycling endosome partnering with the surface Q-SNARE Stx4/SNAP23, which was found to translocate and accumulate on the leading edge of migrating cells. Increasing VAMP3-mediated fusion of the recycling endosome with the surface increased membrane ruffling, while inhibition of VAMP3-mediated fusion showed that incorporation of the recycling endosome is necessary for efficient lamellipodia formation. At the same time, insertion of this recycling endosome membrane also delivers its cargo integrin α5β1 to the cell surface. The loss of this extra membrane for lamellipodia expansion and delivery of cargo in cells resulted in macrophages with a diminished capacity to effectively migrate. Thus, the recycling endosome membrane is incorporated into the leading edge and this aids expansion of the lamellipodia and simultaneously delivers integrins necessary for efficient cell migration.

A new method to detect loss of heterozygosity using cohort heterozygosity comparisons

Background Loss of heterozygosity (LOH) is an important marker for one of the 'two-hits' required for tumor suppressor gene inactivation. Traditional methods for mapping LOH regions require the comparison of both tumor and patient-matched normal DNA samples. However, for many archival samples, patient-matched normal DNA is not available leading to the under-utilization of this important resource in LOH studies. Here we describe a new method for LOH analysis that relies on the genome-wide comparison of heterozygosity of single nucleotide polymorphisms (SNPs) between cohorts of cases and un-matched healthy control samples. Regions of LOH are defined by consistent decreases in heterozygosity across a genetic region in the case cohort compared to the control cohort. Methods DNA was collected from 20 Follicular Lymphoma (FL) tumor samples, 20 Diffuse Large B-cell Lymphoma (DLBCL) tumor samples, neoplastic B-cells of 10 B-cell Chronic Lymphocytic Leukemia (B-CLL) patients and Buccal cell samples matched to 4 of these B-CLL patients. The cohort heterozygosity comparison method was developed and validated using LOH derived in a small cohort of B-CLL by traditional comparisons of tumor and normal DNA samples, and compared to the only alternative method for LOH analysis without patient matched controls. LOH candidate regions were then generated for enlarged cohorts of B-CLL, FL and DLBCL samples using our cohort heterozygosity comparison method in order to evaluate potential LOH candidate regions in these non-Hodgkin's lymphoma tumor subtypes. Results Using a small cohort of B-CLL samples with patient-matched normal DNA we have validated the utility of this method and shown that it displays more accuracy and sensitivity in detecting LOH candidate regions compared to the only alternative method, the Hidden Markov Model (HMM) method. Subsequently, using B-CLL, FL and DLBCL tumor samples we have utilised cohort heterozygosity comparisons to localise LOH candidate regions in these subtypes of non-Hodgkin's lymphoma. Detected LOH regions included both previously described regions of LOH as well as novel genomic candidate regions. Conclusions We have proven the efficacy of the use of cohort heterozygosity comparisons for genome-wide mapping of LOH and shown it to be in many ways superior to the HMM method. Additionally, the use of this method to analyse SNP microarray data from 3 common forms of non-Hodgkin's lymphoma yielded interesting tumor suppressor gene candidates, including the ETV3 gene that was highlighted in both B-CLL and FL.

Inhibition of the JAK2/STAT3 pathway in ovarian cancer results in the loss of cancer stem cell-like characteristics and a reduced tumor burden

Background Current treatment of ovarian cancer patients with chemotherapy leaves behind a residual tumor which results in recurrent ovarian cancer within a short time frame. We have previously demonstrated that a single short-term treatment of ovarian cancer cells with chemotherapy in vitro resulted in a cancer stem cell (CSC)-like enriched residual population which generated significantly greater tumor burden compared to the tumor burden generated by control untreated cells. In this report we looked at the mechanisms of the enrichment of CSC-like residual cells in response to paclitaxel treatment. Methods The mechanism of survival of paclitaxel-treated residual cells at a growth inhibitory concentration of 50% (GI50) was determined on isolated tumor cells from the ascites of recurrent ovarian cancer patients and HEY ovarian cancer cell line by in vitro assays and in a mouse xenograft model. Results Treatment of isolated tumor cells from the ascites of ovarian cancer patients and HEY ovarian cancer cell line with paclitaxel resulted in a CSC-like residual population which coincided with the activation of Janus activated kinase 2 (JAK2) and signal transducer and activation of transcription 3 (STAT3) pathway in paclitaxel surviving cells. Both paclitaxel-induced JAK2/STAT3 activation and CSC-like characteristics were inhibited by a low dose JAK2-specific small molecule inhibitor CYT387 (1 μM) in vitro. Subsequent, in vivo transplantation of paclitaxel and CYT387-treated HEY cells in mice resulted in a significantly reduced tumor burden compared to that seen with paclitaxel only-treated transplanted cells. In vitro analysis of tumor xenografts at protein and mRNA levels demonstrated a loss of CSC-like markers and CA125 expression in paclitaxel and CYT387-treated cell-derived xenografts, compared to paclitaxel only-treated cell-derived xenografts. These results were consistent with significantly reduced activation of JAK2 and STAT3 in paclitaxel and CYT387-treated cell-derived xenografts compared to paclitaxel only-treated cell derived xenografts. Conclusions This proof of principle study demonstrates that inhibition of the JAK2/STAT3 pathway by the addition of CYT387 suppresses the ‘stemness’ profile in chemotherapy-treated residual cells in vitro, which is replicated in vivo, leading to a reduced tumor burden. These findings have important implications for ovarian cancer patients who are treated with taxane and/or platinum-based therapies. Keywords: Ovarian carcinoma, Cancer stem cell, Metastasis, Ascites, Chemoresistance, Recurrence, JAK2/STAT3 pathway