The hypoxic tumor microenvironment regulates the LPA signaling axis to promote cancer cell invasion and metastasis

Le développement des métastases est la cause principale de mortalité des patients atteints de cancer, mais demeure un obstacle majeur aux traitements. L'hypoxie, une caractéristique commune des tumeurs solides, est fortement impliquée dans l'invasion cellulaire et le développement des métastases, mais les mécanismes sous-jacents demeurent méconnus. La signalisation du LPA joue un rôle important au cours de la tumorigenèse et du développement des métastases, les membres de cette voie étant souvent régulés à la hausse dans les cellules tumorales. La signalisation du LPA a également été impliquée dans la production de structures de dégradation, les invadopodes, qui sont nécessaires à la formation de métastases. Des études récentes indiquent que la formation d'invadopodes est également induite par l'hypoxie. Par conséquent, nous avons voulu élucider l'influence du microenvironnement hypoxique sur l'axe de signalisation du LPA et si celui-ci joue un rôle dans la production d'invadopodes et la formation de métastases. Nous avons découvert que le LPA1 est un récepteur utilisé de façon commune et majoritaire pour la production d'invadopodes induite par l'hypoxie, et ce, dans diverses lignées cellulaires cancéreuses. Nous avons démontré que l'hypoxie favorise la formation d'invadopodes en utilisant une voie de signalisation distincte qui implique une communication croisée entre le récepteur LPA1 et l'EGFR, médiée par la kinase Src, Dans ce contexte, l'inhibition combinée du LPA1 et de l'EGFR agit en synergie afin d’empêcher la formation de métastases spontanées. Étant donné que la toxicité et la résistance aux inhibiteurs de l'EGFR représentent un défi important pour les patients atteints de cancer, ce travail permet l’identification d’une cible potentielle, le LPA1, qui pourrait être inhibée conjointement avec l'EGFR dans le but d’améliorer la survie de ces patients. D'autres études sur la régulation hypoxique de l'axe de signalisation du LPA ont démontré que l'hypoxie peut contrôler les niveaux d'expression des enzymes impliqués dans la production (ATX) et la dégradation (LPP1 / LPP3) du LPA, des évènements qui conduisent à une production accrue d'invadopodes. L'hypoxie permet également de modifier la localisation de ces protéines, ce qui pourrait constituer un mécanisme additionnel de régulation de l’axe de signalisation du LPA en hypoxie. Notre travail suggère que l'hypoxie est un régulateur important de l'axe de signalisation du LPA menant à l’invasion et à la formation de métastases. Par conséquent, les thérapies ciblant cet axe pourraient être bénéfiques pour contrer les effets néfastes de l'hypoxie tumorale sur la survie des patients atteints de cancer. De plus, un traitement combiné, ciblant le LPA1 et l’EGFR, pourrait être utile afin de réduire les effets secondaires et la résistance aux inhibiteurs de l'EGFR. Des études supplémentaires seront nécessaires afin de valider le potentiel thérapeutique de ce type de traitement.Abstract : Metastasis is the leading cause of cancer patient mortality yet remains a major hurdle for treatment. Hypoxia, a common feature of solid tumors, has been critically involved in cell invasion and metastasis but the underlying mechanisms remain poorly understood. The LPA signaling axis plays an important role during tumorigenesis and metastasis, with members of this pathway often being upregulated in tumor cells. LPA signaling has also been implicated in production of the degradative structures invadopodia, which are known to be required for metastasis. Interestingly, formation of invadopodia can also be induced by hypoxia. Therefore, we endeavoured to elucidate the influence of the hypoxic tumor microenvironment on the LPA signaling axis and whether this could play a role in invadopodia production and metastasis. We uncovered LPA1 as a common and major receptor used for hypoxia-induced invadopodia production in various cancer cell lines. We demonstrated that hypoxia promotes invadopodia formation through a distinct signaling pathway that involves Src-mediated cross-communication between LPA1 and EGFR, and that combined inhibition of LPA1 and EGFR acts synergistically to impede spontaneous metastasis. Since EGFR inhibitor toxicity and resistance represents a current challenge for cancer patients, this work identifies a potential target, LPA1 that could be inhibited in conjunction with EGFR to improve patient outcomes. Further studies into hypoxic regulation of the LPA signaling axis demonstrated that hypoxia can control the expression levels of LPA producing (ATX) and degrading (LPP1/LPP3) enzymes, events that lead to increased invadopodia production. Hypoxia was also found to alter the localization of these proteins, uncovering an additional mechanism of hypoxic regulation. Our work suggests that hypoxia is a master regulator of the LPA signaling axis that leads to metastasis, therefore therapies targeting this axis could be beneficial to counteract the detrimental effects of tumor hypoxia on cancer patient survival. Furthermore, LPA1-EGFR combination therapy could be a useful strategy to reduce EGFR inhibitor side effects and resistance and therefore warrants further studies to evaluate the potential of combination therapies in cancer patients

Capturing Breathy Voice: Durational Measures of Oral Stops in Marathi

The present study investigates a series of techniques used to capture the durational differences of oral stops in Marathi, an Indic language that exhibits a four-way phonemic distinction among oral stops

The Nature of Optional Sibilant Harmony in Navajo

This thesis represents investigates optional sibilant harmony in Navajo using the first person possessive morpheme, which contains an underlyingly palatal sibilant that may harmonize to alveolar when affixed to noun stems that contain [+anterior] sibilants. The literature commonly describes sibilant harmony as being mandatory in Navajo when sibilants are in adjacent syllables, and optional when there is more distance between sibilants. In other words, sibilant disharmony is ungrammatical, but gradiently rather than categorically; in some instances disharmony is ungrammatical enough that it must be repaired through assimilation, while in other instances it is less ungrammatical and may be tolerated. The statistical nature of the variation in these optional harmony settings is not fully understood, however, and the three studies contained within this thesis were designed to investigate how often assimilation occurs in nonmandatory environments and to identify factors that contribute to the variability observed. In the first study, a Google search was used to evaluate sibilant harmony in online Navajo language use in the Spring of 2008 and again in the Spring of 2010. The findings present a picture of optional sibilant harmony that differs somewhat from the traditional view; harmony seems to be optional even in the environment that has traditionally been described as mandatory, and it occurs far less frequently than anticipated. These results led to the creation of an online survey wherein fluent speakers of Navajo provided grammaticality judgments of both assimilated and unassimilated forms. Almost universally, respondents preferred the unassimilated shi- even in those environments where assimilation would previously have been considered mandatory. The third study involved the recording of data from three speakers of Navajo, none of whom use the assimilated si- either in writing or in speech--at least, not to a degree that is discernible to the naked ear. Acoustic analysis was performed to determine whether the prefix-initial palatal sibilant is acoustically consistent across the board--duration, spectral mean, onset of frication energy, and the second formant of the following vowel were measured to investigate whether the prefixal esh differs acoustically when it appears before words that contain potential triggers than when it does not. Analysis reveals some differences in the spectral mean and duration of the fricative portion of the first person possessive morpheme when it occurs before stems that contain [+anterior] sibilants. Taken together, the findings presented herein suggest that the mandatory sibilant harmony environment no longer exists in Navajo, at least with regards to the first person possessive morpheme. Harmony is far less prevalent than expected overall, and is wholly absent for some speakers. The factors of continuancy and adjacency were found to contribute significantly to the gradience observed in all three studies, however, even for those speakers who do not overtly use assimilation

Phonation Types in Marathi: An Acoustic Investigation

This dissertation presents a comprehensive instrumental acoustic analysis of phonation type distinctions in Marathi, an Indic language with numerous breathy voiced sonorants and obstruents. Important new facts about breathy voiced sonorants, which are crosslinguistically rare, are established: male and female speakers cue breathy phonation in sonorants differently, there are an abundance of trading relations, and--critically--phonation type distinctions are not cued as well by sonorants as by obstruents. Ten native speakers (five male, five female) were recorded producing Marathi words embedded in a carrier sentence. Tokens included plain and breathy voiced stops, affricates, nasals, laterals, rhotics, and approximants before the vowels [a] and [e]. Measures reported for consonants and subsequent vowels include duration, F0, Cepstral Peak Prominence (CPP), and corrected H1-H2*, H1-A1*, H1-A2*, and H1-A3* values. As expected, breathy voice is associated with decreased CPP and increased spectral values. A strong gender difference is revealed: low-frequency measures like H1-H2* cue breathy phonation more reliably in male speech, while CPP--which provides information about the aspiration noise included in the signal--is a more reliable cue in female speech. Trading relations are also reported: time and again, where one cue is weak or absent another cue is strong or present, underscoring the importance of including both genders and multiple vowel contexts when testing phonation type differences. Overall, the cues that are present for obstruents are not necessarily mirrored by sonorants. These findings are interpreted with reference to Dispersion Theory (Flemming 1995; Liljencrants & Lindblom 1972; Lindblom 1986, 1990). While various incarnations of Dispersion Theory focus on different aspects of perceptual and auditory distinctiveness, a basic claim is that one requirement for phonological contrasts is that they must be perceptually distinct: contrasts that are subject to great confusability are phonologically disfavored. The proposal, then, is that the typology of breathy voiced sonorants is due in part to the fact that they are not well differentiated acoustically. Breathy voiced sonorants are crosslinguistically rare because they do not make for strong phonemic contrasts

Life cycle cost analysis for joint elimination retrofits and thermal loading on Colorado bridges

2017 Spring.Includes bibliographical references.Bridge expansion joints are a particularly troublesome component of bridges and many Departments of Transportation (DOTs) are looking for a solution to deteriorating expansion joints on highway bridges. Bridge expansion joints create a break in the structural continuity of a bridge allowing clogging gravels and corroding chlorides to enter. They are designed to absorb thermal movements of the bridge between two bridge elements. There are three main issues regarding expansion joint: maintenance, knowledge about thermal movements, and costs. In order to prevent deterioration due to expansion joints the joints must be cleaned regularly and replaced promptly after failure. However, most DOTs do not have the personnel, time or resources to maintain expansion joints in their districts which leads to bridge deterioration. Other similar maintenance and component issues have been addressed using a Life Cycle Cost Analysis. For this to be used on expansion joints the three main issues of thermal knowledge, maintenance, and costs must first be addressed. The main goal of this project is to help transportation agencies make better decisions about bridge expansion joints. The specific objectives of this study are to 1) expand understanding of thermal loading effects on bridge expansion joints and 2) conduct a LCCA for joint elimination and retrofits for bridges in Colorado. These objectives were accomplished utilizing data from in field instrumentation and finite element models. The study has been developed jointly between the Colorado Department of Transportation (CDOT) and researchers at Colorado State University Three main tasks were conducted to achieve the objectives: 1) collect and analyze long-term thermal loading data from existing bridges to assess thermal loading impacts on joints; 2) perform a parametric study using a calibrated finite element model to further understanding of joint behavior and retrofit options under thermal loads; 3) perform a LCCA for bridge expansion joint retrofitting including impacts on bridge superstructure. The significance of this work includes the results of the data collection and analysis, the parametric study, and the LCCA findings. The preliminary data on the concrete bridge C-17-AT presented in this thesis only accounts for mid-winter temperatures. However, these limited observations do imply that if CDOT is interested in removing an expansion joint, the bridge superstructure and retrofit option would need to support the movement of the bridge. The parametric study and data analysis of thermal gradients indicate a stark need for further research into thermal gradients experienced by bridges. Finally, the LCCA concluded that a retrofit continuous bridge design would provide the most cost effective design by decreasing joint replacement costs and pier cap corrosion

L'autotaxin induit l'invasion des cellules cancéreuses via le récepteur de type 4 de l'acide lysophosphatidique

La formation des métastases est une propriété fondamentale des cellules cancéreuses malignes ainsi que cause principale de décès chez les patients atteints de cancer. Des études récent indique que l'invasion tumorale et la formation des métastases peut être initié par la formation des protrusions riches en actine et capables de dégrader la matrice extracellulaire, appeles des invadopodes. Cependant, malgré les recherches importantes sur la biologie des invadopodes, les informations concernant les initiateurs spécifiques de ces structures lors de la progression tumorale demeurent limitées. L'autotaxin (ATX) est une lysophospholipase sécrétée dont les niveaux d'expression corrèlent avec l'agressivité et le potentiel invasif des tumeurs. L'ATX produit l'acide lysophosphatidique (LPA), un phospholipide impliqué dans la progression tumorale qui agit par l'intermédiaire de récepteurs couplés aux protéines G, LPAt-6. Il a été récemment démontré que la surexpression del' ATX et des récepteurs LPAt-3 cause une augmentation de l'invasion tumorale et de la formation de métastases in vivo, cependant, le rôle d'autres récepteurs, soit les LPA4-6, ainsi que les mécanismes exacts par lesquels l'ATX induit la formation de métastases demeurent peu connus. Afin d'étudier l'influence del' ATX sur la production d'invadopodes, nous avons transfecté des cellules de fibrosarcome, les HT-1080, avec des gène codant soit pour l 'ATX ou des ARN m interférant. Ces cellules ont été testées dans des essais de production d'invadopodes utilisant de la matrice fluorescente et des techniques d'immunofluorescence afm de visualiser de façon simultanée la dégradation de la matrice et les composantes caractéristiques de ces structures. Nos résultats indiquent que l 'ATX est impliquée dans la formation et les fonctions des invadopodes. Par l'ajout du LPC ou du LPA, le substrat et le produit de l' ATX, nous avons montré que la production d'invadopodes est dépendante de la production de LPA du LPC. Parmi les récepteurs du LPA, le LPA4 possède l'expression la plus élevée chez les cellules HT-1080. Par le biais de shARNs spécifiques au LPA4 ainsi que d'agonistes et d'inhibiteurs de la voie de l'AMPc, nos résultats indiquent que la voie de signalisationAMPc-EPAC-Rapl, induite par l'activation du LPA4, régule la formation d'invadopodes en aval de l'ATX. De plus, l'inhibition de Racl, un effecteur connu de Rapl et de la formation d'invadopodes, abolit la production d'invadopodes induite par l'activation d' EPAC, suggérant la participation de Racl en aval de EPAC. Enfm, les résultats d'expériences utilisant des shARNs du LPA4 confrrment l'implication de ce récepteur dans l'invasion des cellules in vitro et la formation de métastases.Abstract: Tumor metastasis is a fundamental property of malignant cancer cells and the major cause of death in cancer patients. Recent studies indicate that tumor cell invasion and metastasis may be initiated by the formation of the actin-rich cell protrusions with ECM degradation activity, invadopodia. However, despite extensive research on the biology of invadopodia, very little is known about their specific inducers during tumor progression. Autotaxin (ATX) is a secreted lysophospholipase whose expression levels within tumors correlates strongly with their aggressiveness and invasiveness. ATX produces lyosophosphatidic acid (LPA), a phospholipid with known tumor promoting functions that acts through the G-protein coupled receptors, LPA[subscript 1-6] . Recently, overexpression of ATX and LPA receptors (LPA[subscript 1-3]) has been linked to increased tumor invasion and metastasis in vivo , however, the role of other LPA receptors (LPA[subscript 4-6]) as well as the exact mechanisms by which ATX induces tumor metastasis remain poorly characterized. In order to determine the involvement of ATX and LPA in invadopodia production, we used the fibrosarcoma HT-1080 cells stably transfected with ATX or shRNA targeting ATX in fluorescent matrix degradation assays. Our results demonstrate that ATX is implicated in the production of invadopodia resulting in an increase in both their formation and function. Using LPC or LPA, the substrate and product of ATX, we further show that invadopodia production is dependent on the production of LPA from LPC. Among the LPA receptors, LPA 4 has the highest expression in HT1080 cells. Using LPA[subscript 4] shRNA as well as agonists and inhibitors of the cAMP pathway, we provide evidence that LPA[subscript 4] signaling through the cAMP-EPAC-Rap1 axis, regulates invadopodia formation downstream of ATX. Furthermore, inhibition of Rac1, a known effector of Rap1 and invadopodia formation, abolished EPAC-induced invadopodia production, suggesting downstream participation of Rac1. Finally, results using LPA[subscript 4] shRNA support the requirement of this receptor for in vitro cell invasion and in vivo metastasis formation. Our results suggest that ATX through LPA[subscript 4] is a strong inducer of invadopodia formation that correlates with the ability of the cells to invade and metastasize. This study also revealed an unexpected signaling pathway for cell invasion involving LPA[subscript 4]-driven cAMP production and subsequent activation of the EPAC-Rap1-Rac1 axis

Homotopy theory of modules over operads in symmetric spectra

We establish model category structures on algebras and modules over operads in symmetric spectra, and study when a morphism of operads induces a Quillen equivalence between corresponding categories of algebras (resp. modules) over operads.Comment: Corrigendu