Overexpression of Bcl-2 is associated with apoptotic resistance to the G-quadruplex ligand 12459 but is not sufficient to confer resistance to long-term senescence

The triazine derivative 12459 is a potent G-quadruplex interacting agent that inhibits telomerase activity. This agent induces time- and dose-dependent telomere shortening, senescence-like growth arrest and apoptosis in the human A549 tumour cell line. We show here that 12459 induces a delayed apoptosis that activates the mitochondrial pathway. A549 cell lines selected for resistance to 12459 and previously characterized for an altered hTERT expression also showed Bcl-2 overexpression. Transfection of Bcl-2 into A549 cells induced a resistance to the short-term apoptotic effect triggered by 12459, suggesting that Bcl-2 is an important determinant for the activity of 12459. In sharp contrast, the Bcl-2 overexpression was not sufficient to confer resistance to the senescence-like growth arrest induced by prolonged treatment with 12459. We also show that 12459 provokes a rapid degradation of the telomeric G-overhang in conditions that paralleled the apoptosis induction. In contrast, the G-overhang degradation was not observed when apoptosis was induced by camptothecin. Bcl-2 overexpression did not modify the G-overhang degradation, suggesting that this event is an early process uncoupled from the final apoptotic pathway

3D collagen type I matrix inhibits the antimigratory effect of doxorubicin

<p>Abstract</p> <p>Background</p> <p>The cell microenvironment, especially extracellular matrix proteins, plays an important role in tumor cell response to chemotherapeutic drugs. The present study was designed to investigate whether this microenvironment can influence the antimigratory effect of an anthracycline drug, doxorubicin, when tumor cells are grown in a matrix of type I collagen, a three-dimensional (3D) context which simulates a natural microenvironment.</p> <p>Methods</p> <p>To this purpose, we studied the migratory parameters, the integrin expression, and the activation state of focal adhesion kinase (FAK) and GTPase RhoA involved in the formation of focal adhesions and cell movement. These parameters were evaluated at non toxic concentrations which did not affect HT1080 cell proliferation.</p> <p>Results</p> <p>We show that while doxorubicin decreased cell migration properties by 70% in conventional two-dimensional (2D) culture, this effect was completely abolished in a 3D one. Regarding the impact of doxorubicin on the focal adhesion complexes, unlike in 2D systems, the data indicated that the drug neither affected β1 integrin expression nor the state of phosphorylation of FAK and RhoA.</p> <p>Conclusion</p> <p>This study suggests the lack of antiinvasive effect of doxorubicin in a 3D environment which is generally considered to better mimic the phenotypic behaviour of cells <it>in vivo</it>. Consistent with the previously shown resistance to the cytotoxic effect in a 3D context, our results highlight the importance of the matrix configuration on the tumor cell response to antiinvasive drugs.</p

Airway Epithelial Cell Migration Dynamics: Mmp-9 Role in Cell–Extracellular Matrix Remodeling

Cell spreading and migration associated with the expression of the 92-kD gelatinase (matrix metalloproteinase 9 or MMP-9) are important mechanisms involved in the repair of the respiratory epithelium. We investigated the location of MMP-9 and its potential role in migrating human bronchial epithelial cells (HBEC). In vivo and in vitro, MMP-9 accumulated in migrating HBEC located at the leading edge of a wound and MMP-9 expression paralleled cell migration speed. MMP-9 accumulated through an actin-dependent pathway in the advancing lamellipodia of migrating cells and was subsequently found active in the extracellular matrix (ECM). Lamellipodia became anchored through primordial contacts established with type IV collagen. MMP-9 became amassed behind collagen IV where there were fewer cell–ECM contacts. Both collagen IV and MMP-9 were involved in cell migration because when cell–collagen IV interaction was blocked, cells spread slightly but did not migrate; and when MMP-9 activation was prevented, cells remained fixed on primordial contacts and did not advance at all. These observations suggest that MMP-9 controls the migration of repairing HBEC by remodeling the provisional ECM implicated in primordial contacts

Evaluation intégrée des mesures agro-environnementales territorialisées à enjeu "qualité des eaux" sur la période 2007 à 2011 : le projet MAEVEAU

The MAEVEAU project has developed an approach for an integrated assessment of effectiveness of regionalized Agro-Environmental Measures (MAET) intended to preserve water quality in relation to pesticides. This approach investigates the concept of efficiency through a triple analysis: the impact (net effects), the environmental cost-effectiveness and the role of organizational factors in the contracting process. The impact is assessed by a quasi-experimental approach by counterfactuals and examines adaptation of the matching method to the regionalized MAET. Cost-effectiveness analysis is based on integrated modeling spatially distributed coupling the agro-hydrological SWAT model, pesticides pressure indicators and a bio-economic model optimizing gross margin. The effectiveness of organizational factors focuses on transaction costs, the role of collective action and preferences for alternative contracts.La recherche conduite dans le projet MAEVEAU a développé une démarche d'évaluation intégrée de l'efficacité des Mesures Agro-Environnementales Territorialisées (MAET) à enjeu préservation de la qualité de l'eau vis-à-vis des pesticides sur la période 2007 à 2011. La question scientifique traite le concept d'efficacité de la politique en s'appuyant sur une triangulation des approches: une évaluation de l'impact (c'est-à-dire des effets propres de la politique), une évaluation coût-efficacité environnementale et une évaluation du rôle des facteurs organisationnels dans le processus d'adhésion. L'impact est évalué par une approche quasi-expérimentale par contrefactuel et questionne l'adaptation de la méthode du matching à la territorialisation des MAET. L'analyse coût-efficacité s'appuie sur une modélisation intégrée spatialisée couplant modèle agro-hydrologique, indicateurs pesticides spatialisés et optimisation économique des marges brutes. L'efficacité des facteurs organisationnels s'est intéressée aux coûts de transaction, au rôle de l'action collective et aux préférences pour des contrats alternatifs

Incidence of Sarcoma Histotypes and Molecular Subtypes in a Prospective Epidemiological Study with Central Pathology Review and Molecular Testing

International audienceBACKGROUND: The exact overall incidence of sarcoma and sarcoma subtypes is not known. The objective of the present population-based study was to determine this incidence in a European region (Rhone-Alpes) of six million inhabitants, based on a central pathological review of the cases. METHODOLOGY/PRINCIPAL FINDINGS: From March 2005 to February 2007, pathology reports and tumor blocks were prospectively collected from the 158 pathologists of the Rhone-Alpes region. All diagnosed or suspected cases of sarcoma were collected, reviewed centrally, examined for molecular alterations and classified according to the 2002 World Health Organization classification. Of the 1287 patients screened during the study period, 748 met the criteria for inclusion in the study. The overall crude and world age-standardized incidence rates were respectively 6.2 and 4.8 per 100,000/year. Incidence rates for soft tissue, visceral and bone sarcomas were respectively 3.6, 2.0 and 0.6 per 100,000. The most frequent histological subtypes were gastrointestinal stromal tumor (18%; 1.1/100,000), unclassified sarcoma (16%; 1/100,000), liposarcoma (15%; 0.9/100,000) and leiomyosarcoma (11%; 0.7/100,000). CONCLUSIONS/SIGNIFICANCE: The observed incidence of sarcomas was higher than expected. This study is the first detailed investigation of the crude incidence of histological and molecular subtypes of sarcomas

Etude de l'implication des récepteurs nicotiniques à l'acétylcholine dans le développement des cancers pulmonaires non à petites cellules

La progression tumorale est caractérisée par deux processus clés, la prolifération et l invasion cellulaires. Les nAChRs, activés par la nicotine et ses nitrosamines dérivées (NNN et NNK), modulent les concentrations calciques intracellulaires et activent in vitro la prolifération, l apoptose, la migration et l invasion de lignées cellulaires tumorales. Dans cette étude, nous montrons, en utilisant des cultures primaires de cellules dérivées de cancers pulmonaires non à petites cellules (carcinomes épidermoïdes et adénocarcinomes), que les nAChRs a7 régulent différemment la prolifération cellulaire en fonction du stade de différenciation des tumeurs. Le nAChR a7 agit comme répresseur de la prolifération cellulaire dans les tumeurs bien différenciées et dans l épithélium respiratoire normal, alors que dans les tumeurs peu différenciées, il stimule la prolifération cellulaire en réponse à la nicotine. A l inverse, le nAChR a3a5b2 n est que partiellement impliqué dans la régulation de la prolifération cellulaire aussi bien dans les tumeurs pulmonaires que dans l épithélium respiratoire normal. Les nAChRs a7 et nAChRs a3a5b2 sont tous les deux impliqués dans la stimulation de l invasion des cellules tumorales des carcinomes épidermoïdes et adénocarcinomes. Le polymorphisme non-synonyme rs16969968 de la sous-unité a5 induit une mutation au niveau d un acide aminé hautement conservé (D398N). De nombreuses études d association pangénomiques lient ce polymorphisme au développement des cancers pulmonaires. Dans cette étude nous montrons que les nAChRs exprimant la sous-unité a5 mutée (D398N) altèrent la prolifération et la différenciation des cellules respiratoires et modulent l invasion des cellules tumorales, en synergie avec les nAChRs a7.Tumor progression is characterized by two key processes, cell proliferation and invasion. Nicotinic receptors, activated by nicotine and its derived nitrosamines (NNK and NNN) modulate intracellular calcium concentrations and activate in vitro proliferation, apoptosis, migration and invasion of tumor cell lines. In this study, we show, by using primary cell cultures from lung cancer tumors, adenocarcinoma and squamous cell carcinoma, that nAChR a7 differently regulates cell proliferation according to the state of tumor differentiation. The a7 nAChRs acts as a repressor of cell proliferation in differentiated lung cancer tissues and in the normal respiratory epithelium, while it stimulates cell proliferation in response to nicotine, in poorly differentiated tumors. Conversely, the a3a5b2 nAChR is only partially involved in the regulation of cell proliferation in lung cancers and in the normal respiratory epithelium. The a7 and a3a5b2 nAChRs are both involved in the in vitro invasion process of adenocarcinoma and squamous cell carcinoma. Non-synonymous polymorphism rs16969968 in the CHRNA5 gene induces a mutation in a highly conserved amino acid (D398N). Many genome-wide association studies have demonstrated the relationship between this polymorphism and the incidence of lung cancer. In this study, we show that nAChRs, expressing the mutated a5 subunit (D398N), are in involved in the alteration of the proliferation and the differentiation state of respiratory epithelial cells, and also modulate tumor cell invasion, in synergy with the a7 nAChRs.REIMS-SCD-Bib. electronique (514549901) / SudocSudocFranceF

Airway epithelial repair, regeneration, and remodeling after injury in chronic obstructive pulmonary disease.

In chronic obstructive pulmonary disease (COPD), exacerbations are generally associated with several causes, including pollutants, viruses, bacteria that are responsible for an excess of inflammatory mediators, and proinflammatory cytokines released by activated epithelial and inflammatory cells. The normal response of the airway surface epithelium to injury includes a succession of cellular events, varying from the loss of the surface epithelium integrity to partial shedding of the epithelium or even complete denudation of the basement membrane. The epithelium then has to repair and regenerate to restore its functions, through several mechanisms, including basal cell spreading and migration, followed by proliferation and differentiation of epithelial cells. In COPD, the remodeling of the airway epithelium, such as squamous metaplasia and mucous hyperplasia that occur during injury, may considerably disturb the innate immune functions of the airway epithelium. In vitro and in vivo models of airway epithelial wound repair and regeneration allow the study of the spatiotemporal modulation of cellular and molecular interaction factors-namely, the proinflammatory cytokines, the matrix metalloproteinases and their inhibitors, and the intercellular adhesion molecules. These factors may be markedly altered during exacerbation periods of COPD and their dysregulation may induce remodeling of the airway mucosa and a leakiness of the airway surface epithelium. More knowledge of the mechanisms involved in airway epithelium regeneration may pave the way to cytoprotective and regenerative therapeutics, allowing the reconstitution of a functional, well-differentiated airway epithelium in COPD

Analyse du comportement dynamique de cellules épithéliales humaines invasives et non invasives dans des modèles de culture bi et tri-dimensionnels

L'invasion tumorale est reliée à des modifications de la cohésion cellulaire analysée par l'étude du comportement sociologique des cellules, grâce à des techniques de vidéomicroscopie. Deux lignées cellulaires bronchiques humaines ont été étudiées : les cellules 16HBE14o- non invasives et les cellules BZR invasives.En culture, les cellules 16HBE14o- sont caractérisées par une distribution en amas qui n'est pas due à une attraction mais à un mouvement aléatoire des cellules combiné à leurs propriétés intrinsèques, alors que les cellules BZR maintiennent une distribution spatiale aléatoire qui n'est pas due à une répulsion cellulaire. A partir d'un modèle de migration cellulaire 3D in vitro nous montrons que la vitesse de migration 3D des cellules BZR est supérieure à celle des cellules 16HBE14o-.Nos résultats ont permis également d'établir une corrélation entre le phénotype migratoire et la localisation du facteur tissulaire qui est potentiellement impliqué dans le processus tumoral@Tumour invasion is mainly related to alterations of the cellular cohesiveness which can be studied through the analysis of cell to cell behaviour by using videomicroscopic techniques. Two human bronchial cell lines were studied: the 16HBE14o- cell line which has no invasive capacity and the BZR cell line which is characterized by a high level of invasiveness.An in vitro model of 2D culture allowed to establish that the 16HBE14o- cell line rapidly formed clusters with a cohesive organization not related to an attraction, whereas the BRZ cell line remained isolated each other and was characterized by a non cohesive organization not related to a repulsion.An in vitro model of 3D culture allowed to observe that the 3D migration speed was significantly higher for the invasive BZR cells compared with the 3D migration speed of the non-invasive 16HBE14o- cells.The localization of Tissue Factor, potentially implicated in tumour process, is correlated to the migratory phenotype of cellsREIMS-BU Santé (514542104) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

Characterizing the spatio-temporal behavior of cell populations through image auto-and cross-correlation microscopy

FULL TEXT http://www.biotechniques.com/default.asp?page=aop&subsection=article_display&id=112478We propose two methods for characterizing the spatio-temporal behavior of cell populations in culture. The first method, image auto-correlation microscopy (IACM), allows us to characterize the variation in the number of objects as a function of time, thus enabling the quantification of the clustering properties of cell populations to be performed. The second method, image cross-correlation microscopy (ICCM), allows us to characterize the migration properties of cell populations. The latter method does not require estimation or measurement of the trajectories of individual cells, which is very demanding when populations of >100 cells are examined. The capabilities of the two methods are demonstrated with simulated cell populations, and their usefulness is illustrated with experiments involving invasive and noninvasive tumor cell populations