Wnt/β-Catenin Signaling Enhances Cyclooxygenase-2 (COX2) Transcriptional Activity in Gastric Cancer Cells

BACKGROUND: Increased expression of the cyclooxygenase-2 enzyme (COX2) is one of the main characteristics of gastric cancer (GC), which is a leading cause of death in the world, particularly in Asia and South America. Although the Wnt/β-catenin signaling pathway has been involved in the transcriptional activation of the COX2 gene, the precise mechanism modulating this response is still unknown. METHODOLOGY/PRINCIPAL FINDINGS: Here we studied the transcriptional regulation of the COX2 gene in GC cell lines and assessed whether this phenomenon is modulated by Wnt/β-catenin signaling. We first examined the expression of COX2 mRNA in GC cells and found that there is a differential expression pattern consistent with high levels of nuclear-localized β-catenin. Pharmacological treatment with either lithium or valproic acid and molecular induction with purified canonical Wnt3a significantly enhanced COX2 mRNA expression in a dose- and time-dependent manner. Serial deletion of a 1.6 Kbp COX2 promoter fragment and gain- or loss-of-function experiments allowed us to identify a minimal Wnt/β-catenin responsive region consisting of 0.8 Kbp of the COX2 promoter (pCOX2-0.8), which showed maximal response in gene-reporter assays. The activity of this pCOX2-0.8 promoter region was further confirmed by site-directed mutagenesis and DNA-protein binding assays. CONCLUSIONS/SIGNIFICANCE: We conclude that the pCOX2-0.8 minimal promoter contains a novel functional T-cell factor/lymphoid enhancer factor (TCF/LEF)-response element (TBE Site II; -689/-684) that responds directly to enhanced Wnt/β-catenin signaling and which may be important for the onset/progression of GC

Regulatory Role of the RUNX2 Transcription Factor in Lung Cancer Apoptosis

Q4Pacientes con Cáncer de pulmónLung cancer is the leading cause of cancer death globally. Numerous factors intervene in the onset and progression of lung tumors, among which the participation of lineage-specific transcription factors stands out. Several transcription factors important in embryonic development are abnormally expressed in adult tissues and thus participate in the activation of signaling pathways related to the acquisition of the tumor phenotype. RUNX2 is the transcription factor responsible for osteogenic differentiation in mammals. Current studies have confirmed that RUNX2 is closely related to the proliferation, invasion, and bone metastasis of multiple cancer types, such as osteosarcoma, breast cancer (BC), prostate cancer, gastric cancer, colorectal cancer, and lung cancer. Thus, the present study is aimed at evaluating the role of the RUNX2 transcription factor in inhibiting the apoptosis process. Loss-of-function assays using sh-RNA from lentiviral particles and coupled with Annexin/propidium iodide (PI) assays (flow cytometry), immunofluorescence, and quantitative PCR analysis of genes related to cell apoptosis (BAD, BAX, BCL2, BCL-XL, and MCL1) were performed. Silencing assays and Annexin/PI assays demonstrated that when RUNX2 was absent, the percentage of dead cells increased, and the expression levels of the BCL2, BCL-XL, and MCL1 genes were downregulated. Furthermore, to confirm whether the regulatory role of RUNX2 in the expression of these genes is related to its binding to the promoter region, we performed chromatin immunoprecipitation (ChIP) assays. Here, we report that overexpression of the RUNX2 gene in lung cancer may be related to the inhibition of the intrinsic apoptosis pathway, specifically, through direct transcriptional regulation of the antiapoptotic gene BCL2 and indirect regulation of BCL-XL and MCL1.https://orcid.org/0000-0003-1555-6661https://orcid.org/0000-0002-5668-0266https://orcid.org/0000-0003-3975-2835https://orcid.org/0000-0001-8528-4433Revista Internacional - IndexadaCN

Enhanced CRAd Activity Using Enhancer Motifs Driven by a Nucleosome Positioning Sequence

Cancer development involves changes driven by the epigenetic machinery, including nucleosome positioning. Recently, the concept that adenoviral replication may be driven by tumor specific promoters (TSPs) gained support, and several conditionally replicative adenoviruses (CRAd) exhibited therapeutic efficacy in clinical trials. Here, we show for the first time that placing a nucleosome positioning sequence (NPS) upstream of a TSP combined with Wnt-responsive motifs (pART enhancer) enhanced the TSP transcriptional activity and increased the lytic activity of a CRAd. pART enhanced the transcriptional activity of the gastrointestinal cancer (GIC)-specific REG1A promoter (REG1A-pr); moreover, pART also increased the in vitro lytic activity of a CRAd whose replication was driven by REG1A-Pr. The pART enhancer effect in vitro and in vivo was strictly dependent on the presence of the NPS. Indeed, deletion of the NPS was strongly deleterious for the in vivo antitumor efficacy of the CRAd on orthotopically established pancreatic xenografts. pART also enhanced the specific activity ofmother heterologous promoters; moreover, the NPS was also able to enhance the responsiveness of hypoxia- and NFκ B-response elements. We conclude that NPS could be useful for gene therapy approaches in cancer as wellas other diseases.Fil: Bravo, Soraya. Universidad Andrés Bello; ChileFil: Núñez Aguilera, Felipe Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Cruzat, Fernando. Universidad de Concepción; ChileFil: Cafferata, Eduardo Gustavo Alfredo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: De Ferrari, Giancarlo V. Universidad Andrés Bello; ChileFil: Montecino, Martín. Universidad Andrés Bello; ChileFil: Podhajcer, Osvaldo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

Widespread loss of the silencing epigenetic mark H3K9me3 in astrocytes and neurons along with hippocampal-dependent cognitive impairment in C9orf72 BAC transgenic mice

Background: Hexanucleotide repeat expansions of the G4C2 motif in a non-coding region of the C9ORF72 gene are the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Tissues from C9ALS/FTD patients and from mouse models of ALS show RNA foci, dipeptide-repeat proteins, and notably, widespread alterations in the transcriptome. Epigenetic processes regulate gene expression without changing DNA sequences and therefore could account for the altered transcriptome profiles in C9ALS/FTD; here, we explore whether the critical repressive marks H3K9me2 and H3K9me3 are altered in a recently developed C9ALS/FTD BAC mouse model (C9BAC). Results: Chromocenters that constitute pericentric constitutive heterochromatin were visualized as DAPI- or Nucblue-dense foci in nuclei. Cultured C9BAC astrocytes exhibited a reduced staining signal for H3K9me3 (but not for H3K9me2) at chromocenters that was accompanied by a marked decline in the global nuclear level of this mark. Similar depletion of H3K9me3 at chromocenters was detected in astrocytes and neurons of the spinal cord, motor cortex, and hippocampus of C9BAC mice. The alterations of H3K9me3 in the hippocampus of C9BAC mice led us to identify previously undetected neuronal loss in CA1, CA3, and dentate gyrus, as well as hippocampal-dependent cognitive deficits. Conclusions: Our data indicate that a loss of the repressive mark H3K9me3 in astrocytes and neurons in the central nervous system of C9BAC mice represents a signature during neurodegeneration and memory deficit of C9ALS/FTD. © 2020 The Author(s).Indexación: Scopu

Primary lung cancer cell culture from transthoracic needle biopsy samples

Artículo de investigación1-14Lung cancer is the leading cause of cancer death in the world. The high mortality rate of this pathology is directly related to its late detection, since its symptoms can be masked by other diseases of lower risk. Although in recent years the number of research related to this subject has increased, molecular mechanisms that trigger this disease remains poorly understood. Experimental models are therefore vital for use in research. Immortalized cell lines have inherent limitations. Explanted tumoral cells obtained by transthoracic needle biopsy can be a potential source of primary culture of human lung tumor cells. Tumor specimens from 14 patients suspected of primary or metastatic lung cancer were obtained by CT-guided transthoracic lung biopsy. Solid tumors were mechanically disaggregated under a stereoscope. Cells were cultured in Base C growth media supplemented with 5% fetal bovine serum in 24-well cell culture plates. Primary lung cancer cell culture was successfully cultured from 12 out of 14 patients. Once a confluent monolayer was obtained, cells were enzymatically harvested and passaged to Petri culture dishes. These primary cell cultures were characterized by cytogenetic tests and gene expression analysis of diagnostic markers. These primary cell cultures revealed chromosome rearrangements and changes in their chromosome complement typical of tumoral cells. Additionally, Fluorescence in situ hybridization analysis demonstrated that three cultures exhibited EGFR amplification. Finally, expression profiles of CK7, NAPSIN A, TTF1, and P63 genes allowed in some cases to confirm sample tumor phenotype. These results demonstrate that primary lung cancer cell culture is possible from percutaneous puncture and provides an important biological source to asses and investigate the molecular mechanisms of lung cancer

EstuPlan: Methodology for the development of creativity in the resolution of scientific and social problems

[EN] Creative thinking is necessary to generate novel ideas and solve problems. "EstuPlan" is a methodology in which knowledge and creativity converge for the resolution of scientific problems with social projection. It is a training programme that integrates teachers, laboratory technicians and PhD students, master and undergraduate students which form working groups for the development of projects. Projects have a broad and essential scope and projection in terms of environmental problems, sustainable use of natural resources, food, health, biotechnology or biomedicine. The results show the success of this significant learning methodology using tools to develop creativity in responding to scientific and social demand for problem-solving to transfer academic knowledge to different professional environments. Bioplastics, Second Life of Coffee, LimBio, Algae oils, Ecomers, Caring for the life of your crop and Hate to Deforestate are currently being developed.Astudillo Calderón, S.; De Díez De La Torre, L.; García Companys, M.; Ortega Pérez, N.; Rodríguez Martínez, V.; Alzahrani, S.; Alonso Valenzuela, R.... (2019). EstuPlan: Methodology for the development of creativity in the resolution of scientific and social problems. En HEAD'19. 5th International Conference on Higher Education Advances. Editorial Universitat Politècnica de València. 711-717. https://doi.org/10.4995/HEAD19.2019.9205OCS71171

Evidence supporting that human-subsidized free-ranging dogs are the main cause of animal losses in small-scale farms in Chile

Abstract We surveyed professionals from the Chilean Ministry of Agriculture working with small-scale farmers to characterize the attacks of free-ranging dogs across Chile. Nationwide, in a single year, free-ranging dogs attacked 25% of the ca. 8500 farms included in the survey, killing or injuring about 10 000 small ruminants. These dogs were ranked as the main cause of animal losses for small-scale farmers, representing a threat to the livelihoods of this vulnerable group. Further, free-ranging dogs attacking small ruminants were considered as human-subsidized, since they would be recruited by irresponsible ownership and abandonment from urban centers. This is the first national assessment reporting that human-subsidized dogs are a main threat to livestock rearing. Policies to control populations of these animals should target their anthropogenic origin as well as cultural shifts in dog ownership and animal welfare. While these policies may be effective mid- to long-term approaches, short-term actions may also be needed

Stochastic Modeling of Complex Systems and Systems Biology: From Stochastic Transition Systems to Hybrid Systems

International audienceModeling complex dynamic systems requires to reuse and to combine models in a non-ambiguous way, integrating processes with different information levels and temporal dependences. System behaviors are consequence of interacting processes, which are affected by external factors often not controlled. In particular, biological functions are the result of processes that connect different hierarchy levels, associated by physical and chemical relations (H. Kitano (2002)). Each process works in different way and it is common to observe that changes in the conditions, such as quantity of nutrients or environment, modify the behavior of the systems. The firstt approach we discuss is the use of Stochastic Transition Systems (STSs, L.D. Alfaro (1998)). It considers the dynamics of system variables given by transitions, changing their values, described by Markov processes with continuous time. Transitions are provoked by specific conditions of system variables, which can be ambiguous and generate non-determinism. Although STSs allow us to incorporate randomness and non-determinism, we do not capture the complexity of behaviors nor the continuity of the variables. To describe the behavior of complex systems over time, it is convenient to combine different types of models: continuous models for gradual changes, discrete models for instantaneous changes, deterministic models for completely predictable behaviors, and stochastic or non-deterministic models to describe behaviors with imprecise or incomplete information. To do that, we use the Hybrid Systems theory and the composition of models. System variables evolve according to continuous models, but deterministic, stochastic and non-deterministic transitions can change the definition of these models. Composition is the action of combining different models into an integrated model. We connect them by using input-output relations, and by processes synchronization (e.g. activation or repression signals). A very intuitive example of hybrid system is the motion of an automobile with a manual gearbox. The dynamics of the velocity and position evolve in a continuous specific way depending of the engaged gear. With an automatic gearbox the gear changes are deterministic, but if it is manual many factors influence the decisions of the driver, and the transitions are stochastic or non-deterministic. According to the form of the model changes, we consider hybrid systems with coefficient switches, or with strong switches. For coefficient switches, the transitions provoke changes in specific coefficients of the continuous model. For strong switches, transitions control the activation of models allowing radical changes. First type favors the interpretation of the effect of transitions on the continuous model, while strong switches are useful for reconciling models with different nature (R. Assar (2011)). This approach allows us to build more complete descriptions of complex biological systems. As application, we built a hybrid model of the osteo-adipo differentiation process. It combines known validated models to predict the bone and fat formation in response to activation of pathways such as the Wnt pathway, stochastic factors, and changes of conditions affecting these functions (R. Assar et al. (2012)). This model is our first phase to simulate physiological responses to treatments of bone mass disorders in silico, and to explore the efficiency of new medical strategies before testing them in vitro or in vivo

Stochastic Modeling of Complex Systems and Systems Biology: From Stochastic Transition Systems to Hybrid Systems

International audienceModeling complex dynamic systems requires to reuse and to combine models in a non-ambiguous way, integrating processes with different information levels and temporal dependences. System behaviors are consequence of interacting processes, which are affected by external factors often not controlled. In particular, biological functions are the result of processes that connect different hierarchy levels, associated by physical and chemical relations (H. Kitano (2002)). Each process works in different way and it is common to observe that changes in the conditions, such as quantity of nutrients or environment, modify the behavior of the systems. The firstt approach we discuss is the use of Stochastic Transition Systems (STSs, L.D. Alfaro (1998)). It considers the dynamics of system variables given by transitions, changing their values, described by Markov processes with continuous time. Transitions are provoked by specific conditions of system variables, which can be ambiguous and generate non-determinism. Although STSs allow us to incorporate randomness and non-determinism, we do not capture the complexity of behaviors nor the continuity of the variables. To describe the behavior of complex systems over time, it is convenient to combine different types of models: continuous models for gradual changes, discrete models for instantaneous changes, deterministic models for completely predictable behaviors, and stochastic or non-deterministic models to describe behaviors with imprecise or incomplete information. To do that, we use the Hybrid Systems theory and the composition of models. System variables evolve according to continuous models, but deterministic, stochastic and non-deterministic transitions can change the definition of these models. Composition is the action of combining different models into an integrated model. We connect them by using input-output relations, and by processes synchronization (e.g. activation or repression signals). A very intuitive example of hybrid system is the motion of an automobile with a manual gearbox. The dynamics of the velocity and position evolve in a continuous specific way depending of the engaged gear. With an automatic gearbox the gear changes are deterministic, but if it is manual many factors influence the decisions of the driver, and the transitions are stochastic or non-deterministic. According to the form of the model changes, we consider hybrid systems with coefficient switches, or with strong switches. For coefficient switches, the transitions provoke changes in specific coefficients of the continuous model. For strong switches, transitions control the activation of models allowing radical changes. First type favors the interpretation of the effect of transitions on the continuous model, while strong switches are useful for reconciling models with different nature (R. Assar (2011)). This approach allows us to build more complete descriptions of complex biological systems. As application, we built a hybrid model of the osteo-adipo differentiation process. It combines known validated models to predict the bone and fat formation in response to activation of pathways such as the Wnt pathway, stochastic factors, and changes of conditions affecting these functions (R. Assar et al. (2012)). This model is our first phase to simulate physiological responses to treatments of bone mass disorders in silico, and to explore the efficiency of new medical strategies before testing them in vitro or in vivo

Seguros obligatorios en la legislacion chilena y su caracter en materia comercial.

181 p.Analiza detalladamente los contratos de Seguro Obligatorio, para luego estudiar los principios dentro de la contratación; posteriormente aborda el tema del seguro desde una perspectiva comercial, para luego centrarse en el análisis de la naturaleza jurídica de este ya citado contrato