The LKB1 Tumor Suppressor as a Biomarker in Mouse and Human Tissues

Germline mutations in the LKB1 gene (also known as STK11) cause the Peutz-Jeghers Syndrome, and somatic loss of LKB1 has emerged as causal event in a wide range of human malignancies, including melanoma, lung cancer, and cervical cancer. The LKB1 protein is a serine-threonine kinase that phosphorylates AMP-activated protein kinase (AMPK) and other downstream targets. Conditional knockout studies in mouse models have consistently shown that LKB1 loss promotes a highly-metastatic phenotype in diverse tissues, and human studies have demonstrated a strong association between LKB1 inactivation and tumor recurrence. Furthermore, LKB1 deficiency confers sensitivity to distinct classes of anticancer drugs. The ability to reliably identify LKB1-deficient tumors is thus likely to have important prognostic and predictive implications. Previous research studies have employed polyclonal antibodies with limited success, and there is no widely-employed immunohistochemical assay for LKB1. Here we report an assay based on a rabbit monoclonal antibody that can reliably detect endogenous LKB1 protein (and its absence) in mouse and human formalin-fixed, paraffin-embedded tissues. LKB1 protein levels determined through this assay correlated strongly with AMPK phosphorylation both in mouse and human tumors, and with mRNA levels in human tumors. Our studies fully validate this immunohistochemical assay for LKB1 in paraffin-embedded formalin tissue sections. This assay should be broadly useful for research studies employing mouse models and also for the development of human tissue-based assays for LKB1 in diverse clinical settings

DAAM is required for thin filament formation and Sarcomerogenesis during muscle development in Drosophila.

During muscle development, myosin and actin containing filaments assemble into the highly organized sarcomeric structure critical for muscle function. Although sarcomerogenesis clearly involves the de novo formation of actin filaments, this process remained poorly understood. Here we show that mouse and Drosophila members of the DAAM formin family are sarcomere-associated actin assembly factors enriched at the Z-disc and M-band. Analysis of dDAAM mutants revealed a pivotal role in myofibrillogenesis of larval somatic muscles, indirect flight muscles and the heart. We found that loss of dDAAM function results in multiple defects in sarcomere development including thin and thick filament disorganization, Z-disc and M-band formation, and a near complete absence of the myofibrillar lattice. Collectively, our data suggest that dDAAM is required for the initial assembly of thin filaments, and subsequently it promotes filament elongation by assembling short actin polymers that anneal to the pointed end of the growing filaments, and by antagonizing the capping protein Tropomodulin

El espacio como instrumento de formación. Arquitectura como estrategia educativa aplicado a un Jardín Infantil

Artículo de GradoEl proyecto se desarrolla como ejercicio académico en un contexto real donde se propone adoptar y entender la transformación de la idea de diseño en las instalaciones educacionales en el país con base en la petición que la secretaria de educación le hizo a el Arq. Frank Locker, donde el arquitecto debe establecer criterios de diseño en búsqueda de cambiar la forma de aprender, todo esto basado en la relación entre el tipo de educación y la arquitectura como lugar de aprendizaje, enfocado en lo que la educación Colombiana busca.1. Resumen 2. Abstract 3. Contenido 4. Introducción 5. Hipótesis 6. Objetivo General 7. Objetivos Específicos 8. Marco teórico 9. Metodología 10. Resultados 11. La relación entre equipamiento y la transformación de espacio público en un entorno urbano no consolidado 12. Espacio para la enseñanza 13. Disposición de elementos estructurales para mayor eficiencia formal y sismo resistente 14. Discusión basada en comparación con particularidades similares 15. Conclusión 16. ReferenciasPregradoArquitect

Immunological assays for chemokine detection in in-vitro culture of CNS cells

Herein we review the various methods currently in use for determining the expression of chemokines by CNS cells in vitro. Chemokine detection assays are used in conjuction with one another to provide a comprehensive, biologically relevant assessment of the chemokines which is necessary for correct data interpretation of a specific observed biological effect. The methods described include bioassays for soluble chemokine receptors, RNA extraction, RT-PCR, Real - time quantitative PCR, gene array analysis, northern blot analysis, Ribonuclease Protection assay, Flow cytometry, ELISPOT, western blot analysis, and ELISA. No single method of analysis meets the criteria for a comprehensive, biologically relevant assessment of the chemokines, therefore more than one assay might be necessary for correct data interpretation, a choice that is based on development of a scientific rationale for the method with emphasis on the reliability and relevance of the method

GU81, a VEGFR2 antagonist peptoid, enhances the anti-tumor activity of doxorubicin in the murine MMTV-PyMT transgenic model of breast cancer

<p>Abstract</p> <p>Background</p> <p>Vascular endothelial growth factor (VEGF) is a primary stimulant of angiogenesis under physiological and pathological conditions. Anti-VEGF therapy is a clinically proven strategy for the treatment of a variety of cancers including colon, breast, lung, and renal cell carcinoma. Since VEGFR2 is the dominant angiogenic signaling receptor, it has become an important target in the development of novel anti-angiogenic therapies. We have reported previously the development of an antagonistic VEGFR2 peptoid (GU40C4) that has promising anti-angiogenic activity <it>in vitro </it>and <it>in vivo</it>.</p> <p>Methods</p> <p>In the current study, we utilize a derivative of GU40C4, termed GU81 in therapy studies. GU81 was tested alone or in combination with doxorubicin for <it>in vivo </it>efficacy in the MMTV-PyMT transgenic model of breast cancer.</p> <p>Results</p> <p>The derivative GU81 has increased <it>in vitro </it>efficacy compared to GU40C4. Single agent therapy (doxorubicin or GU81 alone) had no effect on tumor weight, histology, tumor fat content, or tumor growth index. However, GU81 is able to significantly to reduce total vascular area as a single agent. GU81 used in combination with doxorubicin significantly reduced tumor weight and growth index compared to all other treatment groups. Furthermore, treatment with combination therapy significantly arrested tumor progression at the premalignant stage, resulting in increased tumor fat content. Interestingly, treatment with GU81 alone increased tumor-VEGF levels and macrophage infiltration, an effect that was abrogated when used in combination with doxorubicin.</p> <p>Conclusion</p> <p>This study demonstrates the VEGFR2 antagonist peptoid, GU81, enhances the anti-tumor activity of doxorubicin in spontaneous murine MMTV-PyMT breast tumors.</p

Ten relevant questions for applying biodiversity offsetting in the Pantanal wetland.

Abstract : In Brazil, biodiversity offsetting operates in an ad hoc manner while biodiversity equivalence has mainly been overlooked by public policies. Despite being mandatory since 1965s Forest Code (Law 4.771), we fail to have a robust offsetting framework. The revision of the forest code in 2012 (i.e., Native Vegetation Protection Law—NVPL—Law n° 12,651/2012), maintained the obligation for landowners to set aside a biome-specific percentage of their native vegetation for preservation. Known as Legal Reserves, these set-asides are a precondition for compliance with NVPL's regularization procedure called CAR (Rural Environmental Cadaster). Despite enthusiasm about biodiversity offsetting opportunities following the NVPL enactment in 2012, uncertainties around its implementation remains. Here, we formulated 10 questions that discuss and illustrate how offsetting can be applied to maintain wetland integrity, economic fairness and biodiversity conservation in the Pantanal and Upper Paraguay River basin (UPRB). The aim is to stimulate robust public policies and stimulate wetland offsetting research opportunities. We provided examples of implementation opportunities of the NVPL in integrating the floodplain and highland in Pantanal at UPRB, analyze spatial compliance deficits, and illustrate opportunities that require harmonized legislation and policies between Mato Grosso and Mato Grosso do Sul states in Brazil. No Brasil, a compensação da biodiversidade tem sido implementada de forma não sistematizada, enquanto a equivalência da biodiversidade permanece amplamente negligenciada pelas políticas públicas. Embora sua obrigatoriedade esteja prevista desde o Código Florestal de 1965 (Lei nº 4.771), ainda não há um arcabouço regulatório consolidado para a compensação ambiental. A revisão do Código Florestal em 2012, por meio da Lei de Proteção da Vegetação Nativa (LPVN, Lei nº 12.651/2012), manteve a exigência de que os proprietários rurais destinem um percentual específico de suas propriedades à preservação da vegetação nativa. Essas áreas, denominadas Reservas Legais, constituem um requisito essencial para a regularização ambiental no âmbito do Cadastro Ambiental Rural (CAR). Resumo: Apesar das perspectivas promissoras geradas pela LPVN quanto à compensação da biodiversidade, sua implementação ainda enfrenta desafios e incertezas. Neste estudo, são apresentadas e discutidas dez questões fundamentais que abordam o potencial da compensação ambiental para assegurar a integridade dos ecossistemas úmidos, promover a equidade econômica e fomentar a conservação da biodiversidade no Pantanal e na Bacia do Alto Paraguai (BAP). O objetivo é contribuir para o aprimoramento das políticas públicas e incentivar pesquisas sobre compensação ambiental em ecossistemas úmidos. Adicionalmente, são fornecidos exemplos de oportunidades para a aplicação da LPVN, com foco na integração entre a planície de inundação e as áreas de maior altitude no Pantanal, além da análise de déficits espaciais de conformidade. Por fim, são discutidas oportunidades que demandam a harmonização da legislação e das políticas ambientais entre os estados de Mato Grosso e Mato Grosso do Sul, no Brasil

Locally Administrated Perindopril Improves Healing in an Ovariectomized Rat Tibial Osteotomy Model

Angiotensin-converting enzyme inhibitors are widely prescribed to regulate blood pressure. High doses of orally administered perindopril have previously been shown to improve fracture healing in a mouse femur fracture model. In this study, perindopril was administered directly to the fracture area with the goal of stimulating fracture repair. Three months after being ovariectomized (OVX), tibial fractures were produced in Sprague–Dawley rats and subsequently stabilized with intramedullary wires. Perindopril (0.4 mg/kg/day) was injected locally at the fractured site for a treatment period of 7 days. Vehicle reagent was used as a control. Callus quality was evaluated at 2 and 4 weeks post-fracture. Compared with the vehicle group, perindopril treatment significantly increased bone formation, increased biomechanical strength, and improved microstructural parameters of the callus. Newly woven bone was arranged more tightly and regularly at 4 weeks post-fracture. The ultimate load increased by 66.1 and 76.9% (p<0.01), and the bone volume over total volume (BV/TV) increased by 29.9% and 24.3% (p<0.01) at 2 and 4 weeks post-fracture, respectively. These findings suggest that local treatment with perindopril could promote fracture healing in ovariectomized rats

Eight basic principles for the elaboration of public policies and development projects for the Pantanal.

Abstract: The Pantanal is considered the largest continuous freshwater wetland in the world, and its sustainable use requires a unified conceptual framework. The lengthy process to establish public policies has contributed to the increasing vulnerability of the Pantanal. Given the need for a conceptual basis to help this process, we elaborate a list of eight basic principles based on the accumulated scientific evidence: (i) Consider the Paraguay River Basin a management unity; (ii) Establish rules that follow the concept of restricted use approach; (iii) Ensure the ecologically sustainable use of the Pantanal; (iv) Maintain the environmental heterogeneity and functionality in the Pantanal landscapes; (v) Maintain the hydrological integrity and connectivity; (vi) Ensure the environmental representativeness of the protected areas network; (vii) Provide economic incentives for conservationist use of the land; and (viii) Recognize and protect traditional people, their values, resources, and way of living. However, the elaboration of public policies should be a participatory and inclusive decision‐making process towards a more just and sustainable future.Online first

LKB1 loss promotes endometrial cancer progression via CCL2-dependent macrophage recruitment

Endometrial cancer is the most common gynecologic malignancy and the fourth most common malignancy in women. For most patients in whom the disease is confined to the uterus, treatment results in successful remission; however, there are no curative treatments for tumors that have progressed beyond the uterus. The serine/threonine kinase LKB1 has been identified as a potent suppressor of uterine cancer, but the biological modes of action of LKB1 in this context remain incompletely understood. Here, we have shown that LKB1 suppresses tumor progression by altering gene expression in the tumor microenvironment. We determined that LKB1 inactivation results in abnormal, cell-autonomous production of the inflammatory cytokine chemokine (C-C motif) ligand 2 (CCL2) within tumors, which leads to increased recruitment of macrophages with prominent tumor-promoting activities. Inactivation of Ccl2 in an Lkb1-driven mouse model of endometrial cancer slowed tumor progression and increased survival. In human primary endometrial cancers, loss of LKB1 protein was strongly associated with increased CCL2 expression by tumor cells as well as increased macrophage density in the tumor microenvironment. These data demonstrate that CCL2 is a potent effector of LKB1 loss in endometrial cancer, creating potential avenues for therapeutic opportunities