Early expression of mature αß TCR in CD4-CD8- T cell progenitors enables MHC to drive development of T-ALL bearing NOTCH mutations.

During normal T cell development in mouse and human, a low-frequency population of immature CD4-CD8- double-negative (DN) thymocytes expresses early, mature αß T cell antigen receptor (TCR). We report that these early αß TCR+ DN (EADN) cells are DN3b-DN4 stage and require CD3δ but not major histocompatibility complex (MHC) for their generation/detection. When MHC - is present, however, EADN cells can respond to it, displaying a degree of coreceptor-independent MHC reactivity not typical of mature, conventional αß T cells. We found these data to be connected with observations that EADN cells were susceptible to T cell acute lymphoblastic leukemia (T-ALL) transformation in both humans and mice. Using the OT-1 TCR transgenic system to model EADN-stage αß TCR expression, we found that EADN leukemogenesis required MHC to induce development of T-ALL bearing NOTCH1 mutations. This leukemia-driving MHC requirement could be lost, however, upon passaging the tumors in vivo, even when matching MHC was continuously present in recipient animals and on the tumor cells themselves. These data demonstrate that MHC:TCR signaling can be required to initiate a cancer phenotype from an understudied developmental state that appears to be represented in the mouse and human disease spectrum.

Properties of the PVA-VAVTD KOH Blend as a Gel Polymer Electrolyte for Zinc Batteries.

Rechargeable zinc-air batteries are promising for energy storage and portable electronic applications because of their good safety, high energy density, material abundance, low cost, and environmental friendliness. A series of alkaline gel polymer electrolytes formed from polyvinyl alcohol (PVA) and different amounts of terpolymer composed of butyl acrylate, vinyl acetate, and vinyl neodecanoate (VAVTD) was synthesized applying a solution casting technique. The thin films were doped with KOH 12M, providing a higher amount of water and free ions inside the electrolyte matrix. The inclusion of VAVTD together with the PVA polymer improved several of the electrical properties of the PVA-based gel polymer electrolytes (GPEs). X-ray diffraction (XRD), thermogravimetric analysis (TGA), and attenuated total reflectance- Fourier-transform infrared spectroscopy (ATR-FTIR) tests, confirming that PVA chains rearrange depending on the VAVTD content and improving the amorphous region. The most conducting electrolyte film was the test specimen 1:4 (PVA-VAVTD) soaked in KOH solution, reaching a conductivity of 0.019 S/cm at room temperature. The temperature dependence of the conductivity agrees with the Arrhenius equation and activation energy of ~0.077 eV resulted, depending on the electrolyte composition. In addition, the cyclic voltammetry study showed a current intensity increase at higher VAVTD content, reaching values of 310 mA. Finally, these gel polymer electrolytes were tested in Zn-air batteries, obtaining capacities of 165 mAh and 195 mAh for PVA-T4 and PVA-T5 sunk in KOH, respectively, at a discharge current of -5 mA.

Intervención educativa sobre consecuencias del tabaquismo en adolescentes de Nuevitas / Educational intervention on the smoking consequences in adolescents from Nuevitas

RESUMEN Fundamento: el tabaquismo es un fenómeno social que afecta indistintamente a todos los sectores de la población y representa uno de los mayores problemas para las autoridades sanitarias. Se considera una epidemia de carácter universal y es una gravosa carga para el individuo, la familia y la sociedad. Objetivo: implementar una intervención educativa para modificar los conocimientos de adolescentes acerca de las consecuencias del tabaquismo. Métodos: se realizó un estudio de intervención en el Consultorio 21 del Policlínico Francisco Peña Peña, de Nuevitas, sobre 81 adolescentes que cumplieron con los criterios de inclusión. Se utilizó la estadística descriptiva para analizar los resultados obtenidos al aplicar la encuesta antes y después de la intervención educativa, la cual contenía las siguientes variables: grupo de edades, sexo, nivel de escolaridad, enfermedades asociadas y conocimiento sobre tabaquismo. Resultados: predominó el sexo masculino y las edades de 18 a 20 años, grupo etario que también mostró el mayor conocimiento sobre los efectos del tabaquismo. Prevaleció el nivel de escolaridad de preuniversitario. El nivel de conocimientos evaluado de bien ascendió de 49,38 % antes de la intervención a 80 % después de esta; el de regular descendió de 45,67 % a solo 19 %; al terminar la intervención ningún adolescente se ubicó en la categoría de mal. Conclusiones: se logró modificar el nivel de los conocimientos sobre los efectos nocivos del tabaquismo en adolescentes, mediante la intervención educativa.

ABSTRACT Background: smoking is a social phenomenon that affects all the population sectors indistinctly and represents one of the biggest problems for health authorities. It is considered a universal epidemic and is a heavy burden for the individual, the family and society. Objective: to carry out educational intervention to modify the adolescents' knowledge about the smoking consequences. Methods: an intervention study was carried out in the doctor´s office 21 of the Francisco Peña Peña Polyclinic, in Nuevitas, on 81 adolescents who met the inclusion criteria. Descriptive statistic was used to analyze the results obtained by applying the survey before and after the educational intervention, which contained the following variables: age group, sex, educational level, associated diseases, and knowledge about smoking. Results: male sex and ages 18 to 19 predominated, an age group that also showed the greatest knowledge about the effects of smoking. The pre-university educational level prevailed. The level of knowledge evaluated as good rose from the 49.38% before the intervention to the 80% after it; that of regular fell from the 45.67% to only the 19%; At the end of the intervention, no adolescent was placed in the category of bad. Conclusions: it was possible to modify the knowledge level about the harmful effects of smoking in adolescents, through educational intervention.

Instability of Helios-deficient Tregs is associated with conversion to a T-effector phenotype and enhanced antitumor immunity.

Expression of the transcription factor Helios by Tregs ensures stable expression of a suppressive and anergic phenotype in the face of intense inflammatory responses, whereas Helios-deficient Tregs display diminished lineage stability, reduced FoxP3 expression, and production of proinflammatory cytokines. Here we report that selective Helios deficiency within CD4 Tregs leads to enhanced antitumor immunity through induction of an unstable phenotype and conversion of intratumoral Tregs into T effector cells within the tumor microenvironment. Induction of an unstable Treg phenotype is associated with enhanced production of proinflammatory cytokines by tumor-infiltrating but not systemic Tregs and significantly delayed tumor growth. Ab-dependent engagement of Treg surface receptors that result in Helios down-regulation also promotes conversion of intratumoral but not systemic Tregs into T effector cells and leads to enhanced antitumor immunity. These findings suggest that selective instability and conversion of intratumoral CD4 Tregs through genetic or Ab-based targeting of Helios may represent an effective approach to immunotherapy.

Consideraciones teóricas elementales del método clínico / Basic theoretical considerations about the clinical method

El método clínico surge desde las relaciones humanas más primitivas y aún cuando se encuentra en constante evolución no se consigue su adecuada aplicación. El estudio de este método científico constituye uno de los temas más importantes que guían la praxis médica, ostenta ciertos caracteres que se entrelazan dialécticamente imposibles de desligar, centrados esencialmente en la relación médico-paciente con el objetivo de arribar al diagnóstico de una enfermedad determinada, por lo cual, se brindan algunas consideraciones teóricas a partir del análisis de sus conceptualizaciones y atributos más significativos y se elabora una definición de método clínico (AU)

The clinical method faces constant evolution, in spite of the issues that affect its application. The study of this scientific method constitutes one of the most important topics that lead medical praxis. It entails several characteristics dialectically intertwined that focus mainly on the doctor-patient relation for the diagnosis of a given disease. The objective of this article is to provide some theoretical considerations based on the analysis of its main concepts and attributes (AU)

Consideraciones teóricas elementales del método clínico / Basic theoretical considerations about the clinical method

El método clínico surge desde las relaciones humanas más primitivas y aún cuando se encuentra en constante evolución no se consigue su adecuada aplicación. El estudio de este método científico constituye uno de los temas más importantes que guían la praxis médica, ostenta ciertos caracteres que se entrelazan dialécticamente imposibles de desligar, centrados esencialmente en la relación médico-paciente con el objetivo de arribar al diagnóstico de una enfermedad determinada, por lo cual, se brindan algunas consideraciones teóricas a partir del análisis de sus conceptualizaciones y atributos más significativos y se elabora una definición de método clínico.

The clinical method faces constant evolution, in spite of the issues that affect its application. The study of this scientific method constitutes one of the most important topics that lead medical praxis. It entails several characteristics dialectically intertwined that focus mainly on the doctor-patient relation for the diagnosis of a given disease. The objective of this article is to provide some theoretical considerations based on the analysis of its main concepts and attributes.

Co-potentiation of antigen recognition: A mechanism to boost weak T cell responses and provide immunotherapy in vivo.

Adaptive immunity is mediated by antigen receptors that can induce weak or strong immune responses depending on the nature of the antigen that is bound. In T lymphocytes, antigen recognition triggers signal transduction by clustering T cell receptor (TCR)/CD3 multiprotein complexes. In addition, it hypothesized that biophysical changes induced in TCR/CD3 that accompany receptor engagement may contribute to signal intensity. Nonclustering monovalent TCR/CD3 engagement is functionally inert despite the fact that it may induce changes in conformational arrangement or in the flexibility of receptor subunits. We report that the intrinsically inert monovalent engagement of TCR/CD3 can specifically enhance physiologic T cell responses to weak antigens in vitro and in vivo without stimulating antigen-unengaged T cells and without interrupting T cell responses to strong antigens, an effect that we term as "co-potentiation." We identified Mono-7D6-Fab, which biophysically altered TCR/CD3 when bound and functionally enhanced immune reactivity to several weak antigens in vitro, including a gp100-derived peptide associated with melanoma. In vivo, Mono-7D6-Fab induced T cell antigen-dependent therapeutic responses against melanoma lung metastases, an effect that synergized with other anti-melanoma immunotherapies to significantly improve outcome and survival. We conclude that Mono-7D6-Fab directly co-potentiated TCR/CD3 engagement by weak antigens and that such concept can be translated into an immunotherapeutic design. The co-potentiation principle may be applicable to other receptors that could be regulated by otherwise inert compounds whose latent potency is only invoked in concert with specific physiologic ligands.

Molecular analysis of CD133-positive circulating tumor cells from patients with metastatic castration-resistant prostate cancer.

The function and clinical utility of stem cell markers in metastatic castration-resistant prostate cancer (mCRPC) remains unresolved, and their expression may confer important therapeutic opportunities for staging and therapy. In the adult human prostate, CD133 (PROM1) expression identifies infrequent prostate epithelial progenitor cells and putative cancer stem cells. Previous work demonstrated an association with CD133 and cancer cell proliferation using in vitro model systems. The primary objective here was to investigate the expression of CD133 in circulating tumor cells (CTCs) from patients with mCRPC and to test the hypothesis that patients with mCRPC had CD133-positive CTCs associated with increased cell proliferation, changes in the androgen receptor (AR) protein expression, or AR nuclear co-localization. We utilized ImageStreamX technology, which combines flow cytometry and fluorescence microscopy, to capture and analyze CD45-negative/EpCAM-positive CTCs for CD133, Ki-67, and AR. All patient samples (20/20) contained CD133-positive populations of CTCs, and on average 50.9 ± 28.2% (range of 18.2% to 100%) of CTCs were CD133-positive. CD133-positive CTCs have increased Ki-67 protein expression compared to CD133-negative CTCs, implying that CD133-positive CTCs may have greater proliferative potential when compared to their CD133-negative counterparts. CD133-positive and CD133-negative CTCs have similar levels of AR protein expression and cellular co-localization with nuclear markers, implying that CD133 expression is independent of AR pathway activity and an AR-independent marker of mCRPC proliferation. These studies demonstrate the presence of CD133-positive populations in CTCs from mCRPC with increased proliferative potential.

Quantitative characterization of androgen receptor protein expression and cellular localization in circulating tumor cells from patients with metastatic castration-resistant prostate cancer.

BACKGROUND: Many current therapies for metastatic castration-resistant prostate cancer (mCRPC) are aimed at AR signaling; however, resistance to these therapies is inevitable. To personalize CRPC therapy in an individual with clinical progression despite maximal AR signaling blockade, it is important to characterize the status of AR activity within their cancer. Biopsies of bone metastases are invasive and frequently fail to yield sufficient tissue for further study. Evaluation of circulating tumor cells (CTCs) offers an alternative, minimally invasive mechanism to characterize and study late-stage disease. The goal of this study was to evaluate the utility of CTC interrogation with respect to the AR as a potential novel therapeutic biomarker in patients with mCRPC. METHODS: Fifteen mL of whole blood was collected from patients with progressive, metastatic mCRPC, the mononuclear cell portion was isolated, and fluorescence-activated cell sorting (FACS) was used to isolate and evaluate CTCs. A novel protocol was optimized to use ImageStreamX to quantitatively analyze AR expression and subcellular localization within CTCs. Co-expression of AR and the proliferation marker Ki67 was also determined using ImageStreamX. RESULTS: We found inter-patient and intra-patient heterogeneity in expression and localization of AR. Increased AR expression and nuclear localization are associated with elevated co-expression of Ki-67, consistent with the continued role for AR in castration-resistant disease. Despite intra-patient heterogeneity, CTCs from patients with prior exposure to abiraterone had increased AR expression compared to CTCs from patients who were abiraterone-naïve. CONCLUSIONS: As our toolbox for targeting AR function expands, our ability to evaluate AR expression and function within tumor samples from patients with late-stage disease will likely be a critical component of the personalized management of advanced prostate cancer. AR expression and nuclear localization varies within patients and between patients; however it remains associated with markers of proliferation. This supports a molecularly diverse AR-centric pathobiology imparting castration-resistance.

Lesión cerebral posterior a paro cardiorrespiratorio / Brain injury after cardiac arrest

La lesión cerebral postparo cardíaco es causa común de morbilidad y mortalidad, de ahí la importancia de comprender sus mecanismos fisiopatológicos, su relación con el impacto de las técnicas de resucitación y el pronóstico de los pacientes. La pérdida de la integridad celular dispara la liberación de glutamato generando excitotoxicidad asociada a la disminución de los neurotransmisores moduladores y a elevación del calcio intracelular provocando lesiones en diferentes áreas cerebrales causando síntomas cognitivos con un impacto significativo en la calidad de vida de estas personas. La Categoría de Desempeño Cerebral es el patón de oro para la evaluación de la recuperación neurológica después del paro cardiorrespiratorio (PCR) y el examen neurológico continúa siendo uno de los factores más confiables para determinar la extensión de la lesión y el pronóstico neurológico en las víctimas de PCR. La evidencia demuestra que la hipotermia leve inducida puede disminuir la lesión cerebral, mejorando la supervivencia y el resultado neurológico funcional en pacientes comatosos sobrevivientes de PCR. La lesión cerebral posterior al PCR es sin duda un área importante para la investigación clínica.

Brain injury after cardiac arrest is a common cause of morbidity and mortality, hence the importance of understanding its pathophysiological mechanisms and its relation to the impact of resuscitation and prognosis of patients. The loss of cellular integrity triggers the release of glutamate thus generating excitotoxicity associated with the decreased of modulatory neurotransmitters levels and the elevation of intracellular calcium thense causing lesions in different brain areas and generating cognitive symptoms related to significant impact on quality of life. The Cerebral Performance Category is the gold standard for the assessment of neurological recovery after cardiopulmonary arrest (CPA) and neurological examination remains one of the most reliable factors to determine the extent of the injury and neurological outcome in CPA victims. Evidence shows that mild induced hypothermia may reduce injury, improving survival and functional neurologicals outcomes in comatose survivors of CPA. Brain injury after CPA is an important area for research.

Sox2 is an androgen receptor-repressed gene that promotes castration-resistant prostate cancer.

Despite advances in detection and therapy, castration-resistant prostate cancer continues to be a major clinical problem. The aberrant activity of stem cell pathways, and their regulation by the Androgen Receptor (AR), has the potential to provide insight into novel mechanisms and pathways to prevent and treat advanced, castrate-resistant prostate cancers. To this end, we investigated the role of the embryonic stem cell regulator Sox2 [SRY (sex determining region Y)-box 2] in normal and malignant prostate epithelial cells. In the normal prostate, Sox2 is expressed in a portion of basal epithelial cells. Prostate tumors were either Sox2-positive or Sox2-negative, with the percentage of Sox2-positive tumors increasing with Gleason Score and metastases. In the castration-resistant prostate cancer cell line CWR-R1, endogenous expression of Sox2 was repressed by AR signaling, and AR chromatin-IP shows that AR binds the enhancer element within the Sox2 promoter. Likewise, in normal prostate epithelial cells and human embryonic stem cells, increased AR signaling also decreases Sox2 expression. Resistance to the anti-androgen MDV3100 results in a marked increase in Sox2 expression within three prostate cancer cell lines, and in the castration-sensitive LAPC-4 prostate cancer cell line ectopic expression of Sox2 was sufficient to promote castration-resistant tumor formation. Loss of Sox2 expression in the castration-resistant CWR-R1 prostate cancer cell line inhibited cell growth. Up-regulation of Sox2 was not associated with increased CD133 expression but was associated with increased FGF5 (Fibroblast Growth Factor 5) expression. These data propose a model of elevated Sox2 expression due to loss of AR-mediated repression during castration, and consequent castration-resistance via mechanisms not involving induction of canonical embryonic stem cell pathways.

Growth kinetics of CD133-positive prostate cancer cells.

BACKGROUND: In the adult human prostate CD133 expression is thought to mark rare prostate epithelial stem cells and malignant tumor stem/initiating cells. Such putative stem cell populations are thought to proliferate slowly, but possess unlimited proliferative potential. Based on this, we hypothesized that CD133(pos) prostate cancer cells proliferate slower than CD133(neg) cells. METHODS: Human prostate cancer cell lines were analyzed for CD133 expression and DNA content using flow cytometry. Rates of cell division and DNA synthesis were determined using CFSE cell tracing and BrdU uptake, respectively. Changes in cell cycle distribution and the percentage of CD133(pos) cells were assayed under conditions of different cell density and AR-pathway modulation. Lastly, we over-expressed lentiviral CD133 to measure whether CD133 regulates the cell cycle. RESULTS: The cell cycle distribution differs between CD133(pos) and CD133(neg) cells in all three human prostate cancer cell lines studied. CD133(pos) cells have a greater proportion of cells in G2 and proliferate faster than CD133(neg) cells. High cell density increases the percentage of CD133(pos) cells without changing CD133(pos) cell cycle progression. Treatment with the AR agonist R1881, or the anti-androgen MDV3100, significantly changed the percentage and proliferation of CD133(pos) cells. Finally, ectopic over-expression of CD133 had no effect on cell cycle progression. CONCLUSIONS: Contrary to our hypothesis, we demonstrate that CD133(pos) cells proliferate faster than CD133(neg) cells. This association of CD133 expression with increased cell proliferation is not directly mediated by CD133, suggesting that surface CD133 is a downstream target gene of an undefined pathway controlling cell proliferation.