Técnicas para Detección y Estimación Eficiente de Potenciales Evocados

Los potenciales evocados (PE) en sus distintas modalidades (auditivos, visuales y somatosensoriales), se han convertido actualmente en un estudio de diagnóstico de patologías del sistema nervioso casi rutinario. Estos son señales eléctricas registradas mediante electrodos en el cuero cabelludo, que se producen como respuesta de los sistemas sensoriales a la aplicación de un estímulo adecuado. Sin embargo en su adquisición, además de la respuesta evocada por el estímulo, se registran potenciales generados por diversas fuentes fisiológicas y no fisiológicas; obteniéndose una relación señal a ruido (RSR) muy desfavorable que puede llegar a -20 dB en el caso de los potenciales evocados auditivos de tronco cerebral (PEATC).La técnica comúnmente utilizada para mejorar la RSR, y estimar la señal de PE, es la promediación coherente o sincronizada; la cual consiste en aplicar sucesivos estímulos y promediar la actividad eléctrica registrada en forma sincronizada con el instante de tiempo en que se aplica el estímulo. Si bien la promediación es de uso masivo, presenta limitaciones que justifican buscar alternativas a esta. Una es el tiempo requerido para lograr una estimación confiable del PE, que dependiendo de la modalidad del mismo puede insumir más o menos tiempo; en el caso de los PEATC puede ser de hasta 5 minutos. Otra es que se asume que la señal de PE se mantiene constante de época a época y que el ruido (el resto de los potenciales) es del tipo blanco con media cero, hipótesis que no es cierta para la mayoría de los casos. En consecuencia el resultado obtenido de la promediación es una mala estimación de la señal real de PE. Asimismo, hay situaciones en las que no se necesita estimar la morfología de la señal y solo basta con saber si la señal está presente, como por ejemplo la detección automática de hipoacusias.En este proyecto se propuso revisar, evaluar y desarrollar técnicas y/o algoritmos que permitan detectar la señal de PE así como también estimar de manera más eficiente que la promediación coherent

Modelling human choices: MADeM and decision‑making

Research supported by FAPESP 2015/50122-0 and DFG-GRTK 1740/2. RP and AR are also part of the Research, Innovation and Dissemination Center for Neuromathematics FAPESP grant (2013/07699-0). RP is supported by a FAPESP scholarship (2013/25667-8). ACR is partially supported by a CNPq fellowship (grant 306251/2014-0)

MDMX stability is regulated by p53-induced caspase cleavage in NIH3T3 mouse fibroblasts

MDMX is a p53 binding protein, which shares a high degree of homology with MDM2, a negative regulator of the tumor suppressor p53. MDMX has been shown to counteract MDM2-dependent p53 degradation and to stabilize p53 in its inactive form. In this study: we identify two MDMX proteolytic pathways that control its intracellular levels, and show that MDMX post-translational processing may be regulated by p53. Mouse MDMX is cleaved in vitro and in vivo by caspase activity, between aminoacids 358 and 361, producing a p54 minor form. In addition, MDMX is subjected to proteasome-mediated degradation, which concurs to MDMX proteolysis mainly through degradation of p54. A D361A-MDMX mutant, resistant to caspase cleavage, exhibits prolonged intracellular lifetime in comparison to wild-type protein, indicating that caspase cleavage affects stability of MDMX protein probably by modulating its further degradation. Overexpression of exogenous p53 increases the intracellular levels of p54 product. Similarly, activation of endogenous p53 by adriamycin enhances MDMX cleavage and produces a marked decrease of its intracellular levels, while not affecting the D361A-MDMX mutant. In addition, the D361A-MDMX mutant lacks the ability to inhibit p53 transactivation in respect to wild-type MDMX, suggesting that MDMX caspase cleavage play an important functional role. In conclusion, our results demonstrate that, in analogy to MDM2, MDMX may be subjected to proteolytic modifications that regulate its intracellular levels. Moreover, decrease of MDMX protein levels following p53 activation suggests a p53-dependent regulatory feedback of MDMX function

Identification of an aberrantly spliced form of HDMX in human tumors. a new mechanism for HDM2 stabilization

The HDMX protein is closely related to HDM2 with which it shares different structural domains, particularly the p53 binding domain and the ring finger domain, where the two HDM proteins interact. Several oncogenic forms derived from splicing of HDM2 have been described in cancer. This work aimed at investigating whether analogous forms of HDMX exist in human tumors. Here, we report the characterization of an aberrantly spliced form of HDMX, HDMX211, isolated from the thyroid tumor cell line, ARO. HDMX211 binds and stabilizes the HDM2 protein. Although it lacks the p53 binding domain, HDMX211 also stabilizes p53 by counteracting its degradation by HDM2. However, the resulting p53 is transcriptionally inactive and increasingly associated to its inhibitor HDM2. Expression of HDMX211 strongly enhances the colony-forming ability of human cells in the presence or absence of wild-type p53. Conversely, depletion of HDMX211 by small interfering RNA significantly reduces the growth of ARO cells and increases their sensitivity to chemotherapy. Screening of lung cancer biopsies shows the presence of HDMX211 in samples that overexpress HDM2 protein, supporting a pathologic role for this new protein. This is the first evidence of a variant form of HDMX that has oncogenic potential independently of p53. HDMX211 reveals a new mechanism for overexpression of the oncoprotein HDM2. Most interestingly, it outlines a possible molecular explanation for a yet unclarified tumor phenotype, characterized by simultaneous overexpression of HDM2 and wild-type p53

MDM4 (MDMX) overexpression enhances stabilization of stress-induced p53 and promotes apoptosis.

Rescue of embryonic lethality in MDM4-/- mice through concomitant loss of p53 has revealed a functional partnership between the two proteins. Biochemical studies have suggested that MDM4 may act as a negative regulator of p53 levels and activity. On the other hand, MDM4 overexpression has been reported to stabilize p53 levels and to counteract MDM2-degradative activity. We have investigated the functional role of MDM4 overexpression on cell behaviour. In both established and primary cells cultured under stress conditions, overexpression of MDM4 significantly increased p53-dependent cell death, in correlation with enhanced induction of the endogenous p53 protein levels. This phenomenon was associated with induced p53 transcriptional activity and increased levels of the pro-apoptotic protein, Bax. Further, p53 stabilization was accompanied by decreased association of the protein to its negative regulator, MDM2. These findings reveal a novel role for MDM4 by demonstrating that in non-tumor cells under stress conditions it may act as a positive regulator of p53 activity, by mainly controlling p53 levels. They also indicate a major distinction between the biological consequences of MDM4 and MDM2 overexpressio

Identification of an aberrantly spliced form of HDMX in human tumors: a new mechanism for HDM2 stabilization

The HDMX protein is closely related to HDM2 with which it shares different structural domains, particularly the p53 binding domain and the ring finger domain, where the two HDM proteins interact. Several oncogenic forms derived from splicing of HDM2 have been described in cancer. This work aimed at investigating whether analogous forms of HDMX exist in human tumors. Here, we report the characterization of an aberrantly spliced form of HDMX, HDMX211, isolated from the thyroid tumor cell line, ARO. HDMX211 binds and stabilizes the HDM2 protein. Although it lacks the p53 binding domain, HDMX211 also stabilizes p53 by counteracting its degradation by HDM2. However, the resulting p53 is transcriptionally inactive and increasingly associated to its inhibitor HDM2. Expression of HDMX211 strongly enhances the colony-forming ability of human cells in the presence or absence of wild-type p53. Conversely, depletion of HDMX211 by small interfering RNA significantly reduces the growth of ARO cells and increases their sensitivity to chemotherapy. Screening of lung cancer biopsies shows the presence of HDMX211 in samples that overexpress HDM2 protein, supporting a pathologic role for this new protein. This is the first evidence of a variant form of HDMX that has oncogenic potential independently of p53. HDMX211 reveals a new mechanism for overexpression of the oncoprotein HDM2. Most interestingly, it outlines a possible molecular explanation for a yet unclarified tumor phenotype, characterized by simultaneous overexpression of HDM2 and wild-type p53