Neutron Autoradiography Combined With UV-C Sensitization: Toward the Intracellular Localization of Boron

Our group has reported the imprint formation of biological material on polycarbonate nuclear track detectors by UV-C exposure, which is used as an approach to simultaneously visualize cell imprints and nuclear tracks coming from the boron neutron capture reaction. Considering that the cell nucleus has a higher UV-C absorption than the cytoplasm and that hematoxylin preferentially stains the nucleus, we proposed to enhance the contrast between these two main cell structures by hematoxylin staining before UV-C sensitization. In this study, several experiments were performed in order to optimize UV-C exposure parameters and chemical etching conditions for cell imprint formation using the SK-BR-3 breast cancer cell line. The proposed method improves significantly the resolution of the cell imprints. It allows clear differentiation of the nucleus from the rest of the cell, together with nuclear tracks pits. Moreover, it reduces considerably the UV-C exposure time, an important experimental issue. The proposed methodology can be applied to study the boron distribution independently from the chosen cell line and/or boron compounds.Fil: Gadan, Mario Alberto. Comisión Nacional de Energía Atómica; ArgentinaFil: Lloyd, Rodrigo. Ministerio de Ciencia. Tecnología e Innovación Productiva. Agencia Nacional de Promoción Científica y Tecnológica; Argentina. Comisión Nacional de Energía Atómica; ArgentinaFil: Saint Martin, María Laura Gisela. Comisión Nacional de Energía Atómica; ArgentinaFil: Olivera, María S.. Comisión Nacional de Energía Atómica; ArgentinaFil: Policastro, Lucia Laura. Comisión Nacional de Energía Atómica; Argentina. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Ciudad Universitaria. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia - Nodo Constituyentes | Comision Nacional de Energia Atomica. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia - Nodo Constituyentes.; ArgentinaFil: Portu, Agustina Mariana. Comisión Nacional de Energía Atómica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Silencing peroxiredoxin-2 sensitizes human colorectal cancer cells to ionizing radiation and oxaliplatin

Colorectal cancer (CRC) remains one of the leading causes of cancer-related death worldwide. Antioxidant enzymes decrease the generation of ionizing radiation (IR)-induced free radicals and therefore are associate to radioresistance. The main goal of this work is to study the involvement of peroxiredoxin-2 (Prx2) in the radio and chemoradiotherapy response in CRC cells in vitro and in vivo. We found that Prx2 oxidation state is associated to differential response to ionizing radiation in CRC cell lines. HCT116 radioresistant CRC cell line have lower ROS levels and a higher monomer/dimer Prx2 ratio, compared to halfway resistant Caco-2 and T84, and radiosensitive LoVo cell line. Constitutive and transient Prx2 silencing in CRC cells increase ROS levels, and most importantly, enhance in vitro radiation sensitivity. In addition, we showed that administration of IR plus oxaliplatin in down regulated Prx2 HCT116 cells has higher citotoxic effect than in control cells. Finally, radiosensitizing effect of Prx2 depletion was confirmed in vivo. These results suggest that Prx2 is an important component in tumoral radiation response, and their inhibition could improve radio and chemoradiotherapy protocols in patients with CRC.Fil: Cerda, Maria Belen. Comisión Nacional de Energía Atómica; ArgentinaFil: Lloyd, Rodrigo. Comisión Nacional de Energía Atómica; ArgentinaFil: Batalla, Milena. Comisión Nacional de Energía Atómica; ArgentinaFil: Giannoni, Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Comisión Nacional de Energía Atómica; ArgentinaFil: Casalia, Mariana Laura. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Policastro, Lucia Laura. Comisión Nacional de Energía Atómica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

The tumor microenvironment: characterization, redox considerations and novel approaches for ROS-targeted gene therapy.

The tumor microenvironment is a complex system that involves the interaction between malignant and neighbor stromal cells embedded in a mesh of extracellular matrix (ECM) components. Stromal cells (fibroblasts, endothelial, and inflammatory cells) are co-opted at different stages to help malignant cells invade the surrounding ECM and disseminate. Malignant cells have developed adaptive mechanisms to survive under the extreme conditions of the tumor microenvironment such as restricted oxygen supply (hypoxia), nutrient deprivation, and a prooxidant state among others. These conditions could be eventually used to target drugs that will be activated specifically in this microenvironment. Preclinical studies have shown that modulating cellular/tissue redox state by different gene therapy (GT) approaches was able to control tumor growth. In this review, we describe the most relevant features of the tumor microenvironment, addressing reactive oxygen species-generating sources that promote a prooxidative microenvironment inside the tumor mass. We describe different GT approaches that promote either a decreased or exacerbated prooxidative microenvironment, and those that make use of the differential levels of ROS between cancer and normal cells to achieve tumor growth inhibition.Fil: Policastro, Lucia Laura. Comisión Nacional de Energía Atómica. Gerencia de Área de Investigación y Aplicaciones no Nucleares. Gerencia de Desarrollo Tecnológico y Proyectos Especiales. Departamento de Micro y Nanotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ibañez, Irene Laura. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Notcovich, Cintia Karina. Comisión Nacional de Energía Atómica. Gerencia de Área de Investigación y Aplicaciones no Nucleares. Gerencia de Desarrollo Tecnológico y Proyectos Especiales. Departamento de Micro y Nanotecnología; ArgentinaFil: Duran, Hebe Alicia. Comisión Nacional de Energía Atómica. Gerencia de Área de Investigación y Aplicaciones no Nucleares. Gerencia de Desarrollo Tecnológico y Proyectos Especiales. Departamento de Micro y Nanotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología; ArgentinaFil: Podhajcer, Osvaldo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Instituto Leloir; Argentin

Application of BNCT to the treatment of HER2+ breast cancer recurrences: Research and developments in Argentina

In the frame of the Argentine BNCT Project a new research line has been started to study the application of BNCT to the treatment of locoregional recurrences of HER2+ breast cancer subtype. Based on former studies, the strategy considers the use of immunoliposomes as boron carriers nanovehicles to target HER2 overexpressing cells. The essential concerns of the current stage of this proposal are the development of carriers that can improve the efficiency of delivery of boron compounds and the dosimetric assessment of treatment feasibility. For this purpose, an specific pool of clinical cases that can benefit from this application was determined. In this work, we present the proposal and the advances related to the different stages of current research.Fil: Gadan, M.A.. Comisión Nacional de Energía Atómica; ArgentinaFil: González, Sara Josefina. Comisión Nacional de Energía Atómica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Batalla, M.. Comisión Nacional de Energía Atómica; ArgentinaFil: Olivera, M.S.. Comisión Nacional de Energía Atómica; ArgentinaFil: Policastro, Lucia Laura. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Comisión Nacional de Energía Atómica; ArgentinaFil: Sztejnberg, M.L.. Comisión Nacional de Energía Atómica; Argentin

H2O2 scavenging inhibits G1/S transition by increasing nuclear levels of p27KIP1

The aim of the present study was to evaluate cell cycle regulation by scavenging H(2)O(2) in tumor cells. A significant arrest in the G1 phase of the cell cycle was demonstrated in CH72-T4 carcinoma cells exposed to catalase, associated with a decrease in cyclin D1 and an increase in the CDK inhibitory protein p27(KIP1). Moreover, we found a differential intracellular distribution of p27(KIP1), which remained in the nucleus after catalase treatment. In vivo experiments showed an increase in nuclear levels of p27(KIP1) associated with the inhibition of tumor growth by H(2)O(2) scavenging, confirming in vitro results. To conclude, H(2)O(2) scavenging may induce cell cycle arrest through the modulation of cyclin D1 and p27(KIP1) levels and nuclear localization of p27(KIP1). To our knowledge, this is the first report that demonstrates that the modulation of ROS alters the intracellular localization of a key regulatory protein of G1/S transition.Fil: Ibañez, Irene Laura. Comision Nacional de Energia Atomica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Policastro, Lucia Laura. Comision Nacional de Energia Atomica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Tropper, Ivanna. Universidad Nacional de San Martín; ArgentinaFil: Bracalente, María Candelaria. Comision Nacional de Energia Atomica; ArgentinaFil: Palmieri, Mónica A.. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Rojas, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentina; ArgentinaFil: Molinari, Beatriz Lucia. Comision Nacional de Energia Atomica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Duran, Hebe Alicia. Comisión Nacional de Energía Atómica. Gerencia del Area de Investigación y Aplicaciones No Nucleares. Gerencia de Desarrollo Tecnológico y Proyectos Especiales. Departamento de Micro y Nanotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Enhancement of nucleic acid delivery to hard-to-transfect human colorectal cancer cells by magnetofection at laminin coated substrates and promotion of the endosomal/lysosomal escape

Despite a great diversity of nanomaterials, such as cationic lipid, polymers or inorganic nanoparticles, has been developed in order to carry nucleic acids across plasma membrane, these methodologies have still insufficient efficacy in cells named hard-to-transfect cells, as the colorectal HT29 and Caco-2 cell lines. This paper describes the improvement of plasmid DNA (pDNA) and small interfering RNA (siRNA) transfer in these cells through the combination of magnetofection, a simplified extracellular matrix of laminin and endosomal/lysosomal escape promotion using the endosome-disruptive peptide INF-7. Magnetofection of pDNA complexes using selected vector formulations resulted in up to 2-fold enhancement in luciferase expression, as compared to lipofection. Further enhance in pDNA transfer in HT29 cells was obtained when magnetofection was applied on cells grown on laminin coated substrates, increasing 6-fold the luciferase expression compared to lipofection at uncoated substrates. This technique was also applied to siRNA delivery in cells expressing stably luciferase (Caco-2Luc and HT29Luc), selected magnetic vector formulations resulted in 61±5% and 50±5% of luciferase silencing in HT29Luc and Caco-2Luc, respectively. Further improvement in reporter gene silencing was obtained when the magnetic complexes were modified with INF-7, reaching more than 95% of luciferase silencing in Caco-2Luc cells, while pre-treatment of HT29Luc cells by chloroquine resulted in 80±4% of down regulation of luciferase expression. Thus, magnetofection applied on cells grown over laminin coated substrates and the optimization of endosomal escape of magnetic complexes would be a good alternative to enhance nucleic acid transfer in hard-to-transfect colorectal cancer cells.Fil: Cerda, Maria Belen. Comision Nacional de Energia Atomica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Nanociencias y Nanotecnología; ArgentinaFil: Batalla, Milena. Comision Nacional de Energia Atomica; Argentina. Instituto de Nanociencias y Nanotecnología; ArgentinaFil: Anton, Martina. Technische Universität München; AlemaniaFil: Cafferata, Eduardo Gustavo Alfredo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Podhajcer, Osvaldo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Plank, Christian. Technische Universität München; AlemaniaFil: Mykhaylyk, Olga. Technische Universität München; AlemaniaFil: Policastro, Lucia Laura. Comision Nacional de Energia Atomica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Nanociencias y Nanotecnología; Argentin

Continuous flow generation of magnetoliposomes in a low-cost portable microfluidic platform

We present a low-cost, portable microfluidic platform that uses laminated polymethylmethacrylate chips, peristaltic micropumps and LEGO® Mindstorms components for the generation of magnetoliposomes that does not require extrusion steps. Mixtures of lipids reconstituted in ethanol and an aqueous phase were injected independently in order to generate a combination of laminar flows in such a way that we could effectively achieve four hydrodynamic focused nanovesicle generation streams. Monodisperse magnetoliposomes with characteristics comparable to those obtained by traditional methods have been obtained. The magnetoliposomes are responsive to external magnetic field gradients, a result that suggests that the nanovesicles can be used in research and applications in nanomedicine.Fil: Conde, Alvaro José. Universidad Nacional de Tucuman. Facultad de Cs.exactas y Tecnologia. Departamento de Bioingenieria. Laboratorio de Medios E Interfases; ArgentinaFil: Battalla, Milena. Comision Nacional de Energia Atomica. Gerencia Quimica. CAC; ArgentinaFil: Cerda, Maria Belen. Comision Nacional de Energia Atomica. Gerencia Quimica. CAC; ArgentinaFil: Mykhaylyk, Olga. Technische Universität München. Institute of Experimental Oncology and Therapy Research; AlemaniaFil: Plank, Christian. Technische Universität München. Institute of Experimental Oncology and Therapy Research; AlemaniaFil: Podhajcer, Osvaldo Luis. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; ArgentinaFil: Cabaleiro, Juan Martin. Universidad de Buenos Aires. Facultad de Ingenieria. Departamento de Ingenieria Mecanica. Laboratorio de Fluidodinamica; ArgentinaFil: Madrid, Rossana Elena. Universidad Nacional de Tucuman. Facultad de Cs.exactas y Tecnologia. Departamento de Bioingenieria. Laboratorio de Medios E Interfases; ArgentinaFil: Policastro, Lucia Laura. Comision Nacional de Energia Atomica. Gerencia Quimica. CAC; Argentin