Apoptosis is increased and cell proliferation is decreased in out-of-phase endometria from infertile and recurrent abortion patients

Various endometrial abnormalities have been associated with luteal phase deficiency: a significant dyssynchrony in the maturation of the glandular epithelium and the stroma and a prevalence of out-of-phase endometrial biopsy specimens. Out-of phase endometrium is a controversial disorder related to failed implantation, infertility and early pregnancy loss. Given that the regulation of the apoptotic process in endometrium of luteal phase deficiency is still unknown, the aim of this study was to evaluate cell proliferation, apoptosis and the levels of the main effector caspase, caspase-3 in the luteal in-phase and out-of-phase endometrium.Fil: Meresman, Gabriela Fabiana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Olivares, Carla Noemi. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Vighi, Susana. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Alfie, Margarita. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Irigoyen, Marcela. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Etchepareborda, Juan J.. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; Argentin

Efecto del VEGF sobre la vascularización, foliculogénesis y apoptosis en el ovario de ratonas

Objetivo: El objetivo de este trabajo es investigar los efectos de la inyección ovárica directa de VEGF sobre el desarrollo de folículos antrales, la neoangeogénesis y la apoptosis. Tipo de estudio: Estudio control de laboratorio. Lugar del estudio: Centro de fertilidad afiliado a la Facultad de Medicina de la Universidad de Buenos Aires. Objetos de estudio: Ratonas Balb/c (n=32). Intervención: Las ratonas fueron divididas en 4 grupos: Grupo control (C) n=6, Grupo sin tratamiento; Grupo con hiperestimulacion (HS), n=8, los ovarios fueron hiperestimulados con 7.5 IU de PMSG y 10 IU de HCG; Grupo VEGF (V), n=8, se les realizó una inyección con 0.1 ml de VEGF (0.2 ug) en cada ovario; Grupo V+HS, n=8 se les realizó una inyección con VEGF y luego de 2 semanas se les realizo hiperestimulación ovárica. Medición de resultado: número de folículos antrales y luteinizados, número de vasos y porcentaje de células Bcl-2 positivas. Resultados: El número de folículos antrales en los grupo tratados con VEGF (V y V+HS) fue mayor que en C y HS (16.0 ± 2.5 vs. 6.0 ± 0.9 y 11.3 ± 0.6 respectivamente, p<0.005). Todos los tratamientos incrementaron significativamente el número de vasos (C5.0 ± 0.5 vs. V20.0 ± 4.8, p<0.005 y V+HS 22.2 ± 1.2 p<0.01) así como incrementaron el porcentaje de células positivas para BCl-2 comparados con el grupo control (C=0, V11.8 ± 3.5 p<0.005; V+HS 12.5 ± 3.7 p<0.005). Conclusiones: nuestros hallazgos demuestran que la inyección directa de VEGF en el ovario de ratonas resulta en el desarrollo de una vascularización aumentada promoviendo un incremento del desarrollo folicular y una disminución de la apoptosis.Fil: Quintana, Ramiro. Instituto de Ginecología y Fertilidad; ArgentinaFil: Kopcow, Laura. Instituto de Ginecología y Fertilidad; ArgentinaFil: Young, Edgardo. Instituto de Ginecología y Fertilidad; ArgentinaFil: Sueldo, Carlos. Instituto de Ginecología y Fertilidad; ArgentinaFil: Marconi, Guillermo. Instituto de Ginecología y Fertilidad; ArgentinaFil: Gomez Rueda, Nidia. Instituto de Ginecología y Fertilidad; ArgentinaFil: Vighi, Susana. Instituto de Ginecología y Fertilidad; ArgentinaFil: Barañao, Rosa Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentin

Prenatal Hyperandrogenization Induces Metabolic and Endocrine Alterations Which Depend on the Levels of Testosterone Exposure

Prenatal hyperandrogenism is able to induce polycystic ovary syndrome (PCOS) in rats. The aim of the present study was to establish if the levels of prenatal testosterone may determine the extent of metabolic and endocrine alterations during the adult life. Pregnant Sprague Dawley rats were prenatally injected with either 2 or 5 mg free testosterone (groups T2 and T5 respectively) from day 16 to day 19 day of gestation. Female offspring from T2 and T5 displayed different phenotype of PCOS during adult life. Offspring from T2 showed hyperandrogenism, ovarian cysts and ovulatory cycles whereas those from T5 displayed hyperandrogenism, ovarian cysts and anovulatory cycles. Both group showed increased circulating glucose levels after the intraperitoneal glucose tolerance test (IPGTT; an evaluation of insulin resistance). IPGTT was higher in T5 rats and directly correlated with body weight at prepubertal age. However, the decrease in the body weight at prepubertal age was compensated during adult life. Although both groups showed enhanced ovarian steroidogenesis, it appears that the molecular mechanisms involved were different. The higher dose of testosterone enhanced the expression of both the protein that regulates cholesterol availability (the steroidogenic acute regulatory protein (StAR)) and the protein expression of the transcriptional factor: peroxisome proliferator-activated receptor gamma (PPAR gamma). Prenatal hyperandrogenization induced an anti-oxidant response that prevented a possible pro-oxidant status. The higher dose of testosterone induced a pro-inflammatory state in ovarian tissue mediated by increased levels of prostaglandin E (PG) and the protein expression of cyclooxygenase 2 (COX2, the limiting enzyme of PGs synthesis). In summary, our data show that the levels of testosterone prenatally injected modulate the uterine environment and that this, in turn, would be responsible for the endocrine and metabolic abnormalities and the phenotype of PCOS during the adult life

Apoptosis and expression of Bcl-2 and Bax in eutopic endometrium from women with endometriosis

To evaluate and compare spontaneous apoptosis and Bcl-2 and Bax expression in eutopic endometrium from women with and without endometriosis. Spontaneous apoptosis was significantly lower in eutopic endometrium from patients with endometriosis, compared with healthy controls (2.26 +/- 0.53 and 9.37 +/- 1.69 apoptotic cells/field, respectively) and was independent of cycle phase. An increased expression of Bcl-2 protein was found in proliferative eutopic endometrium from patients with endometriosis. Bax expression was absent in proliferative endometrium, whereas there was an increase in its expression in secretory endometrium from both patients and controls.Women with endometriosis show decreased number of apoptotic cells in eutopic endometrium. The abnormal survival of endometrial cells may result in their continuing growth into ectopic locationsFil: Meresman, Gabriela Fabiana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Vighi, Susana. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Buquet, Ricardo A.. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Contreras Ortiz, Oscar. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Tesone, Marta. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Rumi, Lia S.. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentin

Cytosolic accumulation of HPV16 E7 oligomers supports different transformation routes for the prototypic viral oncoprotein: The amyloid-cancer connection

E7 is the major transforming activity in human papillomaviruses, a causal agent for cervical cancer. HPV16 E7 is a small protein with a natively unfolded domain for which dozens of specific cellular targets were described, and represents a prototypical oncoprotein among small DNA tumor viruses. The protein can form spherical oligomers with amyloid-like properties and chaperone activity. Conformation specific antibodies locate endogenous oligomeric E7 species in the cytosol of 3 model cell lines, strongly colocalizing with amyloid structures and dimeric E7 localizes to the nucleus. The cytosolic oligomeric E7 appear as the most abundant species in all cell systems tested. We show that nuclear E7 levels are replenished dynamically from the cytosolic pool and do not result from protein synthesis. Our results suggest that long-term events related to de-repression of E7 would cause accumulation of excess E7 into oligomeric species in the cytosol. These, together with the known target promiscuity of E7, may allow interactions with many of the non-pRb dependent targets described. This hypothesis is further supported by the detection of E7 oligomers in the cytosol of cancerous cells from tissue biopsies.Fil: Dantur, Karina Ines. 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: Alonso, Leonardo Gabriel. 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: Castaño, Eduardo Miguel. 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: Morelli, Laura. 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: Centeno Crowley, Juan Manuel. 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: Vighi, Susana. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: de Prat Gay, Gonzalo. 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

One step histological detection and staining of the pten tumor suppressor protein by a single strand dna

Antibodies are the most used technological tool in histochemistry. However, even with monoclonal antibodies, their standardization is difficult due to variation of biological systems as well as to variability due to the affinity and amplification of the signal arising from secondary peroxidase detection systems. In this article we combined two synthetic molecules to facilitate the standardization of a detection protocol of protein markers in histological sections. The first molecule was an aptamer, a 50-base single-stranded DNA fragment, which recognizes a PTEN tumor suppressor. The second molecule used was also another single stranded 18-base aptamer DNA fragment, which forms a quadruplex structure guanine box. This G-quadruplex recognizes and attaches a molecule of hemin, increasing the catalytic capacity for the hydrogen peroxide. Our results show how the correct structural design of DNA combining an aptamer together with the peroxidase-like DNAzyme allows to detect proteins in histological sections. This tool offers the standardization of the detection of prognostic markers in cancer, in quality and quantity, due to its synthetic nature and its 1:1 antigen:enzyme ratio. This is the first time that reproducible results have been presented in histological sections staining a cancer marker using a single-stranded DNA molecule with dual function.Fil: Longinotti, María Gloria. Instituto Nacional de Tecnología Industrial; ArgentinaFil: Ybarra, Gabriel Omar. Instituto Nacional de Tecnología Industrial; ArgentinaFil: Vighi, Susana. Universidad de Buenos Aires; ArgentinaFil: Perandones, Claudia. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud "Dr. C. G. Malbrán"; ArgentinaFil: Montserrat, Javier Marcelo. Universidad Nacional de General Sarmiento. Instituto de Ciencias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Yakisich, Juan Sebastian. Hampton University; Estados UnidosFil: Grasselli, Mariano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; ArgentinaFil: Radrizzani Helguera, Martin. Universidad Nacional de San Martín. Escuela de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Multiple stressors in Mediterranean coastal wetland ecosystems : Influence of salinity and an insecticide on zooplankton communities under different temperature conditions

Temperature increase, salinity intrusion and pesticide pollution have been suggested to be among the main stressors affecting the biodiversity of coastal wetland ecosystems. Here we assessed the single and combined effects of these stressors on zooplankton communities collected from a Mediterranean coastal lagoon. An indoor microcosm experiment was designed with temperature variation (20 °C and 30 °C), salinity (no addition, 2.5 g/L NaCl) and the insecticide chlorpyrifos (no addition, 1 μg/L) as treatments. The impact of these stressors was evaluated on water quality variables and on the zooplankton comunity (structure, diversity, abundance and taxa responses) for 28 days. This study shows that temperature is the main driver for zooplankton community change, followed by salinity and chlorpyrifos. The three stressors contributed to a decrease on zooplankton diversity. The increase of temperature contributed to an increase of zooplankton abundance. Salinity generally affected Cladocera, which resulted in a Copepoda increase at 20 °C, and a reduction in the abundance of all major zooplankton groups at 30 °C. The insecticide chlorpyrifos affected primarily Cladocera, altough the magnitude and duration of the direct and indirect effects caused by the insecticide substantially differed between the two temperature scenarios. Chlorpyrifos and salinity resulted in antagonistic effects on sensitive taxa (Cladocera) at 20 °C and 30 °C. This study shows that temperature can influence the direct and indirect effects of salinity and pesticides on zooplankton communities in Mediterranean coastal wetlands, and highlights vulnerable taxa and ecological responses that are expected to dominate under future global change scenarios.</p

Argentophilic nucleolus organizer region as a proliferation marker in cervical intraepithelial neoplasia grade 1 of the uterine cervix

AIM: p16INK4a and argentophilic nucleolus organizer region (AgNOR) can be used as markers for progression of cervical intraepithelial neoplasia grade 1 (CIN1) of the uterine cervix. Our objective was to study the predictive value of the AgNOR technique as a progression marker of CIN1 and its correlation with p16INK4A. MATERIAL AND METHODS: One uterine cervix biopsy from each of 75 patients with diagnosis of CIN1 was selected. All of these patients underwent a second biopsy, and these were also used for the study. RESULTS: The second biopsies showed: regression (20 patients), persistent CIN1 (38 patients), progression to CIN2 (10 patients) and progression to CIN3 (seven patients). p16INK4A showed reactivity in 67 of the 75 first CIN1 biopsies: 12 of the 20 cases that cleared the lesions and the 55 cases with persistent or progressive lesions were positive for p16INK4a (specificity: 40%; sensitivity: 100%; positive predictive value [PPV]: 82%; negative predictive value [NPV]: 100%). Samples with AgNOR areas less than 3.0 μ(2) returned in all cases, but patients whose lesions persisted or progressed to CIN2/CIN3, showed AgNOR areas greater than 3.0 μ(2) in 50/55 cases (specificity: 100%; sensitivity: 91%; PPV: 100%; NPV: 80%). CONCLUSIONS: p16INK4a is expressed in a high percentage of returning lesions. AgNOR might be a better marker of proliferation of CIN1 than p16INK4a (PPV = 100%), which means that a value greater than 3.0 μ(2) indicates the persistence or progression of the lesion. As its NPV is 80%, a value of AgNOR area less than 3.0 μ(2) in CIN1 leaves a margin of doubt about the future behavior of the lesion.Fil: Guerra, Fernando Andres. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Rocher, Adriana Esther. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Villacorta Hidalgo, José. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Díaz, Lilí. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Vighi, Susana. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Cardinal, Lucía. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Tatti, Silvio. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Cúneo, Nicasio. Hospital de Oncología María Curie; ArgentinaFil: de Prat Gay, Gonzalo. 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: Camporeale, Gabriela. 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: Palaoro, Luis Alberto. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; Argentin