Mifepristone increases mRNA translation rate, triggers the unfolded protein response, increases autophagic flux, and kills ovarian cancer cells in combination with proteasome or lysosome inhibitors

The synthetic steroid mifepristone blocks the growth of ovarian cancer cells, yet the mechanism driving such effect is not entirely understood. Unbiased genomic and proteomic screenings using ovarian cancer cell lines of different genetic backgrounds and sensitivities to platinum led to the identification of two key genes upregulated by mifepristone and involved in the unfolded protein response (UPR): the master chaperone of the endoplasmic reticulum (ER), glucose regulated protein (GRP) of 78 kDa, and the CCAAT/enhancer binding protein homologous transcription factor (CHOP). GRP78 and CHOP were upregulated by mifepristone in ovarian cancer cells regardless of p53 status and platinum sensitivity. Further studies revealed that the three UPR-associated pathways, PERK, IRE1α, and ATF6, were activated by mifepristone. Also, the synthetic steroid acutely increased mRNA translation rate, which, if prevented, abrogated the splicing of XBP1 mRNA, a non-translatable readout of IRE1α activation. Moreover, mifepristone increased LC3-II levels due to increased autophagic flux. When the autophagic–lysosomal pathway was inhibited with chloroquine, mifepristone was lethal to the cells. Lastly, doses of proteasome inhibitors that are inadequate to block the activity of the proteasomes, caused cell death when combined with mifepristone; this phenotype was accompanied by accumulation of poly-ubiquitinated proteins denoting proteasome inhibition. The stimulation by mifepristone of ER stress and autophagic flux offers a therapeutic opportunity for utilizing this compound to sensitize ovarian cancer cells to proteasome or lysosome inhibitors.Fil: Zhang, Lei. University Of South Dakota; Estados UnidosFil: Hapon, María Belén. University Of South Dakota; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Goyeneche, Alicia A.. University Of South Dakota; Estados Unidos. McGill University; CanadáFil: Srinivasan, Rekha. University Of South Dakota; Estados UnidosFil: Gamarra Luques, Carlos Diego. University Of South Dakota; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Callegari, Eduardo A.. University Of South Dakota; Estados UnidosFil: Drappeau, Donis D.. University Of South Dakota; Estados UnidosFil: Terpstra, Erin J.. University Of South Dakota; Estados UnidosFil: Pan, Bo. University Of South Dakota; Estados UnidosFil: Knapp, Jennifer R.. University of Kansas; Estados UnidosFil: Chien, Jeremy. University of Kansas; Estados UnidosFil: Wang, Xuejun. University Of South Dakota; Estados UnidosFil: Eyster, Kathleen M.. University Of South Dakota; Estados UnidosFil: Telleria, Carlos Marcelo. University Of South Dakota; Estados Unidos. McGill University; Canadá. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Comparative proteomics of soluble factors secreted by human breast adipose tissue from tumor and normal breast

Tumor progression depends on the tumor-stroma interaction. In the breast, adipose tissue is the predominant stromal type. We have previously demonstrated that conditioned media (CMs) from explants of human adipose tissue of tumor breasts (hATT) increase proliferation and migration of breast cancer epithelial cells when compared to human adipose tissue from normal breasts (hATN). In this work, we aim to identify specific proteins and molecular/biological pathways associated with the secretion profile of hATT and hATN explants. hATT-CMs and hATN-CMs were separated by SDS-PAGE and analyzed by means of two-dimensional nano-liquid chromatography-mass spectrometry. The data was analyzed using ProteoIQ and FunRich software. In addition, 42 cytokines from hATTCMs and hATN-CMs were assayed by a protein antibody assay. Compared to hATNCMs, hATT-CMs showed greater protein diversity. We found that hATT-CMs presented a greater amount of proteins related to complement system activity, metabolism and immune system, as well as proteins involved in a variety of biological processes such as signal transduction and cell communication. Specifically, apolipoprotein AI and AII, complement component 3, and vimentin and desmin were significantly increased in hATT-CMs versus hATN-CMs. Moreover, a multivariate discriminant analysis of the cytokines detected by the array showed that IL-6, MCP-2 and GRO cytokines were sufficient and necessary to differentiate hATT-CMs from hATN-CMs. This analysis also showed that the levels of these three cytokines, taken together, correlated with stage and histological grade of the tumor in the hATT-CMs group, and with body mass index in the hATN-CMs group.Fil: Fletcher, Sabrina Johanna. 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: Hapon, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Callegari, E.. University Of South Dakota; Estados UnidosFil: Crosbie, M. L.. Complejo Médico Policial "Churruca Visca"; ArgentinaFil: Santiso, N.. Complejo Médico Policial "Churruca Visca"; ArgentinaFil: Ursino, A.. Complejo Médico Policial "Churruca Visca"; ArgentinaFil: Amato, A. R.. Complejo Médico Policial "Churruca Visca"; ArgentinaFil: Gutiérrez, A.. Complejo Médico Policial "Churruca Visca"; ArgentinaFil: Sacca, Paula Alejandra. 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: Dreszman, R.. 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: Pérez, A.. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Matemática; ArgentinaFil: Caron, Ruben Walter. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Calvo, Juan Carlos. 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: Pistone Creydt, Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

Effects of 2,4-dichlorophenoxyacetic acid on the ventral prostate of rats during the peri-pubertal, pubertal and adult stage

The herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) is used on a wide variety of terrestrial and aquatic broadleaf weeds. 2,4-D has been shown to produce a wide range of adverse effects on animal and human health. The aim of the current study was to evaluate the effects of pre- and postnatal exposure to 2,4-D on rat ventral prostate (VP). Pregnant rats were exposed daily to oral doses of 70 mg/kg/day of 2,4-D from 16 days of gestation up to 23 days after delivery. Then, the treated groups (n = 8) were fed with a 2,4-D added diet until sacrificed by decapitation on postnatal day (PND) 45, 60, or 90. Morphometric studies were performed and androgen receptor (AR) protein levels in the VP were determined. AR, insulin-like growth factor-I (IGF-1) and insulin-like growth factor-I receptor (IGF-1R) mRNA expression in the VP along with testosterone (T), dihydroxytestosterone (DHT), growth hormone (GH) and IGF-1 serum levels were also determined to ascertain whether these parameters were differentially affected. Results of this study showed that 2,4-D exposure during gestation and until adulthood altered development of the prostate gland in male rats, delaying it at early ages while increasing its size in adults, indicate that 2,4-D could behave as endocrine disruptors (EDs).Fil: Pochettino, Arístides A. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Laboratorio de Toxicología Experimental; Argentina.Fil: Hapon, María Belén. Universidad Nacional de Cuyo. Laboratorio De Reproducción y Lactancia (IMBECU-CONICET); Argentina.Fil: Biolatto, Silvana M. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Laboratorio de Toxicología Experimental; Argentina.Fil: Madariaga, María José. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Laboratorio de Toxicología Experimental; Argentina.Fil: Jahn, Graciela A. Universidad Nacional de Cuyo. Laboratorio De Reproducción y Lactancia (IMBECU-CONICET); Argentina.Fil: Konjuh, Cintia N. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Laboratorio de Toxicología Experimental; Argentina

Hipothyroidism produces change in the uterine vasculature during the implantation process

Hypothyroidism is one of the most common endocrine abnormalities implicated in the recurrent loss of pregnancy. Our laboratory, have shown that hypothyroidism in the rat is associated with a lower number of pups per litter due to a lower number of implantation sites and a decrease in the proliferation of the endothelial and decidual cells during the process of implantation of the embryo. On the other hand, is known that angiogenesis is a critical process in the uterine endometrium for embryo implantation, maintenance of early pregnancy, and development of the placenta. During this period, steroid hormones (E2 and P4) stimulate the synthesis of vascular endothelial growth factor (VEGF-A), the main modulator of angiogenesis during peri-implantation period. Therefore we hypothesize that hypothyroidism affects the normal vascularization of endometrium during implantation. The aim of this work was to study the effect of hypothyroidism on the degree of vascularization of the uterine decidua during the implantation process. Hypothyroidism was induced in female Wistar rats by daily administration of 6-propyl-2-thiouracil (PTU) 0,1 g/L in drinking water. In addition, hormone replacement therapy with T3 was administered simultaneously to the treatment with PTU (PTU+T3), in daily physiological doses of 0.6ug/100g. Both groups were compared to rats that only drink tap water (Control),on day five (G5) and seven (G7) of gestation. Uterine vascularization was evaluated by immunofluorescence. Besides,mRNA expression of PECAM (Platelet endothelial cell adhesion molecule, an indicator of the presence of endothelial cells) and VEGF-A were evaluated during the same peri-implantation periods (G5 y G7)by RTqPCR. Our results demonstrate that hypothyroidism decreases vascularization density of the uterine tissue during the process of implantation of the embryo (p<0.05). On the other hand, our results demonstrated a significant increase of expression of VEGF mRNA when hypothyroid rats were treated with T3 before implantation, in comparison to the control group and hypothyroid group (p< 0.05). However, no changes were found in the levels of PECAM expression among the different groups. In conclusion, the failure of implantation due to hypothyroidism may be directly linked to expression of VEGF-A, and consequently to vascularization of the endometrium before implantation in early gestation. Although are necessary further studies that corroborate the exact mechanism, our results identify molecular targets regulated by thyroid hormones that may link hypothyroidism to implantation failure and recurrent miscarriage.Fil: Troncoso, Mariana Elizabeth. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Rinaldini, Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ezquer, M. E.. Universidad del Desarrollo; ChileFil: Gamarra Luques, Carlos Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Hapon, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaXXXVI Reunión Científica Anual de la Sociedad Cuyo de BiologíaMendozaArgentinaSociedad de Biología de Cuy

Hormone Levels and Ultrasound Evaluation of Caiman latirostris (Crocodylia, Alligatoridae) Ovulation

Although there is much information available about reproduction in Caiman latirostris, knowledge related to steroid hormone levels and follicle development for wild adult females is still lacking. In this study we monitored and assessed the development of follicles and eggs and correlated these results with plasma steroid hormone levels in 32 adult females captured in Santa Fe, Argentina. Fieldwork was carried out over two reproductive seasons (October?January) between 2010 and 2012. Using an ultrasound device to take images of the reproductive structures of adult females, we observed individuals with vitellogenic follicles (n = 5), eggs (n = 4), atretic follicles (n = 11), and inactive reproductive structures (n = 12). We found no reproductive females smaller than 65 cm snout?vent length. High levels of estradiol were found during the ovulation period (November) only in reproductive females. Reproductive females showed no differences in progesterone levels during the study period (November?January) compared to non-reproductive females; however, reproductive females showed higher progesterone levels during nesting (December). We found no differences in progesterone levels between reproductive females and females with atretic follicles at the end of the nesting period (January). Ultrasound imaging was found to be an efficient technique to study reproductive structures at the beginning of reproductive cycle of the broad-snouted caiman. Isolated analyses of hormonal levels are not sufficient to determine the reproductive condition of C. latirostris females.Fil: Costa Gonçalves Portelinha, Thiago. Provincia de Entre Ríos. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción. Universidad Autónoma de Entre Ríos. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción; Argentina. Universidade Federal do Tocantins; Brasil. Universidad Nacional del Litoral; ArgentinaFil: Jahn, Graciela Alma. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Hapon, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Verdade, Luciano M.. Universidade de Sao Paulo; BrasilFil: Piña, Carlos Ignacio. Provincia de Entre Ríos. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción. Universidad Autónoma de Entre Ríos. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Centro de Investigaciones Científicas y Transferencia de Tecnología a la Producción; Argentina. Universidad Nacional del Litoral; Argentina. Universidad Autónoma de Entre Ríos; Argentin