Do GnRH analogues directly affect human endometrial epithelial cell gene expression?

We examined whether Gonadotrophin-releasing hormone (GnRH) analogues [leuprolide acetate (LA) and ganirelix acetate (GA)] modulate gene expression in Ishikawa cells used as surrogate for human endometrial epithelial cells in vitro. The specific aims were: (i) to study the modulatory effect of GnRH analogues by RT –PCR [in the absence and presence of E2 and P4, and cyclic adenosine monophosphate (cAMP)] on mRNA expression of genes modulated during the window of implantation in GnRH analogues/rFSH-treated assisted reproductive technology cycles including OPTINEURIN (OPTN), CHROMATIN MODIFYING PROTEIN (CHMP1A), PROSAPOSIN (PSAP), IGFBP-5 and SORTING NEXIN 7 (SNX7), and (ii) to analyze the 5′ -flanking regions of such genes for the presence of putative steroid-response elements [estrogen-response elements (EREs) and P4-response element (PREs)]. Ishikawa cells were cytokeratin+/vimentin2 and expressed ERa, ERb, PR and GnRH-R proteins. At 6 and 24 h, neither LA nor GA alone had an effect on gene expression. GnRH analogues alone or following E2 and/or P4 co-incubation for 24 h also had no effect on gene expression, but P4 significantly increased expression of CHMP1A. E2 + P4 treatment for 4 days, alone or followed by GA, had no effect, but E2 + P4 treatment followed by LA significantly decreased IGFBP-5 expression. The addition of 8-Br cAMP did not modify gene expression, with the exception of IGFBP-5 that was significantly increased. The GnRH analogues did not modify intracellular cAMP levels. We identified conserved EREs for OPN, CHMP1A, SNX7 and PSAP and PREs for SNX7. We conclude that GnRH analogues appear not to have major direct effects on gene expression of human endometrial epithelial cells in vitro.Fil: Zhang, Xiaomei. Eastern Virginia Medical School; Estados UnidosFil: Bocca, Silvina. Eastern Virginia Medical School; Estados UnidosFil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Anderson, Sandra. Eastern Virginia Medical School; Estados UnidosFil: Kaur, Mandeep. King Abdullah University of Science and Technology; Arabia SauditaFil: Bajic, Vladimir B.. King Abdullah University of Science and Technology; Arabia SauditaFil: Oehninger, Sergio. Eastern Virginia Medical School; Estados Unido

Enseñanza de la biología celular y molecular basada en la práctica científica y el aprendizaje cooperativo

Desde el descubrimiento del ADN, el conocimiento científico en Biología Celular y Molecular (BCM) ha crecido exponencialmente. Este hecho, pone límites a la enseñanza tradicional memorista basada en un aprendizaje pasivo. Por lo tanto, resulta necesario cambiar la forma de enseñanza de la BCM hacia una forma que incluya las prácticas científicas para conseguir, analizar y comprender nuevos conocimientos científicos, mediante el aprendizaje cooperativo, el cual puede mejorar sustantivamente el aprendizaje. El objetivo fue evaluar la eficiencia de una nueva propuesta pedagógica basada prácticas científicas y el aprendizaje cooperativo para introducir al alumno en el pensamiento científico. La propuesta pedagógica proporcionó una plataforma de actividades de prácticas científicas, facilitando que los alumnos desarrollen su capacidad cognitiva, interpretativa e integradora, así como también la capacidad para comprender y comunicar el conocimiento científico en forma escrita y oral. Los resultados de esta propuesta tienen potencial aplicación transversal en otras asignaturas de la carrera, en la vida profesional y a lo largo de la vida.Fil: Giojalas, Laura Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaFil: Guidobaldi, Héctor Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaFil: Cragnolini, Andrea Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaFil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaFil: Garcia, Leticia. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Danelon, Víctor. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. State University of New Jersey; Estados UnidosFil: Moreno Yrusta, A.. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; ArgentinaFil: Dominguez, Esteban Mauricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaFil: Figueras López, María Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Biología Celular; ArgentinaI Jornadas de Experiencias e Investigación Educativas en Ciencias Exactas y NaturalesCórdobaArgentinaUniversidad Nacional de Córdoba. Facultad de ciencias Exactas, Físicas y Naturales. Escuela de Ingeniería Químic

DNA fragmentation of normal spermatozoa negatively impacts embryo quality and intracytoplasmic sperm injection outcome

Objective: To evaluate DNA fragmentation in morphologically normal sperm recovered from the same sample used for intracytoplasmic sperm injection (ICSI) and to correlate DNA damage with embryo quality and pregnancy outcome. Design: Prospective study. Setting: Academic center. Patient(s): 36 infertile men participating in the ICSI program. Intervention(s): Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-fluorescein nick end labeling (TUNEL) assay and morphologic assessment by phase contrast. Main Outcome Measure(s): Simultaneous assessment of sperm morphology and DNA fragmentation by TUNEL assay was performed in the same cell, then the percentage of normal sperm with fragmented DNA (normal SFD) was correlated with embryo quality and pregnancy outcomes. Result(s): A highly statistically significant negative correlation was found between the percentage of normal SFD and embryo quality. This association was confirmed for the transferred embryos and for the total embryo cohort. The receiver operating characteristics curve analysis demonstrated that the percentage of normal SFD and embryo quality were statistically significant predictors of pregnancy. When the percentage of normal SFD was ≤17.6 %, the likelihood of pregnancy was 3.5 times higher. No correlation was found between the percentage of total sperm with fragmented DNA (morphologically normal and abnormal) and ICSI outcomes. Conclusion(s): The DNA fragmentation of morphologically normal sperm negatively impacts embryo quality and probability of pregnancy in ICSI cycles.Fil: Avendaño, Conrado. Eastern Virginia Medical School; Estados UnidosFil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Eastern Virginia Medical School; Estados UnidosFil: Duran, Hakan. Eastern Virginia Medical School; Estados UnidosFil: Oehninger, Sergio. Eastern Virginia Medical School; Estados Unido

Extracellular vesicles from oviductal isthmus and ampulla stimulate the induced acrosome reaction and signaling events associated with capacitation in bovine spermatozoa

Cells can communicate with other neighboring or distant cells through the secretion of extracellular vesicles (EV), composed of a lipid bilayer and bearing surface molecules that allow them to recognize target cells. In this way, EV induce signaling via different mechanisms, modulating the physiological state of the recipient cell. EV have been identified in both male and female reproductive fluids, however, the possible role of EV isolated from female reproductive fluids has become an emerging field only recently. It is known that ejaculated mammalian spermatozoa need to undergo physiological preparation in the female reproductive tract to fertilize the egg. EV secreted by different regions of the female tract constitute signals that may have a key role in regulating sperm functions. The aims of the present study were isolating EV from different regions of the bovine oviduct and analyzing their interaction and physiological effects on spermatozoa. Here, we report the characterization of bovine oviductal fluid EV from the isthmus and ampulla region and their effect on the induced acrosome reaction and signaling events associated with sperm capacitation. EV induced an increase in sperm protein tyrosine phosphorylation, while cell survival of cryopreserved bovine spermatozoa was maintained. We also show that EV uptake regulates the sperm calcium levels by inducing an immediate increase in the intracellular calcium concentration and sperm priming, after a pre-incubation period, of the progesterone-induced intracellular calcium rise. Our data contribute to understand the role of EV in the communication between the female reproductive tract and the sperm physiology, information that may be used to improve the efficiency of reproductive assisted technologies.Fil: Franchi, Nilda Anahi. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Centro de Biología Celular y Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Moreno, Ayelen. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Centro de Biología Celular y Molecular; ArgentinaFil: Dominguez, Esteban Mauricio. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Centro de Biología Celular y Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Adre, Amira Jasmine. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Centro de Biología Celular y Molecular; ArgentinaFil: Giojalas, Laura Cecilia. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Centro de Biología Celular y Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentin

Uterosome-like vesicles prompt human sperm fertilizing capability

STUDY QUESTION: Does the rapid transit through the uterine environment modulate the sperm physiological state?SUMMARY ANSWER: The uterosome-like vesicles (ULVs) secreted by endometrial epithelial cells (EECs) in vitro are able to fuse withhuman spermatozoa, prompting their fertilizing capacity.WHAT IS KNOWN ALREADY: Early studies suggest that sperm capacitation begins in the uterus and ends in the oviduct, and that a synergisticeffect of both female organs may accelerate this process. Although it has been reported that co-incubation of human spermatozoawith endometrial cell-conditioned medium (CM) stimulates sperm capacitation, the mechanism mediating this communication is unknown.STUDY DESIGN, SIZE, DURATION: Human ULVs secreted by EECs were characterized and their effect on human sperm physiologywas analysed. Spermatozoa were incubated with EEC-derived CM or ULV, after which sperm capacitation was evaluated at different timepoints. In addition, the interaction of spermatozoa with ULV was analysed.PARTICIPANTS/MATERIALS, SETTING, METHODS: ULVs were isolated by ultracentrifugation and identified using electronmicroscopy and Western blotting to assess the presence of specific protein markers. Following seminal plasma removal, human spermatozoawere incubated CM or ULV, after which sperm capacitation was evaluated as the ability of the sperm to undergo the induced acrosome reactionand the level of protein tyrosine phosphorylation (PY) determined by Western blot and immunocytochemistry. The interaction ofspermatozoa with labelled ULV was analysed by fluorescence microscopy. In all cases, at least three biological replicates from different spermdonors were performed for each set of experiments. Significant differences between mean values were determined by one-way ANOVA followedby Tukey?s post hoc test. Differences between treatments were considered statistically significant at P ¢¢çÜ 0.05.MAIN RESULTS AND THE ROLE OF CHANCE: The level of capacitated spermatozoa and those recruited by chemotaxis increased3- to 4-fold when spermatozoa were incubated in the presence of CM for 4 h. Even a 15 min incubation of spermatozoa with CM was alsoenough to increase the level of capacitated cells 3- to 4-fold (P < 0.05). Furthermore, a short co-incubation of spermatozoa with ULV stimulatessperm capacitation, as determined by the increase in the level of induced acrosome reaction and the induction of PY. In addition, afterthe co-incubation of spermatozoa with fluorescent labelled ULV, the sperm cells acquired the fluorescent staining which indicates that ULVmight be transferred to the sperm surface by a fusion mechanism.LIMITATIONS, REASONS FOR CAUTION: This is an in vitro study performed with human biological material, spermatozoa and endometrialderived cells; the latter being a cell line originally isolated from a uterine adenocarcinoma.WIDER IMPLICATIONS OF THE FINDINGS: The capability of spermatozoa to briefly interact with ULVs supports the hypothesis thatany step of sperm transport may have physiological consequences, despite the interaction lasting for only a limited period of time. This way ofcommunication of spermatozoa with cell products of uterine origin opens new frontiers of investigation (e.g. the signalling moleculesinvolved), shedding light on the sperm processes that prepare the male gamete for fertilization, which might have implications for human infertilitytreatment.LARGE SCALE DATA: N/A.Fil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Cubilla, Marisa Angélica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Guidobaldi, Héctor Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Bravo, Azucena Anahí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Giojalas, Laura Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentin

Epithelial cell protein milk fat globule-epidermal growth factor 8 and human chorionic gonadotropin regulate stromal cell apoptosis in the human endometrium

To study the regulation of apoptosis in human endometrial cells. The specific aims were to determine whether milk fat globule-epidermal growth factor 8 (MFG-E8), a novel endometrial epithelial protein, modulates caspase activation and DNA fragmentation; and to examine whether hCG, an early embryonic product, regulates Bax and Bcl-2 equilibrium, as well as MFG-E8 expression. Design: Primary cultures of human endometrial epithelial cells (EECs) and endometrial stromal cells (ESCs). Setting: Academic center. Patient(s): Ovulatory women aged 21-30 years. Intervention(s): Treatment with MFG-E8 and hCG. Main Outcome Measure(s): Apoptotic activity was quantified using a luciferase assay. Deoxyribonucleic acid fragmentation was detected by TUNEL assay. Bax, Bcl-2, and MFG-E8 messenger RNA expression levels were determined by quantitative reverse transcription-polymerase chain reaction. Immunocytochemistry was used to establish cell purity and presence of MFG-E8 and hCG-R (receptor) proteins. Result(s): Endometrial epithelial cells were cytokeratin+, vimentin-, MFG-E8+, and hCG-R+, whereas ESC were vimentin+, cytokeratin -, MFG-E8-, and hCG-R+. Treatment of ESC with MFG-E8 resulted in a 13-fold increase in caspase activity and a 30-fold increase in TUNEL. On the other hand, hCG decreased messenger RNA expression of Bax in ESC. Conclusion(s): Milk fat globule-epidermal growth factor 8 has proapoptotic activity, suggesting participation in endometrial remodeling via an epithelial-stromal cell paracrine effect. Conversely, pregnancy levels of hCG has opposite effects on stromal cells.Fil: Riggs, Ryan M.. Eastern Virginia Medical School; Estados UnidosFil: Bocca, Silvina. Eastern Virginia Medical School; Estados UnidosFil: Anderson, Sandra. Eastern Virginia Medical School; Estados UnidosFil: Franchi, Nilda Anahi. Eastern Virginia Medical School; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Rhavi, Bhaskara S.. Old Dominion University; Estados UnidosFil: Oehninger, Sergio. Eastern Virginia Medical School; Estados Unido

Do GnRH analogues directly affect human endometrial epithelial cell gene expression?

We examined whether Gonadotrophin-releasing hormone (GnRH) analogues [leuprolide acetate (LA) and ganirelix acetate (GA)] modulate gene expression in Ishikawa cells used as surrogate for human endometrial epithelial cells in vitro. The specific aims were: (i) to study the modulatory effect of GnRH analogues by RT –PCR [in the absence and presence of E2 and P4, and cyclic adenosine monophosphate (cAMP)] on mRNA expression of genes modulated during the window of implantation in GnRH analogues/rFSH-treated assisted reproductive technology cycles including OPTINEURIN (OPTN), CHROMATIN MODIFYING PROTEIN (CHMP1A), PROSAPOSIN (PSAP), IGFBP-5 and SORTING NEXIN 7 (SNX7), and (ii) to analyze the 5′ -flanking regions of such genes for the presence of putative steroid-response elements [estrogen-response elements (EREs) and P4-response element (PREs)]. Ishikawa cells were cytokeratin+/vimentin2 and expressed ERa, ERb, PR and GnRH-R proteins. At 6 and 24 h, neither LA nor GA alone had an effect on gene expression. GnRH analogues alone or following E2 and/or P4 co-incubation for 24 h also had no effect on gene expression, but P4 significantly increased expression of CHMP1A. E2 + P4 treatment for 4 days, alone or followed by GA, had no effect, but E2 + P4 treatment followed by LA significantly decreased IGFBP-5 expression. The addition of 8-Br cAMP did not modify gene expression, with the exception of IGFBP-5 that was significantly increased. The GnRH analogues did not modify intracellular cAMP levels. We identified conserved EREs for OPN, CHMP1A, SNX7 and PSAP and PREs for SNX7. We conclude that GnRH analogues appear not to have major direct effects on gene expression of human endometrial epithelial cells in vitro.Fil: Zhang, Xiaomei. Eastern Virginia Medical School; Estados UnidosFil: Bocca, Silvina. Eastern Virginia Medical School; Estados UnidosFil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Anderson, Sandra. Eastern Virginia Medical School; Estados UnidosFil: Kaur, Mandeep. King Abdullah University of Science and Technology; Arabia SauditaFil: Bajic, Vladimir B.. King Abdullah University of Science and Technology; Arabia SauditaFil: Oehninger, Sergio. Eastern Virginia Medical School; Estados Unido

Milk fat globule epidermal growth factor 8 (MFG-E8): A novel protein in the mammalian endometrium with putative roles in implantation and placentation

MFG-E8 is a novel endometrial protein with conserved functions in tissue remodeling and angiogenesis in non-uterine tissues. Our aims were: 1. To examine the presence of MFG-E8 protein in the human endometrium during the window of implantation, in human endometrial cell lines, in human placental tissue at different gestational ages, and in murine implantation sites during early gestation; and 2. To study the regulation of MFG-E8 mRNA expression in mice implantation sites. Study design: MFG-E8 protein and its receptor integrin αvβ3 were detected by immunostaining in human endometrial biopsies obtained from normal volunteers, in human endometrial cell lines (epithelial: Ishikawa and HEC-1A, stromal: HESC, and endothelial: HEEC), in human products of conception from all trimesters of gestation, and in murine implantation and inter-implantation sites dissected on days 5 and 8 post-coitus. MFG-E8 gene expression was assessed by RT-PCR. Main outcome measures: Immunohistochemical determination of MFG-E8 in endometrium and products of conception as well as relative MFG-E8 mRNA expression in mice implantation sites. Results: MFG-E8 protein was present almost exclusively in the epithelial compartment of human endometrium. It was also expressed in the cytotrophoblasts and syncytiotrophoblasts outlining chorionic villi of the human placenta at all trimesters of gestation, and in murine implantation sites. MFG-E8 mRNA was significantly up-regulated in murine implantation sites and with increased gestational age. Conclusions: MFG-E8 expression in the endometrial epithelium as well as in chorionic villi suggests its possible role in endometrial reorganization during the receptive phase and in events related to normal pregnancy in mammals.Fil: Bocca, Silvina M.. Eastern Virginia Medical School; Estados UnidosFil: Anderson, Sandra. Eastern Virginia Medical School; Estados UnidosFil: Amaker, Barbara. Sentara Norfolk General Hospital; Estados UnidosFil: Swanson, R. James. Old Dominion Univeristy; Estados UnidosFil: Franchi, Nilda Anahi. Eastern Virginia Medical School; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Lattanzio, Frank A.. Eastern Virginia Medical School; Estados UnidosFil: Oehninger, Sergio Carlos. Eastern Virginia Medical School; Estados Unido

β-microseminoprotein in human spermatozoa and its potential role in male fertility

β-Microseminoprotein (MSMB) is one of the most abundant proteins in human seminal plasma. The objectives of this study were: (1) to purify MSMB from seminal plasma (SP) and generate antibodies against the pure protein; (2) to investigate the interaction of MSMB with ejaculated spermatozoa and its possible effect on the spontaneous acrosome reaction (AR); and (3) to quantify MSMB content in SP and examine its relationship with the clinical sperm parameters. MSMB was purified from SP and its presence on the sperm surface was examined by indirect immunofluorescence using a specific polyclonal antibody. The effect of MSMB on the AR was evaluated using guinea pig epididymal spermatozoa as a model. MSMB quantification assay was performed with a two-site binding ELISA using two polyclonal antibodies against MSMB. MSMB was assessed in semen samples from fertile donors (controls) and subfertile patients according to World Health Organization criteria. MSMB was detected on the sperm surface and mainly localized to the acrosomal region of the head and neck. A significant spontaneous AR inhibition was observed when guinea pig epididymal spermatozoa were preincubated with MSMB. Finally, MSMB was significantly increased in subfertile patients when compared with fertile controls (P<0.02). The association of MSMB to the sperm surface, the inhibitor effect on the spontaneous AR and the increased MSMB levels found in SP in subfertile men suggests a relationship between this protein and semen quality and a possible role in the process of fertilization. © 2008 Society for Reproduction and Fertility.Fil: Franchi, Nilda Anahi. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Avendaño, Conrado. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Molina, Rosa I.. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Tissera, Andrea D.. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Maldonado, Cristina Alicia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Oehninger, Sergio. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Ciencias y Tecnología de los Alimentos; ArgentinaFil: Coronel, Carlos Enrique. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentin

Transcriptome of early embryonic invasion at implantation sites in a murine model

Successful implantation relies on the interaction between a competent embryo and a receptive endometrium. The aim of the present study was to investigate genes differentially expressed in early invasive embryonic tissue versus decidual tissue in mice. Samples were obtained from the ectoplacental cone, the immediately surrounding deciduas and from deciduas from interimplantation sites. Microarray analysis showed that 817 genes were differentially expressed between extra-embryonic tissue and the surrounding decidua and that 360 genes were differentially expressed between the different deciduas, with a high representation of developmental processes. Genes differentially expressed in the maternal compartment included chemokines, lipoproteins, growth factors and transcription factors, whereas the embryonic invasive tissue expressed genes commonly observed in invasive tumour-like processes. These results provide information about genes involved in early embryonic invasion and the control exerted by the surrounding decidua. This information may be useful to find targets involved in pathologies associated with implantation failure and early pregnancy loss.Fil: Moreno Moya, J. M.. Universidad de Valencia; EspañaFil: Franchi, Nilda Anahi. Eastern Virginia Medical School; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Martínez Escribano, S.. Universidad de Valencia; EspañaFil: Martínez Conejero, J. A.. Universidad de Valencia; EspañaFil: Bocca, S.. Eastern Virginia Medical School; Estados UnidosFil: Oehninger, S.. Eastern Virginia Medical School; Estados UnidosFil: Horcajadas, J. A.. Unviersidad Pablo de Olavide; Españ