RESUMEN
Hypoxia-inducible factors (HIFs) are oxygen-dependent transcriptional activators, but there is little information about their role in yak (Bos grunniens) reproduction. The present study, for the first time, investigated the adaptive mechanism of yak reproduction to high-altitude hypoxic stress by comparing the expression of HIF mRNAs between female yaks at high-altitude and cattle at low-altitude. Hypothalamus, anterior pituitary, oviduct, ovary and uterus tissue samples were collected from five adult female yaks and cattle. mRNA expression was determined by the quantitative real-time polymerase chain reaction. Both HIF-1α and HIF-2α were expressed in all five tissues examined from both species, albeit at different levels. In yaks, the highest mRNA levels of HIF-1α and HIF-2α occurred in the oviduct and anterior pituitary, respectively. Both HIF-1α and HIF-2α mRNA levels were higher in yaks than in cattle (p < 0.01). These data provide evidence that adaptation of reproduction to hypoxic conditions is associated with a greater expression of HIF-1α and HIF-2α in the reproductive axis of female yaks than cattle.
Asunto(s)
Adaptación Fisiológica/genética , Altitud , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Subunidad alfa del Factor 1 Inducible por Hipoxia , ARN Mensajero/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/análisis , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Bovinos , Femenino , Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia/análisis , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Especificidad de Órganos , Oviductos/química , Oviductos/metabolismo , Hipófisis/química , Hipófisis/metabolismo , ARN Mensajero/análisis , ARN Mensajero/genética , Estrés Fisiológico/genéticaRESUMEN
This study aimed at investigating the relationship between concentrations of macro and trace elements in blood serum, and fluids from small and large follicles (SFF and LFF, respectively), oviduct (OF), and uterus (UF) of female dromedary camels. Fluids from small (2-6 mm) and large follicles (7-20 mm), oviduct and uterus, and blood samples were collected from 19 camels. The results indicated that the concentrations of serum Mg, Fe, and Mn were significantly higher than their follicular fluid, OF, and UF concentrations. Levels of Zn, Fe, Cu, Cr, and Mn were significantly higher in SFF than in LFF. Se and Mo concentrations were higher in LFF. Co concentration was lower in serum than in reproductive tract fluids. Cr concentration was higher in UF and OF than in the serum, SFF, and LFF. High Ca concentration was observed for serum and SFF, followed by LFF. The concentration of Na was about 1.18-fold higher in SFF than in serum, OF, and LFF, and approximately 4.1-fold higher in serum than in UF. K was present in higher concentration in SFF than in serum and LFF; however, its concentration was low in UF and OF. In conclusion, this study shows the concentrations of certain elements in small and large follicular, uterine, and oviductal fluids, which may be low or high depending on their function in the development and growth of follicles. This information can support the development of new media for in vitro oocyte maturation and fertilization of female camels.
Asunto(s)
Camelus/metabolismo , Líquido Extracelular/química , Oviductos/química , Oligoelementos/metabolismo , Útero/química , Animales , Camelus/sangre , Femenino , Líquido Folicular/química , Oligoelementos/sangreRESUMEN
ADP-ribosylation factor 6 (Arf6) is a small GTPase that regulates endosomal trafficking and actin cytoskeleton remodeling. In the present study, we comprehensively examined the cellular and subcellular localization of Arf6 in adult mouse peripheral tissues by immunofluorescence and immunoelectron microscopy using the heat-induced antigen retrieval method with Tris-EDTA buffer (pH 9.0). Marked immunolabeling of Arf6 was observed particularly in epithelial cells of several tissues including the esophagus, stomach, small and large intestines, trachea, kidney, epididymis, oviduct, and uterus. In most epithelial cells of simple or pseudostratified epithelia, Arf6 exhibited predominant localization to the basolateral membrane and a subpopulation of endosomes. At an electron microscopic level, Arf6 was localized along the basolateral membrane, with dense accumulation at interdigitating processes and infoldings. Arf6 was present in a ring-like appearance at intercellular bridges in spermatogonia and spermatocytes in the testis and at the Flemming body of cytokinetic somatic cells in the ovarian follicle, thymus, and spleen. The present study provides anatomical clues to help understand the physiological roles of Arf6 at the whole animal level.
Asunto(s)
Factores de Ribosilacion-ADP/análisis , Epidídimo/química , Intestino Delgado/química , Riñón/química , Oviductos/química , Testículo/química , Factor 6 de Ribosilación del ADP , Animales , Reacciones Antígeno-Anticuerpo , Femenino , Técnica del Anticuerpo Fluorescente , Células HeLa , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Microscopía InmunoelectrónicaRESUMEN
Avian sperm are stored in the sperm storage tubules (SSTs) of the hen oviduct for a prolonged period. However, the precise mechanisms by which sperm are kept alive in the SSTs are still not fully understood. The aim of this study was to determine whether exosomes are secreted by SST cells and play a role in the survival of sperm. Utero-vaginal junction (UVJ) tissue from approximately 50 wk old White Leghorn hens was collected before (control group) and after intravaginal insemination with seminal plasma (SP group) or semen (AI group). The samples were used to prepare frozen sections and total protein extraction. The localization of the CD63, an exosome marker, was determined by immunohistochemistry and its protein level in the UVJ mucosal tissues was examined by Western blot. Exosomes were isolated from the culture media of UVJ and vaginal mucosa cells by ultracentrifugation and characterized by SDS-PAGE and Western blot. The viability and motility of sperm incubated with exosomes were also examined. CD63 was localized in the apical region of UVJ mucosal epithelium cells and SST cells of control, SP, and AI groups. The CD63 protein decreased in SST cells surrounding resident sperm and tended to appear in the SST lumen in the AI group. The protein level of CD63 in UVJ mucosal tissues was significantly higher in the AI group than control. The CD63 protein (approximately 75 kDa) was detected in ultracentrifugation pellets from the culture medium of UVJ and vagina cells. The viability of sperm incubated with 1 µg/µl vaginal exosomes was significantly decreased but was not affected by UVJ exosomes. These results suggest that exosomes were synthesized by SST cells and may be secreted into SST lumen when sperm were stored in SSTs. The role of SST exosomes in sperm storage needs to be examined further.
Asunto(s)
Pollos , Exosomas/fisiología , Oviductos/ultraestructura , Espermatozoides/fisiología , Tetraspanina 30/análisis , Animales , Western Blotting , Supervivencia Celular , Exosomas/química , Femenino , Inmunohistoquímica , Inseminación Artificial/métodos , Inseminación Artificial/veterinaria , Masculino , Oviductos/químicaRESUMEN
The aim of the present study was to investigate the expression and localization of B-cell lymphoma-2 (Bcl-2) in the oviduct of the Chinese soft-shelled turtle, Pelodiscus sinensis, during the reproductive cycle to analyze the relationship between Bcl-2 and sperm storage. Bcl-2 expression was confirmed in the P. sinensis oviduct by western blot analysis. Hematoxylin-eosin staining showed that female P. sinensis stored sperm from November to April of the following year. The oviduct showed positive immunostaining for Bcl-2 of epithelial ciliated cells, gland ducts, and gland cells. Bcl-2 expression in the oviduct was associated with sperm storage occurrence. This indicates that the survival factor Bcl-2 may play a role in P. sinensis sperm storage.
Asunto(s)
Oviductos/anatomía & histología , Oviductos/química , Proteínas Proto-Oncogénicas c-bcl-2/análisis , Reproducción , Espermatozoides/química , Animales , Femenino , Masculino , Oviductos/citología , Espermatozoides/citología , Tortugas , Útero/anatomía & histología , Útero/química , Útero/citologíaRESUMEN
Apolipoprotein D (APOD) is a glycoprotein which is widely expressed in mammalian tissues. It is structurally and functionally similar to the lipocalins which are multiple lipid-binding proteins that transport hydrophobic ligands and other small hydrophobic molecules, including cholesterol and several steroid hormones. Although multiple functions for APOD in various tissues have been reported, its expression, biological function, and hormonal regulation in the female reproductive system are not known. Thus, in this study, we focused on correlations between APOD and estrogen during development, differentiation, regression, and regeneration of the oviduct in chickens and in the development of ovarian carcinogenesis in laying hens. Results of the present study indicated that APOD messenger RNA (mRNA) expression increased (P < 0.001) in the luminal and glandular (GE) epithelia of the chicken oviduct in response to diethylstilbestrol (a nonsteroidal synthetic estrogen). In addition, the expression of APOD mRNA and protein decreased (P < 0.001) as the oviduct regressed during induced molting, and gradually increased (P < 0.001) with abundant expression in GE of the oviduct during recrudescence after molting. Furthermore, APOD mRNA and protein were predominantly localized in GE of cancerous, but not normal ovaries from laying hens. Collectively, results of the present study suggest that APOD is a novel estrogen-stimulated gene in the chicken oviduct which likely regulates growth, differentiation, and remodeling of the oviduct during oviposition cycles. Moreover, up-regulated expression of APOD in epithelial cell-derived ovarian cancerous tissue suggests that it could be a candidate biomarker for early detection and treatment of ovarian cancer in laying hens and in women.
Asunto(s)
Apolipoproteínas D/genética , Pollos , Dietilestilbestrol/farmacología , Neoplasias Ováricas/veterinaria , Oviductos/fisiopatología , Enfermedades de las Aves de Corral/fisiopatología , Animales , Apolipoproteínas D/análisis , Apolipoproteínas D/fisiología , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Muda/fisiología , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/fisiopatología , Ovario/química , Oviductos/química , Oviductos/crecimiento & desarrollo , Oviposición/fisiología , ARN Mensajero/análisisRESUMEN
Systems for maintaining the viability of ejaculated sperm in the female reproductive tract are widespread among vertebrates and invertebrates. In birds, this sperm storage function is performed by specialized simple tubular invaginations called sperm storage tubules (SSTs) in the uterovaginal junction (UVJ) of the oviduct. Although the incidence and physiological reasons for sperm storage in birds have been reported extensively, the mechanisms of sperm uptake by the SSTs, sperm maintenance within the SSTs, and control of sperm release from the SSTs are poorly understood. In this study, we demonstrated that the highly conserved heat shock protein 70 (HSP70) stimulates sperm motility in vitro and also that HSP70 expressed in the UVJ may facilitate the migration of sperm released from the SSTs. Quantitative RT-PCR analysis demonstrated that the expression of HSP70 mRNA in the UVJ increases before ovulation/oviposition. Gene-specific in situ hybridization and immunohistochemical analysis with a specific antibody to HSP70 demonstrated that HSP70 is localized in the surface epithelium of the UVJ. Furthermore, injection of anti-HSP70 antibody into the vagina significantly inhibited fertilization in vivo. In addition, we found that recombinant HSP70 activates flagellar movement in the sperm and that the binding of recombinant HSP70 to the sperm surface is mediated through an interaction with voltage-dependent anion channel protein 2 (VDAC2). Our results suggest that HSP70 binds to the sperm surface by interacting with VDAC2 and activating sperm motility. This binding appears to play an important role in sperm migration within the oviduct.
Asunto(s)
Coturnix , Proteínas HSP70 de Choque Térmico/farmacología , Oviductos/fisiología , Transporte Espermático/fisiología , Espermatozoides/efectos de los fármacos , Espermatozoides/fisiología , Animales , Anticuerpos/administración & dosificación , Femenino , Fertilización/efectos de los fármacos , Fertilización In Vitro/efectos de los fármacos , Expresión Génica , Proteínas HSP70 de Choque Térmico/genética , Proteínas HSP70 de Choque Térmico/inmunología , Masculino , Oviductos/química , Oviposición , Ovulación , ARN Mensajero/análisis , Motilidad Espermática/efectos de los fármacos , Espermatozoides/química , Útero/efectos de los fármacos , Canal Aniónico 2 Dependiente del Voltaje/fisiologíaRESUMEN
Although viviparity (live birth) has evolved from oviparity (egg laying) at least 140 times in vertebrates, nearly 120 of these independent events occurred within a single reptile taxon. Surprisingly, only squamate reptiles (lizards and snakes) are capable of facilitating embryonic development to increasingly advanced stages inside the mother during extended periods of oviducal egg retention. Viviparity has never evolved in turtle lineages, presumably because embryos enter and remain in an arrested state until after eggs are laid, regardless of the duration of egg retention. Until now, the limiting factor that initiates and maintains developmental arrest has remained elusive. Here, we show that oviducal hypoxia arrests embryonic development. We demonstrate that hypoxia can maintain developmental arrest after oviposition and that subsequent exposure of arrested embryos to normoxia triggers resumption of their development. We discovered remarkably low oxygen partial pressure in the oviducts of gravid turtles and found that secretions produced by the oviduct retard oxygen diffusion. Our results suggest that an extremely hypoxic environment in the oviduct arrests embryonic development and may constrain the evolution of viviparity in turtles, with the reduced diffusive capacity of oviducal secretions possibly creating or contributing to this hypoxia. We anticipate that these findings will allow us to better understand the mechanisms underlying the evolutionary transition between reproductive modes.
Asunto(s)
Evolución Biológica , Hipoxia de la Célula/fisiología , Desarrollo Embrionario/fisiología , Óvulo/química , Tortugas/embriología , Viviparidad de Animales no Mamíferos/fisiología , Análisis de Varianza , Animales , Australia , Difusión , Desarrollo Embrionario/efectos de los fármacos , Femenino , Oviductos/química , Oxígeno/análisis , Oxígeno/farmacología , Presión Parcial , Especificidad de la EspecieRESUMEN
Tight junctions in the mucosal epithelium have essential roles as a mucosal barrier to prevent invasion of microbes into the hen oviduct tissue. The aim of this study was to determine the effects of the egg-laying phase and estradiol on the expression of tight junction molecule "claudins" in the lower oviductal segments in hens. White Leghorn laying and molting hens were used. Molting hens were given either sesame oil (vehicle) or estradiol benzoate (N = 5 per group) via injection. The lower segments of oviduct (isthmus, uterus, and vagina) of these birds were collected. Gene expression of claudin-1, -3, -5, lipopolysaccharide-induced TNFα factor (LITAF), and IFN(γ) was analyzed by quantitative reverse transcription polymerase chain reaction, and localization of claudin-1 was examined by immunohistochemistry. Permeability in the mucosal epithelium was assessed by intrauterine injection of fluorescein isothiocyanate-dextran. Expression of claudin-1, -3, and -5 genes and density of claudin-1 protein in the lower oviductal segments were higher in laying hens than in molting hens (P < 0.01); their expression was upregulated by estradiol (P < 0.01). Expression of LITAF and IFN(γ) genes was higher in molting hens than in laying hens. More fluorescein isothiocyanate-dextran infiltrated into the intercellular space of the uterus mucosal epithelium in molting hens than in laying hens and estradiol-treated molting hens. In conclusion, we inferred that barrier functions of the mucosal epithelium in the lower oviductal segments might be disrupted because of reduced claudin expression in molting hens, which might increase the susceptibility of mucosal tissue during the molting phase.
Asunto(s)
Pollos/fisiología , Claudinas/genética , Estradiol/análogos & derivados , Expresión Génica/efectos de los fármacos , Oviductos/metabolismo , Oviposición/fisiología , Animales , Claudina-1/análisis , Claudina-1/genética , Claudina-3/genética , Claudina-5/genética , Claudinas/análisis , Epitelio/fisiología , Estradiol/farmacología , Femenino , Muda/fisiología , Membrana Mucosa/fisiología , Oviductos/química , Oviductos/efectos de los fármacos , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Archegozetes longisetosus is a parthenogenetic oribatid mite and a chelicerate model organism. We examined the localisation of processes between vitellogenesis and embryogenesis as well as the anatomy and histology of involved structures by means of light- and electron microscopy. The proximal oviduct is differentiated into an oviductal bulb, exhibiting a strong secretory epithelium. Here, solidification of the egg shell instantaneously occurs upon passing of the egg from the perivitelline space into the oviductal lumen. This is interpreted as an internalised oviposition with the generation boundary being effectively located at the ovary-oviduct transition, rendering the oviducts into functional brood chambers. The parity mode combines elements of oviparity and ovolarviparity with facultative egg retention.
Asunto(s)
Oviductos/metabolismo , Partenogénesis , Ácaros y Garrapatas/metabolismo , Ácaros y Garrapatas/ultraestructura , Animales , Femenino , Microscopía Electrónica , Oviductos/química , Oviductos/ultraestructura , VitelogénesisRESUMEN
The control of Newcastle disease (ND) in South Africa has proved difficult since 2002 following the introduction of lineage 5d/VIId Newcastle disease virus (NDV) strain ("goose paramyxovirus" - GPMV) to which commercially available ND vaccines appeared less effective. Most of the ND infections, even in fully vaccinated hens were characterized consistently by a drop in egg production. In this study, commercial and SPF hens-in-lay were vaccinated with La Sota vaccine and challenged with a GPMV isolate. Immunohistochemical labeling was used to determine the distribution of viral antigen in the oviduct of the hens. Following reports that cloacal vaccination offered better protection against egg production losses than the oro-nasal route, the efficacy of cloacal and ocular routes of vaccination against challenge were compared. Results showed that La Sota vaccine offered birds 100% protection against the virulent ND (GPMV) virus challenge from clinical disease and death, but not against infection and replication of the GPMV, as birds showed varying degrees of macropathology. Histopathology of the oviduct of infected birds revealed multifocal lymphocytic inflammation in the interstitium as well as mild glandular ectasia and mild edema. Finely granular NDV-specific immunolabeling was demonstrated in the cytoplasm of epithelial cells and mononuclear (lymphohistiocytic) cells in the interstitium of the oviduct. Both vaccine and virulent GPMV showed greatest tropism for the uterus (versus the magnum and isthmus). There was no clear difference in the protection of the oviduct and in the distribution of oviductal GPMV antigens between the two routes of vaccination.
Asunto(s)
Antígenos Virales/análisis , Enfermedad de Newcastle/prevención & control , Virus de la Enfermedad de Newcastle , Oviductos/virología , Vacunas Virales/uso terapéutico , Animales , Pollos/virología , Ensayo de Inmunoadsorción Enzimática/veterinaria , Femenino , Enfermedad de Newcastle/inmunología , Enfermedad de Newcastle/patología , Enfermedad de Newcastle/virología , Virus de la Enfermedad de Newcastle/inmunología , Oviductos/química , Oviductos/patología , Vacunas Virales/administración & dosificación , Vacunas Virales/inmunologíaRESUMEN
Ovarian cancer is the fifth leading cause of cancer deaths in women and has a 63% mortality rate in the United States(1). The cell type of origin for ovarian cancers is still in question and might be either the ovarian surface epithelium (OSE) or the distal epithelium of the fallopian tube fimbriae(2,3). Culturing the normal cells as a primary culture in vitro will enable scientists to model specific changes that might lead to ovarian cancer in the distinct epithelium, thereby definitively determining the cell type of origin. This will allow development of more accurate biomarkers, animal models with tissue-specific gene changes, and better prevention strategies targeted to this disease. Maintaining normal cells in alginate hydrogels promotes short term in vitro culture of cells in their three-dimensional context and permits introduction of plasmid DNA, siRNA, and small molecules. By culturing organs in pieces that are derived from strategic cuts using a scalpel, several cultures from a single organ can be generated, increasing the number of experiments from a single animal. These cuts model aspects of ovulation leading to proliferation of the OSE, which is associated with ovarian cancer formation. Cell types such as the OSE that do not grow well on plastic surfaces can be cultured using this method and facilitate investigation into normal cellular processes or the earliest events in cancer formation(4). Alginate hydrogels can be used to support the growth of many types of tissues(5). Alginate is a linear polysaccharide composed of repeating units of ß-D-mannuronic acid and α-L-guluronic acid that can be crosslinked with calcium ions, resulting in a gentle gelling action that does not damage tissues(6,7). Like other three-dimensional cell culture matrices such as Matrigel, alginate provides mechanical support for tissues; however, proteins are not reactive with the alginate matrix, and therefore alginate functions as a synthetic extracellular matrix that does not initiate cell signaling(5). The alginate hydrogel floats in standard cell culture medium and supports the architecture of the tissue growth in vitro. A method is presented for the preparation, separation, and embedding of ovarian and oviductal organ pieces into alginate hydrogels, which can be maintained in culture for up to two weeks. The enzymatic release of cells for analysis of proteins and RNA samples from the organ culture is also described. Finally, the growth of primary cell types is possible without genetic immortalization from mice and permits investigators to use knockout and transgenic mice.
Asunto(s)
Alginatos/química , Hidrogeles/química , Técnicas de Cultivo de Órganos/métodos , Ovario/anatomía & histología , Oviductos/anatomía & histología , Animales , Femenino , Ratones , Ovario/química , Ovario/ultraestructura , Oviductos/química , Oviductos/ultraestructuraRESUMEN
A variety of evolutionary processes has led to the development of different organs to ensure that internal fertilization occur successfully. Fallopian tubes are a particularly interesting example of such organs. Some of the key events during fertilization and early embryo development occur in the oviduct. Knowledge of the different components described in the oviduct is extensive. Oviductal components include hormones, growth factors and their receptors that have important roles in the physiology of the oviduct and embryo development. Other oviductal factors protect the gamete and the embryos against oxidative stress and pathogens. Different proteins and enzymes are present in the oviductal fluid and have the ability to interact with the oocyte and the sperm before the fertilization occurs. Of special interest is the oviduct-specific glycoprotein (OVGP1), a glycoprotein that is conserved in different mammals, and its association with the zona pellucida (ZP). Interaction of the oocyte with oviductal secretions leads us to emphasize the concept of 'ZP maturation' within the oviduct. The ZP changes produced in the oviduct result in an increased efficiency of the in vitro fertilization technique in some animal models, contributing in particular to the control of polyspermy and suggesting that a similar role could be played by oviductal factors in human beings. Finally, attention should be given to the presence in the oviductal fluid of several embryotrophic factors and their importance in relation to the in vivo versus in vitro developmental ability of the embryos.
Asunto(s)
Trompas Uterinas/metabolismo , Oviductos/metabolismo , Técnicas Reproductivas Asistidas , Secuencia de Aminoácidos , Animales , Técnicas de Cultivo de Embriones , Desarrollo Embrionario/fisiología , Trompas Uterinas/química , Femenino , Glicoproteínas/química , Glicoproteínas/genética , Glicoproteínas/fisiología , Humanos , Datos de Secuencia Molecular , Oocitos/crecimiento & desarrollo , Oviductos/química , Alineación de Secuencia , Especificidad de la Especie , Interacciones Espermatozoide-Óvulo , Zona Pelúcida/fisiologíaRESUMEN
To investigate the physiological roles of nerve growth factor (NGF) in the oviduct of golden hamsters during the estrous cycle, the localization of NGF and its receptors, trkA and p75, were determined by immunohistochemistry. Positive staining of NGF, trkA, and p75 was present in epithelial cells and muscle cells of the infundibulum, ampulla, and isthmus in the oviduct. The intensities of the immunohistochemical signals for NGF, trkA, and p75 did not markedly change in any segment of the oviduct during the estrous cycle. These results suggest that NGF may play autocrine/paracrine roles in oviductal transport, fertilization, capacitation of spermatozoa and early embryonic development in the oviduct of golden hamsters.
Asunto(s)
Ciclo Estral/metabolismo , Mesocricetus/fisiología , Factor de Crecimiento Nervioso/metabolismo , Oviductos/metabolismo , Receptor de Factor de Crecimiento Nervioso/metabolismo , Receptor trkA/metabolismo , Animales , Biomarcadores/análisis , Biomarcadores/metabolismo , Cricetinae , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Factor de Crecimiento Nervioso/análisis , Oviductos/química , Receptor de Factor de Crecimiento Nervioso/análisis , Receptor trkA/análisisRESUMEN
Mouse sperm-egg binding requires a multiplicity of receptor-ligand interactions, including an oviduct-derived, high molecular weight, wheat germ agglutinin (WGA)-binding glycoprotein that associates with the egg coat at ovulation. Herein, we report the purification and identification of this sperm-binding ligand. WGA-binding, high molecular weight glycoproteins isolated from hormonally primed mouse oviduct lysates competitively inhibit sperm-egg binding in vitro. Within this heterogeneous glycoprotein preparation, a distinct 220 kDa protein selectively binds to sperm surfaces, and was identified by sequence analysis as oviduct-specific glycoprotein (OGP). The sperm-binding activity of OGP was confirmed by the loss of sperm-binding following immunodepletion of OGP from oviduct lysates, and by the ability of both immunoprecipitated OGP and natively purified OGP to competitively inhibit sperm-egg binding. As expected, OGP is expressed by the secretory cells of the fimbriae and infundibulum; however, in contrast to previous reports, OGP is also associated with both the zona pellucida and the perivitelline space of mouse oocytes. Western blot analysis and lectin affinity chromatography demonstrate that whereas the bulk of OGP remains soluble in the ampullar fluid, distinct glycoforms associate with the cumulus matrix, zona pellucida and perivitelline space. The sperm-binding activity of OGP is carbohydrate-dependent and restricted to a relatively minor peanut agglutinin (PNA)-binding glycoform that preferentially associates with the sperm surface, zona pellucida and perivitelline space, relative to other more abundant glycoforms. Finally, pretreatment of two-cell embryos, which do not normally bind sperm, with PNA-binding OGP stimulates sperm binding.
Asunto(s)
Proteínas del Huevo/metabolismo , Glicoproteínas/metabolismo , Glicoproteínas de Membrana/metabolismo , Oviductos/metabolismo , Óvulo/metabolismo , Receptores de Superficie Celular/metabolismo , Espermatozoides/metabolismo , Secuencia de Aminoácidos , Animales , Proteínas del Huevo/genética , Femenino , Glicoproteínas/química , Glicoproteínas/aislamiento & purificación , Masculino , Glicoproteínas de Membrana/genética , Ratones , Datos de Secuencia Molecular , Peso Molecular , Oviductos/química , Unión Proteica , Receptores de Superficie Celular/genética , Glicoproteínas de la Zona PelúcidaRESUMEN
Polyspermy is an important anomaly of fertilization in placental mammals, causing premature death of the embryo. It is especially frequent under in vitro conditions, complicating the successful generation of viable embryos. A block to polyspermy develops as a result of changes after sperm entry (i.e., cortical granule exocytosis). However, additional factors may play an important role in regulating polyspermy by acting on gametes before sperm-oocyte interaction. Most studies have used rodents as models, but ungulates may differ in mechanisms preventing polyspermy. We hypothesize that zona pellucida (ZP) changes during transit of the oocyte along the oviductal ampulla modulate the interaction with spermatozoa, contributing to the regulation of polyspermy. We report here that periovulatory oviductal fluid (OF) from sows and heifers increases (both, con- and heterospecifically) ZP resistance to digestion with pronase (a parameter commonly used to measure the block to polyspermy), changing from digestion times of approximately 1 min (pig) or 2 min (cattle) to 45 min (pig) or several hours (cattle). Exposure of oocytes to OF increases monospermy after in vitro fertilization in both species, and in pigs, sperm-ZP binding decreases. The resistance of OF-exposed oocytes to pronase was abolished by exposure to heparin-depleted medium; in a medium with heparin it was not altered. Proteomic analysis of the content released in the heparin-depleted medium after removal of OF-exposed oocytes allowed the isolation and identification of oviduct-specific glycoprotein. Thus, an oviduct-specific glycoprotein-heparin protein complex seems to be responsible for ZP changes in the oviduct before fertilization, affecting sperm binding and contributing to the regulation of polyspermy.
Asunto(s)
Fertilización , Glicoproteínas/fisiología , Heparina/fisiología , Oviductos/química , Espermatozoides/metabolismo , Zona Pelúcida/metabolismo , Animales , Bovinos , Femenino , Masculino , Pronasa , Unión Proteica , Interacciones Espermatozoide-Óvulo , Espermatozoides/fisiología , Porcinos , Zona Pelúcida/fisiologíaRESUMEN
[(125)I]mesotocin (MT) binding of membrane fractions of the oviduct uterus in laying hens and nonlaying hens was measured by the use of radioligand binding assay to elucidate the presence of MT receptor in the uterine tissue, and whether the binding to the receptor changes before and after oviposition. The uterine tissue of the hen was found to contain a specific [(125)I]MT binding component having properties of MT receptor (i.e., binding specificity, saturable binding, high affinity, and limited capacity). The equilibrium dissociation constant (K(d)) was 0.46 +/- 0.05 nM (x +/- SEM; n = 4) in laying hens holding a hard-shelled egg in the uterus (shell gland) and 0.78 +/- 0.02 nM (n = 4) in nonlaying hens. The maximum binding capacity (B(max)) was 0.28 +/- 0.03 pmol/mg of protein (n = 4) in laying hens and 0.19 +/- 0.01 pmol/mg of protein (n = 4) in nonlaying hens. The K(d) value of the laying hens varied from 0.37 to 0.91 nM during an oviposition cycle showing a decrease 30 min before oviposition and an increase 4 h after oviposition. The B(max) value also varied from 0.15 to 0.38 pmol/mg of protein showing an increase 3 h before oviposition, a decrease 30 min before oviposition, and an increase 4 h after oviposition. In the nonlaying hen, both values were almost constant during a 24-h day. The changes in the binding affinity and capacity of MT receptor of the uterus may be related to oviposition in the hen.
Asunto(s)
Pollos/metabolismo , Oviductos/metabolismo , Oviposición/fisiología , Receptores de la Hormona Hipofisaria/metabolismo , Animales , Unión Competitiva , Relación Dosis-Respuesta a Droga , Femenino , Radioisótopos de Yodo , Cinética , Oviductos/químicaRESUMEN
Our previous studies demonstrated that dietary daidzein improves egg production in ducks during the late period of the laying cycle. The present study was aimed to investigate the effect of daidzein in laying hens, with more focus on eggshell quality. The expression of ER-alpha, GH-R, and IGF-IR mRNA in shell glands was determined to identify the target genes of daidzein action and to reveal the relationship between shell quality and profiles of gene expression in shell glands of laying hens. 1000 ISA hens, at 445 days of age, were allotted at random to two groups and given the basal diet with or without 10 mg of daidzein per kg diet for 9 weeks. Daidzein supplement significantly increased the egg laying rate and the feed conversion ratio. The eggshell thickness increased, while the percentage of cracked eggs decreased in daidzein-treated hens. Serum E2 and phosphate concentrations were not altered, but the level of serum Ca2+ and the tibia bone mineral density were significantly increased in the daidzein-treated group compared with their control counterparts. In parallel with the significant increase of oviduct weight, significant down-regulation of GH-R and IGF-IR mRNA and a trend of decrease in ERalpha mRNA expression in shell glands were observed in daidzein-treated hens. The results indicate that dietary daidzein improves egg laying performance and eggshell quality during the late (postpeak) laying stage of hens, which is associated with modulations in gene expression in the shell gland.
Asunto(s)
Pollos/fisiología , Dieta , Cáscara de Huevo/efectos de los fármacos , Isoflavonas/farmacología , Oviductos/efectos de los fármacos , Oviparidad/efectos de los fármacos , Animales , Densidad Ósea/efectos de los fármacos , Cáscara de Huevo/fisiología , Receptor alfa de Estrógeno/genética , Femenino , Expresión Génica/efectos de los fármacos , Tamaño de los Órganos/efectos de los fármacos , Oviductos/anatomía & histología , Oviductos/química , ARN Mensajero/análisis , Receptor IGF Tipo 1/genética , Receptores de Somatotropina/genéticaRESUMEN
The reproductive remodeling during molting appears to be a complex physiological mechanism regulated by multiple host factors. Lipopolysaccharide (LPS) induced TNF-alpha factor (LITAF) is one of the transcription factors controlling the expression of TNF-alpha and other cytokines. In the present investigation, we studied the involvement of LITAF in the regression of reproductive tissues of molting birds. Semi-quantitative RT-PCR analysis revealed that LITAF mRNA was generally expressed in both ovary and oviduct. In the molting birds, i.e. those subjected to feed withdrawal (FW) or fed high levels of zinc (ZnF) birds, the LITAF expression was upregulated significantly in the ovary after 4 days of molting (DOM). However, LITAF mRNA levels were three-fold higher in ZnF birds, which might be responsible for a greater degree of follicular atresia. In the oviduct of FW birds, peak LITAF expression was noticed on 4DOM and the levels remained significantly higher until the end of the experiment. In ZnF birds, LITAF expression reached its peak on 1DOM and subsequently downregulated to basal levels on 2DOM. This indicated that constantly higher LITAF expression might be required for complete regression of the oviduct during molting. In conclusion, LITAF might be one of the major transcription factors controlling reproductive regression in chicken, as the expression levels were associated with the regression pattern.
Asunto(s)
Pollos/fisiología , Lipopolisacáridos/farmacología , Muda/fisiología , Ovario/química , Oviductos/química , Factor de Necrosis Tumoral alfa/genética , Animales , Femenino , Privación de Alimentos , Expresión Génica , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Zinc/administración & dosificaciónRESUMEN
OBJECTIVE: To compare eighteen samples of Forest frog's oviduct from different regions of northeast China, in order to fomulate the quality evaluation standard. METHODS: According to the documents, comparing the target constituent of Forest frog's oviduct, including the mositure, ash, protein, lipid and expansibility were analysed. RESULTS: It was similar to the chemical constituent in Forest frog's oviduct from different habitiat of northeast China. CONCLUSION: The germplasm of this species is stable.