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ABSTRACT: Abstract. The aim of this study has been to determine the effects of in vivo post-ovulatory ageing (POA) on the distribution of spindle-associated proteins, histone H3/H4 post-translational modifications and v-akt murine thymoma viral oncogene homolog 1 (Akt) expression levels. To this end, oocytes were retrieved 13, 29 and 33 h after human
chorionic gonadotrophin (hCG) treatment. The presence and distribution at the meiotic spindle of acetylated tubulin, 5 g-tubulin, polo kinase-1 and Ser473/Thr308 phosphorylated Akt (pAkt) as well as histone H3 and H4 acetylation and phosphorylation levels were assayed via immunofluorescence. Akt expression levels were determined via reverse
transcription–polymerase chain reaction and western blotting analyses. Spindles from oocytes recovered 13 h and 29 h after hCG treatment showed similar levels of acetylated tubulin but ageing induced: (1) translocation of g-tubulin from spindle poles to microtubules, (2) absence of Thr308- and Ser473-pAkt in 76% and 30% of oocytes, respectively and (3) a 10 significant reduction in phosphorylation levels of serine 10 on histone 3. At 29 h, a significant decrease in Akt mRNA, but not in pAkt or Akt protein expression levels, was recorded. By contrast, protein content significantly decreased 33 h after hCG. We conclude that POA impairs oocyte viability and fertilisability by altering the expression levels and spindle distribution of proteins that are implicated in cell survival and chromosome segregation. Together, these events could play a role in oocyte apoptosis.
Reproduction Fertility and Development 04/2013; · 2.11 Impact Factor
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ABSTRACT: Thyroxine (T4) plus gonadotropins might stimulate ovarian follicular angiogenesis in immature infertile hypothyroid rdw rats by upregulating mRNA expression of major angiogenic factors. Development of growing corpus luteum (CL) is strongly related to angiogenesis and to morphofunctional development of microcirculation. Our aim was to investigate if T4 is involved in CL angiogenesis and in the activation of capillary cells and angiogenic factors after ovulation in a spontaneous model of hypothyroidism, the rdw rat. Rdw rats were treated with T4 plus gonadotropins (equine chorionic gonadotropin plus human chorionic gonadotropin; eCG+hCG) or gonadotropins alone in order to evaluate the effects of T4 on early luteal angiogenesis, on microvascular cells and on expression of major growth factors which are involved in the regulation of angiogenesis. Wistar-Imamichi rats treated with gonadotropins were used as controls. The ovaries were collected 4 days after hCG administration and analyzed using morphologic and molecular approaches. Thyroxine plus gonadotropins stimulated the growth of CLs and follicles as in controls, differently from rdw rats treated only with gonadotropins, in which CLs were not found and only small follicles, often atretic, could be recognized. In T4 plus gonadotropin-treated rdw rats CLs showed increased microvasculature, numerous activated capillaries characterized by sprouting and other angiogenic figures, and associated pericytes. Quantitative analysis revealed that the number of pericytes in T4 plus gonadotropin-treated rdw rats was comparable with that found in control rats and was significantly higher than that found in gonadotropin-treated rdw rats. The mRNA expression of vascular endothelial growth factor and basic fibroblast growth factor was significantly higher in control rats and in T4 plus gonadotropin-treated rdw rats than in gonadotropin-treated rdw rats. mRNA expression of tumor necrosis factor α, transforming growth factor β, and epidermal growth factor did not show significant changes. Our data originally demonstrated that T4 promoted the growth of an active microcirculation in developing CLs of gonadotropin-primed hypothyroid rdw rats, mainly by inducing sprouting angiogenesis, pericyte recruitment, and upregulation of mRNA expression of vascular endothelial growth factor and basic fibroblast growth factor. In conclusion, we suggest that T4 plays a key role in restoring luteal angiogenesis in ovaries of immature hypothyroid rdw rats.
Theriogenology 11/2012; · 1.96 Impact Factor
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ABSTRACT: The heart is composed by a specialized muscle, whose form and function are essentials for an adequate work and shows an amount of connective tissue which support and provide insertion for this muscle, whose collagen fibers are responsible for determination of tissue feature. Our objective was to identify the structural arrangement of the heart collagen fibers in dogs. The hearts of the dogs were submitted to the process of the controlled digestion with NaOH solution and observed by scanning electron microscope. Our results showed that the collagen fibers of the endomysial wall have structural arrangement composed by an irregular network with one layer in normal dogs but in diabetic dogs the network acquires a greater amount of the fibers and layers, looking like a "rug" of fibers modifying the relationships of the stress/strain of the tissue. Ahead of the observed results we are able to conclude that exist increase in the amount and thickness of cardiac collagen fibers, beyond the increase of layers and architectural disarrangement in the endomysial wall in the diabetic dogs.
Microscopy Research and Technique 04/2011; 74(11):1018-23. · 1.79 Impact Factor
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ABSTRACT: In vitro maturation of ovarian follicles, in combination with cryopreservation, might be a valuable method for preserving and/or restoring fertility in mammals with impaired reproductive function. Several culture systems capable of sustaining mammalian follicle growth in vitro have been developed and many studies exist on factors influencing the development of in vitro grown oocytes. However, a very few reports concern the ultrastructural morphology of in vitro grown follicles.
The present study was designed to evaluate, by transmission and scanning electron microscopy, the ultrastructural features of isolated mouse preantral follicles cultured in vitro for 6 days in a standard medium containing fetal calf serum (FCS). The culture was supplemented or not with FSH.
The follicles cultured in FCS alone, without FSH supplementation (FCS follicles), did not form the antral cavity. They displayed low differentiation (juxta-nuclear aggregates of organelles in the ooplasm, a variable amount of microvilli on the oolemma, numerous granulosa cell-oolemma contacts, signs of degeneration in granulosa cell compartment). Eighty (80)% of FSH-treated follicles formed the antral cavity (FSH antral follicles). These follicles showed various ultrastructural markers of maturity (spreading of organelles in ooplasm, abundant microvilli on the oolemma, scarce granulosa cell-oolemma contacts, granulosa cell proliferation). Areas of detachment of the innermost granulosa cell layer from the oocyte were also found, along with a diffuse granulosa cell loosening compatible with the antral formation. Theca cells showed an immature morphology for the stage reached. Twenty (20)% of FSH-treated follicles did not develop the antral cavity (FSH non-antral follicles) and displayed morphological differentiation features intermediate between those shown by FCS and FSH antral follicles (spreading of organelles in the ooplasm, variable amount of microvilli, scattered granulosa cell-oolemma contacts, signs of degeneration in granulosa cell compartment).
It is concluded that FSH supports the in vitro growth of follicles, but the presence of a diffuse structural granulosa cell-oocyte uncoupling and the absence of theca development unveil the incomplete efficiency of the system. The present study contributes to explain, from a morphological point of view, the effects of culture conditions on the development of mouse in vitro grown follicles and to highlight the necessity of maintaining efficient intercellular communications to obtain large numbers of fully-grown mature germ cells.
Reproductive Biology and Endocrinology 01/2011; 9:3. · 2.05 Impact Factor
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ABSTRACT: To ascertain possible cell damage from cryopreservation, the ultrastructure of human oocytes cryopreserved by slow cooling was assessed.
Cryopreservation was performed through two protocols with one-step or two-step propanediol. Fresh control oocytes were examined for comparison. Samples were processed for transmission electron microscopy analysis.
By light microscopy, both fresh and frozen-thawed oocytes appeared regularly rounded, with intact zona pellucida, and homogeneous cytoplasm. By electron microscopy observation, organelles were abundant and uniformly dispersed. Mitochondria-smooth endoplasmic reticulum associations appeared regular. However, both the amount and density of cortical granules appeared abnormally reduced in frozen-thawed samples. Slight to moderate vacuolization was also found in the ooplasm of oocytes of both frozen groups.
Slow cooling ensures a good overall preservation of human oocytes. However, cytoplasmic vacuolization and cortical granule loss appears associated with cryopreservation, irrespective of the protocol used.
Journal of Assisted Reproduction and Genetics 02/2010; 27(4):131-40. · 1.84 Impact Factor
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ABSTRACT: In the mammalian ovary, follicular and corpus luteum cycle is associated with intensive microvascular remodelling. The complex angiogenic dynamics are finely tuned by numerous regulatory factors acting as activators (up-regulators) or inhibitors (down-regulators) of angiogenesis. Alterations of such a tight modulation are involved in several pathologies, including infertility, polycystic ovarian syndrome, ovarian hyperstimulation syndrome and ovarian cancer. We have demonstrated in several experimental models that ovarian function is critically and specifically dependent on angiogenesis for follicular development, ovulation, and corpus luteum growth. The aim of this review is to summarize the results we have obtained on the morphodynamic remodelling of ovarian microvascularization, in polyovulatory (rat, rabbit and pig) and monovulatory species (cow), using scanning electron microscopy of vascular corrosion casts. The knowledge of the morphological expression of the up- and down-regulation of angiogenesis occurring in mono and polyovulatory animals might provide useful information to preserve fertility and to increase of the effectiveness of reproductive management in species of domestic interest.
Italian journal of anatomy and embryology = Archivio italiano di anatomia ed embriologia 01/2010; 115(1-2):109-14.
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ABSTRACT: To improve developmental competence of in vitro matured oocytes, culture medium can be supplemented with hypoxanthine (Hx) and FSH or epidermal growth factor (EGF) to trigger the activation of essential signalling pathways regulating meiotic resumption and progression. Since the serine/threonine kinase, Akt, contributes to the regulation of the meiotic cell cycle, this study analysed its expression level and localization at the meiotic spindle in oocytes matured in vivo or in vitro in the presence of Hx-FSH or Hx-EGF. Independently of culture conditions adopted, Akt mRNA concentration did not vary from germinal vesicle to metaphase I (MI), while at MII a significant decrease in Akt1 mRNA concentration was recorded in oocytes matured in vivo and in those stimulated by Hx-EGF (P < 0.05). Phoshorylated Akt protein content was similar in the different groups of MI oocytes, but it decreased at MII in oocytes matured either in vivo or in vitro with Hx-EGF. Ser-473-phosphorylated Akt was localized uniformly to the meiotic spindle in more than 90% of oocytes. These results indicate that, in mouse oocytes, Akt expression is differentially regulated during in vivo and in vitro maturation and suggest that EGF could be a positive modulator, even stronger than FSH, of oocyte meiotic maturation.
Reproductive biomedicine online 01/2010; 20(1):35-41. · 2.04 Impact Factor
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ABSTRACT: The present research aims to describe the process of vascular readjustment occurring in pig ovary during the periovulatory phase (from LH surge to ovulation) that drives the transformation of the follicle, a limited blood supplied structure, into the corpus luteum, a highly vascularised endocrine gland required to maintain high levels of progesterone in pregnancy. The swine model was chosen because it is characterized by a long periovulatory window (about 40-44 hrs-similar to human) that permits to recover follicles at a precise endocrinological timing.
By validated hormonal protocol (eCG+hCG), able to mimic the physiologic gonadotropin stimulation, preovulatory follicles (PreOFs, 60 h-eCG), follicles in the middle (early periovulatory follicles, EPerOFs, 18 h-hCG) or late (LPerOFs, 36 h-hCG) periovulatory phase were isolated from prepubertal gilts. To understand the angiogenic process, morphological/morphometrical analyses were performed by combining immunohistochemistry (IHC) and SEM of vascular corrosion casts (VCC) techniques.
PreOFs showed a vascular plexus with proliferating endothelial cells (EPI). This plexus was characterized by a dense inner capillary network, with angiogenic figures, connected to the outer network by anastomotic vessels (arterioles and venules of the middle network). EPerOFs decreased their EPI, blood vessel extension in the outer network, and evidenced a reduced compactness of blood vessels. In LPerOFs, a rapid neovascularization was associated to an intensive tissue remodeling: the follicle acquired an undulated aspect presenting arterioles/venules near the basal membrane, increased vascular extension by EPI, sprouting and non-sprouting angiogenesis.The analysis of vascular geometric relations and branching angles evidenced similar values at all stages.
These data allow us to hypothesize that EPerOFs are in a quiescent status. LPerOFs represent the "metamorphic" follicles that rapidly turn-on angiogenesis to sustain a successful corpus luteum formation. Particularly, it is interesting to underlie that the non-sprouting angiogenesis, typical of structures in rapid neovascularization, occurred only in the LPerOFs. Moreover, vascular geometric relations showed as blood vessel remodeling occurs with the "maximum output and the minimum energetic expense".This knowledge will allow to better understand the mechanisms regulating the reproductive success and to clarify the complex physiological angiogenic process in adult tissues.
Reproductive Biology and Endocrinology 02/2009; 7:72. · 2.05 Impact Factor
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ABSTRACT: Abstract
Background
The present research aims to describe the process of vascular readjustment occurring in pig ovary during the periovulatory phase (from LH surge to ovulation) that drives the transformation of the follicle, a limited blood supplied structure, into the corpus luteum, a highly vascularised endocrine gland required to maintain high levels of progesterone in pregnancy. The swine model was chosen because it is characterized by a long periovulatory window (about 40–44 hrs-similar to human) that permits to recover follicles at a precise endocrinological timing.
Methods
By validated hormonal protocol (eCG+hCG), able to mimic the physiologic gonadotropin stimulation, preovulatory follicles (PreOFs, 60 h-eCG), follicles in the middle (early periovulatory follicles, EPerOFs, 18 h-hCG) or late (LPerOFs, 36 h-hCG) periovulatory phase were isolated from prepubertal gilts. To understand the angiogenic process, morphological/morphometrical analyses were performed by combining immunohistochemistry (IHC) and SEM of vascular corrosion casts (VCC) techniques.
Results
PreOFs showed a vascular plexus with proliferating endothelial cells (EPI). This plexus was characterized by a dense inner capillary network, with angiogenic figures, connected to the outer network by anastomotic vessels (arterioles and venules of the middle network). EPerOFs decreased their EPI, blood vessel extension in the outer network, and evidenced a reduced compactness of blood vessels. In LPerOFs, a rapid neovascularization was associated to an intensive tissue remodeling: the follicle acquired an undulated aspect presenting arterioles/venules near the basal membrane, increased vascular extension by EPI, sprouting and non-sprouting angiogenesis.
The analysis of vascular geometric relations and branching angles evidenced similar values at all stages.
Conclusion
These data allow us to hypothesize that EPerOFs are in a quiescent status. LPerOFs represent the "metamorphic" follicles that rapidly turn-on angiogenesis to sustain a successful corpus luteum formation. Particularly, it is interesting to underlie that the non-sprouting angiogenesis, typical of structures in rapid neovascularization, occurred only in the LPerOFs. Moreover, vascular geometric relations showed as blood vessel remodeling occurs with the "maximum output and the minimum energetic expense".
This knowledge will allow to better understand the mechanisms regulating the reproductive success and to clarify the complex physiological angiogenic process in adult tissues.
Reproductive Biology and Endocrinology. 01/2009;
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ABSTRACT: The development of mature ovarian follicles is greatly dependent on healthy thecal angiogenesis. Recent experimental evidence showed that thyroxine (T4) treatment promoted ovarian follicle development in immature hypothyroid (rdw) rats. However, an involvement of thyroid hormone in ovarian follicular angiogenesis has not yet been demonstrated. By morphological and molecular approaches, the present studies demonstrated that antral follicles in untreated, T4- or equine chorionic gonadotropin (eCG)-treated rdw rats were mainly small and/or atretic, and presented a poorly developed thecal microvasculature with ultrastructural evidence of diffuse quiescent or degenerative thin capillaries. However, T4 together with eCG increased the number of large antral and mature follicles with numerous activated capillaries and ultra-structural evidence of rich and diffuse angiogenesis in the theca layer. While T4 alone significantly increased mRNA expression of vascular endothelial growth factor (VEGF) and tumor necrosis factor alpha (TNFalpha), it decreased that of fetal liver kinase compared with those in the untreated group. Combined treatment of T4 and eCG markedly increased mRNA abundance of not only VEGF and TNFalpha, but also basic fibroblast growth factor. These data suggest that T4 may promote ovarian follicular angiogenesis in rdw rats by up-regulating mRNA expression of major angiogenic factors.
Histology and histopathology 12/2008; 23(11):1387-98. · 2.48 Impact Factor
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ABSTRACT: To analyze the ultrastructure of human ovarian follicles after cryopreservation and short-term xenografting.
Prospective experimental study.
Academic gynecology and anatomy research units.
Ovarian cortical biopsy specimens were obtained from 13 patients.
Each ovarian biopsy specimen was dissected into pieces of 1 mm(3) and divided into three groups: [1] fresh tissue, [2] frozen-thawed tissue, and [3] frozen-thawed tissue xenografted onto the peritoneum of nude mice for 3 weeks.
Follicular ultrastructure was assessed by light and transmission electron microscopy in [1] fresh, [2] frozen, and [3] frozen-grafted tissue.
Thirty-five ovarian follicles were analyzed by light and transmission electron microscopy. Twenty-five primordial and primary ovarian follicles were found. Most of them exhibited ultrastructurally well preserved features (fresh [N = 8/10], frozen [N = 7/10], and frozen-grafted [N = 4/5] tissue). Ten secondary follicles were present in xenografts. By transmission electron microscopy, all the healthy-looking secondary follicles (70%) were shown to contain intact oocytes, with features typical of earlier developmental stages, surrounded by several layers of follicular cells.
The present study demonstrates, for the first time, that cryopreservation and xenotransplantation do not appear to greatly affect human primordial/primary follicle ultrastructure. Interestingly, in frozen-thawed xenografts, secondary human ovarian follicles presented a well preserved ultrastructure, but asynchrony between oocyte and granulosa cell development was detected. The possible causes for this asynchrony are discussed.
Fertility and sterility 08/2008; 90(1):23-32. · 3.97 Impact Factor
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ABSTRACT: Collagen extracellular matrix is one of the factors related to high passive stiffness of cardiac muscle. However, the architecture and the mechanical aspects of the cardiac collagen matrix are not completely known. In particular, endomysial collagen contribution to the passive mechanics of cardiac muscle as well as its micro anatomical arrangement is still a matter of debate. In order to investigate mechanical and structural properties of endomysial collagen, we consider two alternative computational models of some specific aspects of the cardiac muscle. These two models represent two different views of endomysial collagen distribution: (1) the traditional view and (2) a new view suggested by the data obtained from scanning electron microscopy (SEM) in NaOH macerated samples (a method for isolating collagen from the other tissue). We model the myocardial tissue as a net of spring elements representing the cardiomyocytes together with the endomysial collagen distribution. Each element is a viscous elastic spring, characterized by an elastic and a viscous constant. We connect these springs to imitate the interconnections between collagen fibers. Then we apply to the net of springs some external forces of suitable magnitude and direction, obtaining an extension of the net itself. In our setting, the ratio forces magnitude /net extension is intended to model the stress /strain ratio of a microscopical portion of the myocardial tissue. To solve the problem of the correct identification of the values of the different parameters involved, we use an artificial neural network approach. In particular, we use this technique to learn, given a distribution of external forces, the elastic constants of the springs needed to obtain a desired extension as an equilibrium position. Our experimental findings show that, in the model of collagen distribution structured according to the new view, a given stress /strain ratio (of the net of springs, in the sense specified above) is obtained with much smaller (w.r.t. the other model, corresponding to the traditional view) elasticity constants of the springs. This seems to indicate that by an appropriate structure, a given stiffness of the myocardial tissue can be obtained with endomysial collagen fibers of much smaller size.
Computer Methods and Programs in Biomedicine 07/2007; 86(3):232-44. · 1.52 Impact Factor
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ABSTRACT: Many human-made chemicals are called endocrine disruptors (EDs) because they have the potential to disrupt endocrine functions in exposed organisms. Many EDs can disrupt hormonal homeostasis by interfering with hormone receptor recognition, binding and activation, while others act by still unknown mechanisms. Among the EDs specifically affecting the female reproductive system, those with steroidogenic/antisteroidogenic effects have been extensively studied and the mechanisms of toxicity clarified also at molecular level. For many others, information is restricted to few epidemiological data and in vivo/in vitro experiments with animal models. This is the case of the dithiocarbamates, and in particular of the fungicide mancozeb, an ethylenedithiocarbamate widely used to protect fruit and vegetables, ginseng included, because of its low acute toxicity in humans. Although the mechanism(s) by which mancozeb may specifically act on female reproductive organs are largely unknown, data on experimental animals in vivo have demonstrated that the fungicide can induce several disturbances on estrus cycle. When used in vitro at concentrations considered too low to cause human health injuries, the fungicide impairs mouse embryo development and meiotic spindle assembly. The possibility that the female germ cell (the oocyte) could be a specific target of mancozeb suggests a role for this fungicide as probable inductor of infertility also in exposed human populations.
Current pharmaceutical design 02/2007; 13(29):2989-3004. · 4.41 Impact Factor
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ABSTRACT: In this study the effects of mancozeb, a widely used ethylenebisdithiocarbamate fungicide, on mouse oocyte meiotic maturation and fertilization were analyzed. Oocyte cumulus cell-complexes were matured in vitro with or without increasing concentrations of the fungicide (from 0.001 to 1 microg/ml) that, due to its different stability in organic solvents and in water, was resuspended either in dimethyl sulfoxide or in culture medium. Although, about 95% of oocytes reached the metaphase II stage; mancozeb-exposed oocytes showed a dose-dependent increase of alterations in spindle morphology, and this negative effect was more evident when the fungicide was resuspended in culture medium. Under the latter culture condition, oocytes matured in the presence of 0.1 and 1 microg/ml mancozeb showed a significant reduction also in the formation of male and female pronuclei. These results indicate that mancozeb can adversely affect mammalian reproductive performance, likely by perturbing microtubular organization during meiotic maturation.
Reproductive Toxicology 08/2006; 22(1):51-5. · 3.23 Impact Factor
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Microscopy Research and Technique 07/2006; 69(6):384-5. · 1.79 Impact Factor
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ABSTRACT: The use of somatic cells for cocultures during in vitro fertilization (IVF) is currently finalized to obtain a higher number of healthy and viable embryos with a high potential of implantation. Among the different cell lines that can be used as feeder cells for cocultures, granulosa cells (GCs) are autologous cells, safe and easy to recover. The aim of the present study was to analyze the fine structure of human GCs used in a coculture system to evaluate, from a morphodynamic point of view, their role in supporting embryo development. GCs were collected during oocyte pick-up, 36 h after human chorionic gonadotropin administration, from patients undergoing IVF procedures, who had given their informed consent to be included in this protocol. After coculture, GCs were fixed and processed for light microscopy (LM) and transmission electron microscopy (TEM). By LM, GCs appeared as clusters of loosely packed cells, irregularly rounded or polyhedral in shape, varying in diameter from 18 to 25 microm. Mitotic cells, as well as regressing elements (with pyknotic nuclei or dense cytoplasm) and cell fragments were occasionally observed. By TEM, the plasma membrane was irregular due to the presence of cytoplasmic evaginations. Linear and annular gap junctions between neighboring GCs were found. GC nuclei, rounded and eccentrically located, contained finely dispersed chromatin, one (often two) prominent nucleoli and, infrequently, peripheral patches of heterochromatin. Numerous organelles populated the GC cytoplasm, among them, mitochondria were rod-shaped or elongated, usually provided with tubular-vesicular cristae but occasionally showing atypical, longitudinally oriented cristae. Membranes of smooth endoplasmic reticulum, Golgi stacks and vesicles, secretory-like granules, cisternae of rough endoplasmic reticulum (RER), free ribosomes and polysomes, lysosomal-like bodies, microfilaments, and lipid droplets were also seen in the GC cytoplasm. In most cells, RER was scarcely represented and numerous lipid droplets filled the perinuclear space. On the contrary, some GCs contained an abundant RER and rare lipid droplets scattered in the cytoplasm. In conclusion, our data demonstrated the presence, in a coculture system, of GCs provided with ultrastructural characteristics typical of healthy, metabolically active, mostly steroidogenic cells. Protein-synthetic cells have also been detected. These data, evaluated at the light of biochemical and clinical studies, sustain the capability of human GCs cocultures to positively affect early embryo development in vitro by the secretion of steroids and proteins, putative "embryotrophic" factors.
Microscopy Research and Technique 07/2006; 69(6):508-16. · 1.79 Impact Factor
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ABSTRACT: To describe the morphological characteristics of the ovarian follicle (F) capillary neoformation and regression, the angiogenic figures were studied by means of scanning electron microscopy of corrosion casts in developing and mature F of rabbit, pig, and cow. Developing F showed gradual neoformation of thecal capillaries characterized by budding and then sprouting, likely from preexisting interstitial vessels. Postcapillary venules frequently showed vasoconstriction rings (sphincters). Vasodilation followed capillary elongation. Mature F, in addition to vessel elongation and dilation, also presented infolding of dilated capillary walls, followed by capillary duplication and sinusoidalization. Periovulatory F mainly showed functional changes, such as capillary dilation, signs of iperpermeabilization, and ischemia, the latter being limited to the apical follicular area. Vessel regression was characterized by thinning of capillaries and presence of avascular areas within the atretic F wall at any stage. This study showed two main types of angiogenic patterns. (a) longitudinal elongation (in series, sprouting angiogenesis) characterizing the initial phase of F development and (b) parallel duplication (in parallel, infolding or intussusceptive angiogenesis), ending in capillary lateral replication or splitting, secondary to functional microvascular changes. Indirect evidence of the establishment of postcapillary resistances contributing to capillary remodeling, was also shown. It is concluded that the sequence of capillary neoformation in mammalian ovarian F occurs in six steps: (1) budding, (2) sprouting (and elongation), (3) dilation, (4) infolding (intussusception), (5) duplication (splitting and elongation), and (6) sinusoidalization. Capillary regression hits F at any stage and characterizes F atresia.
Microscopy Research and Technique 07/2006; 69(6):459-68. · 1.79 Impact Factor
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ABSTRACT: Mancozeb is known to alter reproductive performance in exposed animals, but its specific mechanism of action is still unclear. We investigated whether in female mice of the F1 generation, mancozeb could affect oocyte ability to undergo complete meiotic maturation and fertilization. Female mice were treated with 50 and 500 mg/kg of mancozeb (or vehicle in the controls) from gestational day 2 to postnatal day 20. Results demonstrated that only at the highest dose, mancozeb induced a significant decrease in the number of ovulated eggs. Moreover, at this dose mancozeb caused a significant decrease of fertilizability related to a reduction of the formation of male and female pronuclei.
Reproductive Toxicology 03/2006; 21(2):216-9. · 3.23 Impact Factor
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ABSTRACT: In order to understand the 3-D distribution of collagen in relation to vascularization, chorionic villi of human placentae, belonging to normal pregnancies at term, were studied by scanning electron microscopy (SEM) after alkali maceration techniques, and by transmission electron microscopy (TEM). The villous tree appeared made of an uninterrupted structure of collagen fibres. The collagen fibres connected the chorionic villi axis with their basal plates and organised differently according to the various levels of villous branching. The collagen of stem villi showed copious fibres. The external fibres (facing the villous surface) were arranged mainly longitudinally. The central core of the villi (inner fibres) were arranged concentrically around the wall of the fetal vessels. Both external and internal fibres formed stratified lamellae or small parallel bundles. The inner core of stem villi showed small holes housing capillary spaces. Mature intermediate and terminal villi showed a scarce amount of collagen arranged in thin concentric layer within the villous core, surrounding numerous dilated capillary and sinusoid spaces.These observations demonstrated that the extracellular matrix of human chorionic villi is highly compartmentalised and shows a variable structural 3-D distribution depending on the branching level of the villous tree, such a distribution ensures the most favourable microenvironment for feto-maternal exchanges and it is likely able to provide a modulated support to the developing chorionic fetal vessels and trophoblastic layer as well.
Journal of Reproduction and Development 09/2005; 51(4):433-43. · 1.46 Impact Factor
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ABSTRACT: Human zona pellucida (ZP) is maintained up to the blastocyst stage prior to hatching. In in vitro fertilized (IVF) embryos, it eventually acts as a morphodynamic interface between the cultured embryo and its microenvironment. Ultrastructural data on the ZP of IVF blastocysts are scarce in humans. We employed correlated phase contrast microscopy (PCM) and scanning electron microscopy (SEM) to study retrospectively the ultrastructural morphology of the ZP outer surface of 20 IVF human blastocysts from 16 Japanese patients (28-44 years of age, average 36.7+/-4.2) with a history of infertility. Blastocysts were derived from conventional in vitro fertilization (cIVF) (n = 10) and from intracytoplasmic sperm injection (ICSI) (n = 10). Both cIVF and ICSI groups included "clear blastocysts" (n = 5) and "dark blastocysts" (n = 5). By PCM, the clear blastocysts exhibited a regular, round-shaped contour and consisted of clear and voluminous cells. By SEM, they displayed a spongy ZP with numerous fenestrations formed by networked filaments. By PCM, dark blastocysts appeared irregularly shaped and often collapsed, and comprised dark cells and debris. By SEM, their ZP were smooth with remnants of compact fenestrations. In conclusion, viable blastocysts presented a normal ZP outer surface ultrastructure, whereas unhealthy blastocysts showed an altered ZP outer surface, comparable to that of immature/atretic oocytes. Such alterations could reflect sub-optimal culture conditions and/or could be related to blastocyst degenerative processes. The blastocyst ZP surface ultrastructure was unaffected by the fertilization technique (cIVF or ICSI). These data suggest that blastocyst survival in vitro is related to ZP ultrastructure maintenance.
Archives of Histology and Cytology 07/2005; 68(2):133-41. · 0.57 Impact Factor
