Stage-specific immunolabeling for oviductin in the secretory granules of the oviductal epithelium of the golden hamster during the estrous cycle

Background: We have previously localized an antigen of oviductal origin in the zona pellucida of postovulatory hamster ova. This antigen is a high molecular weight glycoprotein secreted by the non-ciliated secretory cells of the oviduct and is later transferred to the zona pellucida of the oocyte during oviductal transit. This glycoprotein is rich in N-acetyl-D-galactosamine residues and has been designated Hamster Oviductin-1. In the present study, a monoclonal antibody (MAb) raised against this oviductin was used to detect the presence of this antigen in oviductal tissue during the estrous cycle. Methods: Twenty mature female golden hamsters were used and were divided into five groups of five animals each according to the five different stages of the estrous cycle. Quantitative immunocytochemistry was performed on MAb-labeled thin sections of Lowicryl-embedded ampullary region of hamster oviducts. Control experiments were also carried out to assess the specificity of the immunolabeling. Results: Quantitative analysis of the immunogold labeling indicated that maximum labeling for oviductin in the secretory granules of oviductal epithelial secretory cells was found around the time of ovulation, i.e., at estrus. The intensity of immunolabeling decreased from metestrus to diestrus 1, was at a minimum at diestrus 2, and started to increase at proestrus. Conclusion: Together, these quantitative results indicate that expression of oviductin in the secretory granules of the hamster oviductal secretory cells is stage specific. Maximum labeling for the antigen coincides with the time of ovulation suggesting an important role for the oviductal epithelium in contributing its secretory product to the zona pellucida of oocytes freshly released from the ovary. Since the oviduct is the site of sperm-egg interaction and where fertilization and early embryo development take place, the maximal production of oviductin at the time of ovulation may facilitate some of these crucial steps during the intricate process of reproduction. © 1995 Wiley-Liss, Inc.     

Ultrastructural changes in the oviductal epithelium of merino ewes during the estrous cycle

Isthmic and ampullary oviductal epithelia sampled from Merino ewes at days -1, 1, 3, and 10 of the estrous cycle (estrus = day 0) were studied by scanning and transmission electron microscopy after fixation by vascular perfusion. Secretory cells, ciliated cells, and lymphocytelike basal cells were observed in both isthmic and ampullary epithelium at all stages of the estrous cycle studied and their ultrastructural features were analyzed. Synthesis of lamellated secretory granules occurred in the ampullary secretory cells during the follicular and early luteal phases, and their contents were released by exocytosis into the oviductal lumen during the luteal phase. Granule release was associated with nucleated apical protrusion of these cells into the oviductal lumen. No such secretory activity was displayed by isthmic secretory cells even though a few cells contained nonlamellated granules. Apocrine release of apical vesicles and accompanying cytoplasmic material from apical protrusions of ciliated cells occurred in the isthmus around estrus but not in the ampulla. This unexpected feature has not previously been reported in any other mammal. Dendritic basal cells were distinguished in the lower part of the epithelium by their heterochromatic nuclei, electron-lucent cytoplasm, and lack of attachment zones. No migration of basal cells was observed, and their ultrastructural features were similar in the ampulla and isthmus and at all stages of the estrous cycle examined. The function of these lymphocytelike cells in the epithelium is uncertain, but the presence of phagocytic bodies and lysosomes in 20% of them may indicate a phagocytic role.     

Secretory cells of the oviduct of the pig-tailed monkey,Macaca nemestrina,during the menstrual cycle and after estrogen treatment

The secretory cells of the oviductal epithelium in the pig-tailed monkey, Macaca nemestrina, have been studied by light and electron microscopy. Changes during the menstrual cycle and after ovariectomy, with and without subsequent estrogen treatment, have been documented. During the early follicular phase the epithelium is recovering from deciliation and secretory cell atrophy that occur in the late luteal phase. A few fimbrial and a moderate number of ampullar and isthmic cells contain a few electron-dense, homogeneous secretory granules in their apical cytoplasm. During the late preovulatory and early postovulatory periods, secretory cell structure varies considerably. Fimbrial cells typically display apical protrusions that contain no or a few small, mainly homogeneous, secretory granules. The cytoplasm is crowded with elements of the Golgi complex, with granular endoplasmic reticulum profiles often intimately associated with mitochondria, and with variable numbers of polysomes and glycogen granules. In ampullar and isthmic cells secretory granules are more abundant than in fimbrial cells, and electron-lucent vacuoles appear. The granules are of two types: (1) those having an electron-dense, homogeneous matrix, and (2) those possessing lamellar structures within moderately dense matrices. The lamellae of the second type course in parallel arrays separated by a distance of approximately 15.5 nm and exhibit a periodicity of approximately 11.3 nm. Possible transitional stages between the lamellar granules and the vacuoles containing lamellar fragments are observed. Secretion occurs by exocytosis. During the late luteal phase no fimbrial cells have secretory granules. In the ampulla many of the cells have poor development of the organelles involved in secretory activity and have few or no secretory granules. In others, a moderate number of secretory granules are present; in one animal, exocytosis is observed. In the untreated ovariectomized animal no secretory granules occur, and the organelle content is much less than in the cycling and the estrogen-treated monkeys. In ovariectomized, estradiol-treated monkeys, some areas of all three oviductal segments are well stimulated whereas others display little or no secretory activity.     

Lectin binding patterns of porcine oviduct mucosa and endometrium during the oestrous cycle

Important functions of the oviduct during reproduction include the provision of an optimal environment for gametes and zygotes and nutrition of the early embryo. These functions are ensured by the secretion of an oviductal fluid which is known to contain organ-specific glycoproteins. Glycoconjugates of the apical glycocalyx are considered to play a major role in cell recognition and interaction processes. In the present investigation, binding patterns of Con A, HPA, LTA, RCA I, UEA I, and WGA were studied in defined segments of the oviduct (infundibulum, ampulla, isthmus) and in the uterus during the oestrous cycle. The carbohydrate distribution showed regional as well as time dependent differences. LTA, HPA and WGA reacted strongly with Golgi regions and secretory granules in the oviduct epithelium during the follicular phase, indicating high secretory activity. LTA, HPA, and UEA I also revealed a varying carbohydrate composition of the glycocalyx during the oestrous cycle. Prominent regional differences in glycoconjugate expression were shown in oviductal segments by LTA and HPA binding: during the follicular phase, LTA binding sites were only present on epithelial cells of the isthmic segment, the ampulla and infundibulum being unreactive. D-N-acetyl-galactosamine residues were demonstrated on ciliated epithelial cells of the ampulla and infundibulum exclusively during oestrus. The glycocalyx of uterine epithelial cells was clearly defined by HPA, WGA, LTA, RCA I and UEA I; LTA binding was restricted to the secretory phase. The observation of regional and time dependent variability in glycoconjugate distribution strongly indicates their specific physiological functions in reproductive processes.     

Ultrastructural features of goat oviductal secretory cells at follicular and luteal phases of the oestrous cycle

The aim of the present study was to investigate the ultrastructure of secretory cells in the various regions of the goat oviduct during the follicular and luteal phases of the oestrous cycle. During the follicular phase in the fimbriae, the secretory cells contained small secretory granules with electron-dense matrices. In the luteal phase, the secretory granules disappeared and cytoplasmic protrusions, extending beyond the luminal border of the ciliated cells and often containing the nucleus, were predominant. During the follicular phase in ampullary secretory cells, numerous secretory granules with moderately electron-dense matrices were present in the supranuclear cytoplasm and exocytosis of secretory granules was observed. The number of secretory granules was dramatically reduced in the ampullary secretory cells at the luteal phase. Conspicuous cytoplasmic protrusions of secretory cells were observed similar to those of the fimbrial epithelium. Isthmic cells were almost free of secretory granules and lysosome-like bodies were found both at the follicular and luteal phases. In conclusion, our ultrastructural observations of goat oviduct revealed marked cyclic changes in the ultrastructural features of secretory cells and the ultrastructural features and the numbers of secretory granules were distinctive for each particular segment.     

Changes of glycoconjugate contents of the zona pellucida during oocyte growth and development in the golden hamster: a quantitative cytochemical study

We demonstrated here with the high resolution lectin-gold approach and quantitative analysis, changes of glycoconjugates in the hamster zona pellucida (ZP) during oocyte growth and development in the ovary and the oviduct. Glycoconjugates which contain N-acetyl-D-galactosamine as terminal sugar residues are absent in the ovary but are secreted by secretory cells in the oviduct and are added to the ZP of superovulated oocytes during oviductal transit. Glycoconjugates which carry sialic acid as terminal sugar residues appear to be acquired mainly from the ovary. The oviduct contributes little of this particular component to the ZP during the transit of oocytes in the oviduct. On the contrary D-galactose and N-acetylglucosamine associated glycoconjugates, added to the ZP in ovarian follicles, are also secreted by non-ciliated oviductal epithelial cells and these secretory products are transferred to the ZP in significant amount during passage of the oocyte through the oviduct. Lectin-gold labeling of the ZP of superovulated oocytes reveals homogeneous distribution of gold particles throughout the zona matrix. Thus, we conclude that the ZP of hamster superovulated oocytes consists of glycoconjugates that may derive from different origins. Deposition of ZP glycocomponents begins in the ovary. Similar and new glycoconjugates, secreted by oviductal non-ciliated secretory cells, are added to the ZP of oocytes during oviductal transit. At this stage the ZP is made up of a homogeneous matrix of glycoconjugates.     

The Peritoneal Mesothelium Covering the Genital Tract and its Ligaments in the Female Pig Shows Signs of Active Function

The aim of this study was to describe the surface features of the peritoneal mesothelium covering the genital tract and adjacent ligaments of the sow to find signs of biosynthetic activation of cells. Surface features of the serosa covering the genital tract and adjacent ligaments from 14 cyclic sows, 7 in the follicular phase and 7 in the luteal phase of the estrous cycle, were examined by histology and scanning electron microscopy. Five additional sows, three in the follicular phase and two in the luteal phase of the estrous cycle, were examined by transmission electron microscopy (TEM). In this study, the presence of cells of the oviductal epithelium in the serosa of the infundibulum and the ampulla, as well as indications of a high functional activity of the mesothelial cells in the areas studied were two aspects that differed from the findings of previous works. Presence of endosalpingeal cells was observed in the serosal surface, showing cyclical variations with a predominance of either ciliated cells during the follicular phase or secretory cells during the luteal phase. Signs of high functional activity of the mesothelial cells included the predominance of cuboidal over flattened cells, a cytoplasm richly supplied with organelles, a dense microvillous coat, numerous primary cilia, and many secretory structures on the surface of cells. These results indicate that the serosa covering the genital area and the adjacent ligaments in the sow has an active epithelium whose coordinating role between reproductive tissues may be far more significant than previously thought. Anat Rec, 2007. © 2007 Wiley‐Liss, Inc.     

Secreted proteins of the oviduct

During late follicular development and estrus, the mammalian oviduct undergoes specific physiological and biochemical modifications which contribute to an optimization of the microenvironment for fertilization and early cleavage-stage embryonic development. These changes appear to be hormonally regulated by ovarian steroids, most importantly, estrogen. The hundreds of macromolecules found within the oviductal lumen are contributed by selective serum transudation and active biosynthesis and secretion from nonciliated epithelial cells. Recent studies have indicated temporal and regional (infundibulum, ampulla and isthmus) differences in steady-state levels of specific mRNAs and in de novo protein synthesis and secretion by the oviduct. One protein synthesized de novo, the estrogen-dependent oviductal secretory glycoprotein (OSP), has been shown to be unique to the oviduct and is conserved across a number of mammalian species. This protein associates with the zona pellucida, perivitelline space and vitelline or blastomere membrane of ovulated eggs and preimplantation embryos. OSPs have been shown to enhance sperm binding and penetration in oocytes and may regulate development in early preimplantation embryos. Other regulatory molecules, protease inhibitors, growth factors, cytokines, binding proteins, enzymes and immunoglobulins have been identified in the oviductal microenvironment. The identification and potential roles for oviduct-secreted proteins will be reviewed and discussed. Current research focuses on continued identification and characterization of specific oviductal proteins and a determination of the molecular basis of their interactions with the oocyte, sperm or embryo.     

Examination by scanning electron microscopy of oviductal epithelium of the prolific Chinese Meishan pig at follicular and luteal phases.

The luminal surfaces of epithelial cells in the fimbriae, ampulla, isthmus, and utero-tubal junction of the oviducts of the prolific Chinese Meishan pig at follicular and luteal phases of the estrous cycle were examined by scanning electron microscopy. Marked cyclic changes were observed on the surfaces of cells in the fimbriae and ampulla, but little change was found in the isthmus and at the utero-tubal junction. The cells of the fimbrial epithelium in the follicular phase were densely ciliated, and the cilia partially concealed the bulbous processes of the secretory cells. In the luteal phase, the secretory cells predominated in the epithelium, and most of the ciliated cells were hidden by the processes of the secretory cells. The ampullar epithelium showed similar changes, but to a lesser extent. In the isthmus and at the utero-tubal junction, the secretory cells had many microvilli on their bulbous processes at the follicular phase, but they were flat and the microvilli were fewer in number and shorter in length during the luteal phase. Conspicuous solitary cilia protruded from the surfaces of secretory cells in the fimbriae and ampulla during the luteal phase. These results demonstrate that there are regional variations in the cyclic changes associated with the oviductal epithelial cells of the Chinese Meishan pig.     

Ultrastructural and ultracytochemical features of secretory granules in the ampullary epithelium of the hamster oviduct.

The epithelium of mammalian oviducts consists mainly of ciliated and non-ciliated secretory cells. In some mammals, secretory products originating from oviductal secretory cells have been shown to bind to the surface of, or accumulate within, ovulated eggs and/or developing embryos. These findings suggest that the secretions of the oviductal epithelial cells may play an important role in reproductive and developmental events that occur in the oviduct. In the present study, ultrastructural and cytochemical features of secretory cells in the hamster ampullary epithelium were shown by routine electron microscopy, lectin-gold cytochemistry and both conventional freeze-fracture and rapid-freezing techniques with special reference to the organizational aspects of their secretory granules. The use of ferrocyanide-reduced osmium tetroxide as a post-fixative in the Epon embedment of ampullary tissue samples also proved to be advantageous especially in revealing the carbohydrate contents of certain cellular compartments. The most conspicuous characteristic of the secretory cells, based on their staining property, was the presence of two types of secretory granules: those with a homogeneous electron-dense matrix and those with an electron-lucent matrix. Under favorable conditions, distinct features of the organizational arrangement of a crystalline lattice inside the secretory granules were also revealed. This well organized crystalline lattice shown in sections of Epon-embedded oviductal tissue was confirmed by examination of replicas of freeze-fractured oviducts prepared by the rapid-freezing technique. We also demonstrated with high resolution lectin-gold cytochemistry the intracellular distribution of lectin-binding glycoconjugates in the secretory cells of the hamster oviductal ampulla often in a linear array following the crystalline lattice. The results obtained in this study, taken together, provide insight into a possible link of the internal topographical features of oviductal secretory granules along with the cytochemical properties of their contents to the anticipated regulatory mechanism underlying their process of secretions.     

Examination by scanning electron microscopy of oviductal epithelium of the prolific Chinese Meishan pig at follicular and luteal phases

The luminal surfaces of epithelial cells in the fimbriae, ampulla, isthmus, and utero-tubal junction of the oviducts of the prolific Chinese Meishan pig at follicular and luteal phases of the estrous cycle were examined by scanning electron microscopy. Marked cyclic changes were observed on the surfaces of cells in the fimbriae and ampulla, but little change was found in the isthmus and at the utero-tubal junction. The cells of the fimbrial epithelium in the follicular phase were densely ciliated, and the cilia partially concealed the bulbous processes of the secretory cells. In the luteal phase, the secretory cells predominated in the epithelium, and most of the ciliated cells were hidden by the processes of the secretory cells. The ampullar epithelium showed similar changes, but to a lesser extent. In the isthmus and at the utero-tubal junction, the secretory cells had many microvilli on their bulbous processes at the follicular phase, but they were flat and the microvilli were fewer in number and shorter in length during the luteal phase. Conspicuous solitary cilia protruded from the surfaces of secretory cells in the fimbriae and ampulla during the luteal phase. These results demonstrate that there are regional variations in the cyclic changes associated with the oviductal epithelial cells of the Chinese Meishan pig. © 1992 Wiley-Liss, Inc.     

Ultrastructural and ultracytochemical features of secretory granules in the ampullary epithelium of the hamster oviduct

The epithelium of mammalian oviducts consists mainly of ciliated and non‐ciliated secretory cells. In some mammals, secretory products originating from oviductal secretory cells have been shown to bind to the surface of, or accumulate within, ovulated eggs and/or developing embryos. These findings suggest that the secretions of the oviductal epithelial cells may play an important role in reproductive and developmental events that occur in the oviduct. In the present study, ultrastructural and cytochemical features of secretory cells in the hamster ampullary epithelium were shown by routine electron microscopy, lectin‐gold cytochemistry and both conventional freeze‐fracture and rapid‐freezing techniques with special reference to the organizational aspects of their secretory granules. The use of ferrocyanide‐reduced osmium tetroxide as a post‐fixative in the Epon embedment of ampullary tissue samples also proved to be advantageous especially in revealing the carbohydrate contents of certain cellular compartments. The most conspicuous characteristic of the secretory cells, based on their staining property, was the presence of two types of secretory granules: those with a homogeneous electron‐dense matrix and those with an electron‐lucent matrix. Under favorable conditions, distinct features of the organizational arrangement of a crystalline lattice inside the secretory granules were also revealed. This well organized crystalline lattice shown in sections of Epon‐embedded oviductal tissue was confirmed by examination of replicas of freeze‐fractured oviducts prepared by the rapid‐freezing technique. We also demonstrated with high resolution lectin‐gold cytochemistry the intracellular distribution of lectin‐binding glycoconjugates in the secretory cells of the hamster oviductal ampulla often in a linear array following the crystalline lattice. The results obtained in this study, taken together, provide insight into a possible link of the internal topographical features of oviductal secretory granules along with the cytochemical properties of their contents to the anticipated regulatory mechanism underlying their process of secretions. Anat Rec 255:227–239, 1999. © 1999 Wiley‐Liss, Inc.     

The induction of baboon glycodelin expression by progesterone is not through Sp1

Glycodelin is a major secretory product of the uterine glandular epithelial cells of the human and non-human primate during the late luteal phase of the menstrual cycle and early pregnancy. Since progesterone levels are elevated during these periods we sought to determine how progesterone modulates glycodelin gene expression. Co-transfection of various deletions of the baboon glycodelin promoter with the progesterone receptor (PR) into Ishikawa cells, a human endometrial cell line, revealed that full progesterone responsiveness is retained within the region -119/+48. In COS-1 cells, a kidney cell line, progesterone failed to elevate luciferase levels when various deletion constructs and the PR were co-transfected. Mutation of the Sp1 site in the -67/+48 region lowered basal expression but did not affect the ability of progesterone to increase expression of the luciferase reporter in Ishikawa cells. These findings suggest that Sp1 sites are not involved in the progesterone regulation of the baboon glycodelin gene. We propose that progesterone induces a factor that regulates glycodelin gene expression in the uterus since we failed to obtain a similar response in a non-uterine cell line.     

Ultrastructural immunocytochemical studies of the localization and distribution of somatostatin, calcitonin, calcitonin gene-related peptide, and substance P in the bat thyroid follicle

Electron microscope immunocytochemistry was used to determine the intracellular localization and distribution among follicular elements of four peptides: calcitonin, somatostatin, calcitonin gene-related peptide (CGRP), and substance P in the thyroid glands of bats captured in the prehibernation phase of their annual life cycle. Previous studies have shown that this period of the hibernation-activity cycle is characterized by the accumulation and storage of secretory granules in parafollicular cells. Sites of binding of primary antisera to each of the four peptides were identified by means of affinity-purified secondary antisera directly coupled to colloidal gold particles. Calcitonin and somatostatin immunoreactivities were found in all parafollicular cells examined and in every secretory granule within these cells. CGRP was also found in all parafollicular cells examined (n = 75) but only in about half of their secretory granules. In contrast to these peptides, substance P immunoreactivity was not found in any parafollicular cells, but was localized exclusively in nerve endings within the basement membrane of the follicle.     

Control of the secretory cell cycle in cat oviduct by estradiol and progesterone

The control of the secretory cell cycle by estradiol and progesterone in the oviduct of the cat was studied using light and electron microscopy. The epithelium in ovariectomized animals was cuboidal with no evidence of secretory activity. Estradiol treatment induced hypertrophy, hyperplasia, and the differentiation of both secretory and ciliated cells. Differentiation of the secretory cell included the development of an extensive area of basal rough endoplasmic reticulum and a large supranuclear Golgi region. Apical secretory granules were already present after 3 days of estradiol treatment, and after 4 to 5 days maximum hypertrophy and differentiation had occurred. Most cells contained several apical electron-dense granules; however, no large accumulation of granules in any one cell was ever observed. Occasional release of secretory product by exocytosis occurred during chronic treatment with estradiol. Rapid elevation of the serum levels of estradiol or progesterone by means of IV injection did not enhance exocytosis or result in any ultrastructural alterations. The chronic administration of progesterone to estradiol-primed animals resulted in rapid cell atrophy, dedifferentiation, and death (apoptosis) within the epithelium of the oviduct. Secretory granules were no longer observed after 2 days of estradiol and progesterone treatment, and after 7 days the epithelium was approximately the same height as that measured in ovariectomized animals. These data illustrate that estradiol induces the differentiation and maintains the mature state of the secretory cell within the oviductal epithelium of the cat, and that progesterone has an immediate antiestrogenic effect on these cells. This study also suggests that the secretory product is released gradually, as the granules form and mature during chronic estradiol administration.     

Stromal cell interaction and relevance to predecidual events and menstruation

Endometrial stromal cells play a vital role during decidual-izationand implantation. The aim of this study was to analyse the cyclicultrastructural variations of stromal fibroblasts, granulocytesand blood-derived cells which invade the stroma during variousstages of the menstrual cycle. Diverse opinions exist in theliterature regarding the origin, fate and function of the endometrialgranulocytes. Our study is in agreement with available immunologicaldata and shows that (i) fibroblasts and granulocytes are distinctcell types in the stroma, (ii) they exhibit distinct changesacross the menstrual cycle and (iii) fibroblasts are not thecommon progenitor for granulocytes and predecidual cells inthe secretory phase endometrium, as suggested by previous investigators.The infiltration of eosinophils, platelets, macrophages, neutrophils,etc., during the menstrual phase makes the stroma a potentialsource of growth factors and cytokines which may regulate theprocesses of regression and regeneration of the endometrium.Furthermore, we propose that endometrial granulocytes couldbe the source of decidual prolactin because the ultrastructuralmorphology of their secretory granules closely resembles thatof the prolactin-secreting cells in the pituitary.     

Differences between mature ovarian and oviductal oocytes: a study using the golden hamster

To investigate whether mature ovarian oocytes are physiologically identical with recently ovulated oocytes, hamster oocytes collected from ovaries approximately 1 h before ovulation were compared with the oocytes which had been in the oviduct for 3-4 h. Although these two groups of oocytes were at the metaphase of the second meiosis, there were quantitative differences between the two with respect to (i) sensitivity of cumulus matrix to hyaluronidase and spermatozoa (oviductal greater than ovarian), (ii) size of the perivitelline space (oviductal greater than ovarian), (iii) viscosity of the ooplasm (ovarian greater than oviductal), (iv) responsiveness to Ca2+-ionophore (oviductal greater than ovarian), and (v) time needed in completing meiosis (ovarian greater than oviductal). The most prominent difference was found in the zona pellucida. The zona of the oviductal oocyte was 'heterogeneous' in its optical density and had a stronger acrosome-reaction-inducing ability than that of the ovarian oocyte. When cultured in artificial media (e.g. F-10 medium with serum) for 4 h, the ooplasm of ovarian oocytes became like that of oviductal oocytes. However, their zonae remained unchanged. Zonae of ovarian oocytes became like those of oviductal oocytes only when they were exposed to ampullary and/or isthmic fluids. The zona-altering factors in the oviductal fluid (oviductal glycoproteins), which are apparently integrated into the native zona, may act to enhance the various functions of the zona.     

Cyclic changes of granulocyte colony-stimulating factor (G-CSF) mRNA in the human follicle during the normal menstrual cycle and immunolocalization of G-CSF protein

BACKGROUND: Ovulation has several similarities with inflammation and is closely connected to the activity of leukocytes and inflammatory cytokines. Since granulocytes are one of the major leukocytes, we focused our attention on the presence and local production of granulocyte colony-stimulating factor (G-CSF) in the human ovary. METHODS: The presence of G-CSF protein in the follicular fluid and perifollicular tissues was examined by Western blot analysis (n = 5) and immunohistochemical staining (n = 10). The relative expression levels of G-CSF mRNA in relation to GAPDH in granulosa, theca and luteal cells during the menstrual cycle were measured by quantitative RT-PCR using TaqMan technology (n = 15). RESULTS: G-CSF protein was detected in all follicular fluid and located mainly in granulosa cells of the follicle and luteal cells. The expression level of G-CSF mRNA in the late follicular phase was 137.6 +/- 18.5, which was approximately 10-fold greater than other phases during the menstrual cycle (P < 0.05). CONCLUSIONS: These results demonstrate that G-CSF is produced in the human follicle shortly before the ovulatory phase and may play an important role in the mechanism of ovulation.     

Large scale validation of human N-myc downstream-regulated gene (NDRG)-1 expression in endometrium during the menstrual cycle

A major challenge in the comprehension of the endometrial transformations leading to the completion of each menstrual cycle in humans is in the identification of specific molecular pathways underlying these monthly turnovers. Towards this goal we compared, by the differential display technique, the relative expression of mRNA in endometrial biopsies harvested in individuals (n = 48) either at the proliferative or the secretory phase of the menstrual cycle. We isolated a cDNA fragment homologous to NDRG1 (N-myc Downstream-Regulated Gene-1) that is present in markedly higher amounts in the secretory phase. Northern blot analysis and quantitative real time PCR experiments confirmed this result in distinct cohorts of individuals (44 and 560 respectively). A closer examination of data showed that the highest mRNA levels were found during the range of 25-28 days of the uterine cycle. Consistent with the mRNA data, the temporal profile of the NDRG1 protein showed a 15-fold increase during the secretory phase, as demonstrated by using semi-quantitative dot blot analyses (n = 92). Immunohistochemical localization revealed that NDRG1 was expressed both in epithelial and stromal cells. This large scale validation of the NDRG1 mRNA and protein increase in endometrium during the secretory phase is consistent with its differentiation-related function described in other tissues and its potential involvement in the window of implantation of the human endometrium, as suggested by previous chip-based evidence.     

Composition and morphology of lipid droplets from oviduct epithelial cells

This study was undertaken to determine the composition and morphology of lipid droplets in situ and isolated from oviductal epithelial cells and oviductal fluid. Oviductal epithelial cells were harvested enzymatically from oviducts of cows in either the luteal or the follicular stages of the ovarian cycle. Lipid droplets were isolated from cellular homogenates and characterized biochemically using thin layer chromatography. The morphology of lipid droplets in oviductal epithelial cells and in fractions isolated by ultracentrifugation from cellular homogenates was examined by electron microscopy. Lipid droplets isolated from oviduct epithelial cells varied in composition with the ovarian cycle and the oviductal region. There was more total lipid in droplets isolated from cows in the luteal than follicular phase of the ovarian cycle. Most of this difference was due to large amounts of esterified cholesterol present in the samples from luteal-stage animals. The most esterified cholesterol was found in droplets isolated from the oviductal isthmus of luteal cows. Droplets similar in lipid composition to those isolated from epithelial cells were found in oviductal fluid. Four distinct types of lipid inclusions were evident in electron micrographs of oviductal epithelia and characterized as osmiophilic droplets, lipofuscin-like clusters, lamellar structures, and composite bodies. All of the lipid inclusions were found in droplet isolates except for the extracted lipid portion of the composite body. The presence and diversity of oviduct epithelial lipid inclusions suggest that the oviductal epithelium may be very active in lipid metabolism, particularly cholesterol dynamics. © 1993 Wiley-Liss, Inc.