Factor VII deficiency rescues the intrauterine lethality in mice associated with a tissue factor pathway inhibitor deficit.
Mice doubly heterozygous for a modified tissue factor pathway inhibitor (TFPI) allele (tfpi delta) lacking its Kunitz-type domain-1 (TFPI+/delta) and for a deficiency of the factor VII gene (FVII+/-) were mated to generate 309 postnatal and 205 embryonic day 17.5 (E17. 5) offspring having all the predicted genotypic combinations. Progeny singly homozygous for the tfpidelta modification but with the wild-type fVII allele (FVII+/+/TFPIdelta/delta), and mice singly homozygous for the fVII deficiency and possessing the wild-type tfpi allele (FVII-/-/TFPI+/+), displayed previously detailed phenotypes (i.e., a high percentage of early embryonic lethality at E9.5 or normal development with severe perinatal bleeding, respectively). Surprisingly, mice of the combined FVII-/-/TFPIdelta/delta genotype were born at the expected mendelian frequency but suffered the fatal perinatal bleeding associated with the FVII-/- genotype. Mice carrying the FVII+/-/TFPIdelta/delta genotype were also rescued from the lethality associated with the FVII+/+/TFPIdelta/delta genotype but succumbed to perinatal consumptive coagulopathy. Thus, the rescue of TFPIdelta/delta embryos, either by an accompanying homozygous or heterozygous FVII deficiency, suggests that diminishment of FVII activity precludes the need for TFPI-mediated inhibition of the FVIIa/tissue factor coagulation pathway during embryogenesis. Furthermore, the phenotypes of these combined deficiency states suggest that embryonic FVII is produced in mice as early as E9.5 and that any level of maternal FVII in early-stage embryos is insufficient to cause a coagulopathy in TFPIdelta/delta mice. (+info)
Progesterone inhibits estrogen-induced cyclin D1 and cdk4 nuclear translocation, cyclin E- and cyclin A-cdk2 kinase activation, and cell proliferation in uterine epithelial cells in mice.
The response of the uterine epithelium to female sex steroid hormones provides an excellent model to study cell proliferation in vivo since both stimulation and inhibition of cell proliferation can be studied. Thus, when administered to ovariectomized adult mice 17beta-estradiol (E2) stimulates a synchronized wave of DNA synthesis and cell division in the epithelial cells, while pretreatment with progesterone (P4) completely inhibits this E2-induced cell proliferation. Using a simple method to isolate the uterine epithelium with high purity, we have shown that E2 treatment induces a relocalization of cyclin D1 and, to a lesser extent, cdk4 from the cytoplasm into the nucleus and results in the orderly activation of cyclin E- and cyclin A-cdk2 kinases and hyperphosphorylation of pRb and p107. P4 pretreatment did not alter overall levels of cyclin D1, cdk4, or cdk6 nor their associated kinase activities but instead inhibited the E2-induced nuclear localization of cyclin D1 to below the control level and, to a lesser extent, nuclear cdk4 levels, with a consequent inhibition of pRb and p107 phosphorylation. In addition, it abrogated E2-induced cyclin E-cdk2 activation by dephosphorylation of cdk2, followed by inhibition of cyclin A expression and consequently of cyclin A-cdk2 kinase activity and further inhibition of phosphorylation of pRb and p107. P4 is used therapeutically to oppose the effect of E2 during hormone replacement therapy and in the treatment of uterine adenocarcinoma. This study showing a novel mechanism of cell cycle inhibition by P4 may provide the basis for the development of new antiestrogens. (+info)
Daidzein and genistein glucuronides in vitro are weakly estrogenic and activate human natural killer cells at nutritionally relevant concentrations.
Daidzein and genistein glucuronides (DG and GG), major isoflavone metabolites, may be partly responsible for biological effects of isoflavones, such as estrogen receptor binding and natural killer cell (NK) activation or inhibition. DG and GG were synthesized using 3-methylcholanthrene-induced rat liver microsomes. The Km and Vmax for daidzein and genistein were 9.0 and 7.7 micromol/L, and 0.7 and 1.6 micromol/(mg protein. min), respectively. The absence of ultraviolet absorbance maxima shifts in the presence of sodium acetate confirmed that the synthesized products were 7-O-glucuronides. DG and GG were further purified by a Sephadex LH-20 column. DG and GG competed with the binding of 17beta-(3H) estradiol to estrogen receptors of B6D2F1 mouse uterine cytosol. The concentrations required for 50% displacement of 17beta-(3H) estradiol (CB50) were: 17beta-estradiol, 1.34 nmol/L; diethylstilbestrol, 1.46 nmol/L; daidzein, 1.6 micromol/L; DG, 14.7 micromol/L; genistein, 0.154 micromol/L; GG, 7.27 micromol/L. In human peripheral blood NK cells, genistein at <0.5 micromol/L and DG and GG at 0.1-10 micromol/L enhanced NK cell-mediated K562 cancer cell killing significantly (P < 0.05). At > 0.5 micromol/L, genistein inhibited NK cytotoxicity significantly (P < 0.05). The glucuronides only inhibited NK cytotoxicity at 50 micromol/L. Isoflavones, and especially the isoflavone glucuronides, enhanced activation of NK cells by interleukin-2 (IL-2), additively. At physiological concentrations, DG and GG were weakly estrogenic, and they activated human NK cells in nutritionally relevant concentrations in vitro, probably at a site different from IL-2 action. (+info)
An ultrastructural study of implantation in the golden hamster. II. Trophoblastic invasion and removal of the uterine epithelium.
Sixty six implantation sites from 18 golden hamsters were examined with light and electron microscopy between 4 and 5 1/2 days of pregnancy (post-ovulation). At 4 days some blastocysts began to invade the uterine epithelium, with trophoblastic processes penetrating and engulfing portions of the uterine epithelium. The majority of epithelial cells appeared normal before invasion, although at two implantation sites three or four adjoining epithelial cells were necrotic before penetration by the trophoblast. In general the epithelial cells were degenerating at the time the trophoblast invaded the epithelium. Inclusions, representing portions of the engulfed epithelium, and varying in size and electron density, were present throughout the invading trophoblast cells at 4 1/2 and 5 days of pregnancy. At 5 1/2 days the uterine epithelium had disappeared and the embryo was now almost completely surrounded by blood lacunae. (+info)
Molecular and cellular aspects of endometrial receptivity.
Endocrine and paracrine controls regulate the endometrium during the luteal phase of the cycle to permit implantation. Part of this differentiation process is the production of a specific secretion which fills the intrauterine cavity and glandular lumen. Its molecular composition originates from the gland secretion, from transudations from stroma, from the endometrial blood vessels, and last, but not least, from cellular components of apoptotic and exfoliated cells. We have studied the secretions of all phases during the menstrual cycle using patterns evaluated by SDS-PAGE, by laser densitometry or Western blots. Uterine secretion electrophoresis (USE) permits detailed analyses of the intrauterine micromilieu and allows clinical assessment of the receptive stage of endometrium during the luteal phase. Several individual protein bands have been defined as characteristic markers for such receptive pattern. We have isolated and identified the molecular structure of several of these proteins, e.g. histones, cyclophilin, transthyretin, haptoglobin and uteroglobin. Investigations on the endocrine regulation of these proteins, were carried out on the uterine secretions of patients treated with progesterone antagonists (mifepristone and onapristone). The results demonstrate how progesterone-dependent components produce a receptive pattern, which can serve as a useful and precise marker in the clinical diagnosis of the luteal phase. Essential progesterone-dependent components differentiating during the luteal phase may provide new targets for contraceptive interventions by preventing the physiological changes typical of receptivity. (+info)
Mucin expression and function in the female reproductive tract.
Reproductive tract epithelia are characterized by the presence of a thick, apical glycocalyx. This glycoprotein coat is drastically reduced in the uterus of many species during the time of embryo implantation. Recent studies indicate that mucin glycoproteins constitute a large proportion of the apical glycocalyx. One of these mucins, Muc-1, has particularly important functions at the luminal surface of the uterus and other female reproductive tract tissues. Muc-1 appears to play a dominant role in maintaining a functionally non-receptive uterine surface with regard to blastocyst attachment. Conversion to a receptive uterine state is brought about by the concerted actions of ovarian steroid hormones that in several species also strongly modulate Muc-1 protein and mRNA expression. Muc-1 also appears to serve a general function in protecting reproductive tract mucosa since Muc-1 null mice are particularly prone to bacterial infection. Collectively, these studies indicate that mucins, including Muc-1, play important barrier roles in reproductive processes and protection from bacterial pathogenesis in the female reproductive tract. (+info)
Human uterine lymphocytes.
During the luteal phase and the early months of pregnancy, there is a dense mucosal infiltration of CD56+ natural killer (NK) cells. These uterine NK cells have a phenotype (CD56bright, CD16-, mCD3-) which distinguishes them from peripheral blood NK cells (CD56dim, CD16bright, mCD3-). The uterine NK cells are in close association with extravillous trophoblast (EVT) cells which infiltrate into the decidua and maternal spiral arteries. This subpopulation of trophoblast expresses two human leukocyte antigen (HLA) class I molecules, HLA-G and HLA-C. Circulating NK cells express receptors for HLA class I molecules. We have recently found evidence that similar receptors are present on decidual NK cells belonging to both the Killer Inhibitory Receptor (KIR) and CD94 families. The repertoire of NK receptors expressed varies between different women. The findings indicate that decidual NK cells do have receptors for trophoblast HLA class I molecules. Experiments are underway to determine the effects of this interaction on NK cell function. (+info)
Myometrial zonal differentiation and uterine junctional zone hyperplasia in the non-pregnant uterus.
Human non-gravid myometrium differentiates in response to ovarian sex steroids into a subendometrial layer or junctional zone and an outer myometrial layer. Compared to the outer myometrial layer, the junctional zone myocytes are characterized by higher cellular density and lower cytoplasmic-nuclear ratio. These structural differences allow in-vivo visualization of the myometrial zonal anatomy by T2-weighted magnetic resonance (MR) imaging. The human myometrium is also functionally polarized. Video-vaginosonography studies have shown that propagated myometrial contractions in the non-pregnant uterus originate only from the junctional zone and that the frequency and orientation of these contraction waves are dependent on the phase of the menstrual cycle. The mechanisms underlying zonal myometrial differentiation are not known, but growing evidence suggests that ovarian hormone action may be mediated through cytokines and uterotonins locally released by the basal endometrial layer and endometrio-myometrial T-lymphocytes. Irregular thickening of the junctional zone due to inordinate proliferation of the inner myometrium, junctional zone hyperplasia, is a common MR finding in women suffering from menstrual dysfunction. Preliminary data suggest that junctional zone hyperplasia is further characterized by loss of normal inner myometrial function. Although irregular thickening of the junctional zone has been associated with diffuse uterine adenomyosis, the precise relationship between subendometrial smooth muscle proliferation and myometrial invasion by endometrial glands and stroma remains to be established. (+info)