Localization and distribution of tissue type and urokinase type plasminogen activators and their inhibitors type 1 and 2 in human and rhesus monkey fetal membranes

Fetal membranes consist of 10 distinct layers including components of amnion, chorion and decidua, the latter being of maternal origin. They form mechanically integrated sheets capable of retaining amniotic fluid and play an essential role in protecting fetal growth and development in the pregnant uterus. The extracellular matrix, substrate for plasminogen activators (PAs), is an important supportive framework of the fetal membranes. :Fetal membranes from women with preterm premature rupture of membranes may differ in their protease activity compared with normal membranes. To identify the presence of PAs and their inhibitors (PAI) and their possible role in the process of fetal membrane rupture, this study in investigated the distribution and localization of both protein and mRNA for tissue (t) and urokinase (u) PA and their inhibitors type 1 (PAI-1) and type 2 (PAI-2) in amniochorion of human and rhesus monkey using conventional and. confocal immunofluorescence microscopy. In situ hybridization analysis showed that the distribution and localization of mRNAs for tPA, uPA, PAI-I and PAI-2 were similar in the fetal membranes of human and rhesus monkey; no obvious species difference was observed. Evidence of tPA mRNA was detected in amniotic epithelium, trophoblast cells and nearly all cells of the decidual layer. Strong expression of uPA mRNA was noted in the decidual cells which increased in intensity as the abscission point was approached. Weak staining in chorion laeve trophoblast was also detected. In situ hybridization experiments showed PAI-1 mRNA to be concentrated mainly in the decidual cells, some of which were interposed into the maternal-facing edge of the chorion laeve. Maximal labelling of the decidua occurred towards the zone of abscission. Weak expression of PAI-1 mRNA nas also noted in some cells of the chorion laeve. The distribution of PAI-2 mRNA in amniochorion was also concentrated in the cells of the decidual layer, maximum expression of the mRNA was in the level of abscission. No detectable amount of mRNAs for tPA, uPA, PAI-1 and PAI-2 was found in the fibroblast, reticular and spongy layers. Distribution of the proteins of tPA, uPA and PAI-1 in the fetal membranes of these two species was consistent with the distribution of their mRNA. Anti-PAI-2 immunofluorescence was found to be strongly concentrated in the amniotic epithelium, but PAI-2 mRNA was negative in this layer, suggesting that the epithelium-associated PAI-2 is not of epithelial origin. These findings suggest that a local fibrinolysis in fetal membranes generated by precisely balanced expression of PAs and their inhibitors via paracrine or autocrine mechanisms may play an essential role in fetal membrane development, maturation and in membrane rupture. Following an analysis of the distribution and synthesis of activators and inhibitors it was found that they may play a role in abscission during the third stage of labour. (C) 1998 W. B. Saunders Company Ltd.

Os histricomorfos sul-americanos: uma análise comparativa do desenvolvimento embriológico

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Desenvolvimento e composição química do útero gravídeo, da glândula mamária e as mudanças corporais em cabras durante a gestação

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Desenvolvimento do útero grávido e da glândula mamária e mudanças corporais em cabras leiteiras durante a gestação

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Transporte placentário via cavéola na placenta de bovinos clonados e transgênicos

Pós-graduação em Ciência e Tecnologia Animal - FEIS

Proliferação celular em gestações naturais e de conceptos bovinos transgênicos e clonados, que expressam a proteína fluorescente verde (GFP)

Pós-graduação em Ciência e Tecnologia Animal - FEIS

Caracterização das proteínas caveolinas -1 e -2 na placenta de conceptos bovinos clonados transgênicos

The transgenic application of green fluorescent protein (GFP) as fetal cell marker on cattle cloned placenta could provide an exclusive model for studying the morphologic and immunologic maternal-fetal interactions, providing information about its mapping, distinguishing the fetal from maternal cells. This model will have direct application, mainly because these animals present problems during its development. With this model's support, we intend to verify the substances transport between mother and fetus during endocytosis, through the immunolocalization of protein named caveolae. For these, we used 06 cloned bovine and 30 cattle samples of artificial insemination (AI) with 90 days of pregnancy, which had been their development interrupted by humanitarian slaughter of the recipient and recovery of the pregnant uterus. We collected the placentome and the chorion. A part of the samples was cut and fixed, by immersion, on a solution containing 4% of parafomaldehyde or 10% of formaldehyde on a sodium phosphate buffer (PBS), at 0,1 M pH 7.4, Zamboni solution (4% of paraformaldehyde, 15% of picric acid, on sodium phosphate buffer 0,1 M pH 7.4), metacarn (60% of metanol, 30% of chloroform, and 10% glacial acetic acid), for morphologic and immunohistochemistry verification for caveolinas proteins -1 and -2 (CAV -1 and CAV-2). The caveolins -1 were found in fetal and maternal villi, but its strongest staining was observed in the endometrial stroma. The caveolins -2 had positive staining in trophoblast and chorioallantoic membrane, and specifically in giant trophoblastic binucleated cell. Therefore the results were compared between cloned cattle and from AI or natural mating, for assisting on detection of the reason of many placental alterations, embryonic losses, spontaneous abortion, post-natal mortality and large offspring syndrome on laboratory-manipulated animals. The result suggests that the proteins caveolins -1 and -2 (CAV-1 and CAV-2) are part of the caveolae composition and important structures related to the molecule transfer to the fetus, nourish it through endocytosis and pinocytosis.

Apoptose em placentônios bovinos de gestações de conceptos naturais e de transgênicos clonados

Pós-graduação em Ciência e Tecnologia Animal - FEIS

Progesterone (PR), Oestrogen (ER-alpha and ER-beta) and Oxytocin (OTR) Gene Expression in the Oviduct and Uterus of Pregnant and Non-pregnant Bitches

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anatomical modelling of the pregnant radiotherapy patient

This study aimed to take existing anatomical models of pregnant women, currently used for radiation pro-tection and nuclear medicine dose calculations, and adapt them for use in the calculation of fetal dose from external beam radiotherapy (EBRT). The models investigated were ‘KATJA’, which was provided as an MCNPX geometry file, and ‘RPI-P6’, which was provided in a simple, voxelized bina-ry format. In-house code was developed, to convert both mod-els into an `egsphant’ format, suitable for use with DOSXYZnrc. The geometries and densities of the resulting phantoms were evaluated and found to accurately represent the source data. As an example of the use of the phantoms, the delivery of a cranial EBRT treatment was simulated using the BEAMnrc and DOSXYZnrc Monte Carlo codes and the likely out-of-field doses to the fetus in each model was calculated. The results of these calculations showed good agreement (with-in one standard deviation) between the doses calculated in KATJA and PRI-P6, despite substantial anatomical differ-ences between the two models. For a 36 Gy prescription dose to a 233.2 cm3 target in the right brain, the mean doses calcu-lated in a region of interest covering the entire uterus were 1.0 +/- 0.6 mSv for KATJA and 1.3 +/- 0.9 mSv for RPI-P6. This work is expected to lead to more comprehensive studies of EBRT treatment plan design and its effects on fetal dose in the future.

Functional and molecular characterization of Tachykinins and tachykinin receptors in the mouse uterus

The aim of this study was to analyze the function and expression of tachykinins, tachykinin receptors, and neprilysin (NEP) in the mouse uterus. A previous study showed that the uterotonic effects of substance P (SP), neurokinin A (NKA), and neurokinin B (NKB) in estrogen-treated mice were mainly mediated by the tachykinin NK, receptor. In the present work, further contractility studies were undertaken to determine the nature of the receptors mediating responses to tachykinins in uteri of late pregnant mice. Endpoint and real-time quantitative RTPCR were used to analyze the expression of the genes that encode the tachykinins SP/NKA, NKB, and hemokinin-1 (HK-1) (Tac1, Tac2, and Tac4); and the genes that encode tachykinin NK1 (Tacr1), NK2 (Tacr2), and NK3 (Tacr3) receptors in uteri from pregnant and nonpregnant mice. The data show that the mRNAs of tachykinins (particularly NKB and HK-1), tachykinin receptors, and NEP are locally expressed in the mouse uterus, and their expression changes during the estrous cycle and during pregnancy. The tachykinin INK, receptor is the predominant tachykinin receptor in the nonpregnant and early pregnant mouse and may mediate tachykinin-induced uterine contractions in the nonpregnant mouse. The tachykinin NK, receptor is predominant in the late pregnant mouse and is the main receptor mediating uterotonic responses to tachykinins at late pregnancy. The tachykinin NK, receptor is expressed in considerable amounts only in uteri from nonpregnant diestrous animals, and its physiological significance remains to be clarified.

Effect of Oral Supplementation of the Linoleic and Gammalinolenic Acids on the Diabetic Pregnant Rats

The aim of this work was to evaluate the direct protective action of oral fatty acid supplementation against the deleterious effect of hyperglycemia on maternal reproductive outcomes; fetal growth and development on female Wistar rats. The animals were distributed into four experimental groups: G1= non-diabetic without supplementation (Control group); G2= non-diabetic treated with linoleic (LA) and gammalinolenic acid (GLA) (1 mL of Gamaline-V/day); G3= diabetic without supplementation and G4= diabetic treated with LA and GLA. Diabetes was induced by streptozotocin (40 mg/kg). At day 21 of pregnancy, the gravid uterus was weighed and dissected to count the dead and live fetuses, resorption, implantation, and corpora lutea numbers. The fetuses were analyzed for external and internal anomalies. The treatment with Gamaline-V supplementation to diabetic rats interfered in the maternal reproductive outcome (reduced number of live fetuses and embryonic implantation); however, it protected the deleterious on the incidence of congenital anomalies caused by hyperglycemia.

Uterine and luteal hemodynamic evaluation of the non pregnant mare

Background: Studies with Doppler ultrassonography started at the end of the 90s for the determination of physiological and pathological alterations in the reproductive tract of the mare. Uterine alterations caused by inflammation, response from seminal plasma infusion, hormonal variations during estrous and diestrus, pregnancy and action of various vasoactive factors influence on the vascular perfusion detected by Doppler ultrasound. The development of efficient methods for uterine quality evaluation is of big importance for field equine reproduction veterinarians, once uterine environment is responsible for pregnancy maintenance. Review: Nowadays, the most used methods of uterine evaluation are the mode B ultrassonography, cytology, culture and biopsy. Hemodynamic evaluation of the uterus can be done by spectral data collected from large vessels, as A. uterine and its ramifications, or from subjective or objective evaluations from endometrium, miometrium and mesometrium attachment, which provide data referent to local and specific alterations of the evaluated area. Alterations in uterine vascular perfusion has been detected during estrous cycle, during pregnancy and in cases of infusion of inflammatory substances. These alterations happen because of vasoactive substances that act in the uterus during these events, however, most of these vasoactive substances are probably not even known. Also, important hemodynamic alterations in old mares, as an increase in vascular resistance, have been described. This increase might result from fibrosis of the uterus and in women it is considered to be a cause of infertility. In mares, periglandular fibrosis of the endometrium is considered to be the major diagnosable cause of embryonic and fetal loss in older mares. For the CL, ovarian artery of the mare supplies the ovary as well as the oviduct and therefore can be used for evaluation of these areas. The CL evaluation can also be done by the percentage of luteum area with colored signals as an indicator of the extent of blood flow. The percentage of the CL area with colored signals is determined subjectively by images observations in real time and/or by a freezing Power Doppler cross-section image with the maximum number of color pixels taped and the total number of color pixels is assessed by a computer analysis system. Therefore, a high correlation between plasmatic progesterone and CL vascularization also allows the CL evalution by this technique. In a first report, CL circulation reached its maximum on D5, the progesterone concentration in peripheral blood increased until D7 and in a posterior report, maximum perfusion was achieved two days after the maximum progesterone concentration (D8). Blood flow reduced between D10-D14 some days before the plasma progesterone decrease and, during the luteolytic period (D15-D17), the decline in CL blood-flow area was greater than blood flow decrease. Conclusion: Doppler ultrassonography add knowledge about uterine viability and CL functionality can be easily used by veterinarians in the field. It is a noninvasive method that provides real time results. However, because of the short time studies in this area have been done, many other answers still need to be found until normal and pathological patterns will be established.

Mineral metabolism in singleton and twin-pregnant dairy goats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)