No data are available from human embryos during the peri-attachment period, and future studies should investigate the presence of GnRH and receptor during human embryonic development. Nevertheless, after embryo-endometrial adhesion, as the embryo invades the uterine endometrial stroma, trophoblastic differentiation into cyto- and syncytiotrophoblast layers occurs, and both are known to produce GnRH as well as to express the GnRH receptor in the first-trimester placenta.
The role of GnRH in controlling placental hCG production and secretion has been fully demonstrated both in vitro and in vivo, especially in first-trimester placenta. The stimulatory effect appears to be receptor-dependent, since GnRH antagonist (GnRH-ant) blocks both the GnRH- and GnRH-a-induced effects. Furthermore, GnRH-ant reduces the amplitude of spontaneous placental hCG pulses, suggesting a direct blockage of endogenous placental GnRH. Recently, an antibody against GnRH has been described in the maternal circulation of pregnant women with previous miscarriages and low levels of hCG, reinforcing the important role of GnRH in human implantation and placentation.
This enigmatic process is the result of an embryonic-maternal dialogue, in which the embryo and the endometrium induce changes in each other to promote receptivity. Growth factors and cytokines are secreted by the developing blastocyst to enhance uterine receptivity, directly by influencing the endometrial epithelial cells to undergo cellular changes such as down-regulation of cell polarity, or indirectly by stimulation of ovarian steroidogenesis through hCG, until placental progesterone production is sufficient to maintain the continuing pregnancy. buy yasmin online
We have shown for the first time that both GnRH and its receptor are expressed at the mRNA level in vivo by the human endometrium throughout the entire menstrual cycle of fertile patients. Isolated endometrial stromal and epithelial cells expressed both GnRH and its receptor, with greater GnRH mRNA expression in stromal cells compared to epithelial cells during proliferative phase. However, increased epithelial expression of GnRH mRNA occurred during the luteal phase. This is consistent with the immunohistochem-ical data, which showed affinity-purified antibody to GnRH bound to the endometrial stroma of each section of every patient studied both in the follicular and luteal phases of the menstrual cycle, with the most intense reactivity in the luteal phase. We also localized the GnRH staining to both the glandular and luminal epithelium, with increased signal in the luteal phase.
To examine the quantitative amount of GnRH mRNA in the endometrial cells from individual subjects throughout the menstrual cycle, we coamplified the patient sample for 40 cycles in the presence of defined amounts of internal standard cDNA for GnRH. Figure 2 shows the standard curve obtained for GnRH mRNA by plotting the logarithmically transformed ratios of the densities of target cDNA to competitive cDNA against the log amount of initially added target cDNA in each PCR. The quantitative PCR of the different endometrial samples studied throughout the menstrual cycle showed a progressive increase in the mRNA levels from the early proliferative phase to the midluteal phase, which was statistically significant (p < 0.05) between these two phases (Fig. 4).
RT-PCR was employed to increase the sensitivity of detection, and a sequence of 399 bp of GnRH mRNA was amplified in all the endometrial samples analyzed from fertile women in both the follicular and luteal phases of the menstrual cycle. buy ortho tri-cyclen online
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Five-microliter aliquots from each RT product were mixed with 95 ^l of the PCR-mastermix described above with 4 ^l 3′ + 5′ primer-mix (5 ^M each) for p-actin. Program parameters were 30 cycles of 94°C for 45 sec, 54°C for 45 sec, and 72°C for 60 sec. For this PCR, no competitor was added. flovent inhaler