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AL Mikkelsen and S Lindenberg

The aim of this study was to determine whether the rates of in vitro oocyte maturation, fertilization and cleavage, as well as implantation rate and pregnancy rate, could be improved by low-dose priming with FSH in vivo before retrieval of immature oocytes in patients with polycystic ovary syndrome (PCOS). From March 1998 to June 2000, a total of 28 women underwent 36 completed treatment cycles, randomized sequentially in one of two groups. Women in group 1 (n = 12 cycles) received no stimulation and women in group 2 (n = 24 cycles) received 150 iu recombinant FSH day(-1) for 3 days, initiated on day 3 after menstruation. Aspiration was performed transvaginally between day 9 and day 17 in the unstimulated group and on day 8 or day 9 in the FSH-primed group after FSH deprivation for 2 or 3 days. All cumulus-enclosed oocytes of healthy appearance were matured in culture medium (TCM-199) in vitro for 28-36 h before intracytoplasmic sperm injection (ICSI). After oocyte retrieval the women were given oestradiol (6 mg day(-1)) and progesterone administration (300 mg day(-1)) was initiated 2 days later. Suitable embryos (maximum two embryos) were transferred on day 3 after ICSI. The percentage of oocytes reaching metaphase II was significantly higher (P < 0.05) in the FSH-primed group (59%, 92/156) compared with the non-primed group (44%, 36/81). There were no significant differences in the rates of oocyte fertilization and cleavage between these groups. No pregnancies were obtained in group 1 (0%, 0/12), whereas seven clinical pregnancies were obtained in group 2 (29%, 7/24) (P < 0.05). In group 2, 37 embryo transfers resulted in eight implantations (21.6%). Three healthy singleton children have been born at term; the remaining pregnancies ended with spontaneous abortions in the first trimester. These results indicate that priming with recombinant FSH before harvesting of immature oocytes from patients with PCOS may improve the maturational potential of the oocytes and the implantation rate of the cleaved embryos.

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S. J. Kimber and S. Lindenberg

Summary. Ovariectomy and hormone replacement of mice were used to examine the hormonal control of expression on the uterine surface of a carbohydrate determinant (lacto-N-fucopentaose I, LNF I), involved in the initial interaction between the blastocyst and the endometrial epithelium at implantation. Pseudopregnant mice mated with sterile males were also used to elucidate the impact of embryonic signals on the expression of this antigen on the uterine surface. Two groups of fucosylated structures could be distinguished; one group was predominantly dependent on maternal oestrogen and progesterone, while the other group appeared to be less influenced by the hormonal milieu.

Keywords: implantation; uterus; hormones; carbohydrates; receptors; mouse

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S. Lindenberg, S. J. Kimber and E. Kallin

Summary. Mouse blastocysts bound LNF I conjugated to BSA-FITC or HSA-FITC and binding was inhibited by LNF I-HSA and to some extent by free LNF I, suggesting that the trophectoderm carries receptors specific for LNF I-like structures previously shown to be involved in implantation.

Keywords: embryo; implantation; neoglycoprotein; receptor; trophoblast, mouse

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AL Mikkelsen, E Host and S Lindenberg

The aim of this study was to investigate the incidence of apoptosis in granulosa cells from immature human follicles undergoing in vitro maturation (IVM) and to compare the incidence of apoptotic granulosa cells (i) between FSH-primed and unprimed normal ovaries and (ii) between polycystic and normal ovaries. Furthermore, the incidence of apoptosis was related to maturation and subsequent fertilization and cleavage of the oocytes from the corresponding ovary. Seventy women undergoing 70 IVM cycles were included. Group 1 consisted of patients with normal ovaries (n = 52) and group 2 consisted of patients with polycystic ovaries (n = 18). Patients in group 1 were subdivided into two groups according to priming with FSH before aspiration. In group 1a (n = 27 cycles) oocytes were obtained in unstimulated cycles. In group 1b (n = 25 cycles) oocytes were obtained after priming with recombinant FSH for 3 days initiated on day 3 after spontaneous menstruation. In group 2 all patients were primed with recombinant FSH for 3 days before aspiration. Aspiration was performed transvaginally and cumulus-enclosed oocytes were matured for 28-30 h before fertilization. Granulosa cells were collected from follicular aspirates. An APOPTAG detection kit was used to stain the granulosa cells and to detect apoptosis. The incidence of apoptosis in granulosa cells was decreased in follicles from FSH-primed normal ovaries compared with follicles from unprimed normal ovaries and FSH-primed polycystic ovaries. No difference was found between granulosa cells from FSH-primed polycystic ovaries and granulosa cells from unstimulated normal ovaries. No differences in maturation rate, fertilization rate, cleavage rate and implantation rate were observed when oocytes from a polycystic ovary were compared with oocytes from an unstimulated normal ovary. In unstimulated cycles, the ovaries were grouped according to the presence of a dominant follicle. The incidence of apoptosis was significantly higher in granulosa cells from an ovary without a dominant follicle compared with granulosa cells from an ovary with a dominant follicle. The rates of maturation, fertilization and cleavage did not differ between the two groups.

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S. Lindenberg, K. Sundberg, S. J. Kimber and A. Lundblad

Summary. Seven oligosaccharides isolated from human milk were tested for their effect in an in-vitro model of mouse blastocyst adhesion and trophoblast outgrowth on endometrial epithelial monolayers. One compound, lacto-N-fucopentaose I (LNF I), produced a significant reduction in the percentage of attached and outgrown blastocysts after co-culture for 72 h (P < 0·001). No significant effect of any other tested oligosaccharide was obtained.

Keywords: trophectoderm; uterine–epithelial cells; blastocyst; oligosaccharides; attachment; in vitro

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U. Bentin-Ley, B. Pedersen, S. Lindenberg, J. Falck Larsen, L. Hamberger and T. Horn

A cell culture system was established in which endometrial stromal cells were embedded in a collagen matrix and separated from the endometrial epithelium by basement membrane material (Matrigel). The epithelium, seeded on top of the collagen matrix, grew in a monolayer. The cultures were evaluated by light microscopy and transmission and scanning electron microscopy. Light and transmission electron microscopy indicated a polarized columnar epithelium in monolayer with basally positioned nuclei. Scanning electron microscopy revealed a confluent epithelium with an abundance of microvilli and cilia, as well as pinopodes on the apical surface. Immunohistochemical staining for cytokeratin confirmed the epithelial origin of the surface cells, and staining for human collagen IV demonstrated its presence underneath the epithelial cells. This culture system represents a three-dimensional system that imitates the normal endometrium.