Raymond D. Lambert

Expression of Cyclooxygenase-2 and Granulocyte-Macrophage Colony-Stimulating Factor in the Endometrial Epithelium of the Cow Is Up-Regulated During Early Pregnancy and in Response to Intrauterine Infusions of Interferon-τ

Abstract On the basis of results obtained in vitro, we previously proposed a model in which signals from the conceptus, namely interferon-tau (IFN-τ) and prostaglandin E2, increase the expression of cyclooxygenase (COX)-2 or granulocyte-macrophage colony-stimulating factor (GM-CSF) in immune and nonimmune cells of the bovine endometrium. Two experiments were conducted to verify the validity of this hypothesis in vivo. In experiment 1, the in vivo expression of COX-2 and GM-CSF during early pregnancy was monitored. Uteri from heifers were collected at different days (d) of the estrous cycle and pregnancy (P). In experiment 2, the effects of intrauterine infusions of IFN-τ on the expression of COX-2 and GM-CSF were analyzed. Immunohistochemistry was performed on uterine sections, and image analysis was used to evaluate the staining intensity in the conceptus, the luminal epithelium (LE), and the subepithelial stroma. In experiment 1, staining for COX-2 was maximal between d18P and d24P, both in the LE and in the conceptus, whereas staining for GM-CSF reached a plateau between d18P and d30P in the LE. In experiment 2, in response to IFN-τ, COX-2 was up-regulated in the LE of the ipsilateral horn, whereas GM-CSF was enhanced in both uterine horns. The current report supports the view that the conceptus, through its secretion of IFN-τ, stimulates maternal epithelial expression of COX-2 and GM-CSF during the peri-attachment period in the cow.

In vitro fertilization of bovine oocytes matured in vivo and collected at laparoscopy

Abstract Laparoscopy was used to collect 1618 in vivo-matured bovine follicular oocytes. Inexperienced laparoscopists encountered difficulties such as intestine perforation or failure to enter the peritoneal cavity. The recovery rate of oocytes was lower for beginners (53–59%) than for experienced laparoscopists (72–79%). 975 follicular oocytes were inseminated in vitro using spermatozoa exposed to a high ionic strength medium. Oocytes surrounded by expanded cumuli were fertilized and cleaved more frequently (P

Interferon-Tau Stimulates Granulocyte-Macrophage Colony-Stimulating Factor Gene Expression in Bovine Lymphocytes and Endometrial Stromal Cells

Abstract Interferon-tau (IFN-τ), the antiluteolytic signal produced by the trophoblast prior to implantation in ruminants, exhibits immunomodulatory properties. It stimulates the production of prostaglandin (PG) E2 in bovine endometrial cells via the induction of cyclooxygenase-2 (COX-2). We previously demonstrated that preconditioning lymphocytes with PGE2 increases the expression of granulocyte–macrophage colony-stimulating factor (GM-CSF), a cytokine that promotes conceptus growth and survival. Our goal in the present study was to evaluate the impact of IFN-τ on the expression of GM-CSF in bovine peripheral blood lymphocytes (PBL) and endometrial epithelial and stromal cells. Changes in PGE2 production and mRNA levels of COX-2 were also studied in PBL in response to IFN-τ. Gene expression was estimated by semiquantitative reverse transcription–polymerase chain reaction and Northern analysis. The expression of GM-CSF in PBL was stimulated by treatment with IFN-τ. Furthermore, GM-CSF mRNA levels were increased after preconditioning PBL for 3 days with IFN-τ, followed by a 12-h restimulation without IFN-τ. Inhibition rather than stimulation of PGE2 production and COX-2 expression in PBL during treatment with IFN-τ suggests a direct effect on GM-CSF expression. Moreover, GM-CSF expression was stimulated in uterine stromal cells in response to IFN-τ. This study provides the first evidence for stimulation of GM-CSF expression by IFN-τ in both leukocytes and endometrial stromal cells. In view of the role of GM-CSF on fetal growth and survival, these results support the hypothesis that the conceptus mediates accommodation mechanisms in the uterus during early pregnancy by modulating the expression of beneficial cytokines at the fetomaternal interface.

Endoscopy in cattle by the paralumbar route: Technique for ovarian examination and follicular aspiration

Abstract The ovary and follicular aspiration, or both, were observed in 50 heifers by endoscopy via the right paralumbar fossa. A total of 129 laparoscopies were performed. Eight animals underwent more than four interventions. Occasionally adhesions were observed, but they never interfered with ovary examination and follicular aspiration. The rate of ovum recovery was higher when a suction device was used rather than a syringe system (P

TGF-β2 and PGE2 in rabbit blastocoelic fluid can modulate GM-CSF production by human lymphocytes

PROBLEM: During normal pregnancy, major changes occur in the production of Th2/Th1 cytokines at the feto‐maternal interface. Th2 cytokines such as interleukin‐4 (IL‐4) or interleukin‐10 (IL‐10) are predominantly produced locally in the uterine and placental tissues, whereas the production of Th1 cytokines such as tumor necrosis factor alpha (TNF‐α) and interferon gamma (IFN‐γ) are decreased. Because these modulations might be induced by the embryo, the current study was carried out to test the effect of rabbit blastocoelic fluid on the production of Th2/Th1 cytokines by lymphocytes, and to investiate the possible implication of transforming growth factor β2 (TGF‐β2) prostaglandin E2 (PGE2) as modulators of the production of these cytokines.

Prediction of human ovulation by rapid luteinizing hormone (LH) radioimmunoassay and ovarian ultrasonography

Accurate prediction of the time of ovulation is essential for the recovery of a mature oocyte for in vitro fertilization (IVF). The preovulatory luteinizing hormone (LH) surge can be considered the most reliable hormonal change closely related to ovulation. With the use of a rapid LH assay and ovarian ultrasonography, it now appears possible to predict quite accurately the time of ovulation, provided that these technologies are appropriately applied. Therefore, we have used a newly standarized rapid (3-hour) LH radioimmunoassay (RIA) for serial estimation of preovulatory serum LH surge. So far, 21 women have come to our center daily during their 5-day preovulatory period. Blood samples were taken daily every 3 to 5 hours between 7:00 A.M. and 11:00 P.M. Ovarian ultrasonic scans were performed one to three times daily depending on the day of the cycle and the serum LH level. Data show that there is a significant variation in the mean preovulatory follicular size, in the duration of the LH surge, and in the time interval between the initial rise of LH surge and the estimated time of ovulation. Results obtained from six women studied during two to four cycles indicate that each woman has a distinctive hormonal and ultrasonic pattern that appears to be reproducible. Thus it is recommended that such a pattern be appropriately evaluated before attempting laparoscopy for recovery of a mature human oocyte.

CD8 Membrane Expression Is Down‐regulated by Transforming Growth Factor (TGF)‐β1, TGF‐β2, and Prostaglandin E2

PROBLEM: CD8 T‐cells are present at a lower frequency in human decidua than in peripheral blood. Because transforming growth factor (TGF)‐β2 down‐regulates CD4 membrane expression, its contribution, as well as the contribution of TGF‐β1 and prostaglandin (PG) E2, to the modulation CD8 expression was studied using human peripheral blood lymphocytes (PBLs).

TGFβ2 in Rabbit Blastocoelic Fluid Regulates CD4 Membrane Expression: Possible Role in the Success of Gestation

PROBLEM: During pregnancy, major changes occur in the decidual cell population. One of these changes involves some phenotypical transformations of lymphocyte sub‐populations. Since these variations might be due to the presence of the embryo, the current study was designed to investigate the implication of blastocoelic fluid (BF) in these changes and to determine the mechanism by which this phenomenon occurs.

Immunosuppressive effects of rabbit blastocoelic fluid and embryo culture medium.

In order to understand the mechanism of the feto-maternal immune relationship, we assayed the immunosuppression activities of fresh blastocoelic fluid and decomplemented peripheral serum collected from day-9 pregnant white New Zealand rabbits and of rabbit embryo culture medium (ECM). Because the viability of the human lymphocytes was not affected by either of these biological fluids and since they were easy to obtain in sufficient quantities, they were used uniformly in all the experiments. Immunosuppressive effect was calculated by the relative inhibition of proliferation of Con A-stimulated lymphocytes. The immunosuppressive effect of blastocoelic fluid of the 9-day pregnant rabbits was significantly higher than that of autologous decomplemented serum (P less than 0.001). The inhibition by the serum was non-specific because sera from non-pregnant animals as well as sera from different stages of pregnancy and pseudo-pregnancy showed the same level of inhibition. The ECM of 6.5-7-day-old embryo showed a pronounced immunosuppressive effect. When embryos of 1,3 and 5 days were cultured and their culture media were assayed only with 5-day-old embryo the effect had begun to appear, but it was far less than that of 7-day-old embryo (P less than 0.02). The suppressive activity of both the blastocoelic fluid and ECM was not due to cytotoxic effect, since this fluid supported the in vitro growth of single-cell rabbit embryos up to the stage of blastocyst. These results suggest that the immunologic tolerance of the embryo might be due to the immunosuppressors secreted by the embryo and that there might be a localized effect at the implantation site rather than a maternal systemic immunosuppressive effect.

Laparoscopic investigation of the bovine ovary in the periovulatory phase of the cycle

Laparoscopy, in combination with a rapid radioimmunoassay for plasma-LH determination, has been used to predict and observe ovulation in heifers. Experiments on three animals with typical progesterone levels, LH levels and apex formation are described. Ovulation was observed from 25 h to 29 h after the beginning of LH rise and from 17 h to 19 h after LH peak. The LH peak lasted for 9 h to 11 h. Cow ovulation was observed and photographed. The preovulatory follicle, apex formation, ovulation and freshly ruptured follicles are illustrated. The results presented here demonstrate that laparoscopy could offer valuable diagnostic assistance in clinical veterinary medicine.