From the germinal cells to the newborn animal: the transmission of genes and life through the generations.

The technology of reproduction progressed considerably during the last decade, leading to a certain availability of in vitro methods for fertilisation, oocyte maturation and embryo culture. The most spectacular manipulations are cloning and transgenesis. This review focuses on the early appearance of germinal cell precursors and the long-standing fate of gametes in mammals. The evident complexity and long-term programming of events in gametes and early embryos explain part of the difficulties encountered during the development of in vitro and in vivo methods such as multiple ovulation and embryo transfer (MOET), oestrus synchronisation, ovulation induction, superovulation, in vitro maturation and fertilisation, cryopreservation, transgenesis, nuclear transfer and cloning) and the occurrence of unexpected alterations of development, e.g. embryonic or fetal mortality, large-weight newborn syndrome and other dysregulations in imprinting or DNA transmission.

Pregnancy associated glycoproteins in ruminants: inactive members of the aspartic proteinase family.

The Pregnancy Associated Glycoproteins (PAGs) presented in this paper are largely expressed in the ruminant placenta. These proteins are classified as probably inactive members of the aspartic proteinase family. Pepsinogen, renin, cathepsin E & D and chymosine are typical members of this family, characterised by the presence of aspartic acids boarding the recognition sites. Secreted in the peripheral blood of the pregnant female from early pregnancy, these proteins can be used in serological tests for establishing different diagnoses. In the veterinary practice, these diagnoses are useful for both pregnancy confirmation and follow-up of trophoblastic function. The first aspect can help breeders in the management of reproduction, while the second one more specifically concerns clinicians and researchers wishing to establish a differential diagnosis of pathologic conditions affecting pregnancy.

Myeloid hypoxia-inducible factor 1α prevents airway allergy in mice through macrophage-mediated immunoregulation.

Hypoxia-inducible factor (HIF) has important roles in promoting pro-inflammatory and bactericidal functions in myeloid cells. Conditional genetic ablation of its major subunit Hif1α in the myeloid lineage consequently results in decreased inflammatory responses in classical models of acute inflammation in mice. By contrast, we report here that mice conditionally deficient for Hif1α in myeloid cells display enhanced sensitivity to the development of airway allergy to experimental allergens and house-dust mite antigens. We support that upon allergen exposure, MyD88-dependent upregulation of Hif1α boosts the expression of the immunosuppressive cytokine interleukin (IL)-10 by lung interstitial macrophages (IMs). Hif1α-dependent IL-10 secretion is required for IMs to block allergen-induced dendritic cell activation and consequently for preventing the development of allergen-specific T-helper cell responses upon allergen exposure. Thus, this study supports that, in addition to its known pro-inflammatory activities, myeloid Hif1α possesses immunoregulatory functions implicated in the prevention of airway allergy.

Evolution of anti-eCG antibodies in response to eCG doses and number of injections. Relationship with productivity of rabbit does.

The aim of this experiment was to study the kinetics of anti-eCG (equine chorionic gonadotrophin) antibodies in relation to eCG dose (8 or 25 IU) and number of injections (n = 11) in comparison with a control group (no injection), and to relate antibody production to sexual receptivity and productivity of rabbit does. In all, 124 lactating primiparous rabbit does were inseminated every 35 days for a year. Just before eCG injection (48 h before insemination), blood samples were collected from all the does to assay anti-eCG antibodies. The anti-eCG antibody binding rate, regardless of the injected dose, shows that none of the does developed detectable anti-eCG antibodies before the 7th injection. The level of detectable anti-eCG antibodies began to show an increase at the 7th injection and was significant only for the 25 IU dose at the 11th injection. At the end of the experiment, 15% and 39% of does treated with 8 and 25 IU, respectively, developed immunity to eCG (binding rate >6%: higher binding rate of the control group). Consequently, the immune response depends on the eCG dose and on the number of injections. Moreover, productivity of does estimated from the number of weaned rabbits produced per insemination is not influenced by the level of eCG antibodies (7.0 and 6.9 for binding rate <6% and binding rate 6%, respectively). Only 19 inseminations (n = 6 and n = 13 for 8 and 25 IU, respectively) were made on hyperimmune does. Consequently, the immune response to eCG seems to be marginal for rabbit does. Moreover, under the described experimental conditions, reproductive performances of hyperimmune does were not affected.

Influence of the Ixodes ricinus tick blood-feeding on the antigen-specific antibody response in vivo.

The blood meal of hard ticks such as Ixodes ricinus lasts several days. This is made possible by tick salivary factors that inhibit inflammation, haemostasis and the host immune response. We assessed the latter on a model of immune response in vivo. A significant reduction of specific IgM and IgG levels was observed in BALB/c mice infested 5 days before injection with bovine serum albumin (BSA) and QuilA but not in mice infested 5 days after the immunization. This effect was not observed in mock-infested mice and could not be attributed to the use of anesthetics. The antibody response was not merely delayed and the Th(1)/Th(2) balance appeared not altered. T-dependent zones and germinal centers in lymph nodes draining the tick bite site showed no apparent morphological alterations or shift in T cell subpopulations. However, the spleens of tick-infested mice had also an enlarged red pulp, indicating an increased extramedullary haematopoietic activity.

Anti-PMSG antibody levels in sheep subjected annually to oestrus synchronisation.

Estimation of the long-term consequences on reproduction performance of the oestrus synchronisation treatments that are annually applied to ewes was carried out on nine officially controlled dairy flocks in the Roquefort region of France. A hormonal treatment combining the insertion of a vaginal fluoro-gestone acetate (FGA) sponge for 14 days and the injection of about 500 IU of pregnant mare serum gonadotropin (PMSG) at withdrawal was applied to the ewes in seven of the nine flocks. The ewes in the two other flocks were used as controls. Blood samples were taken from each female just before the treatment (to test for the presence of residual antibodies) and 20 days after the PMSG injection. Anti-PMSG antibody binding rates were calculated for each blood sample. The residual binding rate increased with age and induce negative effects on the following years reproduction performances, i.e., they increased the probability that the ewes would not become pregnant.

Comparison of the ability of three radioimmunoassay to detect pregnancy-associated glycoproteins in bovine plasma.

CONTENTS: Pregnancy-associated glycoproteins (PAGs) constitute a large family of glycoproteins that are synthesized in the superficial layer of the ruminant placenta according to a spatial and temporal expression pattern. When PAGs are released in the maternal blood they can be used for pregnancy diagnosis, pregnancy follow-up and for the monitoring of the trophoblastic function. Three different radioimmunoassay systems (RIA 1, RIA 2 and RIA 3) using antisera produced against PAG I67 (RIA 1), PAG55+62 (RIA 2) and PAG55+59 (RIA 3) were used in this investigation in order to measure the PAG concentration in plasma samples withdrawn from pregnant cows and heifers during different periods following artificial insemination (AI). These systems were able to detect PAG molecules in the maternal blood as early as 21 days after AI in different concentrations (RIA 1: 0.43 +/- 0.24 ng/ml, mean +/- SD; RIA 2: 0.48 +/- 0.24 ng/ml; RIA 3: 0.64 +/- 0.37 ng/ml). On days 32 and 42 RIA 2 (4.30 +/- 1.32 ng/ml and 5.56 +/- 1.95 ng/ml) and RIA 3 (4.17 +/- 1.15 ng/ml and 5.60 +/- 1.89 ng/ml) presented significantly (p < 0.0001) higher PAG concentrations than those of RIA 1 (2.43 +/- 0.81 ng/ml and 4.01 +/- 1.48 ng/ml), respectively. After day 21, significant correlations (p < 0.0001; r >/= 0.929) were determined between the three systems. Additionally the three individual PAG profiles presented in this study showed that PAG molecules secreted in the maternal blood between 21 and 50 days after AI were better recognized by the RIA 2 and RIA 3 systems. This study clearly indicated that the ability of a RIA test to recognize PAG molecules in the maternal blood can be improved by carefully selecting the antiserum.

Aspartic proteinase members secreted by the ruminant placenta: specificity of three radioimmunoassay systems for the measurement of pregnancy-associated glycoproteins.

Pregnancy-associated glycoproteins (PAGs) isolated from the placenta of various ruminant species are enzymatically inactive members of the aspartic proteinase family. The measurement of these proteins in the maternal blood can be a good indicator of the presence of a live embryo. As certain aspartic proteinases are present in biological fluids in physiological and pathological conditions at various concentrations, it was necessary to determine the specificity of three radioimmunoassay (RIA) systems currently used for the detection of PAG molecules. Commercially available members of the aspartic proteinase family like pepsinogen, pepsin, chymosin, rennet, cathepsin D and renin were tested in a wide concentration range (10 ng/ml - 1 mg/ml). Pepsinogen cross-reacted in RIA 1, RIA 2 and RIA 3 over 1 mg/ml, 50 microg/ml and 500 microg/ml concentrations, respectively. In the presence of pepsin, cross-reaction was observed in RIA 1, RIA 2 and RIA 3 over 1 mg/ml, 500 microg/ml and 1 mg/ml concentrations, respectively. Chymosin and rennet could cross-react in RIA 2 and RIA 3, while renin and cathepsin D did not decrease the binding of the tracer to antisera more, than that of the minimal detection limit. As the plasma/serum concentrations of the examined aspartic proteinases reported in the literature were outside the concentration range where cross-reaction was observed, it can be concluded that these RIA systems were specific for the detection of PAGs in biological fluids.

Increase of plasma eCG binding rate after administration of repeated high dose of eCG to cows.

Equine chorionic gonadotrophin (eCG) is still used to promote follicular growth in cattle and, more recently with an increased frequency of administration, in ovum pick-up protocols. The aim of this experiment was to verify the possible effect of high frequency of administration on the immune response to eCG. The profiles of eCG binding rate, in the blood of two groups (A, B) of 4 primiparous cross breed beef cows (3-3.5 years old) submitted weekly for 5 to 10 weeks to repeated high doses (1000-2000 IU) of equine chorionic gonadotrophin, are presented in this paper. A sensitive radiometric method was used to detect antibodies in plasma. The profiles clearly indicated a marked increase of eCG binding rate after 3 to 5 injections of the exogenous hormone to the females. The statistical analysis of the results established that treatments induced a significant increase (P < 0.01) in binding rates after 6 and 3 injections in group A and B respectively. These binding rates remained elevated for at least 1 week following the last injection and decreased afterwards. The values of plasma binding rates following repeated eCG administration differed significantly between groups (0.90+/-1.04 and 1.04+/-0.11 for groups A and B before treatment versus 11.77+/-0.92, 6.70+/-0.85 for groups A and B after treatment, P < 0.01) and from one cow to another (P < 0.01) with some cows presenting no significant immune response while others were more reactive against the hormone (at least 3 injections).

Four years of induction/synchronization of estrus in dairy goats: effect on the evolution of eCG binding rate in relation with the parameters of reproduction.

Ninety-eight Alpine goats of two herds were followed over 4 years in a program of annual artificial insemination after estrus induction/synchronization, including progestagen administration (vaginal sponge) followed by prostaglandin analog and equine chorionic gonadotrophin (eCG) 48 h before sponge removal. Goats were sampled every 4 hours from the 16th to the 56th following sponge removal, for determination of LH surge and tested for estrus by the presence of a buck. Seven days after AI, endoscopic examination of the ovaries was performed to determine the number of corpus lutea. Pregnancy diagnosis was performed at day 21-22 post AI by determination of plasma progesterone and at day 40-45 by ultrasonography. Parturition, number and sex of kids were recorded. All the goats were sampled before and after each treatment, for anti-eCG antibodies screening. Statistical analysis of the results clearly established a significant effect of the treatments on anti-eCG antibodies. Time of estrus and LH surge were significantly different between herd. The antibodies significantly delayed the time of coming out of estrus as well as the time of LH surge. Two antagonistic effects were evidenced: first, the delayed of time of estrus and time of LH surge in relation with the immune reaction to eCG; secondly, the ahead of time of estrus and time of LH surge during the years of treatment, identical to both herd. The antibodies negatively influenced the percentage of ovulating females as well as kidding rate. Finally, no effect of antibodies on prolificacy was found.