Isolation and characterization of eight pregnancy-associated glycoproteins present at high levels in the ovine placenta between day 60 and day 100 of gestation

in Reproduction Nutrition Development (2004), 44(3, May-Jun), 169-181

Pregnancy-associated glycoproteins (PAG), structurally related to aspartic proteinases, are expressed in the outer epithelial cell layer (chorion/trophectoderm) of the ungulate placenta. The aim of the ... [more ▼]

Pregnancy-associated glycoproteins (PAG), structurally related to aspartic proteinases, are expressed in the outer epithelial cell layer (chorion/trophectoderm) of the ungulate placenta. The aim of the present study was to isolate as many PAG molecules as possible from placentae collected between day 60 and day 100 of gestation and to characterize their amino-terminal amino-acid sequences. Three heterologous radioimmunoassays were used to monitor PAG immunoreactivity throughout the isolation procedures. Sequential use of DEAE-cellulose, gel filtration, and CM ceramic chromatographies led to the isolation of several fractions rich in PAG immunoreactivity. The fractions with a large amount of proteins were also purified by chromatofocusing. The analysis of immunoreactive fractions by SDS-PAGE, Western blotting and two-dimensional electrophoresis followed by amino-terminal microsequencing on PVDF membranes allowed to identify eight different ovPAG with apparent molecular masses ranging from 55 to 66 kDa and isoelectric points from 4.0 to 6.8. The N-terminal sequences were determined and their comparison to those previously identified revealed that four of them are identical to those encoded by previously known cDNA, while the additional four sequences appear to be novel since they have not yet been described. [less ▲]

Short-term effects of spermine ingestion on the small intestine: a comparison of suckling and weaned rats

in Reproduction Nutrition Development (2004), 44(4, Jul-Aug), 353-364

We have previously shown that spermine, shortly after its ingestion, can induce the alteration of the morphology of the small intestine of suckling rats. It was proposed that this alteration is due to ... [more ▼]

We have previously shown that spermine, shortly after its ingestion, can induce the alteration of the morphology of the small intestine of suckling rats. It was proposed that this alteration is due to polyamine accumulation inside the epithelial cells. This could also be related to the fact that the intestine of the suckling rat is in an immature state. To shed light on this issue, disaccharidase and alkaline phosphatase activity assays, protein, DNA and RNA content measurements and polyamine concentration analysis were performed on the small intestine of suckling and weaned Wistar rats treated with spermine. Spermine did not induce the same intestinal alterations in weaned rats compared to suckling animals. Indeed, in sucklings, spermine administration induced a decrease of the protein, DNA, putrescine and spermidine intestinal content, suggesting a cell loss. The cell loss impaired the activity of intestinal enzymes: lactase, maltase and alkaline phosphatase. In weaned rats, the same treatment did not alter these parameters. Exogenous spermine by itself is not sufficient to induce the alterations described here and previously. The maturity degree of the small intestine could be the basis of this process. [less ▲]

Chorionic expression of heterogeneous products of the PAG (Pregnancy-Associated Glycoprotein) gene family secreted in vitro throughout embryonic and foetal development in the pig

in Reproduction Nutrition Development (2003), 43(6), 497-516

Porcine PAG (pPAG) are placental products of a multigene family that is strongly expressed in the chorionic epithelium (trophoblast and trophectoderm). The objective of this study was to define a pattern ... [more ▼]

Porcine PAG (pPAG) are placental products of a multigene family that is strongly expressed in the chorionic epithelium (trophoblast and trophectoderm). The objective of this study was to define a pattern of the pPAG proteins, secreted in vitro by chorionic explants harvested on 16-77 days of pregnancy. Trophoblastic and trophectodermal explants were collected from pregnant (PR) gilts (n = 27) and used for protein in vitro production (8-261 h). Endometrial explants of luteal-phase gilts (E10, n = 4) and pseudopregnant gilts (PsE, n = 2) were used as negative controls for protein immunoblotting. Proteins (PR, E10, PsE) were isolated mainly from incubation media, fractionated, dialysed and separated by SDS-PAGE. Heterogeneous Western blotting with various polyclonal anti-PAG sera raised against bovine or ovine antigens (anti-bPAG, or anti-oPAG) initially identified the pPAG proteins. Such blotting of fractionated chorionic proteins allowed for the isolation of porcine antigens that were employed as immunogens to raise several homologous antisera (anti-pPAG). Crude antisera were adsorbed on endometrial extracts or proteins of non-PR pigs, to remove non-relevant antibodies. The patterns of pPAG proteins secreted in Vitro varied throughout pregnancy (35-72 kDa). During implantation, similar to43 kDa (Day 16) or similar to68.1 kDa (Days 17-25) pPAG proteins were detected. During placentation and as pregnancy advanced (Days 31-77), similar to72.3 kDa pPAG proteins were observed. The secretions of parallel multiple smaller proteins (35.4-47.2 kDa), presumably, as forms of processed pPAG precursors, increased with the progress of gestation. In conclusion, the pPAG protein family plays a very important role during implantation, placenta formation and embryonic/foetal development in the pig. [less ▲]

Characterization of pregnancy-associated glycoproteins extracted from zebu (Bos indicus) placentas removed at different gestational periods

in Reproduction Nutrition Development (2002), 42(3), 1-15

In the present work, two biochemical approaches were used to characterize PAGs isolated from Bos indicus fetal cotyledons removed at different gestational ages. The first procedure included acidic and ... [more ▼]

In the present work, two biochemical approaches were used to characterize PAGs isolated from Bos indicus fetal cotyledons removed at different gestational ages. The first procedure included acidic and ammonium sulfate precipitations, anion and cation exchange chromatographies and the second included pepstatin-agarose affinity chromatography. A bovine PAG radioimmunoassay was used to monitor the immunoreactivity throughout the isolation procedures. The most immunoreactive fractions issued from cation exchange and affinity chromatographies were analyzed by SDS-PAGE and Western blotting, before transfer to a polyvinylidene difluoride (PVDF) membrane for NH2-microsequence determination. Use SDS-PAGE and Western blotting, different isoforms of PAG with apparent molecular masses of 51 to 69 kDa and isoelectric points varying from 4.4 to 6.7 were identified in the placentas from different gestational ages. N-terminal microsequencing (10 to 25 aa long) indicates the expression of one single terminal amino acid sequence in the Bos indicus placenta, which is 100% identical to the bovine PAG-1. [less ▲]

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

in Reproduction Nutrition Development (2001), 41(5), 401-412

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 ... [more ▼]

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. [less ▲]

Towards Marker Assisted Selection in Livestock

in Reproduction Nutrition Development (1999), 39(5-6, Sep-Dec), 555-61

In recent years, genomic tools have become available for most livestock species and are now being used routinely to map quantitative trait loci underlying the genetic variance for numerous economically ... [more ▼]

In recent years, genomic tools have become available for most livestock species and are now being used routinely to map quantitative trait loci underlying the genetic variance for numerous economically important traits. Fine-mapping methods are being devised to refine the initially coarse map positions of the quantitative trait loci to the point required for marker assisted selection and, eventually, the positional cloning of the underlying genes. Mapping information on QTL is beginning to be used to increase genetic response by enhancing genetic variance, selection accuracy, selection intensity and by reducing the generation interval. Optimal use of MAS will require the development of more robust methods for the routine genotyping of preimplantation embryos for multiple markers. [less ▲]

The Ovine Callipyge Locus: A Paradigm Illustrating the Importance of Non-Mendelian Genetics in Livestock

in Reproduction Nutrition Development (1996), 36(6), 651-7

An inheritable muscular hypertrophy was recently described in sheep and shown to be determined by the callipyge (CLPG) gene mapped to ovine chromosome 18. We demonstrate in this work that the callipyge ... [more ▼]

An inheritable muscular hypertrophy was recently described in sheep and shown to be determined by the callipyge (CLPG) gene mapped to ovine chromosome 18. We demonstrate in this work that the callipyge phenotype is characterized by a non-Mendelian inheritance pattern, referred to as polar overdominance, in which only heterozygous individuals having inherited the CLPG mutation from their sire express the phenotype. The possible role of parental imprinting in the determinism of polar overdominance is envisaged. [less ▲]

Evaluation of cryopreservation techniques for goat embryos

in Reproduction Nutrition Development (1995), 35

Evaluation des coefficients d'utilisation digestive des constituants du triticale par le belier adulte: comparaison de deux methodes.

in Reproduction Nutrition Development (1988), 28(1), 95-96

The regression method was compared with the difference method to determine the nutritive value of triticale in sheep. More accurate results were obtained with the regression method and with the difference ... [more ▼]

The regression method was compared with the difference method to determine the nutritive value of triticale in sheep. More accurate results were obtained with the regression method and with the difference method when the ration included at least 60 % of triticale. [less ▲]

Prévision de la quantité d'herbe ingérée par la vache laitière au pâturage. Comparaison de méthodes d'estimation de la digestibilité et du mode d'échantillonnage des fécès.

in Reproduction Nutrition Development (1988), 28(Suppl. 1), 61-62

Herbage intake of grazing dairy cows was estimated with several techniques. Grab sampling introduced an upwards bias in the estimate of faecal output comparison with field sampling. Estimates of OM ... [more ▼]

Herbage intake of grazing dairy cows was estimated with several techniques. Grab sampling introduced an upwards bias in the estimate of faecal output comparison with field sampling. Estimates of OM digestibility by NIR and faecal indexes methods were similar except with high level of concentrates. [less ▲]