Tissue-specific profiling reveals modulation of cellular and mitochondrial oxidative stress in normal- and low-birthweight piglets throughout the peri-weaning period.

Weaning is known to induce important nutritional and energetic stress in piglets. Low-birthweight (LBW) piglets, now frequently observed in swine production, are more likely to be affected. The weaning period is also associated with dysfunctional immune responses, uncontrolled inflammation and oxidative stress conditions that are recognized risk factors for infections and diseases. Mounting evidence indicates that mitochondria, the main cellular sources of energy in the form of adenosine 5' triphosphate (ATP) and primary sites of reactive oxygen species production, are related to immunity, inflammation and bacterial pathogenesis. However, no information is currently available regarding the link between mitochondrial energy production and oxidative stress in weaned piglets. The objective of this study was to characterize markers of cellular and mitochondrial energy metabolism and oxidative status in both normal-birthweight (NBW) and LBW piglets throughout the peri-weaning period. To conduct the study, 30 multiparous sows were inseminated and litters were standardized to 12 piglets. All the piglets were weighted at day 1 and 120 piglets were selected and assigned to 1 of 2 experimental groups: NBW (n = 60, mean weight of 1.73 ± 0.01 kg) and LBW piglets weighing less than 1.2 kg (n = 60, 1.01 ± 0.01 kg). Then, 10 piglets from each group were selected at 14, 21 (weaning), 23, 25, 29 and 35 days of age to collect plasma and organ (liver, intestine and kidney) samples. Analysis revealed that ATP concentrations were lower in liver of piglets after weaning than during lactation (P < 0.05) thus suggesting a significant impact of weaning stress on mitochondrial energy production. Oxidative damage to DNA (8-hydroxy-2'-deoxyguanosine, 8-OHdG) and proteins (carbonyls) measured in plasma increased after weaning and this coincides with a rise in enzymatic antioxidant activity of glutathione peroxidase (GPx) and superoxide dismutase (SOD) (P < 0.05). Mitochondrial activities of both GPx and SOD are also significantly higher (P < 0.05) in kidney of piglets after weaning. Additionally, oxidative damage to macromolecules is more important in LBW piglets as measured concentrations of 8-OHdG and protein carbonyls are significantly higher (P < 0.05) in plasma and liver samples, respectively, than for NBW piglets. These results provide novel information about the nature, intensity and duration of weaning stress by revealing that weaning induces mitochondrial dysfunction and cellular oxidative stress conditions which last for at least 2 weeks and more severely impact smaller piglets.

Whole-body propionate and glucose metabolism of multiparous dairy cows receiving folic acid and vitamin B12 supplements.

This study was undertaken to evaluate the effect of supplementation of folic acid and vitamin B12 on glucose and propionate metabolism. Twenty-four multiparous cows were assigned according to a complete block design in a 2 × 2 factorial arrangement to one of the following treatments: (1) saline 0.9% NaCl, (2) 320 mg of folic acid, (3) 10 mg of vitamin B12, or (4) 320 mg of folic acid and 10 mg of vitamin B12. Intramuscular injections were given weekly from 3 wk before the expected calving date until 9 wk postpartum. At 63 d in milk, d-[6,6-2H2]-glucose (16.5 mmol/h; jugular vein) and [1-13C]-sodium propionate (13.9 mmol/h; ruminal vein) were simultaneously infused for 4 h; blood samples were collected from 2 to 4 h of the infusion period. Liver biopsies were carried out the following day. Supplements of folic acid and vitamin B12 respectively increased folate and vitamin B12 concentrations, both in milk and liver. Although dry matter intake was unaffected by treatments, milk and milk lactose yields tended to be lower by 5.0 and by 0.25 kg/d, respectively, for cows receiving the folic acid supplement. Plasma ß-hydroxybutyrate concentration with the folic acid supplement followed the same tendency. Hepatic gene expression of methylmalonyl-CoA mutase and S-adenosylhomocysteine hydrolase was higher for cows receiving the combined folic acid and vitamin B12 supplement compared with cows receiving only the supplement of folic acid, whereas no treatment effect was noted for cows not receiving the folic acid supplement. Whole-body glucose rate of appearance and the proportion of whole-body glucose rate of appearance secreted in milk lactose decreased by 229 g/d and 5%, respectively, for animals receiving the folic acid supplement, concomitant with the lower milk lactose synthesis in these cows, indicating that supplementary folic acid may alter energy partitioning in cows. The absence of treatment effect on plasma concentrations of methylmalonic acid as well as on the proportion of glucose synthesized from propionate, averaging 60%, supports the fact that vitamin B12 supply was sufficient in control cows in the current study. Our results suggest that the folic acid supplement reduced glucose-derived lactose synthesis by redirecting glucose for other metabolic activity in the mammary gland or in other tissues.

Effect of dietary organic and inorganic selenium on antioxidant status, embryo development, and reproductive performance in hyperovulatory first-parity gilts.

This project aimed to determine the effect of Se as inorganic Na-selenite (MSe) or organic Se-yeast (OSe) on antioxidant status, hormonal profile, reproductive performance, and embryo development in first-parity gilts. Forty-nine gilts were allocated to 1 of the 3 dietary treatments starting at first pubertal estrus and lasting up to 30 d after AI: control [CONT: basal diet (Se = 0.2 mg/kg) without added Se; n = 16], MSe (CONT + 0.3 mg/kg of MSe; n = 16), and OSe (CONT + 0.3 mg/kg of OSe; n = 17). Blood was collected from all gilts on the day after each onset of estrus and on d 30 after AI. Blood was also collected daily from d -4 to d +4 of the third onset of estrus (d 0) in 8 CONT, 9 MSe, and 8 OSe cannulated gilts. Gilts had received, after d 14 and 15 of their third estrus, a hormonal challenge to induce super-ovulation. At slaughter, embryos and corpora lutea (CL) were weighed and measured. Blood Se was less (P < 0.01) in CONT than in Se gilts and greater in OSe than in MSe (P < 0.01) from the first estrus until d 30 of gestation. At the same time, blood Se-dependent glutathione peroxidase (GSH-Px) decreased for CONT gilts, whereas it increased for both Se groups. The increase was greater in MSe than in OSe gilts (treatment × time, P = 0.02). Plasma 3,3',5-triiodothyronine and thyroxine concentrations for MSe tended to be less than for OSe gilts (P < 0.06). In cannulated gilts, plasma FSH tended to change among treatments (treatment × time, P = 0.06), and plasma estradiol-17ß (E(2)) was less (P = 0.01) for MSe than for OSe. There was no treatment effect on mean litter size or embryonic antioxidant status. The Se content of individual embryos was greater for Se-treated than for CONT gilts (P = 0.03), and Se content of individual embryos and total litter was greater for OSe than for MSe gilts (P < 0.01). The length, weight, and protein content of embryos were greater in OSe than in MSe gilts (P < 0.05). There was no treatment effect on weight, length, Se content, and ferric reducing antioxidant power of CL, but GSH-Px in CL was greater for Se than for CONT gilts (P = 0.02). In summary, the Se status response of gilts to dietary Se was affected by both the quantity and the source of Se dietary supplements. Moreover, the uterine transfer of Se to embryos was improved with OSe as compared with MSe, and this was concomitant with an enhanced development of embryos.

Effects of dietary vitamin supplementation and semen collection frequency on hormonal profile during ejaculation in the boar.

To evaluate the effect of dietary and management factors on boar hormonal status during ejaculation, 39 boars were canulated to determine the profiles of luteinizing hormone (LH), follicle-stimulating hormone (FSH), 17beta-estradiol (E2), and testosterone (T) in blood plasma and seminal fluid. Prior to canulation, 18 boars were fed a basal diet (control), whereas the remainder (n=21) were fed a basal diet supplemented with extra vitamins (supplemented). Within each dietary treatment, two regimens of semen collection were used over the 3mo preceding the hormonal evaluation: three times per 2wk (3/2) or three times per wk (3/1). Plasma E2 was lower (P<0.01) before ejaculation (232.5+/-22.6pg/mL) than at the onset of ejaculation (255.2+/-27.1ng/mL). Plasma T increased from 5.14+/-0.72, before ejaculation to 5.87+/-0.86ng/mL at the onset of ejaculation in supplemented boars, whereas it decreased from 5.15+/-0.65 to 4.87+/-0.70ng/mL in controls (diet by time, P<0.05). At the onset of ejaculation, plasma FSH was higher in 3/2 boars (0.436+/-0.06ng/mL) than in 3/1 boars (0.266+/-0.04ng/mL; P<0.05). During ejaculation, plasma LH increased linearly (P<0.01) from 0.59+/-0.07 to 0.97+/-0.10ng/mL, and plasma E2 and T concentrations were correlated (r=0.62, P<0.01). Plasma FSH before and during ejaculation was negatively correlated with sperm production (r=-0.60, P<0.01) and testicular weight (r=-0.50, P<0.01). In conclusion, dietary and management factors had few impacts on hormonal profiles during ejaculation, but homeostasis of some hormones was related to some criteria of reproductive performance in boars.

Effect of oestrous cycle and early pregnancy on uterine production and expression of immune regulatory factors in gilts.

This study was undertaken to characterize uterine immune factors involved in the establishment of pregnancy in gilts. Thirty crossbred Yorkshire-Landrace gilts of similar age and weight were observed twice a day for oestrous behaviour with intact boars. On the day of first standing oestrus (Day 0) and 12h later, 15 gilts were inseminated with pooled semen from Duroc boars of proven fertility. Pregnant gilts were slaughtered either on Days 10, 15 or 25 of gestation (n=5 per day). The other 15 gilts were not inseminated and were slaughtered on either Days 0, 10 or 15 of the oestrous cycle (n=5 per day). Immediately after slaughter, endometrial tissue samples from the mesometrial side were removed for gene expression using RNase protection assay and in situ hybridization methodologies. The other uterine horn was flushed with 20 ml of PBS to collect the uterine fluid. In pregnant gilts, endometrial interleukin (IL)-6 mRNA expression was higher on Day 15 than on Days 10 and 25 (P<0.01 and P<0.1, respectively). On Day 15, IL-6 expression was also significantly higher (P<0.01) in pregnant gilts than in cyclic gilts. In both pregnant and cyclic gilts, transforming growth factor (TGF)-beta2 in uterine fluid was significantly higher (P<0.0001) on Day 15 than on Day 10. At the gene expression level, TGF-beta2 also increased between Days 10 and 15 in both cyclic and pregnant gilts but differences were not significant. On Day 15, concentrations of interferon-gamma and prostaglandin E(2) (PGE(2)) in uterine fluid were markedly higher (P<0.001) in pregnant gilts than in cyclic gilts, whereas the total amount of TGF-beta2 in uterine fluid and its endometrial expression were approximately 70% higher although this increase was not significant. Finally, tumour-necrosis factor-alpha and granulocyte-macrophage/colony-stimulating factor mRNA expressions were undetectable in all endometrial samples. In conclusion, production and/or expression of uterine TGF-beta2, IL-6 and PGE(2) increased during the embryonic attachment period and are coincidental with embryonic interferon-gamma production.

Uterine immune reaction and reproductive performance of sows inseminated with extended semen and infused with pooled whole dead semen.

The objective of this study was to investigate the effect of infusing whole dead semen (WDS) after AI with diluted commercial semen on uterine inflammatory reaction and embryonic survival rate in gilts. Sixty Yorkshire-Landrace gilts were assigned at their second estrus to one of the following AI treatments: 1) commercial semen adjusted to 1 x 10(9) sperm cells (S1) per dose, followed by an infusion of 80 mL of WDS (S1-WDS); 2) S1 followed by an infusion of 80 mL of Beltsville Thawing Solution (S1-BTS); 3) commercial semen adjusted to 3 x 10(9) sperm cells (S3) per dose, followed by an infusion of 80 mL of BTS (S3-BTS); and 4) a negative control group, in which gilts received two infusions of 80 mL of BTS (BTS). Two days after the first AI, eight gilts from Groups 1, 2, and 4 were slaughtered and reproductive tracts were collected. One horn was cut open longitudinally along the antimesometrial aspect and endometrial samples were taken and immediately frozen for analysis of messenger RNA (mRNA) abundance for inflammatory cytokines and growth factors. The other horn was flushed with 20 mL of PBS, and the contents of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta1 (TGF-beta1) were determined by ELISA. On d 25 after AI, gilts from Groups 1, 2, and 3 were slaughtered and their reproductive tracts were collected to evaluate the number of fetuses and corpora lutea. On d 2 after the first AI, only TGF-beta1 was detected in the flush of all gilts, and no difference was observed between S1-WDS, S1-BTS, and BTS gilts. Endometrial levels of IFN-gamma and interleukin (IL)-6 mRNA were marked in all gilts, but they were not affected by the AI treatments, whereas the mRNA abundances for IL-1 and IL-2 were negligible. Infusions of WDS or BTS after a fertile AI did not affect IGF-I, IGF-I receptor, or IGF-II mRNA levels compared with gilts infused with BTS only, whereas the mRNA abundance for the IGF-II receptor was decreased (P < 0.05) in WDS-infused gilts. In gilts inseminated with S1 doses, infusion of WDS did not affect the number of live embryos. Although infusions of WDS did not affect the mRNA level and secretion of the cytokines measured and did not improve embryonic survival rates, further studies are needed to better understand the influence of semen composition on the uterine response after mating.

Effect of dietary fat sources on systemic and intrauterine synthesis of prostaglandins during early pregnancy in gilts.

The present experiment was conducted to determine the influence of dietary fatty acids C18:2n-6 and C18:3n-3 on the modulation of intrauterine synthesis of prostaglandin E2 (PGE2) and F2alpha (PGF2alpha) during early pregnancy in pigs. Prostaglandin E2 in uterine fluid has been previously reported to be associated with embryo survival and development. Thirty-two Yorkshire-Landrace nulliparous gilts were randomly allocated to four diets containing 5% supplemental fat. The four dietary treatments were: HT, hydrogenated tallow (26.5% C16:0 and 54.8% C18:0); SO, sunflower oil (61.3% C18:2n-6); LO, linseed oil (50.4% C18:3n-3); and SO(CLA), a mixture of sunflower oil and conjugated linoleic acids to provide 20% CLA. Treatments started 2 d after the first pubertal estrus (d -21) and lasted for 36 d (slaughter), which was 15 d after the second estrus (d 0; insemination). Fatty acids and PGE2 were measured in the peripheral blood plasma on d -19, d -7, d 0, and d 14. Fatty acids in endometrial tissues and PGE2 and PGF2alpha in the uterine fluid collected on d 15 were also measured. Concentrations of fatty acids in the plasma reflected the content of fatty acids in the diet as early as d -7. From d -7, PGE2 concentrations in the plasma were higher in gilts fed SO compared with HT (P < 0.05). Plasma PGE2 concentrations were lower (P < 0.01) on d 14 in gilts fed LO compared with HT. Total PGF2alpha contents in the uterine fluid of gilts fed LO were more than 70% lower (P < 0.05) than for the HT group. A similar trend was observed for total PGE2 content and for the ratio PGF2alpha:PGE2, but the effect (LO vs HT) was less marked (P < 0.07 and P < 0.10, respectively). There was no effect of SO or SO(CLA) on total PGE2 contents in the uterine fluid. Dietary enrichment in C18:2n-6 and/or C18:3n-3 for early pregnant gilts can influence fatty acids in plasma and endometrial tissue and can modulate circulatory and intrauterine prostaglandins.

Effect of folic acid and glycine supplementation on embryo development and folate metabolism during early pregnancy in pigs.

The present work aimed to determine if different levels of prolificacy either by parity or by genetic origin are linked to folate metabolism. Nulliparous Yorkshire-Landrace (YL) and multiparous YL, and multiparous Meishan-Landrace (ML) sows were randomly assigned to two treatments: 0 ppm or 15 ppm folic acid+0.6% glycine. Supplements were given from the estrus before mating until slaughter on d 25 of gestation. At slaughter, embryo and endometrial tissues were collected to determine concentrations of DNA, protein, and homocysteine. Allantoic fluid samples were also collected to determine concentrations of folates, vitamin B12 and amino acids. Blood samples were taken at first estrus, at mating, and on d 8, 16, and 25 of gestation to determine serum concentrations of folates, vitamin B12, and relative total folate binding capacity (TFBC). Over the entire experiment, multiparous YL sows had higher average serum concentrations of folates than nulliparous YL sows (P < 0.05) but had similar serum concentrations of relative TFBC. Concentrations of folates and relative TFBC averaged higher in ML measured over the entire experiment than in multiparous YL sows (P < 0.05). Concentrations of serum vitamin B12 were higher in multiparous YL than in ML sows or YL nulliparous sows (P < 0.05) over the entire experiment. In allantoic fluid, folates, vitamin B12, and essential amino acids contents were significantly lower in ML than in YL multiparous sows (P < 0.05). The folic acid+glycine supplement increased concentrations of serum folates, but the increase was more marked in nulliparous YL sows (nulliparous x folic acid+glycine, P < 0.05). The folic acid+glycine supplement had no effect on litter size and embryo survival, but it tended to increase embryo DNA in multiparous YL sows (P = 0.06) but not in ML and nulliparous YL sows. Homocysteine was decreased by folic acid+glycine supplement in embryos from all sows, but in endometrium, the folic acid+glycine effect was dependent on parity (nulliparous x folic acid+glycine, P < 0.05). The effects of folic acid+glycine on litter size and embryo development and survival and some aspects of folate metabolism suggest that the basal dietary content of folic acid+glycine was adequate for ML and nulliparous YL sows but not to optimize embryo development in YL multiparous sows.

Effects of breed, parity, and folic acid supplement on the expression of leptin and its receptors' genes in embryonic and endometrial tissues from pigs at day 25 of gestation.

Recent evidence has pointed toward a possible role of leptin (Lep) and its receptor (Lepr) in early gestation materno-fetal cross-talk. However, in gestating sows, exhaustive characterization of leptin mRNA expression in backfat and leptin-receptor mRNA expression in endometrial and embryonic tissues is still pending. The objectives of this study were to characterize the Lep, Lepr, and long Lepr-L isoform mRNA expression according to the breed and parity of gestating sows or to specific folic acid (B(9)) + glycine dietary treatments. To this end, nulliparous (GT) and multiparous occidental Yorkshire-Landrace (YL) sows as well as multiparous Chinese Meishan-Landrace (ML) sows were used. These sows were randomly assigned to two different dietary treatments: 0 or 15 ppm of B(9) + 0.6% glycine, given from the estrous preceding mating until slaughter on Day 25 of gestation. Jugular blood samples were collected at mating and on Day 25 of gestation and assayed for circulating leptin concentrations. Expression levels of Lep in backfat and of Lepr and Lepr-L in endometrial and embryonic tissues were performed using semiquantitative reverse transcription-polymerase chain reaction. Results demonstrated that on Day 25 of pregnancy, the ML sows showed higher concentrations of circulating leptin along with higher backfat thickness and higher expression of Lep in backfat tissue. Moreover, in embryonic tissues, the mRNA expression levels of Lepr and Lepr-L genes were higher in ML than in YL sows. Parity effects were observed for mRNA expression of Lepr in both endometrial and embryonic tissues, whereas mRNA levels were higher in YL than in GT sows. In addition, embryonic Lepr-L mRNA levels were higher in GT than in YL sows, and B(9) + glycine dietary supplement decreased the mRNA expression levels of Lep in backfat and of Lepr in embryonic tissues. These decreases were independent of breed or parity of the sows. The effect of B(9) + glycine on Lepr-L mRNA expression levels was only seen in YL sows, whereas the treatment lowered Lepr-L expression levels in both endometrial and embryonic tissues. These results indicate that leptin and its receptor may play a role during early stages of development of the pig embryo-fetus, and that these roles could be modulated according to the breed and parity of the sows. Moreover, the effects of B(9) + glycine on expression levels of embryonic and endometrial Lepr-L mRNA in YL sows may explain the previously reported effects of B(9) on embryo survival rate and litter size observed in occidental multiparous sows.

The importance of porcine sperm parameters on fertility in vivo.

It would be desirable to use semen parameters to predict the in vivo fertilizing capacity of a particular ejaculate. In animal production, an ejaculate is divided into multiple doses for artificial insemination (AI); therefore, it would be economically beneficial to know the functional quality (i.e., fertility) of the semen before it is inseminated. To identify a predictive assay of the fertilizing capacity of a porcine ejaculate, we performed 4 rapid assays of sperm quality (motility, viability, physiological status as assessed by chlortetracycline fluorescence, and ATP content) on samples from 9 ejaculates, before and after a thermal stress test (42.5 degrees C, 45 min). These parameters were subsequently correlated with in vivo fertility resulting from AI with 2 sperm doses, 3 x 10(9) or 0.3 x 10(9) motile cells in 70 mL (optimal or suboptimal sperm number per insemination, respectively) from these same ejaculates. No parameter was correlated to the fertility rates obtained after inseminating with the optimal semen doses, either before or after the thermal stress test (P > 0.05). However, with respect to the animals inseminated with the suboptimal semen dose, sperm motility (the percentage of motile spermatozoa as assessed visually by microscopy) prior to thermal stress was well-correlated to fertility rates (r = 0.783, P = 0.01). The percentage of spermatozoa displaying the chlortetracycline Pattern AR (acrosome reaction) was also statistically related to fertility (r = 0.05, P = 0.04), but the biological importance of this relationship is questionable given the small variation among ejaculates (range: 0 to 2%). No other sperm parameter was significantly related to fertility rates in this group (P > 0.05). These data, therefore, indicate that sperm motility is a useful indicator of sperm fertilizing capacity in vivo. Moreover, to identify a predictor of semen fertility it is critical that the number of spermatozoa used during insemination is sufficiently low to detect differences in sperm fertilizing efficiency.

Effect of intrauterine application of oestradiol-17 beta and prostaglandin E-2 on the porcine oestrous cycle and uterine endocrinology.

Silastic beads were inserted into the uterine lumen on Day 10 after oestrus. Gilts received beads containing oestradiol-17 beta only, oestradiol benzoate, or oestradiol-17 beta+prostaglandin (PG) E-2. Oestrous cycles were slightly longer in treated than in untreated pigs (20.2 +/- 0.4 days), and durations were 22.6 +/- 1.3, 26.2 +/- 1.7 and 23.2 +/- 1.8 days for oestradiol-17 beta, oestradiol benzoate and oestradiol-17 beta+PGE-2 treatments, respectively (P greater than 0.05). Thus, PGE-2 and an oestrogen such as oestradiol benzoate that persist for a longer period cannot prolong the cycle more than oestradiol-17 beta alone. Additional cyclic gilts underwent similar treatments with beads containing oestradiol-17 beta, oestradiol-17 beta+PGE-2 or cholesterol, and cannulation of one utero-ovarian vein on Day 10. Blood samples were collected from the catheter every 15 min from 08:00 until 11:00 h and from 20:00 until 23:00 h for 5 consecutive days starting the day after surgery and peripheral plasma samples were also collected daily. On Day 16, beads containing oestradiol-17 beta were surrounded by endometrial folds whereas cholesterol beads were free. Concentrations of plasma progesterone did not vary significantly from Days 11 to 16 in gilts treated with oestradiol-17 beta or oestradiol-17 beta+PGE-2, but decreased in cholesterol-treated gilts. Concentrations of plasma oestrone and oestradiol-17 beta were more than ten times higher in gilts treated with oestradiol-17 beta or oestradiol-17 beta+PGE-2 than in cholesterol-treated gilts on the day after bead insertion, but decreased rapidly to values comparable to those in cholesterol-treated gilts by Day 14. In contrast, concentrations of oestrone sulphate remained high until Day 16. Concentrations of PGE-2 in the utero-ovarian vein plasma did not differ (P greater than 0.05) between treatments but those of PGF-2 alpha were higher (P less than 0.004) in gilts treated with cholesterol than in those treated with oestradiol-17 beta or oestradiol-17 beta+PGE-2. It is postulated that insufficient oestradiol-17 beta is released by the beads toward the end of a 'recognition period' to prolong the cycle for more than 3-6 days.

Structural and functional aspects of porcine endometrial capillaries on days 13 and 15 after oestrus or mating.

The permeability of subepithelial capillaries in porcine endometrium was studied during midcycle and early pregnancy. Gilts were slaughtered on Day 13 or Day 15 of the oestrous cycle or pregnancy, 15 min after injection through the ear vein of 11.1 MBq of 125I-labelled human albumin in phosphate-buffered saline. The radioactivity of endometrial strips taken along the mesometrial and antimesometrial aspects of the uterine horn varied on average from 270 to 701 c.p.m./g and no difference (P greater than 0.05) was found between reproductive status, days of slaughter or sampling sites. The majority of the subepithelial capillaries showed ultrastructural evidence of increased vascular permeability, such as marked thinning of the capillary walls, especially on the side proximal to the epithelial basal lamina, multilayering and partial disparition of the endothelial basal lamina and abundant endothelial vesicles. Fenestrated pores were observed, but were rare. There was no obvious difference between reproductive status, days of sampling or sampling sites inside the uterus, suggesting that on Days 13-15 after oestrus the ultrastructural characteristics of porcine endometrial capillaries are little affected by the presence of attaching blastocysts and supporting the results obtained with radioactive albumin. Ferritin injected directly into a uterine artery of one gilt on Day 15 of pregnancy was carried through the capillary wall by endothelial vesicles, showing ultrastructural evidence of increased permeability.

Effect of topical application of estradiol-17 beta and PGE2 on PGE-binding sites in the porcine endometrium.

High- and low-affinity prostaglandin E2 (PGE2) binding sites were found on day 15 after estrus in the endometrium of cycling (Cy) and pregnant (Pr) gilts as well as gilts treated with intra-uterine Silastic beads containing estradiol-17 beta (E2) alone or in combination with PGE2 (E and PG gilts respectively) and inserted into the uterine lumen on day 10 of the cycle. The average apparent dissociation constants (Kd) and binding site concentrations (Bmax) for the high- and low-affinity sites were respectively (mean +/- SEM): 8.4 +/- 0.7 pM and 3.28 +/- 0.38 fmol/mg of protein and 5.3 +/- 0.8 nM and 71 +/- 9 fmol/mg of protein. Samples collected along the meso- and antimesometrial aspects did not differ (P greater than 0.05), although the low-affinity Bmax was higher on the antimesometrial aspect for Pr and Cy gilts only. No difference in Kd (P greater than 0.10) was found between treatments for high-affinity binding sites. For the low-affinity binding sites, Kd was higher for Pr compared to PG and E but not to Cy gilts (P less than 0.05). The high-affinity Bmax was higher (P less than 0.05) for PG, followed by E, Pr and Cy gilts (respectively: 5.50 +/- 0.26; 4.19 +/- 0.46; 1.78 +/- 0.40; 1.64 +/- 0.23 fmol/mg of protein), although Pr and Cy gilts were not different (P greater than 0.05). These results suggest that the localized presence of conceptuses in the uterus in early pregnancy does not markedly affect PGE binding sites but that intrauterine applications of Silastic beads containing E2 and PGE2 increase high-affinity Bmax and decrease low-affinity Kd.

Autofluorescence of the porcine endometrium during early pregnancy.

Uteri recovered from cyclic gilts (n = 5) on Days 15-19 and pregnant animals (n = 34) on Days 13-40 were opened and examined under UV light. A line of greenish fluorescence was present in the mesometrial region in contact with embryonic membranes at Day 13. Small patches of reddish fluorescence subsequently appeared on the uterine mucosa near the embryonic disc, and these increased in intensity and size until they encompassed the entire area of contact between each conceptus and the endometrium, for lengths of about 20 cm, by Day 29. Fluorescence then diminished gradually and was almost totally absent by Day 40. Three additional gilts were unilaterally hysterectomized on Day 15 and treated with Evans blue dye 10 min before removal of the second uterine horn. Both horns were opened and compared under UV light, but no difference in the pattern of fluorescence could be detected. All fluorescence was associated with uterine rather than conceptus tissues. The occurrence of autofluorescence in uteri of pregnant pigs precludes use of Evans blue dye as an indicator of vascular permeability.