J. Anim Sci.
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Chartrand, R.
Right arrow Articles by Laforest, J. P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Chartrand, R.
Right arrow Articles by Laforest, J. P.
J. Anim. Sci. 2003. 81:726-734
© 2003 American Society of Animal Science

Effect of dietary fat sources on systemic and intrauterine synthesis of prostaglandins during early pregnancy in gilts1,2

R. Chartrand*, J. J. Matte{dagger},3, M. Lessard{dagger}, P. Y. Chouinard*, A. Giguère{dagger} and J. P. Laforest*

* Département des Sciences Animales, Faculté des Sciences de l’Agriculture et de l’Alimentation, Université Laval, Ste-Foy, Québec, Canada, G1K 7P4 and and {dagger} Dairy and Swine R&D Centre, Agriculture and Agri-Food Canada, Lennoxville, Québec JIM 1Z3

3 Correspondence:
Fax: 819-564-5507, E-mail:
mattej{at}agr.gc.ca.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
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 F2{alpha} (PGF2{alpha}) 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 SOCLA, 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 PGF2{alpha} 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 PGF2{alpha} 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 PGF2{alpha}: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 SOCLA 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.

Key Words: Fatty Acids • Pig • Pregnancy • Prostaglandins


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 
During early pregnancy in pigs, estradiol from embryonic origin is involved in the antiluteolytic shift of PGF2{alpha} secretion from an endocrine to an exocrine pathway (Bazer et al., 1984). The conceptus and the endometrium can also secrete other PG, such as PGE2 (Lewis and Waterman, 1983; Bazer et al., 1984). Prostaglandin E2 is critical, in early gestation, for vascular permeability, placental development, and immune response of pigs (Kennedy, 1977; Geisert et al., 1990). In the allantoic fluid, it has also been related to larger litter size and weight (Giguère et al., 2000).

Arachidonic acid (C20:4n-6) is the immediate precursor of the 2 species PG. It is bioavailable either directly from the diet or via desaturation and elongation reactions (Levine, 1988) and is one of the most abundant fatty acids in the phospholipids of mammalian cellular membrane (Shapiro et al., 1993). The synthesis of 2 sp. PG is controlled by the presence and activity of the cyclooxygenase (COX) enzymes (Dubois et al., 1993). Eicosapentaenoic acid (C20:5n-3), a precursor of the 3 sp. PG, and conjugated linoleic acids (CLA; Li and Watkins, 1998) are known to compete at the COX level with C20:4n-6. The balance among these fatty acids determines the quantity and type of PG synthesized. Linolenic acid (C18:3n-3) is a precursor of C20:5n-3, whereas C20:4n-6 is derived from linoleic acid (C18:2n-6). In this last case, an intermediate metabolite, dihomo-{gamma}-linolenic acid (C20:3n-6) is precursor of both C20:4n-6 and the 1 sp. PG.

This experiment was aimed to determine the effect of pre- and postmating dietary enrichments in n-6 and n-3 fatty acids and in CLA on some factors related to reproductive efficiency in early pregnant gilts. The fatty acid profile and PGE2 were monitored in plasma during the experimental period. At slaughter, fatty acids were measured in endometrial tissue, PGE2 and PGF2{alpha}, in uterine fluid, and the in vitro synthesis of PGE2 after incubation of endometrial explants.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Implications
 Literature Cited
 
Animals and Treatments
Thirty-two Yorkshire-Landrace nulliparous gilts were randomly allocated to a basal diet containing 5% supplemental fat. The four dietary fat treatments (Table 1Go) were: HT, hydrogenated tallow (JEFO Nutrition inc., St-Hyacinthe, Québec, Canada); SO, sunflower oil (Pokonobe Industries, Montréal, Québec, Canada); LO, linseed oil (Swimco Canada Inc., Georgetown, Ontario, Canada); and SOCLA, mixture of sunflower oil and CLA to provide 20% CLA. The CLA supplement (Natural Lipids Ltd., Hovdebygda, Norway) contained 33.1% cis-9, trans-11 C18:2 and 32.2% trans-10, cis-12 C18:2 in the form of methyl esters. Once a day at 0900, gilts received their treatment as a top dress at a rate of 5% (by weight) on 2.8 kg of the basal diet (Table 2Go). No feed refusals were observed. To reduce the risk of oil oxidation, butylated hydroxyanisole (0.01%) and butylated hydroxytoluene (0.01%) were previously added to SO, LO and SOCLA. Oils and tallow were kept at 4°C until they were fed to the animals.


View this table:
[in this window]
[in a new window]
  		Table 1. Fatty acid composition (g/100 g of fatty acids) of the basal dietand the fat supplementsab
 

View this table:
[in this window]
[in a new window]
  		Table 2. Composition of the basal dieta
 
On arrival in the experimental room, gilts were grouped in pens (1.5 m x 2.5 m) of two to three, next to a boar, in order to stimulate estrus. At the first pubertal estrus, gilts were placed into individual stalls (0.6 m x 2.2 m). Eight blocks were established based on the total body fat of the animals estimated from their backfat thickness and BW (Whittemore and Yang, 1989). Gilts were then randomly assigned to one of the four experimental groups within each block.

Treatments started 2 d after the first estrus (d -21) and lasted for 36 d. Gilts were inseminated at the second estrus and slaughtered 15 d later (d 15), between 1100 and 1300. Estrus was detected twice a day by introducing a boar into the pen between 0800 and 0900 and from 1600 to 1700. Gilts were inseminated twice with 85 mL of semen (3 x 109 live sperm cells from pooled semen of three Duroc boars) provided by a local AI centre (CIPQ Inc., St-Lambert, Québec, Canada). When estrus was detected in the morning, gilts were inseminated twice, 8 and 24 h later. When estrus was detected in the afternoon, the two inseminations were done 16 and 24 h later. The day of the second insemination was considered d 0 of the trial.

Out of the 40 gilts that started the project, 32 were pregnant at slaughter on d 15. Gilts were removed from the experiment due to health problems (two HT and one LO), insemination failure (one HT and one LO), immature uterine tract on d 15 (one HT), nonpregnancy on d 15 (one LO), and anestrus (one case) before assignment of treatments.

All animals were cared for and slaughtered according to the recommended code of practice of Agriculture Canada (1993), and the procedure was approved by the local Animal Care Committee following the guidelines of the Canadian Council on Animal Care (1993).

Sampling and Analyses
Blood samples were collected between 0800 and 0900 in tubes containing potassium EDTA (15%, wt/wt) (Vacutainer, Becton Dickinson and Co., Rutherford, NJ) following a 16-h feed withdrawal. Samples used for the PGE2 assay were preacidified with hydrochloric acid (0.1 N; 0.1 mL/mL of blood) as described by Lewis et al. (1978). Blood was collected on the mornings of d -19, -14, -7, 0, and 14 by a jugular catheter (Matte, 1999). For technical reasons, some (n = 19) of the blood samplings on d 0 were done without a catheter by Vacutainer (Tremblay et al., 1989). The sampling procedure did not have any effect on the different variables measured in plasma on d 0. Tubes were centrifuged for 5 min at 1,800 x g and the plasma was separated into aliquots and frozen at -20°C for PGE2 and at -80°C for the fatty acid assays.

Immediately after slaughter, the reproductive tract was collected, placed on ice, and transported to the laboratory. The tract was carefully separated from the mesometrium, mesosalpinx, and mesovarium. The uterus and cervix were separated and both horns were flushed with 20 mL of PBS (GIBCO BRL, No. 70011-044, Burlington, Ontario, Canada) within 1 h after slaughter as described by Laforest and King (1992). The solution was introduced at the oviduct end using a blunt needle and a syringe. The other end of the horn was fitted with a plastic tube and firmly held on a cotton string by a tie rap to channel the fluid into a graduated cylinder. The total fluid collected from both horns was considered to be the "uterine fluid," and its volume was recorded to determine the total content of hormones and metabolites collected from the uterine lumen. The uterine fluid was centrifuged (86 x g for 5 min) and the supernatant was collected and stored at -20°C. Conceptuses were washed twice with 15 mL of PBS and then stored at -80°C for further DNA and protein analyses.

Both horns were also used to collect the endometrial tissues to determine the production of PGE2 in vitro. The sampling technique was adapted from Duquette et al. (1997), and the explant culture procedure followed a method proposed by Rosenkrans et al. (1990). After opening the horn along the antimesometrial aspect, longitudinal strips of endometrium (20 x 1 cm) were taken from the mesometrial side of the horns and then minced with a scalpel blade under sterile conditions in samples of 90 to 110 mg each. Explants were incubated in 24-well culture plates, each well containing one explant and 2 mL of a modified Krebs Henseleit buffer (Sigma K 3753, St. Louis, MO) supplemented with vitamins and amino acids (essential and nonessential) in minimum Eagle’s essential medium (MEM) (GIBCO, Grand Island, NY). The incubation mixture was rocked gently (two oscillations/min) under controlled temperature (37°C) and atmosphere (95% air and 5% CO2) in a humidified chamber.

Incubations were conducted for a total of 6 h. During the first 2-h period, no treatment was applied to the medium culture. At the beginning of the second period (lasting 4 h), the medium was replaced with fresh medium containing 0, 20, or 60 -g of C20:4n-6 or C20:5n-3 per milliliter. At the end of the experiment, supernatants were collected and stored at -20°C.

Fatty acids were methylated according to the method described by Park and Goins (1994) and measured in plasma on d -19, -7, 0, and 14, in the endometrium tissue collected on d 15, in the basal diet, and in the supplemental fats. Endometrium tissue (2 g) was homogenized in 4 mL of demineralized water. The in situ transesterification was done on 200 -L of endometrium homogenates, on 200 -L of plasma, on 200 mg of finely ground meal (1 mm), and on 20 mg of supplemental fats. Methylene chloride (200, 150, 200, and 100 -L, respectively) and 1 mL of 0.5 N NaOH in methanol were added to each aliquot of lipid containing samples.

Fatty acid methyl ester profiles were measured by gas chromatography on a Hewlett-Packard 6890 chromatograph (Hewlett-Packard Ltd., Montreal, Québec, Canada) fitted with a 60 m x 0.25 mm Supelco SP-2380 capillary column (Sigma-Aldrich Canada Ltd., Ontario, Canada) with a film thickness of 0.20 -m. Helium at 0.8 mL/min was used as the carrier gas and also at 29 mL/min as the make-up gas. Average carrier gas velocity was 20 cm/s. The split:split less ratio was 100:1 and the injection volume was 1 -L. Injector and detector temperatures were 260°C and 275°C, respectively. After an initial isothermal period of 5 min at 140°C, the temperature was programmed to 240°C, rising by 5°C/min with a hold of 5 min at that temperature, and then programmed to 258°C at 9°C/min with a final hold of 4 min at that temperature. The hydrogen flow was 40 mL/min and the airflow was 450 mL/min. The data were processed with HP Chemstation software version A.06.01. Values were expressed as a percentage (g/100 g) of total fatty acids.

The PGE2 concentrations were measured in plasma by RIA (Jaffe and Behrman, 1974) on d -19, -14, -7, 0, and 14, in uterine fluid collected on d 15, and in the supernatant of endometrial explant from cell culture plot. Plasma PGE2 had been previously extracted with absolute ethyl alcohol (1:10 water; Aldrich 27,074-1, Milwaukee, WI) according to the methods of Laforest and King (1992). The samples from uterine fluid and supernatants were diluted with PBS-BSA without any extraction procedure. For PGE2 assays, the PGE2 rabbit anti-PGE2-BSA serum (ICN Biomedicals, Inc., Aurora, OH) cross-reacted 270% or less with PGE1, 7.7% or less with PGF1{alpha}, 6.8% or less with PGF{alpha} and less than 5% with other PG. No specific information was available on antibody cross-reaction with PGE3. The validation tests showed a satisfactory parallelism among dilutions (CV of 4.7% between 1:10 and 1:200) and recovery (104%). Inter- and intraassay CV were 5.9 and 4.3%, respectively.

The PGF2{alpha} was determined using an ELISA system (Cayman Chemical Co., Ann Arbor, MI). This assay cross-reacted 100% with PGF1{alpha}, PGF2{alpha}, and PGF3{alpha}, and 51% with PGD2. No extraction of the samples was necessary since the 1:500 and 1:1,000 dilutions differed by less than 10%. The validation tests showed a satisfactory parallelism among dilutions (CV of 9.9% between 1:200 and 1:500) and recovery (111%). Inter- and intraassay CV were 5.3 and 2.7%, respectively.

In spite of the different cross-reactions with the 1 sp. PG, the PG determinations are likely to represent mostly the 2 sp. PG. Indeed, the fatty acid profiles in plasma and in endometrial tissue indicated that the amount of C20:3n-6, a precursor of both 1 sp. PG and C20:4n-6, corresponded to a small fraction of C20:4n-6 content, less than 1/25 (plasma) and 1/12 (endometrial tissue) in SO- and SOCLA-enriched diets. Moreover, as reported recently by Levin et al. (2002), dietary strategies to increase the cellular ratio of C20:3n-6:C20:4n-6 are not effective in altering endogenous cellular PGE1 synthesis, which was substantially lower than PGE2; this was particularly marked in tissues or cells where COX-1 (constitutive) predominates over COX-2 (inducible). In this way, we have recently shown that endometrial messenger RNA expressions of COX-1 were double that of COX-2 in nulliparous sows at 15 d of gestation (Guay et al., unpublished data). Therefore, the terms PGE2 and PGF2{alpha} were used throughout the text in reference to the PG determinations in the present experiment.

Protein and DNA contents were measured on the conceptuses, previously homogenized (glass tissue grinder) in 8 mL of PBS. Protein concentrations were determined by colorimetric assay (Bio-Rad DC protein assay No. 500-0116, Mississauga, Ontario, Canada), DNA concentrations, and a fluorimetric assay as validated by Labarca and Paigen (1980).

Statistical Analyses
The data were analyzed using the SAS procedure for MIXED models (SAS Inst., Inc., Cary, NC) as a randomized complete block design with four treatments in eight blocks. Specific a priori contrasts (SO vs HT, LO vs HT, and SOCLA vs SO) were used to establish comparisons among different treatments. For repeated measurements, such as body fat (estimated from backfat thickness), plasma PGE2, and fatty acids, polynomial contrasts were used to estimate time effects along with their interactions with treatments or specific treatment contrasts. The covariance structure used was spatial power.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Implications
 Literature Cited
 
Performance
Body weight and backfat thickness were (means ± SEM) 123.5 ± 2.0 kg and 13.2 ± 0.5 mm, respectively, at the first estrus; 133.1 ± 1.9 kg and 14.6 ± 0.5 mm, respectively, at insemination; and 143.6 ± 1.9 kg and 16.2 ± 0.5 mm, respectively, at slaughter. The increase in the estimated body fat content of the gilts between d -19 to 15 (Table 3Go) was more marked (33 vs 24%) in LO than in HT gilts (interaction of fat x time was linear, P < 0.02).


View this table:
[in this window]
[in a new window]
  		Table 3. Estimated body fat content (%) in gilts fed differentfat supplements (5% of the diet)ab
 
Fatty Acids in Maternal Plasma
Compared with gilts fed HT, plasma of gilts fed SO contained more (P < 0.001) C18:2n-6 and less (P < 0.01) C16:0, C18:1, C20:3n-6, and C20:5n-3, whereas in gilts fed LO, there was less (P < 0.05) C16:0, C18:1, C18:3n-6, C20:4n-6, C22:6n-3, and more (P < 0.001) C18:2n-6, C18:3n-3, and C20:5n-3 (Table 4Go). Average plasma values of {gamma}-linolenic acid (C18:3n-6) and C20:4n-6 were lower (P < 0.05), and those of C16:0, C20:5n-3, and CLA were higher (P < 0.05) in gilts fed SOCLA compared with SO (Table 4Go). In fact, CLA was detected only in gilts fed SOCLA.


View this table:
[in this window]
[in a new window]
  		Table 4. Effect of dietary fat supplements (5% of the diet) given to gilts from 19 d before insemination and throughout a 15-d gestation, on the average (d-19, -14, -7, 0, and 14) fatty acid composition of maternal blood plasma
 
For most fatty acids, the treatment effects (fat x time linear or time quadratic [plateau]) were established (P < 0.02) after 14 d of supplementation (d -7) and remained constant up to the last blood sampling (data not shown).

Fatty Acids in Endometrial Tissues
In the endometrial tissue, the fatty acid profiles of gilts fed SO vs HT showed less (P < 0.001) C18:1 and more (P < 0.001) C18:2n-6 (Table 5Go). In the case of gilts fed LO vs HT, there was less (P < 0.01) C18:1 and C20:4n-6 and more (P < 0.05) C18:2n-6, C18:3n-3, C20:3n-6, and C20:5n-3. No C18:3n-6 was detected in endometrial tissue, whatever the treatments. For gilts fed SOCLA compared with SO, there was less (P < 0.05) C18:1 and C18:3n-3 and more (P < 0.05) C18:2n-6 and CLA (Table 5Go).


View this table:
[in this window]
[in a new window]
  		Table 5. Effect of dietary fat supplements (5% of the diet) given to gilts, from 19 d before insemination and throughout a 15-d gestation, on fatty acid composition of endometrial tissues on d 15 of gestation.
 
Prostaglandins in Blood Plasma and Uterine Fluids
The PGE2 content of peripheral blood plasma was higher (P < 0.05) in gilts fed SO compared with HT (values for d -14, -7, and 14 are shown in Figure 1Go), the effect was apparent as early as d -7 of the experiment (interaction of fat [SO vs HT] x time cubic, P < 0.01). Maternal plasma PGE2 was lower in gilts fed LO compared with HT, the effect being particularly marked on d 14 (interaction of fat [LO vs HT] x time quadratic, P < 0.02).



View larger version (37K):
[in this window]
[in a new window]
  		Figure 1.

Plasma PGE2 concentrations of gilts receiving a diet supplemented with 5% fat served as a top dress meal once a day during the pre- and postmating periods. The four fat treatments were: HT = hydrogenated tallow, SO = sunflower oil, LO = linseed oil, SOCLA = mixture of sunflower oil and conjugated linoleic acids. Specific contrasts (SO vs HT, LO vs HT, and SOCLA vs SO) were used to establish comparisons among treatments. Values are means ± SEM (n = 8). *SO vs HT, P < 0.05; **LO vs HT, P < 0.01.

 
There was a marked decrease of over 70% (P < 0.05) of the total amount of PGF2{alpha} in the uterine fluid in gilts fed LO compared with HT (Table 6Go). A similar trend was observed for PGE2 and for the ratio PGF2{alpha}:PGE2, but the effect (HT vs LO) was less marked (P < 0.07 and P < 0.10, respectively).


View this table:
[in this window]
[in a new window]
  		Table 6. Effect of dietary fat supplements (5% of the diet) given to gilts, from 19 d before insemination and throughout a 15-d gestation, on composition of uterine fluid and embryo development on d 15 of gestation.
 
In Vitro Prostaglandin E2 Production by Endometrial Tissues
Average PGE2 production by endometrial explants (data not shown) increased from 1.3 ± 0.1 to 4.0 ± 0.5 pg/mL in the presence of C20:4n-6 at 20 and 60 -g/mL, respectively (quadratic effect of C20:4n-6 concentration, P < 0.001), whereas the concentration of C20:5n-3 had no effect. With C20:4n-6 as a precursor in the culture media, the pattern was similar among the fat treatments, whereas the response to C20:5n-3 was slightly different, a maximum being reached at 20 -g/mL for LO and 60 -g/mL for HT (interaction of fat [LO vs HT] x C20:5n-3 concentration quadratic, P < 0.05).

Weight, Protein, and DNA Measurements from the Conceptus
No treatment effect was found on weight, protein, or DNA contents of the conceptus, or on the protein:DNA ratios (Table 6Go). Nevertheless, the ratio of protein:DNA was 30% lower (P < 0.09) in LO than in HT gilts.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Implications
 Literature Cited
 
Performances
Although the treatments were designed to provide similar amounts of dietary fat, gilts fed LO gained more body fat (8.0%) than those fed HT (5.9%). Such a difference could be explained by the higher available ME of vegetable oil compared with tallow (8,500 and 7,500 kcal/kg, respectively; INRA, 1984). In accordance with this higher available ME of vegetable over animal fat, the body fat gain was also higher, although not significantly, in SO vs HT gilts.

Fatty Acids in the Maternal Blood Plasma
The fatty acid profiles in plasma were readily altered by the dietary fat source. For most fatty acids, the delay between the initiation of treatments and stabilization of the proportions of individual fatty acids in the plasma was as short as 14 d. Such an early response is in agreement with Warnants et al. (1999), who showed that in pigs fed full-fat soybeans instead of tallow, 50% of the increase in C18:2n-6 in backfat occurred within 2 wk of the diet change.

Regardless of the fat treatment, C18:2n-6 was the most abundant fatty acid in the plasma (Table 4Go). Plasma C18:2n-6 varied according to the dietary proportions of this fatty acid, but this was not the case for C18:3n-3. Indeed, the higher proportions of C18:2n-6 in SO and LO vs HT diets induced an increase in concentrations of plasma C18:2n-6, whereas for C18:3n-3, it increased only in LO gilts. It seems that the actual small amounts of C18:3n-3 in SOCLA (0.7%) and SO (1.2%) diets were not sufficient to influence the concentration of this fatty acid in circulation. These results are partly consistent with those of Bee et al. (1999), who reported a dose-response relationship between dietary PUFA supply and tissue content. However, in contrast to the present results (Table 4Go), the response reported by Bee et al. (1999) was due to a lower tissue incorporation of monounsaturated fatty acids without any effect of the tissue incorporation of saturated fatty acids. This difference between the two experiments might be due to the metabolic pools involved; tissue lipids were studied in Bee et al. (1999), compared to plasma, in the present experiment.

The lower C18:3n-6 (0.3 vs 0.4%) and C20:4n-6 (4.9 vs 9.0%) and the higher C20:5n-3 (2.3 vs 0.3%) contents in LO than in HT gilts are in agreement with the increase of total n-3 PUFA (Cunnane et al., 1990) and the decrease of n-6 PUFA (Calder, 1996) observed after the addition of flax in the diet of pigs.

Compared with SO, SOCLA induced an increase of plasma C20:5n-3 and CLA and a decrease in plasma C18:3n-6 and C20:4n-6. As proposed by Banni and Martin (1998), such results suggest that CLA may selectively promote the elongation of n-3 PUFA and increase the degradation of n-6 PUFA. Li and Watkins (1998) suggested that for rats given CLA, the relative sparing effect of n-3 fatty acids might be explained either by an increase in utilization of n-6 or conservation of n-3 fatty acids.

Fatty Acids in Endometrial Tissues
As observed for plasma values, an increase of dietary PUFA lowered endometrial monounsaturated fatty acids, but without any effect on saturated fatty acid content. The treatment differences for endometrial C18:2n-6 content suggest, as proposed by James et al. (1993), that the rate of esterification of C18:2n-6 into various tissue fractions was not saturated by the provision of C18:2n-6 from the basal diet. The proportions of C20:4n-6 in the endometrium were approximately twice the corresponding plasma values. However, none of the n-6-enriched diets was efficient in modulating C20:4n-6 incorporation in endometrium. This lack of SOCLA and SO effects on total endometrial C20:4n-6 occurred in spite of an endometrial C18:2n-6 increase. Possibly, the conversion of C18:2n-6 to C20:4n-6 was inhibited in porcine endometrial cells, as suggested by Artzen et al. (1998) for human decidual cells, and/or the conversion and incorporation of C20:4n-6 into tissue phospholipids is a saturable process (Whelan et al., 1992). This lack of effect of the n-6-enriched diets could also be related to the fact that endometrial tissues, like the placenta, selectively or preferentially take up the long-chain PUFA (Campbell et al., 1998) from endogenous (liver) sources of fatty acids at the expenses of exogenous (diet) sources (McBride and Burton, 1964; Thomas and Lowry, 1987). In such cases, binding proteins for a specific fatty acid (Campbell et al., 1998) might be involved in the regulation of this long-chain PUFA uptake within the endometrium in order to support the implantation and development of the embryo. Furthermore, James et al. (1993) indicated that dietary fish oil, which contains C20:5n-3, is effective in decreasing cellular C20:4n-6 content, whereas a decrease in dietary C18:2n-6 intake has little or no influence on tissue C20:4n-6.

In LO-fed gilts, a 30% decrease in C20:4n-6 was observed in endometrial tissues; the fatty acid shift being mostly in favor of C20:5n-3. It seems that endometrial C18:3n-3 in those gilts induced a competitive inhibition of {Delta}6-desaturase, the rate-limiting reaction for the conversion of C18:2n-6 to the intermediary metabolite C18:3n-6 toward the synthesis of C20:4n-6 (Amusquivar et al., 2000).

The inefficiency of SOCLA over SO to lower n-6 PUFA and to increase the C20:5n-3 content of endometrial tissues could be partly explained by the weak incorporation of CLA in these tissues compared with CLA incorporation in plasma (0.8 and 1.5%, respectively). Therefore, at such a low tissue concentration of CLA, the endogenous high levels of C20:4n-6 in the endometrial tissue could have prevented or masked any CLA effect to lower n-6 PUFA and promote n-3 PUFA elongation into C20:5n-3.

Maternal Plasma and Uterine Prostaglandin E2 Production
Plasma PGE2 in HT gilts remained stable from d -14 to 14, whereas on d 0, a peak was observed for SO and LO gilts and to a lesser degree for SOCLA gilts. These results are in agreement with Burke et al. (1996), who showed that a component of olive oil, probably C18:2n-6, stimulated PG synthesis and the release of PGFM, a stable metabolite of PGF2{alpha}, and PGE2 in circulation on d 13 to 15 of the estrous cycle in ewes. Nevertheless, it cannot be ruled out that the treatment effects on d 0 could be indirectly due to differences in the timing and/or the length of ovulation.

The treatment effects on circulating PGE2 were established as early as d -7. Unexpectedly, plasma PGE2 are not always related to the corresponding C20:4n-6 content. Such a lack of a direct relationship between PGE2 and its precursor in the plasma pool is possibly due to the fact that the concentration of PGE2 in the plasma reflects the different metabolic responses to fatty acids from other metabolic pools. Nevertheless, there seems to be some relationship between plasma PGE2 and the content of n-6 and n-3 PUFA in the diet as shown by the higher plasma PGE2 in SO vs HT gilts on d -7 and 14. It has been previously demonstrated that a diet enriched with n-6 PUFA enhances PGE2 production by different cellular types (Calder, 1996).

From d -14, plasma PGE2 in LO gilts was lower than in HT gilts, the effect being particularly marked on d 14. Such an effect suggests that the high content of n-3 PUFA in LO lowered the C20:4n-6 metabolism in favor of C20:5n-3 by competing for {Delta}6-desaturase, and afterward by lowering PGE2 production in favor of PGE3 at the cyclooxygenase level (Miles and Calder, 1998). Nevertheless, such competition appears limited at the systemic level because the effect of LO over HT, although significant on the intermediary metabolite C18:3n-6, was small (0.3 vs 0.4%).

In spite of their effect in circulation, the influence of dietary fats on PGE2 in uterine fluid was less marked with only a tendency for a decrease in LO gilts. Nevertheless, the decrease was numerically important (43%) compared with HT gilts. In the early stages of pregnancy, it is known that endometrial tissues of gilts can produce and secrete large amounts of PGE2 in the uterine lumen (Laforest and King, 1992). The conceptuses also produce large amounts of those PG (Lewis and Waterman, 1983), although the overall contribution within uterine fluid is likely to be limited by the small amount of embryo tissue compared with the endometrium at that stage of gestation. The rate of incorporation of C20:4n-6 into phospholipids and the ability of phospholipase A2 to release it, could influence PGE2 production, as proposed for PGF2{alpha} by Norman and Poyser (2000). Considering that C20:4n-6 is the primary substrate for production of PGE2 and PGF2{alpha} (Arntzen et al., 1998), it seems likely that the similar endometrial content of C20:4n-6 in HT-,SOCLA-, and SO-fed gilts was a critical factor for the lack of effect among these three treatments on local PGE2 production from both endometrium and conceptuses. Moreover, the in vitro capacity of the endometrial tissue to produce PGE2 in the presence of C20:4n-6 seems to be independent of the dietary treatments, as demonstrated by the culture of endometrial explants. The lack of effect of dietary CLA on PGE2 in uterine secretions contrasted with previous results showing an inhibitory effect of CLA on circulating and spleen PGE2 (Sugano et al., 1997) in rats. In rat bone, dietary CLA also lowered ex vivo production of PGE2 beyond that of n-3 fatty acid feeding (Li and Watkins, 1998). Differences in the form of CLA isomers used (methyl esters in the current trial vs free fatty acids in previous experiments) could be involved in such a discrepancy between studies.

As mentioned previously with the endometrial fatty acids profiles, it seems that n-3 PUFA from LO had interfered directly with intrauterine PGE2 and PGF2{alpha} synthesis by competing for the {Delta}6-desaturase and/or for the cyclooxygenase. The stronger LO effect on PGF2{alpha} than on PGE2 suggests that, beyond synthesis from fatty acids, other physiological events could have affected differentially the metabolism of these two different forms of PG. In this way, LO might have affected the efficiency of the shift of PGF2{alpha} endometrial secretion, which is directed at the expense of local circulation toward the uterine lumen (Bazer et al., 1984); this mechanism is critical for inhibition of luteolysis at the time of implantation. Such a hypothesis remains to be investigated. Nevertheless, an analytical artifact due to cross-reaction of PGE3 with PGE2 determinations cannot be ruled out, but such information was not available in the present conditions.

Although not significant (P < 0.09), the 30% decrease in blastocyst protein:DNA ratio of gilts fed LO vs HT suggests a tendency for LO to impair embryonic growth. Furthermore, such an effect is corroborated by observations showing, in four LO-fed gilts, several alterations of the endometrium (pale sites of attachment, small uterine horns with low elasticity, distended and purple uterus [gilt considered not pregnant]) or ovaries (large CL with petechiae on the tissue surface). Taken together, these results suggest that LO and/or C18:3n-3 could have a detrimental effect on the uterine environment and the well-being of the embryos in early pregnancy. Further studies are required to confirm and explain these results. Amusquivar et al. (2000) demonstrated that an excess of n-3 fatty acids in the maternal diet could induce a specific deficiency of C20:4n-6 in the fetuses. In such cases, the effect is either due to the direct inhibitory action of n-3 fatty acids on {Delta}6 desaturation within the fetus and/or indirectly to an altered proportion of fatty acids for placental transfer. Sayre and Lewis (1993) demonstrated that, as early as d 4 of development, ovine embryos can convert C20:4n-6 into several compounds, including PG, and hypothesized that PGE2 and PGF2{alpha} may be involved in the transition from spherical to elongated embryos. In pigs, it has been proposed recently that PGE2 might play a key role in embryo development during early stages of gestation (Giguère et al., 2000).


   Implications
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
Literature Cited
 
The dietary enrichment in C18:2n-6 and C18:3n-3 can influence the fatty acid composition of plasma and endometrial tissues and modulate systemic prostaglandin E2 concentration during early pregnancy in gilts. However, C18:2n-6-enriched diets were not efficient in increasing total prostaglandin E2 and prostaglandin F2{alpha} recovered in the uterine fluid, whereas C18:3n-3 decreased both prostaglandins drastically. This difference seems to be due to the fact that C18:2n-6 is not efficient in modulating local C20:4n-6 concentrations, whereas C18:3n-3 decreased C20:4n-6 by 30% and increased by more than fivefold the C20:5n-3 content in the endometrial tissues.


   Footnotes
 
1 The authors would like to thank F. Guay, M. Guillette, M. Lachance-Cloutier, and L. St-James for technical support; E. Bérube, C. Mayrand, and D. Morissette for animal care; D. Fontaine and S. Gagné Giguère for their library assistance; S. Méthot for statistical advice; CORPAQ, Pig Improvement Co., and CIPQ for boar semen. Back

2 Contribution No. 764. Back

Received for publication June 20, 2002. Accepted for publication October 25, 2002.


   Literature Cited
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Implications
 Literature Cited
 


Agriculture Canada. 1993. Recommended Code of Practice for Care and Handling of Pigs. Pub. No. 1771E. Agriculture Canada, Ottawa, ON, Canada.

Amusquivar, E., F. J. Rupérez, C. Barbas, and E. Herrera. 2000. Low arachidonic acid rather than {alpha}-tocopherol is responsible for the delayed postnatal development in offspring of rats fed fish oil instead of olive oil during pregnancy and lactation. J. Nutr. 130:2855–2865.

Arntzen, K. J., O. L. Brekke, L. Vatten, and R. Austgulen. 1998. Reduced production of PGE2 and PGF2{alpha} from decidual cell cultures supplemented with n-3 polyunsaturated fatty acids. Prostaglandins Other Lipid Mediat. 56:183–195.

Banni, S., and J. C. Martin. 1998. Conjugated linoleic acid and metabolites. Pages 261–302 in Trans-fatty Acids in Human Nutrition. J. L. Sebedio and W. W. Christie, ed. Oily Press, Dundee.

Bazer, F. W., S. R. Marengo, R. D. Geisert, and W. W. Thatcher. 1984. Exocrine versus endocrine secretion of prostaglandin F2{alpha} in the control of pregnancy in swine. Anim. Reprod. Sci. 17:115–132.

Bee, G. 2000. Dietary conjugated linoleic acids alter adipose tissue and milk lipids of pregnant and lactating sows. J. Nutr. 130:2292–2298.

Burke, J. M., D. J Carroll, K. E. Rowe, W. W. Thatcher, and F. Stormshak. 1996. Intravascular infusion of lipid into ewes stimulates production of progesterone and prostaglandin. Biol. Reprod. 55:169–175.

Calder, P. C. 1996. Immunomodulatory and anti-inflammatory effects of n-3 polyunsaturated fatty acids. Proc. Nutr. Soc. 55:737–774.

Campbell, F. M., A. M. Clohessy, M. J. Gordon, A. K. Dutta Roy, and R. A. Riemersma. 1998. Preferential uptake of long chain polyunsaturated fatty acids by human placental cells. Pages 19–22 in Essential Fatty Acids and Eicosanoids: Invited Papers from the 4th Int. Cong., Edinburgh, Scotland.

Canadian Council on Animal Care. 1993. Guide to the Care and Use of Experimental Animals. Vol. 1. Canadian Council on Animal Care, Ottawa, ON, Canada.

Cunnane, S. C., P. A. Stitt, S. Ganguli, and J. K. Armstrong. 1990. Raised omega-3 fatty acid levels in pigs fed flax. Can. J. Anim. Sci. 70:251–254.

Dubois, D. H., L. C. Smith, and F. W. Bazer. 1993. Determination of porcine endometrial phospholipase A2 activity and detection of immunoreactive cyclooxygenase during the oestrous cycle and early pregnancy. Reprod. Fertil. Dev. 5:531–543.

Duquette, ., J. J. Matte, C. Farmer, C. L. Girard, and J. P. Laforest. 1997. Pre and post mating dietary supplements of folic acid and uterine secretory activity in gilts. Can. J. Anim. Sci. 77: 415–420.

Geisert, R. D., M. T. Zavy, R. J. Moffatt, R. M. Blair, T. Yellin, and D. J. A. Cole. 1990. Embryonic steroids and the establishment of pregnancy in pigs. J. Reprod. Fertil. (Suppl.)140:293–305.

Giguère, A., C. L. Girard, R. Lambert, J. P. Laforest, and J. J. Matte. 2000. Reproductive performance and uterine prostaglandin secretion in gilts conditioned with dead semen and receiving dietary supplements of folic acid. Can. J. Anim. Sci. 80:467–472.

INRA. 1984. L’alimentation des Animaux Monogastriques: Porc, Lapin, Volailles. Institut National de la Recherche Agronomique, Paris.

Jaffe, B. M., and H. R. Behrman. 1974. Pages 19–34 in Methods of Hormone Radioimmunoassay. Academic Press, New York.

James, M. J., R. A. Gibson, M. D’Angelo, M. A. Neumann, and L. G. Cleland. 1993. Simple relationship exists between dietary linoleate and the n-6 fatty acids of human neutrophils and plasma. Am. J. Clin. Nutr. 58:497–500.

Kennedy, T. G. 1977. Evidence for a role for prostaglandins in the initiation of blastocyst implantation in the rat. Biol. Reprod. 16:286–291.

Labarca, C., and K. Paigen. 1980. A simple, rapid and sensitive DNA assay procedure. Anal. Biochem. 102:344–352.

Laforest, J.-P., and G. J. King. 1992. Effect of intrauterine application of oestradiol-17ß and prostaglandin E2 on the porcine oestrous cycle and uterine endocrinology. J. Reprod. Fertil. 94:381–394.

Levin, G., K. L. Duffin, M. G. Obukowicz, S. L. Hummer, H. Fujiwara, P. Needleman, and A. Raz. 2002. Differential metabolism of dihomo-{gamma}-linolenic acid (DGLA) and arachidonic acid (AA) by cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2): Implications for cellular synthesis of PGE1 and PGE2. Biochem. J. (In press).

Levine, L. 1988. Introduction. Prog. Allergy. Vol. 44: 1–9. Edition Karger, Basel, Switzerland.

Lewis, G. S., P. E. Jenkins, R. L. Fogwell, and E. K. Inskeep. 1978. Concentrations of prostaglandins E2 and F2{alpha} and their relationship to luteal function in early pregnant ewes. J. Anim. Sci. 47:1314–1323.

Lewis, G. S., and R. A Waterman. 1983. Metabolism of arachidonic acid in vitro by porcine blastocysts and endometrium. Prostaglandins 25:871–880.

Li, Y., and B. A. Watkins. 1998. Conjugated linoleic acids alter bone fatty acid composition and reduce ex vivo prostaglandin E2 biosynthesis in rats fed n-6 or n-3 fatty acids. Lipids 33:417–425.

Matte, J. J. 1999. A rapid and non-surgical procedure for jugular catheterization of pigs. Lab. Anim. 33:258–264.

Miles, E. A., and P. C. Calder. 1998. Modulation of immune function by dietary fatty acids. Proc. Nutr. Soc. 57:277–292.

Norman, S. J., and N. L. Poyser. 2000. Effects of inhibitors of arachidonic acid turnover on the production of prostaglandins by the guinea-pig uterus. J. Reprod. Fertil. 118:181–186.

Park, P. W., and R. E. Goins. 1994. In situ preparation of fatty acid methyl esters for analysis of fatty acid composition in foods. J. Food Sci. 59:1261–1266.

Rosenkrans, C. F., Jr., B. C. Paria, D. L. Davis, and G. Milliken. 1990. In vitro synthesis of prostaglandin E and F2{alpha} by pig endometrium in the presence of estradiol, catechol estrogen and ascorbic acid. J. Anim. Sci. 68:435–443.

Sayre, B. L., and G. S. Lewis. 1993. Arachidonic acid metabolism during early development of ovine embryos: A possible relationship to shedding of the zona pellucida. Prostaglandins 45:557–569.

Shapiro, A. C., D. Wu, and S. N. Meydani. 1993. Eicosanoids derived from arachidonic and eicosapentaenoic acids inhibit T cell proliferative response. Prostaglandins 45:229–240.

Sugano, M., A. Tsujita, M. Yamasaki, K. Yamada, I. Ikeda, and D. Kritchevsky. 1997. Lymphatic recovery, tissue distribution, and metabolic effects of conjugated linoleic acid in rats. J. Nutr. Biochem. 8:38–43.

Tremblay, G. F., J. J. Matte, J. J. Dufour, and G. J. Brisson. 1989. Survival rate and development of fetuses during the first 30 days of gestation after folic acid addition to a swine diet. J. Anim. Sci. 67:724–732.

Warnants, N., M. J. van Oeckel, and C. V. Boucqué. 1999. Incorporation of dietary polyunsaturated fatty acids into pork fatty tissues. J. Anim. Sci. 77:2478–2490.

Whelan, J., K. S. Broughton, M. E. Surette, and J. E. Kinsella. 1992. Dietary arachidonic and linoleic acids: Comparative effects on tissue lipids. Lipids 27:85–88.

Whittemore, C. T., and H. Yang. 1989. Physical and chemical composition of the body of breeding sows with differing body subcutaneous fat depth at parturition, differing nutrition during lactation and differing litter size. Anim. Prod. 48:203–212.


This article has been cited by other articles:


Home page
 J DAIRY SCIHome page
E. Castaneda-Gutierrez, B. C. Benefield, M. J. de Veth, N. R. Santos, R. O. Gilbert, W. R. Butler, and D. E. Bauman
Evaluation of the Mechanism of Action of Conjugated Linoleic Acid Isomers on Reproduction in Dairy Cows
J Dairy Sci, September 1, 2007; 90(9): 4253 - 4264.
[Abstract] [Full Text] [PDF]

Home page
 J DAIRY SCIHome page
C. Caldari-Torres, C. Rodriguez-Sallaberry, E. S. Greene, and L. Badinga
Differential effects of n-3 and n-6 fatty acids on prostaglandin F2alpha production by bovine endometrial cells.
J Dairy Sci, March 1, 2006; 89(3): 971 - 977.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Chartrand, R.
Right arrow Articles by Laforest, J. P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Chartrand, R.
Right arrow Articles by Laforest, J. P.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS