Metritis and the uterine disease microbiome are associated with long-term changes in the endometrium of dairy cows†.

Cows with metritis (uterine disease) during the first 1 to 2 wk postpartum have lower pregnancy rates when inseminated later postpartum (typically >10 wk). We hypothesized that metritis and the disease-associated uterine microbiome have a long-term effect on endometrial gene expression. Changes in gene expression may inform a mechanism through which disease lowers pregnancy rates. A total of 20 cows were enrolled at 1 to 2 wk postpartum to either metritis (clinical disease; n = 10) or healthy (control; n = 10) groups and randomly assigned to be slaughtered at approximately 80 d and 165 d postpartum (mid-lactation). The microbiome of the reproductive tract was sampled to confirm the presence of pathogens that are typical of metritis. In addition to the original clinical diagnosis, study cows were retrospectively assigned to uterine-disease and control groups based on the composition of their microbiome. There was no effect of early postpartum uterine disease on the uterine microbiome at mid-lactation (time of slaughter). Nonetheless, early postpartum metritis and the disease microbiome were associated with a large number of differentially-expressed genes at mid-lactation primarily in the caruncular compared with the inter-caruncular endometrium. Gene enrichment analysis identified oxidative phosphorylation as the primary pathway increased in caruncular endometrium of diseased cows whereas growth factor signaling pathways were reduced. The current study demonstrated that metritis and a uterine disease microbiome leave a sustained imprint on gene expression in the caruncular endometrium that may explain lower fertility in cows with postpartum uterine disease.

Conceptus Estrogen and prostaglandins provide the maternal recognition of pregnancy signal to prevent Luteolysis during early pregnancy in the pig†.

Conceptus estrogens and prostaglandins have long been considered the primary signals for maternal recognition of pregnancy (MRP) in the pig. However, loss-of-function studies targeting conceptus aromatase genes (CYP19A1 and CYP19A2) and prostaglandin-endoperoxide synthase 2 (PTGS2) indicated that conceptuses can not only signal MRP without estrogens or prostaglandins but can maintain early pregnancy. However, complete loss of estrogen production leads to abortion after day 25 of gestation. Although neither conceptus estrogens nor prostaglandins had a significant effect on early maintenance of CL function alone, the two conceptus factors have a biological relationship. To investigate the role that both conceptus estrogens and prostaglandins have on MRP and maintenance of pregnancy, a triple loss-of function model (TKO) was generated for conceptus CYP19A1, CYP19A2 and PTGS2. In addition, a conceptus CYP19A2-/- model (A2KO) was established to determine the role of placental estrogen during later pregnancy. Estrogen and prostaglandin synthesis were greatly reduced in TKO conceptuses which resulted in a failure to inhibit luteolysis after day 15 of pregnancy despite the presence of conceptuses in the uterine lumen. However, A2KO placentae not only maintained functional CL but were able to maintain pregnancy to day 32 of gestation. Despite the loss of placental CYP19A2 expression, the allantois fluid content of estrogen was not affected as the placenta compensated by expressing CYP19A1 and CYP19A3, which are normally absent in controls. Results suggest conceptuses can signal MRP through production of conceptus PGE or stimulating PGE synthesis from the endometrium through conceptus estrogen. Failure of conceptuses to produce both factors results in failure of MRP and loss of pregnancy.

Conceptus interferon gamma is essential for establishment of pregnancy in the pig†.

Establishment and maintenance of pregnancy in the pig is a complex process that relies on conceptus regulation of the maternal proinflammatory response to endometrial attachment. Following elongation, pig conceptuses secrete interferon gamma (IFNG) during attachment to the endometrial luminal epithelium. The objective here was to determine if conceptus production of IFNG is important for early development and establishment of pregnancy. CRISPR/Cas9 gene editing and somatic cell nuclear transfer technologies were used to create an IFNG loss-of-function study in pigs. Wild-type (IFNG+/+) and null (IFNG-/-) fibroblast cells were used to create embryos through somatic cell nuclear transfer. IFNG expression was not detected in IFNG-/- conceptuses on either day 15 or day 17 of pregnancy. Ablation of conceptus IFNG production resulted in the reduction of stromal CD3+ and mast cells, which localized to the site of conceptus attachment on day 15. The uteri of recipients with IFNG-/- conceptuses were inflamed, hyperemic and there was an abundance of erythrocytes in the uterine lumen associated with the degenerating conceptuses. The endometrial stromal extracellular matrix was altered in the IFNG-/- embryo pregnancies and there was an increased endometrial mRNA levels for collagen XVII (COL17A1), matrilin 1 (MATN1), secreted phosphoprotein 1 (SPP1), and cysteine-rich secretory protein 3 (CRISP3), which are involved with repair and remodeling of the extracellular matrix. These results indicate conceptus IFNG production is essential in modulating the endometrial proinflammatory response for conceptus attachment and survival in pigs.

Evaluation of cervical and uterine size, at 4 weeks postpartum, as a predictor of subsequent fertility in Jersey cattle.

Uterine and cervical size of Holstein dairy cows is reported among reasons for a decline in dairy cow fertility. Therefore, the objectives of this study were to (a) determine whether size of the cervix and uterus at 4 weeks postpartum impacted subsequent fertility at first service in Jersey cattle, (b) determine whether progesterone level at 4 weeks postpartum impacted cyclicity and (c) the association of the presence of corpus luteum and uterus and cervix size. Body condition scores at calving, presence of postpartum diseases, parity number and milk weights were taken from lactating Jersey dairy cows (N = 147) for 28 days postpartum. During the fourth week postpartum, a blood sample was obtained for progesterone concentration, and transrectal ultrasonography was performed by a high-resolution ultrasound machine to determine cervical and uterine horn diameter, as well as ovarian structures measurements. Correcting for parity number, BCS at calving, presence of diseases and milk yield, cows with a cervix >2.54 ± 0.63 cm and uterine horn >2.25 ± 0.59 cm were less likely to become pregnant at first service (p = .04 and p = .003, respectively). The cows with larger cervix had a trend to be less likely to have a corpus luteum present at the 4th week of lactation (p = .067). Cows with larger uterine horn size were less likely to have a corpus luteum present at the 4th week of lactation (p = .015). It is concluded that a larger cervix and/or uterus during the postpartum was associated negatively with fertility and cyclicity in Jersey cows.

Concurrent and long-term associations between the endometrial microbiota and endometrial transcriptome in postpartum dairy cows.

BACKGROUND: Fertility in dairy cows depends on ovarian cyclicity and on uterine involution. Ovarian cyclicity and uterine involution are delayed when there is uterine dysbiosis (overgrowth of pathogenic bacteria). Fertility in dairy cows may involve a mechanism through which the uterine microbiota affects ovarian cyclicity as well as the transcriptome of the endometrium within the involuting uterus. The hypothesis was that the transcriptome of the endometrium in postpartum cows would be associated with the cyclicity status of the cow as well as the microbiota during uterine involution. The endometrium of first lactation dairy cows was sampled at 1, 5, and 9 weeks postpartum. All cows were allowed to return to cyclicity without intervention until week 5 and treated with an ovulation synchronization protocol so that sampling at week 9 was on day 13 of the estrous cycle. The endometrial microbiota was measured by 16S rRNA gene sequencing and principal component analysis. The endometrial transcriptome was measured by mRNA sequencing, differential gene expression analysis, and Ingenuity Pathway Analysis. RESULTS: The endometrial microbiota changed from week 1 to week 5 but the week 5 and week 9 microbiota were similar. The endometrial transcriptome differed for cows that were either cycling or not cycling at week 5 and cyclicity status depended in part on the endometrial microbiota. Compared with cows cycling at week 5, there were large changes in the transcriptome of cows that progressed from non-cycling at week 5 to cycling at week 9. There was evidence for concurrent and longer-term associations between the endometrial microbiota and transcriptome. The week 1 endometrial microbiota had the greatest effect on the subsequent endometrial transcriptome and this effect was greatest at week 5 and diminished by week 9. CONCLUSIONS: The cumulative response of the endometrial transcriptome to the microbiota represented the combination of past microbial exposure and current microbial exposure. The endometrial transcriptome in postpartum cows, therefore, depended on the immediate and longer-term effects of the uterine microbiota that acted directly on the uterus. There may also be an indirect mechanism through which the microbiome affects the transcriptome through the restoration of ovarian cyclicity postpartum.

The transcriptome of the endometrium and placenta is associated with pregnancy development but not lactation status in dairy cows.

Infertility in lactating dairy cows is explained partially by the metabolic state associated with high milk production. The hypothesis was that lactating and nonlactating cows would differ in endometrial and placental transcriptomes during early pregnancy (day 28 to 42) and this difference would explain the predisposition for lactating cows to have embryonic loss at that time. Cows were either milked or not milked after calving. Reproductive [endometrium (caruncular and intercaruncular) and placenta] and liver tissues were collected on day 28, 35, and 42 of pregnancy. The hypothesis was rejected because no effect of lactation on mRNA abundance within reproductive tissues was found. Large differences within liver demonstrated the utility of the model to test an effect of lactation on tissue gene expression. Major changes in gene expression in reproductive tissues across time were found. Greater activation of the transcriptome for the recruitment and activation of macrophages was found in the endometrium and placenta. Changes in glucose metabolism between day 28 and 42 included greater mRNA abundance of rate-limiting genes for gluconeogenesis in intercaruncular endometrium and evidence for the establishment of aerobic glycolysis (Warburg effect) in the placenta. Temporal changes were predicted to be controlled by CSF1, PDGFB, TGFB1, and JUN. Production of nitric oxide and reactive oxygen species by macrophages was identified as a mechanism to promote angiogenesis in the endometrium. Reported differences in pregnancy development for lactating vs. nonlactating cows could be explained by systemic glucose availability to the conceptus and appeared to be independent of the endometrial and placental transcriptomes.

Heat stress in pregnant sows: Thermal responses and subsequent performance of sows and their offspring.

Seasonal infertility is a significant problem in the swine industry, and may be influenced by photoperiod and heat stress. Heat stress during gestation in particular affects pregnancy, resulting in long-term developmental damage to the offspring. This review summarizes what is known about how heat stress on the pregnant sow affects lactation and her offspring. Sows responded to heat stress during gestation with increased rectal temperature, respiration rate, and skin temperature, and tended to reduce their activity-which may have changed their body composition, increasing the adipose-to-muscle ratio. Heat stress during gestation caused temporary insulin resistance during lactation, but this metabolic state did not seem to affect health, lactation, or rebreeding performance of the sow. Heat-stressed sows also presented with a shorter gestation period and reduced litter birth weight, although weaning weights are not affected when these sows are moved to thermoneutral conditions for lactation. The offspring of gestational heat-stressed sows, however, possessed unique phenotypes, including elevated body temperature, greater fat deposition, and impaired gonad development. Thus, gestational heat stress may significantly impact a herd through its effects on sows and their offspring. Further work is necessary to determine the magnitude of the effects across fa cilities and breeds.

Rapid conceptus elongation in the pig: An interleukin 1 beta 2 and estrogen-regulated phenomenon.

Establishment and maintenance of pregnancy in the pig involves activating many physiological, cellular, and molecular signaling pathways between the developing conceptus and hormonally regulated maternal endometrium. Rapid elongation of the pig trophoblast allows for the establishment of sufficient placental surface area for the transport of nutrients to the fetus throughout pregnancy. Estrogens secreted by the conceptus during elongation act on uterine epithelia to induce secretion of uterine factors required for conceptus development and for preventing endocrine secretion of prostaglandin F2α, which would cause luteolysis. Thus, trophoblast expansion within the uterine lumen during early gestation is an essential process for implantation and maintenance of pregnancy in species with an epitheliochorial form of placentation. In the pig, rapid conceptus elongation involves the unique expression of interleukin-1 beta 2 (IL1B2), which establishes pro-inflammatory effects that may be tempered by the spatiotemporal secretion of estrogen from the conceptuses. The present review provides current information on pig conceptus remodeling and signaling via estrogen and IL1B2 pathways, as well as endometrial responses to those conceptus factors leading to establishment of pregnancy.

Interleukins, interferons, and establishment of pregnancy in pigs.

Early pregnancy in mammals requires complex and highly orchestrated cellular and molecular interactions between specialized cells within the endometrium and the conceptus. Proinflammatory cytokines are small signaling proteins released by leukocytes that augment innate and adaptive immune responses. They are also released by the mammalian trophectoderm as the conceptus apposes the uterine surface for implantation. On approximately day 12 of development in pigs, the conceptus undergoes a rapid morphological transformation referred to as elongation while simultaneously releasing estrogens and a novel conceptus form of interleukin-1 beta (IL1ß). Following elongation, pig conceptuses express interferon gamma (IFNγ) and, in lesser amounts, interferon delta (IFNδ). Significant IFN signaling takes place within the endometrium between day 14 and 18 of pregnancy as the conceptus intimately associates with the uterine epithelium. Based on studies carried out in pigs and other mammals, the combined spacio-temporal activities of conceptus estrogens, IL1ß, and IFN set in motion a series of coordinated events that promote establishment of pregnancy. This is achieved through enhancement of conceptus development, uterine receptivity, maternal-fetal hemotropic exchange, and endometrial leukocyte function. These events require activation of specific signaling pathways within the uterine luminal epithelium, glandular epithelium, and stroma. Here, we review proinflammatory cytokine expression by pig conceptuses and the hypothesized actions of these molecules during establishment of pregnancy.

Invited review: Recommendations for reporting intervention studies on reproductive performance in dairy cattle: Improving design, analysis, and interpretation of research on reproduction.

Abundant evidence from the medical, veterinary, and animal science literature demonstrates that there is substantial room for improvement of the clarity, completeness, and accuracy of reporting of intervention studies. More rigorous reporting guidelines are needed to improve the quality of data available for use in comparisons of outcomes (or meta-analyses) of multiple studies. Because of the diversity of factors that affect reproduction and the complexity of interactions between these, a systematic approach is required to design, conduct, and analyze basic and applied studies of dairy cattle reproduction. Greater consistency, clarity, completeness, and correctness of design and reporting will improve the value of each report and allow for greater depth of evaluation in meta-analyses. Each of these benefits will improve understanding and application of current knowledge and better identify questions that require additional modeling or primary research. The proposed guidelines and checklist will aid in the design, conduct, analysis, and reporting of intervention studies. We propose an adaptation of the REFLECT (Reporting Guidelines for Randomized Controlled Trials for Livestock and Food Safety) statement to provide guidelines and a checklist specific to reporting intervention studies in dairy cattle reproduction. Furthermore, we provide recommendations that will assist investigators to produce studies with greater internal and external validity that can more often be included in systematic reviews and global meta-analyses. Such studies will also assist the development of models to describe the physiology of reproduction.

Activation of the transcription factor nuclear factor-kappa B in uterine luminal epithelial cells by interleukin 1 Beta 2: a novel interleukin 1 expressed by the elongating pig conceptus.

Conceptus mortality is greatest in mammals during the peri-implantation period, a time when conceptuses appose and attach to the uterine surface epithelium while releasing proinflammatory molecules. Interleukin 1 beta (IL1B), a master proinflammatory cytokine, is released by the primate, rodent, and pig blastocyst during the peri-implantation period and is believed to be essential for establishment of pregnancy. The gene encoding IL1B has duplicated in the pig, resulting in a novel gene. Preliminary observations indicate that the novel IL1B is specifically expressed by pig conceptuses during the peri-implantation period. To verify this, IL1B was cloned from mRNA isolated from Day 12 pig conceptuses and compared with IL1B cloned from mRNA isolated from pig peripheral blood leukocytes (PBLs). The pig conceptuses, but not the PBLs, expressed a novel IL1B, referred to here as interleukin 1 beta 2 (IL1B2). Porcine endometrium was treated with recombinant porcine interleukin 1 beta 1 (IL1B1), the prototypical cytokine, and IL1B2 proteins. Immunohistochemistry and real-time RT-PCR were used to measure activation of nuclear factor-kappa B (NFKB) and NFKB-regulated transcripts, respectively, within the endometrium. Both IL1B1 and IL1B2 activated NFKB in the uterine luminal epithelium within 4 h. The NFKB activation and related gene expression, however, were lower in endometrium treated with IL1B2, suggesting that the conceptus-derived cytokine may have reduced activity within the uterus. In conclusion, the peri-implantation pig conceptus expresses a novel IL1B that can activate NFKB within the uterine surface epithelium, likely creating a proinflammatory microenvironment during establishment of pregnancy in the pig.

Systemic antibiotic treatment of cows with metritis early postpartum does not change the progression of uterine disease or the uterine microbiome at 1 month postpartum.

Background: Postpartum uterine disease (metritis) is common in dairy cows. The disease develops within 1 week after calving and is associated with microbial dysbiosis, fever, and fetid uterine discharge. Cows with metritis have a greater likelihood of developing endometritis and infertility later postpartum. Antibiotic treatment is used to relieve symptoms of metritis but the capacity of antibiotic treatment to improve fertility later postpartum is inconsistent across published studies. We hypothesized that an antibiotic has only a short-term effect on the uterine microbiome and does not change the progression of disease from metritis to endometritis. To test this hypothesis, we studied the effects of systemic antibiotic given to cows diagnosed with metritis and healthy cows early postpartum on the development of endometritis and the uterine microbiome at 1 month postpartum. Results: Cows diagnosed with metritis were compared to healthy ones in a 2 × 2 factorial design, where they were either treated with an antibiotic (ceftiofur hydrochloride) at 7 to 10 days postpartum or left untreated. Cows were slaughtered at one month postpartum and the uterus was assessed for endometritis (presence of purulent material in the uterine lumen and inflammation in the endometrium) and uterine samples were collected for bacteriology and metagenomics (16S rRNA gene sequencing). As expected, the uterine microbiome at disease diagnosis had dysbiosis of typical metritis pathogens (e.g., Fusobacterium, Bacteroides, and Porphyromonas) in diseased compared with healthy cows. At one month postpartum, there was a tendency for more endometritis in metritis cows compared with healthy but antibiotic treatment had no effect on endometritis prevalence regardless of the original disease diagnosis. Likewise, when bacteria were cultured or sequenced, there were a greater number of species (culture) or amplicon sequence variants (ASV; sequencing) in the uterine lumen of cows with metritis. However, antibiotic treatment had no effect on the prevalence of cultured species or the composition of the detected ASV. The uterine microbiome at 1 month postpartum was associated with the clinical observation of the uterus (endometritis or healthy). Conclusions: Early postpartum antibiotic treatment only provides temporary resolution of uterine dysbiosis that is not sustained long-term. Failure to resolve the dysbiosis is associated with a greater prevalence of endometritis in cows with metritis, and the occurrence of endometritis significantly impacts fertility later postpartum.

Establishment of the uterine microbiome following artificial insemination in virgin heifers.

Introduction: The concept of a sterile uterus was challenged by recent studies that have described the microbiome of the virgin and pregnant uterus for species including humans and cattle. We designed two studies that tested whether the microbiome is introduced into the uterus when the virgin heifer is first inseminated and whether the origin of the microbiome is the vagina/cervix. Methods: The uterine microbiome was measured immediately before and after an artificial insemination (AI; Study 1; n = 7 AI and n = 6 control) and 14 d after insemination (Study 2; n = 12 AI and n = 12 control) in AI and non-AI (control) Holstein heifers. A third study (Study 3; n = 5 Holstein heifers) that included additional negative controls was subsequently conducted to support the presence of a unique microbiome within the uterus despite the low microbial biomass and regardless of insemination. Traditional bacteriological culture was performed in addition to 16S rRNA gene sequencing on the same samples to determine whether there were viable organisms in addition to those detected based on DNA sequencing (16S rRNA gene sequence). Results and discussion: Inseminating a heifer did not lead to a large change in the microbiome when assessed by traditional methods of bacterial culture or metataxonomic (16S rRNA gene) sequencing (results of Studies 1 and 2). Very few bacteria were cultured from the body or horn of the uterus regardless of whether an AI was or was not (negative control) performed. The cultured bacterial genera (e.g., Bacillus, Corynebacterium, Cutibacterium, Micrococcus, Staphylococcus, and Streptococcus) were typical of those found in the soil, environment, skin, mucous membranes, and urogenital tract of animals. Metataxonomic sequencing of 16S rRNA gene generated a large number of amplicon sequence variants (ASV), but these larger datasets that were based on DNA sequencing did not consistently demonstrate an effect of AI on the abundance of ASVs across all uterine locations compared with the external surface of the tract (e.g., perimetrium; positive control samples for environment contamination during slaughter and collection). Major genera identified by 16S rRNA gene sequencing overlapped with those identified with bacterial culture and included Cutibacterium, Staphylococcus, and Streptococcus.

The microbiome of the pregnant uterus in Holstein dairy heifers and cows assessed by bacterial culture and 16S ribosomal RNA gene sequencing.

Introduction: The possibility that there is a resident and stable commensal microbiome within the pregnant uterus has been supported and refuted by a series of recent studies. One element of most of the initial studies was that they were based primarily on 16S rRNA gene sequencing from bacteria. To account for this limitation, the current study performed both bacterial culture and 16S rRNA gene sequencing in a side-by-side manner (e.g., same tissues isolated from the same animal). Methods: The uteruses of 10 mid-pregnant (156 ± 5 d of gestation) Holstein heifers and cows were collected following slaughter. The external surface of the reproductive tract (positive control for contamination during tissue collection) as well as tissues within the pregnant uterus (placentome, inter-cotyledonary placenta, inter-caruncular endometrium, amnionic fluid, allantoic fluid, fetal abomasum content, and fetal meconium) were sampled for bacterial culture and 16S rRNA gene sequencing. Results: There were 87 unique bacterial species cultured from the external surface of the pregnant reproductive tract (contamination control) and 12 bacterial species cultured from pregnancy tissues. Six out of 10 cattle (60%) exhibited bacterial growth in at least one location within the pregnant uterus. For the metataxonomic results (16S rRNA gene sequencing), a low targeted microbial biomass was identified. Analyses of the detected amplicon sequence variants (ASV) revealed that there were: (1) genera that were prevalent on both the external surface and within the pregnant uterus; (2) genera that were prevalent on the external surface but either not detected or had very low prevalence within the pregnant uterus; and (3) genera that were either not detected or had low prevalence on the external surface but found with relatively high prevalence within the pregnant uterus. Conclusion: There are a small number of viable bacteria in the pregnant uterus. The 16S rRNA gene sequencing detected a microbial community within the pregnant uterus but with a low biomass. These results are consistent with recent studies of the pregnant bovine uterus and leave open the question of whether there is adequate microbial mass to significantly affect the biology of the normal healthy bovine pregnancy.

Factors influencing pregnancy per artificial insemination (AI) and embryonic mortality in Nelore females subjected to timed-AI in Brazil.

A greater understanding of factors influencing fertility is essential to improve pregnancy rates and reduce the occurrence of embryonic mortality in beef herds. The objective of the current study was to evaluate retrospective data of pregnancy per artificial insemination (P/AI) and pregnancy loss in Nelore females subjected to timed-AI (TAI) in Brazil. Data from 40,104 TAI collected from six breeding seasons (2016-2022) were analyzed, and the effects of animal category (e.g., classification based on age and parity), farm, month of parturition, sire, sire breed (Nelore vs Angus), estrus expression at TAI, animal temperament, and body condition scores (BCS) were evaluated. P/AI and pregnancy loss were affected (P < 0.001) by animal category. There was also an effect of farm (P = 0.0013) on P/AI and pregnancy loss (P = 0.001), as P/AI ranged from 49.28% and 55.58% and pregnancy loss from 3.37% to 6.89% across the herds evaluated. Month of parturition also affected (P < 0.001) P/AI and was higher for cows that became pregnant at the beginning of the previous breeding season. Calmer animals, presenting lower velocity scores while exiting the chute following TAI, achieved higher P/AI (P < 0.001). Lower BCS at TAI was associated (P < 0.001) with increased pregnancy loss, and BCS gain following AI was associated (P < 0.001) with reduced rates of embryonic mortality. There was a major effect (P < 0.001) of sire on P/AI and pregnancy loss, as P/AI ranged from 11% to 79%, and embryonic mortality from 0% to 40% for the bulls used in the study, highlighting the importance of the sire fertility on overall pregnancy success. Results from the current study reinforce the idea that animal age and parity at the beginning of the breeding season, BCS at the onset of estrous synchronization, BCS gain following AI, estrus expression at TAI, sire, and month of parturition are important factors influencing P/AI and rates of embryonic mortality in beef herds.

Pregnancy development from day 28 to 42 of gestation in postpartum Holstein cows that were either milked (lactating) or not milked (not lactating) after calving.

The objective was to determine if lactation affects fetal and placental development from day 28 to 42 of gestation. Bos taurus Holstein cows were assigned to one of the two treatments immediately after parturition (lactating (n=23) or nonlactating (dried off immediately after calving; n=20)). Cows were inseminated at ~60 days postpartum with semen from a single ejaculate. Pregnant cows were slaughtered at 1 of 3 days of gestation (day 28, 35, or 42) and tissues were collected. The interval to first insemination, services per conception, and days to pregnancy were similar for lactating and nonlactating cows. Lactating cows had greater plasma GH and nonesterified fatty acids. Nonlactating cows had greater plasma glucose, insulin, and IGF1. There was no effect of lactation on plasma progesterone or estradiol concentrations. Lactation had a negative effect on the weight of the fetus and placenta (weights were less in lactating cows). Fetuses collected from cows that became pregnant after first insemination were heavier than fetuses collected from cows that became pregnant after second or third insemination. Pregnancy after first insemination was associated with greater blood glucose and IGF1 during the first 30 days postpartum. The conclusions were that lactation negatively affects the growth of fetal and placental tissues perhaps through a mechanism that involves hormones and metabolites that are affected by lactation. Fetal growth within cows conceiving at first insemination compared to second or third insemination was more rapid and was associated with greater blood glucose and IGF1 early postpartum (before day 30).

Uterine progesterone receptor expression, conceptus development, and ovarian function in pigs treated with RU 486 during early pregnancy.

Establishment of pregnancy in the pig depends on down-regulation of progesterone receptor (PGR) in the uterine luminal and glandular epithelium during the first week after breeding. The present study evaluated the regulation of endometrial PGR by progesterone and the possible role of endometrial tumor necrosis factor (ligand) superfamily member 11 (TNFSF11) and nuclear factor-kappa B (NFKB) activation in PGR expression. Mature, cycling gilts were inseminated (Day 0) and assigned to either untreated control (n = 9) or one of two treatments that employed RU 486 to block progesterone action either before (treatment 1 [T1]) or after (treatment 2 [T2]) the initiation of PGR down-regulation. The T1 gilts were treated with RU 486 (400 mg/day) on Days 3-5 of pregnancy (n = 9), and T2 gilts were treated with RU 486 on Days 6 and 7 of pregnancy (n = 9). Uteri and ovaries were collected on Day 8 or 12 of gestation. The diameter of the conceptuses in T1 gilts was approximately half that in controls by Day 8, and normal conceptuses were not collected from any T1 gilts on Day 12. Endometrial PGR mRNA was more abundant in T1 and T2 gilts compared with control gilts. The PGR-B protein decreased from Day 8 to Day 12 in the luminal epithelium and, to some extent, in superficial glandular epithelium in control and T2 gilts. In T1 gilts, the PGR-B protein remained elevated (i.e., failed to undergo down-regulation) on Day 12. Blocking PGR action early in the cycle (i.e., on or before Day 5), therefore, prevented normal conceptus development, caused elevated PGR mRNA, and prevented the decrease in PGR protein that typically occurs in pigs. We could not confirm a role for NFKB activation in PGR down-regulation, because pigs with extreme differences in PGR and TNFSF11 expression (T1 and controls) had similar NFKB activation on Day 8. Activated NFKB within the luminal epithelium and glandular epithelium (both superficial and deep) was observed in T2 and control pigs on Day 12 when elongating conceptuses (presumably releasing interleukin 1 beta to activate NFKB) were recovered. Gilts treated with RU 486 had greater ovarian follicular growth and greater plasma estradiol concentrations. We conclude that the mechanisms controlling PGR down-regulation are progesterone-dependent and occur between Day 3 and Day 6 of pregnancy. NFKB activation did not appear to have a role in PGR down-regulation within the period that we studied. Blocking progesterone action after Day 6 did not reverse the process of PGR down-regulation, nor did it appear to affect the development of conceptuses collected on Day 12.