Non-pregnant Mares Research Articles

Transcription of equine chorionic gonadotropin/luteinizing hormone b subunit gene in the endometrium of non-pregnant mares

Transcription of equine chorionic gonadotropin/luteinizing hormone b subunit gene in the endometrium of non-pregnant mares

Features of ventrolateral abdominal wall muscles in non-pregnant mares and pregnant mares with and without abdominal wall disease

Abdominal Wall Disease an uncommon poorly understood, condition of late pregnantmares. The study objective was to establish reference measurements for abdominal wall muscle thicknesses and echotexture by breed, age, pregnancy and disease status as there were no published guidelines. Bilateral transabdominal muscle thickness measurements and observations of muscle echotexture were obtained using a 5MHz linear array probe in 4 anatomic locations: i).10 cm caudal to the sternum, ii). lateral to the umbilicus, iii). 10 cm cranial to the udder, iv). >10 cm lateral to the udder where 1 or more of the following muscles were present: rectus abdominis, pectoralis, and cutaneous trunci. Means of 3 ultrasound measurements of each muscle per location were obtained and an overall mean generated for each muscle. Twenty – six healthy mares were examined including 13 (9 light, 4

Next-generation sequencing of small non-coding RNAs (ncRNA) in equine endometrial tissue during maternal recognition of pregnancy (MRP)

Since the mechanism by which equine maternal recognition of pregnancy occurs remains unknown, and gene expression evaluations have failed to uncover any obvious candidates, we have begun to explore the role of small ncRNAs. Small ncRNAs (< 200 nt) do not code for protein and are involved in regulation of RNA processing and function. We postulate these small ncRNA, i.e., microRNA (miRNA) and small nucleolar RNA (snoRNA), play a role in regulation of equineMRP. For example, miRNAs are known to regulate mRNA through inhibition of translation or RNA degradation. SnoRNAs are responsible for rRNA processing through methylation and / or pseudouridylation, as well as alternative splicing. Our previous data revealed differences in miRNA content in serum of pregnant versus non-pregnant mares during the period of maternal recognition of pregnancy. The objective of this study was to identify all small ncRNA present in equine endometrial tissue during maternal recognition of pregnancy. This experiment utilized a crossover design with each mare serving as a pregnant and non-mated control (n 1⁄4 3/day) at days 11 and 13 post-ovulation. Mares were randomly assigned to a collection day and each provided endometrial samples from a pregnant and non-pregnant cycle. Endometrial biopsies were snap frozen and stored until RNA isolation. Total RNA was isolated using homogenization in Tri Reagent, and precipitated with chloroform

Hemostatic profile during late pregnancy and early postpartum period in mares

Hemostasis is a physiological process that prevents excessive blood loss and represents a protective mechanism at the time of delivery. Peripartum hemorrhage is a recurring hazardous condition to mare's health; therefore, we aimed to study mares' hemostatic profile to investigate whether physiological adjustments occur during late pregnancy and early postpartum. Fifteen pregnant mares have been monitored from the 34th week of pregnancy until the third week after foaling. Fifteen nonpregnant mares were used as control group. Jugular blood samples were analyzed for platelet count (Plt), prothrombin time (PT), activated partial thromboplastin time (aPTT), and fibrinogen (Fb). Platelet count showed significant changes at foaling (P < 0.05) and a negative correlation (r = −0.968; P = 0.032) with postpartum. Prothrombin time changed (P < 0.05) showing a significant correlation (r = 0.675; P = 0.016) with late pregnancy. Fibrinogen concentrations changed throughout the experimental period (P < 0.0001). The linear regression model revealed a positive correlation (r = 0.9210; P < 0.0001) between Fb and late pregnancy and a negative correlation (r = −0.9583; P = 0.042) between Fb and early postpartum. The shortening in PT recorded in the imminence of parturition along with the increase in Plt and Fb at foaling might reflect a physiological hypercoagulable state that constrains excessive bleeding, enhancing mares' odds of surviving. Our research improves the knowledge about blood coagulation in periparturient mares providing specific information on routine coagulation tests that may support in monitoring mare's hemostatic profile during late pregnancy and early postpartum.

The luteal blood flow, area and pixel intensity of corpus luteum, levels of progesterone in pregnant and nonpregnant mares in the period of 16 days after ovulation

The objective of the present study was to examine if luteal blood flow (LBF) monitoring could be used as an additional prognostic tool for early pregnancy diagnosis, and we particularly focused on the differences in LBF between pregnant and nonpregnant mares. Furthermore, other possible developmental differences of corpus luteum (CL) between pregnant and nonpregnant mares were evaluated. The CL (n = 119) of 27 mares were monitored once daily in B- and Power-Doppler Mode on days 1, 2, 9, 12, and 16 after ovulation (day 0 = ovulation). The data were evaluated using the MIXED Linear Model with repeated measures, and parameters were estimated by the REML method. The course of LBF, area of CL, and pixel intensity differed in nonpregnant mares on a day-to-day basis in contrast to more stable values in pregnant mares. Further, the profiles of the courses were identical until day 9, but since day 12 the differences between pregnant and nonpregnant mares started to be prominent. The LBF, pixel intensity, and level of progesterone (P4) were similar in all mares until day 16, when smaller LBF, lower pixel intensity, and lower levels of P4 were found in nonpregnant mares (P = 0.04, P = 0.02, P &amp;lt; 0.05, respectively). In pregnant and nonpregnant mares the LBF was weakly (r = 0.29 in both) and pixel intensity strongly (r = 0.48 and 0.59, respectively) correlated to the levels of P4. LBF was strongly correlated to the area of CL in pregnant as well as nonpregnant mares (r = 0.72 and 0.64, respectively). In accordance with the results presented in our study we can state that LBF monitoring is not a suitable tool for early pregnancy diagnosis or prognosis as the differences between pregnant and nonpregnant mares are notable &amp;ndash; similarly to other indicators of CL status &amp;ndash; just after the onset of luteolysis (day 16) when embryo itself is detectable. &amp;nbsp;

Antibody Responses to Natural Rattlesnake Envenomation and a Rattlesnake Toxoid Vaccine in Horses

Antivenom antibody titers following administration of rattlesnake venom for antivenom production in horses are well documented; however, antivenom antibody titers following natural rattlesnake envenomation in horses are not. Antibody titers produced in response to the commercially available rattlesnake venom vaccine are also not published. Our study objectives were to measure antivenom antibody titers in rattlesnake-bitten horses and compare them to titers in horses vaccinated with the rattlesnake venom vaccine. Additionally, titers were compared in pregnant versus nonpregnant horses to assess the affect of pregnancy on vaccine response and were measured pre- and postsuckle in foals of vaccinated mares to detect passive transfer of vaccine immunoglobulins. Blood samples were collected from 16 rattlesnake-bitten horses. Thirty-six horses (11 pregnant mares, 12 nonpregnant mares, 13 geldings) were vaccinated using a Crotalus atrox venom toxoid vaccine. Blood was collected before administering each vaccination and 30 days following the third vaccination. Blood was collected from foals of vaccinated mares pre- and postsuckle. All serum was assayed for anti-Crotalus atrox venom antibodies using an enzyme-linked immunosorbent assay (ELISA). Rattlesnake-bitten horses had higher (P = 0.001) titers than vaccinated horses. There was no significant difference between titers in vaccinated pregnant versus nonpregnant horses. One mare had a positive titer at foaling, and the foals had positive postsuckle titers. Antivenom antibody titer development was variable following natural envenomation and vaccination, and vaccine-induced titers were lower than natural envenomation titers. Further studies are required to determine if natural or vaccine antivenom antibody titers reduce the effects of envenomation.

Equine chorionic gonadotropin in the uterus of non-pregnant mares?

Equine chorionic gonadotropin in the uterus of non-pregnant mares?

Review of seven cases of granulosa cell tumour of the equine ovary

This paper reviews the equine granulosa cell tumour (GCT) and describes the clinicopathological features, treatment and outcome in seven cases of GCT in mares. Mares were presented with unilateral ovarian...

122 OXYTOCIN-INDUCED PROLONGED DIOESTRUS DOES NOT DOWNREGULATE THE EXPRESSION OF PROSTAGLANDIN-ENDOPEROXIDASE SYNTHASE 2 OR OXYTOCIN RECEPTOR IN EQUINE ENDOMETRIUM

Lifespan of the corpus luteum is regulated through the release of PGF2α from the endometrium. The endocrine factors regulating the timed release of prostaglandin during luteolysis and the abrogation thereof during pregnancy is only partially understood in the mare. Vanderwall et al. (2007 J. Am. Vet. Med. Assoc. 231, 1864–1867) reported that twice-daily administration of oxytocin during early dioestrus results in prolongation of luteal function. We hypothesised that the aforementioned repeated administration of oxytocin during early dioestrus results in downregulation of oxytocin receptor transcript while prostaglandin-endoperoxide synthase 2 transcript abundance is unaffected. The current study, therefore, compared the expression oxytocin receptor (OXTR) and prostaglandin-endoperoxide synthase 2 (PTGS2) in endometrial samples obtained 16 days after ovulation from nonpregnant mares (n = 5), pregnant mares (n = 5), and mares displaying prolonged dioestrus after the repeated administration of oxytocin (n = 6). Prolonged dioestrus was observed in all mares treated with oxytocin, evident by plasma progesterone levels above 6 ng mL–1 at the time of sample collection. Using quantitative real-time PCR analysis, PTGS2 showed a 4.37-fold lower abundance in pregnant v. nonpregnant mares (P = 0.009), while OXTR abundance was not affected by pregnancy status. Repeated administration of oxytocin did not lower expression levels of PTGS2 or OXTR when compared to levels observed in nonpregnant mares. As hypothesised, PTGS2 abundance was not affected by the treatment with oxytocin. Furthermore, we failed to prove the hypothesis that repeated administration of oxytocin leads to lowered expression of OXTR 16 days after ovulation in mares displaying prolonged dioestrus following repeated administration of oxytocin. A regulation of the oxytocin receptor at the post-transcriptional level, such as uncoupling of the receptor from its G-protein or possibly phosphorylation of the receptor, cannot be excluded and might account for the observed prolongation of luteal lifespan.

Expression of Aquaporin Water Channels in Equine Endometrium is Differentially Regulated During the Oestrous Cycle and Early Pregnancy

The expression of 12 different aquaporin subtypes in equine endometrium was examined at the mRNA and protein level. Endometrial samples were obtained during anoestrus, oestrus, 8, and 14days after ovulation in non-pregnant mares, and 14days after ovulation in pregnant mares. Quantitative PCR revealed a time-dependent pattern for all aquaporin subtypes examined except for AQP10 and 12. AQP3, 5 and 7 showed highest mRNA abundance 8days after ovulation, while AQP0 and 2 were most abundant at Day 14 of the cycle in non-pregnant mares. At 14days of pregnancy, AQP1, 4, 8, 9 and 11 displayed highest expression levels. Western blot analysis confirmed protein expression of AQP0, 2 and 5. Immunohistochemistry localized protein expression to luminal and glandular epithelial and stromal cells. AQP0 staining intensity was highest in samples obtained on Day 14 of the oestrous cycle. AQP2 immunoreactivity seemed to be stronger in samples collected 14days after ovulation from non-pregnant animals, in particular luminal epithelial staining. Samples collected 8days after ovulation from cyclic animals were characterized by intense AQP5 staining of glandular epithelium, predominantly in the deeper glands. Progesterone treatment of anoestrous mares did not enhance expression of AQPs, indicating that factors other than progesterone are required for the up-regulation of certain AQP subtypes during dioestrus. In conclusion, it seems that an equine-specific collaboration of aquaporin subtypes contributes to changes in endometrial fluid content occurring throughout the oestrous cycle and contributes to endometrial receptivity during early pregnancy in the mare.

Gene profiling of inflammatory genes in day 18 endometria from pregnant and non‐pregnant mares

Maternal recognition of pregnancy is a physiological process that primarily describes endometrial responses to a conceptus. Recognition of a conceptus prevents the release of prostaglandin F(2α) , thereby ensuring survival of the corpus luteum and continued progesterone production. Exactly how this occurs in the mare is poorly understood. Because prostaglandin F(2α) is a pro-inflammatory hormone, we hypothesized that differential gene expression in the endometrium at the time of maternal recognition reflects an anti-inflammatory event leading to decreased prostaglandin F(2α) secretion. Mares were inseminated, and endometrial biopsies were recovered from pregnant mares on Day 18 post-ovulation. In subsequent estrous cycles, mares were not inseminated and Day 18 post-ovulation endometrial biopsies were collected (non-pregnant control, matched per individual). Endometrial gene expression profiles were examined by screening an Affymetrix equine GeneChip containing probes specific for genes related to inflammatory processes. Microarray analysis revealed 118 genes that were up-regulated and 93 genes that were down-regulated (P < 0.001) at least 1.5-fold in the endometrium of pregnant versus non-pregnant mares. Quantitative, real-time RT-PCR confirmed the microarray results for three up-regulated genes homologous to TSC22D3, PPAPDC2, and KLF6, and three down-regulated genes homologous to ESR1, MARCKSL1, and EPSTI1 (P < 0.05). It is concluded that the presence of the equine embryo induces differential gene expression in the endometrium of Day 18 pregnant mares, and that these genes are associated with inflammatory processes and pathways involving cellular growth and proliferation. The results from this study provide important new insights into endometrial gene expression in response to early equine pregnancy.

Evidence for an involvement of regulatory T-lymphocytes (Treg cells) in tolerance of pregnancy in mares

treatment-associated endometrial changes were seen at Day 19 than at Day 20. Cloprostenol treatment in pregnant mares markedly increased (10-100x) expression of many endometrial genes at Day 20. Of these, several genes involved in signaling (IL6, GABRP, IGFBP-1, IL1RN, CHGA, PENK), lipid metabolism (SPLA2, SCGB1A1, SAA1, EquC1), extracellular matrix modification (MMP3, HYAL4) and lipid transport (SAA1) were most increased in treated pregnant mares compared with control pregnant mares. The changes in expression of secretory phospholipase A2 (SPLA2), uteroglobin (SCGB1A1), and a lipocalin (EquC1) were also higher in non-pregnant mares than in control pregnant mares so these changes could partly reflect transition to the non-pregnant state after luteolysis. Endometrial expression of ST6GALNAC1, PLA2G6, PLA2G4, NCAM1 and GZMB were most reduced in pregnant mares in response to cloprostenol. By comparison, gene expression in Day 19 or 20 trophoblasts from control or treated mares were similar with only 14 significant differences in the 2-5 fold range observed. These included increases in serpins B2 and B10 after cloprostenol. Major alterations in proteins in uterine flush fluids and yolk-sac fluids were assessed in a separate series in which cloprostenol was administered 3 or 4 days before collection on Days 15-21. Identified proteins were ranked in relative amounts from trypsin digested peptides identified by LC/MS/MS and Scaffold 2 software performed at the Advanced Protein Technology Centre, Toronto. Proteins corresponding to SERPINA14, ANXA1, MUC6, SPLA2, and B2Mweremost increased in uterine flush fluids after cloprostenol treatment, with SERPINA14 and SPLA2 also being increased in yolk-sac fluid. By comparison, VNN2, VNN2, GZMB, CD109 and A2Mwere consistently reduced in uterine flush fluid from treated mares. These studies reveal several alterations in the endometrium and conceptus that might beharmful effectors, or biomarkers of adverse uterine conditions, useful for clarifying the mechanisms of pregnancy establishment and pregnancy failure in mares.

Equine fetal sex determination using circulating cell-free fetal DNA ( ccffDNA)

In this study, polymerase chain reaction (PCR) reamplification of the first PCR product (2nd-PCR) and a qPCR assay were used to detect the sex determining region Y (SRY) gene from circulating cell-free fetal DNA (ccffDNA) in blood plasma of pregnant mares to determine fetal sex. The ccffDNA was isolated from plasma of 20 Thoroughbred mares (5–13 y old) in the final 3 mo of pregnancy (fetal sex was verified after foaling). For controls, plasma from two non-pregnant mares and two virgin mares were used, in addition to the non-template control. The 182 bp nucleotide sequence corresponding to the SRY-PCR product was confirmed by DNA sequencing. Based on SRY/PCR, 8 of 11 male and 9 of 9 female fetuses were correctly identified, resulting in a sensitivity of 72.7% (for male fetuses) and an overall accuracy of 85%. Furthermore, using SRY/2nd-PCR and qPCR techniques, sensitivity and accuracy were 90.9 and 95%, respectively. In conclusion, this study is apparently the first report of fetal sex determination in mares using ccffDNA.

Review of seven cases of granulosa cell tumour of the equine ovary

This paper reviews the equine granulosa cell tumour (GCT) and describes the clinicopathological features, treatment and outcome in seven cases of GCT in mares. Mares were presented with unilateral ovarian...

Concurrent Lactation and Pregnancy: Pregnant Domestic Horse Mares Do Not Increase Mother-Offspring Conflict during Intensive Lactation

Lactation is the most energy demanding part of parental care in mammals, so parent-offspring conflict arises over milk provided by the mother. In some species females commonly become pregnant shortly after parturition of previous young. This further intensifies mother-offspring conflict due to concurrent pregnancy and lactation. In equids it has been well established that pregnant females wean their foals earlier than non-pregnant ones. Intensified mother-offspring conflict was presumed to associate with pregnancy also during the period of intensive lactation, i.e., before the weaning process starts. We investigated the effect of pregnancy on suckling behaviour characteristics as indicators of mother-offspring conflict in domestic horses. Contrary to expectation, here we provide evidence of a decreased mother-offspring conflict related to pregnancy in lactating females during first two trimesters of pregnancy. Pregnant mares provided longer suckling bouts and did not reject or terminate suckling of their foals more often than non-pregnant mares. Our results suggest that pregnant mares cope with parallel investment into a nursed foal and a foetus through enhancing nursing behaviour in early stages of pregnancy before the initially low requirements of the foetus increase. They compensate their suckling foal with the perspective of its early weaning due to ongoing pregnancy.

Infection of peripheral blood mononuclear cells with neuropathogenic equine herpesvirus type-1 strain Ab4 reveals intact interferon-α induction and induces suppression of anti-inflammatory interleukin-10 responses in comparison to other viral strains

The recent increase in incidence, morbidity, and mortality of neurological disease induced by equine herpesvirus type 1 (EHV-1) has suggested a change of virulence of the virus. The exact mechanisms by which EHV-1 induces neurologic disease are not known. Environmental, viral, and host risk factors might contribute to neurological manifestation. Here, we investigated innate interferon-α (IFN-α), interleukin-10 (IL-10) and IL-4 responses after infection of equine peripheral blood mononuclear cells (PBMC) with EHV-1 using an available cytokine multiplex assay. Three viral strains representing an older isolate (RacL11), a recent abortigenic (NY03) and a neuropathogenic isolate (Ab4) were compared to identify differences in cytokine induction that might explain the increased pathogenicity of Ab4. Cytokine concentrations were also compared between foals, mares after birth, pregnant and non-pregnant mares to investigate whether immune responses to EHV-1infection are influenced by age or pregnancy status. PBMC from all groups secreted high concentrations of anti-viral IFN-α in response to EHV-1. A reduced response was observed in foals compared to non-pregnant mares. EHV-1 infection induced moderate IL-10 and overall low IL-4 secretion. Ab4 infection resulted in a significant reduction of IL-10 responses in adult horses. IL-10 and IL-4 responses were lower in foals than in most mare groups. These data suggested that EHV-1 induces robust IFN-α secretion without major differences between viral strains. However, anti-inflammatory IL-10 production was significantly reduced after infection with neuropathogenic Ab4. The ability of this EHV-1 isolate to down-regulate IL-10 production might contribute to increased local inflammation and a higher risk for neurological manifestation of the disease after infection with Ab4.

Evaluation of the effects of pregnancy on insulin sensitivity, insulin secretion, and glucose dynamics in Thoroughbred mares

To characterize the effects of pregnancy on insulin sensitivity (SI) and glucose dynamics in pasture-maintained mares fed supplemental feeds of differing energy composition. Pregnant (n = 22) and nonpregnant (10) healthy Thoroughbred mares. Pregnant and nonpregnant mares underwent frequently sampled intravenous glucose tolerance tests at 2 times (period 1, 25 to 31 weeks of gestation; period 2, 47 weeks of gestation). Following period 1 measurements, mares were provided a high-starch (HS; 39% starch) or high-fat and -fiber (14% fat and 70% fiber) supplemental feed. From a subset of mares (n = 12), blood samples were collected hourly for 24 hours to assess glycemic and insulinemic response to feeding while pastured. The minimal model of glucose and insulin dynamics was used to estimate SI, glucose effectiveness, and acute insulin response to glucose from tolerance testing data. Pregnant mares during period 1 had a lower SI and glucose effectiveness and higher acute insulin response to glucose than did nonpregnant mares. The SI value decreased in nonpregnant but not pregnant mares from periods 1 to 2. Pregnant mares fed HS feed had a greater glycemic and insulinemic response to feeding than did any other group. Pregnant mares had slower glucose clearance and greater insulin secretion at 28 weeks of gestation than did nonpregnant mares. Glucose and insulin responses to meal feeding, particularly with HS feed, were greater in pregnant mares, indicating that pregnancy enhanced the postprandial glycemic and insulinemic effects of starch-rich feed supplements.

Effect of Dietary Energy Manipulation on Mares and their Foals: Determination of Voluntary Dry Matter Intake of Mares During Late Pregnancy Using a Dual Marker System

s / Journal of Equine Veterinary Science 31 (2011) 230-356 265 energy and protein requirements (NRC, 2007). There was a significant increase in VDMI in the Lactation Phase compared to the Gestation Phases (P < .05). On average, mares consumed 10.97 .13 kg/d during gestation compared to 14.58 .42kg/d during lactation. The average VDMI of 2.45 .07% BW for the lactating mares was greater (P < .05) than the average VDMI of 1.92 .11% BW for the non-pregnant mares during the Lactation Phase. Conclusion: High forage diets can be used to meet the energy and protein needs of mares in mid-gestation. Mares in late gestation and early lactation can also be fed high forage diets if high quality forage is used.

Pattern of faecal 20-oxopregnane and oestrogen concentrations during pregnancy in wild plains zebra mares

Regulative endocrine mechanisms influence the reproductive behaviour and success of mammals, but they have been studied predominantly in domestic and captive animals. The study aims at describing the pattern of faecal 20-oxopregnane and oestrogen concentrations during pregnancy in wild plains zebra Equus quagga chapmani. Data were collected during wet and dry seasons 2007–2009. Enzyme Immunoassays were used to determine 20-oxopregnane and oestrogen concentrations in faecal samples (n=74) collected from individual mares (n=32) whose dates of foaling were known through long-term monitoring. Hormonal profiles were described with a General Additive Model (GAM: Hormone ∼ Days to Foaling). Faecal 20-oxopregnanes have a complex cycle during pregnancy (GAM, n=70, R2=0.616, p<0.001). From −250days to foaling, faecal 20-oxopregnane concentrations were above the baseline levels found in non-pregnant mares, peaking in the last 50days. Faecal oestrogen levels showed a clear peak in mid-pregnancy (GAM, n=62, R2=0.539, p<0.001). The sex of the foetus and season had no detectable effect on hormone concentrations during pregnancy. High levels (>200ng/g DW) of faecal 20-oxopregnanes associated with high (>160ng/g DW) faecal oestrogen levels indicate mid-pregnancy in c.90% of cases (16/17). High faecal 20-oxopregnanes (>200ng/g DW) and low faecal oestrogen levels (<160ng/g DW) indicate late pregnancy, again in c.90% of cases. Two faecal samples would allow the stage of pregnancy to be determined with confidence.

Functions of ectopically transplanted invasive horse trophoblast

The invasive and fully antigenic trophoblast of the chorionic girdle portion of the equine fetal membranes has the capacity to survive and differentiate after transplantation to ectopic sites. The objectives of this study were to determine i) the survival time of ectopically transplanted allogeneic trophoblast cells in non-pregnant recipient mares, ii) whether equine chorionic gonadotropin (eCG) can be delivered systemically by transplanted chorionic girdle cells, and iii) whether eCG delivered by the transplanted cells is biologically active and can suppress behavioral signs associated with estrus. Ectopically transplanted chorionic girdle survived for up to 105 days with a mean lifespan of 75 days (95% confidence interval 55-94) and secreted sufficient eCG for the hormone to be measurable in the recipients' circulation. Immunohistochemical labeling of serial biopsies of the transplant sites and measurement of eCG profiles demonstrated that graft survival was similar to the lifespan of equine endometrial cups in normal horse pregnancy. The eCG secreted by the transplanted cells induced corpora lutea formation and sustained systemic progesterone levels in the recipient mares, effects that are also observed during pregnancy. This in turn caused suppression of estrus behavior in the recipients for up to 3 months. Thus, ectopically transplanted equine trophoblast provides an unusual example of sustained viability and function of an immunogenic transplant in a recipient with an intact immune system. This model highlights the importance of innate immunoregulatory capabilities of invasive trophoblast cells and describes a new method to deliver sustained circulating concentrations of eCG in non-pregnant mares.