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The bovine oviduct provides the site for fertilization and early embryonic development. Modifications to this physiological environment, for instance the presence of pathogenic bacterial species, could diminish reproductive success at early stages of pregnancy. The aim of this study was to elucidate the inflammatory responses of bovine oviductal epithelial cells (BOEC) to a pathogenic bacterial species (Trueperella pyogenes) and a potentially pathogenic bacterium (Bacillus pumilus). BOEC from four healthy animals were isolated, cultured in passage 0 (P0) and passaged until P3. Trypan blue staining determined BOEC viability during 24 h co-culture with different multiplicities of infection (MOI) of T. pyogenes (MOI 0.01, 0.05, 0.1 and 1) or B. pumilus (MOI 1 and 10). BOEC remained viable when co-cultured with T. pyogenes at MOI 0.01 and with B. pumilus at MOI 1 and 10. Extracted total RNA from control and bacteria co-cultured samples was subjected to reverse transcription-quantitative polymerase chain reaction (RTq-PCR) to determine mRNA expression of various studied genes. The rate of release of interleukin 8 (IL8) and prostaglandin E2 (PGE2) from BOEC was measured by ELISA after 24 h co-culture with bacteria. RT-qPCR of various selected pro-inflammatory factors revealed similar mRNA expression of pro-inflammatory factors in BOEC co-cultured with T. pyogenes and in the controls. Higher mRNA expression of IL 1A, -1B, tumor necrosis factor alpha and CXC ligand (CXCL) 1/2, -3, -5 and IL8 and PG synthesis enzymes in BOEC co-cultured with B. pumilus was observed. In the presence of B. pumilus a higher amount of IL8 and PGE2 was released from BOEC than from controls. The viability and pro-inflammatory response of P3 BOEC incubated with bacteria was lower than in P0 BOEC. These findings illustrate the pathogenicity of T. pyogenes towards BOEC in detail and the potential role of B. pumilus in generating inflammation in oviductal cells. Culturing conditions influenced the pro-inflammatory responses of BOEC towards bacteria. Therefore, researchers conducting epithelial-bacterial in vitro co-culture should not underestimate the effects of these parameters.

In the uterus, the first pathogen confrontations take place at the luminal endometrial epithelium. Therefore, it is required that these cells have the potential to recognize and respond to a bacterial infection. Antimicrobial peptides (AMP), part of the innate immune system in addition to cytokines, are principal effector molecules of mucosal immunity against pathogens. One important family of AMP that can permeabilize bacterial membranes is the beta-defensin (DEFB) family, which includes the following members: DEFB1, DEFB4A, and DEFB5, lingual AMP, and tracheal AMP. The bactericidal/permeability-increasing protein is also a cationic AMP that results in the death of bacteria. Another AMP family is the S100 calcium-binding protein (S100A) family including the following members: S100A8, S100A9, S100A11, and S100A12. These AMP exert their antimicrobial action through chelation of several ions. The aim of the present study was to evaluate mRNA expression patterns of selected AMP in bovine endometrial cells collected (1) at different stages of the estrous cycle (postovulatory, early-to-mid luteal, late luteal, and pre-ovulatory phase); (2) during the puerperium depending on uterine health status (healthy, subclinical, or clinical endometritis) starting on Day 24 to 30 postpartum for 3 weeks on a weekly basis; and (3) in vitro after co-culturing with Bacillus pumilus at three different multiplicities of infection (MOI 1, 5, and 10) up to 6 hours. The results reported that the mRNA expression of all candidate AMP, except DEFB1, S100A8, and S100A9, was estrous cycle dependent. In particular, around the time of ovulation, the transcription level of most AMP was higher (P 0.05) compared with the luteal phase. Almost all candidate AMP mRNA expression was dependent on uterine health status, with a higher transcription level (P 0.05) in inflamed endometrial tissues, especially during the late stage of the puerperium (Day 45-51 postpartum). Members of the DEFB family were nearly unaffected in their mRNA expression in primary endometrial cells co-incubated with B. pumilus. However, S100A8 and S100A9 mRNA contents were higher after 4 and 6 hours of co-incubation with B. pumilus compared with untreated controls. In conclusion, higher mRNA expression of the candidate AMP around ovulation or in inflamed endometrial tissue during the puerperium suggests their crucial role in uterine innate immunity in the defense against invading bacteria.

Progesterone (P4), pregnanediol glucuronide (PdG), estradiol-17β (E2), and estrone sulfate (E1S) were measured in the feces of four female two-toed sloths (Choloepus didactylus) for early pregnancy diagnosis. For individual feces assignment, the examined female sloths were fed with a turquoise food colorant every second day. Fecal samples were collected one to four times per week, depending on the defecation rate throughout the pregnancies and the postpartum periods. The complete course of pregnancy was subdivided into three 16-week intervals (trimester of pregnancy, TP1-3) and a 5-week post-partum period after birth. Progesterone and PdG concentrations started to increase above luteal phase levels 3 weeks after conception (P = 0.028 and 0.005, respectively). At the beginning of TP1, P4 concentrations averaged 345.0 ± 283.0 ng/g and increased approximately 100- to 300-fold to a peak of 7588.0 ± 6717.0 ng/g over the TP3. Progesterone concentrations were considerably lower than PdG concentrations that started with 3206.0 ± 1500.0 ng/g at TP1 and increased up to 12.8556.0 ± 53.744.0 ng/g until birth. In contrast, mean concentrations of E2 (8.2 ± 2.4-11.7 ± 4.2 ng/g) and E1S (12.2 ± 6.7-22.9 ± 13.0 ng/g) elevated insignificantly and were not suitable for pregnancy detection. All hormones analyzed decreased rapidly within the first weeks after birth. Progesterone and PdG, as well as E2 and E1S, highly significantly correlated (r = 0.602, P < 0.001 and r = 0.497, P < 0.001, respectively) at TP1. During the TP2, only P4 and PdG significantly correlated (TP2: r = 0.661, P < 0.001 and postpartum period: r = 0.616, P = 0.009). In summary, only P4 metabolite concentrations were suitable to determine the status of reproduction in the two-toed sloth. Thereby, PdG was ideally suited to diagnose early pregnancy because it was more sensitive and detected pregnancy 2 weeks earlier than P4.

Hot season is a major constraint to production and reproduction in buffaloes. The present work aimed to investigate the effect of season on ovarian function, developmental competence, and the relative abundance of gene expression in buffalo oocytes. Three experiments were conducted. In experiment 1, pairs of buffalo ovaries were collected during cold season (CS, autumn and winter) and hot season (HS, spring and summer), and the number of antral follicles was recorded. Cumulus oocyte complexes (COCs) were aspirated and evaluated according to their morphology into four Grades. In experiment 2, Grade A and B COCs collected during CS and HS were in vitro matured (IVM) for 24 hours under standard conditions at 38.5 °C in a humidified air of 5% CO2. After IVM, cumulus cells were removed and oocytes were fixed, stained with 1% aceto-orcein, and evaluated for nuclear configuration. In vitro matured buffalo oocytes harvested during CS or HS were in vitro fertilized (IVF) using frozen-thawed buffalo semen and cultured in vitro to the blastocyst stage. In experiment 3, buffalo COCs and in vitro matured oocytes were collected during CS and HS, and then snap frozen in liquid nitrogen for gene expression analysis. Total RNA was extracted from COCs and in vitro matured oocytes, and complementary DNA was synthesized; quantitative Reverse Transcription-Polymerase Chain Reaction was performed for eight candidate genes including GAPDH, ACTB, B2M, GDF9, BMP15, HSP70, and SOD2. The results indicated that HS significantly (P < 0.01) decreased the number of antral follicles and the number of COCs recovered per ovary. The number of Grade A, B, and C COCs was lower (P < 0.05) during HS than CS. In vitro maturation of buffalo oocytes during HS significantly (P < 0.01) reduced the number of oocytes reaching the metaphase II stage and increased the percentage of degenerated oocytes compared with CS. Oocytes collected during HS also showed signs of cytoplasmic degeneration. After IVF, cleavage rate was lower (P < 0.01) for oocytes collected during HS, and the percentage of oocytes arrested at the two-cell stage was higher (P < 0.01) than oocytes IVF during CS. Oocytes matured during CS showed a higher (P < 0.01) blastocyst rate than those matured during HS. Also, COCs recovered in HS showed significant (P < 0.05) upregulation of HSP70 mRNA expression compared with those recovered in CS. For in vitro matured oocytes, CS down regulated the transcript abundance of ACTB and upregulated GAPDH and HSP70 mRNA levels compared with HS condition. In conclusion, HS could impair buffalo fertility by reducing the number of antral follicles and oocyte quality. In vitro maturation of buffalo oocytes during HS impairs their nuclear and cytoplasmic maturation, fertilization, and subsequent embryo development to the morula and blastocyst stages. This could be in part because of the altered gene expression found in COCs and in vitro matured oocytes.

Little is known about reproductive physiology in the two-toed sloth (Choloepus didactylus). Therefore, the aim of this study was to obtain detailed information about the ovarian cycle. Measurements of reliable gonadal steroids in the feces of this species were undertaken. For this purpose, fecal samples were collected one to three times per week from nonpregnant captive females (n = 2) over a 16-month period. Before assay analysis, the fecal samples were extracted with methanol. Radioimmunoassays and enzyme immunoassays for fecal progesterone, estradiol-17β, pregnanediol-glucuronide (PdG), and estrone sulfate were tested for their ability to detect the ovarian activity. Using the lowest and highest progesterone values, the ovarian cycle length was comparatively analyzed. The ovarian cycle (n = 26) averaged between 31.4 ± 9.1 days (lowest progesterone) and 32.5 ± 7.5 days (highest progesterone) throughout the whole year. The length of the follicular phase, as indicated by low progesterone levels, was 18.1 ± 4.4 days (range 12-25 days), and the length of the luteal phase, as characterized by elevated progesterone levels, was 13.2 ± 1.8 days (range 11-16 days). In contrast, estradiol-17β and estrone sulfate were not suitable to detect the cycle due to irregular collection intervals. Fecal progesterone and PdG, as well as estradiol-17β and estrone sulfate, significantly correlated (r = 0.621, P < 0.01 and r = 0.606, P < 0.01). PdG concentrations (dilution factor (DF) 1:40) were considerably higher than progesterone concentrations (DF 1:10), PdG amounted in the range of 1326.7 ± 320.2 ng/g wet feces (animal S1) and 1373.8 ± 468.3 ng/g wet feces (animal S2) compared with progesterone concentrations in the range of 98.0 ± 17.0 ng/g (S1) and 105.9 ± 30.0 ng/g (S2). The estrone sulfate levels (DF 1:2) were similar to estradiol-17β (DF 1:22). The mean fecal estradiol-17β concentrations were 6.7 ± 0.9 ng/g for animal S1 and 7.5 ± 1.6 ng/g for animal S2. In conclusion, the ovarian activity of the two-toed sloth was studied using the noninvasive method by means of the fecal steroid monitoring. Progesterone was the most reliable fecal steroid hormone to determine the duration of the ovarian cycle independent of the weekly defecation rate. The course of progesterone concentrations resulted in a cycle length of 4-5 weeks length in the two-toed sloth. Thus, the reproductive activity of the two-toed sloth does not show any seasonality like the three-toed sloth.

Shortened estrous cycles, embryonic death, and abortion might be associated with insufficient secretion of progesterone by the canine corpora lutea. The aim of this study was to investigate the concentration of progesterone, prolactin, and relaxin in hypoluteoid and control bitches during pregnancy and the nonpregnant cycle. Moreover, canine antibodies against progesterone were analyzed because of a possible connection between embryonic loss associated with hormone changes related to a hormone antibody response. An enzyme-linked immunosorbent assay (ELISA) was developed and optimized for these purposes. Serum samples from 20 short-cycling and 18 control bitches were analyzed. Animals were assigned to pregnant and nonpregnant groups after ultrasound examination. The results show that the nonpregnant, short-cycling bitches had significantly lower progesterone concentrations than the animals of the control group. In German Shepherd dogs, prolactin concentrations were significantly higher in the hypoluteoid pregnant bitches compared with the control group. Relaxin concentrations did not differ significantly between the pregnant hypoluteoid and pregnant control group at any time of measurement. Moreover, increased levels of IgE antibodies against progesterone were found in the serum of six bitches (five short-cycling bitches, one control animal) out of a total of 38 animals. The results indicated a specific binding of the antibodies to progesterone, but a cross-reaction with estradiol and testosterone might occur. The presented data verify the existence of antibodies against progesterone and suggest that these antibodies might play a role in some cases of unexplained pregnancy failure and shortened cycle length.

It is known that insulin-like growth-factor I (IGF-I) promotes early embryonic development from the morula to the blastocyst stage in rabbits (28). Therefore we used autoradiography to investigate whether IGF-I binds to preimplantation embryos and its coats. From Day 3 after mating onwards, a clear binding of IGF-I to the embryos was observed. There was no difference in binding to the embryoblast or trophoblast cells. Using ligand blot, several IGF-binding proteins (IGFBP; 31 kDa, 33 kDa, 36 kDa, three overlapping bands at 40 to 55 kDa) were obvious in the embryoblast and trophoblast. A 120 to 130 kDa protein was observed exclusively in the embryoblast. Significant binding of (125)I IGF-I to the coats of embryos older than 3 d was detected, and IGF-I was bound via a 38 kDa protein, as detected by ligand blot. To investigate the origin of this protein, the patterns of IGFBP were determined in the oviductal and uterine fluids of pregnant animals (Days 0 to 6). The following binding proteins were observed regularly in the oviductal and uterine flushings: 28 kDa, 32 kDa and 3 overlapping bands in the area of 40 to 55 kDa. In the oviduct the main IGF binding protein was the 32 kDa band (38.7% to 45.9%), while in the uterus it was the 3 overlapping bands at 40 to 55 kDa (42.5% to 24.1%). Because IGF-I is produced in the oviduct and uterus (27), IGFBPs are found in oviductal and uterine fluids, IGF-I is stored in the coats, IGF-I binds to preimplantation embryos and IGF-I promotes early embryonic development (28), the IGF system seams to have a function in the maternal-embryonic interaction.

The success of breeding programs is limited by the sparse knowledge about endocrine regulation and biochemical reactions in the psittacine male tract. The immunocytochemical analysis of parrots' testicular tissues provides an insight into their reproductive system but is often hampered by the lack of reliable antibodies. In the present study, we tested a large panel of antibodies raised against steroid receptors, steroidogenic enzymes, relaxin peptides including their receptors, and proliferation markers on paraffin sections of testicular tissue from eight psittacine genera representing three continents. All investigated species displayed the tested markers in somatic and germ cells of testis and epididymis, even though cell distribution was partly heterogenous and in species-specific patterns. The 17β-hydroxysteroid-dehydrogenase-2, 3β-hydroxysteroid-dehydrogenase, and smooth muscle actin allowed the cross-species differentiation between active and nonactive gonads. The remaining steroidogenic enzymes, steroid receptors, relaxin peptides, and Ki67 proved to be suitable to define reproductive activity depending on the parrot species. Adapting immunocytochemical methods to different psittacines was successful, though various cellular expression patterns do not allow the transfer of results among different parrot species. However, the availability of a reliable repertory of sexual markers is important to examine reproductive biology of psittacine birds.

Male chicks are an unwanted by-product when producing laying hens. The common practice to kill them directly after they have hatched gives rise to ethical concerns worldwide. The aim of this study was to develop an endocrine method to determine the sex of domestic chicken before hatch. On Days 7 to 10 of incubation, the allantoic fluid from brown layers' eggs (n = 750) was analyzed via enzyme immunoassay for their content of estradiol, estrone sulfate, and testosterone in order to detect gender differences. We successfully established a reliable method for in ovo sex identification on Day 9 of incubation by estrone sulfate measurement in the allantoic fluid. Female embryos displayed significantly higher hormone levels in the allantoic fluid than males (female: median = 0.312 ng/mL; male: median = 0.110 ng/mL; P ≤ 0.001). Our method allows the sexing of domestic chicken at a very early stage of embryonic development, even before the onset of pain perception. The possibility to eliminate eggs containing male embryos on Day 9 of incubation represents a vast improvement compared with culling day-old chicks.

The 13 kDa acidic seminal fluid protein (aSFP) is a major component of bovine semen exerting growth factor-like activity. The influence of the pure protein on sperm viability was observed by evaluating sperm motility using computer-assisted semen analysis. Furthermore, mitochondrial dehydrogenase activity as a parameter of sperm metabolism and the integrity of sperm membranes using a metal catalyzed lipid peroxidation assay were measured. Over a wide physiological range (0.003 to 4 g/l) aSFP did not influence motility and average-path velocity of sperm, but at the highest concentration (6 g/l) a significant reduction in motility could be observed. Mitochondrial activity was significantly stimulated at medium concentrations (0.125 to 2 g/l), whereas a 40% suppression was observed at maximum levels (4 g/l). A dose-dependent inhibition of lipid peroxidation could be demonstrated for medium and high concentrations of aSFP (0.125 to 4 g/l). Compared with other reducing agents, aSFP showed the highest potency in preventing oxidative stress. Such effects might be explained by the remarkable redox behavior of the protein. We suggest that in the bull aSFP may play a role in the regulation of sperm metabolism and the protection of sperm membranes from oxidative damage.

Somatotropin and FSH act synergystically on insulin-like growth factor-I (IGF-I) synthesis in ovarian follicles; IGF-I regulates several granulosa cell specific functions and may thereby be beneficial in bovine superovulation. In a series of 3 experiments we investigated the effects of recombinant bovine somatotropin (rBST) on several parameters of the superovulatory response in dairy cows. A total of 81 Holstein Friesian crossbred dairy cows received either 640 mg rBST or the vehicle (controls) on Day 4 or 13 of the superovulation schedule. Superovulation was induced with 2500 IU PMSG on Day 9. The cows were artificially inseminated on Day 13. In Experiment 1, on Days 4, 8, 11, 13 and 17 4 to 5 animals each were slaughtered to obtain follicular fluid, endometrium and plasma. The rBST application increased IGF-I contents in plasma and follicular fluid on Days 8, 11 and 13 (P0.05) in the treated cows when compared with that of the controls. Plasma and follicular IGF-I contents were correlated closely (rBST: r=0.90, n=10; control: r=0.94, n=9). The number of antral follicles increased following rBST treatment, and on the day of artificial insemination (AI) twice as many follicles4 mm were counted in the rBST treated animals than in the control group. In Experiment 2, the flushing of 38 donors on Day 7 after AI resulted in more transferable embryos in the rBST group than in the control group (4.2+/-1.0 vs 2.5+/-0.7; P0.05). In contrast, in Experiment 3 involving 21 animals when rBST was administered at the time of AI the superovulation response was not altered. It is concluded that rBST increases follicular and plasma IGF-I contents and thereby has profound effects on follicular and early embryonic development.

For exploring the processes leading to successful reproduction, differentiated long-term in vitro systems modelling the mammalian oviduct are needed. Therefore, in the present study culture conditions for primary porcine oviductal epithelial cells were optimized with regard to morphological differentiation and usability for extended cultivation periods. To evaluate different growth media for the primary cells, we used morphological criteria as well as real-time impedance measurement. After an initial media testing, the cells were grown on hanging membranes and the culture settings (conventionally cultured, serum gradient over the membrane and air-liquid interface) were assessed by histology and electron microscopy. We proved long-term expression of an oviduct specific marker (oviductal glycoprotein 1) and showed a hormone responsiveness of the culture system by means of quantitative reverse transcription-PCR. Differentiated epithelial cells could reproducibly be cultured up to 6 weeks in an air-liquid interface. After 3 weeks of culturing, the cells were clearly polarized and exhibited cilia. The model maintains physiological properties such as morphological features (mixed cell population of ciliated and secretory cells, apical cell-cell contacts typical for columnar epithelial cells) and oviduct-specific markers showing hormone responsiveness. We established a polarized long-term in vitro-system of the porcine oviductal epithelium preserving detailed features of the porcine oviduct. Therefore, we provide a useful tool to elucidate unsolved scientific questions concerning reproductive physiology.

Insulin-like growth factors 1 and 2 (IGF-1 and 2), oxytocin, progesterone, estradiol and ubiquitin were measured in bovine follicle-lutein-cysts and in follicular fluid after the classification of ovarian follicles by size (Class I =4 mm; Class II = 5-8 mm; Class III = 9-12 mm; Class IV = preovulatory; Class V = cystic). It was found that IGF-1 concentrations increased during growth from 280 ng/ml in small follicles to 489 ng/ml in preovulatory follicles; IGF-2 appeared to remain constant in follicular fluid and in cysts (275 ng/ml). Oxytocin values were low in Class I, II and III follicles (30 pg/ml) but increased in preovulatory and cystic follicles (75 pg/ml). Estradiol increased significantly only in preovulatory follicles. Ubiquitin, a protein reflecting cellular replicative activity, could be found in bovine follicular fluid in high concentrations: 1.6 mug/ml in Class I,II and III follicles with the highest amounts in preovulatory follicles (2.3 mug/ml). In contrast with normal follicles, cysts were found to have a minimal concentration of ubiquitin (0.3 mug/ml). Progesterone levels were 5 times higher in cysts (325 ng/ml) and IGF-1 concentrations were markedly higher in cystic follicles (881 ng/ml) than in the other follicles. Simultaneously, maximum gene expression for IGF-1 was found in granulosa/lutein cells of cystic follicles (Class V), suggesting de novo synthesis of IGF-1. Between the different follicle classes progesterone, oxytocin and IGF-1 concentrations correlated positively (r=0.82). Hormonal levels in follicle-lutein-cysts indicated an arrested stage of insufficient luteinization as a possible result from the premature release of LH or from the release of amounts of LH inadequate to cause ovulation.

Somatic cloning in cattle is associated with impaired embryo development, caused by inappropriate epigenetic reprogramming during embryogenesis; however, there is a paucity of data regarding gene expression at the critical elongation and peri-implantation stages. The objective of the present study was to identify genes differentially expressed in bovine cloned embryos at Day 17 of development (Day 0=day of nucleus transfer or IVF). Day 7 blastocysts (Hand Made Cloned or IVP) were transferred to recipient cattle and collected at Day 17. The efficiency of recovery of elongated embryos was similar, however cloned embryos elongated less than IVP embryos (91.8+/-45.8 vs. 174+/-50mm) and fewer had embryonic discs (63 vs. 83%). Qualitative and quantitative PCR detected expression of OCT4, NANOG, IFNtau, EOMES, FGF4, SOX2, and CDX2 in all IVP embryos. In most cloned embryos, NANOG and FGF4 were absent (verified by qPCR); NANOG, EOMES, and FGF4 were underexpressed, whereas IFNtau was overexpressed in cloned embryos. Based on qPCRs, other genes, i.e., SPARC, SNRB1, and CBPP22, were down-regulated in cloned embryos, whereas HSP70 and TDKP1 were overexpressed. In bovine microarrays, 47 genes (3.6%) were deregulated in cloned embryos, including several involved in trophoblast growth and differentiation. In conclusion, we inferred that these data were indicative of incomplete epigenetic reprogramming after cloning; this could lead to aberrant gene expression and subsequently early pregnancy loss. There was an apparent association between incomplete morphological elongation and aberrant reprogramming of a subset of genes critical for early embryonic development.

The objective of this study was to identify microRNAs (miRNAs) expressed in bovine (Bos Taurus) cloned embryos at Day 17 of development (Day 0=day of nucleus transfer or in vitro fertilization) during elongation. Day 7 bovine expanded blastocysts produced by hand made cloning (HMC) or in vitro fertilization were bulk-transferred to synchronized recipient cattle (48 HMC embryos to 10 recipients and 28 in vitro-produced embryos to four recipients). Elongated embryos were retrieved at Day 17; miRNAs were isolated and subjected to microarray screening using custom composite slides spotted with human, mouse, and rat and in silico-predicted miRNAs. An initial profile of expressed miRNAs was determined in cloned embryos and somatic donor cells; this profile changed after somatic cell nucleus transfer, identifying differentially expressed miRNAs between cloned and in vitro-produced bovine embryos. Furthermore, microarray data were validated using a miRNA-specific quantitative reverse transcription-polymerase chain reaction (qRT-PCR) approach (miR-Q). There was an 83% correlation (P=0.01) between microarray and qPCR data. Based on qRT-PCR, correct reprogramming of some miRNAs from the donor cells was confirmed in cloned bovine embryos, whereas other somatic miRNAs were not appropriately reprogrammed. Some of the miRNAs that were equally reprogrammed clustered on the same chromosomal location in the bovine genome. In conclusion, reprogramming of miRNAs seemed to occur in cloned bovine embryos. This could have profound implications for elucidating nuclear reprogramming in somatic cloning, as well as for the role of miRNAs in preimplantation mammalian development.

Several cytokines and prostaglandins play an important role in preparing the endometrium for implantation and mediating pro-inflammatory events. The aim of the present study was to examine mRNA expression of interleukin 1alpha (IL-1alpha), interleukin receptor antagonist (IL-1-RN), cytosolic prostaglandin E synthase (cPGES), microsomal PGES (mPGES-1 and mPGES-2) and lipocalin-type PGDS (L-PGDS) in the bovine endometrium. Endometrial epithelium samples were collected ex vivo from cows with different status of health at day 21-27 postpartum on a dairy farm. Three groups (n=9 animals each) were defined: (1) healthy cows with no signs of endometritis (control group), (2) cows with subclinical endometritis, and (3) cows with signs of clinical endometritis. Oestrous cycle-dependent mRNA expression pattern was investigated using bovine endometrial epithelial cells from healthy uteri collected at the abattoir. These uteri were classified into post-ovulatory, early-to-mid luteal, late luteal or pre-ovulatory phase (n=8 animals for each cycle phase). After collecting endometrial epithelium using the cytobrush-method, mRNA analysis was performed by real-time RT-PCR. L-PGDS, IL-1alpha and IL-1-RN mRNA were expressed significantly higher (P0.05) in the endometrium of cows with subclinical or clinical endometritis compared with healthy cows. A twofold lower cPGES mRNA expression (P0.05) was detected in cows with subclinical endometritis compared to healthy cows. L-PGDS and IL-1-RN mRNA expression was increased (P0.05) after ovulation compared with the pre-ovulatory or luteal phase, respectively. These results support the hypothesis that a dys-regulated cytokine and/or prostaglandin profile in the uterus could be induced by subclinical endometritis or clinical endometritis.

The aim of the present study was to investigate the effects of two medications on two subsequent abortions and plasma hormone concentrations of dogs. For this purpose, two groups of bitches (n=5 each), received the antiprogesterone aglepristone (Alizine) at 10mg/kg body weight on two subsequent days around day 30 after mating. In group II, the antiprolactin cabergoline (Galastop) was additionally administered po at 5 microg/kg body weight until the start of abortion. The plasma concentrations of relaxin, progesterone (P4) and estradiol-17beta (E2) were measured before, during and after each abortion. During the next cycle after the abortion, the same bitches were mated again and in pregnant animals, induction of abortion was performed as before. During the third cycle, pregnant bitches were allowed to whelp. Termination of first pregnancy occurred significantly earlier after the combined treatment (6.8 versus 10.6 days, p0.05). In both groups and during both abortions, relaxin varied between individuals; however, there was a continuous decrease after the abortions and no significant differences between groups (p0.05). In one bitch with high relaxin concentrations before treatment (11.6 ng/ml), a cystic endometrial hyperplasia was diagnosed. In the aglepristone only group, P4 concentrations increased significantly after the first application (p0.05), then decreased continuously until day 45 after the beginning of abortion. In the combined group, there was a continuous decrease until day 45 (p0.05). At this time, P4 concentrations between 0.47 and 84.9 nmol/l were measured in both groups. The level of E2 over time was not influenced by any medication. We therefore note that the two medications mainly influenced plasma concentrations of P4 in different ways, probably due to specific treatment-hormone interactions. However, all measurements fell within the range considered normal.

Twenty-two nonpregnant and 19 pregnant German Shepherd dogs were assigned to either a control group or a suspected short-cycling group, based on the interestrous interval (> or = 6 month and < 5 month, respectively) and data from previous pregnancies. Blood serum concentrations of progesterone and prolactin were determined from days 5 to 60 (day 0 = ovulation) for characterization of luteal function. In pregnant bitches, placental integrity was additionally assessed by relaxin concentrations. The nonpregnant, suspected short-cycling bitches had significantly lower progesterone concentrations than the controls, indicating decreased luteal activity both in the autonomous and prolactin-dependent period. In the pregnant suspected short-cycling bitches, unavoidable progesterone supplementation prevented assessment of luteal function; it may have suppressed prolactin secretion (significantly lower prolactin concentrations from days 20 to 60, compared with the pregnant control group), but deficient prolactin secretion affecting luteal function cannot be excluded. The significantly lower relaxin concentrations, together with a high incidence of embryonic death found in the pregnant, suspected short-cycling group, may indicate loss of placental integrity and may have caused decreased prolactin concentrations.