Immunohistochemical expression of proliferating cell nuclear antigen (PCNA) was evaluated in the endometrium of mares during estrus and at early diestrus. Three samples were collected by endometrial biopsy from 10 mares, on estrus/ second day, in the ovulation day and seven days after the ovulation day. PCNA expression was high in luminal epithelium and low in endometrial glands on samples taken on estrus/second day and on the ovulation day (p < 0.05). For samples collected on the seventh day following ovulation, the averaged PCNA immunostaining was higher in glandular epithelium (p < 0.05). The study revealed that luminal epithelial cells exhibit higher proliferation during estrus and glandular epithelial cells exhibited higher proliferation during diestrus.

Introduction: Bovine postpartum metritis causes great losses. Mast cell (MC)-released mediators participate in uterine inflammation and immune response, but their role in postpartum metritis in cows has not been reported. This study investigated the effect of endometrial MC on the disorder.Material and Methods: Ten dairy cows, at 6 to 10 days postpartum and with acute purulent metritis made up the experimental group, and 10 comparable healthy cows the control group. Endometrial histamine and IgE levels were determined by ELISA, and the MC particle state and expression of histamine H₁ (H₁R) and H₂ (H₂R) mRNA receptors were examined by transmission electron microscope and real-time quantitative PCR, respectively.Results: Endometrial histamine and IgE levels were significantly higher in the experimental group. In the control group, homogenously distributed size-varied granules were seen in MC cytoplasm of endometrium of lamina propria. In the experimental group however, these showed degranulation with features of reduction. The level of H₁R mRNA was lower in the experimental group, but that of H₂R mRNA was higher.Conclusion: The results suggest MC type I hypersensitivity characteristics during metritis, and histamine provocation of local inflammation. High expression of H₂R and low expression of H₁R inhibited the inflammatory response and prevented excessive uterine tissue damage.

Objective: To determine the effects of 2 conjugated linoleic acid (CLA) isomers (cis-9, trans-11 and trans-10, cis-12) on synthesis of prostaglandin (PG) E2 and F2α and expression of prostaglandin H synthase-2 (PGHS-2) of adult and fetal bovine endometrial epithelial cells in vitro. Sample: Primary cultures of endometrial epithelial cells obtained from 4 adult cows and 4 fetal bovine carcasses. Procedures: Cells were exposed to 0, 50, 100, or 200μM cis-9, trans-11 or trans-10, cis-12 CLA isomers for 24 hours. Culture media collected before and after 6 hours of stimulation of cells with phorbol 12-myristate 13-acetate were assayed to detect PGE2 and PGF2α via ELISA. After stimulation, cells were collected for western blot analysis to quantify PGHS-2. Results: Concentrations of PGF2α and PGE2 were significantly lower in culture media of adult and fetal endometrial epithelial cells exposed to any concentration of either CLA than they were in media of cells not exposed to CLAs. The trans-10, cis-12 CLA isomer seemed to decrease PG production more markedly than did the cis-9, trans-11 CLA isomer. Most concentrations of both CLAs significantly reduced culture media PGE2:PGF2α concentration ratios of cells. Exposure of cells to CLAs did not affect expression of PGHS-2 protein. Conclusions and Clinical Relevance: Results of this study indicated CLAs significantly decreased PGF2α and PGE2 concentrations and PGE2:PGF2α concentration ratios for cultures of adult and fetal endometrial epithelial cells with no apparent effect on PGHS-2 expression. Similar effects in cows could have effects on maternal recognition of pregnancy and immune function.

Chloramphenicol was administered by constant IV infusion to 7 healthy postpartum cows at rates predicted to approach a steady-state plasma concentration of 5 micrograms/ml. After 8 hours of constant IV infusion, uterine tissues were removed surgically and were assayed for chloramphenicol concentrations. Mean plasma-to-tissue ratios of chloramphenicol concentrations were 3.05, 3.63 (6 cows only), and 3.22 for caruncles, endometrium, and uterine wall, respectively. Plasma-to-tissue ratios of the 3 tissues were not significantly different (P greater than 0.10). Intrauterine (IU) injections of chloramphenicol (20 mg/kg of body weight) were administered to 3 healthy postpartum cows. The mean value of the fraction of the drugabsorbed from the uteri of these cows was 0.04. Mean concentrations of chloramphenicol were 43.8 micrograms/g in caruncles, 34.6 micrograms/g in endometrium, 2.8 micrograms/g in uterine wall, and 2.9 micrograms/ml in plasma 8 hours after IU injections. Chloramphenicol has now been banned for use in food-producing animals in the United States because of its potential for causing toxicosis in human beings. It is illegal to use chloramphenicol in food-producing animals in the United States and in some other countries as well. This includes use by the IU route of administration because chloramphenicol and most drugs are absorbed from the uterus into the bloodstream and are distributed to milk and tissues.

Objective: To develop a broad-range 28S ribosomal DNA quantitative PCR (qPCR) assay for detection of fungal DNA in equine endometrial samples. Sample: 12 fungal samples from a clinical diagnostic laboratory and 29 samples obtained from 17 mares. Procedures: The qPCR assay was optimized with commercially acquired fungal organisms and validated with samples obtained from the clinical diagnostic laboratory. Subsequently, 29 samples from 17 mares suspected of having fungal endometritis were evaluated via the qPCR assay and via traditional fungal culture and endometrial cytology. Amplicons from the qPCR assay were subjected to genetic sequencing to identify the organisms. Results: The qPCR assay theoretically had a detection threshold of 2 organisms of Candida albicans. Fungal DNA was amplified from all 12 fungal samples from the commercial diagnostic laboratory. Fungal identification by use of genetic sequencing was successful for 34 of 36 amplicons from the 12 samples assayed. A fungal agent was identified via qPCR assay and genetic sequencing in all 12 samples; in contrast, a fungal agent was identified in only 8 of 12 samples via standard fungal culture and biochemical analysis. The qPCR assay detected fungal DNA in samples from 12 of 17 mares suspected of having fungal endometritis. Conclusions and Clinical Relevance: A rapid, sensitive, and repeatable qPCR assay was developed for detection of fungal DNA from equine endometrial samples. The qPCR may prove to be clinically useful as an adjunct to microbial culture and cytologic examination to provide identification of fungal organisms in a timely manner.

Two experiments were performed to study the relationship between leukotriene B4 (LTB4) synthesis and placental separation and uterine involution in the cow. In experiment I, the concentration and synthesis of LTB4 by caruncular tissue was lower in cows with retained fetal membranes (RFM cows, n = 11) than in cows that expelled the fetal membranes normally (NFM cows, n = 19). The presence of bacterial cell wall, especially of alpha-hemolytic streptococci and coagulase positive staphylococci enhanced LTB4 synthesis by allantochorion only in NFM cows. In the RFM group, Escherichia coli lipopolysaccharide decreased allantochorionic LTB4 synthesis. With caruncle, only epidermal growth factor increased LTB4 production in NFM cows. In experiment II, the caruncular and endometrial secretion of LTB4 was lower in cows with subuterine involution (SUI cows, n = 5) or cows with SUI and RFM (SUI+RFM cows, n = 4) than in cows with normal uterine involution (NUI cows, n = 8). This decrease was especially noticeable in the previously gravid horn. In the three uterine involution groups, there were no differences in LTB4 synthesis by caruncular tissue taken from the previously gravid horn. However, progesterone and a bacterial suspension of E. coli reduced the synthesis of LTB4. Estradiol had no effect on LTB4 synthesis at the end of the postpartum period. These results suggest that LTB4 may play an important role in both placental separation and uterine involution in cattle and LTB4 synthesis may be modulated by endocrine and bacterial factors.

Cephapirin (20 mg/kg of body weight, IV) was administered before and after 3 doses of probenecid (25, 50, or 75 mg/kg, intragastrically, at 12-hour intervals) to 2 mares. Clearance and apparent volume of distribution, based on area under the curve, were negatively correlated with probenecid dose. Clearance of cephapirin was decreased by approximately 50% by administration of 50 mg of probenecid/kg. Serum, synovial fluid, peritoneal fluid, CSF, urinary and endometrial concentrations of cephapirin were determined after 5 doses of cephapirin (20 mg/kg, IM, at 12-hour intervals) without and with concurrently administered probenecid (50 mg/kg, intragastrically) to 6 mares, including the 2 mares given cephapirin, IV. Highest mean serum cephapirin concentrations were 16.1 +/- 2.16 micrograms/ml at 0.5 hour after the 5th cephapirin dose [postinjection (initial) hour (PIH) 48.5] in mares not given probenecid and 23.7 +/- 1.30 micrograms/ml at 1.5 hours after the 5th cephapirin dose (PIH 49.5) in mares given probenecid. Mean peak peritoneal fluid and synovial fluid cephapirin concentrations were 6.2 +/- 0.57 micrograms/ml and 6.6 +/- 0.58 micrograms/ml, respectively, without probenecid administration and 12.3 +/- 0.46 micrograms/ml and 10 +/- 0.78 micrograms/ml, respectively, with concurrent probenecid administration. Mean trough cephapirin concentrations for peritoneal and synovial fluids in mares given probenecid were 2 to 3 times higher than trough concentrations in mares not given probenecid. Overall mean cephapirin concentrations were significantly higher for serum, peritoneal fluid, synovial fluid, and endometrium when probenecid was administered concurrently with cephapirin (P less than 0.01). Cephapirin was not detected in CSF samples. Overall mean urinary cephapirin concentrations (2.47 mg/ml without concurrent probenecid administration and 3.06 mg/ml with concurrent probenecid) were not significantly different (P greater than 0.05). Mean trough serum probenecid concentration was 61.2 +/- 5.28 micrograms/ml. Highest serum probenecid concentration was 148.8 +/- 5.97 micrograms/ml, 2 hours after the 5th cephapirin dose (PIH 50). Probenecid administration increased serum, synovial fluid, peritoneal fluid, and endometrial concentrations of cephapirin in mares.

Ultrasonography was used to determine whether there is embryo reduction in mares iwth unilaterally fixed twins when a major portion of the vascularized area of the wall of one of the embryonic vesicles is in apposition with the wall of the adjacent vesicle, rather than with the endometrium (deprivation hypothesis). In addition, the effect of ovulatory pattern (synchronous and asynchronous) on the incidence of embryo reduction was studied. Twin vesicles were ultrasonically detected on days 11 to 15 (ovulation = day 0) and were examined daily until there was embryo reduction or until day 40. In 31 mares with twin embryonic vesicles, unilateral fixation (71%) was more frequent (P less than 0.05) than was bilateral fixation (29%). In 28 mares with known ovulatory patterns, synchronous ovulations did not affect the type of fixation (9/17 unilateral and 8/17 bilateral); however, for asynchronous ovulators the frequency of unilateral fixation (10/11) was greater (P less than 0.01) than the frequency of bilateral fixation (1/11). The incidence of embryo reduction was greater (P less than 0.01) for unilateral fixation (14/19) than for bilateral fixation (0/9) and was greater (P less than 0.05) for asynchronous ovulators (9/11) than for synchronous ovulators (5/17). In mares with embryo reduction, the reduction was complete before detection of both embryo propers (early reduction) in 10/14 and after detection of both embryo propers (late reduction) in 4/14. For 17 synchronous ovulators, fewer underwent early embryo reduction (0 mares) than late reduction (5 mares) or no reduction (12 mares; 4 unilateral and 8 bilateral), whereas in the 11 asynchronous ovulators, more underwent early reduction (8 mares) than late reduction (1 mare) or no reduction (2 mares; 1 unilateral and 1 bilateral; P less than 0.01). In mares with early embryo reduction, the orientation and spatial relationship of one vesicle relative to the other was not determinable until the embryo proper was detected. In the 2 mares in which the embryo proper of the survivor was detected before embryo reduction was complete, the embryo proper was located opposite to the site of reduction, ie, the vesicle that was to be eliminated impinged on the thin-walled area of the yolk sac wall of the survivor. The position of the embryo proper and its emerging allantoic sac seemed to determine whether a given conceptus survived or underwent late embryo reduction. The embryo proper, the vascularized wall of the yolk sac adjacent to the embryo proper, and the emerging allantoic sac were exposed to the endometrium (uterine lumen) in the surviving vesicles; in the vesicles that underwent reduction, much of the corresponding area of the vesicle wall was covered by the wall of the adjacent survivor. The results supported the deprivation hypothesis.

Persistence of the oxytocin receptor (OTR) in the bovine uterus during the first 7 d after calving was investigated by means of immunohistochemical staining. Immunoreactive OTRs were present in different locations in the uterus on almost all days except day 2, when staining was seen only in the endothelium of blood vessels in the endometrium and myometrium. This finding supports the hypothesis that oxytocin may be ecbolic in cows through the 1st week post partum, but further studies are required to assess the receptor functionality during this period.

Equine endometria representative of Kenney's categories I, II, and III were incubated in vitro with phosphate buffer, Streptococcus pneumoniae, or S zooepidemicus. Endometrial tissues from mares in estrus and diestrus were first categorized according to Kenney's classification, then were tested for adherence of S pneumoniae and S zooepidemicus to the epithelia. Bacteria were not observed when the endometrial tissue was incubated with phosphate buffer or S pneumoniae. There was no statistical difference in attachment of S zooepidemicus to endometrial tissue from mares in estrus or diestrus if endometrial classification was ignored. However, bacterial attachment was significantly (P less than or equal to 0.05) higher in category III endometrium during estrus.