Endometrial nitric oxide production and nitric oxide synthases in the equine endometrium: Relationship with microvascular density during the estrous cycle

Nitric oxide (NO) plays an important role in angiogenesis and in the regulation of the blood flow. This study was carried out to investigate (i) the effects of endogenous estrogens and progestins and exogenous progesterone (P₄) (5 ng/ml or 1 μg/ml) or estradiol 17β (E₂β) (50 pg/ml or 1 μg/ml) on in vitro endometrial NO synthesis; (ii) the presence of different isoforms of NO synthase; (iii) and their relationship to microvascular density in the equine endometrium during the estrous cycle. NOS expression was also evaluated in the myometrium. Expression of endothelial and inducible forms of NOS in the uterus was assessed by Western blot and immunocytochemistry. Vascular density in endometrial tissue was determined on histologic sections. In the luteal phase, compared to the follicular phase, endometrial NO production increased without exogenous hormones and with exogenous E₂β (1 μg/ml). Although immunocytochemistry revealed iNOS and eNOS expression in the endometrium, no positive signal for iNOS was detected by Western blot. Endothelial NOS was observed in endometrial glands, endothelial cells, fibroblasts, blood and lymphatic vessels. Endometrial eNOS expression was the highest in the follicular and mid-luteal phases while it was found to be the lowest in the early luteal phase. In the follicular phase, hyperplasia of endometrial tissue with respect to myometrium was detected. No difference in vascular density was present between phases. All together, NO may play some roles in both proliferative and secretory phases of endometrial development in the mare.     

The efficacy of a single chain recombinant equine luteinizing hormone (reLH) in mares: induction of ovulation, hormone profiles, and inter-ovulatory intervals

The objectives of this study were to determine the efficacy of recombinant equine luteinizing hormone (reLH) in shortening the time to ovulation in cycling mares and to determine the effects of treatment on endogenous hormones and inter-ovulatory intervals. In study 1, mares of light horse breeds (3–20 years) were treated with either a vehicle, various doses of reLH, or human chorionic gonadotropin (hCG). Cycling mares were examined by palpation and ultrasound per rectum daily or every 12 h from the time of treatment to ovulation. In studies 2 and 3, jugular blood samples were collected daily or every 12 h from the time of treatment to ovulation for analysis of LH, follicle stimulating hormone (FSH), estradiol-17β (E₂), and progesterone (P₄) by radioimmunoassays (RIA). Increasing doses of reLH (0.3, 0.6, 0.75, and 0.9 mg) showed increasing effectiveness at inducing ovulation within 48 h of treatment. Treatments with the 0.75 and 0.9 mg doses of reLH resulted in 90% and 80% ovulation rates, which were similar to hCG treatment (85.7%). Except for the early rise in LH after treatment with 0.5, 0.65, and 1.0 mg of reLH, hormone profiles appeared to be similar between control and treated cycles. Inter-ovulatory intervals were similar between control and treatment cycles. In conclusion, reLH is a reliable and effective ovulatory agent that does not significantly alter endogenous hormone profiles or affect inter-ovulatory intervals.     

The post-ovulatory rise in progesterone is lower and the persistence of oestrous behaviour longer during the first compared with the second cycle of the breeding season in mares

Mares are seasonally polyoestrous breeders. Therefore, the first ovulation of the season, following winter anoestrus, is the only cycle in which mares ovulate without the presence of an old CL from the previous cycle. The objective of this study was to compare the length of oestrous behaviour, and plasma progesterone concentrations during the early post-ovulatory period between mares after the first and second ovulation of the breeding season. Overall, 38 mares and 167 oestrous periods were used in the study. From those, 11 mares were used during the first and subsequent oestrous period to measure and compare the post-ovulatory rise in progesterone concentration, whereas all the mares were used to compare the length of the post-ovulatory oestrous behaviour between the first and subsequent cycles of the breeding season. The persistence of the post-ovulatory oestrus was longer (p < .001) following the first ovulation of the year (median of 52 h) compared with the subsequent ovulations (median of 36 h for second and later ovulations groups; n = 38 mares). The progesterone concentration at any of the four 8 h-intervals analysed (28, 36, 76 and 84 h post-ovulation) was lower (p < .01) following the first versus the second ovulation of the year. By 36 h post-ovulation the progesterone concentration of mares at the second ovulation of the year had passed the threshold of 2 ng/ml (2.1 ± 0.33 ng/ml), whereas in the first cycle it was 1.2 ± 0.13 ng/ml. In conclusion, mares had lower progesterone concentrations in their peripheral circulation and longer persistence of oestrous behaviour following the first ovulation of the year compared with the second and subsequent ovulatory periods of the breeding season.     

Thecoma in a mare

A 4-year-old crossbred Thoroughbred mare was referred to the University of Tehran Veterinary Medical Teaching Hospital for evaluation of a mass in the pelvic cavity. The mare had been partially anorectic and pyrexic. On clinical examination, the mare was thin and depressed, with body temperature of 38°C, a pulse rate of 38 beats/minute, and a respiratory rate of 10 breaths/minute. Palpation per rectum revealed a large, smooth and tense mass in the anterior part of the pelvic cavity. Ultrasound examination confirmed the presence of a soft tissue mass with a thick wall. The inner surface of the mass had a villous-like appearance. The stroma of the mass was mottled, with accumulation of speckled and hypoechoic to slightly hyperechoic fluid. Hormone assays revealed low serum concentration of testosterone. Serum progesterone concentration indicated that active luteal tissue and serum estradiol concentration was 24 pg/ml. On histological examination, the sections from the tumor were composed of oval- or spindle-shaped cells loosely arranged in diffuse sheets or irregularly interlacing fascicles. On the basis of these histological findings, the large tumor mass of the left ovary was considered to be a thecoma. To our knowledge, the details of the clinical, ultrasonographic, endocrinologic, and histologic findings of this tumor in the mare have not been described in the veterinary literature.     

Bladder Eversion Caused by Chronic Cystitis in an Arabian Racehorse: A Case Report

Bladder eversion is a rare condition and may occur in mares as a result of excessive straining during pregnancy or in the postpartum period. In the present case, bladder eversion was caused by chronic cystitis in a nonpregnant mare. An Arabian racehorse (mare, 3 years old) was admitted to The Racehorse Hospital of the Turkish Jockey Club with a history of lumbar pain, excessive straining, and frequently assuming the urination position. Physical examination revealed the presence of tenesmus, stranguria, passing of small amounts of urine, and a visible mucosal structure at the ventral vulvar commissure during tenesmus. Laboratory findings revealed leucocytosis, increased urine pH, proteinuria, pyuria, and hematuria. Streptococcus equi subsp. zooepidemicus and Escherichia coli were isolated and identified in urine culture. Transrectal ultrasonography revealed thickening of the bladder wall and prolapse of the bladder corpus into the bladder. In the cystoscopic examination, performed following bladder reduction, severe hyperemia, erosion, and ulcers were determined in the bladder mucosa. Chronic cystitis was treated using antibiotics, based on urine culture test results, together with steroids and non-steroidal anti-inflammatory drugs. Twenty-four hours after the start of treatment, the severity of straining was observed to have decreased, ceasing completely on Day 4, and the bladder returned to its normal position.     

Exercising the Pregnant Mare from Day 16 to Day 80 of Gestation

Our previous research has demonstrated moderate exercise can be detrimental to early pregnancy in the mare, but little work has examined exercise after pregnancy has been detected. We exercised mares (n = 8) 6 days a week for 45 min from Day 16 until Day 80 of gestation. Color Doppler ultrasonography was used to evaluate embryonic vesicle size, fetal length, and uterine blood flow. Blood was sampled every other day to analyze cortisol and progesterone concentrations. Results indicated that exercising pregnant mares (n = 4) led to greater (P < .01) cortisol concentrations 30 min after the exercise period. No overall treatment effect could be detected in progesterone concentrations; however, following Day 60 of gestation, progesterone concentrations were lower (P < .05) in exercised mares. Additionally, progesterone concentrations peaked earlier in exercised mares at Day 52 of gestation compared to peak levels at Day 68 of gestation for control mares. No significant effects were detected in embryonic vesicle size. Fetal length tended (P = .06) to be longer in the conceptus of exercised mares. Uterine blood flow did not differ between groups but increased as pregnancy progressed (P < .001) in both groups. All mares in this study went on to deliver healthy foals and suffered no difficulties during parturition. These results indicated that moderate exercise was not detrimental to mare pregnancy.     

The Use of Equine Follicle Stimulating Hormone to Increase Equine Chorionic Gonadotropin in the Pregnant Mare

Equine chorionic gonadotropin (eCG), obtained from pregnant mares, is used for assisted reproductive technologies in laboratory rodents and livestock. The objective of the present study was to use equine follicle-stimulating hormone (eFSH) to increase the incidence of twin pregnancies, through multiple ovulations, and increase eCG. Nineteen light horse–type mares were enrolled in the study. The control group (n = 9) was bred with fresh or cooled semen and given human chorionic gonadotropin (hCG) at the time of breeding. The second group (n = 10) was given 12.5 mg of eFSH intramuscularly twice a day beginning 5–7 days after ovulation. Prostaglandin F2α was administered intramuscularly the second day of eFSH treatment. Treatment with eFSH continued until follicles were >35 mm in diameter, and mares were then given no treatment for 36 hours. The mares were then bred with fresh or cooled semen from the same stallion as the control group and given hCG. Blood samples were taken weekly from day 35 to day 105 after ovulation. Serum concentration of eCG was obtained, and data were analyzed with multivariate analysis using the mixed procedure. Significance was set at P < .05. Data were combined for all mares carrying twins and compared with those carrying singletons. The group of mares carrying twins had higher peak concentrations of eCG and higher values for area under the curve compared with mares carrying singletons (P < .05). These results suggest inducing twins could be a method used to increase eCG production.     

The Relationship Between the Positive Identification of the Embryo Proper in Equine Pregnancies Aged 18-28 Days and its Future Viability: A Field Study

Early embryonic loss (EEL) negatively affects the reproductive efficiency of equine reproduction. A sign of future EEL is when the embryo proper (EP) fails to develop within the embryonic vesicle after 30 days of gestation. The earlier the identification of impending EEL, the earlier the mare can be rebred to allow a second chance of pregnancy. The objectives of this study were to determine the percentage of embryonic vesicles with a visible EP at 18-28 days of gestation and to study the association between the presence/absence of the EP at different days of gestation and the future viability of the pregnancy. A total of 1,256 pregnancies were identified and followed by transrectal B-mode ultrasonography 12-45 days post ovulation in mares of the same Thoroughbred farm. The identification of the EP was attempted once during days 18-28. Pregnancy reconfirmation was performed on days 35-45. The percentage of vesicles with an EP increased gradually from days 18 (2.8%) to 21 (86.9%) (P < .05). From day 20 onward, the EEL rate of mares with vesicles without an EP was significantly higher (P < .05) than that of vesicles with a positive identification of an EP. In conclusion, the EP of the equine vesicle can be identified reliably with B-mode ultrasonography in the majority of mares (>71%) on day 20 of gestation. The lack of a positive identification of an EP from day 20 onward is associated with a higher EEL rate.     

A Modified One-Stage Repair of Third-Degree Rectovestibular Lacerations in Mares

A modified one-stage surgical repair was performed on eight mares that suffered from third-degree rectovestibular lacerations. The rectovestibular septum was reconstructed by three lines of sutures using polydioxanone. Primary healing was achieved in seven mares. Four mares got pregnant after the surgery, and no further injury was noticed at subsequent delivery. Complications of the currently used technique included pneumorectum (in one mare) and rectovestibular fistula (in another mare). The obtained results indicated that mares with third-degree rectovestibular lacerations are candidates for this modified one-stage technique with satisfactory outcome for subsequent fertility.