H. S. Choi

Pregnancy diagnosis in cows and heifers by determination of oestradiol-17α in faeces

Abstract The concentration of unconjugated oestradiol-17α was measured in faeces of 21 non-pregnant animals and of 39 cows and heifers between the 10th and 25th week of gestation. After the 14th week of gestation all pregnant animals secreted significantly more oestradiol-17α in the faeces than non-pregnant animals. This is a possible method therefore for the confirmation of pregnancy not only in cows but also in heifers.

Pregnancy diagnosis in sows by estimation of oestrogens in blood, urine or faeces☆

Abstract Concentrations of oestrone sulphate in blood and of total unconjugated oestrogens in faeces were measured during the oestrous cycle and early gestation. The level of these hormones in blood and faeces was consistently higher (blood, 100%; faeces, 69–100%) between Days 24 and 30 in pregnant than in non-pregnant sows. Therefore, early pregnancy diagnosis based on the concentration of oestrogens in blood, urine and faeces was performed on Day 27, 28 or 29 after service for a total of 290 sows. Blood samples (25 μl) were taken from the tail vein of 289 sows. One hundred urine samples (1 ml) were collected by inserting a cellulose sponge into the vagina and faeces samples (2 g) were taken from the rectum of 290 sows. Pregnancy diagnosis was made on the assumption that more than 2.79 ng oestrone sulphate/ml blood, 19.7 ng oestrogens/g faeces and 65.6 ng oestrogens/ml urine indicated a pregnant animal. The efficiency of the tests was determined by comparing the diagnosis based on oestrogen concentrations with the farrowing results. The oestrone sulphate concentration in blood, as measured by enzyme immunoassay on microtitre plates, showed that 265 of the examined sows were correctly diagnosed, an accuracy of 91.7%. Seven results were false negative and 17 false positive. The determination of oestrogens in urine by radioimmunoassay (RIA) showed that 98 of 100 examined sows were correctly diagnosed and 2 were false negative. According to the oestrogen concentration in faeces, measured by RIA, 271 sows (93.5%) were correctly diagnosed, 16 were false negative and 3 false positive.

Progesterone, 17α-hydroxyprogesterone, androgens and oestrogens in bovine ovarian cysts

The concentration of steroid hormones in cystic follicular fluid was determined in cows with cystic ovaries. There was a significant difference in concentrations of the hormones from the cysts with granulosa cell layers, and the cysts without granulosa cell layers or only 2 to 3 layers with pycnotic nuclei in the granulosa cells (P < 0.01). The cystic follicles that consist of both thecal- and granulosa cell layers contained a low amount of progesterone and high levels of 17α-hydroxyprogesterone, androgens and oestrogens, not different from preovulatory follicles. In contrast, cysts that consist only of thecal cell layers contained a very high amount of progesterone, but very low levels of 17α-hydroxyprogesterone, androgens and oestrogens. That is functionally similar to the bovine corpus luteum which produces high concentrations of progesterone but has no or very low 17α-hydroxylase activity. In conclusion, the cystic follicles without granulosa cells are not capable of secreting 17α-hydroxyprogesterone, androgens and oestrogens, in spite of high levels of progesterone. It may be suggested that in these cysts there is a blockade of 17α-hydroxylase activity.

Rapid and efficient extraction of steroids from microlitre quantities of plasma using extrelut® mini-columns

Abstract The radioimmunoassay (RIA) of steroid hormones in body fluids normally involves extraction procedures which are both time consuming and costly with respect to special equipment (glassware, shakers, centrifuges etc.). The extraction stages can be omitted when direct RIA is used, but then highly specific antisera are required and only relatively high concentrations of steroids can be measured. Recently, as an alternative to conventional extraction procedures, the use was proposed [1] of columns of granular Kieselgur (Extrelut ®, Merck). We have modified such an extraction method using mini-columns of Extrelut®, and have tested the reliability and repeatability of this extraction method with respect to several steroids. Our findings are reported below.

Conversion of steroids in bovine blood in vitro

Blood samples collected from eight Braunvieh cows between the sixth and eighth month of gestation were allowed to stand with and without anticoagulant at 20 degrees C and 0 degrees C for different time periods. In these samples the degree of in vitro conversion of gestagens, androgens and estrogens was investigated. The concentrations were measured by radioimmunoassay. After 24 h at 20 degrees C, the levels of pregnenolone, progesterone, 17alpha-hydroxyprogesterone, androstenedione, dehydroepiandrosterone and estrone decreased to 62, 29, 25, 10, 34 and 44%, respectively, of the initial value and those of 17alpha, 20beta-dihydroxyprogesterone, epitestosterone and estradiol-17alpha increased to 385, 800 and 852%, respectively. The conversion was slower in clotted blood. The concentrations of testosterone and estradiol-17beta were consistent over the 24 h period. There was no marked decrease of the steroid concentration after 24 h of incubation of whole blood at 0 degrees C and of plasma at 20 degrees C. After the addition of (3)H-steroids, conversion could be demonstrated by thin-layer chromatography and autoradiography. These results demonstrate that all investigated hormones except testosterone and estradiol-17beta were metabolized by bovine blood cells.

In vitro conversion of steroid hormones in bovine ovarian follicles.

In bovine ovarian follicles an in vitro conversion of androstenedione and oestrone into oestradiol-17β, and of oestradiol-17β into oestrone was observed. There was no in vitro conversion of pregnenolone, progesterone, dehydroepiandrosterone and testosterone.

Effect of immunization of male rabbits against androstenedione.

After active immunization of male rabbits against androstenedione and with increasing antibody titre the concentration of androstenedione and testosterone in peripheral blood increased more than 1000-fold resp. 20-fold above values of control animals. On histological examination of the testes a marked Leydigs cell hyperplasia was found, though there was no difference in testis weight.

Epitestosterone and oestrogen concentration in plasma of cows during induction of parturition with prostaglandin F2α

Abstract Administration of 25 mg Dinoprost (PGF 2α -THAM) to six cows on day 270 of gestation induced parturition in five animals within 48–72 h. Six other cows served as controls. Blood samples were collected from the jugular vein three times a day and the concentrations of epitestosterone, unconjugated oestrogens (ether extractable) and conjugated oestrogens (not ether extractable) were measured by radioimmunoassay. Epitestosterone and conjugated oestrogen levels were similar to those of the control group (5–7 and 11–24 nmol/l respectively). After parturition, the concentration decreased to basal levels. The concentration of unconjugated oestrogens reached maximum levels at birth but the absolute values were lower in the group with induced parturition (2.8 ± 0.7 nmol/l) compared to the control group (9.2±1.0 nmol/l).