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The aim of this work was to investigate the secretion of dehydroepiandrosterone (DHEA), testosterone (T), dihydrotestosterone (DHT) and oestradiol (E) as biological markers in response to illegal administration of testosterone, 19-nortestosterone (N) and oestradiol, either alone or in combination. Twenty male Friesian calves (age 13–14 months) were allotted to a control group (n= 5), and five experimental groups (n= 3) each. Each experimental animal was repeatedly injected with one of the following hormonal treatments: E, T, N, T+E and N+E. Circulating DHEA, T, DHT and E were determined by radioimmunoassay. The administration of T alone did not induce any variation in plasma DHEA, T, DHT and E, which were similar to those in the control group. In contrast, DHEA, T and DHT were on average significantly lower in the T+E and N-treated groups (χ<0.01), whereas the administration of N+E resulted in the reduction of plasma T and DHT without any modification of plasma DHEA. The administration of E alone or in combination increased circulating levels of E but did not affect androgen plasma profiles. The results indicate that plasma levels of T do not permit detection of illegal treatments because plasma androgens always remained within the physiological range. Illegal E treatment could be detected in blood samples when they were collected at least every 20 days.

We evaluated the agreement between ultrasonographic characteristcs of the corpus luteum (CL) and plasma progesterone (P4) concentration in dairy cows. In Phase 1 of the study, the ovaries of 8 cows were ultrasonographically examined, and P4 was analyzed daily from estrus (Day 0) to Day 4, then at Day 7 and Day 10, and again daily from Day 17 to the onset of next estrus. In Phase 2, the ovaries of 157 randomly selected Friesian cows were examined once by ultrasonography, and blood samples collected concurrently were analyzed for plasma P4. On the basis of the P4 values, the function of CLs was classified as follows: 1) non-secretory CL when plasma P4 was lower than 1 ng/mL (n=41); 2) evolving CL when plasma P4 was between 1 and 4 ng/mL (n=55); and 3) mid-cycle CL when plasma P4 was more than 4 ng/mL (n=61). On the basis of ultrasonographic examination, 3 additional groups were established (absence of CL, evolving CL, midcycle CL). Ultrasonographic characteristics and size of Day 3 to 4 CLs and their respective plasma P4 concentrations were not distinguishable from those of CLs observed 3 to 4 d before the susequent estrus. The degree of agreement between the two classification was 72%. The data indicate that the functional classification of CLs is difficult to determine based on ultrasonography alone.

The ability of site-specific antipeptide antisera to enhance the biological activity of ovine FSH (oFSH) in vivowas investigated using hypopituitary Snell dwarf mice. These animals were shown to respond to increasing doses of oFSH (3·3–90 μg/day), administered in two daily injections over a 5-day treatment period, in a highly significant dose-dependent fashion. The responses measured were increases in uterine weight, ovarian weight and the index of keratinisation in vaginal smears. The dose-dependent response to oFSH confirmed the suitability of this animal model for these investigations and suggested the suboptimal dose of oFSH (20 μg/day) for use in enhancement studies.Five peptides derived from the β subunit of bovine FSH (bFSH) (A, residues 33–47; B, 40–51; C, 69–80; D, 83–94; E, 27–39) were used to generate polyclonal antipeptide antisera. Of these peptides, only A and B produced an antiserum (raised in sheep) capable of recognising 125I-bFSH in a liquid phase RIA. Antisera prepared against peptide A or peptide B were found to significantly enhance the biological activity of 20 μg oFSH/day over a 5-day treatment period. The response to antipeptide antisera alone did not differ significantly from that observed in PBS-injected control animals, neither did the response to FSH alone differ from that observed in animals treated with FSH plus preimmune serum. Thus the enhanced responses are dependent upon the presence of FSH plus antipeptide antiserum. Peptides A and B are located in a region thought to be involved in receptor recognition, this may have implications for the mechanism underlying this phenomenon and/or the structure/function relationships of FSH.That FSH-enhancing antisera can be generated by immunisation of animals with peptides A and B suggests that it may be possible to develop these peptides as vaccines capable of increasing reproductive performance, such as ovulation rate. The high degree of sequence homology between ovine, bovine and porcine (and to a lesser extent human and equine) FSH in the region covered by peptides A and B suggests that these peptides could also be used to promote and regulate ovarian function in all of these species.Journal of Endocrinology(1997) 152,355–363

After the incubation of minced mammary tissues from non-lactating/non-pregnant (NL/NP), non-lactating/pregnant (NL/P), fully lactating (FL) and late-lactating (LL) cows with [14C]-labelled pregnenolone or progesterone and dehydroepiandrosterone (DHEA), the following metabolites were identified at all stages: 20α-dihydropregnenolone, progesterone (from pregnenolone), 5α-pregnanedione, 5α-pregnan-3β-ol-20-one, 20α- and 20β-dihydroprogesterone (from progesterone), 5-androstene-3β,17β-diol, 5α-androstanedione, 5α-androstan-3β-ol-17-one, androstenedione, testosterone and DHEA acyl ester (from DHEA). These products indicate the occurrence of 3β-hydroxysteroid dehydrogenaseΔ5−Δ4 isomerase, 17β-hydroxysteroid oxidoreductase (17β-HOR), 20α- and 20β-hydroxysteroid dehydrogenases, steroid 5α-reductase and acyl transferase activities. Incubation of mammary tissue homogenates with [1,2,6,7-3H]androstenedione and testosterone confirmed the presence of a 17β-HOR acting prevalently in a reductive way but failed to show evidence of any aromatase activity beyond background level. When total RNA from mammary tissues of NL/NP and LL cows was reverse-transcribed and amplified by polymerase chain reaction (PCR) with three sets of primers specific for bovine P450scc, P450c17 and P450arom cDNAs, no fragment of the expected size could be detected on gel. Southern analysis with corresponding digoxigenin-labelled ovarian probes, however, gave a positive signal for P450arom cDNA in five out of eight samples of LL mammary tissue. These data indicate that the bovine mammary gland has very limited steroidogenic capabilities that are essentially compatible with the terminal activation of circulating steroids from steroidogenic endocrines. It is uncertain, however, whether this conclusion applies to anestrous or ovariectomized lactating cows as well.

125I-Labelled ovine prolactin was infused for 15 min into a pudic artery supplying one mammary gland of lactating goats (n=17). Between 0 and 4·25 h significandy more total (P<0·01) and trichloroacetic acid (TCA)-precipitable (P<0·001) radioactivity appeared in the milk of the infused compared with the non-infused gland. Gel chromatography and antibody precipitation indicated the presence of undegraded 125I-labelled prolactin in milk whey. Maximum transfer occurred 60–80 min after the end of infusion suggesting passage via a transcellular route. High plasma prolactin concentrations, resulting from infusion of cold prolactin with labelled prolactin in late lactation or from seasonally elevated prolactin at peak lactation, reduced the specific activity of infused prolactin and depressed the difference in secretion of 125I-labelled prolactin into milk of infused and non-infused glands. This suggests the operation of a competitive and saturable mechanism. Together with the increase in the milk to blood ratio of prolactin in goats given long-term (3 week) bromocriptine treatment, the results suggest that the goat mammary gland has a high avidity for prolactin especially when circulating prolactin is low. There was also evidence from TCA precipitation that prolactin may be protected from degradation in these circumstances. These mechanisms may contribute to the resistance of ruminant lactation to reduction in plasma prolactin and protect lactation from seasonal prolactin fluctuations.Journal of Endocrinology(1995) 146,411–420

Luteinizing hormone (LH) was isolated from camel hypophyses by extraction, precipitation and successive ion exchange chromatography, and gel filtration. Gel electrophoresis showed three main bands at 27·5-29, 17-18 and 15-16 kDa. Biological activity of the purified product (camLH) was assayed by evaluating testosterone production of rat Leydig cells after stimulation with the isolated camLH and with porcineLH. Dose response curves indicated that the camLH and the reference porcineLH had the same potency. Immunoreactivity was tested in a heterologous radioimmunoassay (RIA) system using an anti-ovineLH serum. The camLH standard curve showed close parallelism with that of the ovineLH used as a reference. The two curves had median values (B/B_o = 50%) of 449·5 ± 17·4 and 822·0 ± 69·2 pg tube^-1 (mean ± S.E.M.) with sensitivities (B/B_o = 90%) of 58·2 ± 7·0 and 44·8 + 8·8 pg tube^-1. The purified product obtained after gel-filtration was injected into rabbits to produce the specific anti-camLH serum which was then applied to the development of the new RIA which uses the isolated camLH as a standard reference and iodinated ovineLH as labelled antigen. Sensitivity was 57·7 ± 6·4 pg tube^-1 with a median value of 238·0 ± 15·5 pg tube^-1. Specificity was tested using ovine follicle stimulating hormone (FSH) as interferant. The cross-reactivity was 100% for camLH and 0·3% for ovineFSH. The anti-camLH serum showed good sensitivity and specificity in the heterologous RIA and appeared suitable for the development of a more specific homologous RIA system. Copyright 1994, 1999 Academic Press

AbstractA simple and rapid method based on the in vitrobovine lymphocyte proliferation was developed to assess the immunomodulatory activity of natural compounds extracted from plant and foods. The proliferation of lymphocytes was measured by the MTT [3-(4, 5- dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide] assay. This test was used to measure the immuno-enhancing activity of 2 plant extracts and 2 samples of fermented milk after 48 h of incubation. To evaluate the effect of the cow physiological phase on the assay response, the tests were repeated using lymphocytes taken from non-pregnant dry, pregnant dry and post partum cows. The assay was not affected by the cow physiological phase and the results obtained were comparable. Both plant extracts enhanced lymphocyte growth, in particular the extract 2 was slightly more potent than extract 1. The fermentation of milk with proteolytic starter cultures, such as FAIR E-63 and LA 2, can generate peptidic compounds from milk protein with potential immunostimulating activity. These samples tested in cell culture inhibited lymphocyte proliferation because they showed an acidic pH of 4.5. However, this bioassay was enough sensitive to detect biological activity of different compounds also at low concentrations.