Search PubMed⌕ Search

Biomedical subjects

W R Allen

Publications and source records attributed to W R Allen.

At least 91 records · Page 5Linked to original sources
PubMed1996

The ovarian follicular wave pattern and induction of ovulation in the mated and non-mated one-humped camel (Camelus dromedarius).

Ovarian follicular wave patterns were studied ultrasonographically in three groups of dromedary. Group 1 camels (n = 20) were teased daily with a vasectomized male but mating was prevented; group 2 camels (n = 8) ran freely with a vasectomized male camel for 10 h each day and group 3 camels (n = 8) were kept completely separate from any males. In a second experiment (n = 63), when the diameter of the dominant follicle reached 0.5-0.9 cm, 1.0-1.9 cm, 2.0-2.9 cm or > 3 cm, the camel was given one of three treatments to induce ovulation: (i) natural mating; (ii) 20 micrograms of the GnRH analogue, buserelin; or (iii) 3000 iu hCG. The ovaries were re-scanned regularly to monitor ovulation, and daily blood samples were assayed for progesterone and oestradiol concentrations. The follicular cycle was divisible into a growth phase (10.5 +/- 0.5 days), a mature phase (7.6 +/- 0.8 days) and a regression phase (11.9 +/- 0.8 days). The dominant follicle reached a mean +/- SEM maximum diameter of 2.0 +/- 0.1 cm (range 1.5-2.5 cm) in 34 cycles (52%) before it began to regress. In the other 32 cycles (48%), however, the dominant follicle continued to grow to 4.2 +/- 0.2 cm (range 4.0-6.0 cm) before regression commenced. Group 2 camels were mated when their follicles reached 1.3 +/- 0.1 cm in diameter and the mean interval between successive matings was 13.8 +/- 1.0 days. Mean +/- SEM serum concentrations of oestradiol reached peak values at 39.0 +/- 1.8 pg ml-1, when the dominant follicle measured 1.7 +/- 0.1 cm and, after ovulation, mean serum concentrations of progesterone reached peak values at 2.6 +/- 0.3 ng ml-1 on day 8, before decreasing to < 1 ng ml-1 by day 10 or 11. When the dominant follicle measured 0.5-0.9 cm in diameter, 70%, 60% and 60% of them ovulated in response to mating, or treatment with buserelin or hCG, respectively. These ovulation rates increased to 85% (mating), 81% (buserelin) and 67% (hCG) when the follicle measured 1-1.9 cm, but they decreased again to 12.5% (mating), 29% (buserelin) and 13% (hCG) when the diameter had increased to 2.0-2.9 cm at the time of treatment. No follicles measuring > 3.0 cm ovulated in response to any of the treatments. These results indicated that the optimum time to mate or attempt to induce ovulation in the female dromedary is when the growing follicle measures 0.9-1.9 cm in diameter.

Animals↗
PubMed1996

Failure of endometrial cup development in the donkey-in-horse model of equine abortion.

The mature preinvasive chorionic girdles of horse, mule, donkey and extraspecies donkey-in-horse conceptuses, and the very young endometrial cups on d 37 of gestation in mares carrying horse, mule and transferred donkey-in-horse conceptuses, were compared histologically and ultrastructurally to determine possible mechanisms underlying failure of endometrial cup development in the donkey-in-horse model of equine abortion. The progenitor chorionic girdle from the failing donkey-in-house pregnancy was similar in size to the normal donkey chorionic girdle but the trophoblast cells within the former were smaller, less organised and showed definite signs of degeneration and pyknosis. In the 37 d endometrial cups both in the horse and mule pregnancies, the recently invaded, differentiated and enlarging endometrial cup cells had penetrated deeply into the endometrial stroma and were becoming tightly packed between the persisting endometrial glands. In the donkey-in-horse pregnancy, on the other hand, relatively few donkey chorionic girdle cells had begun the invasion process and the majority of these, having penetrated and dislodged the horse luminal epithelium, did not penetrate the basement membrane beneath. Very few cells had reached the endometrial stroma and these had already attracted considerable numbers of lymphocytes to the area. It is concluded that unknown factors in the horse uterus affect adversely all phases of the development, attachment and invasion of donkey chorionic girdle cells, thereby leading to very little or no endometrial cup development and equine chorionic gonadotropin secretion in the extraspecific donkey-in-horse pregnancy created by embryo transfer.

Abortion, Veterinary↗
PubMed1995

Cloning and analysis of the cDNA encoding the horse and donkey luteinizing hormone beta-subunits.

The coding regions of the horse (Equus caballus) and donkey (E. asinus) luteinizing hormone (LH) beta-subunit transcripts were cloned from pituitary gland RNA, in order to investigate their relationships to the corresponding equine chorionic gonadotropin (CG) beta-subunits and to further understand the unusual receptor-binding properties of equine LH and CG. The horse and donkey LH beta-subunit sequences were very similar (97% identity at the nucleotide (nt) level; 93% at the amino acid (aa) level), confirming their very close evolutionary linkage and also indicating that the C-terminal extension in both subunits occurred prior to the divergence of horse and donkeys. Furthermore, sequence comparisons and Southern blot analysis confirmed that in donkeys, as in horses, the pituitary (LH) and placental (CG) beta-subunits are almost certainly derived from a single gene. This contrasts with primates which have a single LH beta gene expressed in the pituitary and a family of separate CG beta genes that are expressed in the placenta. The deduced aa sequences also revealed several differences between the horse and donkey LH/CG beta-subunits that could explain their differences in biological activity. In addition, the study confirmed that the donkey subunit contains an additional Cys residue that is not present in other gonadotropin beta-subunits.

Amino Acid Sequence↗
PubMed1995

Transforming growth factor beta 1 expression in the endometrium of the mare during placentation.

In situ hybridization, Northern blotting, and immunohistochemical techniques were used to study the expression of transforming growth factor beta 1 (TGF beta 1) in the endometrium of the mare during the first 150 days of pregnancy (term = 330-340 days). In situ hybridization using an oligonucleotide (45mer) probe, based on a homologous region within all known mammalian TGF beta 1 DNA sequences, demonstrated TGF beta 1 mRNA accumulation in the glandular and lumenal epithelial cells of the endometrium from day 33 onwards which corresponds to the time of implantation (day 33-45). Expression in the endometrium remained at a high level to the end of the sampling period (day 150). There was also marked expression of TGF beta 1 in the mononuclear cells accumulated around the periphery of the specialized trophoblast cells of the endometrial cup within the endometrium and in the mononuclear cells accumulated in the endometrial stroma of mares carrying failing donkey-in-horse pregnancies created by embryo transfer. The sense (control) oligonucleotide probe exhibited no hybridization to any tissue at any stage. Northern blot analysis demonstrated that the oligonucleotide probe and a porcine TGF beta 1 cDNA clone hybridized to a single 2.5 kb transcript in horse endometrial and lymphocyte RNA, thus validating the oligonucleotide probe for detection of horse TGF beta 1 mRNA. Furthermore, both probes demonstrated an increased signal in the pregnant endometrium from day 33 onwards, thereby confirming the in situ hybridization results. Immunostaining with a specific anti-bovine TGF beta 1 serum also showed increasing TGF beta 1 accumulation in endometrial epithelia during pregnancy, and localization of the protein in endometrial stroma and in the trophoblast layer of the placenta after day 60 of pregnancy. These results show that TGF beta 1 expression increases in the maternal endometrium of the mare at the time of implantation and that it may play a role in regulating endometrial and/or trophoblast growth and differentiation during placentation in this species. It may also influence fetal development (via placental transfer) at a later stage of gestation.

Animals↗
PubMed1995

The maternal leucocyte response to the endometrial cups in horses is correlated with the developmental stages of the invasive trophoblast cells.

Invading trophoblasts form endometrial cups in the endometrium of the pregnant mare. In the present study we characterized the maternal leucocyte response to endometrial cups from their formation to their regression. The maternal leucocyte response was correlated with the stages of trophoblast development. (1) Aggregates of CD4+ and CD8+ cells were present between the migrating and differentiating endometrial cup trophoblasts and surrounding the forming endometrial cups. (2) Numbers of CD4+ cells within the mature endometrial cups were much reduced. At the periphery of the endometrial cups CD4+ and CD8+ cells were found in patchy accumulations around endometrial glands; small clusters of CD79+ B lymphocytes were present as well. (3) Scattered CD4+ and CD8+ cells were found within dying endometrial cups; areas of cell death were infiltrated with neutrophils. Large aggregates of CD4+ cells and CD8+ cells, and small but numerous clusters of CD79+ cells and eosinophils, were found outside of the dying endometrial cups. The CD4+ or CD8+ cells were mostly CD3+ T cells; some were probably macrophages which can express both of these markers in horses. The correlation between the developmental stages of the endometrial cup trophoblast and the maternal leucocyte response suggests a complicated cytokine-mediated regulatory network.

Animals↗
PubMed1995

An 80-kDa syncytiotrophoblast alloantigen bound to maternal alloantibody in term placenta.

PROBLEM: We have shown that most of the IgG present on term syncytiotrophoblast, membrane, microvesicles is bound to an 80 kDa protein antigen (R80K). METHODS: Microvesicles were prepared from term human placenta, and the IgG eluted at pH3. RESULTS: When IgG antibody was eluted at pH3 and reacted with acid-treated vesicles of other placentae, the alloantibody always bound to the preparation from which it was obtained, but only to about 10% of acid-treated preparations from other placentae. A similar polymorphic protein found in association with IgG antibody was found in term horse placentae. Cross-reactivity of the antibodies between species was not found. Using binding of labelled antibody, complement dependent cytotoxicity and FACS two-color analysis, the human polymorphic antigen was present on peripheral blood monocytes and B-lymphocytes. The R80k antigen on intact microvesicles was resistant to trypsin, but after acid elution of IgG, trypsin released a soluble 50 kDa fragment which reacted with the acid-eluted IgG antibody. CONCLUSION: The presence of antibodies to R80K in all term placentae studied, including first pregnancies, suggests that development of this alloantibody may be a normal requirement for successful pregnancy.

Animals↗
PubMed1995

Immunohistochemical localisation of steroidogenic enzymes and phenylethanolamine-N-methyl-transferase (PNMT) in the adrenal gland of the fetal and newborn foal.

An increase in fetal adrenal cortisol output signals the onset of parturition in many animal species but, in the fetal horse, plasma concentrations of cortisol remain low for much of late pregnancy, with a rise occurring only very close to the time of birth (term 320-360 days). Immunohistochemistry was used to determine the localisation and changes in distribution of key steroidogenic enzymes for cortisol production; P450scc, P450C17 and 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) in adrenal tissue from fetal and newborn horses and these findings were correlated with the appearance of immunoreactive (IR)-phenylethanolamine-N-methyl-transferase (PNMT), a cortisol-dependent enzyme. Five micron sections of adrenal tissue from fetuses at Day 100-156 (n = 5), Day 244-295 (n = 8), greater than Day 300 (n = 4) and from newborn foals (n = 6), were stained using specific antibodies and the avidin-biotin-peroxidase technique. All 3 steroidogenic enzymes were present by Day 150, but in less than 20% of the cortical cells. By late gestation the steroidogenic enzymes were present in approximately 30% of the cells, but the distribution varied. P450SCC and P450C17 predominated in cortical cells proximal to the medulla; 3 beta HSD was present throughout the cortex, but more in the zona fasciculata. In foals after birth, IR-3 beta HSD and IR-P450SCC had increased substantially throughout the adrenal cortex, and IR-P450C17 was present in most cells of the presumptive zonae fasciculata and reticularis. IR-PMNT was localised to nuclei of scattered medullary cells at the medullary-cortical interface by Day 150.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Hydroxysteroid Dehydrogenases↗
PubMed1995

Localisation of 15-hydroxy prostaglandin dehydrogenase (PGDH) and steroidogenic enzymes in the equine placenta.

15-hydroxy prostaglandin dehydrogenase (PGDH) is the critical enzyme that determines metabolism of primary prostaglandins. Its expression is determined in part by steroid hormones, particularly progesterone, formed from delta(5) steroids through 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity. To assess whether the regulation of PGDH might occur in a paracrine, autocrine or intracrine fashion, we used immunohistochemistry (IHC) to determine the localisation of key steroidogenic enzymes in the equine placenta and compared these patterns to the distribution of immunoreactive (IR-) PGDH. Placental tissue was obtained from pony or Thoroughbred mares at about Days 150, 250-280 and >300 of pregnancy (term 320 to 360 days; n=5-8 each group). IR-PGDH, 3beta-HSD, cholesterol side chain cleavage enzyme (P450(scc)) and 17-hydroxylase/lyase (P450(C17)) were localised using specific antibodies and the avidin-biotin peroxidase technique and visualised using diaminobenzidine as substrate. IR-P450(scc) was present in trophoblast cells, but not in maternal tissues of the microcotyledons. In contrast, at Days 150 and 280, IR-PGDH was present in maternal epithelial and interstitial cells in the microcotyledons, but was not detected in trophoblast epithelium, chorioallantois or endometrial glands. After Day 300, IR-PGDH was present in the maternal epithelium and interstitial cells of the placenta and it was also present in trophoblast cells in some specimens.

3-Hydroxysteroid Dehydrogenases↗
PubMed1995

Serial measurement of peripheral oestrogen and progesterone concentrations in oestrous mares to determine optimum mating time and diagnose ovulation.

Rapid enzyme-based immunoassays were used to measure concentrations of oestradiol-17 beta and progesterone in daily blood samples recovered throughout oestrus and for a few days after ovulation from 34 Thoroughbred and 8 pony-type maiden, barren and foaling mares. The first detectable fall in oestradiol-17 beta levels occurred in 88% of the mares within the interval -72 to 0 h with respect to ovulation and in 65% of mares within the interval of -48 to 0 h. The results indicated that serial daily hormone assays of this type could, in a high proportion of animals, predict a correct time for a single mating and diagnose the occurrence of ovulation in Thoroughbred mares at stud in the absence of the present management system which includes repeated veterinary examinations of the ovaries. However, the absolute need for serial daily sampling from all mares to produce interpretable hormone profiles, and the cost of this sampling and the hormone assays, make the method impractical at the present time.

Animals↗
PubMed1995

Insulin-like growth factor II gene expression in the fetus and placenta of the horse during the first half of gestation.

Placentation in equids involves two types of trophoblast: a minor invasive component, the chorionic girdle, that gives rise to transient endocrine structures known as endometrial cups, and a major non-invasive component, the allantochorion, that forms the diffuse, microcotyledonary placenta. Growth factors are likely to be important in controlling these complex events at implantation and this study describes the use of in situ hybridization and northern blotting techniques to monitor expression of insulin-like growth factor II (IGF-II) in the fetus and placenta of the horse (Equus caballus), using 12 conceptuses recovered between 14 and 150 days of gestation (term is about 340 days). The anti-sense, but not the sense, ovine IGF-II oligonucleotide probe (45 mer) hybridized to a total of seven IGF-II mRNA transcripts (6.2-1.3 kb) in RNA extracted from horse fetal liver, demonstrating the specificity and validity of the probe for equine IGF-II mRNA. In situ hybridization demonstrated that the IGF-II gene was expressed intensely in the fetus at all stages examined, predominantly in tissues of mesodermal origin, but also in the endoderm-derived liver and epithelia of the gut and lung bronchioles, and the ectoderm-derived facial mesenchyme and choroid plexus. High concentrations of IGF-II mRNA were also detected in the extraembryonic mesoderm, invasive chorionic girdle and mature endometrial cup tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Accumulation of chromotrope 2R positive cells in equine endometrium during early pregnancy and expression of transforming growth factor-beta 2 (TGF-beta 2).

Endometrial tissue from the gravid uterine horn of pregnant mares was examined by northern analysis and in situ hybridization for mRNA that hybridized to cDNA and RNA probes generated from a mouse TGF-beta 2 1.2 kb cDNA clone. The mouse cDNA probe hybridized to characteristic TGF-beta 2 mRNA transcripts on a northern blot of total RNA isolated from horse endometrium collected at day 45 of gestation. Two major 4.0 and 3.5 kb transcripts and possibly a minor 1.6 kb transcript were observed, consistent with specific hybridization to equine TGF-beta 2 mRNA. By in situ hybridization, riboprobes transcribed from the same fragment used in northern analysis hybridized to clusters of cells scattered between endometrial glands at days 38, 40, 42, 43, 78 and 81 of gestation. Positive cells were absent before day 38. From day 38 to day 43 there was marked hybridization over maternal leucocytes in the region of the developing endometrial cups, and at later stages (day 78 and day 81) clusters of cells positive for mRNA encoding TGF-beta 2 were localized within the dense band of leucocytes at the periphery of the degenerating endometrial cups. There was no hybridization to invasive or non-invasive trophoblast or to fully differentiated endometrial cup cells. Approximately 90% of the TGF-beta 2 positive cells detected in the sections taken at day 78 also stained with chromotrope 2R used to detect eosinophils and the morphology of approximately 50% of these cells was characteristic of eosinophils.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

A novel uterine protein that associates with the embryonic capsule in equids.

An apparently unique protein produced in large quantities by the endometrium of the mare which adheres to, or is incorporated into, the acellular capsule that surrounds the equine conceptus in early pregnancy, has been characterized and partially sequenced. It has a molecular mass of approximately 18 kDa on SDS-PAGE gels and is nonglycosylated as assessed by a sensitive carbohydrate detection kit. Comparison of its first 24 amino-terminal amino acids with all entries in the databases failed to show any significant identity with any other protein sequence. Secretion of the protein appears to be progesterone dependent, as its presence in uterine flushings correlates with peripheral serum progesterone profiles during the oestrous cycle and its secretion can be induced in anoestrous mares by administration of a synthetic progestagen. However, in pregnant mares, the protein disappears from the uterus after about day 20 (term = 320-340 days), despite the persistence of high serum concentrations of progesterone, indicating that additional mechanisms control its synthesis and secretion. The strong association of the protein with the glycoprotein capsule that surrounds the equine blastocyst suggests that it may be incorporated into the capsule as the capsule expands from day 11 after ovulation. Alternatively, or additionally, it may be involved in the transport of nutrients or other substances through the capsule, and may therefore play an important role in the maintenance of pregnancy.

Amino Acid Sequence↗
PubMed1994

Report of the First International Workshop on Equine Leucocyte Antigens, Cambridge, UK, July 1991.

The First International Workshop on Equine Leucocyte Antigens was organized and convened for the purposes of identifying immunologically relevant cell surface molecules of equine leucocytes and establishing a system of nomenclature for those molecules. Participating members of the workshop represented the majority of laboratories world-wide engaged in the tasks of production and characterization of equine leucocyte and lymphocyte markers using monoclonal antibodies. The workshop confirmed the identification of several equine CD molecules described previously by individual laboratories, and in addition recognized antibodies identifying new CD molecules. The workshop also succeeded in fostering co-operation between laboratories around the world which study equine immunobiology. Equine CD molecules identified by the current battery of monoclonal antibodies include EqCD2, EqCD4, EqCD5, EqCD8, EqCD11a/18, EqCD13 and EqCD44. Other antibodies are markers for MHC class I and class II molecules, for B cells, granulocytes, macrophages, T cell subsets distinct from those defined by CD4 and CD8, and other sub-populations of horse leucocytes that do not have obvious counterparts in humans, rodents, or other species. Despite the progress made in the first workshop, there are still substantial gaps in the armory of reagents available to study equine leucocyte biology, and further definition of the structure, function, and genetics of the antigens identified by the workshop clusters (WC1, WC2 etc.) and other molecules of immunological importance will be a goal of future workshops. The study of equine immunobiology and resistance to disease also urgently requires the development of tools to study equine immunoglobulins and cytokines, and these needs will provide ample scope for future studies.

Animals↗
PubMed1994

Oestrogen synthesis by the peri-implantation conceptus of the one-humped camel (Camelus dromedarius).

The ability of the embryonic tissues of the dromedary camel (Camelus dromedarius) to synthesize oestrogens in vitro was studied in 15 conceptuses recovered by non-surgical uterine lavage between 10 and 33 days after ovulation. Definitive evidence for strong aromatase activity with synthesis of considerable quantities of oestrogens was obtained at all stages when conceptus tissues were incubated with [3H]androstenedione. A high proportion of the oestrogens was in the form of oestradiol, which contrasts to the higher ratio of oestrone:oestradiol in the oestrogens synthesized by embryonic tissues of horses and pigs. Biopsies of endometrial tissue recovered from pregnant and nonpregnant camels showed great ability to conjugate both oestradiol and oestrone when incubated with tritium-labelled forms of these two hormones.

Androstenedione↗