Specialist dental training.
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Education, Dental, Graduate↗
Biomedical subjects
W R Allen
Publications and source records attributed to W R Allen.
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↗
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↗
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↗
Immunohistochemical localisation of met-enkephalin in the adrenal gland of the fetal and newborn horse.
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Adrenal Glands↗
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↗
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↗
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↗
The ovarian follicular wave pattern in the mated and non-mated dromedary camel (Camelus dromedarius).
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Animals↗
Videoendoscopic evaluation of the mare's uterus: I. Findings in normal fertile mares.
The new generation of videoendoscopes uses an electronic, instead of an optical, system for image transmission. Advantages over conventional fibre-optic endoscopes include increased image quality, handling robustness and direct display of the image on a TV monitor for multiple simultaneous viewing. In the present study, hysteroscopy was performed on 14 normal fertile Welsh Pony and Thoroughbred mares at various times during the annual and ovarian breeding cycles. Oestrus was characterised by an oedematous, relaxed cervix lying on the floor of the vagina, diffuse oedema of the endometrium and the occasional accumulation of small amounts of clear secretions in the uterine lumen. In dioestrus, the cervix appeared tight and pale and was located in the centre of the vaginal fornix. The endometrium appeared thinner due to the reduction in interstitial fluid, which allowed the underlying blood vessels to become visible, but it still had a glistening surface as a result of endometrial gland secretions. The appearance of the uterotubal papillae reflected the changes in the cervix, being relaxed, pinkish and oedematous during oestrus but remaining pale, tight and erect in dioestrus (between Days 7 and 12 after ovulation). A flexible polythene cannula was passed into the uterine lumen through the working channel of the endoscope for directed collection of uterine fluid for cytological and bacteriological examinations. Site-directed biopsies of the endometrium were taken, with flexible forceps inserted through the working channel of the endoscope, or with rigid forceps passed through the cervix alongside the endoscope. Both methods enabled accurate visual selection of the sampling area but the rigid forceps were preferred due to the larger size of the piece of endometrium recovered.
Anestrus↗
Videoendoscopic evaluation of the mare's uterus: II. Findings in subfertile mares.
Videoendoscopy of the reproductive tract was performed in 87 Thoroughbred mares with histories of reduced fertility. During hysteroscopy samples for cytological, microbiological and histological examinations were obtained under visual control. Common findings in these broodmares included: (a) endometrial degeneration, as assessed by an uneven distribution or atrophy of endometrial folds and/or a scarred appearance of the endometrium (49 mares, 56%); (b) endometrial cysts of various sizes and locations within the uterus with the most common location being at the base of the uterine horns (48 mares, 55%); (c) fluid accumulation in the uterine lumen (28 mares, 32%). A few mares had transluminal adhesions (7 mares, 8%) and in 2 mares the adhesions appeared to obstruct one uterine horn completely. A solitary discrete lump was detected in the wall of the uterine body in one mare and the suspicion of it being a leiomyoma was confirmed histologically with the aid of a visually directed biopsy sample. Free intraluminal structures were present in the uterine lumen in 3 mares, including one inspissated blood clot and two suspected remnants of resorbing pregnancies. Flexible biopsy forceps and scissors passed through the working channel of the endoscope were used to sever small thin adhesions, but this method proved inadequate for multiple adhesions or cysts. Solitary endometrial cysts were removed by means of conventional rigid biopsy forceps passed alongside the endoscope, although bleeding from the operation site usually limited this type of intervention.
Anestrus↗
Videoendoscopic evaluation of the mare's uterus: III. Findings in the pregnant mare.
During a 5-year period 65 Pony and 20 Thoroughbred pregnant mares were subjected to videoendoscopic hysteroscopy from 10 to 266 days of gestation. The aims of these examinations were to 1) observe foetal and placental development in vivo (60 Pony and 10 Thoroughbred mares); 2) eliminate one of unicornuate twin conceptuses (9 Thoroughbred mares); 3) recover embryonic foetal and placental tissues non-surgically for experimental purposes (47 Pony mares); 4) induce focal separation of the placenta in late gestation as an experimental model of placentitis (5 pony mares and 1 Thoroughbred mare). It was possible to view the embryo and all constituent extra-embryonic membranes of the conceptus between Days 10 and 87 of gestation without having to perforate the allantochorion. This allowed study of physiological processes such as the coordinated uterine contractions responsible for conceptus motility between Days 7 and 17, active foetal movements, which began as early as Day 34, and invasion of chorionic girdle cells into the endometrium, which occurred between Days 34 and 38. From Day 90, vision of the foetus was reduced or prevented by the increased thickness of the allantochorion. Transendoscopic recovery of the conceptus was successful in all of 10 mares under 30 days of gestation, whereas only 10 of 18 attempts between Days 30 and 45 produced the conceptus without resorting to uterine lavage after initial rupture of the allantochorion. All 9 attempts to eliminate one of unilateral twin conceptuses were unsuccessful and the technique was abandoned as clinically unsuitable. Nevertheless, 9 of 22 (41%) single conceptuses remained viable after one or more hysteroscopic examinations.(ABSTRACT TRUNCATED AT 250 WORDS)
Abortion, Induced↗
A gelonin-containing immunotoxin directed against human breast carcinoma.
Toxins may be specifically directed to tumor cells and the toxins' potency greatly increased by covalent conjugation to monoclonal antibodies recognizing tumor-associated antigens. Antibody 15A8, an immunoglobulin G1 (IgG1) subclass anti-human breast carcinoma murine monoclonal antibody and gelonin, a plant toxin, were covalently modified with N-succimindyl 3-(2-pyridyldithio) proprionate and iminothiolane, respectively, and allowed to cross-link. 15A8-gelonin conjugates were purified from unreacted antibody and free gelonin by gel filtration and blue sepharose chromatography. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the final product contained two bands corresponding to antibody:gelonin conjugates of 1:1 (predominant) and 1:2. There were no contaminating amounts of free antibody or free toxin in the preparation. The yield of the final purified 15A8-gelonin conjugate was approximately 20% based on the amount of starting antibody. The protein synthesis inhibitory activity of the immunoconjugate was assessed by in vitro rabbit reticulocyte translation assay. This functional activity was normalized to that of unmodified gelonin for use in in vitro antiproliferative assays against antigen-negative (Hs294t human melanoma) and antigen-positive (ME-180 human cervical carcinoma) cell lines. Antigen-negative Hs294t cells incubated for 72 hours with 15A8-gelonin immunotoxin showed no increased cytotoxicity compared with HS294t cells exposed to free gelonin alone. However, the immunotoxin was preferentially toxic to antigen-positive ME-180 cells; over 5 logs greater cell kill was observed after 72 hours exposure to 15A8-gelonin than after the same exposure to gelonin alone. Various lysosomotropic agents augmented 15A8-gelonin cytotoxicity; the most effective potentiating agent appeared to be monensin. In addition, the chemotherapeutic agents L-phenylalanine mustard (L-PAM), 5-fluorouracil, vincristine, and bleomycin, and the biological response modifiers interferon-alpha and tumor necrosis factor-alpha were shown to augment 15A8-gelonin cytotoxicity. Should in vivo pharmacology and therapeutic studies confirm these in vitro findings, 15A8-gelonin conjugate may be a potent agent for therapy of cancer in man.
Antibodies, Neoplasm↗
Oestrogen production by the preimplantation donkey conceptus compared with that of the horse and the effect of between-species embryo transfer.
Aromatase distribution in membranes of preimplantation horse and donkey conceptuses was compared by measuring the incorporation of [3H]androstenedione into oestrone and oestradiol-17 beta. In the donkey conceptus, aromatase activity was similar in all the tissues examined (yolk sac, chorionic girdle and allantochorion), whereas in the horse it was generally lower and showed the relationship chorionic girdle greater than yolk sac greater than allantochorion. A higher proportion of labelled precursor was incorporated into oestradiol-17 beta by extra-embryonic tissues of the donkey compared with those of the horse. In contrast to previous results, aromatase in the chorionic girdle did not decline progressively before its migration into the endometrium on Day 36 to form the endometrial cups. The chorionic girdle of a donkey conceptus carried in the uterus of a mare failed to invade the surrogate horse endometrium and aromatase activity was still high in this tissue at Day 42. Aromatase distribution in 2 transferred donkey-in-horse conceptuses resembled that of the fetal, rather than the maternal, genotype indicating a lack of effect of the maternal environment.
Amnion↗
Effects of placental pathology on maternal plasma progestagen and mammary secretion calcium concentrations and on neonatal adrenocortical function in the horse.
Between 1985 and 1990, 25 Thoroughbred mares were classified as having an abnormal pregnancy on the basis of precocious signs of impending parturition. Throughout the remainder of their pregnancies, plasma progestagen and mammary secretion calcium concentrations were monitored. After parturition, evidence of placental pathology was recorded and their foals were assessed with regard to adrenocortical function and behaviour. Seven pony mares were used to establish a model of placental pathology and, following induced placental separation, were subjected to the same protocol. Control animals consisted of 35 Thoroughbred mares and 39 pony mares, all considered to have normal pregnancies. Of the 25 abnormal mares, 16 had precociously increased plasma progestagen concentrations, 17 had precociously increased mammary calcium concentrations and 24 had an abnormal placenta. Eighteen live foals resulted, of which 11 appeared abnormal on the basis of behaviour. Adrenocortical function was assessed in 13; 4 had normo- and 3 hypofunction and 6 were intermediate. These results were independent of gestational age at delivery. Of 13 foals examined for plasma progestagens, 7 had normal and 6 abnormal concentrations. Following placental separation in the pony mares, maternal plasma progestagen concentrations increased in 4 of the 7 mares within 4-11 days and in 1 after 42 days; there was placental pathology in 3 of these mares, 1 of which aborted at 287 days gestation. There was no increment in progestagens in the remaining 2 mares, which aborted at 240 days and 271 days. It was concluded that placental dysfunction may be diagnosed from early plasma progestagen increments prior to 308 days gestation; and that foals that survive birth in these circumstances may be physiologically immature but often have normal or intermediate adrenocortical function, irrespective of gestational age. This is in contrast to foals born prematurely without maternal plasma progestagen or mammary secretion calcium increments.
Adrenocorticotropic Hormone↗
Expression of major histocompatibility complex (MHC) class 1 molecules on early trophoblast.
The expression of class I Major Histocompatibility Complex (MHC) molecules by early trophoblast of intraspecific horse and donkey, interspecific mule and extraspecific donkey-in-horse conceptuses was determined using a rat monoclonal antibody (MAC 291) in a peroxidase anti-peroxidase immunohistochemical technique. Most non-invasive allantochorion of horse, donkey and mule conceptuses did not express class I MHC molecules at any stage of gestation except in small isolated patches of pseudostratified trophoblast lying adjacent to the openings of endometrial glands. In contrast, MHC class I molecules were expressed strongly on horse chorionic girdle cells at Days 33 and 34 of gestation, just prior to their invasion. However, class I MHC was down-regulated with the differentiation of these girdle cells into mature gonadotrophin-secreting endometrial cup cells between Days 40 and 45 so that by Days 55-65, class I molecules were no longer detected on endometrial cups. Similarly, all endometrial cups originating from 3 intraspecific donkey conceptuses at Days 41, 59 and 82 and 2 interspecific mule conceptuses at Days 46 and 47 were negative for class I molecules. A total of 7 extraspecific donkey-in-horse pregnancies, in which no endometrial cups from and implantation is abnormal, were established by embryo transfer. The chorionic girdle recovered from a single donkey-in-horse conceptus at Day 35 of gestation stained strongly for MHC class I molecules. Later in gestation (Days 73-91) and in contrast to the other forms of equine pregnancy examined, most unimplanted, failing donkey allantochorion was strongly stained for MHC class I molecules and had large numbers of lymphocytes in the adjacent endometrial stroma. The hypothesis is raised that the mechanisms that normally suppress the expression of MHC class I molecules by the epithelial trophoblast layer of the equine placenta can only function if the apical surface of the cells is in close and stable contact with other tissues such as the endometrial epithelium.
Animals↗
Prolactin secretion in lactating mares before and after treatment with bromocriptine.
The effect of treating lactating mares with the dopamine agonist bromocriptine was investigated. Seven pony and 4 Thoroughbred lactating mares were given a single intramuscular (i.m.) injection of 100 mg bromocriptine between Days 18 and 28 after foaling when the secretion rate of prolactin was elevated. Prolactin and luteinizing hormone (LH) concentrations were measured in serial peripheral plasma samples taken before and after the treatment and, in 5 of the pony mares, levels of these hormones were also measured in pituitary venous effluent obtained by cannulation of the cavernous sinus. In all the mares, prolactin concentrations fell sharply after treatment (Day 0) and they started to increase again on Day 2 in the Thoroughbreds and Day 3 in the ponies. Pituitary LH secretion rate also declined significantly for 2 days after treatment but there were no measureable influences on cyclic ovarian activity.
Animals↗
Sociodemographic and environmental correlates of racial socialization by black parents.
The present study focused on the content and practice of racial socialization by black parents. Sociodemographic correlates of patterns of racial socialization were examined using data from a national probability sample of 2,107 respondents (National Study of Black Americans). Multivariate analysis revealed that gender, age, marital status, region, and racial composition of neighborhood predicted whether or not black parents imparted racial socialization messages to their children. Black parents envision racial socialization as involving several components, including messages regarding their experience as minority group members, themes emphasizing individual character and goals, and information related to black cultural heritage. The findings highlighted the critical importance of sociodemographic and environmental influences on the socialization process. Implications for future research on racial socialization are discussed.
Adolescent↗