Bilateral sertoli and interstitial cell tumours in abdominal testes of a goat with polled intersex syndrome (PIS).

An 8-year-old, mixed breed, polled goat was presented for evaluation of male-like behaviour. Clinical findings included clitoromegaly, a heavily muscled neck, pronounced beard, and erect dorsal guard hairs, which are phenotypic characteristics commonly observed in intersex animals. Transrectal ultrasonography revealed the presence of two abdominal masses caudolateral to the uterine horns. Serum concentration of estradiol was elevated. Genetic evaluation was compatible with polled intersex syndrome defined by an XX karyotype without a Y chromosome or SRY gene. Based on gross and histologic evaluation, the abdominal masses were determined to be intra-abdominal testes, each of which was effaced by Sertoli cell and interstitial (Leydig) cell tumours. The Sertoli cell tumours (SCTs) represented two unique histologic patterns. Regardless of pattern, neoplastic Sertoli cells were consistently lipid laden and positive for vimentin. Interstitial cell tumours (ICTs) were negative for vimentin. Clinical and histopathologic findings suggest that prolonged exposure to steroids secreted by neoplastic Sertoli cells contributed to virilization. In addition, results from immunohistochemistry indicated that vimentin may be a valuable immunodiagnostic tool for differentiation between interstitial and Sertoli cell tumours in goats.

Diseases of the canine uterus.

Lesions most commonly found in the canine and feline uteri fall primarily into two categories: (i) abnormalities of endometrial growth and repair and (ii) uterine infections with associated endometritis. Neoplastic conditions of the tubular genitalia of the bitch, with the possible exception of leiomyomas (tumours that arise from smooth muscle cells), are uncommon. Congenital lesions involving the uterus are relatively rare. The primary object of this paper is to provide a summary of the most common uterine lesions found in the canine and feline uterus using a series of images that underscore the most prominent and important gross diagnostic features of each.

Diagnostic value of transcervical endometrial biopsies in domestic dogs compared with full-thickness uterine sections.

Transcervical endometrial biopsy is a useful tool for obtaining information about uterine health in some species. The clinical application of information gained from histopathological interpretations of endometrial biopsies in the bitch has not been validated. We hypothesized that transcervical endometrial biopsy samples would be as diagnostic as full-thickness uterine sections in identifying cystic endometrial hyperplasia (CEH), inflammation and periglandular fibrosis. Endometrial biopsies were obtained from 20 female adult dogs. Vaginal swabs, gross appearance of the vulva and vaginal tract, and serum progesterone values were used to determine the stage of the oestrous cycle at the time of sampling. The uteri were removed between 1 and 6 days after the biopsy procedure, and full-thickness sections were collected from each uterine horn and ovary and processed for histopathology. Two pathologists, blinded to the origin of each sample, compared full-thickness sections from the excised uteri to the biopsy samples collected via the transcervical technique. Pathologic features noted included: CEH, inflammation and periglandular fibrosis. Pathological diagnoses obtained from the biopsy sections were compared with those obtained from the full-thickness sections, as well as comparing diagnoses between the two pathologists, using McNemar's test. Of the 59 total biopsy samples obtained, 54 were considered diagnostic. All stages of the canine oestrous cycle were represented (anoestrus, proestrus, oestrus and dioestrus). Pyometra was not noted in any of the transcervical biopsy sections, but was noted in many of the full-thickness sections collected from dogs in dioestrus, suggesting either that biopsy is not a sensitive indicator of pyometra or that the procedure may induce pyometra in dioestrous dogs. Transcervical endometrial biopsy showed similar sensitivity as full-thickness sections in detecting CEH, inflammation and fibrosis. No differences in describing lesions were detected between pathologists.

A case of SRY-positive 38,XY true hermaphroditism (XY sex reversal) in a cat.

Investigation of abnormal sexual development in companion animals can allow for the elimination of inherited disorders from breeding populations while contributing to the understanding of the complex process of mammalian sexual development and differentiation. A 1-year-old mixed-breed cat, presented for neutering, was tentatively diagnosed as a male with bilateral cryptorchidism. During surgery, the surgeon identified gonads in an ovarian position and a complete bicornuate uterus. Both testicular and ovarian architecture in the gonads and Mullerian and Wolffian duct derivatives were identified histologically. The karyotype was that of a normal male (38,XY), and no causative mutation was identified in the feline SRY coding sequence amplified from genomic DNA. All features of the case were compatible with a diagnosis of SRY-positive 38,XY sex reversal, true hermaphrodite phenotype. To the authors' knowledge, this is the first report of this disorder in a domestic cat.

Canine and feline abortion diagnostics.

Knowledge of the causes of canine or feline pregnancy loss is limited and the success rate for making a definitive diagnosis is disappointingly low. Although these facts are discouraging, there are some things that can be done to improve success rates. This paper will address limitations and explore ways for improvement. For abortions caused by microbial infections, there are many reasons why it may not possible to identify the agents. "Non-infectious" causes are much more difficult to diagnose, and their relative importance is unknown. These include endocrine failure, underlying endometrial disease, genetic abnormalities, nutritional deficiencies, and toxicosis from drugs or environmental sources. Genetic abnormalities are a major cause of human pregnancy loss, yet we have little specific information about genetic diseases leading to abortion in animals. This paper addresses ways clinicians and diagnosticians can work together to improve diagnostic success. Necropsy techniques for fetal and placental examination and sampling are briefly reviewed. It is hoped that this series of papers will stimulate discussion on the causes and pathogenesis of pregnancy failure, and focus attention on areas where abortion diagnostics can be improved.

Cystic endometrial hyperplasia, pseudo-placentational endometrial hyperplasia, and other cystic conditions of the canine and feline uterus.

Cystic lesions in the uteri of bitches and queens arise from the uterine serosa, myometrium, or endometrium and include: serosal inclusion cysts, adenomyosis, endometrial polyps, cystic remnants of mesonephric ducts, and cysts associated with endometrial hyperplasia (both cystic glands and "pseudo-placentational" hyperplasia). Of these, "cystic endometrial hyperplasia (CEH)" is the most common and is frequently associated with pyometra. A second form of endometrial hyperplasia occurs in the bitch; although it was first described over 100 y ago, it is not widely recognized by clinicians or diagnostic pathologists. In this form, the endometrium proliferates in a highly organized manner, remodeling the uterine lining to closely resemble the histology of the endometrium at placentation sites in normal pregnancy. Although this lesion is very different from CEH, it is quite easy to induce in dogs during the luteal phase of their cycles and has been perhaps inappropriately proposed as modeling CEH. This lesion has been referred variously as "deciduoma", endometrial hyperplasia in pseudocyesis, and "maternal placental-like endometrial hyperplasia". An alternative name is suggested that is descriptive and draws attention to the difference between this lesion and CEH; the term pseudo-placentational endometrial hyperplasia (PEH) is proposed. The histopathology and pathogenesis of CEH and PEH are discussed. The objectives of this paper are to review the pathophysiology of cystic lesions of canine uterus, to demonstrate these using subgross photomicrographs taken from natural cases, and to present key diagnostic features of each.

Equine endometrial biopsy: enhancement of clinical value by more extensive histopathology and application of new diagnostic techniques?

During the 1960s and 1970s, the clinical value of equine endometrial histopathology was firmly established after it was shown that fertility outcome was correlated with the presence and severity of specific microscopic lesions. The objective of this paper is to summarize reports from the veterinary literature published after the mid 1980s that describe new diagnostic methods of assessing equine uterine health using material collected by endometrial biopsy.

Temporal and regional regulation of major histocompatibility complex class I expression at the bovine uterine/placental interface.

In most mammals trophoblast cells do not express major histocompatibility complex (MHC) antigens. This probably protects the placenta from immune attack. We have used immunohistochemistry to study the ontogeny of MHC class I expression by bovine trophoblast and endometrial epithelial cells. The interplacentomal, placentomal arcade and placentomal villous/crypt regions were studied. In the interplacentomal region a substantial proportion of trophoblast cells were class I positive from the sixth month on and about half of the endometrial epithelium was class I positive throughout pregnancy. In the arcade region trophoblast class I expression was first observed in the sixth month, increased slowly and peaked at term. Here there was no endometrial epithelial class I expression until term and then only a small percentage of cells were positive. In contrast, in the placentomal villous/crypt region both trophoblast and endometrial epithelium were class I negative throughout gestation. This study shows that cattle have extensive trophoblast class I expression. Moreover class I expression on placentomal, cryptal endometrial epithelium is shut down. Because binucleate trophoblast cells migrate and fuse with endometrial epithelial cells, total shut down of class I expression in areas of intimate interdigitation may be critical for avoidance of immunological rejection.

Maternal dexamethasone increases endothelin-1 sensitivity and endothelin a receptor expression in ovine foetal placental arteries.

Despite National Institutes of Health recommendations to administer antenatal steroids as a single course to women threatening preterm delivery, repeated treatments are often given. We investigated effects of repeated dexamethasone (DM) administered to the ewe on small maternal and foetal placental arteries. We hypothesized that DM would increase responsiveness to endothelin-1 (ET-1) and norepinephrine (NE) and that foetal arteries would react differently to ET-1 and NE compared to maternal arteries. Ewes received three treatments beginning at 103, 110, and 117 days of gestation (dGA). Each treatment consisted of four IM injections of 2mg DM or saline at 12-h intervals. At 119 dGA, in vitro functional studies were performed using Mulvany wire myography and endothelin receptor (ETR) expression was quantified using real-time RTPCR and receptor ligand autoradiography. Foetal placental arteries demonstrated greater maximal contractility to ET-1 and lesser maximal contractility to NE compared to maternal arteries. DM increased the maximal contraction elicited by ET-1 and NE in foetal but not maternal placental arteries. DM also increased the abundance of type-A ETR but not type-B ETR mRNA in foetal but not maternal placental arteries. However, within the whole placentome, DM increased the abundance of type-B ETR and decreased type-A ETR mRNA, which was confirmed by similar changes in ETR binding specifically within the labyrinth region. In summary, repeated DM treatment results in agonist and vascular bed specific responses within the placenta.

Ethylene glycol monomethyl ether effects on health and reproduction in male rabbits.

Male Dutch rabbits were weighed and randomly assigned within each weight group to five groups of six animals each (plus one more in the highest dose group). They received 0, 12.5, 25.0, 37.5, or 50.0 mg of ethylene glycol monomethyl ether (EGME) per kg of body weight in the drinking water 5 d/week for 12 weeks. Feed and water consumption were monitored daily and body weight weekly. All animals consumed the water and feed, maintained body weight, and were in good health throughout the experiment. Semen was collected twice weekly for 12 weeks, and 96% of the ejaculates were obtained. By weeks 6 and 9, most males in groups receiving 50.0 or 37.5 mg of EGME per kg were oligospermic. Only minor changes in other characteristics of sperm obtained from treated animals were found, as measured by computer-assisted sperm analysis. Fertility of the males still producing sufficient sperm during week 12 to use for insemination was tested with 96 does producing 2839 oocytes, and fertility of treated males (41%) was not lower (P > 0.05) than 47% in controls. At necropsy, all vital organs were grossly normal, with no notable histopathology. However, the groups of animals receiving 37.5 and 50 mg of EGME per kg of body weight produced fewer sperm and had smaller testes than controls (P < 0.05). Although all rabbits appeared grossly normal, there was a marked disruption of spermatogenesis as ingestion of EGME increased above 25 mg/kg of body weight. Rabbit testes appear to be more sensitive to EGME than testes of rats or mice.

Ontogeny of inflammatory cell responsiveness: superoxide anion generation by phorbol ester-stimulated fetal, neonatal, and adult bovine neutrophils.

Newborn calves, like human infants, are uniquely susceptible to bacterial infections. Part of this increased susceptibility may be related to defects in newborn polymorphonuclear leukocyte (PMN) defensive functions. It remains unclear whether reported deficits in newborn PMN function represent maturational disorders or are manifestations of some form of perinatal suppression phenomenon. We therefore compared the ability of bovine newborn PMNs (less than 24 h old), newborn PMNs (7-10 days of age), fetal PMNs (210-220 days gestational age), and adult PMNs to generate superoxide anion (O2-) as an indicator of respiratory burst activity. Citrated blood was collected, and PMNs were isolated to greater than 95% purity and 98% viability. O2- generation was measured as the superoxide dismutase-inhibitable (10 micrograms/ml) reduction of ferricytochrome c (2 mg/ml) after activation of PMNs with phorbol myristate acetate (PMA, 2 micrograms/ml) to directly stimulate protein kinase C. The reaction kinetics were measured (37 degrees C, 550 nm) using a spectrophotometer and chart recorder for continuous monitoring. O2- generation was measured for 5 min after the initial lag period and the total nanomoles of O2- generated calculated using the extinction coefficient for ferricytochrome c. Newborn PMNs (N = 10) generated significantly less O2- (5.7 +/- 0.8 nmol O2-/10(6) cells/5 min, P less than 0.01) than did adult PMNs (N = 14) (9.6 +/- 2.1 nmol O2-/10(6) cells/5 min) or fetal PMNs (N = 4) (10.7 +/- 0.7 nmol O2-/10(6) cells/5 min). PMNs from 7- to 10-day-old calves (N = 9) generated almost identical amounts of O2- as newborn PMNs (5.7 +/- 1.6 nmol O2-/10(6) cells/5 min). There was no difference in measured lag time period between newborn and adult PMNs, but fetal PMNs had significantly reduced (P less than 0.01) mean lag time. The data indicated that bovine newborn PMNs have a decreased ability to generate O2- in response to PMA stimulation, which persists for at least 7-10 days, and that this functional decrement may be a manifestation of some form of perinatal PMN suppression phenomenon rather than a developmental abnormality since fetal PMNs produced O2- as well as adult PMNs.

Minute virus of canines (MVC, canine parvovirus type-1): pathogenicity for pups and seroprevalence estimate.

Minute virus of canines (MVC, canine parvovirus type-1) caused inapparent to severe illness in neonatal specific-pathogen-free pups exposed by the oronasal route. The experimental disease was generally mild. Four of 21 infected pups had clinical signs of respiratory illness, but only 2 pups, not euthanized during the early postinoculation period, developed severe illness or died. Principal pathologic changes included bronchitis and interstitial pneumonia with various degrees of lymphadenitis. In contrast to the reported field cases, enteric signs were absent in the experimentally infected animals. Histopathologic changes in the small intestine were mild or absent. Bronchial, bronchiolar, and alveolar epithelial cells appeared to be the sites of initial and most extensive viral growth, reflecting the pattern of histopathologic changes. The disease caused by MVC was mild in comparison to that caused by canine parvovirus-type 2. MVC now appears to be established as a cause of illness in young pups and of transplacental infections with embryo resorption. The prevalence of MVC hemagglutination-inhibiting antibodies was high (approximately 50%) in adult dog sera from widely separated geographic areas of the United States.

A comparison of three avidin-biotin complex immunoenzyme systems for detection of African swine fever virus antigen in paraffin-embedded tissues.

The sensitivity and specificity of 3 avidin-biotin complex (ABC) immunostaining systems were compared on paraffin-embedded tissues from African swine fever virus (ASFV)-infected pigs. Results were also compared with immunofluorescent detection on cryosections of the same tissue for optimal detection of ASFV antigen. The ABC-alkaline phosphatase (ABC-AP) and ABC-peroxidase (ABC-PO) systems were at least as sensitive as direct fluorescent antibody (FA) and 10-fold more sensitive than the ABC-glucose oxidase system. Three ABC-AP and 2 ABC-PO chromagens with different counterstains were compared. In addition, 2 fixatives, 2 biotinylation procedures, 7 endogenous peroxidase blocking regimes, 6 tissue adhesives, and 3 mounting media were compared. The ABC-AP system with a red chromagen and hematoxylin counterstaining was preferred and most closely approximated routinely stained pathologic sections. Fixation in paraformalde-hydelysine-periodate fixative preserved ASFV antigen for research studies for at least 3 years. Formalin-fixed tissues retained some staining for up to 10 years.

Early infection of interdigitating dendritic cells in the pig lymph node with African swine fever viruses of high and low virulence: immunohistochemical and ultrastructural studies.

The role of interdigitating dendritic cells (IDCs) in the early pathogenesis of African swine fever (ASF) was investigated using mandibular lymphoid tissue from normal pigs and pigs inoculated oronasally with highly virulent Lisbon 60 (L-60) and moderately virulent Dominican Republic 1979 (DR-2) ASF virus (ASFV) isolates. Paraffin-embedded tissue sections were immunostained for ASFV antigen and S-100 protein, a marker of IDCs, using an avidin-biotin alkaline phosphatase procedure. Swine IDCs were identified morphologically by light microscopy, electron microscopy, and S-100 immunostaining. Infection with ASFV caused a marked reduction in S-100 staining by 3 days postinfection (DPI) that persisted through 14 DPI. Early ASFV infection of IDCs was demonstrated at 3 DPI by double immunohistochemical staining of cryosections and by transmission electron microscopy. These results support the hypothesis that the failure of a humoral immune response to virulent ASFV may be due to a primary infection of IDCs and the inability of IDCs to initiate an immune response. Infection of IDCs has also been demonstrated with human immunodeficiency virus (HIV-1), and these infections have some aspects in common.

African swine fever interference with foot-and-mouth disease infection and seroconversion in pigs.

Initial oral infection of pigs with either highly virulent (L-60) or moderately virulent (DR-2) African swine fever virus (ASFV), followed in 3 days with exposure to foot-and-mouth disease virus (FMDV) (tongue inoculation and contact), failed to cause FMDV infection or seroconversion in 18 of 22 L-60-infected pigs and 13 of 34 DR-2-infected pigs. Of the 13 DR-2-infected pigs remaining free of foot-and-mouth disease (FMD), 2 pigs survived to 24 days without antibody to FMDV, despite constant contact with clinically infected pigs with FMD. Three other DR-2-infected pigs never developed FMD lesions but did develop low levels of antibody to FMDV by day 17. A group of larger pig (in which DR-2 is less virulent) infected with DR-2 and then FMDV had a rapid but suppressed immune response to FMDV. Contact pigs introduced 3 days postinoculation and inoculated with FMDV only all became infected with ASFV by contact and died. This remarkably long lasting 1-way interference with FMD infection during acute and subacute African swine fever was not anticipated. Infection with ASFV may have blocked the initial target cells (possibly dendritic cells) necessary for establishment of FMDV infection.

African swine fever virus infection of skin-derived dendritic cells in vitro causes interference with subsequent foot-and-mouth disease virus infection.

Highly purified skin-derived dendritic cells (SDDCs) isolated from swine skin by a simple novel method were cultured for 24 hours before independent or sequential inoculation with African swine fever virus (ASFV) and foot-and-mouth disease virus (FMDV). By avidin-biotin immunohistochemical staining, ASFV antigen was detected in 50% of SDDCs as early as 1.5 hours postinfection (HPI) and in 80% by 3 HPI when cytopathic effect was noted. Cell lysis was detected with FMDV infection as early as 8 HPI; immunostaining for FMDV antigen was found in 10% of the cells. African swine fever virus replication was detected by transmission electron microscopy in a high percentage of SDDCs by 11 HPI. Sequential infection with FMDV 3 hours after ASFV inoculation blocked FMDV infection. These findings demonstrate that both ASFV and FMDV infect dendritic cells of Langerhans cell type in vitro and ASFV interferes with FMDV infection.

Trypanosoma theileri in direct tissue cultures prepared from naturally infected cattle.

Trypanosoma theileri was identified in 44 of 125 (35.2%) buffy coat cultures prepared from peripheral blood samples collected from 11 Holstein heifers. Nine of these animals had received steroid treatment, but trypanosomes were also recovered from two untreated controls. When direct tissue cultures were prepared from tissues collected from eight of these naturally infected heifers, T. theileri appeared in cultures from seven of them. This suggests that T. theileri can survive the procedures employed during direct tissue culture preparation.

The bovine placenta before and after birth: placental development and function in health and disease.

This paper reviews bovine placental development, anatomy (microscopic and gross), nomenclature and classification. The paper focuses on the biology of those specialized cells that arise from the outermost layer of very early embryos, the trophoblast cells, and on placental macrophages, cells that play a key role in fetal/placental defense. Data is presented from an immunohistochemical quantitative study that characterizes the ontogeny of placental macrophages using placental tissues from 21 cows (sampled from 4 months of pregnancy through the post partum period). Understanding of bovine placental development is essential for veterinarians, pathologists, diagnosticians and researchers. Lesions of diagnostic significance can be recognized for many economically important infectious abortifacient diseases, and there is growing evidence that pregnancy failure of cloned calves is due in part to unexplained placental failure. Placentology and placental pathology are becoming of increasing importance.

Major histocompatibility antigen expression on the bovine placenta: its relationship to abnormal pregnancies and retained placenta.

In viviparous animals, regulation of expression of major histocompatibility complex (MHC) class I antigens by the trophoblast cells, which constitute the outermost layer of the placenta, seems to be critical for maternal immunological acceptance of an allogeneic fetus. Cattle are unusual in this regard, since the bovine trophoblast cells, in specific regions of the uterine/placental interface, normally express MHC class I antigens during the third trimester of gestation. This expression appears to be biologically relevant as MHC class I compatibility between a cow and her fetus has been associated with an increased incidence of placental retention. We have found significant differences in lymphocyte populations, cytokine production, and trophoblast cell apoptosis in the placentomes of MHC-compatible and -incompatible pregnancies at parturition. This suggests that maternal immunological recognition of fetal MHC class I proteins triggers an immune/inflammatory response that contributes to placental separation at parturition in cattle. Early in pregnancy, a complete shutdown of MHC class I expression by trophoblast cells appears to be critical for normal placental development and fetal survival. In bovine somatic cell nuclear transfer (SCNT) pregnancies, there is an extremely high rate of fetal loss between days 30 and 90 of pregnancy. We have shown that in bovine SCNT pregnancies, between days 34 and 63 of gestation, there is both abnormal expression of MHC class I antigens by trophoblast cells and an abnormal accumulation of lymphocytes within the uterine stroma. Consequently, it is likely that activation of the maternal mucosal immune system, within the uterus at the same time when placentomes are being established, interferes with the process of placentome development and leads to immune-mediated abortion. Our data suggest that bovine MHC-compatible pregnancies provide a unique model for studying regulation of the uterine immune system, as well as immune-mediated placental rejection.

Endotoxemia in pregnant cows: Comparisons of maternal and fetal effects utilizing the chronically catheterized fetus.

We utilized the chronically catheterized bovine fetus to compare maternal and fetal responses to maternal lipopolysaccharide (LPS) infusion. Our hypothesis was that LPS-induced abortion was primarily a maternal luteolytic event with minimal transplacental fetal exposure. Fetal tibial arteries, amniotic, allantoic cavities and maternal carotid arteries were catheterized. Three cows had patent catheters with viable fetuses (190 to 200 days of gestation) 1 week after operation and were included in the study. Following a 2-day maternal and fetal baseline, 0.5 mug Salmonella typhimurium LPS/kg was infused into a maternal jugular vein over a 2-hour period. Maternal and fetal responses were monitored clinically, biochemically and hormonally. The maternal response consisted of marked increases in plasma prostaglandin F(2alpha) metabolite (PGFM), tumor necrosis factor (TNF), ACTH and cortisol with a dramatic maternal leucopenia within 2 hours. Progesterone concentrations decreased within 7 hours (P<0.05). The LPS was rapidly cleared from maternal circulation and no transplacental exposure was detected in the fetuses. Fetal responses to maternal endotoxemia consisted of increased ACTH and cortisol concentrations with delayed increases in PGE(2); TNF did not change in fetal fluids following maternal endotoxemia. There was a fetal leucocytosis within 2 hours. The results indicate that the fetus does not appear to play a major role in the pathogenesis of LPS-induced abortions. However, the role of maternal TNF in endotoxin abortion requires further study.