Ultrastructure of the chorioallantoic placenta and chorionic vesicles of the lesser bush baby (Galago senegalensis).

The ultrastructure of the chorioallantoic placenta of the lesser bush baby (Galago senegalensis) has been studied. The placenta was shown to be of the diffuse, epitheliochorial and adeciduate type. The trophoblasts of the chorionic villi, other than those lining the chorionic vesicles, were characterized by the presence of many lipid droplets. In the later stage of gestation, the fetal capillaries indented the trophoblastic epithelium reducing the distance between fetal and maternal circulations. In addition chorionic vesicles were observed. The trophoblasts lining the chorionic vesicles have outward bulging apical surfaces. There are clefts between these cells and this region is occupied by microvilli of adjacent cells. Several layers of fusiform cells that did not extend up into the cores of the chorionic vesicle villi formed the outer component of the vesicular wall. Granulated cells were observed within the maternal connective tissue and their possible role is discussed.

The origin and development of the upper lateral incisor and premaxilla in normal and cleft lip/palate monkeys induced with cyclophosphamide.

OBJECTIVE: Cleft lip/palate (CLP) is a common human congenital defect in which the maxillary lateral incisors are often absent, malformed, and malpositioned. The present study was designed to examine the origin of the upper primary lateral incisor relative to the medial nasal process (MNP) and maxillary process (MP) fusion area and to the premaxillary/maxillary (incisive) suture in monkeys. METHOD: Scanning electron microscopy, histology, skeletal staining, and drying techniques were used to study facial development in embryo and fetal monkey specimens. A teratogenic dose of cyclophosphamide was administered to pregnant monkeys prior to fusion of the MNP and MP and fetuses were examined for CLP. RESULTS: Formation of the anterior maxilla involved fusion of the MNP and MP at stages 14-18. At stages 18-20, the palatal portion of the MNP had formed the medial and lateral incisive mounds. By stage 22, the upper primary lateral incisor has formed within the MP, lateral to the MNP/MP fusion area and to the ossifying premaxilla. Ossification of the premaxilla begins in the MNP and subsequently spreads laterally across the MNP/MP fusion area into the MP. Accordingly, the lateral incisor undergoes a complex positional shift (mainly medial) relative to the incisive suture both prenatally and postnatally and is finally located medial to the suture. Examination of the cyclophosphamide-induced CLP fetuses showed that the lateral incisor is located lateral to the alveolar cleft and does not shift medial to the incisive suture. CONCLUSION: Understanding the origin of the lateral incisor (the tooth closest to the cleft) and the shift after its formation provides clues to high incidence of malformations and ectopia of this incisor in cleft patients.

Morphological characterization of the seminiferous cycle in the goat (Capra hircus): a histological and ultrastructural study.

The cycle of spermatogenesis/seminiferous cycle was investigated in the goat testis using both light and electron microscopy techniques. Using the various cell associations and the accompanying changes in spermatid shape and location, the cycle was divided into eight (8) successive stages. The cycle began with the accomplishment of spermiation (stage 1) and ended with apical migration and close attachment of late maturation phase spermatids at the Sertoli cell apex accompanied by adluminal retention of residual bodies with dense staining inclusions (stage 8). The early stages of the cycle (stages 1-4) were therefore characterized by the presence of only one generation of spermatids, the second one appearing only after the division of secondary spermatocytes in stage 4. Consequently, stages 5-8 had two generations of spermatids; Golgi or cap phase as well as maturation phase spermatids. Although stages 5 to 7 appeared as distinct entities, stages 6 and 7 were rather short-lived and considered as continuations of stage 5. Therefore, the 8 stages of the cycle in the goat were further condensed into 6 main divisions. The duration of each stage was estimated by the frequency of occurrence in sections. Among these, stage 1 had the highest frequency (34%) followed by stages 5-7 (27%). Stages 8 and 4 had the shortest frequency (up to 9%) while stages 2 and 3 had 13% and 12% respectively. These results indicate that, like most domestic species, goats have a cycle of 8 stages with 6 main divisions, the longest being stage 1.

Characterization of antigens expressed in normal baboon trophoblast and cross-reactive with HIV/SIV antibodies.

Electron microscopic studies have revealed the presence of endogenous retroviral (ERV) particles in normal primate placental tissues. These particles have ultrastructural similarities to type C retroviral particles and are mainly associated with the trophoblast. In normal human placental tissues, they have antigenic similarity with exogenous retroviruses, such as the human immunodeficiency virus (HIV), and may have a role to play in the regulation of cellular gene expression, syncytiotrophoblast formation or pregnancy-related immunosuppression. In this study, a panel of antibodies (polyclonal and monoclonal antibodies) against viral proteins (anti-HIV and anti-SIV) and endogenous retroviral (ERV) proteins were assessed by immunohistochemistry and immunoblotting, for their cross-reactivity with ERV particles isolated from normal baboon placental tissues. The antibodies (anti-HERV-K RT, anti-ERV3 env, anti-HIV-1 p17, anti-HIV-2 gp120) reacted positively with the syncytiotrophoblast and each antibody recognized one or two proteins of molecular weights (MW) 38, 58 or 64 kDa present in the baboon placental villous tissues and SIV-infected molt-4 Cl8 cells, but not in uninfected cells. The results of this study confirm the specific expression of retroviral cross-reactive antigens in normal baboon placental tissues and suggest placental cellular proteins may have antigenic similarity with those recognized by anti-HIV/SIV antibodies. The role of these retroviral-related proteins expressed at the maternal-fetal interface remain unclear.

Immunohistochemical localization of retroviral-related antigens expressed in normal baboon placental villous tissue.

Endogenous retroviral particles (ERVs) have been detected in the genome of all eukaryotes. They are generally non-pathogenic except in mice where they have been found to induce tumors and immunological disorders. The ERVs have morphological features consistent with type-C retroviral particles and are commonly expressed in normal placental villous tissues. ERVs may have a role in the regulation of placental gene expression, syncytiotrophoblast formation, or pregnancy-related immunosuppression. In this study, well-characterized antibodies (monoclonal and polyclonal antibodies) raised against retroviral proteins (anti-HIV and anti-SIV) and endogenous retroviral (ERV) particles were assessed for their cross-reactivity (by using immunohistochemistry) with normal baboon placental and other adult tissues. The monoclonal antibodies to exogenous retroviral proteins (anti-HIV-2 gp120, anti-HIV-1 gp41, anti-SIVmac p27, anti-HIV-1 RT, and anti-HIV-2 core protein) showed specific immunohistochemical reactivity with the syncytiotrophoblast. Antibodies to endogenous retroviral gene products (anti-ERV3 env, anti-HERV-K RT, and anti-HERV-K env) also reacted in a similar manner and did not cross-react with other adult tissues. These studies have shown that retroviral-cross-reactive proteins are expressed in baboon placental syncytiotrophoblast and may have a role to play at the feto-maternal interface.

Characterisation of retroviral-related antigens expressed in normal baboon placental tissues.

The genomes of all eukaryotes contain multiple copies of DNA sequences that are related to sequences found in infectious retroviruses. Endogenous retroviruses (ERVs) are generally non-pathogenic although they have been implicated in the induction of tumours and immunological disorders. The ERVs have morphological features consistent with type-C retroviral particles and are expressed in normal placental tissue in most mammals. They have antigenic similarity with exogenous retroviruses such as HIV-1 and may have a role to play in the regulation of cellular gene expression, syncytiotrophoblast formation or pregnancy-related immunosuppression. Some of the human endogenous retroviruses have been well-characterised. Among the non-human primates, the baboon endogenous virus (BaEV) is the only endogenous retrovirus so far which has been shown to be effective in vitro. The entire nucleotide sequence of BaEV has been determined. It has been shown to have a chimeric genomic structure of about 8 kb long. BaEV particle expression in placental tissues has been demonstrated using electron microscopy. However, to date, very little work has been done to evaluate the expression of retroviral-related antigens in normal baboon tissues. In this study, mouse polyclonal antibodies were produced against isolated baboon placental ERV particles and characterised using immunohistochemistry and immunoblotting techniques. Most of the anti-BERV antibodies displayed specific immunoperoxidase staining on placental syncytiotrophoblast and cross-reacted with exogenous retroviral proteins on immunoblot analyses. Reverse transcriptase (RTase) activity associated with sucrose gradient-purified placental retroviral-like particles were also demonstrated. These studies indicate that endogenous retroviral particles are expressed in baboon placental villous tissue and suggest retroviral proteins may play an immunomodulatory role at the maternal-foetal interface.

Morphology of a six-legged goat with duplication of the intestinal, lower urinary, and genital tracts.

BACKGROUND: An adult female goat with rare malformations, which consisted of duplication of the intestinal, lower urinary, and genital tracts as well a pair of parasitic appendages, is presented. METHODS: A complete dissection was performed on a moribund female goat (Capra hircus). RESULTS: The animal had a normal body with a parasitic attachment located within the pelvic region. This attachment was represented by an ovoid, trunk-like, adipose mass that lacked internal organs or vertebrae but that had two fairly well-developed limbs with the normal components of hind limbs. There was duplication involving the external and internal genitalia, the urethra, the urinary bladder, and portions of the small intestine as well as the large bowel, including the anal openings. CONCLUSIONS: An autosite with a duplication involving the hindgut and paramesonephric anlages was identified. These features were compatible with life in utero and postutero and emanated from incomplete twinning (heteropagus twins). A review of the literature also suggests that heteropagus twins are a very rare abnormality in both domestic animals and humans.

Use of biometric embryonic growth parameters as indicator of exposure to a teratogen.

Disturbances in embryonic growth were studied in 233 foetuses harvested on day 14.5 of gestation, after the administration of various doses of 5 Fluoro-2' deoxyuridine (FUdR) to pregnant mice on day 11.0 of gestation. Measurements of crown-rump length (CRL) and mean wet body weights showed a significant retardation of embryonic growth (p < 0.001), following doses of 30, 80 and 100 mg FUdR per kg maternal body weight. Compared to the controls, whole FUdR-treated embryos that had been macerated, cleared and double stained with alcian blue 8GX plus alizarin red S for skeletal anlage, showed that ossification had not commenced in the vertebral bones of tail. All bones in the craniofacial region and limbs including the girdles, were smaller, while there were distortions of the long bones. The severity of the changes were dependent on the concentration of FUdR dose administered. Among the live FUdR-treated foetuses harvested, 95% had mesomelic limb defects. The incidence of delay or prevention of palatal processes elevation was 79%, 49%, 21% and 30% respectively for 0 mg (control), 30 mg, 80 mg and 100 mg FUdR doses. The results show that administration of a teratogenic agent (FUdR) causes retardation of growth which correlates with abnormalities of the secondary palate and limbs. It is proposed that the initial screening of potential teratogenic substances in food, such as preservatives or colourings, may be carried out by monitoring changes in secondary palate and limb development, including biometric growth parameters of an animal model.

Holoprosencephaly in a long-tailed macaque (Macaca fascicularis): a case report.

A long-tailed macaque fetus with ethmocephaly, alobar holoprosencephaly, and arhinia is reported. This fetus was exposed to an antiprogestational agent, RU 486 (2.5 mg/kg intramuscular, once daily), during gestational days 15-18. Three hypotheses are proposed to explain these malformations: (1) they are a direct result of drug exposure, (2) they are a secondary effect of treatment and the result of decreased blood supply to the developing embryo due to an incomplete abortion, and (3) they represent a spontaneous occurrence.

Ultrastructural study of the testis of non-breeding naked mole-rat (Heterocephalus glaber, Ruppell).

The testicular structure of the wild caught naked mole rat was studied. It comprises of a large volume of lipid-rich interstitial cells of Leydig among which are few scattered seminiferous tubules. In addition, the interstitial cells possess elongated mitochondria and vast network of smooth endoplasmic reticulum (sER). The Golgi apparatus (GA) apparently is not conspicuous or well developed. All stages of spermatogenesis occur in the seminiferous tubules although the mature forms (secondary spermatocytes, spermatids and spermatozoa) are few. Sertoli cells show an irregular nucleus, mitochondria oriented perpendicular to the basement membrane, a vast network of endoplasmic reticulum with sER as the predominant form and lipid droplets. The ultrastructural features of Leydig cells seem to suggest a steroidogenic capacity although the vast accumulation of lipid droplets may imply impaired utilisation of cholesterol reservoir as a result of pituitary hormonal imbalance or (and) the local paracrine influence by Sertoli cells. The cause of slow-down in spermatogenesis is still unclear but may also be under the influence of pheromonal cues or the local paracrine control. Sertoli cell features point towards a role of synthesis and secretion.

The relationship of serum estradiol and progesterone concentrations to the enzyme immunoassay measurements of urinary estrone conjugates and immunoreactive pregnanediol-3-glucuronide in Macaca mulatta.

Paired urine and serum samples from four conceptive and six nonconceptive ovarian cycles of seven adult Macaca mullatta were analyzed by radioimmunoassay (RIA) for circulating estradiol (E2) and progesterone (Po), and urinary estrone conjugates (E1C) and immunoreactive preganediol-3-glucuronide (iPDG) using enzyme immunoassay (EIA). Nonconceptive cycles exhibited a fivefold increase in urinary E1C and serum E2 levels from follicular phase levels to the preovulatory peak. Linear correlation between urinary E1C and serum E2 nonconceptive cycle hormone levels was significant (P <0.01, r = 0.69). Luteal phase levels of iPDG and serum Po levels were approximately parallel in nonconceptive cycles. Similarly, conceptive cycle urinary E1C levels and serum E2 measurements had a correlation coefficient that was significant (P<0.01, r = 0.45). Nonconceptive and conceptive cycle iPDG and Po levels were significantly correlated (P = 0.05, r = 0.63, and P<0.01, r = 0.66, respectively). These data demonstrate that EIA measurements of ovarian hormones in daily urine samples can be used to accurately monitor ovarian function and early pregnancy in Macaca mulatta.

The effect of the anti-progestin RU 486 on early pregnancy in the long-tailed macaque (Macaca fascicularis).

The efficacy of various doses of RU 486 in terminating pregnancy before and after the luteal-placental shift (LPS) in the long-tailed macaque (Macaca fascicularis) was assessed through sonographic examination and measurements of steroid hormones and their metabolites. Intramuscular injection of 1.0, 2.5, 12.5, or 25.0 mg/kg was administered either from gestational day (GD) 15-18 (Group 1; N = 11) or GD 23-26 (Group 2; N = 9). The timing of treatment was determined by the detection of the preovulatory estrogen peak via daily urinary estrone conjugate (E1C) measurements. In Group 1, a 90.9% pregnancy loss was observed (10/11); seven animals aborted during GD 15-20, two animals indicated early embryonic death with retained gestational sacs, one animal aborted on GD 56, and one pregnancy was maintained. In Group 2, an 88.9% pregnancy loss was observed (8/9); eight animals aborted between GD 26-29, and one pregnancy was unaffected. Hormone profiles appeared to fall secondarily to the loss of trophoblast function. These results indicate: (a) RU 486 was more effective after the LPS; and (b) the primary effect of RU 486 appeared to be at the level of the products of conception.

Fetal membranes and placenta of the African green monkey (Cercopithecus aethiops).

This study examined developmental changes in fetal membranes and placenta of Cercopithecus aethiops from a Carnegie developmental stage 12 embryo to near-term fetuses. Ultrastructurally, yolk sac cells (endoderm and mesothelium) were similar to comparable stages in other primates. Endodermal cells had few apical microvilli, abundant rough-endoplasmic reticulum, electron dense mitochondria and dense bodies. In contrast, mesothelial cells were squamous with numerous microvilli, small mitochondria and a few short strands of rough endoplasmic reticulum. Amnion cells early in gestation were squamous with few microvilli, large glycogen deposits and poorly developed cytoplasmic components. Tight junctions and desmosomes held adjacent cells together. The basal surface was smooth and the basal lamina was distinct. As development proceeded the amniotic cells became cuboidal and possessed numerous microvilli. Cytoplasmic organelles were better developed and glycogen deposits increased by mid-gestation. A thick layer of microfibrils and collagen fibers was prominent below the basal lamina. Near-term, the glycogen had virtually disappeared and the amount of lipid droplets increased. Basal infoldings and podocytic processes and the extracellular matrix had increased. The smooth chorion consisted of pseudostratified columnar cells. Cells had short microvilli, numerous granules and vesicles of variable size and electron density in early gestation. With increasing age, amounts of granules and vesicles decreased, as the endoplasmic reticulum became prominent. The chorionic trophoblast was a continuous layer in mid-pregnancy and its cells had well-developed organelles and inclusions. Late in gestation, the trophoblastic layer became discontinuous and wide intercellular spaces and channels were present. In the placenta, the trophoblastic elements showed features characteristic of primate placenta.

Early placentation in the African green monkey (Cercopithecus aethiops).

Placentation in the green monkey (Cercopithecus aethiops) from gestational ages of 18-30 days was studied by light and electron microscopy. The placentae show many features in common with that of the rhesus monkey and other related macaques. Implantation is superficial and bidiscoid, and the placenta is classified as villous and hemomonochorial. Epithelial plaque formation and a moderate stromal reaction occur. Plaque cells are large, with massive deposits of glycogen and marginal pleomorphic dense bodies. Observations further support the concept that chorionic blood vessels, angioblastic and mesenchymal cells are not derived by delamination from cytotrophoblast. Capillary formation is by attachment of one or more mesenchymal cells and/or their processes. Embryonic blood was not observed in the villi until after day 22.