A morphological analysis of the transition between the embryonic primitive intestine and yolk sac in bovine embryos and fetuses.

The yolk sac (YS) is the main source of embryonic nutrition during the period when the placenta has not yet formed. It is also responsible for hematopoiesis because the blood cells develop from it as part of the primitive embryonic circulation. The objective of this study was to characterize the transitional area between the YS and primitive gut using the techniques of light microscopy, transmission electron microscopy, and immunohistochemistry to detect populations of pluripotent cells by labeling with Oct4 antibody. In all investigated embryos, serial sections were made to permit the identification of this small, restricted area. We identified the YS connection with the primitive intestine and found that it is composed of many blood islands, which correspond to the vessels covered by vitelline and mesenchymal cells. We identified large numbers of hemangioblasts inside the vessels. The mesenchymal layer was thin and composed of elongated cells, and the vitelline endodermal membrane was composed of large, mono- or binucleated cells. The epithelium of the primitive intestine comprised stratified columnar cells and undifferentiated mesenchymal cells. The transitional area between the YS and the primitive intestine was very thin and composed of cells with irregular shapes, which formed a delicate lumen containing hemangioblasts. In the mesenchyme of the transitional area, there were a considerable number of small vessels containing hemangioblasts. Using Oct4 as a primary antibody, we identified positive cells in the metanephros, primordial gonad, and hepatic parenchyma as well as in YS cells, suggesting that these regions contain populations of pluripotent cells.

Genital morphology of the male South American fur seal (Arctocephalus australis) and biological implications / Morfologia dos órgãos genitais do macho do Lobo marinho (Arctocephalus australis) e implicações biológicas

Male capacity for spreading genes to a great number of descendents and to determine population dynamics depend directly on the genital organs. Morphological studies in pinnipeds are scarce and the functional meaning of some characteristics has never been discussed. We hypothesized that Arctocephalus australis (A. australis) shows morphophysiological adaptations in order to guarantee the perpetuation of the species in the unique annual mating season. Seven males, dead from natural causes, had their genital organs collected and fixed for morphological description. Some features differ from other described mammalian males and are closely related to the biology and reproductive cycle of this species, as the scrotal epidermis, absence of glandular portion in the ductus deferens and spermatogenic epithelium suggest a recrudescent testis period. The corona glandis exhibits a singular arrangement: its erectile border looks like a formation of petals and its association with the os penis gives a "lily-flower" form to this region. We propose the name margo petaliformis to this particular erectile border of the corona glandis because of its similarity to a flower corola. The male genital organs of A. australis show morphological features compatible with adaptation to environment requirements and reproductive efficiency.(AU)

A capacidade do macho de espalhar seus genes a um grande número de descendentes e determinar a dinâmica populacional depende diretamente dos seus órgãos genitais. Estudos morfológicos em pinípedes são escassos e o significado funcional de algumas de suas características ecológicas ainda foi pouco discutido. Nossa hipótese é que Arctocephalus australis (A. australis) apresenta adaptações morfofisiológicas em seus órgãos genitais capazes de interagir com o meio e garantir a perpetuação da espécie que apresenta apenas uma época de acasalamento que ocorre uma vez a cada ano. Sete A. australis machos, mortos recentes por causas naturais, tiveram seus órgãos genitais coletados e fixados para a descrição macro, micro e ultraestrutural. Algumas características diferem de outros machos já descritos e estão intimamente relacionados com a biologia e ciclo reprodutivo da espécie, dentre elas podemos citar a alta queratinização da epiderme escrotal que pode se relacionar com as rotineiras lesões por atrito desta região nas pedras; a ausência da porção glandular do ducto deferente aqui descrita pela primeira vez, o epitélio espermatogênico sugere um período de testículo recrudescente. A glande apresenta um arranjo singular: a coroa da glande apresenta porção lateral de tecido esponjoso que são bordas livres com capacidade de intumescencia. O osso peniano se encontra no centro destas bordas e representa a extremidade mais distal do penis, levando consigo o óstio uretral externo. As bordas associadas ao osso peniano, dão uma forma de "Flor de lírio" a esta região. Utilizamos o nome margo petaliformis a margem erétil liliforme a particular morfologia da glande, pela sua semelhança a uma corola de flor. Os órgãos genitais masculinos de A. australis mostram características morfológicas compatíveis com uma adaptação aos requisitos ambientais e de eficiência reprodutiva.(AU)

Goat scrotal-testicular biometry: Influence of the season on scrotal bipartition / Biometria escroto-testicular em caprinos: influência de período do ano na bipartição escrotal

The scrotal-testicular biometry was evaluated in goats raised in Piaui state, Brazil, presenting different levels of scrotal division, in rainy and dry periods of the year. For this study, eighteen male goats at mating age were accomplished and arranged into three groups (6 animals each), obeying the classification as goats with no scrotal bipartition (GI), goats showing scrotal bipartition up to 50 percent of testicular length (GII), and goats with more than 50 percent of scrotal bipartition (GIII). The biometry of the scrotal-testicular was made evaluating the scrotal length (SL), scrotal circumference (SC), testicular length (TL) and testicular volume (TV). The results were evaluated following the variance analysis (ANOVA) and the SNK test applied on the average comparisons. The analysis of the data demonstrated high values, in dry and rainy periods, of SC (24.63cm/ 26.97cm), SL (16.61cm/ 18.24cm), TL (5.32cm/ 5.93cm), TV (173.81cm³/ 203.01cm³). This supports the hypothesis of the influence of the period of the year and of the scrotal bipartition on the scrotal-testicular biometry in goat.

Objetivou-se neste trabalho avaliar a biometria escroto-testicular em caprinos com escroto simples e bipartido, criados no Estado do Piauí, Brasil, nos períodos seco e chuvoso do ano. Foram utilizados 18 caprinos machos, divididos em três grupos de seis caprinos. O grupo I (escroto sem bipartição), o grupo II (escroto bipartido até 50 por cento de comprimento testicular) e o grupo III (bipartição escrotal superior a 50 por cento do comprimento testicular). A biometria escroto-testicular consistiu do comprimento escrotal (COE), circunferência escrotal (CE), comprimento testicular (CT) e volume testicular (V). Os resultados foram submetidos à análise de variância (ANOVA), seguido do teste SNK para comparação das médias. Os dados mostraram que os animais do grupo GIII apresentaram, no período seco, os valores de CE, COE, CT e V de 24,63cm, 16,61cm, 5,32cm, e 173,81cm³, respectivamente e de 26,97cm para CE, 18,24cm para COE, 5,93cm para CT e 203,01cm³ para V, no período chuvoso. Todos esses valores foram superiores (p<0,05) aos encontrados para os animais dos demais grupos. Conclui-se que o grau de bipartição escrotal e o período do ano interferem na biometria escroto-testicular de caprinos.

Ultrastructural characterization of bovine umbilical cord blood cells / Caracterização ultra-estrutural das células sanguíneas do cordão umbilical bovino

The umbilical cord blood (UCB) is an important source of pluripotent stem cells, which motivated researches on ontogeny and transplantation. The morphological characterization of umbilical cord cells is the first step to establish subsequent experiments on these areas. Although some information on humans can be found, no data on UCB is available for bovines. Therefore, this work is the first attempt to conduct an ultrastructural characterization of bovine umbilical cord blood. Blood was collected from the umbilical cord of twenty fetuses by punction of the umbilical vein. Samples were processed for whole leucocytes observation by centrifugation and the buffy coat was collected. Cells were washed and pelleted and prepared according to the standard protocol of the transmission electron microscopy. The presence of cells with morphologic characteristics compatible with the precursors from the erythrocytic, neutrophilic, eosinophilic, basophilic, and lymphocytic lineages was observed. Atypical cells with peculiar morphological features, strongly similar to apoptotic cells, were seen. Bovine neutrophils with three types of cytoplasmic granules were also found in the blood. The ultrastructural characteristics of observed bovine UCB cells where similar to those found in other species, suggesting that bovines could possibly constitute an experimental model for approaches on UCB cells research.

O sangue de cordão umbilical (SCU) é uma importante fonte de células progenitoras pluripotentes, que motiva pesquisas em ontogenia e transplantes. A caracterização morfológica das células de cordão umbilical é o primeiro passo para se estabelecer experimentos subsequentes nessas áreas. Embora algumas informações sobre SCU em humanos possam ser encontradas, não existe nenhuma informação disponível sobre elas em bovinos. Portanto, este trabalho é a primeira tentativa de se conduzir uma caracterização ultra-estrutural do sangue de cordão umbilical bovino. O sangue foi coletado do cordão umbilical de 20 fetos por punção da veia umbilical. As amostras foram processadas para observação dos leucócitos totais por centrifugação pela coleta do botão leucocitário. As células foram lavadas, peletizadas e preparadas de acordo com protocolo padrão para microscopia eletrônica de transmissão. A presença de células com características morfológicas compatíveis com precursores das linhagens eritrocítica, neutrofílica, eosinofílica, basofílica e linfocítica foram observadas. Células atípicas com características morfológicas peculiares muito semelhantes a células em apoptose foram observadas. No sangue do cordão umbilical também foi encontrado neutrófilos bovinos apresentando três tipos de grânulos citoplasmáticos. As características ultraestruturais do SCU bovino foram semelhantes às encontradas em outras espécies, sugerindo que esta espécie possa servir como modelo experimental para abordagens em pesquisa sobre sangue de cordão umbilical.

Influence of the bipartite scrotum on the testicular and scrotal temperatures in goats / Influência do escroto bipartido sobre as temperaturas dos testículos e escroto em caprinos

The influence of the scrotal bipartition and of the year period on the scrotal-testicular thermal regulation was evaluated in male goats raised in Piaui State, Brazil. Eighteen male goats at mating age were accomplished in this study and arranged into three Groups (6 animals each) obeying the classification as goats presenting no scrotal bipartition (Group I), goats showing scrotal bipartition at 50 percent of the testicular length (Group II), and goats with more than 50 percent of scrotal bipartition (Group III). The scrotal, testicular and spermatic funiculi temperatures were evaluated invasively with the aid of a digital thermometer and non-invasive with a pyrometer in the proximal, medial and distal portion. The data were acquired during the dry (October-November) and rainy (February-March) period of the year, measured in two shifts: morning (6h00-7h00) and afternoon (14h00-15h00). The results were submitted to variance analysis (ANOVA) following the SNK test for average comparison (p<0.05). The year period interfered on the scrotal-testicular thermal regulation, due to increased temperatures of the scrotal, testicular and spermatic funiculi during the dry period in comparison with the rainy period. The bipartition level was also a factor which contributed to the influence of scrotal-testicular temperature regulation, due to lower average scrotal-testicular temperature rates observed during both periods in the goats with higher levels of scrotal bipartition (>50 percent). It is possible to conclude that with the experimental conditions applied on this study, the level of scrotal bipartition and the climatic conditions interfere with the scrotal-testicular thermal regulation in goats.

O objetivo deste trabalho foi avaliar a influência do grau de bipartição escrotal e do período do ano sobre a termorregulação escroto-testicular em caprinos criados no Estado do Piauí. Foram utilizados 18 reprodutores caprinos machos, divididos em três grupos de seis animais: O Grupo I contendo caprinos com escroto simples, o Grupo II, caprinos com escroto bipartido até 50 por cento do comprimento testicular e o Grupo III, caprinos com bipartição superior a 50 por cento do comprimento testicular. Os parâmetros avaliados foram as temperaturas do escroto, testículo e funículo espermático, obtidas de forma invasiva, com um termômetro digital termoacoplável, e não invasiva, com um pirômetro, nos terços proximal, médio e distal. Os dados foram coletados nos períodos seco (outubro-novembro) e chuvoso (fevereiro-março) do ano, bem como, nos turnos da manhã (6h00 às 7h00) e tarde (14h00 às 15h00). Os resultados foram submetidos à análise de variância (ANOVA) seguida do teste SNK para comparação das médias (p<0,05). O período do ano interferiu na termorregulação escroto-testicular, pois no período seco as temperaturas do escroto, testículo e funículo espermático foram mais elevadas que as observadas no período chuvoso. O grau de bipartição do escroto foi outro fator que modificou a temperatura escroto-testicular, já que os caprinos que apresentaram escroto com maior grau de bipartição demonstraram as menores médias das temperaturas escroto-testiculares em ambos os períodos e turnos avaliados. Conclui-se, portanto, que tanto o período do ano quanto o grau de bipartição do escroto influenciaram o processo de termorregulação escroto-testicular em caprinos.

Evaluation of differences between breeds for substance P, vasoactive intestinal polypeptide, and neurofilament 200 in the abomasal wall of cattle.

OBJECTIVE: To compare the content of substance P, vasoactive intestinal polypeptide, and neurofilament 200 in biopsy specimens taken from the abomasal wall of healthy cows of 2 breeds. SAMPLE POPULATION: Biopsy specimens taken from different sites of the abomasal wall from 20 German Holstein cows and 20 German Fleckvieh cows. PROCEDURES: Biopsy specimens were examined immunohistochemically, and the content of substance P, vasoactive intestinal polypeptide, and neurofilament 200 was determined by measuring the immunoreactive areas. RESULTS: Significant differences between the breeds were detected. Substance P-immuno-reactive area in the corpus abomasi was significantly smaller in the German Holsteins (geometric mean +/- geometric SD, 679 +/- 1.83 microm2) than in the German Fleckvieh cows (1,020 +/- 1.65 microm2). Concerning vasoactive intestinal polypeptide, differences between breeds were not significant. Overall nerve density in the antral abomasal wall was significantly greater in German Holsteins than in German Fleckvieh cows (immunoreactive areas for neurofilament 200 in German Holsteins was 4,842 +/- 1.29 microm2 and in German Fleckvieh cows was 3,333 +/- 1.63 microm2). Conclusions and Clinical Relevance-The significantly lower content of substance P in the corpus abomasi could explain why German Holstein cows are predisposed to abomasal displacement, compared with German Fleckvieh cows, in which this disease is a rare finding.

Integrin activation in bovine placentomes and in caruncular epithelial cells isolated from pregnant cows.

In the bovine synepitheliochorial placenta, restricted trophoblast invasion requires complex interactions of integrin receptors with proteins of the extracellular matrix (ECM) and integrin receptors of neighboring cells. Activated integrins assemble to focal adhesions and are linked to the actin cytoskeleton via signaling molecules including alpha-actinin (ACTN), focal adhesion kinase (PTK2 or FAK), phosphotyrosine, and talin (TLN1). Aims of this study were to assess integrin activation and focal adhesion assembly within epithelial cells of bovine placentomes and low-passage (not transformed) placentomal caruncular epithelial cells cultured on dishes coated with ECM proteins. Immunofluorescence analysis was performed to colocalize the signaling molecules ACTN, PTK2, phosphotyrosine, and TLN1 with each other and with beta(1)-integrin (ITGB1) in placentomal cryosections throughout pregnancy and in caruncular epithelial cells in vitro. Antibody specificity was confirmed by Western blot. Cells were cultured on uncoated dishes, and the dishes were coated with fibronectin (FN), laminin (LAMA), and collagen type IV (COL4), thereby statistically assessing cell number and qualitatively assessing the expression pattern of ITGB1, phosphotyrosine, and TLN1. Results demonstrated integrin activation and focal adhesion assembly in the placentome and that low-passage caruncular epithelial cells maintain integrin-associated properties observed in vivo. Expression and/or colocalization of signaling molecules with ITGB1 confirmed, for the first time, integrin activation and participation in "outside-in" and "inside-out" signaling pathways. The prominent role of ECM, and FN in particular, in integrin signaling is supported by the in vitro enhancement of proliferation and focal adhesion expression. Thus, this in vitro model provides excellent potential for further mechanistic studies designed to elucidate feto-maternal interactions in the bovine placentome.

Age-dependent changes in the pigeon bursa of Fabricius vasculature: a comparative study using light microscopy and scanning electron microscopy of vessel casts.

The present study was carried out to analyse the vascularization of the pigeon bursa cloacalis of Fabricius and to determine whether it undergoes age-dependent changes during its functionally most important growth period after hatching of the pigeon. Morphological assessment of vascular corrosion casts, studied qualitatively and quantitatively, was applied for the first time to investigate the vascularization of the pigeon pigeon bursa of Fabricius. This also allowed us to analyse the microvasculature and morphological aspects of the vessel interrelationships as occurring in the natural state. The casts were compared with histological sections stained by haematoxylin-eosin and by binding of the lectin e-PHA (Phaseolus vulgaris, erythroagglutinin) to blood vessels. The vascular architecture of the bursa of Fabricius of the pigeon revealed that the organ is irrigated via two pathways, first through the terminal capillary system of lymphoid follicles arising from the internal pudendal artery, and secondly through arteries originating from the cloacal vasculature of the collum of the organ supplying the periluminal capillary system of the pigeon bursa of Fabricius. Both systems are drained by a venous system which is collateral to the system of the internal pudendal artery and clearly functions as a direct link between the lumen and vasculature of the cloaca or gut, respectively, and the bursa fabricii. This could allow the lymphocytes to be confronted with antigens from the contents of the gut, and their subsequent transport into the secondary lymphoid organs of the organism. Our results demonstrate that the blood vessels, as major and supplying part of the lymphoid system of the bursa Fabricii, clearly reflect three different phases of development: the evolution phase from about day 20 until day 50 post-hatching, the mature phase from days 50 to 90, and the involution phase after day 90. During the evolution phase the density of the vessel system rapidly increases, while in the mature phase the vascular architecture is maintained. The involution phase is dominated by vascular degeneration combined with shrinkage of the whole organ. Therefore, the morphology of the vasculature distinctly reflects the functional status of this primary lymphoid organ during its lifespan.

The carnivore pregnancy: the development of the embryo and fetal membranes.

The aim of this research was to compare the morphological aspects during the development of pregnancy in dogs and cats, distinguishing features of the fetal membranes, such as yolk sac evolution and differentiation of hemangioblasts, and the degree of elaboration of the amnion and allantois. Canine and feline placentae from 20, 24, 35, 45 and 55 d of pregnancy were perfusion-fixed for histological investigation and vascular corrosion casts were produced. The casts were prepared for scanning electron microscopy (SEM) and the embryo and fetal membrane development was analyzed. The growth patterns of the conceptuses were compared with the organization of the placentation process, and changes of the morphology during pregnancy were recorded. In feline placentae, an incomplete zonary shape was present in 62.5% out of 60 studied cases. This was located distal to the insertion of the umbilical cord. In the lamellar zone, the interhemal membrane or placental barrier resembled endotheliochorial conditions, and the maternal-fetal microvascular blood flow interrelationship was of simple crosscurrent type. Dogs have a zonary placenta, completely surrounding the fetus, and complex lamellar organization of maternal and fetal tissues. At the border, two marginal hematomes with green colouration delimited the central placental girdle. The yolk sac consisted of one large sacculation with an inverted "T" shape and an enormous number of blood vessels; it had hemangioblast cells in contact with the epithelium. The amnion was avascular in early stages, but became vascularized by blood vessels of the internal allantoic membrane in later stages of pregnancy by intrinsic relation.

Platelet-activating factor receptor (PAF-R) and acetylhydrolase (PAF-AH) are co-expressed in immature bovine trophoblast giant cells throughout gestation, but not at parturition.

Platelet-activating factor (PAF) was associated with successful implantation in the cow, trophoblast invasiveness and angiogenesis. Bovine placentation is characterized by the limited invasion of trophoblast giant cells (TGC) into the maternal caruncular epithelium. TGC exhibit both endocrine activity and properties of tumor cells and may thus be targets of and mediators for the action of PAF. We examined PAF-receptor (PAF-R) and PAF-acetylhydrolase (PAF-AH) gene expression and localized mRNA and corresponding proteins in bovine placentomes throughout gestation and at parturition. PAF-R and PAF-AH protein and mRNA were highly expressed and colocalized in immature TGC from early gestation until near term, while mature TGC were negative. After the onset of parturition both PAF-R and PAF-AH were expressed in the maternal stroma, predominantly endothelial cells. The expression of PAF-R and PAF-AH in immature but not mature TGC during gestation implicates a role for PAF in the differentiation, maturation and function of bovine placentomal TGC. Placentomal angiogenesis could be mediated by binding of PAF to PAF-R present in endothelial cells. The parturition-related "switch" of PAF-R and PAF-AH from TGC to the maternal stroma suggests that PAF may participate in the regulation of parturition and in prepartum tissue programming.

Analysis of fetal and maternal microvasculature in ruminant placentomes by corrosion casting.

Vascular corrosion casting is a useful tool for studying the vascular architecture of complex organs. The synepitheliochorial placenta of ruminants is composed of two closed blood circuits, a fetal and a maternal one. The microvasculature of each circuit has the shape of the corresponding cotyledon (villous trees) and caruncle (crypts). These two compartments interdigitate with each other in a complementary fashion. Understanding three-dimensional vascular arrangements is facilitated by scanning electron microscopy of vascular corrosion casts. Methods to be used in the generation of vascular casts from fetal and maternal placentomal blood vessels are described, with special emphasis on casting resins and corrosion using potassium hydroxide. The procedure of splitting larger casts following gelatin embedding and freezing is also presented.

Pregnancy associated glycoprotein-1, -6, -7, and -17 are major products of bovine binucleate trophoblast giant cells at midpregnancy.

Pregnancy associated glycoproteins (PAGs) are extensively glycosylated secretory proteins of ruminant trophoblast cells. In cattle placenta several PAG cDNAs are expressed, but the variety of correspondent proteins and their degree of glycosylation are not well characterized. Thus, we purified PAGs by using a protocol which included a lectin (Vicia villosa agglutinin) affinity chromatography. Due to their specific glycosylation pattern, PAGs derived from binucleate trophoblast giant cells were highly enriched by this protocol. PAGs were purified from cotyledons of 2 day 100 placentas and from a single placenta at day 155 and 180. In all samples three major bands (75; 66; 56 kDa) were detected by one-dimensional SDS-PAGE. Mass-spectrometric analysis identified the 75 kDa band as a mixture of PAG-7 and PAG-6, the 66 kDa band as PAG-1 and the 56 kDa band as PAG-17. N-terminal sequencing of the day 100 sample confirmed the mass spectrometric identifications. Enzymatic release of N-glycans with peptide-N-glycanase-F from PAGs reduced the molecular weight to approximately 37 kDa which corresponds to the theoretical molecular mass of PAGs. Limited peptide-N-glycanase-F treatment revealed that all four N-glycosylation sites are quantitatively occupied in PAG-1. Compared to PAG-1 the number of potential N-glycosylation sites is lower in PAG-17 (three sites) and higher in PAG-6 and -7 (five and six sites, respectively). This suggests that the number of attached N-glycans is the main determinant of molecular mass of bovine PAGs. The degree of glycosylation may be a major factor regulating the plasma half life of PAGs.

Genome multiplication is a generalised phenomenon in placentomal and interplacentomal trophoblast giant cells in cattle.

The frequency of polyploidisation in bovine binucleate trophoblast giant cells (TGC) from placentomes (PL) and the interplacentomal allantochorion (AL) of six male fetuses with a crown-rump length between 3.5 and 103 cm was determined by in situ hybridisation with a chromosome-7-specific probe, using a probe specific for the Y chromosome to distinguish between maternal and fetal nuclei. The results showed that polyploid nuclei were essentially always of fetal origin. The frequency of tetraploid nuclei varied between 3% and 15% in both the placentomal and interplacentomal samples, with mean frequencies of 8.8% and 10.0% respectively. Octoploid nuclei were observed with a mean frequency of 1.1% in the interplacentomal samples, but were absent in samples from placentomes. Subsequent determination of nuclear DNA content by cytophotometric measurement of Feulgen-stained nuclei revealed that the frequency of nuclei with an 8C DNA content was several fold higher (AL 5.4%; PL 7.8%) than the frequency of octoploidy, suggesting that tetraploid TGC cells are arrested in the G2 phase of the cell cycle.

Microvascular architecture of the fetal cotyledons in water buffaloes (Bubalus bubalis) during different stages of pregnancy.

To elucidate the morphological background of physiological differences between bovine and buffalo gestation forty-two placentae ranging from the 3rd to 10th month of pregnancy were used to study the microvascular architecture of the fetal cotyledons in the buffalo. The tissues were prepared for light and scanning electron microscopy by paraformaldehyde fixation and corrosion casting of the fetal cotyledonary vascular system. Histology and vascular casts revealed the buffalo fetal cotyledons to consist of a series of conical villous trees changing from a wide to slender shape during pregnancy, and with a base strictly facing the fetal side. The branches of these trees, intermediate and terminal villi, projected radially from the stem, thus representing a rough type of villous tree. A second type of tree lacked these branches and was therefore termed smooth villus. The rough type was most prevalent, and the intermediate and terminal villi showed capillary complexes arranged in stories by the 4th to 5th month of gestation. The stories became broader and denser with the progress of pregnancy (6th to 7th month of gestation), due to extensive growth of new capillaries and simultaneous development of convolutions causing the vascular ridges of the terminal villi to appear bushy. Near term (9th to 10th month) the capillary system became very dense, particularly at the margin of the vascular ridges, leaving only narrow spaces for the corresponding maternal septal tissue. In detail, at its base the trunk of each villous tree contained a single central stem artery which originated from the allantochorionic arterial system, and 1-3 parallel peripheral stem veins. When approaching the cone tip, these vessels branched into new stem arteries and veins, each giving rise to arterioles and venules according to the principle vascularization of the stem villus first, followed by intermediate and terminal villi. The capillary complex of the terminal villi consisted of arterial capillary limbs, capillary loops with sinusoidal dilatations and anastomoses, and venous capillary limbs. The latter converged into venules of terminal and intermediate villi which joined stem veins leading into allantochorionic veins. In conclusion, the fetal vasculature of the buffalo placentome was greatly increased from early pregnancy to near term. This was denoted by the general development of stem villous trees and an increase in the volume and density of the capillary system of the terminal villi, reflecting the increasing need of materno-fetal substance exchange in the buffalo placenta, particularly near term.

Placentation in the alpaca Lama pacos.

Reproduction in South American camelids is poorly studied. To extend our knowledge of the development and cellular physiology of the placenta in the alpaca Lama pacos, we have examined specimens from day 150 of pregnancy to term. Morphological investigations using light, transmission and scanning electron microscopy, the histochemical localization of iron, alkaline and acid phosphatase activity, and the immunodetection of placental lactogen hormone were performed. Throughout pregnancy there was a progressive increase in the depths of folds on the uterine mucosa surface together with a thickening of the endometrium. Glandular cells exhibited PAS and acid phosphatase (AcP) positive secretion granules. In the chorion, giant trophoblast polyploid cells gradually became more numerous and larger. Non-giant cells exhibited positive granules for PAS, alkaline phosphatase (AkP) reaction and immunostaining for bovine placental lactogen hormone (PLH). SDS -PAGE electrophoresis and Western blotting procedures also confirmed the presence of a bovine PLH-like glycoprotein in the fetal alpaca placenta. Over the glandular openings, the chorion formed typical areolae, where the trophoblast exhibited AcP and PAS positive reactions. At these sites, the fetal endothelial cells contained iron-storage granules in their cytoplasm. The trophoblast-epithelial interface exhibited a complex microvillous interdigitation, in which an AkP reaction was very prominent. The chorionic capillaries progressively indented adjacent trophoblast cells. These data suggest that although the epitheliochorial alpaca placenta is diffuse, various trophoblast cell types and specialized areas of the maternofetal interface give the placenta micro-regional functions where histiotrophic nutrition, hormone production and molecular exchange are prevalent.

In bovine binucleate trophoblast giant cells, pregnancy-associated glycoproteins and placental prolactin-related protein-I are conjugated to asparagine-linked N-acetylgalactosaminyl glycans.

Bovine binucleate trophoblast giant cells (BNCs) produce large amounts of PAS-positive cytoplasmic granules. After fusion of BNCs with uterine epithelial cells, the contents of these granules are released into the maternal stroma which underlies the uterine epithelium. Histochemically, the granules can be labeled with N-acetylgalactosamine-specific lectins ( Dolichos biflorus, Vicia villosa, and Wisteria floribunda agglutinins) and with Phaseolus vulgaris leucoagglutinin. In this study, we used lectin western blot analysis of proteins from fetal cotyledons to characterize the lectin binding glycoproteins. Lectin western blots showed several bands. A main band of approximately 65 kDa was identified as pregnancy-associated glycoproteins (PAGs) and a double band at 34-35 kDa as prolactin-related protein-I (PRP-I) by their crossreactivity with specific antisera. Enzymatic cleavage of N-linked glycans with peptide- N-glycanase F abolished the lectin binding to PRP and PAGs in western blots, revealing that the lectins bound to asparagine-linked glycans. The high specificity of the lectins was used for the enrichment of PRP-I and PAGs from placental cotyledons with Vicia villosa lectin affinity chromatography. The occurrence of the relatively uncommon asparagine-linked N-acetylgalactosaminyl glycans on secretory proteins of the BNCs suggests a functional role of this specific glycosylation pattern.

Morphological characteristics of the tongue and its papillae in the donkey (Equus asinus): a light and scanning electron microscopical study.

The morphology of the donkey tongue and its papillae were investigated by macroscopy and by light and scanning electron microscopy in ten adult animals (six males and four females). The spatula-shaped tongues measured about 28 cm in length, 4.5 cm in breadth and 3.5 cm in thickness. Samples from different areas of four tongues were grossly examined and pieces were processed for light and scanning electron microscopy. Filiform papillae were distributed mainly on the dorsum of the tongue, being thin and relatively short at the apex, conical and scaly in the main part (triangular zone) of the body, and thin and longer at the caudal part of the body. Few of them were found on the lateral surfaces. Fungiform papillae appeared scattered mainly on the lateral surfaces. They were mostly rounded (about 1.0 mm in diameter), but lobulated forms were also observed. Filiform and fungiform papillae were both completely devoid of taste buds, indicating a more mechanical function. The vallate papillae were 2-3 in number, located at the most caudal part of the body. They were three to four times as large as the fungiform papillae (about 5.6 mm in diameter) each with a wide circular groove around the central bulbous projection. Secondary grooves originating from the primary one were also demonstrated. The vallate papillae contained many taste buds with taste pores opening deeply into the papillary groove. Fine filiform papillae were demonstrated on the bulb-like part of the vallate papillae. The donkey tongue had sinister and dexter well-developed sets of foliate papillae close to the basis of the palatoglossal arch. They were arranged in the form of numerous leaves separated by deep, variably wide grooves and contained a very large number of taste buds. It is believed that the existence of well developed foliate papillae in donkey may substitute the comparatively few vallate papillae in this species.

Occurrence of estrogen receptor alpha in bovine placentomes throughout mid and late gestation and at parturition.

The bovine placenta produces estrogens from the first trimester until the end of its life span. However, with the exception of the immediate prepartal and intrapartal phases, in which an involvement of placental estrogens has been suggested for the preparation of parturition, their function has not been elucidated yet. To test for a role of placental estrogens as local factors regulating placental growth and differentiation, placentomes from cows that were pregnant for 150, 220, 240, and 270 days, and parturient cows (3 animals per group) were screened immunohistochemically for the expression of estrogen receptor alpha (ERalpha). Indirect immunoperoxidase staining methods were applied using primary monoclonal antibodies (pmAbs) directed against the C-terminus (AER311, HT277) or the N-terminus (AER314, 1D5) of the ERalpha molecule. Both types of pmAbs identified ERalpha in stromal cells and capillary pericytes of the maternal caruncular septae. Using pmAb 1D5, the mean percentage of ERalpha-positive caruncular stromal cells decreased from 39.0% +/- 5.9% in pregnant cows to 17.5% +/- 8.3% at parturition (P = 0.011). Only pmAb recognizing the C-terminus identified ERalpha in the caruncular epithelium, in which positive reactions were found in all cells, with the exception of areas adjacent to the chorionic plate and to major chorionic villi, where the specific signal gradually faded and occasionally disappeared. No positive reactions were observed in the fetal part of the placentomes. The expression of ERalpha in bovine placentomes was further confirmed by the detection of ERalpha-specific mRNA by reverse transcriptase-polymerase chain reaction and by Western blot analysis. The results suggest a role for placental estrogens as paracrine factors involved in the regulation of placental growth and differentiation.