A Phyllodes-like Mammary Tumor in a Breeding Galago (Otolemur garnettii).

In humans, phyllodes tumors of the breast are rare fibroepithelial tumors that are further characterized as benign, borderline, or malignant according to their histomorphologic features. Phyllodes tumors are poorly responsive to treatment other than excision. NHP have a much lower frequency of mammary neoplasia than do humans, and none of the lesions reported previously in NHP are consistent with phyllodes tumors. Here we present the case of a mammary tumor in a northern greater galago (Otolemur garnettii) that was histologically characteristic of a malignant phyllodes tumor. An 11-y-old, multiparous, pregnant galago presented with a mass in the right middle mammary gland. A fine-needle aspirate yielded neoplastic epithelial cells. Because the animal was pregnant and showed no signs of skin ulceration, pain, or distress, she was allowed to deliver and nurse the infant. At 20 wk after initial presentation, the infant was weaned and the mother was euthanized. At necropsy, the mammary mass measured 3.5 × 2.5 × 1.5 cm, a 13-fold increase in volume since initial presentation. There was no evidence of metastasis in draining lymph nodes, lungs, or any other tissue examined. The tumor was composed of neoplastic stromal, glandular, and adipose tissues and was diagnosed as a malignant phyllodes tumor in light of its high stromal cellularity, high mitotic rate, and marked atypia. This tumor also exhibited liposarcomatous differentiation, which occurs frequently in malignant phyllodes tumors. To our knowledge, this report represents the first described case involving an NHP of a mammary tumor with characteristics consistent with human phyllodes tumors.

Female reproductive activity and its endocrine correlates in the African lesser bushbaby, Galago moholi.

Steroid hormones play an important role in female reproductive physiology and behaviour and are often used to monitor important female reproductive events. However, such studies are often attempted on captive populations alone, delivering limited data. One such example is the African lesser bushbaby, Galago moholi, for which contradicting observational data exist between captive and free-ranging populations, while hormonal analyses have only been obtained from a single captive population. To extend and rectify the limited information, we monitored faecal progestagen and oestrogen metabolite levels across various important life history stages of both captive and free-ranging G. moholi. We additionally recorded changes in vaginal state as well as the occurrence of reproductive and aggressive behaviour throughout the study. Data from our captive population revealed an ovarian cycle length of 33.44 ± 0.59 days (mean ± SD), with follicular and luteal phases of 14.2 ± 1.0 and 19.1 ± 1.5 days, respectively, and an average pregnancy length of 128 ± 3.3 days. The initiation of female reproductive activity was closely linked to an oestrus-related increase in faecal oestrogen metabolite levels. Four of the seven captive females monitored in our study conceived during the May mating period, with one additional female fertilised in September, supporting the idea that the September mating period functions as a back-up for female G. moholi. Identified benchmark faecal progestagen metabolite levels (non-pregnant: >1 µg/g dry weight (DW), pregnant: >9 µg/g DW) should help researchers to determine pregnancy status of randomly wild-caught females in even a cross-sectional study setup.

Uterine gland microstructure in the pregnant and the non-pregnant lesser galago (galago senegalensis) / Microestructura de la glándula uterina en la gálago menor (galago senegalensis) preñada y no preñada

The histology and the ultrastructure of the uterine glands of the lesser bushbaby (Galago senegalensis) were studied in six specimens (5 pregnant and one non pregnant) which were fixed partly with bouin's fixative and part with 2.5% glutaraldehyde in 0.1 M cacodylate buffer. An overview of the main results revealed uterine glands in the non-pregnant uterus are rudimentary and scarce within the mucosa. In early pregnancy (first trimester) the uterine glands profiles appear in clusters. In late stage pregnancy (third trimester) the uterine gland profiles appear opposite chorionic vesicles. In the later stages of gestation maternal glandular epithelium consisted mainly of simple columnar epithelium. The cells had abundant flattened cisternae of granular endoplasmic reticulum usually with an apical-basal orientation. Their nuclei had abundant euchromatin relative to the amount of heterochromatin. They also had a prominent Golgi apparatus quite characteristic of protein synthesizing cells. The basal plasmalemma was thrown into infoldings that have the effect of increasing the surface area across which nutrients could pass from the maternal circulation and are secreted by the cell as histiotrophe. Both physiologic hypertrophy and hyperplasia of the uterine glands are observed to occur with advancement of pregnancy.

La histología y ultraestructura de las glándulas uterinas de la gálago menor (Galago senegalensis) fueron estudiadas en seis ejemplares (5 preñadas y 1 no preñada). Una parte de las glándula se fijó con Bouin y otra con glutaraldehído al 2,5% en tampón cacodilato 0,1 M. Una visión general de los principales resultados reveló que las glándulas uterinas en el útero no gestante son rudimentarias y escasas dentro de la mucosa. Al principio de la preñez (primer trimestre) las glándulas uterinas aparecen en racimos. En la última etapa de la preñez (tercer trimestre) las glándulas uterinas aparecen opuestas a las vesículas coriónicas. En las últimas etapas de gestación el epitelio glandular materno consiste principalmente en epitelio cilíndrico simple. Las células tenían abundantes cisternas aplanadas en el retículo endoplásmico rugoso, por lo general con una orientación apico-basal. Sus núcleos tenían abundante eucromatina en relación con la cantidad de heterocromatina. También tenían un aparato de Golgi prominente bastante característico de células que sintetizan proteínas. El plasmalema basal fue rechazado en repliegues que psoeen el efecto de aumentar el área de superficie a través del cual los nutrientes podrían pasar desde la circulación materna y son secretadas por las célula como histiotrofo. Hipertrofia fisiológica e hiperplasia de las glándulas uterinas se observaron con el avance de la preñez.

Gonadoblastoma in the ovaries of a lesser galago (Galago senegalensis braccatus).

An enlarged right ovary was removed from a 14-year-old lesser galago (Galago senegalensis braccatus). Cytological preparations consisted of a heterogeneous population of neoplastic cells admixed with extracellular hyaline structures and cell-free nuclei. Microscopically, the ovary was replaced with gonadoblastoma and was composed of nests of germinal cells, including large oocyte-like cells, and sex cord-stromal cells arranged in palisading patterns around the germinal cells, the periphery of the nests and around extracellular hyaline material. The animal died 2 years after initial diagnosis. Necropsy examination revealed gonadoblastoma in the left ovary. The germinal cells of the tumour in the right and left ovaries were immunoreactive for calretinin, OCT3/4, PGP 9.5, Ki67 and/or faintly for cytokeratins. Sex cord-stromal cells were immunoreactive for calretinin, OCT3/4, GATA-4, E-cadherin and vimentin. Luteinized sex cord-stromal cells were immunoreactive for inhibin-alpha. The extracellular hyaline material was immunoreactive for laminin. This is the first case of gonadoblastoma in a non-human primate.

First description of demodicosis in 12 galagos (Galago senegalensis).

Twelve Galago senegalensis from the Moscow Zoo were presented with papular to nodular (2-11 mm) lesions on the pinnae, containing a white, waxy material. Microscopic examination revealed large numbers of mites consistent with the morphology of Demodex spp. mites. Nine animals were treated with ivermectin, 600 µg/kg/day topically, orally or subcutaneously for 3-10 months, while one remained untreated. All the treated animals achieved clinical remission. The control animal was still affected and died 11 months later due to pneumonia and possible eosinophilic leukaemia. No adverse effects were noted in any animals during the treatment. No animal relapsed in the 13-19 months follow-up period. To the authors' knowledge, this is the first report of demodicosis in G. senegalensis. The use of ivermectin in G. senegalensis was safe, although its effectiveness in the treatment of demodicosis needs further investigation.

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.

Structural analysis and mapping of DNase I hypersensitivity of HS5 of the beta-globin locus control region.

The beta-globin locus control region (LCR) is a cis regulatory element that is located in the 5' part of the locus and confers high-level erythroid lineage-specific and position-independent expression of the globin genes. The LCR is composed of five DNase I hypersensitive sites (HSs), four of which are formed in erythroid cells. The function of the 5'-most site, HS5, remains unknown. To gain insights into its function, mouse HS5 was cloned and sequenced. Comparison of the HS5 sequences of mouse, human, and galago revealed two extensively conserved regions, designated HS5A and HS5B. DNase I hypersensitivity mapping revealed that two hypersensitive sites are located within the HS5A region (designated HS5A(major) and HS5A(minor)), and two are located within the HS5B region (HS5B(major), HS5B(minor)). The positions of each of these HSs colocalize with either GATA-1 or Ap1/NF-E2 motifs, suggesting that these protein binding sites are implicated in the formation of HS5. Gel retardation assays indicated that the Ap1/NF-E2 motifs identified in murine HS5A and HS5B interact with NF-E2 or similar proteins. Studies of primary murine cells showed that HS5 is formed in all hemopoietic tissues tested (fetal liver, adult thymus, and spleen), indicating that this HS is not erythroid lineage specific. HS5 was detected in murine brain but not in murine kidney or adult liver, suggesting that this site is not ubiquitous. The presence of GATA-1 and NF-E2 motifs (which are common features of the DNase I hypersensitive sites of the LCR) suggests that the HS5 is organized in a manner similar to that of the other HSs. Taken together, our results suggest that HS5 is an inherent component of the beta-globin locus control region.

Identity of the beta-globin locus control region binding protein HS2NF5 as the mammalian homolog of the notch-regulated transcription factor suppressor of hairless.

Previously, we characterized a DNA-binding protein, HS2NF5, that bound tightly to a conserved region within hypersensitive site 2 (HS2) of the human beta-globin locus control region (LCR) (Lam, L. T. , and Bresnick, E. H. (1996) J. Biol. Chem. 271, 32421-32429). The beta-globin LCR controls the chromatin structure, transcription, and replication of the beta-globin genes. We have now purified HS2NF5 to near-homogeneity from fetal bovine thymus. Two polypeptides of 56 and 61 kDa copurified with the DNA binding activity. The two proteins bound to the LCR recognition site with an affinity (3.1 nM) and specificity similar to mouse erythroleukemia cell HS2NF5. The amino acid sequences of tryptic peptides of purified HS2NF5 revealed it to be identical to the murine homolog of the suppressor of hairless transcription factor, also known as recombination signal binding protein Jkappa or C promoter binding factor 1 (CBF1). The CBF1 site within HS2 resides near sites for hematopoietic regulators such as GATA-1, NF-E2, and TAL1. An additional conserved, high affinity CBF1 site was localized within HS4 of the LCR. As CBF1 is a downstream target of the Notch signaling pathway, we propose that Notch may modulate LCR activity during hematopoiesis.

Evolutionary sequence comparisons using high-density oligonucleotide arrays.

We explored the utility of high-density oligonucleotide arrays (DNA chips) for obtaining sequence information from homologous genes in closely related species. Orthologues of the human BRCA1 exon 11, all approximately 3.4 kb in length and ranging from 98.2% to 83.5% nucleotide identity, were subjected to hybridization-based and conventional dideoxysequencing analysis. Retrospective guidelines for identifying high-fidelity hybridization-based sequence calls were formulated based upon dideoxysequencing results. Prospective application of these rules yielded base-calling with at least 98.8% accuracy over orthologous sequence tracts shown to have approximately 99% identity. For higher primate sequences with greater than 97% nucleotide identity, base-calling was made with at least 99.91% accuracy covering a minimum of 97% of the sequence. Using a second-tier confirmatory hybridization chip strategy, shown in several cases to confirm the identity of predicted sequence changes, the complete sequence of the chimpanzee, gorilla and orangutan orthologues should be deducible solely through hybridization-based methodologies. Analysis of less highly conserved orthologues can still identify conserved nucleotide tracts of at least 15 nucleotides and can provide useful information for designing primers. DNA-chip based assays can be a valuable new technology for obtaining high-throughput cost-effective sequence information from related genomes.

Gastrointestinal tumors observed in nonhuman primates at the German primate center.

Twenty-six gastrointestinal tumors were observed in twenty-three nonhuman primates during routine necropsies at the German Primate Center, Göttingen. The majority (15 cases) were colorectal mucoid adenocarcinomas in cotton-top tamarins (Saguinus oedipus), which in two animals were associated with gastric adenomas. Three cases of small intestinal mucoid adenocarcinomas occurred in common marmosets (Callithrix jacchus). One colonic leiomyoma was observed in a dwarf galago (Galagoides demidovii) and another one in a cotton top tamarin. Singular findings were a tubular adeno-carcinoma of the ileo-caecal valve in a saddle-backed tamarin (Saguinus fuscicollis) and a lymphosarcoma of jejunum, ileum, and colon in another saddle-backed tamarin. Multiple tubular adeno-carcinomas of the colonic diverticles occurred in an aged rhesus monkey (Macaca mulatta). The findings are discussed in comparison to the situation in man.

Effects of estradiol-17beta and progesterone on mating behavior in female lesser bushbabies (Galago moholi) in captivity.

The effects of ovariectomy, estradiol-17beta, and progesterone on female sexuality were investigated in a prosimian primate, the lesser bushbaby (Galago moholi). Each of eight ovariectomized, adult bushbabies was pair-tested with a male partner in the absence of hormone treatment, during estradiol-17beta (E2) treatment (3.125 microg/0.1 ml), and during E2 + progesterone (0.125 mg/0.1 ml) treatment. Pretreatment females were sexually nonreceptive and nonattractive toward the males. In contrast, E2 treatment elicited vaginal opening, partial or complete epithelial cell cornification, and female receptivity in all females and elicited the complete mating repertoire in seven pairs. Progesterone treatment opposed the facilitatory effects of E2 by inhibiting female receptivity and epithelial cell cornification and inducing vaginal closure. Behaviors were quantified following changes in hormone treatment, vaginal physiology, and the presence or absence of intromission. Female attractiveness (male sexual arousal) was initiated during vaginal swelling and was maintained for the duration of vaginal opening, while female receptivity was manifested exclusively during the period of vaginal opening. Female proceptive behavior was mainly associated with the period of receptivity. This study provided evidence of a strict hormonal regulation of both behavioral and nonbehavioral aspects of female sexuality in a prosimian.

Cholinergic axonal dystrophy and mitochondrial pathology in prosimian primates.

Progressive cholinergic axonal dystrophy, cholinergic denervation, and generalized gliosis begin in the prosimian primate species Otolemur at 10% of maximum life span. In these same animals, extensive cerebral beta-amyloidosis follows relatively more abruptly at 50% of maximum life span. In contrast, even at maximum life span, the prosimian primate species Galago senegalensis Moholi, Microcebus murinus, and Eulemur fulvus collaris and insectivore species T. belangeri are either spared or much less affected. In this report, we further document this progressive cholinergic denervation in Otolemur which involves first projections of the pedunculopontine nucleus (PPN, CH5-6) and later projections of CH1-4 cholinergic nuclei, as well as other noncholinergic pathways. Affected cholinergic cell bodies and axons contain abnormal mitochondria with increased content of manganese superoxide dismutase (MnSOD). This syndrome correlates with moderate copper deficiency marked by diminished liver copper levels and cuproenzyme activities, carnitine deficiency possibly secondary to renal Fanconi syndrome, and evidence for stress inflammatory response activation. Mitochondrial pathology was observed in pancreatic islet cells, proximal renal tubule epithelial cells, and choroid plexus epithelial cells, and it involved central cholinergic neurons. In Otolemur garnetti, the degree of central cholinergic injury directly correlated to depression of liver copper stores. The Otolemur syndrome involves "sentinel" central cholinergic injury and selective mitochondrial pathology in cell classes defined by high mitochondrial content and/or metabolic activity and high content of nitric oxide synthetase and MnSOD. Environmental factors affecting copper and carnitine metabolism could interact with genetic defects or traits to produce abnormal and aggressive aging of Otolemur. Subclinical, cell-class specific mitochondrial dysfunction in these prosimian primates may be a model for human neurodegenerative diseases.

A preliminary investigation of the relationship between ovarian steroids, LH, reproductive behaviour and vaginal changes in lesser bushbabies (Galago moholi).

Urinary hormone profiles, determined by means of radioimmunoassays, were aligned with changes in mating behaviour and vaginal morphology and cytology during the ovarian cycles of adult, female lesser bushbabies (Galago moholi), a prosimian species. Intromission occurred in all seven females, 2.0 +/- 1.1 days (mean +/- SEM, median = 0) after the occurrence of vaginal opening, and lasted for 4.1 +/- 0.7 days. Three females subsequently gave birth. Vaginal swelling and labial reddening were initiated at least 2.5 +/- 0.5 days before vaginal opening and lasted 10.4 +/- 0.9 days. Pro-oestrous and vaginal oestrous smears coincided with vaginal opening, specifically during the period of mating. Concentrations of immunoreactive oestradiol increased during the first few days of vaginal opening in one pregnant female and in the nonpregnant females, and coincided with mating. Concentrations of immunoreactive progesterone did not show any temporal pattern for either pregnant or nonpregnant females. Concentrations of immunoreactive testosterone were generally higher during vaginal swelling and opening than during the remainder of the cycle in one pregnant female, whereas in nonpregnant females, no specific temporal pattern was evident. In both pregnant and nonpregnant females, immunoreactive LH concentrations increased during the period of vaginal swelling and opening, while in nonpregnant females increases were also apparent after vaginal closure. The data reported here are preliminary, and further research is necessary to establish conclusively patterns of excreted hormones during the reproductive cycles of Galago moholi and in other prosimian species.

Differential phylogenetic footprinting as a means to identify base changes responsible for recruitment of the anthropoid gamma gene to a fetal expression pattern.

Expression of the anthropoid (simian) gamma gene in fetal life contrasts with the exclusively embryonic expression pattern of the gamma-like genes of other eutherian mammals. To elucidate the factors responsible for this change in expression pattern, we utilized a strategy called differential phylogenetic footprinting (DPF). This strategy entails the following: (a) identification, within regulatory regions, of the gamma promoter, of individual nucleotides that differ between human (fetal expression), and galago (embryonic expression) gamma genes, (b) analysis of the effect of these nucleotide differences on the binding of nuclear proteins to human and galago sequences, and (c) assessment of the functional consequences of these binding changes in expression assays. The DPF analysis revealed several proteins that bind upstream from the CCAAT motif in the galago gamma promoter but do not bind to the corresponding region of the human gamma promoter. In transfection assays, binding of these proteins is associated with erythroid-specific repression of promoter strength. Binding sites for these proteins also occur near the CCAAT box of other embryonically expressed genes, including rabbit, mouse, and dwarf lemur gamma genes and the human epsilon globin gene. These data are consistent with the hypothesis that sequence changes near the proximal CCAAT box in the ancestral simian gamma gene may have facilitated a novel expression pattern by reducing the binding of repressors that act in the fetal stage.

Serotonin and serotonin receptors in the central auditory system.

Immunohistochemical and ligand-binding techniques were used to visualize the neurotransmitter serotonin and one of its receptors, the 5-HT1A subtype, in auditory nuclei of the brainstem. Serotonergic fibers and terminal endings were found in all auditory nuclei extending from the cochlear nucleus to the inferior colliculus, including the superior olivary complex and the nuclei of the lateral lemniscus. The density of the innervation varied between and within each nucleus. All serotonergic cell bodies were located outside the auditory nuclei. The 5-HT1A receptor subtype was found in the cochlear nucleus as well as in the inferior colliculus. With no serotonergic cell bodies present in the auditory nuclei, the present neuroanatomic and neurochemical findings support behavioral and neurophysiologic findings that the serotonergic system may modulate central auditory processing.

The effects of age on glutathione synthesis enzymes in lenses of Old World simians and prosimians.

The activities of gamma-glutamylcysteine synthetase and glutathione synthetase, the two enzymes required for glutathione synthesis, were determined as a function of age in lenses of three species of Old World higher primates: orangutan, pigtail monkey and olive baboon. These were compared to enzyme activities in lenses of two prosimians: mouse lemur and galago. gamma-Glutamylcysteine synthetase activity decreased as a function of age in all three Old World simians. The rate of decrease was greatest in the juvenile lenses. In contrast, the enzyme activity increased continuously with age in the galago lens. In the mouse lemur the enzyme activity increased per lens, but was constant when expressed as specific activity or as units per gram of lens. The loss of enzyme activity with age was limited to Old World higher primates apparently representing genetic change. Glutathione synthetase activity decreased logarithmically with age in the lenses of all five species.

Single copy sequences in galago DNA resemble a repetitive human retrotransposon-like family.

Galago DNA contains a few single copy sequences that are homologous to the human THE 1 family of repeats. Two of these galago loci have been isolated as genomic clones and their structures are compared to the THE 1 consensus sequence. Whereas the human sequence resembles a proretroviral transposon, the galago sequences provide no evidence for a proretroviral sequence organization. The two galago clones share a common repeat sequence, which is homologous to the U5 region of the THE 1 long terminal repeat. Immediately 3' to this repeat, each galago clone contains sequences that are homologous to mutually exclusive regions of the internal THE 1 sequence. Thus, the human THE 1 sequence can be represented as a mosaic of the two ancestrally related galago loci. The galago loci are transcribed in vivo, so that their conservation in the primate genome could be selected. Human THE 1 repeats apparently resulted by recruiting preexisting cellular sequences via a retrovirally mediated process.

Progesterone levels during pregnancy in the greater thick-tailed galago (Galago crassicaudatus).

Serum progesterone concentrations in Galago crassicaudatus were quantified at 3-week intervals throughout the 136-day pregnancy. Progesterone concentrations were significantly elevated over those of nonpregnant controls as early as 6 weeks after conception. Progesterone continued to increase throughout gestation. The progesterone profile in pregnant G. crassicaudatus quantitatively resembles that of chimpanzees and qualitatively resembles that of humans. In two animals that aborted, progesterone concentrations after abortion decreased to values comparable to those seen in nonpregnant animals.